minocycline and Brain-Ischemia

Various randomized-controlled clinical trials (RCTs) have investigated the neuroprotective role of minocycline in acute ischemic stroke (AIS) or acute intracerebral hemorrhage (ICH) patients. We sought to consolidate and investigate the efficacy and safety of minocycline in patients with acute stroke.. Literature search spanned through November 30, 2017 across major databases to identify all RCTs that reported following efficacy outcomes among acute stroke patients treated with minocycline vs. placebo: National Institute of Health Stroke Scale (NIHSS), Barthel Index (BI), and modified Rankin Scale (mRS) scores. Additional safety, neuroimaging and biochemical endpoints were extracted. We pooled mean differences (MD) and risk ratios (RR) from RCTs using random-effects models.. We identified 7 RCTs comprising a total of 426 patients. Of these, additional unpublished data was obtained on contacting corresponding authors of 5 RCTs. In pooled analysis, minocycline demonstrated a favorable trend towards 3-month functional independence (mRS-scores of 0-2) (RR = 1.31; 95% CI 0.98-1.74, p = 0.06) and 3-month BI (MD = 6.92; 95% CI - 0.92, 14.75; p = 0.08). In AIS subgroup, minocycline was associated with higher rates of 3-month mRS-scores of 0-2 (RR = 1.59; 95% CI 1.19-2.12, p = 0.002; I. Although data is limited, minocycline demonstrated efficacy and seems a promising neuroprotective agent in acute stroke patients, especially in AIS subgroup. Further RCTs are needed to evaluate the efficacy and safety of minocycline among ICH patients.

Topics: Brain Ischemia; Cerebral Hemorrhage; Humans; Minocycline; Neuroprotective Agents; Randomized Controlled Trials as Topic; Stroke

Minocycline, in animal models and 2 small randomized controlled human trials, is a promising neuroprotective agent in acute stroke. We analyzed the efficacy and safety of intravenous minocycline in acute ischemic and hemorrhagic stroke.. A multicenter prospective randomized open-label blinded end point evaluation pilot study of minocycline 100 mg administered intravenously, commenced within 24 hours of onset of stroke, and continued 12 hourly for a total of 5 doses, versus no minocycline. All participants received routine stroke care. Primary end point was survival free of handicap (modified Rankin Scale, ≤2) at day 90.. Ninety-five participants were randomized; 47 to minocycline and 48 to no minocycline. In the intention-to-treat population, 29 of 47 (65.9%) allocated minocycline survived free of handicap compared with 33 of 48 (70.2%) allocated no minocycline (rate ratio, 0.94; 95% confidence interval, 0.71-1.25 and odds ratio, 0.73; 95% CI, 0.31-1.71). A meta-analysis of the 3 human trials suggests minocycline may increase the odds of handicap-free survival by 3-fold (odds ratio, 2.99; 95% CI, 1.74-5.16) but there was substantial heterogeneity among the trials.. In this pilot study of a small sample of acute stroke patients, intravenous minocycline was safe but not efficacious. The study was not powered to identify reliably or exclude a modest but clinically important treatment effect of minocycline. Larger trials would improve the precision of the estimates of any treatment effect of minocycline.. http://www.anzctr.org.au. Unique identifier: ACTRN12612000237886.

Topics: Administration, Intravenous; Aged; Aged, 80 and over; Anti-Bacterial Agents; Brain Ischemia; Cerebral Hemorrhage; Female; Humans; Male; Middle Aged; Minocycline; Pilot Projects; Severity of Illness Index; Stroke; Treatment Outcome

Neuroprotection seeks to restrict injury to the brain parenchyma following an ischaemic insult by preventing salvageable neurons from dying. The concept of neuroprotection has shown promise in experimental studies, but has failed to translate into clinical success. Many reasons exist for this including the heterogeneity of human stroke and the lack of methodological agreement between preclinical and clinical studies. Even with the proposed Stroke Therapy Academic Industry Roundtable criteria for preclinical development of neuroprotective agents for stroke, we have still seen limited success in the clinic, an example being NXY-059, which fulfilled nearly all the Stroke Therapy Academic Industry Roundtable criteria. There are currently a number of ongoing trials for neuroprotective strategies including hypothermia and albumin, but the outcome of these approaches remains to be seen. Combination therapies with thrombolysis also need to be fully investigated, as restoration of oxygen and glucose will always be the best therapy to protect against cell death from stroke. There are also a number of promising neuroprotectants in preclinical development including haematopoietic growth factors, and inhibitors of the nicotinamide adenine dinucleotide phosphate oxidases, a source of free radical production which is a key step in the pathophysiology of acute ischaemic stroke. For these neuroprotectants to succeed, essential quality standards need to be adhered to; however, these must remain realistic as the evidence that standardization of procedures improves translational success remains absent for stroke.

Topics: Acute Disease; Animals; Benzenesulfonates; Brain Ischemia; Chelating Agents; Clinical Trials as Topic; Combined Modality Therapy; Diffusion of Innovation; Disease Models, Animal; Drug Evaluation, Preclinical; Egtazic Acid; Hematopoietic Cell Growth Factors; Humans; Hydroxymethylglutaryl-CoA Reductase Inhibitors; Hypothermia, Induced; Magnesium; Minocycline; NADPH Oxidases; Neuroprotective Agents; Pregnatrienes; Serum Albumin; Stroke; Thrombolytic Therapy; Translational Research, Biomedical

Sphingolipids represent a major class of lipids in which selected family members act as bioactive molecules that control diverse cellular processes, such as proliferation, differentiation, growth, senescence, migration and apoptosis. Emerging evidence reveals that sphingomyelinase/ceramide pathway plays a pivotal role in neurodegenerative diseases that involve mitochondrial dysfunction, oxidative stress and apoptosis. Minocycline, a semi-synthetic second-generation tetracycline derivative in clinical use for infection control, is also considered an effective protective agent in various neurodegenerative diseases in pre-clinical studies. Acting via multiple mechanisms, including anti-inflammatory, anti-oxidative and anti-apoptotic effects, minocycline is a desirable candidate for clinical trials in both acute brain injury as well as chronic neurodegenerative disorders. This review is focused on the anti-apoptotic and anti-oxidative mechanisms of minocycline against neurotoxicity induced by sphingomyelinase/ceramide in relation to neurodegeneration, particularly Alzheimer's disease and cerebral ischemia.

Topics: Alzheimer Disease; Antioxidants; Apoptosis; Brain Ischemia; Ceramides; Cyclic GMP; Humans; Minocycline; Mitochondria; Nitric Oxide; Oxidative Stress; Proto-Oncogene Proteins c-bcl-2; Signal Transduction; Sphingomyelin Phosphodiesterase; Thioredoxins

Due to a lack of efficient treatments, searching for novel therapies against acute ischemic stroke represents one of the main fields in neuropharmacology. In this review we summarize and discuss the role of mitochondrial participation in ischemia-induced neuronal death. Mitochondria are regarded as the main link between cellular stress signals and the execution of programmed death of nerve cells. Since it was described that the release of mitochondrial proteins such as cytochrome c, apoptosis inducing factor and endonuclease G are key elements in cell death pathways, they have been the focus of cell death studies. Changes in the permeability of the mitochondrial outer membrane result in a non-reversible step in cell death processes. Cytochrome c released from mitochondria binds in the cytoplasm to Apaf-1 to initiate the formation of an apoptosome, which then binds pro-caspase-9. Active caspase-9 cleaves "executioner" caspases, which in turn proceed to cleave key substrates in the cell. Thus, the identification of new targets might enable establishment of novel strategies for therapeutic research, in this case based on the molecular mechanisms of mitochondrial pathways, to improve the development of compounds for treatment of ischemia.

Topics: Anti-Bacterial Agents; Apoptotic Protease-Activating Factor 1; Brain Ischemia; Calcium; Cell Death; Cytochromes c; Energy Metabolism; Homeostasis; Minocycline; Mitochondria; Mitochondrial Membranes; Neurons; Permeability; Reactive Oxygen Species

Tissue plasminogen activator (tPA) is the only drug approved by the United States Food and Drug Administration for treatment of acute ischemic stroke. Because the drug must be used soon after symptom onset and is associated with intracerebral hemorrhage, tPA remains underutilized. Research has therefore focused on identifying other drugs that can be used concomitantly with tPA to improve the odds of a favorable recovery and to reduce the risk of intracerebral hemorrhage. Minocycline is a broad-spectrum antibiotic that has been found to be a neuroprotective agent in preclinical ischemic stroke models. Minocycline inhibits matrix metalloproteinase-9, a biomarker for intracerebral hemorrhage associated with tPA use. Minocycline is also an antiinflammatory agent and inhibits poly(ADP-ribose) polymerase-1. Minocycline has been safe and well tolerated in clinical trials. Additional safety and efficacy data are needed, and a phase III trial of minocycline with tPA in patients experiencing acute ischemic stroke is planned.

Topics: Animals; Brain Ischemia; Drug Repositioning; Female; Fibrinolytic Agents; Humans; Male; Matrix Metalloproteinase 9; Mice; Minocycline; Neuroprotective Agents; Rats; Stroke; Time Factors; Tissue Plasminogen Activator

An argument is made that small-vessel stroke, which usually results in lacunar infarction, is a serious medical problem. Therefore, it is surprising that only a few animal models exist that mimic small-vessel stroke and that these models have not been used for a systematic investigation of the genesis of lacunar infarctions. We make a case that the modified pial vessel class II disruption model mimics certain important aspects of lacunar infarctions, namely cavitation caused specifically by ischemia of smaller vessels. We found evidence that upregulation of inflammatory properties within a few days of inducing lesions prevents repopulation of the lesion with reactive astrocytes. We propose that this is the key mechanism by which cavitation occurs weeks later. We also found that treatment with minocycline after induction of lesions but before cavitation prevented the formation of the fluid-filled cavity. Rather than being walled off, the lesion apparently became part of the brain parenchyma and consisted of reactive astrocytes. We conclude that this new model can be used to investigate the mechanism of lacune formation and its prevention.

Topics: Animals; Anti-Bacterial Agents; Brain Infarction; Brain Ischemia; Cerebral Arteries; Disease Models, Animal; Encephalitis; Gliosis; Humans; Microcirculation; Minocycline; Pia Mater

The vascular events that happen during ischemic stroke worsen outcomes in patients by causing edema, hemorrhagic transformation, and general neurologic tissue compromise. In the past 2 decades, clinical trials in patients after ischemic stroke focused on neuroprotection, but these strategies have failed in providing actual benefit. Vascular protection represents a new field to be explored in acute ischemic stroke in order to develop new approaches to therapeutic intervention.. We identified tactics likely to provide vascular protection in patients with ischemic stroke. These tactics are based on knowledge of the molecular processes involved.. The pathologic processes due to vascular injury after an occlusion of a cerebral artery can be separated into acute (those occurring within hrs), subacute (hrs to days), and chronic (days to mo). Targets for intervention can be identified for all three stages. In the acute phase, superoxide is the predominant mediator, followed by inflammatory mediators and proteases in the subacute phase. In the chronic phase, proapoptotic gene products have been implicated. Many already-marketed therapeutic agents (statins, angiotensin modulators, erythropoietin, minocycline, and thiazolidinediones), with proven safety in patients, have been shown to have activity against some of the key targets of vascular protection.. Currently available pharmacologic agents are poised for clinical trials of vascular protection after acute ischemic stroke.

Topics: Acute Disease; Angiotensin-Converting Enzyme Inhibitors; Brain; Brain Edema; Brain Ischemia; Cerebral Hemorrhage; Chronic Disease; Erythropoietin; Humans; Hydroxymethylglutaryl-CoA Reductase Inhibitors; Minocycline; Stroke; Thiazolidinediones; Thrombolytic Therapy

Topics: Animals; Anti-Inflammatory Agents; Brain Ischemia; Enzyme Inhibitors; Humans; Minocycline; Mitogen-Activated Protein Kinases; Neuroprotective Agents; p38 Mitogen-Activated Protein Kinases; Stroke; Tetracyclines

Minocycline, in animal models and 2 small randomized controlled human trials, is a promising neuroprotective agent in acute stroke. We analyzed the efficacy and safety of intravenous minocycline in acute ischemic and hemorrhagic stroke.. A multicenter prospective randomized open-label blinded end point evaluation pilot study of minocycline 100 mg administered intravenously, commenced within 24 hours of onset of stroke, and continued 12 hourly for a total of 5 doses, versus no minocycline. All participants received routine stroke care. Primary end point was survival free of handicap (modified Rankin Scale, ≤2) at day 90.. Ninety-five participants were randomized; 47 to minocycline and 48 to no minocycline. In the intention-to-treat population, 29 of 47 (65.9%) allocated minocycline survived free of handicap compared with 33 of 48 (70.2%) allocated no minocycline (rate ratio, 0.94; 95% confidence interval, 0.71-1.25 and odds ratio, 0.73; 95% CI, 0.31-1.71). A meta-analysis of the 3 human trials suggests minocycline may increase the odds of handicap-free survival by 3-fold (odds ratio, 2.99; 95% CI, 1.74-5.16) but there was substantial heterogeneity among the trials.. In this pilot study of a small sample of acute stroke patients, intravenous minocycline was safe but not efficacious. The study was not powered to identify reliably or exclude a modest but clinically important treatment effect of minocycline. Larger trials would improve the precision of the estimates of any treatment effect of minocycline.. http://www.anzctr.org.au. Unique identifier: ACTRN12612000237886.

