Contribution of calpains to photoreceptor cell death in N-methyl-N-nitrosourea-treated rats
Contribution of calpains to photoreceptor cell death in N-methyl-N-nitrosourea-treated rats
复制标题
DOI：
10.1016/j.expneurol.2006.09.011
复制
发表时间：
2007-03-01
影响因子：
5.3
通讯作者：
Azuma, Mitsuyoshi
中科院分区：
文献类型：
--
作者：
Oka, Takayuki;Nakajima, Takeshi;Azuma, Mitsuyoshi
本研究的目的是在腹腔注射n -甲基-n -亚硝基脲（MNU）诱导的大鼠光感受器变性模型中，确定钙依赖性酶calpain （EC 3.4.22.17）的蛋白水解是否导致视网膜细胞死亡。H&E染色观察视网膜变性，TUNEL法观察细胞死亡。用原子吸收分光光度法测定视网膜总钙含量。酪蛋白酶谱法和免疫印迹法测定钙蛋白酶的活化。免疫印迹法检测a-spectrin和p35 （Cdk5的调节因子）的蛋白水解情况。将Calpain抑制剂SNJ-1945口服mn - u处理大鼠，观察其药效。MNU降低了由外核层（ONL）和外段（OS）组成的感光细胞层的厚度。ONL中大量细胞TUNEL染色阳性。MNU后视网膜总钙含量升高。注射MNU后观察calpain的活化和a-spectrin的特异性蛋白水解。口服SNJ-1945给药m - nu处理的大鼠显示出对光感受器细胞丧失的显著保护作用，证实钙蛋白酶参与光感受器变性。p35向p25的转化与calpain的激活密切相关，提示Cdk5/p25的长时间激活可能是mnu诱导的光感受器细胞死亡的下游机制。SNJ-1945减少了光感受器细胞的死亡，尽管MNU是光感受器细胞退化最严重的模型之一。口服钙蛋白酶抑制剂SNJ-1945可能作为抗视网膜色素变性的候选药物进行试验。(c) 2006爱思唯尔公司版权所有。
The purpose of the present study was to determine if proteolysis by the calcium-dependent enzyme calpains (EC 3.4.22.17) contributed to retinal cell death in a rat model of photoreceptor degeneration induced by intraperitoneal injection of N-methyl-N-nitrosourea (MNU). Retinal degeneration was evaluated by H&E staining, and cell death was determined by TUNEL assay. Total calcium in retina was measured by atomic absorption spectrophotometry. Activation of calpains was determined by casein zymography and immunoblotting. Proteolysis of a-spectrin and p35 (regulator of Cdk5) were evaluated by immunoblotting. Calpain inhibitor SNJ-1945 was Orally administrated to MN-U-treated rats to test drug efficacy. MNU decreased the thickness of photoreceptor cell layer, composed of the outer nuclear layer (ONL) and outer segment (OS). Numerous cells in the ONL showed positive TUNEL staining. Total calcium was increased in retina after MNU. Activation of calpains and calpain-specific proteolysis of a-spectrin were observed after MNU injection. Oral administration of SNJ-1945 to M-NU-treated rats showed a significant protective effect against photoreceptor cell loss, confirming involvement of calpains in photoreceptor degeneration. Conversion of p35 to p25 was well correlated with calpain activation, suggesting prolonged activation of Cdk5/p25 as a possible downstream mechanism for MNU-induced photoreceptor cell death. SNJ-1945 reduced photoreceptor cells death, even though MNU is one of the most severe models of photoreceptor cell degeneration. Oral calpain inhibitor SNJ-1945 may be a candidate for testing as a medication against retinal degeneration in retinitis pigmentosa. (c) 2006 Elsevier Inc. All rights reserved.