minocycline and 3-nitrotyrosine

This study aimed to elucidate the protective effect of minocycline against streptomycin-induced damage of cochlear hair cells and its mechanism. Cochlear membranes were isolated from newborn Wistar rats and randomly divided into control, 500μmol/L streptomycin, 100μmol/L minocycline, and streptomycin and minocycline treatment groups. Hair cell survival was analyzed by detecting the expression of 3-nitrotyrosine (3-NT) in cochlear hair cells by immunofluorescence and an enzyme-linked immunosorbent assay. Expression of 3-NT and inducible nitric oxide synthase (iNOS), and poly (ADP-Ribose) polymerase (PARP) and caspase-3 activation were evaluated by western blotting. The results demonstrated hair cell loss at 24h after streptomycin treatment. No change was found in supporting cells of the cochleae. Minocycline pretreatment improved hair cell survival and significantly reduced the expression of iNOS and 3-NT in cochlear tissues compared with the streptomycin treatment group. PARP and caspase-3 activation was increased in the streptomycin treatment group compared with the control group, and pretreatment with minocycline decreased cleaved PARP and activated caspase-3 expression. Minocycline protected cochlear hair cells from injury caused by streptomycin in vitro. The mechanism underlying the protective effect may be associated with the inhibition of excessive formation of nitric oxide, reduction of the nitration stress reaction, and inhibition of PARP and caspase-3 activation in cochlear hair cells. Combined minocycline therapy can be applied to patients requiring streptomycin treatment.

Topics: Animals; Anti-Bacterial Agents; Caspase 3; Cell Death; Cochlea; Enzyme Activation; Hair Cells, Auditory; Minocycline; Neuroprotective Agents; Nitric Oxide; Nitric Oxide Synthase Type II; Poly(ADP-ribose) Polymerases; Rats, Wistar; Streptomycin; Transcription, Genetic; Tyrosine

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

Surgical correction of congenital cardiac malformations mostly implies the use of cardiopulmonary bypass (CPB). However, a possible negative impact of CPB on cerebral structures like the hippocampus cannot be neglected. Therefore, we investigated the effect of CPB on hippocampus CA1 and CA3 regions without or with the addition of epigallocatechin-3-gallate (EGCG) or minocycline. We studied 42 piglets and divided them into six experimental groups: control without or with EGCG or minocycline, CPB without or with EGCG or minocycline. The piglets underwent 90 minutes CPB and subsequently, a 120-minute recovery and reperfusion phase. Thereafter, histology of the hippocampus was performed and the adenosine triphosphate (ATP) content was measured. Histologic evaluation revealed that CPB produced a significant peri-cellular edema in both CA regions. Moreover, we found an increased number of cells stained with markers for hypoxia, apoptosis and nitrosative stress. Most of these alterations were significantly reduced to or near to control levels by application of EGCG or minocycline. ATP content was significantly reduced within the hippocampus after CPB. This reduction could not be antagonized by EGCG or minocycline. In conclusion, CPB had a significant negative impact on the integrity of hippocampal neural cells. This cellular damage could be significantly attenuated by addition of EGCG or minocycline.

Topics: Adenosine Triphosphate; Animals; Apoptosis Inducing Factor; Brain Edema; CA1 Region, Hippocampal; CA3 Region, Hippocampal; Cardiopulmonary Bypass; Caspase 3; Catechin; Chromatography, High Pressure Liquid; Disease Models, Animal; Hypoxia-Inducible Factor 1, alpha Subunit; Minocycline; Neuroprotective Agents; Poly Adenosine Diphosphate Ribose; Swine; Tyrosine

Cardiopulmonary bypass (CPB) often is required for the operative correction of congenital heart defects in small infants. Unfortunately, CPB is associated with injury of inner organs such as the brain, kidney, lung, and liver. Renal failure and increase in liver enzymes are typical side effects observed after CPB. Here, we investigate whether organ protection of the kidney and liver can be achieved with the application of minocycline, which is known-besides its anti-infective effects-to act as a poly-ADP-ribose-polymerase inhibitor. Twenty-nine 4-week-old Angler Sattelschwein-piglets (8-15 kg) were divided into four groups: control group (n = 8), CPB group (n = 9), minocycline-control group (n = 6), and the minocycline-CPB group (n = 6). CPB groups were thoracotomized and underwent CPB for 120 min (cross-clamp, 90 min; reperfusion, 30 min) followed by a 90-min recovery time. The control groups also were thoracotomized but not connected to CPB. The minocycline group received 4 mg/kg minocycline before and 2 mg/kg after CPB. In the kidneys, CPB histologically resulted in widening of Bowman's capsule, and-mainly in tubules-formation of poly-ADP-ribose, nitrosylation of tyrosine-residues, nuclear translocation of hypoxia-induced factor HIF-1α, and of apoptosis-inducing factor (AIF). In addition, we found significantly less ATP in the kidney and significantly increased plasma urea and creatinine. Similar but gradually attenuated changes were found in the liver together with significantly elevated de-Ritis coefficient. These changes in the kidney and liver were significantly diminished by minocycline (except AIF in the liver which was similar in all groups). In conclusion, CPB causes damage in the kidney and-to a lower degree-in the liver, which can be attenuated by minocycline.