Topics: Administration, Intravenous; Aged; Aged, 80 and over; Anti-Bacterial Agents; Brain Ischemia; Cerebral Hemorrhage; Female; Humans; Male; Middle Aged; Minocycline; Pilot Projects; Severity of Illness Index; Stroke; Treatment Outcome

Plasma matrix metalloproteinase-9 levels predict posttissue plasminogen activator (tPA) hemorrhage.. The authors investigated the effect of minocycline on plasma matrix metalloproteinase-9 in acute ischemic stroke in the Minocycline to Improve Neurological Outcome in Stroke (MINOS) trial and a comparison group.. Matrix metalloproteinase-9 level decreased at 72 hours compared with baseline in MINOS (tPA, P=0.0022; non-tPA, P=0.0066) and was lower than in the non-MINOS comparison group at 24 hours (tPA, P<0.0001; non-tPA, P=0.0019).. Lower plasma matrix metalloproteinase-9 was seen among tPA-treated subjects in the MINOS trial. Combining minocycline with tPA may prevent the adverse consequences of thrombolytic therapy through suppression of matrix metalloproteinase-9 activity.

Topics: Aged; Anti-Bacterial Agents; Brain Ischemia; Female; Fibrinolytic Agents; Humans; Infusions, Intravenous; Male; Matrix Metalloproteinase 9; Matrix Metalloproteinase Inhibitors; Middle Aged; Minocycline; Stroke; Thrombolytic Therapy; Tissue Plasminogen Activator

Animal models of cerebral ischemia have improved our understanding of the pathophysiology and mechanisms involved in stroke, as well as the investigation of potential therapies. The potential of zebrafish to model human diseases has become increasingly evident. The availability of these models allows for an increased understanding of the role of chemical exposure in human conditions and provides essential tools for mechanistic studies of disease. To evaluate the potential neuroprotective properties of minocycline against ischemia and reperfusion injury in zebrafish and compare them with other standardized models. In vitro studies with BV-2 cells were performed, and mammalian transient middle cerebral artery occlusion (tMCAO) was used as a comparative standard with the zebrafish stroke model. Animals were subjected to ischemia and reperfusion injury protocols and treated with minocycline. Infarction size, cytokine levels, oxidative stress, glutamate toxicity, and immunofluorescence for microglial activation, and behavioral test results were determined and compared. Administration of minocycline provided significant protection in the three stroke models in different parameters analyzed. Both experimental models complement each other in their particularities. The proposal also strengthens the findings in the literature in rodent models and allows the validation of alternative models so that they can be used in further research involving diseases with ischemia and reperfusion injury.

Topics: Animals; Brain Ischemia; Disease Models, Animal; Humans; Infarction, Middle Cerebral Artery; Mammals; Minocycline; Neuroprotective Agents; Reperfusion Injury; Stroke; Zebrafish

Ischemic stroke is the most common cause of adult disability and one of the leading causes of death worldwide, with a serious socio-economic impact. In the present work, we used a new thromboembolic model, recently developed in our lab, to induce focal cerebral ischemic (FCI) stroke in rats without reperfusion. We analyzed selected proteins implicated in the inflammatory response (such as the RNA-binding protein HuR, TNFα, and HSP70) via immunohistochemistry and western blotting techniques. The main goal of the study was to evaluate the beneficial effects of a single administration of minocycline at a low dose (1 mg/kg intravenously administered 10 min after FCI) on the neurons localized in the penumbra area after an ischemic stroke. Furthermore, given the importance of understanding the crosstalk between molecular parameters and motor functions following FCI, motor tests were also performed, such as the Horizontal Runway Elevated test, CatWalk™ XT, and Grip Strength test. Our results indicate that a single administration of a low dose of minocycline increased the viability of neurons and reduced the neurodegeneration caused by ischemia, resulting in a significant reduction in the infarct volume. At the molecular level, minocycline resulted in a reduction in TNFα content coupled with an increase in the levels of both HSP70 and HuR proteins in the penumbra area. Considering that both HSP70 and TNF-α transcripts are targeted by HuR, the obtained results suggest that, following FCI, this RNA-binding protein promotes a protective response by shifting its binding towards HSP70 instead of TNF-α. Most importantly, motor tests showed that reduced inflammation in the brain damaged area after minocycline treatment directly translated into a better motor performance, which is a fundamental outcome when searching for new therapeutic options for clinical practice.

Topics: Animals; Brain Ischemia; Disease Models, Animal; Ischemic Stroke; Minocycline; Neurons; Rats; Rats, Sprague-Dawley; Stroke; Tumor Necrosis Factor-alpha

Hypoxia caused by ischemia induces acidosis and neuroexcitotoxicity, resulting in neuronal death in the central nervous system (CNS). Monoacylglycerol lipase (MAGL) is a modulator of 2-arachidonoylglycerol (2-AG), which is involved in retrograde inhibition of glutamate release in the endocannabinoid system. In the present study, we used positron emission tomography (PET) to monitor MAGL-positive neurons and neuroinflammation in the brains of ischemic rats. Additionally, we performed PET imaging to evaluate the neuroprotective effects of an MAGL inhibitor in an ischemic injury model.

Topics: Animals; Arachidonic Acids; Benzodioxoles; Brain; Brain Ischemia; Carbon Radioisotopes; Cell Hypoxia; Disease Models, Animal; Endocannabinoids; Glycerides; Infarction, Middle Cerebral Artery; Ischemic Stroke; Male; Minocycline; Monoacylglycerol Lipases; Neuroprotective Agents; Piperidines; Positron-Emission Tomography; Rats; Rats, Sprague-Dawley; Stroke; Tomography, X-Ray Computed

Increasing evidence suggests that microglia experience two distinct phenotypes after acute ischemic stroke (AIS): a deleterious M1 phenotype and a neuroprotective M2 phenotype. Promoting the phenotype shift of M1 microglia to M2 microglia is thought to improve functional recovery after AIS. Minocycline, a tetracycline antibiotic, can improve functional recovery after cerebral ischemia in pre-clinical and clinical research. However, the role and mechanisms of minocycline in microglia polarization is unclear.. Using the transient middle cerebral artery occlusion - reperfusion (MCAO/R) model, we treated mice with saline or different minocycline concentration (10, 25, or 50 mg/kg, i.p., daily for 2 wk) at 24 h after reperfusion. Neurobehavioral evaluation, rotarod test, and corner turning test were carried out on day 14 after reperfusion. Then, neuronal injury, reactive gliosis, and microglia polarization were performed on day 7 following MCAO/R. Finally, we treated primary microglial cultures with LPS (Lipopolysaccharide; 100 ng/mL) plus IFN-γ (20 ng/mL) 24 h to induce M1 phenotype and observed the effects of minocycline on the M1/M2-related mRNAs and the STAT1/STAT6 pathway.. We found that a 14-day treatment with minocycline increased the survival rate and promoted functional outcomes evaluated with neurobehavioral evaluation, rotarod test, and corner turning test. Meanwhile, minocycline reduced the brain infarct volume, alleviated neuronal injury, and suppressed reactive gliosis on day 7 following MCAO/R. Moreover, we observed an additive effect of minocycline on microglia polarization to the M1 and M2 phenotypes in vivo and in vitro. In the primary microglia, we further found that minocycline prevented neurons from OGD/R-induced cell death in neuron-microglia co-cultures via regulating M1/M2 microglia polarization through the STAT1/STAT6 pathway.. Minocycline promoted microglial M2 polarization and inhibited M1 polarization, leading to neuronal survival and neurological functional recovery. The findings deepen our understanding of the mechanisms underlying minocycline-mediated neuroprotection in AIS.

Topics: Animals; Anti-Bacterial Agents; Brain Ischemia; Cell Polarity; Cells, Cultured; Ischemic Stroke; Male; Mice; Mice, Inbred C57BL; Microglia; Minocycline; Recovery of Function; Signal Transduction; STAT1 Transcription Factor; STAT6 Transcription Factor

The poor clinical relevance of experimental models of stroke contributes to the translational failure between preclinical and clinical studies testing anti-inflammatory molecules for ischemic stroke. Here, we (i) describe the time course of inflammatory responses triggered by a thromboembolic model of ischemic stroke and (ii) we examine the efficacy of two clinically tested anti-inflammatory drugs: Minocycline or anti-CD49d antibodies (tested in stroke patients as Natalizumab) administered early (1 h) or late (48 h) after stroke onset. Radiological (lesion volume) and neurological (grip test) outcomes were evaluated at 24 h and 5 days after stroke. Immune cell responses peaked 48 h after stroke onset. Myeloid cells (microglia/macrophages, dendritic cells, and neutrophils) were already increased 24 h after stroke onset, peaked at 48 h, and remained increased-although to a lesser extent-5 days after stroke onset. CD8

Topics: Animals; Anti-Inflammatory Agents; Brain; Brain Ischemia; Disease Models, Animal; Ischemic Stroke; Male; Mice; Minocycline; Natalizumab; Reperfusion Injury; Thromboembolism

Mammalian target of rapamycin (mTOR) has been evidenced as a multimodal therapy in the pathophysiological process of Acute Ischemic Stroke (AIS). However, the pathway that minocycline targets mTOR signaling is not fully defined in the AIS pathogenesis. This study aims at the roles of minocycline on the mTOR signaling in the AIS process and further discovers the underlying mechanisms of minocycline involved in the following change of mTOR signaling-autophagy.. Cerebral ischemia/reperfusion (CIR) rat animal models were established with the transient suture occlusion into the middle cerebral artery. Minocycline (50mg/kg) was given by intragastric administration. The Morris water maze was used to test the cognitive function of animals. Immunohistochemistry and immunofluorescence were introduced for testing the levels of synaptophysin and PSD-95. Western blot was conducted for investigating the levels of mTOR, p-mTOR (Ser2448), p70S6, p-p70S6 (Thr389), eEF2k, p-eEF2k (Ser366), p-eIF4B (Ser406), LC3, p62, synaptophysin and PSD-95.. Minocycline prevents the cognitive decline of the MCAO stroke rats. Minocycline limits the expression of p-mTOR (Ser2448) and the downstream targets of mTOR [p70S6, p-p70S6 (Thr389), eEF2k, p-eEF2k (Ser366) and p-eIF4B (Ser406)] (P<0.01), while minocycline has no influence on mTOR. LC3-II abundance and the LC3-II/I ratio were upregulated in the hippocampus of the MCAO stroke rats by the minocycline therapy (P<0.01). p62 was downregulated in the hippocampus from the MCAO stroke rats administrated with minocycline therapy(P<0.01). The levels of SYP and PSD-95 were upregulated in the brain of the MCAO stroke rats administrated with minocycline therapy.. Minocycline prevents cognitive deficits via inhibiting mTOR signaling and enhancing the autophagy process, and promoting the expression of pre- and postsynaptic proteins (synaptophysin and PSD-95) in the brain of the MCAO stroke rats. The potential neuroprotective role of minocycline in the process of cerebral ischemia may be related to mitigating ischemia-induced synapse injury via inhibiting the activation of mTOR signaling.

Topics: Animals; Autophagy; Brain; Brain Ischemia; Male; Minocycline; Neuroprotective Agents; Rats; Rats, Wistar; Reperfusion Injury; Signal Transduction; TOR Serine-Threonine Kinases

Stroke induced white matter injury can induce marked neurological deficits even after relatively small infarcts, due to the tightly packed nature of white matter tracts especially in certain areas in the brain. Many drugs which were successful in the pre-clinical trials failed in clinical trials, which was attributed in part to the focus on grey matter injury completely and ignoring their effect on white matter. In this work we selected two known neuroprotective drugs (minocycline and progesterone) and examined their effect on white matter injury after focal cerebral ischemia/reperfusion injury in rats. Focal cerebral ischemia was induced in male Wistar rats (one-hour ischemia followed by reperfusion). Progesterone and minocycline were administered immediately after reperfusion onset. Infarct size, microglial activation and white matter injury were assessed and compared between the treatment and no-treatment groups and Sham operated animals. Our data showed that both progesterone and minocycline reduced infarct size, microglial activation and white matter injury. This work shows a new neuroprotective mechanism of both drugs, via white matter injury reduction, that can be exploited for stroke management. While the utility of either drugs as a sole agent in the management of stroke is questionable, there is a value of using either drugs as an adjuvant therapy to traditional stroke therapy, making use of the white matter protective effect that would improve outcome and facilitate healing after stroke.

Topics: Animals; Brain Ischemia; Female; Male; Minocycline; Neuroprotective Agents; Progesterone; Rats; Rats, Wistar; Reperfusion Injury; Stroke; White Matter

Topics: Brain Ischemia; Cerebral Hemorrhage; Humans; Minocycline

Minocycline, an anti-infective agent of a tetracycline derivative, is reported to improve behavioral functional recovery after cerebral ischemia via enhancing the levels of brain-derived neurotrophic factor (BDNF). However, the precise mechanisms that minocycline targets to enhance the expression of BDNF are not fully defined. In the present study, we observed the neuroprotective effect and its potential mechanisms of minocycline using oxygen-glucose deprivation/reoxygenation (OGD/R)-treated N2a cells. We found that 50 µM minocycline protected against neuronal apoptosis induced by OGD/R injury, with increased expression ratio of Bcl-2/Bax and reduced expression of caspase-3. Interestingly, minocycline resulted in the up-regulation of only BDNF protein, not BDNF mRNA in N2a cells treated with OGD/R. Furthermore, we found that minocycline inhibited OGD/R-induced up-regulation of miR-155 targeted BDNF transcripts. Moreover, miR-155 mimic could partially abolish the neuroprotective effects of minocycline via inhibiting the levels of BDNF protein. These findings suggest that minocycline is neuroprotective against ischemic brain injury through their modulation of miR-155-mediated BDNF repression.

