Itaro Hattori

Intravenous Administration of Thioredoxin Decreases Brain Damage Following Transient Focal Cerebral Ischemia in Mice

Thioredoxin (TRX) is induced by a variety of oxidative stimuli and shows cytoprotective roles against oxidative stress. To clarify the possibility of clinical application, we examined the effects of intravenously administered TRX in a model of transient focal cerebral ischemia in this study. Mature male C57BL/6j mice received either continuous intravenous infusion of recombinant human TRX (rhTRX) over a range of 1-10 mg/kg, bovine serum albumin, or vehicle alone for 2 h after 90-min transient middle cerebral artery occlusion (MCAO). Twenty-four hours after the transient MCAO, the animals were evaluated neurologically and the infarct volumes were assessed. Infarct volume, neurological deficit, and protein carbonyl contents, a marker of protein oxidation, in the brain were significantly ameliorated in rhTRX-treated mice at the dose of 3 and 10 mg/kg versus these parameters in control animals. Moreover, activation of p38 mitogen-activated protein kinase, whose pathway is involved in ischemic neuronal death, was suppressed in the rhTRX-treated mice. Further, rhTRX was detected in the ischemic hemisphere by western blot analysis, suggesting that rhTRX was able to permeate the blood-brain barrier in the ischemic hemisphere. These data indicate that exogenous TRX exerts distinct cytoprotective effects on cerebral ischemia/reperfusion injury in mice by means of its redox-regulating activity.

Thioredoxin suppresses 1-methyl-4-phenylpyridinium-induced neurotoxicity in rat PC12 cells

Thioredoxin (TRX) is a redox-active protein which plays a cytoprotective role against oxidative stress. Geranylgeranylacetone (GGA), used widely as an anti-ulcer drug, has been reported to induce TRX as well as heat shock protein 70 (HSP70) in hepatocytes and other cells. 1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP), causes dopaminergic denervation and Parkinsonism in humans. The 1-methyl-4-phenylpyridinium ion (MPP(+)), an active metabolite of MPTP, induces cell death in a rat pheochromocytoma cell line (PC12 cells). We found that MPP(+) suppresses TRX expression in PC12 cells. Overexpression or administration of TRX attenuates MPP(+)-induced neurotoxicity on PC12 cells. Moreover, GGA induces expression of TRX and HSP70 and attenuates MPP(+)-induced toxicity in PC12 cells. These results indicate that TRX and GGA have a possible potential as new therapeutic agents for Parkinson disease.

Phosphorylation of c-Jun NH2-terminal kinase and p38 mitogen-activated protein kinase after transient forebrain ischemia in mice

We investigated the activation of c-Jun NH(2)-terminal kinase (JNK) and p38 mitogen-activated protein kinase (MAPK) during transient forebrain ischemia to clarify the roles of these stress kinases during brain ischemia. Mice were subjected bilateral common carotid artery (BCCA) occlusion for 20 min followed by reperfusion. Immunohistochemical analysis and Western blot analysis for active JNK and active p38 MAPK were performed at 0, 5, 10, 30 and 150 min after reperfusion. After 5 min of reperfusion, active JNK and p38 MAPK immunoreactivities were enhanced in neurons in the cerebral cortex and hippocampus; this activation peaked at 30 min of reperfusion. Stress kinases activation dominantly occurred in the similar regions, in which neurons with fragmented DNA were detected at 72 h after reperfusion. Western blot analysis indicated that JNK 1, JNK 2 and p38 MAPK were activated at 10 and 30 min after reperfusion. These findings indicate that JNK and p38 MAPK pathways may play important roles in neuronal death during brain ischemia.

Hypoxia-ischemia induces thioredoxin expression and nitrotyrosine formation in new-born rat brain

Abstract Thioredoxin (TRX) is a 13 kDa protein with antioxidant effect and redox regulating functions. Peroxynitrite is a strong oxidizing and nitrating agent which can react with all classes of biomolecules. In the present study, we focused on the association between TRX and nitrotyrosine, which served as a marker of peroxynitrite formation, in the neonatal hypoxia-ischemia (HI) rat brain. At 4-16 h after HI, the immunoreactivity for TRX was diminished in the injured region in the cortex and striatum, whereas nitrotyrosine immunoreactivity was enhanced. In contrast, around the injured region, TRX immunoreactivity was enhanced in survival neurons at 4-24 h after HI, while the immunoreactivity for nitrotyrosine was mostly not detected. Northern blot analysis showed increased TRX mRNA induction in the cerebral hemisphere ipsilateral to the carotid ligation from 4-24 h after HI but not in the contralateral hypoxic hemisphere. These findings suggest that production of peroxynitrite is involved in HI brain injury, and that induced TRX plays a neuroprotective role against oxidative stress resulting from HI.

Excitotoxic hippocampal injury is attenuated in thioredoxin transgenic mice.

Thioredoxin is a small, multifunctional protein with a redox-active disulfide/dithiol in the active site. Thioredoxin plays several important biologic roles both in intracellular and extracellular compartments with its redox-regulating and reactive oxygen intermediates scavenging activities. We assayed the seizure response and the excitotoxic hippocampal injury in thioredoxin transgenic and wild-type C57BL/6 mice. Seizure score after kainic acid treatment was significantly lower in thioredoxin transgenic mice. Seven days after kainic acid administration, the damage in the hippocampal CA1 and CA3 regions was significantly attenuated in thioredoxin transgenic mice. Thioredoxin and redox regulation play an important role in excitotoxic brain damage.

Expression and distribution of redox regulatory protein, thioredoxin after metabolic impairment by 3-nitropropionic acid in rat brain

Thioredoxin (TRX) is a small, multifunctional protein with a redox-active site and multiple biological functions that include reducing activity for reactive oxygen intermediates. We assayed TRX by immunohistochemical methods in the rat brain after intraperitoneal injection of 3-nitropropionic acid (3-NP), an irreversible inhibitor of succinate dehydrogenase. Systemic administration of 3-NP produced lateral striatal, hippocampal CA1 and CA3 lesions in the present study. The immunoreactivity for TRX was enhanced in hippocampal CA3, dentate gyrus and lateral striatum, but not detected in hippocampal CA1 subfield after 3-NP intoxication. The data suggest that TRX may play an important role in the pathogenesis of 3-NP neurotoxicity.

Redox regulation in neuronal damage during brain ischemia

Reactive oxygen species has been proved not only to damage cells and tissues but to play as second messenger in signal transduction pathways. Increasing evidence has indicated that the cellular redox status modulates various aspects of cellular events, including proliferation and apoptosis. Thioredoxin (TRX) plays important biological roles both in intra- and extracellular compartments including regulation of various intracellular molecules via thiol redox control. We have described the role of TRX after brain ischemia. TRX was induced in neurons after middle cerebral artery occlusion. Moreover TRX overexpressing mice has the resistance against focal ischemic brain damage. In addition, we found about 20% reduction of infarct volume in recombinant human TRX-treated mice versus control animals. Neurological deficit was also ameriorated significantly in rhTRX-treated mice compared with control mice. In this section, we describe the role of TRX in ischemic neuronal death. TRX and the redox state modified by TRX play a crucial role in brain damage during stroke.