The Neuroprotective Effects And Mechanisms Of Catechin On Neuron Injury Induced By Acidosis In Rats

Objectives:Cerebral ischemia or stroke can do great harm to human health,with high rate of mutilation and high mortality.It is one of the leading causes of death and long-term disability in aged populations,now the morbility has been increasing in the youth,often results in irreversible brain damage and subsequent loss of neuronal function.At present,there is no specific effective medical therapy for this disease. Therefor,it is very important to use neuronal protective drugs accompanying with thrombolytic treatment.Lots of studies had proved that(+)-catechin may have protective effects on CNS injury by inhibiting oxidative stress.Then,(+)-catechin inhibited oxidative stress through scavenging of reactive oxygen pieces(ROS) and increasing the activation of antioxidative enzymes.There were more homoplastic activities in catechins and their derivants,but the difference of their activities contributed to the change of chemical and stereo-structure.(+)-catechin processed the more efficient antioxidative effect and the lower cytotoxity,the bioactivity was related with its stereo-structure.Recently,Lots of evidence has highlighted the role of acid-sensing ion channels(ASICs) in the central nervous system in various physiological and pathological processes such as learning and memory,emotion, ischemic brain injury and brain tumor.Especially in cerebral ischemia ASICs may play important role in the course of pathophysiology.It has become new pharmacal target that being researched.ASICs are ligand-sensitive and voltage-insensitive cation channels that activated by extracellular H⁺.But whether(+)-catechin can affect ASICs in ischemia-reperfusion injury is not seen internal and abroad.The objective of this study is to investigate the influence of(+)-catechin on the expression of ASIC1a of cortical neuron after acidosis in rats,so as to further explore the neuro-protective mechanisms of(+)-catechin and provide theoretical base for application of neuro-protective drugs in clinic.Methods:Cortical neurons of neonatal SD rats(within 24 hours) were cultured in vitro.The cells were collected,purified and incubated in 50cm² culture bottles(the cell density was adjusted to 1×10⁶ cells/ml with DMEM/F-12 culture liquid).Cultured cortical neurons of neonatal SD rats were divided five groups:normal group;injured group;high dose group(1.0mmol/L);middle dose group(0.5mmol/L);low dose group(0.25mol/L).Hydrogen Chloride(HCL) and sugar free were used to replicate the ischemia injury model.Cell death was also evaluated morphologically by phase-contrast light microscopy.Hydrogen peroxide(H₂O₂) and malondialdehyde (MDA) production were assessed by colorimetric micro assay.The mRNA levels of the genes ASIC1a of the cortical neurons injured by acidosis were evaluated by reverse transcription polymerase chain reaction(RT-PCR).Results:(1) Compared with normal group(73.15±2.81 mmol/L),the content of H₂O₂ was strongly increased in injured group(115.57±6.16 mmol/L)(P＜0.01),while the content of H₂O₂ in(+)-catechin treated groups were lower than that in injured group(P＜0.05);the content of H₂O₂ had no difference between low dose(102.39±5.56 mmol/L)and middle dose group(100.50±4.08 mmol/L)(P＞0.05).(2) Compared with normal group(4.21±0.51 nmol/ml),the content of MDA was strongly increased in injured group(11.22±0.85 nmol/ml )(P＜0.01),while the content of MDA in(+)-catechin treated groups were lower than that in injured group(P＜0.05),the content of MDA had difference among three groups(P＜0.05)(3) Compared with normal group(0.14±0.03),the ASIC1a mRNA expression was increased obviously in injured group(0.79±0.10 )(P＜0.01);Compared with injured group,the mRNA expression of ASIC1a was down-regulated in(+)-catechin treated groups(P＜0.05),the mRNA expression of ASIC1a had no difference between high dose(0.26±0.05) and middle dose group(0.31±0.07 )(P＞0.05).(4)(+)-catechin could obviously improve neuronic morphology in acidosis.Conclusions:(1)(+)-catechin strongly protect primary cultured neurons against acidosis-induced oxidative stress by decreasing the production of H₂O₂ and MDA in neurons.(2)(+)-catechin can also improve neuronic injury by decreasing the mRNA level of ASIC1a in neurons.