Topics: Animals; Apoptosis; Brain; Brain Ischemia; Brain-Derived Neurotrophic Factor; Glucose; Mice; MicroRNAs; Minocycline; Neurons; Neuroprotective Agents; Oxygen; Signal Transduction

LKB1/AMPK signaling pathway, as a metabolic checkpoint, is involved in the pathogenesis of cerebral ischemia injury. Minocycline, a tetracycline derivative, protects against cerebral ischemia via reducing inflammation, oxidative stress, and apoptosis. The aim of the study was to evaluate the influence of minocycline on oxidative biomarkers and LKB1/AMPK signaling pathway in Wistar rats with focal cerebral ischemia injury and to clarify the neuroprotective mechanism of minocycline against focal cerebral ischemia injury.. The focal cerebral ischemia injury of Wistar rats was established by inserting a thread into the left middle cerebral artery. 2,3,5-Triphenyltetrazolium chloride (TTC) staining was used to label infarct volume. The levels of MDA and LPO were measured with a biochemical assay. All other items were determined by Western blotting.. Minocycline decreased cerebral infarct volume, but had no effects on neurological scores. Minocycline improved the biological activity of GPx-1/2, GSS and GR, while limited the GGT1 activity in the hippocampus of cerebral ischemia-reperfusion rats. Minocycline also elevated the biological activity of SOD and counteracted lipid peroxidation. Minocycline enhanced the activity of both LKB1 and the levels of the three AMPK subunits in the hippocampus of cerebral ischemia-reperfusion rats.. Minocycline effectively inhibits oxidative stress via modulating antioxidative enzymes and activating the LKB1/AMPK signaling pathway in the process of acute cerebral infarct.

Topics: AMP-Activated Protein Kinase Kinases; AMP-Activated Protein Kinases; Animals; Brain Ischemia; Male; Minocycline; Oxidative Stress; Protein Serine-Threonine Kinases; Rats; Rats, Wistar; Signal Transduction

Memory deficit is the most visible symptom of cerebral ischemia that is associated with loss of pyramidal cells in CA1 region of the hippocampus. Oxidative stress and inflammation may be involved in the pathogenesis of ischemia/reperfusion (I/R) damage. Minocycline, a semi-synthetic tetracycline derived antibiotic, has anti-inflammatory and antioxidant properties. We evaluated the neuroprotective effect of minocycline on memory deficit induced by cerebral I/R in rat. I/R was induced by occlusion of common carotid arteries for 20min. Minocycline (40mg/kg, i.p.) was administered once daily for 7days after I/R. Learning and memory were assessed using the Morris water maze test. Nissl staining was used to evaluate the viability of CA1 pyramidal cells. The effects of minocycline on the microglial activation was also investigated by Iba1 (Ionized calcium binding adapter molecule 1) immunostaining. The content of malondialdehyde (MDA) and pro-inflammatory cytokines (IL-1β and TNF-α) in the hippocampus were measured by thiobarbituric acid reaction substances method and ELISA, respectively. Minocycline reduced the increase in escape latency time and in swimming path length induced by cerebral I/R. Furthermore, the ischemia-induced reduction in time spent in the target quadrant during the probe trial was increased by treatment with minocycline. Histopathological results indicated that minocycline prevented pyramidal cells death and microglial activation induced by I/R. Minocycline also reduced the levels of MDA and pro-inflammatory cytokines in the hippocampus in rats subjected to I/R. Minocycline has neuroprotective effects on memory deficit induced by cerebral I/R in rat, probably via its anti-inflammatory and antioxidant properties.

Topics: Animals; Anti-Inflammatory Agents; Antioxidants; Brain Ischemia; Cognitive Dysfunction; Disease Models, Animal; Hippocampus; Inflammation; Ischemic Attack, Transient; Learning; Male; Memory; Memory Disorders; Minocycline; Neuroprotective Agents; Oxidants; Oxidative Stress; Pyramidal Cells; Rats; Rats, Wistar; Reperfusion; Reperfusion Injury; Tumor Necrosis Factor-alpha

Cerebral ischemia leads to multifaceted injury to the brain. A polytherapeutic drug that can be administered immediately after reperfusion may increase protection to the brain by simultaneously targeting multiple deleterious cascades. This study evaluated efficacy of the combination of three clinically approved drugs: lamotrigine, minocycline, and lovastatin, using two mouse models: global and focal cerebral ischemia induced by transient occlusion of the common carotid arteries or the middle cerebral artery, respectively. In vitro, the combination drug, but not single drug, protected neurons against oxygen-glucose deprivation (OGD)-induced cell death. The combination drug simultaneously targeted cell apoptosis and DNA damage induced by ischemia. Besides acting on neurons, the combination drug suppressed inflammatory processes in microglia and brain endothelial cells induced by ischemia. In a transient global ischemia model, the combination drug, but not single drug, suppressed microglial activation and inflammatory cytokine production, and reduced neuronal damage. In a transient focal ischemia model, the combination drug, but not single drug, attenuated brain infarction, suppressed infiltration of peripheral neutrophils, and reduced neurological deficits following ischemic stroke. In summary, the combination drug confers a broad-spectrum protection against ischemia/reperfusion (I/R) injury and could be a promising approach for early neuroprotection after out-of-hospital cardiac arrest or ischemic stroke.

Topics: Animals; Apoptosis; Brain; Brain Ischemia; Drug Therapy, Combination; Female; Lamotrigine; Lovastatin; Mice; Mice, Inbred C57BL; Minocycline; Neurons; Neuroprotective Agents; Reperfusion Injury; Triazines

Brain ischemia causes irreversible damage to functional neurons in cases of infarct. Promoting endogenous neurogenesis to replace necrotic neurons is a promising therapeutic strategy for ischemia patients. The neuroprotective role of sevoflurane preconditioning implies that it might also enhance endogenous neurogenesis and functional restoration in the infarct region. By using a transient middle cerebral artery occlusion (tMCAO) model, we discovered that endogenous neurogenesis was enhanced by sevoflurane preconditioning. This enhancement process is characterized by the promotion of neuroblast proliferation within the subventricular zone (SVZ), migration and differentiation into neurons, and the presence of astrocytes and oligodendrocytes at the site of infarct. The newborn neurons in the sevoflurane preconditioning group showed miniature excitatory postsynaptic currents (mEPSCs), increased synaptophysin and PSD95 staining density, indicating normal neuronal function. Furthermore, long-term behavioral improvement was observed in the sevoflurane preconditioning group consistent with endogenous neurogenesis. Further histological analyses showed that sevoflurane preconditioning accelerated microglial activation, including migration, phagocytosis and secretion of brain-derived neurotrophic factor (BDNF). Intraperitoneal injection of minocycline, a microglial inhibitor, suppressed microglial activation and reversed neurogenesis. Our data showed that sevoflurane preconditioning promoted microglial activities, created a favorable microenvironment for endogenous neurogenesis and accelerated functional reconstruction in the infarct region.

Topics: Animals; Axons; Brain Injuries; Brain Ischemia; Brain-Derived Neurotrophic Factor; Cell Differentiation; Cell Movement; Cell Proliferation; Disease Models, Animal; Ischemic Preconditioning; Male; Methyl Ethers; Microglia; Minocycline; Neurogenesis; Neurons; Phagocytosis; Rats; Sevoflurane

Cerebral ischemia leads to memory impairment that is associated with loss of hippocampal CA1 pyramidal neurons. Neuroinflammation and oxidative stress may be implicated in the pathogenesis of ischemia/reperfusion damage. Minocycline has anti-inflammatory and antioxidant properties. We investigated the neuroprotective effects of minocycline in rats subjected to cerebral ischemia/reperfusion injury. Thirty male rats were divided into three groups: control, sham, and minocycline-pretreated group. Minocycline (40 mg/kg) was injected intraperitoneally immediately before surgery, and then ischemia was induced by occlusion of common carotid arteries for 20 min. Seven days after reperfusion, the Morris water-maze task was used to evaluate memory. Nissl staining was also performed to analyze pyramidal cell damage. We measured the contents of malondialdehyde and proinflammatory cytokines in the hippocampus by the thiobarbituric acid method and enzyme-linked immunosorbent assay, respectively. Microglial activation was also investigated by Iba1 immunostaining. The results showed that pretreatment with minocycline prevented memory impairment induced by cerebral ischemia/reperfusion. Minocycline pretreatment also significantly attenuated ischemia-induced pyramidal cell death and microglial activation in the CA1 region and reduced the levels of malondialdehyde and proinflammatory cytokines (interleukin-1β and tumor necrosis factor-α) in the hippocampus of ischemic rats. Minocycline showed neuroprotective effects on cerebral ischemia-induced memory deficit probably through its anti-inflammatory and antioxidant activities.

Topics: Animals; Anti-Inflammatory Agents; Antioxidants; Brain Ischemia; Disease Models, Animal; Enzyme-Linked Immunosorbent Assay; Inflammation; Male; Maze Learning; Memory Disorders; Minocycline; Neuroprotective Agents; Oxidative Stress; Rats; Reperfusion Injury

Chronic alcohol intoxication (CAI) increases both morbidity and mortality of stroke patients. Despite the high prevalence of CAI and ischemic stroke, studies addressing their comorbidity and/or protective alternatives remain scarce. Thus, the influence of CAI on both stroke outcome and minocycline treatment (recognized for its neuroprotective effect) was investigated. Female Wistar rats (35 days old) were treated with water or ethanol (6.5 g/kg/day, 22.5% w/v) for 55 days. Then, focal ischemia was induced by endothelin-1 in the motor cortex. Two hours later, four doses of 50 mg/kg of minocycline every 12 hours followed by five doses of 25 mg/kg every 24 hours were administered. Behavioral performance (open field and rotarod tests) and immunohistochemical (cellular density, neuronal death, and astrocytic activation) and biochemical (lipid peroxidation and nitrite levels) analyses were performed. CAI increased motor disruption, nitrite and lipid peroxidation levels, and neuronal loss caused by ischemia, whereas it reduced the astrogliosis. Minocycline was effective in preventing the motor and tissue damage caused by stroke. However, these effects were attenuated when CAI preceded stroke. Our data suggest that CAI beginning in adolescence contributes to a worse outcome in ischemic stroke survivors and reduces the benefits of minocycline, possibly requiring adjustments in therapy.

Topics: Alcoholic Intoxication; Animals; Behavior, Animal; Brain Ischemia; Disease Models, Animal; Endothelin-1; Female; Immunohistochemistry; Lipid Peroxidation; Minocycline; Motor Cortex; Neuroprotective Agents; Nitrites; Rats; Rats, Wistar

Minocycline, a tetracycline antibiotic, has shown anti-inflammatory effects in cerebral ischemia and neurodegenerative disease; however, the molecular mechanisms underlying this effect have not been clearly identified. Since NLRP3 inflammasome activation controls the maturation and release of proinflammatory cytokines, especially interleukin-1β (IL-1β) and IL-18 in ischemia stroke, we suppose that minocycline may be involved in the regulation of NLRP3 inflammasome activation.. We investigated the effects of minocycline on NLRP3 inflammasome activation using the transient middle cerebral artery occlusion (tMCAO) mouse model and an in vitro oxygen-glucose deprivation/reoxygenation injury model in BV2 microglial cells.. We found that minocycline administrated 1 h after reperfusion can improve neurological disorder, reduce infarct volume, and alleviate cerebral edema. Meanwhile, we showed that minocycline prevented the activation of microglias and attenuated NLRP3 inflammasome signaling after tMCAO injury. Furthermore, we found that the pretreatment of minocycline significantly inhibited signal 1 and signal 2 of NLRP3 inflammasome activation in BV2 cells.. We demonstrated that minocycline can ameliorate ischemia-induced brain damage via inhibiting NLRP3 inflammasome activation.

Topics: Animals; Anti-Bacterial Agents; Brain Ischemia; Cell Line; Dose-Response Relationship, Drug; Inflammasomes; Male; Mice; Mice, Inbred C57BL; Microglia; Minocycline; NLR Family, Pyrin Domain-Containing 3 Protein; Stroke

Even after decades of intensive studies, therapeutic options for patients with stroke are rather limited. Thrombolytic drugs effectively treat the very acute stage of stroke, and several neuroprotectants that are designed to treat secondary injury following stroke are being tested in clinical trials. However, these pharmacological approaches primarily focus on acute stroke recovery, and few options are available for treating chronic stroke patients. In recent years, stem cell-mediated regenerative approaches have emerged as promising therapeutic strategies for treating the chronic stage of stroke. In this study, we examined whether systemically administered bone marrow cells (BMCs) could have beneficial effects in a rat model of chronic ischemia. Our transplantation experiments using BMCs obtained from ischemic donor rats showed functional and structural recovery during the chronic stage of stroke. BMC-mediated neural proliferation was prominent in the brains of rats with chronic stroke, and most of the new cells eventually became neurons instead of astrocytes. BMC-mediated enhanced neural proliferation coincided with a significant reduction (∼50%) in the number of activated microglia, which is consistent with previous reports of enhanced neural proliferation being linked to microglial inactivation. Strikingly, approximately 57% of the BMCs that infiltrated the chronic ischemic brain were CD25(+) cells, suggesting that these cells may exert the beneficial effects associated with BMC transplantation. Based on the reported anti-inflammatory role of CD25(+) regulatory T-cells in acute experimental stroke, we propose a working model delineating the positive effects of BMC transplantation during the chronic phase of stroke; infiltrating BMCs (mostly CD25(+) cells) reduce activated microglia, which leads to enhanced neural proliferation and enhanced recovery from neuronal damage in this rat model of chronic stroke. This study provides valuable insights into the effect of BMC transplantation in the chronic ischemic brain, which may lead to the development of effective therapy for chronic stroke patients who currently lack satisfactory therapeutic options.

