摘要
目的以即刻早期基因c-fos和c-jun为标志物研究抑肽酶预处理对凝血酶诱导的星形胶质细胞内信号转导的改变,初步探讨大剂量凝血酶对星形胶质细胞损伤以及抑肽酶拮抗作用的机制,为进一步探讨抑肽酶的临床应用打下基础。方法以体外培养的星形胶质细胞为实验模型,采用在细胞培养基中混入抑肽酶和凝血酶的加药方式,利用免疫组织化学和免疫荧光技术,在显微镜下观测c-fos和c-jun基因在星形胶质细胞中的表达,计算两种基因在细胞内的阳性表达率,数据采用SPSS10.0进行卡方检验。结果体外原代培养的小鼠星形胶质细胞经纯化后纯度达到(93.5±3)%,药物处理24h后,T20组细胞阳性表达率为c-jun(71.74±0.74)%,c-fos(69.27±0.69)%,c-fos和c-jun二者表达趋势基本一致,其中A100组细胞阳性表达率非常显著地高于T20组(P<0.01),A300组显著高于T20组(P<0.05),A500与T20组无显著差异,A700组c-jun表达阳性率与T20组无显著性差别,而c-fos表达阳性率非常显著地低于T20组。结论小剂量抑肽酶可显著增加凝血酶诱导的即刻早期基因表达的细胞数目,但单个细胞表达的强度减弱,而大剂量的抑肽酶预处理使即刻早期基因表达的数目减少,表达强度增强,推测小剂量的抑肽酶可以逆转发生损伤的细胞,而大剂量时有促进已发生严重损伤的细胞进一步加重的可能。
Objective To investigate changes of action in astrocytes induced by thrombin after pretreatment with aprotinin,immediate-early gene c-fos and c-jun were being as the tools to reveal the changes of intercellular signal transduction. Damnification of thrombin to astrocytes and antagonization of aprotinin was also discussed in this article. And these maybe helpful to the next research in clinical. Methods Primary-cultured astrocytes after purification was used as the model, and astrocytes were treated with aprotinin and thrombin mixed in the culture medium for 24h, then immunohistochemical technique（ABC method） was employed to detect the expression of c-fos and c-jun. The data was analyzed in the software of SPSS10.0 with crosstable. Results The proportion of astrocyte was to （93. 5 ±3）% after purification of the primary-cultured neurocytes. After treatment with aprotinin and thrombin of 24h,the ratio of negative cells in the group of T20 was （71.74±0.74）% （c-jun） and （69.27±0.69）% （c-fos）. The two trends were matched with each other. Whether in the group of c-fos or c-jun, the ratio of negative cell in the group were： which in A100 markedly higher than that in the group of T20（P〈0.01）, A300 higher than T20. There was no difference among A500, A700 and T20 of c-jun, About c-fos, A700 was markedly lower than T20, and no difference between A500 and T20. Conclusion The measure of pretreatment with low-dose Aprotinin can markedly increase the amount of astrocytes expressing of c-fos and c-jun, but weaken the intensity. On the contrary, high-dose aprotinin can markedly reduce the amount, but enhance the intensity. We draw a conclusion that low-dose aprotinin maybe protective to the astrocytes but high-dose will embitter the cell which suffer the badly injury.
出处
《重庆医学》
CAS
CSCD
2005年第9期1365-1367,共3页
Chongqing medicine
