minocycline and thiazolyl-blue

Alzheimer's disease (AD), a common neurodegenerative disorder, recognized to be a major cause of dementia. The aim of the present study was to investigate the neuroprotective mechanisms of clove oil in intracerebroventricular (icv)-colchicine induced cognitive dysfunction in rats.. Single bilateral icv-colchicine (15μg/5μl) was administered, followed by drug treatment with clove oil (0.05ml/kg and 0.1ml/kg, ip), minocycline (25 and 50mg/kg, ip) and their combinations for a period of 21 days. Various neurobehavioral parameters followed by biochemical, acetylcholinesterase (AChE) level and mitochondrial respiratory enzyme complexes (I-IV) were assessed.. Colchicine icv administration significantly impaired cognitive performance in Morris water maze (MWM) causes oxidative stress, raised AChE level, caused neuroinflammation and mitochondrial dysfunction as compared to sham treatment. Treatment with clove oil (0.05ml/kg and 0.1ml/kg) and minocycline (25 and 50mg/kg) alone significantly improved cognitive performance as evidenced by reduced transfer latency and increased time spent in target quadrant (TSTQ) in MWM task, reduced AChE activity, oxidative damage (reduced lipid peroxidation levels, nitrite level and restored glutathione levels) and restored mitochondrial respiratory enzyme complex (I-IV) activities as compared to icv-colchicine treatment. Further, combinations of clove oil (0.1ml/kg) with minocycline (50mg/kg) significantly modulate the neuroprotective effect of clove oil as compared to their effect alone.. The present study highlights that the major neuroprotective effect of clove oil due to its mitochondrial restoring and anti-oxidant properties along with a microglial inhibitory mechanism.

Topics: Acetylcholinesterase; Animals; Brain; Clove Oil; Cognition Disorders; Colchicine; Drug Synergism; Electron Transport Complex IV; Infusions, Intraventricular; Male; Minocycline; Mitochondria; NADH Dehydrogenase; Neuroprotective Agents; Oxidative Stress; Rats; Succinate Dehydrogenase; Tetrazolium Salts; Thiazoles

A 3-antibiotic combination (3Mix) has been widely used in regenerative endodontics. Recent studies recommend that a safe concentration of 3Mix is in the range of 0.39 μg/mL and 1 mg/mL because higher concentrations may limit tissue regeneration. The aim of this study was to determine the regenerative capacity of isolated human dental pulp cells (DPCs) and apical papilla cells (APCs) after a 7-day treatment with selected doses of 3Mix.. Primary human DPCs/APCs from the third passage were divided into control and experimental groups. In the control group, cells were cultured in regular complete media. In the experimental group, cells were cultured in complete media containing 0.39 μg/mL or 1 mg/mL of 3Mix for 7 days. After the treatment period, the media were changed, and the cells were further tested for proliferation and differentiation potential. For cell proliferation, a colorimetric qualification of 3-[4, 5-dimethylthiazol-2-yl]-2,5-diphenyl tetrazolium bromide was used on days 1, 3, 5, and 7. For differentiation potential, a dentinogenic differentiation medium was added into treated cells and cultured for 7, 14, and 21 days. Results were analyzed using quantitative alizarin red S staining and real-time reverse-transcription polymerase chain reaction.. After 7 days of treatment, 100% cell death was discovered in the 1-mg/mL 3Mix group. The proliferative capacity of 0.39 μg/mL 3Mix-treated DPCs and APCs was significantly lower than that of untreated cells at all time points (P < .05). Mineralized nodule formation was found both in the 3Mix-treated and control groups, but it was significantly less in the 3Mix-treated groups at 7, 14, and 21 days (P < .01). Quantitative reverse-transcription polymerase chain reaction showed no statistically significant difference (95% confidence interval) in bone sialoprotein, alkaline phosphatase, and dentin matrix protein 1 gene expression in either 3Mix-treated DPCs or APCs compared with control groups.. One milligram per milliliter of 3Mix had strong toxicity to DPCs/APCs when applied for 7 days, whereas 0.39 μg/mL 3Mix showed no toxicity but still affected cell proliferation and mineralization potential. However, no differences in dentinogenic gene expressions were observed between the 3Mix-treated and untreated groups.

Topics: Adolescent; Adult; Alkaline Phosphatase; Anthraquinones; Anti-Bacterial Agents; Calcification, Physiologic; Cell Culture Techniques; Cell Death; Cell Differentiation; Cell Proliferation; Cells, Cultured; Ciprofloxacin; Coloring Agents; Dental Papilla; Dental Pulp; Dentinogenesis; Extracellular Matrix Proteins; Humans; Integrin-Binding Sialoprotein; Metronidazole; Minocycline; Osteogenesis; Phosphoproteins; Regeneration; Tetrazolium Salts; Thiazoles; Time Factors; Young Adult