Topics: Animals; Behavior, Animal; Bone Marrow Cells; Bone Marrow Transplantation; Brain; Brain Ischemia; Cell Differentiation; Cell Proliferation; Disease Models, Animal; Immunophenotyping; Interleukin-2 Receptor alpha Subunit; Male; Microglia; Minocycline; Neurons; Rats; Rats, Sprague-Dawley; Recovery of Function; T-Lymphocytes, Regulatory; Transplantation, Homologous

The protective effect of ischemic postconditioning (IPostC) against stroke has been well-established, and the underlying mechanisms are known to involve inhibited-inflammation and free radical production. Nevertheless, how IPostC affects protein expression of iNOS, nitrotyrosine, and COX-2 has not been characterized. In addition, the role of the galectin-9/Tim-3 cell signaling pathway--a novel inflammatory pathway--in IPostC has not been studied. We examined whether iNOS, nitrotyrosine, and COX-2, as well as galectin-9/Tim-3 are involved in the protective effects of IpostC in a rat focal ischemia model. Western blot and confocal immunofluoresent staining results indicate that IPostC significantly inhibited Tim-3 expression, and that galectin-9 expression was also inhibited. In addition, IPostC attenuated production of iNOS and nitrotyrosine, but not COX-2, suggesting that IPostC has distinct effects on these inflammatory factors. Furthermore, the inflammation inhibitor minocycline blocked Tim-3 and iNOS expression induced by stroke. Taken together, we show that the galectin-9/Tim-3 cell signaling pathway is involved in inflammation induced by stroke, and IPostC may reduce infarction by attenuating this novel pathway as well as the inflammatory factors iNOS and nitrotyrosine, but not COX-2.

Topics: Animals; Anti-Inflammatory Agents; Brain Ischemia; Cyclooxygenase 2; Ischemic Postconditioning; Male; Minocycline; Nitric Oxide Synthase Type II; Rats; Rats, Sprague-Dawley; Receptors, Cell Surface; Signal Transduction; Tyrosine

To investigate the effects of minocycline in promoting the survival of pheochromocytoma (PCI2) cells exposed to oxygen glucose deprivation (OGD) and explore the underlying mechanisms.. An in vitro cell model of cerebral ischemia was established by OGD for 6 h in PCI2 cells with pretreatment with minocycline or an ERK1/2 inhibitor. At 24 h after OGD injury, the cells were evaluated for cell viability by MTT assay and expressions of heme oxygenase-I (HO-I) and phospholylated extracellular signal-regulated protein kinase 1/2 (ERK1/2) by Western blotting.. The cell viability decreased dramatically following OGD. Pretreatment with minocycline (O.I-IO JJ.mol/L) induced a significant increase in the cell viability after OGD and caused up-regulation of HO-I protein and enhanced ERK1/2 phospholylation, and the effects were especially obvious with 1 JJ.mol/L minocycline and were abolished by inhibition of ERK1/2 activity with UOI26 (IO JJ.mol/L).. Minocycline can protect PCI2 cells against OGD-induced toxicity by up-regulating HO-I protein expression through ERKl/2 signaling pathways.

Topics: Animals; Brain Ischemia; Cell Hypoxia; Cell Survival; Glucose; Heme Oxygenase (Decyclizing); MAP Kinase Signaling System; Minocycline; Oxygen; PC12 Cells; Rats; Up-Regulation

It has been suggested that propofol can modulate microglial activity and hence may have potential roles against neuroinflammation following brain ischemic insult. However, whether and how propofol can inhibit post-cardiac arrest brain injury via inhibition of microglia activation remains unclear. A rat model of asphyxia cardiac arrest (CA) was created followed by cardiopulmonary resuscitation. CA induced marked microglial activation in the hippocampal CA1 region, revealed by increased OX42 and P2 class of purinoceptor 7 (P2X7R) expression, as well as p38 MAPK phosphorylation. Morris water maze showed that learning and memory deficits following CA could be inhibited or alleviated by pre-treatment with the microglial inhibitor minocycline or propofol. Microglial activation was significantly suppressed likely via the P2X7R/p-p38 pathway by propofol. Moreover, hippocampal neuronal injuries after CA were remarkably attenuated by propofol. In vitro experiment showed that propofol pre-treatment inhibited ATP-induced microglial activation and release of tumor necrosis factor-α and interleukin-1β. In addition, propofol protected neurons from injury when co-culturing with ATP-treated microglia. Our data suggest that propofol pre-treatment inhibits CA-induced microglial activation and neuronal injury in the hippocampus and ultimately improves cognitive function. We proposed a possible mechanism of propofol-mediated brain protection after cardiac arrest (CA). CA induces P2X7R upregulation and p38 phosphorylation in microglia, which induces release of TNF-α and IL-1β and consequent neuronal injury. Propofol could inhibit microglial activation and alleviate neuronal damage. Our results suggest propofol-induced anti-inflammatory treatment as a plausible strategy for therapeutic intervention in post-CA brain injury.

Topics: Adenosine Triphosphate; Animals; Asphyxia; Brain Ischemia; CA1 Region, Hippocampal; Cardiopulmonary Resuscitation; CD11b Antigen; Cells, Cultured; Heart Arrest; Interleukin-1beta; Male; MAP Kinase Signaling System; Maze Learning; Mice; Microglia; Minocycline; Models, Animal; Nerve Tissue Proteins; Neuroprotective Agents; p38 Mitogen-Activated Protein Kinases; Phosphorylation; Propofol; Protein Processing, Post-Translational; Random Allocation; Rats; Rats, Sprague-Dawley; Receptors, Purinergic P2X7; Tumor Necrosis Factor-alpha

To observe the effects of minocycline on morphology and the expression of synaptophysin in cortical tissues of rats after cerebral ischemia reperfusion injury.. 36 male SD rats were randomly divided into 3 groups: sham group, model group [ischemia reperfusion (I/R)] and minocycline (Min) group (treated with minocycline for 14 d, 3 mg/kg, 2 times/d ). Middle cerebral artery occlusion (MCAO) was used to established as focal cerebral I/R model. At 14 d after I/R. Neurological functional recovery was evaluated using the staircase test, the cell morphology in cortex was evaluated by HE staining, the neurite growth was observed by immunostaining with anti-microtubule-associated protein-2 (MAP-2) antibody, the expression of synaptophysin in pei-infarct region was tested by Western blot.. In the sham group, the rats did not show any neurological deficits. The neurons in the cortex were arranged in neat rows and the morphology were normal, the MAP-2 positive neurons showed longer neuronal processes than the model group. Compared to the model group, minocycline significantly improved forelimb motor function, increased the expression of synaptophysin and the number of MAP-2-positive cells in peri-infarct region (P < 0.05).. Minocycline could improve the neurite regrowth and the expression of synaptophysin of neuron in ischemic cortex, promote neurological functional recovery of rats after MCAO, which is related to regulate the neuronal plasticity.

Topics: Animals; Brain Ischemia; Male; Minocycline; Neuronal Plasticity; Neurons; Rats; Rats, Sprague-Dawley; Recovery of Function; Reperfusion Injury; Synaptophysin

Ethanol is an important risk factor for the occurrence of cerebral ischemia contributing to poor prognosis and inefficacy of drug treatments for stroke-related symptoms. Females have a higher lifetime risk for stroke than males. Moreover, female gender has been associated with increased ethanol consumption during adolescence. In the present study, we investigated whether chronic ethanol exposure during adolescence may potentiate the motor impairments and cortical damage induced by focal ischemia in female rats. We also addressed whether these effects can be mitigated by minocycline, which has been shown to be neuroprotective against different insults in the CNS. Female rats were treated with distilled water or ethanol (6.5 g/kg/day, 22.5% w/v) for 55 days by gavage. Focal ischemia was induced by microinjections of endothelin-1 (ET-1) into the motor cortex. Animals of both groups were treated daily with minocycline (25-50 mg/kg, i.p.) or sterile saline (i.p.) for 5 days, and motor function was assessed using open field, inclined plane and rotarod tests. Chronic ethanol exposure exacerbated locomotor activity and motor coordination impairments induced by focal ischemia in rats. Moreover, histological analysis revealed that microinjections of ET-1 induced pyramidal neuron loss and microglial activation in the motor cortex. Minocycline reversed the observed motor impairments, microglial activation and pyramidal neuron loss in the motor cortex of ischemic rats even in those exposed to ethanol. These results suggest that minocycline induces neuroprotection and functional recovery in ischemic female rats intoxicated with ethanol during adolescence. Furthermore, the mechanism underlying this protective effect may be related to the modulation of neuroinflammation.

Topics: Alcohol-Related Disorders; Animals; Brain Ischemia; Central Nervous System Depressants; Disease Models, Animal; Endothelin-1; Ethanol; Female; Microglia; Minocycline; Motor Activity; Motor Cortex; Movement Disorders; Neurons; Neuroprotective Agents; Pyramidal Cells; Rats, Wistar; Recovery of Function

The aim of this study was to assess the effects of minocycline on cerebral ischemia-reperfusion (I/R) injury in rats. The study was carried out on 24 male Wistar albino rats, weighing 200-250 g, which were divided into three groups: (i) control (n = 8), (ii) I/R (n = 8) and (iii) I/R + minocycline (n = 8). Minocycline was administrated at a dose of 90 mg/kg p.o. to the I/R group 48, 24 and 1 h before ischemia. Following bilateral exposure of the common carotid arteries by anterior cervical dissection and separation of the vagus nerve, I/R injury was performed by occlusion. Following reperfusion, malondialdehyde (MDA), superoxide dismutase, glutathione peroxidase and catalase levels in the blood and brain tissue, and creatine kinase (CK), CK-BB, lactate dehydrogenase (LDH), neuron-specific enolase (NSE) and protein S100β levels in the blood were measured and the histopathological changes were monitored. Regarding histopathological evaluation, symptoms of degeneration were significantly improved in the I/R + minocycline group compared to the I/R-only group. Statistical analysis of the biochemical parameters revealed significant differences in MDA (p < 0.001), nitric oxide (p < 0.05), CK (p < 0.05) and CK-MB (p < 0.05) levels between the I/R + minocycline group and the I/R group. According to the literature, the effect of minocycline is firstly assessed by LDH, CK-MB, NSE and S-100β analysis in addition to antioxidant status and histopathological analysis.

Topics: Animals; Brain; Brain Ischemia; Male; Minocycline; Neuroprotective Agents; Rats; Rats, Wistar; Reperfusion Injury; Time Factors

We explored whether the modulation of microglia activation with minocycline is beneficial to the therapeutic actions of bone marrow mononuclear cells (BMMCs) transplanted after experimental stroke. Male Wistar adult rats were divided in four experimental groups: ischemic control saline treated (G1, N = 6), ischemic minocycline treated (G2, N = 5), ischemic BMMC treated (G3, N = 5), and ischemic minocycline/BMMC treated (G4, N = 6). There was a significant reduction in the number of ED1+ cells in G3 animals (51.31 ± 2.41, P < 0.05), but this effect was more prominent following concomitant treatment with minocycline (G4 = 29.78 ± 1.56). There was conspicuous neuronal preservation in the brains of G4 animals (87.97 ± 4.27) compared with control group (G1 = 47.61 ± 2.25, P < 0.05). The behavioral tests showed better functional recovery in animals of G2, G3, and G4, compared with G1 and baseline (P < 0.05). The results suggest that a proper modulation of microglia activity may contribute to a more permissive ischemic environment contributing to increased neuroprotection and functional recovery following striatal ischemia.

Topics: Animals; Bone Marrow Cells; Bone Marrow Transplantation; Brain Ischemia; Cells, Cultured; Endothelin-1; Macrophage Activation; Macrophages; Male; Microglia; Minocycline; Neuroprotective Agents; Rats; Rats, Wistar; Recovery of Function; Stroke

Minocycline has been recently implicated in protection against focal cerebral ischemia reperfusion (I/R), but the protective effects on neurobehavioral abnormalities remains contradictory. In the present study, we investigate whether minocycline improves axonal regeneration and neurological function recovery by inhibiting the expression of the repulsive guidance molecular A (RGMa) after focal cerebral ischemia reperfusion. Male Sprague-Dawley (SD) rats were subjected to occlusion of the right middle cerebral artery (MCAO) for 2 h and 3 mg kg⁻¹ minocycline was injected intravenously immediately after reperfusion twice a day for 14 days. The staircase test and modified neurological severity score (mNSS) were performed to evaluate functional outcome and blood-brain barrier (BBB) permeability was assessed by Evan's blue dye extravasation (EB) at the expected time point. The expression of RGMa in ischemic cortex was measured by immunohistochemical staining and Western blot 2 weeks after MCAO. Neurofilament protein 200 (NF-200) immunohistochemical staining was used to assess axonal damage. Treatment with minocycline at a dose of 3 mg kg⁻¹ via the caudal vein significantly reduced the extravasation of EB, elevated mNSS and improved forelimb motor function as assessed by the staircase test when compared to the I/R group (P < 0.05). Moreover, axonal regrowth was enhanced in the minocycline treatment group when compared to the I/R group (P < 0.05). In addition, minocycline significantly reduced the expression of RGMa protein 2 weeks after MCAO as assessed by both immunostaining and Western blot. Our studies suggest that early minocycline treatment promotes neurological functional recovery and axonal regeneration in rats after MCAO, which might be mediated by down-regulating RGMa expression.

