脑蛋白水解物对神经细胞损伤修复活力测定方法的研究

Study of the Activity Determination of Cerebroprotein Hydrolysate on Nerve Cell Damage

关键词:  脑蛋白水解物  PC12细胞  活力测定

Key words:  cerebroprotein hydrolysate  PC12 cell  activity determination

摘要:

目的 评价脑蛋白水解物对H₂O₂诱导的PC12细胞损伤的修复效果，建立一种脑蛋白水解物注射液的生物活性检测方法。方法 以脑蛋白水解物的进口代表药物注射用脑蛋白水解物为例，采用PC12细胞培养，分析不同细胞接种浓度、不同浓度的H₂O₂损伤细胞、不同浓度脑蛋白水解物对抗干预的影响；采用MTT比色法对样品组、对照组和损伤组的细胞进行染色后，用酶标仪测定OD值，计算修复率，以评价脑蛋白水解物对模型细胞PC12损伤的保护作用；同时将该损伤修复评价方法用于国产脑蛋白水解物的生物活性检测。结果 在8×10⁴~12×10⁴·mL^-1细胞接种量、0.5 mmol·L^-1 H2O2损伤浓度下、60 μg·L^-1药物浓度(以含氮量计)下可成功的建立H₂O₂诱导的PC12细胞损伤模型。结论 采用该方法对8批国产脑蛋白水解物注射液样品及注射用脑蛋白水解物进行对比检测，发现对H₂O₂损伤的细胞均具有良好修复作用。

Abstract:

OBJECTIVE To evaluation the restoration of PC12 cell by cerebroprotein hydrolysate and provide criterion for the biological activity detection method of cerebroprotein hydrolysate. METHODS Cerebrolysin, the representative cerebroprotein hydrolysate imported, was adopted as repair agent in this study. PC12 cell was adopted as the indicator of nerve cell. Effects of concentration of cell inoculations, H₂O₂ and cerebroprotein hydrolysate on PC12 restoration were studied respectively. The protection of cerebroprotein hydrolysate on PC12 model cell was determined by MTT colorimetry which colorized the sample group, control group and damage group. Restoration rates were calculated by OD values. Biological activities of domestic products were evaluated by the method also. RESULTS The PC12 cell damage model could be established by the condition of 8×10⁴-12×10⁴ per·mL^-1, 0.5 mmol·L^-1 H₂O₂ and 60 μg·L^-1 cerebroprotein hydrolysate (calculated by nitrogen content). Contrast detections were carried out using this method. CONCLUSION Domestic cerebroprotein hydrolysate of 8 batch samples and cerebrolysin were proved to be effective on the restoration of PC12 cell damaged by H₂O₂.