Ischemic brain injury is widely modeled in vitro with paradigms of oxygen-glucose deprivation (OGD), which leads to cell death. The prevention and attenuation of brain injury by the tetracycline antibiotic minocycline has been attributed largely to suppression of microglial activation, but its benefits in oligodendrocyte cells have not been well characterized. Using primary cultures of rat oligodendroglial precursor cells (OPC) exposed to OGD, we investigated the direct effects of minocycline on the survival, proliferation, and maturation of oligodendroglial lineage cells. OGD for 2 hr caused a decrease in the total number of OPC and the amount of proliferating progenitors by 50%, which was attenuated by inclusion of minocycline. The reduced numbers of immature oligodendroglial cells at 72 hr and of mature oligodendrocytes at 120 hr after OGD were partially restored by minocycline. In OPC, OGD caused an increase of reactive oxygen species (ROS) and production of TUNEL-positive cell numbers, which was abolished by minocycline. Minocycline preferentially increased the expression of superoxide dismutase under OGD but not in control OPC. Minocycline also prevented the OGD-induced downregulation of the transcription factors Sox10 and Olig2 and of myelin-specific genes 2'3' cyclic nucleotide phosphodiesterase (CNP) and myelin basic protein (MBP) in response to OGD. These studies demonstrate direct protective actions of minocycline on oligodendroglial-lineage cells, suggesting potential benefit in white matter injury involving OGD.

Topics: Analysis of Variance; Animals; Apoptosis; Cell Differentiation; Cell Proliferation; Cells, Cultured; Embryo, Mammalian; Embryonic Stem Cells; Female; Gangliosides; Gene Expression Regulation; Glucose; Hypoxia; Ki-67 Antigen; Minocycline; Myelin Basic Protein; O Antigens; Oligodendroglia; Oxidative Stress; Pregnancy; Rats; Rats, Sprague-Dawley; Tetrazolium Salts; Thiazoles; Time Factors; Transcription Factors

The purpose of this study was to determine the effect of the active substance of three types of local delivery systems, doxycycline hyclate 10% (DOXY), chlorhexidine gluconate, 2.5 mg (CHX), and minocycline hydrochloride, 1 mg (MINO), on osteoblastic cell proliferation and differentiation.. There were four groups: control osteoblastic cells (OB) alone, OB + DOXY, OB + CHX, and OB + MINO. Trypan blue and MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] assays were used to test osteoblastic cell viability. Cell differentiation was tested by measuring alkaline phosphatase levels. Osteoblast morphology was investigated by light and scanning electron microscopy.. At a concentration of 0.5 mg/ml, the Trypan blue test showed that DOXY, MINO, and CHX had significant toxicity effects on osteoblast cells compared to the control group, with a mean cell viability of 84%, 74%, and 51%, respectively (P <0.05). The MTT test showed that the control and DOXY groups were statistically significantly different (P <0.05) compared to CHX and MINO groups. The DOXY group showed a significantly higher alkaline phosphatase activity ( approximately 56%) than the control and MINO groups, and it was nearly 178% higher than the CHX group (P <0.05). The morphology of the osteoblasts seemed to be slightly altered when they were incubated with DOXY; however, with MINO, they appeared rounded with minimal attachment. In the CHX group, the osteoblasts assumed a shape of a very thin filopodia with a volcano-like nucleus.. At a concentration of 0.5 mg/ml, CHX and, to a lesser extent, MINO had a cytotoxic effect on osteoblast proliferation in vitro. However, DOXY seemed to enhance maturation and differentiation rather than proliferation. In addition to DOXY's beneficial effect as an adjunctive therapy to mechanical debridement in the treatment of periodontal disease, it may have an effect on periodontal regeneration.

Topics: Alkaline Phosphatase; Anti-Bacterial Agents; Anti-Infective Agents, Local; Biomarkers; Cell Adhesion; Cell Differentiation; Cell Nucleus; Cell Proliferation; Cell Shape; Cell Survival; Cells, Cultured; Chlorhexidine; Coloring Agents; Doxycycline; Drug Carriers; Female; Gels; Humans; Male; Microscopy, Electron, Scanning; Microspheres; Middle Aged; Minocycline; Osteoblasts; Pseudopodia; Tetrazolium Salts; Thiazoles; Trypan Blue

Recently, we have reported that minocycline, a semi-synthetic tetracycline with neuroprotective effects, inhibits the in vitro ischemic-like injury and 5-lipoxygenase (5-LOX) activation in PC12 cells. In the present study, we further determined whether minocycline protects PC12 cells from excitotoxicity via inhibiting 5-LOX activation. We used N-methyl-d-aspartate (NMDA, 200 microM) to induce early (exposure for 6 h) and delayed (exposure for 6 h followed by 24 h recovery) injuries. We found that NMDA receptor antagonist ketamine, 5-LOX inhibitor caffeic acid and minocycline concentration dependently attenuated NMDA-induced early and delayed cell injuries (viability reduction and cell death). However, only ketamine (1 microM) inhibited NMDA-evoked elevation of intracellular calcium. In addition, immunohistochemical analysis showed that NMDA induced 5-LOX translocation to the nuclear membrane after 1- to 6-h exposure which was confirmed by Western blotting, indicating that 5-LOX was activated. Ketamine, caffeic acid and minocycline (each at 1 microM) inhibited 5-LOX translocation after early injury. After delayed injury, PC12 cells were shrunk, and 5-LOX was translocated to the nuclei and nuclear membrane; ketamine, caffeic acid and minocycline inhibited both cell shrinking and 5-LOX translocation. As a control, 12-LOX inhibitor baicalein showed a weak effect on cell viability and death, but no effect on 5-LOX translocation. Therefore, we conclude that the protective effect of minocycline on NMDA-induced injury is partly mediated by inhibiting 5-LOX activation.