Topics: Animals; Axons; Brain Ischemia; Cerebral Cortex; GPI-Linked Proteins; Male; Membrane Proteins; Middle Cerebral Artery; Minocycline; Nerve Regeneration; Nerve Tissue Proteins; Neurofilament Proteins; Neuroprotective Agents; Rats; Rats, Sprague-Dawley; Recovery of Function; Reperfusion Injury

Normobaric hyperoxia (NBO), which maintains penumbral oxygenation, reduces brain injury during cerebral ischemia, and minocycline, a tetracycline derivative, reduces reperfusion injury, including inflammation, apoptosis and matrix metalloproteinases (MMPs) activation. Since they have different mechanisms of action, we hypothesized that combining them would provide greater neuroprotection. To test the hypothesis, we evaluated the neuroprotective effects of the combination of NBO with minocycline. Male Sprague-Dawley rats were exposed to NBO (95% O(2)) or normoxia (21% O(2)) during 90-min filament occlusion of the middle cerebral artery, followed by 48 h of reperfusion. Minocycline (3 mg/kg) or vehicle was intravenously administered to rats 15 min after reperfusion onset. Treatment with NBO and minocycline alone resulted in 36% and 30% reductions in infarction volume, respectively. When the two treatments were combined, there was a 68% reduction in infarction volume. The combination therapy also significantly reduced hemispheric swelling, which was absent with monotherapy. In agreement with its greater neuro- and vasoprotection, the combination therapy showed greater inhibitory effects on MMP-2/9 induction, occludin degradation, caspase-3 and -9 activation and apoptosis inducing factor (AIF) induction in ischemic brain tissue. Our results show that NBO plus minocycline effectively reduces brain injury in transient focal cerebral ischemia with protection due to inhibition on MMP-2/9-mediated occludin degradation and attenuation of caspase-dependent and independent apoptotic pathways.

Topics: Animals; Anti-Bacterial Agents; Atmospheric Pressure; Brain Ischemia; Disease Models, Animal; Hyperoxia; Infarction, Middle Cerebral Artery; Male; Minocycline; Oxygen Inhalation Therapy; Rats; Rats, Sprague-Dawley

We aimed to investigate the effects of bone marrow derived mesenchymal stem cells (MSCs), minocycline, and these two therapies combined on functional and histological improvement in cerebral ischemic injury created rats.. Twenty-eight Sprague Dawley female rats, weighing 250-300 g, were included in the study. Two male rats with similar properties were sacrificed for bone marrow derived MSC production. Group 1 was established as the control group. Group 2 was the group of only minocycline administered rats. Group 3 was the one of only MSCs administered rats. Group 4 was composed of the rats given the combination of MSCs and minocycline. Hematoxylin and eosin staining was done to assess the degeneration of the cells. Immunohistochemical staining was performed to evaluate the regeneration. Motor functions were examined by using Bederson's score.. Cell degeneration was the least in group 4. The cells stained with GFAP were observed mostly in group 4. The cells stained with Neu N in group 1 were statistically lower than in other groups. When the groups were ordered in terms of functional improvement at the end of the second week, group 4 had the most and group 1 had the least.. Bone marrow derived MSCs can lead to more histological and functional improvement when administered with minocycline, which is a neuroprotective agent as early as 24 h following the ischemic injury in a rat model. Minocycline therapy alone can be as effective as bone marrow derived MSCs therapy alone in ischemic cerebral rat model.

Topics: Animals; Anti-Bacterial Agents; Biomarkers; Bone Marrow Cells; Brain Injuries; Brain Ischemia; Combined Modality Therapy; Female; Glial Fibrillary Acidic Protein; Male; Mesenchymal Stem Cell Transplantation; Mesenchymal Stem Cells; Minocycline; Motor Activity; Neuroprotective Agents; Rats; Rats, Sprague-Dawley; Treatment Outcome

Transplantation of neural stem cells (NSCs) offers a novel therapeutic strategy for stroke; however, massive grafted cell death following transplantation, possibly due to a hostile host brain environment, lessens the effectiveness of this approach. Here, we have investigated whether reprogramming NSCs with minocycline, a broadly used antibiotic also known to possess cytoprotective properties, enhances survival of grafted cells and promotes neuroprotection in ischemic stroke. NSCs harvested from the subventricular zone of fetal rats were preconditioned with minocycline in vitro and transplanted into rat brains 6 h after transient middle cerebral artery occlusion. Histological and behavioral tests were examined from days 0-28 after stroke. For in vitro experiments, NSCs were subjected to oxygen-glucose deprivation and reoxygenation. Cell viability and antioxidant gene expression were analyzed. Minocycline preconditioning protected the grafted NSCs from ischemic reperfusion injury via upregulation of Nrf2 and Nrf2-regulated antioxidant genes. Additionally, preconditioning with minocycline induced the NSCs to release paracrine factors, including brain-derived neurotrophic factor, nerve growth factor, glial cell-derived neurotrophic factor, and vascular endothelial growth factor. Moreover, transplantation of the minocycline-preconditioned NSCs significantly attenuated infarct size and improved neurological performance, compared with non-preconditioned NSCs. Minocycline-induced neuroprotection was abolished by transfecting the NSCs with Nrf2-small interfering RNA before transplantation. Thus, preconditioning with minocycline, which reprograms NSCs to tolerate oxidative stress after ischemic reperfusion injury and express higher levels of paracrine factors through Nrf2 up-regulation, is a simple and safe approach to enhance the effectiveness of transplantation therapy in ischemic stroke.

Topics: Animals; Brain Ischemia; Cell Death; Cells, Cultured; Ischemic Preconditioning; Male; Minocycline; Neural Stem Cells; Neuroprotective Agents; Rats; Rats, Sprague-Dawley; Rats, Transgenic; Stem Cell Transplantation; Stroke

Many new candidate pharmaceuticals designed to improve recovery after stroke have been proposed recently, but there are still too few molecular imaging methods capable to assess their efficacy. A hallmark of the inflammatory reaction that follows focal cerebral ischemia is overexpression of the mitochondrial peripheral benzodiazepine receptor/18 kDa translocator protein (PBR/TSPO) in the monocytic lineage and astrocytes. This overexpression can be imaged with positron emission tomography (PET) using PBR/TSPO-selective radioligands such as [(18)F]DPA-714.. Here, we tested whether PET with [(18)F]DPA-714 would evidence the effect of minocycline, a broad spectrum antibiotic presently tested as neuroprotective agent after stroke, on the inflammatory reaction induced in an experimental model of stroke.. Ten rats were subjected to a 2-h transient middle cerebral artery occlusion with reperfusion. Minocycline or saline was intravenously administrated 1 h after reperfusion and daily during the following 6 days. PET studies were performed using [(18)F]DPA-714 at 7 days after cerebral ischemia.. In vivo PET imaging showed a significant decrease in [(18)F]DPA-714 uptake at 7 days after cerebral ischemia in rats treated with minocycline with respect to saline-treated animals. Minocycline treatment had no effect on the size of the infarcted area.. Minocycline administered daily during 7 days after ischemia decreases [(18)F]DPA-714 binding, suggesting that the drug exerts an anti-inflammatory activity. [(18)F]DPA-714 PET is a useful biomarker to study novel anti-inflammatory strategies in experimental cerebral ischemia.

Topics: Animals; Brain Ischemia; Disease Models, Animal; Fluorine Radioisotopes; Male; Minocycline; Pyrazoles; Pyrimidines; Radionuclide Imaging; Radiopharmaceuticals; Rats; Rats, Sprague-Dawley; Receptors, GABA-A

The study investigated a possible neuroprotective potency of minocycline in an experimental asphyxial cardiac arrest (ACA) rat model. Clinically important survival times were evaluated thus broadening common experimental approaches.. Adult rats were subjected to 5 min of ACA followed by resuscitation. There were two main treatment groups: ACA and sham operated. Relating to minocycline treatment each group consisted of three sub-groups: pre-, post-, and sans-mino, with three different survival times: 4, 7, and 21 days. Neurodegeneration and microgliosis were monitored by immunohistochemistry. Alterations of microglia-associated gene expression were analyzed by quantitative RT-PCR.. ACA induced massive nerve cell loss and activation of microglia/macrophages in hippocampal CA1 cell layer intensifying with survival time. After 7 days, minocycline significantly decreased both, neuronal degeneration and microglia response in dependence on the application pattern; application post ACA was most effective. After 21 days, neuroprotective effects of minocycline were lost. ACA significantly induced expression of the microglia-associated factors Ccl2, CD45, Mac-1, F4-80, and Tnfa. Independent on survival time, minocycline affected these parameters not significantly. Expression of iNOS was unaffected by both, ACA and minocycline.. In adult rat hippocampus microglia was significantly activated by ACA. Minocycline positive affected neuronal survival and microglial response temporary, even when applied up to 18 h after ACA, thus defining a therapeutically-relevant time window. As ACA-induced neuronal cell death involves acute and delayed events, longer minocycline intervention targeting also secondary injury cascades should manifest neuroprotective potency, a question to be answered by further experiments.

Topics: Animals; Asphyxia; Biomarkers; Brain; Brain Ischemia; Cell Death; Disease Models, Animal; Heart Arrest; Hippocampus; Immunohistochemistry; Microglia; Minocycline; Rats; Resuscitation; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Time Factors

Nicotinamide adenine dinucleotide phosphate oxidase (NOX) is widely expressed in brain tissue including neurons, glia, and endothelia in neurovascular units. It is a major source of oxidants in the post-ischemic brain and significantly contributes to ischemic brain damage. Inflammation occurs after brain ischemia and is known to be associated with post-ischemic oxidative stress. Post-ischemic inflammation also causes progressive brain injury. In this study we investigated the role of NOX2 in post-ischemic cerebral inflammation using a transient middle cerebral artery occlusion model in mice. We demonstrate that mice with NOX2 subunit gp91(phox) knockout (gp91 KO) showed 35-44% less brain infarction at 1 and 3 days of reperfusion compared with wild-type (WT) mice. Minocycline further reduced brain damage in the gp91 KO mice at 3 days of reperfusion. The gp91 KO mice exhibited less severe post-ischemic inflammation in the brain, as evidenced by reduced microglial activation and decreased upregulation of inflammation mediators, including interleukin-1β (IL-1β), tumor necrosis factor-α, inducible nitric oxide synthases, CC-chemokine ligand 2, and CC-chemokine ligand 3. Finally, we demonstrated that an intraventricular injection of IL-1β enhanced ischemia- and reperfusion-mediated brain damage in the WT mice (double the infarction volume), whereas, it failed to aggravate brain infarction in the gp91 KO mice. Taken together, these results demonstrate the involvement of NOX2 in post-ischemic neuroinflammation and that NOX2 inhibition provides neuroprotection against inflammatory cytokine-mediated brain damage.

Topics: Analysis of Variance; Animals; Blotting, Western; Brain; Brain Ischemia; Cytokines; Encephalitis; Immunohistochemistry; Mice; Mice, Knockout; Minocycline; NADPH Oxidases; Oxidative Stress; Reverse Transcriptase Polymerase Chain Reaction

Topics: Anti-Bacterial Agents; Brain Ischemia; Female; Humans; Male; Matrix Metalloproteinase 9; Minocycline; Stroke

A neuroprotective strategy through a combination therapy is always being superior to any other singular therapeutic interventions, as these acts through a multifauceted approach within the brain during cerebral ischemia. Therefore, the development of a potential new combination of drug is necessitated which can bring about desirable improved neuroprotection targeting different pathways against ischemic stroke. Numerous past studies have enumerated the neuroprotective roles of minocycline and magnesium administered in single against cerebral ischemia in animal model hence we hypothesized that by using magnesium with minocycline in combination would provide additive neuroprotection than either of the agents used alone. In this article, we discuss our hypothesis regarding the possibility of minocycline and magnesium as a potent combination which may have a positive therapeutic role in treatment of cerebral ischemia through its anti-inflammatory, anti-apoptotic and anti-oxidative characteristics with magnesium contributing as a regulator of increased calcium influx.

Topics: Animals; Brain Ischemia; Drug Therapy, Combination; Humans; Magnesium; Minocycline; Models, Biological; Neuroprotective Agents; Rats

We previously reported that minocycline attenuates acute brain injury and inflammation after focal cerebral ischemia, and this is partly mediated by inhibition of 5-lipoxygenase (5-LOX) expression. Here, we determined the protective effect of minocycline on chronic ischemic brain injury and its relation with the inhibition of 5-LOX expression after focal cerebral ischemia.. Focal cerebral ischemia was induced by 90 min of middle cerebral artery occlusion followed by reperfusion for 36 days. Minocycline (45 mg/kg) was administered intraperitoneally 2h and 12h after ischemia and then every 12h for 5 days. Sensorimotor function was evaluated 1-28 days after ischemia and cognitive function was determined 30-35 days after ischemia. Thereafter, infarct volume, neuron density, astrogliosis, and 5-LOX expression in the brain were determined.. Minocycline accelerated the recovery of sensorimotor and cognitive functions, attenuated the loss of neuron density, and inhibited astrogliosis in the boundary zone around the ischemic core, but did not affect infarct volume. Minocycline significantly inhibited the increased 5-LOX expression in the proliferated astrocytes in the boundary zone, and in the macrophages/microglia in the ischemic core.. Minocycline accelerates functional recovery in the chronic phase of focal cerebral ischemia, which may be partly associated with the reduction of 5-LOX expression.

