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Analysis of apoptotic mechanism of cerebellar granular cells

小脑颗粒细胞凋亡机制分析

基本信息

批准号：
09470058
负责人：
NAGASHIMA Kazuo
金额：
$ 6.53万
依托单位：
Hokkaido University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
1997
资助国家：
日本
起止时间：
1997 至 1999
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-09470058/
关键词：
apoptosis methylmercury chloride Western blotting 小脳顆粒細胞 Zicl 転写因子 ICE 有機水銀

项目摘要

We have established a rat model of injury of cerebellar granule cells by intoxication of methylmercury chloride (MMC). To elucidate the molecular mechanisms of brain damage induced by MMC we initial ly examined the pathologic examination by H&E staining, electron microscope, TUNEL staining, and DNA fragmentation. These examination revealed that cerebellar granule cell death by organic mercury intoxication occurs via an apoptotic process and was restricted to the granule cells and that the distributional pattern of apoptotic cerebellar granule cells was specific in rat cerebellar vermis. Secondly, we investigated expression levels of apoptosis related molecules such as; BDNF, Trk B, Fas ligand, RIP, Akt, Bad, Bcl-2, Caspase 9, Caspase 3, CAS, and TIAR using Western blot and also evaluated the function of Purkinje cells by immunostaining with anti-calbindin D. Moreover, we investigated the alteration of gene expression of rat brain after MMC intoxication. These results demonstrated that … More expression of BDNF and Trk B were up-regulated, while RIP, CAS, Fas ligand, Bad, and Caspase 9 were down-regulated. The findings suggest that the up-regulated molecules function to suppress apoptosis at this stage. Gene expression analysis disclosed that among up-regulated genes 1) those related to apoptosis were protein kinase C γ, interleukin 13, insulin receptor substrate Z, cyclophilin, protein phosphatase 1, and RL/IF-1, 2) those related to hormone were vasopressin and oxytocin, 3) those related to enzyme were serum and glucocorticoid-regulated kinase, mast cell protease, serine protease, glycoprotein specific UDP-glucuronyl transferase, protein tyrosine phosphatase, 4) 7 miscellaneous molecules, and 5) 7 unknown genes. In contrast, down-regulated genes were 2α1 globin gene and 2 unknown genes. The results could be summarized that apoptosis suppressive genes were activated at the beginning of cerebellar apoptosis. Thus, it could be concluded that in the initial stage of MMC intoxication the expression of anti-apoptotic molecules were more enhanced to protect cell death than apoptosis-inducing molecules. Less

本文建立了氯化甲基汞（MMC）中毒大鼠小脑颗粒细胞损伤模型。为探讨MMC致脑损伤的分子机制，我们采用HE染色、电镜、TUNEL染色、DNA片段化等方法对MMC致脑损伤的病理组织学进行了初步研究。这些检查表明，有机汞中毒小脑颗粒细胞死亡发生通过凋亡过程，并仅限于颗粒细胞和凋亡小脑颗粒细胞的分布模式是特定的大鼠小脑蚓部。第二，采用Western blot检测细胞凋亡相关分子BDNF、Trk B、Fas配体、RIP、Akt、Bad、Bcl-2、Caspase 9、Caspase 3、CAS和TIAR的表达水平，并采用抗calbindin D的免疫组化染色评价浦肯野细胞的功能。此外，我们还研究了MMC中毒后大鼠脑组织基因表达的变化。这些结果表明 ...更多信息脑源性神经营养因子（BDNF）和Trk B的表达上调，RIP、CAS、Fas配体、Bad和Caspase 9的表达下调。研究结果表明，上调分子的功能，以抑制细胞凋亡在这个阶段。基因表达分析显示，在上调的基因中，1）与细胞凋亡相关的有蛋白激酶C γ、白细胞介素13、胰岛素受体底物Z、亲环素、蛋白磷酸酶1和RL/IF-1，2）与激素相关的有加压素和催产素，3）与酶相关的有血清和糖皮质激素调节激酶、肥大细胞蛋白酶、丝氨酸蛋白酶，糖蛋白特异性UDP-葡糖醛酸转移酶、蛋白酪氨酸磷酸酶、4）7种杂类分子和5）7种未知基因。下调基因为2个α1珠蛋白基因和2个未知基因。结果表明，在小脑细胞凋亡的早期，凋亡抑制基因被激活。因此，可以得出结论，在MMC中毒的初始阶段，抗凋亡分子的表达比凋亡诱导分子更增强，以保护细胞死亡。少