Hayashi Tetsuo, Yoshida Soichiro, Yoshinaga Atsushi, Ohno Rena, Ishii Nobuyuki, Yamada Takumi
Department of Urology, Saitama Medical Center, Saitama Medical School, Kawagoe, Saitama, Japan.
Int J Urol. 2006 Feb;13(2):157-64. doi: 10.1111/j.1442-2042.2006.01250.x.
The aim of the present study was to elucidate the role of high temperature requirement A2 (HtrA2) in germ cell loss in the heat-stressed testis.
We examined the expression of HtrA2, caspase-9 activity and proteolytic activity of HtrA2 in the rat testis, and their in vivo responses to experimental cryptorchid treatment.
Northern analysis revealed the expression of HtrA2 mRNA peaked at days 1 and 7 after cryptorchid treatment. While expression of HtrA2 mRNA was seen in the spermatogonium, spermatocytes and some spermatids in normal adult rat testis, experimental cryptorchidism treatment resulted in a marked increase in its signal intensity in spermatocytes and some spermatids, and the layers of spermatogonium and early primary spermatocytes became negative at days 1 and 7 after the treatment. However, the spermatogonium, Sertoli cells and interstitial cells appeared to have strong intensities at days 14, 28 and 56 after the treatment. Western analysis revealed the expression of HtrA2 protein peaked at day 2 coinciding with the increase of positive spermatogonium, the appearance of protein-positive interstitial cells, and day 28 coinciding with the reappearance of protein-positive interstitial cells. Caspase-9 activity peaked at day 2 and HtrA2 proteolytic activity peaked at day 28. Consequently, the first peak of HtrA2 mRNA expression was followed by the peak of caspase-9 activity and the second peak was followed by the peak of proteolytic activity; however, the second peak of mRNA expression had considerable chronological difference from that of the protein.
These findings suggest the probabilities that the heat stress results in germ cell death by a caspase-independent manner with the elevation of HtrA2 proteolytic activity, as well as a caspase-dependent manner with the elevation of caspase-9 activity.
本研究旨在阐明高温需求蛋白A2（HtrA2）在热应激睾丸生殖细胞丢失中的作用。
我们检测了大鼠睾丸中HtrA2的表达、半胱天冬酶-9活性及HtrA2的蛋白水解活性，以及它们对实验性隐睾治疗的体内反应。
Northern分析显示，隐睾治疗后第1天和第7天，HtrA2 mRNA的表达达到峰值。在正常成年大鼠睾丸的精原细胞、精母细胞和一些精子细胞中可见HtrA2 mRNA的表达，而实验性隐睾治疗导致精母细胞和一些精子细胞中其信号强度显著增加，且治疗后第1天和第7天，精原细胞层和早期初级精母细胞变为阴性。然而，治疗后第14天、28天和56天，精原细胞、支持细胞和间质细胞的信号强度似乎较强。Western分析显示，HtrA2蛋白的表达在第2天达到峰值，此时阳性精原细胞增加，蛋白阳性间质细胞出现；在第28天达到峰值，此时蛋白阳性间质细胞再次出现。半胱天冬酶-9活性在第2天达到峰值，HtrA2蛋白水解活性在第28天达到峰值。因此，HtrA2 mRNA表达的第一个峰值之后是半胱天冬酶-9活性的峰值，第二个峰值之后是蛋白水解活性的峰值；然而，mRNA表达的第二个峰值与蛋白的峰值在时间上有相当大的差异。
这些发现提示，热应激可能通过HtrA2蛋白水解活性升高导致的非半胱天冬酶依赖性方式以及半胱天冬酶-9活性升高导致的半胱天冬酶依赖性方式引起生殖细胞死亡。
