Effect of Tongxinluo on basic fibroblast growth factor mRNA after cerebral ischemia/reperfusion injury

摘要

BACKGROUND:Basic fibroblast growth factor(Bfgf)can cause proliferation and differentiation of plentiful cells which are derived from mesoblast and neural ectoblast.Previous researches suggested that Tongxinluo could promote differentiation from neural stem cells(NSCs)to neurons and astrocytes after cerebral ischemia/repefusion injury.OBJECTIVE:TO observe the effect of Tongxinluo on Bfgf Mrna of rats after cerebral ischemia/reperfusion injury and analyze its dosage-dependence.DESIGN:Randomized controlled study.SETTING:Department of Neurology,Nanfang Hospital of Southern Medical University.MATERIALS: The experiment was carried out in Neurological Department of Nanfang Hospital affiliated to Southern Medical University from May 2005 to March 2006.A total of 192 healthy Wistar rats, weighing (200±20)g,of SPF grade,of both gender by half,were provided by Experimental Animal Center of Nanfang Hospital Affiliated to Southern Medical University.Tongxinluo was consisted of renshen,wugong,quanxie.shuizhi chishao, bingpian, etc.and was provided by Shijiazhuang Yiing Pharmaceutical Co.,Ltd.(batchnumber:9906115).Main reagents:Agarose,DEPC-treated water,total RNA purification kit and one-step reverse transcription polymerase chain reaction(RT-PCR) kit were provided by Beijing Saibaisheng Gene Technique Co.,Ltd.METHODS:All rats were randomly divided into 4 groups,including high-dosage Tongxinluo group,low-dosage Tongxinluo group,model group and sham operation group with 48 in each group.Rats in the former three groups were observed at 3,5,7,14,21 and 30 days after ischemia/reperfusion.In addition,rats in sham operation group were selected at the relevant time points.Every 8 rats were selected from each group for once.①Right middle cerebral artery occlusion-reperfusion models were induced by Koizumi method and modified;moreover,neurological impairment was evaluated with Zealonga method.Except rats in sham operation group.rats in other three groups were involved in the final analysis.if scores of neurological impairment were higher than 2 at 6 hours after modeing.②Administration:Rats in high-dosage Tongxinluo group were perfused with 150 g/L 1 g/(kg·d) Tongxinluo suspension;rats in low-dosage Tongxinluo group were perfused with 75 g/L 0.5 g/(kg·d) Tongxinluo suspension;rats in sham operation group and control group were perfused with the same volume of distilled water.Rats in each group were administrated at the wakeful time after reperfusion immediately.③Preparation of brain tissue:Rats were anesthetized at 3,5,7,14,21 and 30 days after perfusion to obtain their brains,separete brain tissue on ischemic site,extract total RNA,perform RT-PCR and semi-quantitative analysis and defect expression of bF-GF mRNA at ischemic brain tissue.④Statistical analysis:Intensity at each time point was measured with analysis of variance and Ftest.MAIN OUTCOME MEASURES: Changes of bFGF mRNA during proliferation and differentiation of NSCs in ischemic brain tissue at each time point after cerebral ischemia/reperfusion injury.RESULTS:All 192 rats were involved in the final analysis.Expression of bFGF mRNA was hardly measured at focal side of rats in sham operation group.Otherwise.expression of bFGF mRNA was measured at various time points at ischemic side of rats in model group;moreover,the expression was the srongest on the 7th day,decreased on the 14th day and fallen down to the basic value on the 30th day.At 5,7,14.21 and 30 days after modeing,expressions of bFGF mRNA at focal side were 0.86±0.04,0.87±0.04:0.88±0.02,0.87±0.02;0.79±0.02, O.81±0.03;0.52±0.04, 0.54±0.04;0.46±O.03, 0.45±0.03 in high-dosage Tongxinluo group andlow-dosage Tongxinluo group,respectively,which were stronger than those in model group(0.62±0.06,0.63±O.05,0.52±0.06,0.43±0.06,0.19±0.04,P＜0.05).Expression of bFGF mRNA at each time point was similar in high-dosage Tongxinluo group to that in low-dosage Tongxinluo group.In addition.expression of bFGF mRNA was the strongest at focal side in high-dosage Tongxinluo group and low-dosage Tongxinluo group on the 7th day after modeing;meanwhile,the expression was still stronger within 30 days after ischemia.CONCLUSION:①Tongxinluo can strengthen the expression of bFGF mRNA at ischemic side;meariwhile,the expression may last for 30 days.②This effect does not show as dosage-dependence.