Topics: Agnosia; Animals; Astrocytes; Brain; Brain Ischemia; Cell Count; Cell Proliferation; Chronic Disease; Immunohistochemistry; Injections, Intraperitoneal; Lipoxygenase Inhibitors; Macrophages; Male; Maze Learning; Microglia; Minocycline; Neurons; Neuroprotective Agents; Rats; Rats, Sprague-Dawley; Recovery of Function

Minocycline is a promising anti-inflammatory and protease inhibitor that is effective in multiple preclinical stroke models. We conducted an early phase trial of intravenous minocycline in acute ischemic stroke.. Following an open-label, dose-escalation design, minocycline was administered intravenously within 6 hours of stroke symptom onset in preset dose tiers of 3, 4.5, 6, or 10 mg/kg daily over 72 hours. Minocycline concentrations for pharmacokinetic analysis were measured in a subset of patients. Subjects were followed for 90 days.. Sixty patients were enrolled, 41 at the highest dose tier of 10 mg/kg. Overall age (65±13.7 years), race (83% white), and sex (47% female) were consistent across the doses. The mean baseline National Institutes of Health Stroke Scale score was 8.5±5.8 and 60% received tissue plasminogen activator. Minocycline infusion was well tolerated with only 1 dose limiting toxicity at the 10-mg/kg dose. No severe hemorrhages occurred in tissue plasminogen activator-treated patients. Pharmacokinetic analysis (n=22) revealed a half-life of approximately 24 hours and linearity of parameters over doses.. Minocycline is safe and well tolerated up to doses of 10 mg/kg intravenously alone and in combination with tissue plasminogen activator. The half-life of minocycline is approximately 24 hours, allowing every 24-hour dosing. Minocycline may be an ideal agent to use with tissue plasminogen activator.

Topics: Aged; Aged, 80 and over; Anti-Inflammatory Agents; Brain Ischemia; Chromatography, High Pressure Liquid; Drug Administration Schedule; Drug Therapy, Combination; Female; Fibrinolytic Agents; Half-Life; Humans; Male; Middle Aged; Minocycline; Severity of Illness Index; Stroke; Time Factors; Tissue Plasminogen Activator; Treatment Outcome

Recently, the beneficial role of minocycline on endogenous neurogenesis after cerebral ischemia has been contradicted by many reports. We examined whether minocycline influences post-ischemic neurogenesis in the subventricular zone. Adult male Sprague-Dawley rats were subjected to focal cerebral ischemia for 2 h, and divided into a minocycline-treated (90 mg/Kg on reperfusion and 45 mg/Kg daily for maintenance) and a saline-treated group. Bromodeoxyuridine was injected to determine levels of cell proliferation. Inflammation was assessed by counting polymorphonuclear cell and activated microglia and by measuring myeloperoxidase activity. Endogenous neurogenesis was quantified by immunohistochemical staining and functional outcome was measured by infarct size and behavioral tests. Minocycline treatment decreased inflammation on 1st and 4th days after ischemia. BrdU-positive cells on 7th day (saline vs. minocycline: 602.80+/-146.96 vs. 399.40+/-109.69) and the number of double labeling cells of BrdU/NeuN on 7th day (13.00+/-4.36 vs. 6.40+/-2.07) and BrdU/DCx on 4th day (17.00+/-5.00 vs. 7.50+/-1.91) were significantly decreased in minocycline-treated rats. Infarct size and behavioral tests were not different. Our results indicate that minocycline may reduce immediate post-ischemic neurogenesis despite adequately suppressed inflammation.

Topics: Animals; Anti-Inflammatory Agents, Non-Steroidal; Brain; Brain Ischemia; Bromodeoxyuridine; Cell Count; Doublecortin Domain Proteins; Doublecortin Protein; Enzyme Activation; Ischemic Attack, Transient; Male; Microglia; Microtubule-Associated Proteins; Minocycline; Motor Activity; Neurogenesis; Neuroimmunomodulation; Neurons; Neuropeptides; Neutrophils; Peroxidase; Rats; Rats, Sprague-Dawley; Treatment Outcome

Minocycline is neuroprotective in clinical and experimental stroke studies, due in part to its ability to inhibit poly (ADP-ribose) polymerase. Previous preclinical data have shown that interference with poly (ADP-ribose) polymerase signaling leads to sex-specific neuroprotection, reducing stroke injury only in males. In this study, we show that minocycline is ineffective at reducing ischemic damage in females after middle cerebral artery occlusion, likely due to effects on poly (ADP-ribose) polymerase signaling. Clinical trials must consider possible sex differences in the response to neuroprotective agents, if we hope to translate promising therapies to stroke patients of both sexes.

Topics: Animals; Brain Ischemia; Drug Evaluation, Preclinical; Female; Infarction, Middle Cerebral Artery; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Minocycline; Neuroprotective Agents; Poly(ADP-ribose) Polymerase Inhibitors; Sex Factors; Stroke; Treatment Failure

New treatment strategies for acute ischemic stroke must be evaluated in the context of effective reperfusion. Minocycline is a neuroprotective agent that inhibits proteolytic enzymes and therefore could potentially both inactivate the clot lysis effect and decrease the damaging effects of tissue-type plasminogen activator (t-PA). This study aimed to determine the effect of minocycline on t-PA clot lysis and t-PA-induced hemorrhage formation after ischemia.. Fibrinolytic and amidolytic activities of t-PA were investigated in vitro over a range of clinically relevant minocycline concentrations. A suture occlusion model of 3-hour temporary cerebral ischemia in rats treated with t-PA and 2 different minocycline regimens was used. Blood-brain barrier basal lamina components, matrix metalloproteinases (MMPs), hemorrhage formation, infarct size, edema, and behavior outcome were assessed.. Minocycline did not affect t-PA fibrinolysis. However, minocycline treatment at 3 mg/kg IV decreased total protein expression of both MMP-2 (P=0.0034) and MMP-9 (P=0.001 for 92 kDa and P=0.0084 for 87 kDa). It also decreased the incidence of hemorrhage (P=0.019), improved neurologic outcome (P=0.0001 for Bederson score and P=0.0391 for paw grasp test), and appeared to decrease mortality. MMP inhibition was associated with decreased degradation in collagen IV and laminin-alpha1 (P=0.0001).. Combination treatment with minocycline is beneficial in t-PA-treated animals and does not compromise clot lysis. These results also suggest that neurovascular protection by minocycline after stroke may involve direct protection of the blood-brain barrier during thrombolysis with t-PA.

Topics: Animals; Anti-Bacterial Agents; Blood-Brain Barrier; Brain Ischemia; Cerebral Hemorrhage; Drug Evaluation, Preclinical; Fibrinolysis; Gene Expression Regulation; Male; Matrix Metalloproteinase 2; Matrix Metalloproteinase 9; Minocycline; Neuroprotective Agents; Rats; Rats, Wistar; Stroke; Tissue Plasminogen Activator

Degenerative changes in areas remote from the primary lesion site have been linked to the clinical outcome of focal brain damage, and inflammatory mechanisms have been considered to play a key role in the pathogenesis of these remote cell death phenomena. Minocycline is a tetracycline derivative, therapeutically effective in various experimental models of central nervous system (CNS) injuries that include inflammatory and apoptotic mechanisms, although recent findings have yielded mixed results. In this study, we investigated the effectiveness of minocycline treatment in reducing remote cell death. Glial activation and neuronal loss in precerebellar stations following cerebellar lesion were investigated using immunohistochemistry and Western blot techniques. Our results show that minocycline was effective in reducing microglial activations in axotomized precerebellar nuclei, but failed to mitigate either astrocytic response or neuronal loss. This finding supports the role of minocycline in modulating inflammatory response after CNS lesion and suggests its ineffectiveness in influencing degenerative phenomena in areas remote from the primary lesion site.

Topics: Animals; Brain Ischemia; Cerebellum; Inflammation; Microglia; Minocycline; Nerve Degeneration; Olivary Nucleus; Rats

Thrombolysis with tPA is the only FDA-approved therapy for acute ischemic stroke. But its widespread application remains limited by narrow treatment time windows and the related risks of cerebral hemorrhage. In this study, we ask whether minocycline can prevent tPA-associated cerebral hemorrhage and extend the reperfusion window in an experimental stroke model in rats.. Spontaneously hypertensive rats were subjected to embolic focal ischemia using homologous clots and treated with: saline at 1 hour; early tPA at 1 hour, delayed tPA at 6 hours; minocycline at 4 hours; combined minocycline at 4 hours plus tPA at 6 hours. Infarct volumes and hemorrhagic transformation were quantified at 24 hours. Gelatin zymography was used to measure blood levels of circulating matrix metalloproteinase-9 (MMP-9).. Early 1-hour thrombolysis restored perfusion and reduced infarction. Late 6-hour tPA did not decrease infarction but instead worsened hemorrhagic conversion. Combining minocycline with delayed 6-hour tPA decreased plasma MMP-9 levels, reduced infarction, and ameliorated brain hemorrhage. Blood levels of MMP-9 were also significantly correlated with volumes of infarction and hemorrhage.. Combination therapy with minocycline may extend tPA treatment time windows in ischemic stroke.

Topics: Animals; Biomarkers; Blood-Brain Barrier; Brain Ischemia; Cerebral Hemorrhage; Cerebral Infarction; Drug Administration Schedule; Drug Evaluation, Preclinical; Fibrinolytic Agents; Intracranial Embolism; Male; Matrix Metalloproteinase 9; Matrix Metalloproteinase Inhibitors; Minocycline; Neuroprotective Agents; Rats; Rats, Inbred SHR; Recombinant Proteins; Reperfusion; Thrombolytic Therapy; Time Factors; Tissue Plasminogen Activator

Reperfusion injury is a complication of recanalization therapies after focal cerebral ischemia. The disruption of the blood-brain barrier (BBB) caused by up-regulated metalloproteinases (MMPs) can lead to edema and hemorrhage. Middle cerebral artery occlusion (MCAO=90 min) and reperfusion (R=24 h vs. 5 days) was induced in male Wistar rats. Rats were randomized in four groups: (1) control (C), (2) twice daily minocycline (30 mg/kg bodyweight) every day (M), (3) hypothermia (33 degrees C) for 4 h starting 60 min after occlusion (H), (4) combination of groups 2 and 3 (MH). Serial MRI was performed regarding infarct evolution and BBB disruption, MMP-2 and MMP-9 were assessed by zymography of serum and ischemic brain tissue, and a functional neuroscore was done at 24 h and 5 days. M and H reduced both infarct sizes, volume and signal intensity of BBB breakdown and improved neuroscore at all points in time to the same extent. This was most likely due to inhibition of MMP-2 and MMP-9. The presence of MMP-9 at 24 h or MMP-2 at 5 days in brain tissue correlated with BBB breakdown whereas serum MMP-2- and -9 showed no relationship with BBB breakdown. The combination MH had a small but not significantly additional effect over the single treatments. Minocycline seems to be as neuroprotective as hypothermia in the acute and subacute phase after cerebral ischemia. One essential mechanism is the inhibition of MMPs. The combination therapy is only slightly superior. The net effect of MMPs inhibition up to 5 days after focal cerebral ischemia is still beneficial.

Topics: Acute Disease; Animals; Anti-Bacterial Agents; Blood-Brain Barrier; Brain Edema; Brain Ischemia; Disease Models, Animal; Disease Progression; Hypothermia, Induced; Infarction, Middle Cerebral Artery; Intracranial Hemorrhages; Magnetic Resonance Imaging; Male; Matrix Metalloproteinase 2; Matrix Metalloproteinase 9; Metalloproteases; Minocycline; Neuroprotective Agents; Rats; Rats, Wistar; Reperfusion Injury; Time Factors

We previously reported that delayed administration of the general cyclin-dependent kinase inhibitor flavopiridol following global ischemia provided transient neuroprotection and improved behavioral performance. However, it failed to provide longer term protection. In the present study, we investigate the ability of delayed flavopiridol in combination with delayed minocycline, another neuroprotectant to provide sustained protection following global ischemia. We report that a delayed combinatorial treatment of flavopiridol and minocycline provides synergistic protection both 2 and 10 weeks following ischemia. However, protected neurons in the hippocampal CA1 are synaptically impaired as assessed by electrophysio logical field potential recordings. This is likely because of the presence of degenerated processes in the CA1 even with combinatorial therapy. This indicates that while we have addressed one important pre-clinical parameter by dramatically improving long-term neuronal survival with delayed combinatorial therapy, the issue of synaptic preservation of protected neurons still exists. These results also highlight the important observation that protection does not always lead to proper function.