Topics: Analysis of Variance; Animals; Arachidonate 5-Lipoxygenase; Blotting, Western; Calcium; Dose-Response Relationship, Drug; Drug Interactions; Enzyme Activation; Enzyme Inhibitors; Excitatory Amino Acid Agonists; Excitatory Amino Acid Antagonists; Flavanones; Immunohistochemistry; Ketamine; Minocycline; N-Methylaspartate; Neuroprotective Agents; PC12 Cells; Rats; Tetrazolium Salts; Thiazoles; Time Factors

Minocycline, a semisynthetic derivative of tetracycline, displays beneficial activity in neuroprotective in models including, Parkinson disease, spinal cord injury, amyotrophic lateral sclerosis, Huntington disease and stroke. The mechanisms by which minocycline inhibits apoptosis remain poorly understood. In the present report we have investigated the effects of minocycline on mitochondria, due to their crucial role in apoptotic pathways. In mitochondria isolated suspensions, minocycline failed to block superoxide-induced swelling but was effective in blocking mitochondrial swelling induced by calcium. This latter effect might be mediated through dissipation of mitochondrial transmembrane potential and blockade of mitochondrial calcium uptake. Consistently, minocycline fails to protect SH-SY5Y cell cultures against reactive oxygen species-mediated cell death, including malonate and 6-hydroxydopamine treatments, but it is effective against staurosporine-induced cytotoxicity. The effects of this antibiotic on mitochondrial respiratory chain complex were also analyzed. Minocycline did not modify complex IV activity, and only at the higher concentration tested (100 microM) inhibited complex II/III activity. Other members of the minocycline antibiotic family like tetracycline failed to induce these mitochondrial effects.

Topics: Animals; Calcium; Cell Count; Cell Death; Cell Line, Tumor; Cell Survival; Dose-Response Relationship, Drug; Drug Interactions; Enzyme Inhibitors; Glutathione; Humans; Membrane Potentials; Minocycline; Mitochondria; Mitochondrial Swelling; NADP; Neuroblastoma; Neuroprotective Agents; Rats; Rats, Sprague-Dawley; Spectrophotometry; Staurosporine; Tetrazolium Salts; Thiazoles

Minocycline has been shown to have remarkably neuroprotective qualities, but underlying mechanisms remain elusive. We reported here the robust neuroprotection by minocycline against glutamate-induced apoptosis through regulations of p38 and Akt pathways. Pre-treatment of cerebellar granule neurons (CGNs) with minocycline (10-100 microm) elicited a dose-dependent reduction of glutamate excitotoxicity and blocked glutamate-induced nuclear condensation and DNA fragmentations. Using patch-clamping and fluorescence Ca2+ imaging techniques, it was found that minocycline neither blocked NMDA receptors, nor reduced glutamate-caused rises in intracellular Ca2+. Instead, confirmed by immunoblots, minocycline in vivo and in vitro was shown to directly inhibit the activation of p38 caused by glutamate. A p38-specific inhibitor, SB203580, also attenuated glutamate excitotoxicity. Furthermore, the neuroprotective effects of minocycline were blocked by phosphatidylinositol 3-kinase (PI3-K) inhibitors LY294002 and wortmannin, while pharmacologic inhibition of glycogen synthase kinase 3beta (GSK3beta) attenuated glutamate-induced apoptosis. In addition, immunoblots revealed that minocycline reversed the suppression of phosphorylated Akt and GSK3beta caused by glutamate, as were abolished by PI3-K inhibitors. These results demonstrate that minocycline prevents glutamate-induced apoptosis in CGNs by directly inhibiting p38 activity and maintaining the activation of PI3-K/Akt pathway, which offers a novel modality as to how the drug exerts protective effects.

Topics: Activating Transcription Factor 2; Animals; Animals, Newborn; Apoptosis; Blotting, Western; Calcium; Cell Count; Cell Survival; Cells, Cultured; Cerebellum; Chromatin; Cyclic AMP Response Element-Binding Protein; Dizocilpine Maleate; DNA Fragmentation; Dose-Response Relationship, Drug; Drug Interactions; Enzyme Inhibitors; Glutamic Acid; Humans; Membrane Potentials; Microscopy, Confocal; Minocycline; N-Methylaspartate; Neurons; Neuroprotective Agents; p38 Mitogen-Activated Protein Kinases; Patch-Clamp Techniques; Protein Serine-Threonine Kinases; Proto-Oncogene Proteins; Proto-Oncogene Proteins c-akt; Rats; Rats, Sprague-Dawley; Serine; Signal Transduction; Tetrazolium Salts; Thiazoles; Time Factors; Transcription Factors