Topics: Animals; Apoptosis Inducing Factor; Cardiopulmonary Bypass; Hypoxia-Inducible Factor 1, alpha Subunit; Kidney; Liver; Minocycline; Poly(ADP-ribose) Polymerases; Protective Agents; Swine; Tyrosine

We previously reported that the precursor form of nerve growth factor (pro-NGF) and not mature NGF is liberated in the CNS in an activity-dependent manner, and that its maturation and degradation occur in the extracellular space by the coordinated action of proteases.Here, we present evidence of diminished conversion of pro-NGF to its mature form and of greater NGF degradation in Alzheimer disease (AD) brain samples compared with controls. These alterations of the NGF metabolic pathway likely resulted in the increased pro-NGF levels. The pro-NGF was largely in a peroxynitrited form in the AD samples. Intrahippocampal injection of amyloid-beta oligomers provoked similar upregulation of pro-NGF in naive rats that was accompanied by evidence of microglial activation (CD40), increased levels of inducible nitric oxide synthase, and increased activity of the NGF-degrading enzyme matrix metalloproteinase 9. The elevated inducible nitric oxide synthase provoked the generation of biologically inactive, peroxynitrite-modified pro-NGF in amyloid-beta oligomer-injected rats. These parameters were corrected by minocycline treatment. Minocycline also diminished altered matrix metalloproteinase 9, inducible nitric oxide synthase, and microglial activation (CD40); improved cognitive behavior; and normalized pro-NGF levels in a transgenic mouse AD model. The effects of amyloid-beta amyloid CNS burden on NGF metabolism may explain the paradoxical upregulation of pro-NGF in AD accompanied by atrophy of forebrain cholinergic neurons.

Topics: Aged; Aged, 80 and over; Alzheimer Disease; Amyloid beta-Peptides; Amyloid beta-Protein Precursor; Animals; Brain; CD40 Antigens; Disease Models, Animal; Female; Humans; Immunoprecipitation; Male; Matrix Metalloproteinase 9; Maze Learning; Mice; Mice, Transgenic; Minocycline; Nerve Growth Factor; Nerve Growth Factors; Nitric Oxide Synthase Type II; Peptide Fragments; Peroxynitrous Acid; Protein Precursors; Rats; Rats, Inbred F344; Reaction Time; Tyrosine; Up-Regulation

This work has examined levels of 3-nitrotyrosine (3-NT, a marker for peroxynitrite formation) and intactness of blood-brain barrier (BBB) in amyloid beta-peptide (Abeta(1-42))-injected rat hippocampus. Immunohistochemical analysis demonstrated 3-NT immunoreactivity in microglia/macrophages and astrocytes were significantly increased at 7 days post-Abeta(1-42) injection. Administration of the broad spectrum anti-inflammatory agent minocycline or the selective iNOS inhibitor 1400W markedly reduced 3-NT levels. Double immunofluorescence staining showed that 3-NT was prominently expressed in microglia/macrophages and astrocytes located in proximity to blood vessels. Additionally, Abeta(1-42) injection caused a marked increase in permeability of the BBB to immunoglobulin G (IgG); both minocycline and 1400W were highly effective in decreasing the leakiness of the BBB. Our results suggest the involvement of glial-derived reactive nitrogen species in mediating increased BBB permeability in Abeta(1-42) injected rat hippocampus.

Topics: Amidines; Amyloid beta-Peptides; Animals; Astrocytes; Benzylamines; Blood-Brain Barrier; CD11b Antigen; Cell Count; Drug Interactions; Enzyme Inhibitors; Fluorescent Antibody Technique; Gene Expression Regulation; Glial Fibrillary Acidic Protein; Hippocampus; Male; Microglia; Minocycline; Nitric Oxide Synthase Type II; Peptide Fragments; Rats; Rats, Sprague-Dawley; Tyrosine

1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) damages the nigrostriatal dopaminergic pathway as seen in Parkinson's disease (PD), a common neurodegenerative disorder with no effective protective treatment. Consistent with a role of glial cells in PD neurodegeneration, here we show that minocycline, an approved tetracycline derivative that inhibits microglial activation independently of its antimicrobial properties, mitigates both the demise of nigrostriatal dopaminergic neurons and the formation of nitrotyrosine produced by MPTP. In addition, we show that minocycline not only prevents MPTP-induced activation of microglia but also the formation of mature interleukin-1beta and the activation of NADPH-oxidase and inducible nitric oxide synthase (iNOS), three key microglial-derived cytotoxic mediators. Previously, we demonstrated that ablation of iNOS attenuates MPTP-induced neurotoxicity. Now, we demonstrate that iNOS is not the only microglial-related culprit implicated in MPTP-induced toxicity because mutant iNOS-deficient mice treated with minocycline are more resistant to this neurotoxin than iNOS-deficient mice not treated with minocycline. This study demonstrates that microglial-related inflammatory events play a significant role in the MPTP neurotoxic process and suggests that minocycline may be a valuable neuroprotective agent for the treatment of PD.

Topics: 1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine; Animals; Anti-Bacterial Agents; Astrocytes; Corpus Striatum; Disease Models, Animal; Dose-Response Relationship, Drug; Interleukin-1; Mice; Mice, Inbred C57BL; Mice, Knockout; Microglia; Minocycline; NADPH Oxidases; Neurons; Neuroprotective Agents; Nitric Oxide Synthase; Nitric Oxide Synthase Type II; Parkinsonian Disorders; Substantia Nigra; Tyrosine; Tyrosine 3-Monooxygenase; Up-Regulation