Topics: Animals; Anti-Bacterial Agents; Brain Infarction; Brain Ischemia; Dendrites; Disease Models, Animal; Drug Administration Schedule; Drug Synergism; Drug Therapy, Combination; Flavonoids; Hippocampus; Male; Minocycline; Nerve Degeneration; Neural Pathways; Neurons; Neuroprotective Agents; Piperidines; Protein Kinase Inhibitors; Rats; Rats, Wistar; Synaptic Transmission; Time Factors; Treatment Outcome

Minocycline, a semisynthetic tetracycline antibiotic, has been reported to ameliorate brain injury and inhibit microglial activation after focal cerebral ischemia. However, the cerebroprotective mechanism of minocycline remains unclear. In the present study, we investigated that mechanism of minocycline in a murine model of 4-hour middle cerebral artery (MCA) occlusion.. One day after 4-hour MCA occlusion, minocycline was administered intraperitoneally for 14 days. Neurologic scores were measured 1, 7, and 14 days after cerebral ischemia. Motor coordination was evaluated at 14 days by the rota-rod test at 10 rpm. Activated microglia and high-mobility group box1 (HMGB1), a cytokine-like mediator, were also evaluated by immunostaining and Western blotting. In addition, terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling immunostaining was carried out 14 days after cerebral ischemia.. Repeated treatment with minocycline (1, 5, and 10 mg/kg) for 14 days improved neurologic score, motor coordination on the rota-rod test, and survival in a dose-dependent manner. Minocycline decreased the expression of Iba1, a marker of activated microglia, as assessed by both immunostaining and Western blotting. Moreover, minocycline decreased the activation of microglia expressing HMGB1 within the brain and also decreased both brain and plasma HMGB1 levels. Additionally, minocycline significantly decreased the number of terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling-positive cells and prevented ischemic brain atrophy 14 days after cerebral ischemia.. Our results suggest that minocycline inhibits activated microglia expressing HMGB1 and decreases neurologic impairment induced by cerebral ischemia. Minocycline will have a palliative action and open new therapeutic possibilities for treatment of postischemic injury via an HMGB1-inhibiting mechanism.

Topics: Animals; Apoptosis; Atrophy; Brain; Brain Ischemia; Dose-Response Relationship, Drug; Drug Administration Schedule; HMGB1 Protein; In Situ Nick-End Labeling; Infarction, Middle Cerebral Artery; Injections, Intraperitoneal; Male; Mice; Microglia; Minocycline; Nervous System Diseases; Neuroprotective Agents; Psychomotor Performance; Survival Analysis

Evidence suggests that activated microglia are detrimental to the survival of new hippocampal neurons, whereas blocking inflammation has been shown to restore hippocampal neurogenesis after cranial irradiation and seizure. The aim of this current study is to determine the effect of minocycline on neurogenesis and functional recovery after cerebral focal ischemia.. Four days after temporary middle cerebral artery occlusion, minocycline was administered intraperitoneally for 4 weeks. BrdU was given on days 4 to 7 after middle cerebral artery occlusion to track cell proliferation. The number of remaining new neurons and activated microglia were quantified in the dentate gyrus. Infarct volume was measured to assess the treatment effect of minocycline. Motor and cognitive functions were evaluated 6 weeks after middle cerebral artery occlusion.. Minocycline delivered 4 days after middle cerebral artery occlusion for 4 weeks did not result in reduction in infarct size but significantly decreased the number of activated microglia in the dentate gyrus. Minocycline also significantly increased the number of newborn neurons that coexpressing BrdU and NeuN without significantly affecting progenitor cell proliferation in the dentate gyrus. Lastly, minocycline significantly improved motor coordination on the rotor rod, reduced the preferential use of the unaffected limb during exploration, reduced the frequency of footfalls in the affected limb when traversing on a horizontal ladder, and improved spatial learning and memory in the water maze test.. Minocycline reduces functional impairment caused by cerebral focal ischemia. The improved function is associated with enhanced neurogenesis and reduced microglia activation in the dentate gyrus and possibly improved neural environment after chronic treatment with minocycline.

Topics: Animals; Anti-Bacterial Agents; Brain Ischemia; Cell Differentiation; Cell Proliferation; Cerebral Infarction; Disease Models, Animal; Drug Administration Schedule; Infarction, Middle Cerebral Artery; Male; Memory; Minocycline; Motor Activity; Nerve Degeneration; Nerve Regeneration; Neurons; Neuroprotective Agents; Nootropic Agents; Rats; Rats, Sprague-Dawley; Stem Cells

Considering that several pathways leading to cell death are activated in cerebral ischemia, we tested in mouse models of transient and permanent ischemia a drug cocktail aiming at distinct pharmacological targets during the evolution of ischemic injury. It consists of minocycline--an antibiotic with anti-inflammatory properties, riluzole--a glutamate antagonist, and nimodipine--a blocker of voltage-gated calcium channels. Administered 2 h after transient or permanent MCAO, it significantly decreased the size of infarction, by approximately 65% after transient and approximately 35% after permanent ischemia and markedly improve clinical recovery of mice. In both experimental models a three-drug cocktail achieved significantly more efficient neuroprotection than any of the components tested alone. However, some interesting observation emerged from the single-drug studies. Treatment with minocycline alone was efficient in both experimental models while treatment with glutamate antagonist riluzole conferred neuroprotection only after transient MCAO. Immunohistochemical analysis following three-drug treatment revealed reduced microglia/macrophages and caspase-3 activation as well as preserved GFAP immunoreactivity following transient ischemia. No detectable differences in the levels of Mac-2, GFAP and caspase-3 immunoreactivities were observed 72 h after permanent MCAO. These marked differences in the brain tissue responses to ischemic injury and to treatments suggest that different pathological mechanisms may be operating in transient and permanent ischemia. However, the three-drug cocktail exerted significant neuroprotection in both experimental models thus demonstrating that simultaneous targeting of several pathophysiological pathways involved in the evolution of ischemic injury may represent a rational therapeutic strategy for stroke.

Topics: Animals; Brain; Brain Ischemia; Caspase 3; Drug Combinations; Enzyme Activation; Glial Fibrillary Acidic Protein; Immunohistochemistry; Ischemic Attack, Transient; Macrophages; Male; Mice; Mice, Inbred C57BL; Microglia; Minocycline; Myocardial Infarction; Neuroprotective Agents; Nimodipine; Riluzole; Time Factors

To determine whether the anti-inflammatory effect of minocycline on postischemic brain injury is mediated by the inhibition of 5-lipoxygenase (5-LOX) expression and enzymatic activation in rats.. Focal cerebral ischemia was induced for 30 min with middle cerebral artery occlusion, followed by reperfusion. The ischemic injuries, endogenous IgG exudation, the accumulation of neutrophils and macrophage/microglia, and 5-LOX mRNA expression were determined 72 h after reperfusion. 5-LOX metabolites (leukotriene B4 and cysteinyl leukotrienes) were measured 3 h after reperfusion.. Minocycline (22.5 and 45 mg/kg, ip, for 3 d) attenuated ischemic injuries, IgG exudation, and the accumulation of neutrophils and macrophage/microglia 72 h after reperfusion. It also inhibited 5-LOX expression 72 h after reperfusion and the production of leukotrienes 3 h after reperfusion.. Minocycline inhibited postischemic brain inflammation, which might be partly mediated by the inhibition of 5-LOX expression and enzymatic activation.

Topics: Animals; Anti-Bacterial Agents; Arachidonate 5-Lipoxygenase; Behavior, Animal; Brain; Brain Ischemia; Encephalitis; Enzyme Activation; Infarction, Middle Cerebral Artery; Lipoxygenase Inhibitors; Male; Minocycline; Rats; Rats, Sprague-Dawley

Many neurological insults are accompanied by a marked acute inflammatory reaction, involving the activation of microglia. Using a model of exogenous application of fluorescence-labeled BV2 microglia in pathophysiologically relevant concentrations onto organotypic hippocampal slice cultures, we investigated the specific effects of microglia on neuronal damage after ischemic injury. Neuronal cell death after oxygen-glucose deprivation (OGD) was determined by propidium iodide incorporation and Nissl staining. Migration and interaction with neurons were analyzed by time resolved 3-D two-photon microscopy. We show that microglia protect against OGD-induced neuronal damage and engage in close physical cell-cell contact with neurons in the damaged brain area. Neuroprotection and migration of microglia were not seen with integrin regulator CD11a-deficient microglia or HL-60 granulocytes. The induction of migration and neuron-microglia interaction deep inside the slice was markedly increased under OGD conditions. Lipopolysaccharide-prestimulated microglia failed to provide neuroprotection after OGD. Pharmacological interference with microglia function resulted in a reduced neuroprotection. Microglia proved to be neuroprotective even when applied up to 4 h after OGD, thus defining a "protective time window." In acute injury such as trauma or stroke, appropriately activated microglia may primarily have a neuroprotective role. Anti-inflammatory treatment within the protective time window of microglia would therefore be counterintuitive.

Topics: Animals; Anisomycin; Anti-Bacterial Agents; Brain Ischemia; CD11a Antigen; Cell Death; Cell Line; Glucose; Granulocytes; Hippocampus; HL-60 Cells; Humans; Hypoxia; Mice; Mice, Transgenic; Microglia; Minocycline; Neurons; Rats; Rats, Wistar

Blood-brain barrier (BBB) disruption after stroke can worsen ischemic injury by increasing edema and causing hemorrhage. We determined the effect of microglia on the BBB and its primary constituents, endothelial cells (ECs) and astrocytes, after ischemia using in vivo and in vitro models.. Primary astrocytes, ECs, or cocultures were prepared with or without added microglia. Primary ECs were more resistant to oxygen-glucose deprivation/reperfusion than astrocytes. ECs plus astrocytes showed intermediate vulnerability. Microglia added to cocultures nearly doubled cell death. This increase was prevented by minocycline and apocynin. In vivo, minocycline reduced infarct volume and neurological deficits and markedly reduced BBB disruption and hemorrhage in mice after experimental stroke.. Inhibition of microglial activation may protect the brain after ischemic stroke by improving BBB viability and integrity. Microglial inhibitors may prove to be an important treatment adjunct to fibrinolysis.

Topics: Acetophenones; Animals; Antioxidants; Astrocytes; Blood-Brain Barrier; Brain; Brain Ischemia; Cell Death; Cells, Cultured; Cerebral Hemorrhage; Cerebral Infarction; Coculture Techniques; Endothelial Cells; Glucose; Hydrogen Peroxide; Hypoxia; Male; Mice; Mice, Inbred C57BL; Microglia; Minocycline; Nervous System Diseases; Superoxides; Tumor Necrosis Factor-alpha

Minocycline, a second-generation tetracycline, has been shown to possess neuroprotective effects in animal models of stroke. Pial vessel disruption in adult Wistar rats leads to a cone-shaped cortical lesion and turns into a fluid-filled cavity surrounded by a GFAP+ glia limitans 21 days after injury. This mimics the clinical situation in lacunar infarcts. Minocycline was given intraperitoneally at a dose of 45 mg/kg 1 and 12 h after lesioning, followed by 22.5 mg/kg twice daily until 6 days after lesioning. Control rats received intraperitoneal injections of equivalent volumes of saline. Cavitation was prevented in five out of six minocycline-treated animals and the glia limitans did not appear as the space was filled with GFAP+ reactive astrocytes. However, the number of activated microglia showed no difference between minocycline-treated and -untreated groups. Minocycline did not reduce the number of infiltrating leukocytes, predominately polymorphonuclear neutrophils (PMNs) determined by myeloperoxidase immunoreactivity, or infiltration of CD3+ lymphocytes. The pial vessel occlusion induced a significant upregulation of IL-1beta expression; however, minocycline treatment did not significantly alter this upregulation of IL-1beta. In this study, we found minocycline facilitated the repopulation of the lesion by reactive astrocytes and therefore prevented cavitation; however, we could not identify the molecular signal.

Topics: Animals; Anti-Bacterial Agents; Astrocytes; Brain Ischemia; Cerebral Arteries; Cerebral Cortex; Cerebral Infarction; Chemotaxis, Leukocyte; Disease Models, Animal; Dose-Response Relationship, Drug; Drug Administration Schedule; Glial Fibrillary Acidic Protein; Gliosis; Interleukin-1; Leukocytes; Male; Microglia; Minocycline; Nerve Degeneration; Neuroprotective Agents; Rats; Rats, Wistar; Treatment Outcome; Up-Regulation

Early reperfusion after an ischemic stroke can cause blood-brain barrier injury with subsequent cerebral edema and devastating brain hemorrhage. These complications of early reperfusion, which result from excess production of reactive oxygen species, significantly limit the benefits of stroke therapies. In this article, we use a novel animal model that facilitates identification of specific components of the reperfusion injury process, including vascular injury and secondary brain damage, and allows assessment of therapeutic interventions.. Knock-out (KO) mice containing 50% manganese-superoxide dismutase activity (SOD2-KO) and transgenic mice overexpressing SOD2 undergo transient focal ischemia and reperfusion followed by assessment of infarct, edema, hemorrhage rates, metalloproteinase activation, and microvascular injury.. SOD2-KO mice demonstrate delayed (>24h) blood-brain barrier breakdown associated with activation of matrix metalloproteinases, inflammation, and high brain hemorrhage rates. These adverse consequences are absent in wild-type littermates and minocycline-treated SOD2-KO animals. Increased hemorrhage rates also are absent in SOD2 overexpressors, which have reduced vascular endothelial cell death. Finally, we show that the tight junction membrane protein, occludin, is an early and specific target in oxidative stress-induced microvascular injury.. This model is ideal for studying ischemia/reperfusion-induced vascular injury and secondary brain hemorrhage and offers a unique opportunity to evaluate antioxidant-based neurovascular protective strategies as potential adjunct treatments to currently approved stroke therapies such as thrombolysis and endovascular clot retrieval.

Topics: Animals; Blood-Brain Barrier; Blotting, Western; Brain; Brain Ischemia; Cerebral Hemorrhage; Disease Models, Animal; Enzyme Inhibitors; In Situ Nick-End Labeling; Matrix Metalloproteinase 9; Mice; Mice, Knockout; Mice, Transgenic; Minocycline; Oxidative Stress; Reperfusion Injury; Superoxide Dismutase

The absence of effective treatments for stroke presents a critical need for novel strategies that can reduce ischemic injury. Neuroinflammation following focal ischemia induces secondary injury in the region surrounding the insult, thus anti-inflammatory agents are potential neuroprotectants. Minocycline is one such agent possessing neuroprotective properties, however many studies examining minocycline after ischemia have used minimal delays between ischemia and treatment, short survival periods, and lack measures of functional outcome. Such studies do not distinguish whether minocycline provides sustained protection or merely delays cell death. This study was designed to address some of these concerns. Male Sprague-Dawley rats were treated with multiple doses of minocycline (45 mg/kg i.p.) or vehicle beginning 2.5 h after endothelin-1-induced focal ischemia. Measures of forelimb asymmetry and skilled reaching (staircase test) were used to determine functional outcome 7, 15 and 28 days after ischemia. Long-term functional assessment indicates that minocycline provides limited benefit in the staircase test, but confers long-term benefit in the forelimb asymmetry test. Subcortical and whole hemisphere infarct volumes were reduced by 41 and 39% respectively in minocycline-treated animals. Further analysis revealed that minocycline attenuated long-term white matter damage adjacent to the striatal injury core, which correlated with sustained functional benefits. This study indicates that delayed minocycline treatment improves long-term functional outcome which is linked to protection of both white and gray matter.

Topics: Analysis of Variance; Animals; Behavior, Animal; Brain Infarction; Brain Ischemia; Disease Models, Animal; Drug Administration Schedule; Endothelin-1; Functional Laterality; Male; Minocycline; Neuroprotective Agents; Psychomotor Performance; Rats; Rats, Sprague-Dawley; Recovery of Function; Severity of Illness Index; Time Factors; Treatment Outcome

Matrix metalloproteinases 2 and 9 (MMP-2 and MMP-9) are increased in the brain after experimental ischemic stroke in rats. These two proteases are involved with the degradation of the basal lamina and loss of stability of the blood brain barrier that occurs after ischemia and that is associated with thrombolytic therapy in ischemic stroke. Minocycline is a lipophilic tetracycline and is neuroprotective in several models of brain injury. Minocycline inhibits inflammation, apoptosis and extracellular matrix degradation. In this study we investigated whether delayed minocycline inhibits brain MMPs activated by ischemia in a model of temporary occlusion in Wistar rats.. Both MMP-2 and MMP-9 were elevated in the ischemic tissue as compared to the contra-lateral hemisphere after 3 hours occlusion and 21 hours survival (p < 0.0001 for MMP-9). Intraperitoneal minocycline at 45 mg/kg concentration twice a day (first dose immediately after the onset of reperfusion) significantly reduced gelatinolytic activity of ischemia-elevated MMP-2 and MMP-9 (p < 0.0003). Treatment also reduced protein concentration of both enzymes (p < 0.038 for MMP-9 and p < 0.018 for MMP-2). In vitro incubation of minocycline in concentrations as low as 0.1 mug/ml with recombinant MMP-2 and MMP-9 impaired enzymatic activity and MMP-9 was more sensitive at lower minocycline concentrations (p < 0.05).. Minocycline inhibits enzymatic activity of gelatin proteases activated by ischemia after experimental stroke and is likely to be selective for MMP-9 at low doses. Minocycline is a potential new therapeutic agent to acute treatment of ischemic stroke.

Topics: Animals; Anti-Inflammatory Agents; Apoptosis; Blood-Brain Barrier; Brain; Brain Ischemia; Disease Models, Animal; Drug Administration Schedule; Encephalitis; Enzyme Activation; Enzyme Inhibitors; Extracellular Matrix; Male; Matrix Metalloproteinase 2; Matrix Metalloproteinase 9; Matrix Metalloproteinase Inhibitors; Matrix Metalloproteinases; Minocycline; Nerve Degeneration; Neuroprotective Agents; Rats; Rats, Wistar; Stroke; Time Factors; Treatment Outcome

Minocycline is protective in models of transient middle cerebral artery occlusion (MCAO). We studied whether minocycline and doxycycline, another tetracycline derivative, provide protection in permanent MCAO. Because minocycline inhibits matrix metalloprotease-9 (MMP-9), we also compared minocycline's protective effect in wild type (wt) and MMP-9 knock-out (ko) mice. Wt FVB/N, Balb/C, and two lines of MMP-9 ko and their wt C57Bl/6 control mice were subjected to 24- or 72-hour permanent MCAO. Drug administration was started either 12 hours before or 2 hours after the onset of MCAO. Infarct size was determined by triphenyltetrazolium staining or T2-weighted MRI. Zymography was used to study the expression of MMPs. In wt strains, tetracycline treatments started before MCAO reduced the infarct size by 25% to 50%, whereas the treatment started after MCAO was not protective. Minocycline inhibited ischemia-provoked pro-MMP-9 induction in wt mice, but was not protective in MMP-9 ko mice. Pro-MMP-2 was induced by MCAO in wt and MMP-9 ko mice. MCAO-induced pro-MMP-2 was downregulated by minocycline treatment in wt mice but remained in MMP-9 ko mice at the same level as in saline-treated wt mice. Tetracyclines are protective in permanent MCAO when the treatment is started before the insult. Minocycline may provide protection by interfering with MMPs.

Topics: Animals; Anti-Bacterial Agents; Brain Ischemia; Cerebral Cortex; Down-Regulation; Doxycycline; Infarction, Middle Cerebral Artery; Male; Matrix Metalloproteinase 9; Matrix Metalloproteinase Inhibitors; Mice; Mice, Inbred BALB C; Mice, Knockout; Minocycline

To establish an in vitro model of hippocampal slice to detect electrophysiological alteration after oxygen/glucose deprivation (OGD), and to observe the effects of edaravone, minocycline and ONO-1078 [pranlukast, 4-oxo-8-[p-(4-phenylbutyloxy) benzoyl-amino]-2-(tetrazol-5-yl)-4H-1-benzopyran hemihydrate].. Hippocampal slices from rats were perfused with artificial cerebrospinal fluid lacking oxygen and glucose for 3, 4, 7 and 10 min. The population spike (PS) was recorded, and 2,3,5-triphenyltetrazolium chloride (TTC) staining was performed in some experiments, to detect the slice viability in the presence or absence of drugs in the perfusion solution.. Four min of OGD treatment was the most suitable duration for induction of slice injury, and PS amplitudes were recovered to (29 +/- 6)% of baseline values within 1 h after 4 min OGD. Edaravone, a free radical scavenger, at 1 and 10 mumol.L-1 significantly increased the recovery rate to (56 +/- 13)% and (69 +/- 12)% of baseline respectively 1 h after OGD. However, the anti-inflammatory drug minocycline (10 mumol.L-1) and leukotriene receptor antagonist ONO-1078 (1 mumol.L-1) did not increase the recovery. NMDA receptor antagonist ketamine, as a positive control, also promoted the recovery concentration-dependently.. OGD for 4 min was a feasible in vitro ischemia model for determination on electrophysiological alteration in hippocampal slices. Edaravone showed concentration-dependent protective effect on OGD injury, and anti-inflammatory drugs minocycline and ONO-1078 showed no effect.

Topics: Animals; Anti-Bacterial Agents; Antipyrine; Brain Ischemia; Chromones; Edaravone; Evoked Potentials; Female; Free Radical Scavengers; Glucose; Hippocampus; Leukotriene Antagonists; Minocycline; Neuroprotective Agents; Rats; Rats, Sprague-Dawley

Hypoxic-ischemic encephalopathy (HIE) is a leading cause of mortality and morbidity during the perinatal period, and currently no therapeutic drug is available. Minocycline, an antibiotic, has recently been shown to have neuroprotective effects distinct from its antimicrobial effect in several neurological disorders including ischemic brain injury. We examined the effect of minocycline on neonatal hypoxic-ischemic brain injury by using histologic scoring in both mouse and rat models. Mouse (C57Bl/6) and rat (SD) pups were exposed to a unilateral hypoxic-ischemic insult at 8 and 7 days of age, respectively. Minocycline hydrochloride was administered according to protocols that were reported to provide neuroprotection in adult or neonatal rats. Seven days after the insult, we examined brain injury in Nissl stained sections. Although minocycline ameliorated brain injury in the developing rat, it increased injury in the developing mouse. This detrimental effect in the mouse was consistent across different regions (cortex, striatum, and thalamus), with both single and multiple injection protocols and with both moderate and high-dose treatment (P < 0.05). The mechanism of the contrasting effects in mouse and rat is not clear and remains to be elucidated. Minocycline has been used as an antibiotic in the clinical setting for decades; therefore, it may be considered for use in infants with hypoxic-ischemic brain damage, based on prior reports of neuroprotection in the rat. However, it is important to examine this drug carefully before clinical use in human infants, taking our data in the mouse model into consideration.

Topics: Analysis of Variance; Animals; Animals, Newborn; Brain Infarction; Brain Ischemia; Disease Models, Animal; Dose-Response Relationship, Drug; Drug Administration Schedule; Female; Hypoxia-Ischemia, Brain; Male; Mice; Mice, Inbred C57BL; Minocycline; Random Allocation; Rats; Sensitivity and Specificity; Statistics, Nonparametric

Minocycline has been reported to exert neuroprotection through inhibition of inflammatory processes and of mitochondrial cell death pathway. To further characterize the neuroprotective effect of minocycline, we determined its efficacy in different neuronal damage paradigms involving inflammation or mitochondrial dysfunction. In transient global ischaemia in gerbils, minocycline reduced hippocampal neuronal damage measured by peripheral type benzodiazepine binding sites density, a marker of microglial activation. The antiinflammatory properties of minocycline were confirmed on the model of carrageenan-induced paw oedema in rats. The use of two experimental animal models involving administration of mitochondrial toxins inhibiting a different complex of the mitochondrial respiratory chain permitted the exploration of the mitochondrial impact of minocycline. Although minocycline exhibited a marked efficacy in 1-methyl-4-phenyl-1,2,5,6-tetrahydropyridine (MPTP; complex I inhibitor)-induced neurotoxicity in mice, it was ineffective in malonate (complex II inhibitor)-induced striatal lesion in rats. In vitro investigations on energized mitochondria isolated from rat liver showed that minocycline (1 microM) did not inhibit the swelling induced by MPP+(1-methyl-4-phenylpyridinium). Moreover, higher concentrations of minocycline induced swelling. From these experiments, the neuroprotective activity of minocycline appears more related to its antiinflammatory activity than to a direct beneficial action on mitochondria.

Topics: Animals; Binding, Competitive; Brain Ischemia; Carrageenan; Corpus Striatum; Disease Models, Animal; Dopamine; Dose-Response Relationship, Drug; Edema; Gerbillinae; Hindlimb; Isoquinolines; Male; Malonates; Mice; Mice, Inbred C57BL; Minocycline; Mitochondrial Swelling; MPTP Poisoning; Neuroprotective Agents; Rats; Rats, Sprague-Dawley; Time Factors; Tritium

Inflammatory reactions occurring in the brain after ischemia may contribute to secondary damage. In the present study effects of minocycline, an anti-inflammatory agent, alone or in combination with mild hypothermia, on focal embolic brain ischemia have been examined. Focal ischemic injury was induced by embolizing a preformed clot into the middle cerebral artery (MCA). Infarct volume was measured at 48 h after the injury. Administration of minocycline alone or minocycline plus mild hypothermia reduced infarct volume significantly. However, mild hypothermia in combination with minocycline did not show any additive effect. These results suggest that minocycline is beneficial in focal ischemic brain injury, and the lack of the enhanced neuroprotection may be due to the brief exposure to hypothermia.

Topics: Animals; Anti-Inflammatory Agents, Non-Steroidal; Brain Ischemia; Combined Modality Therapy; Hypothermia, Induced; Infarction, Middle Cerebral Artery; Intracranial Embolism; Male; Minocycline; Rats; Rats, Wistar; Stroke; Survival Analysis

The only treatment of patients with acute ischemic stroke is thrombolytic therapy, which benefits only a fraction of stroke patients. Both human and experimental studies indicate that ischemic stroke involves secondary inflammation that significantly contributes to the outcome after ischemic insult. Minocycline is a semisynthetic second-generation tetracycline that exerts antiinflammatory effects that are completely separate from its antimicrobial action. Because tetracycline treatment is clinically well tolerated, we investigated whether minocycline protects against focal brain ischemia with a wide therapeutic window. Using a rat model of transient middle cerebral artery occlusion, we show that daily treatment with minocycline reduces cortical infarction volume by 76 +/- 22% when the treatment is started 12 h before ischemia and by 63 +/- 35% when started even 4 h after the onset of ischemia. The treatment inhibits morphological activation of microglia in the area adjacent to the infarction, inhibits induction of IL-1beta-converting enzyme, and reduces cyclooxygenase-2 expression and prostaglandin E(2) production. Minocycline had no effect on astrogliosis or spreading depression, a wave of ionic transients thought to contribute to enlargement of cortical infarction. Treatment with minocycline may act directly on brain cells, because cultured primary neurons were also salvaged from glutamate toxicity. Minocycline may represent a prototype of an antiinflammatory compound that provides protection against ischemic stroke and has a clinically relevant therapeutic window.

Topics: Animals; Anti-Inflammatory Agents, Non-Steroidal; Base Sequence; Brain Ischemia; Cyclooxygenase 2; Dinoprostone; DNA Primers; Immunohistochemistry; Inflammation; Isoenzymes; Male; Minocycline; Prostaglandin-Endoperoxide Synthases; Rats; Rats, Sprague-Dawley; Rats, Wistar; Receptors, N-Methyl-D-Aspartate