Stress response of the rat testis: In situ hybridization and immunohistochemical analysis of heme oxygenase-1 (HSP32) induction by hyperthermia
Stress response of the rat testis: In situ hybridization and immunohistochemical analysis of heme oxygenase-1 (HSP32) induction by hyperthermia
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DOI：
10.1095/biolreprod54.5.1070
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发表时间：
1996-05-01
影响因子：
3.6
通讯作者：
Ewing, JF
中科院分区：
文献类型：
--
作者：
Maines, MD;Ewing, JF
采用原位杂交和免疫组织化学方法，研究了热应激条件下血红素氧合酶（HO）-1（HSP 32）转录本和蛋白质的分布规律及其对热应激的反应。用地高辛标记的HO-1cDNA探针和多克隆抗体检测了正常和热休克睾丸组织中HO-1 mRNA和蛋白的表达水平。地高辛标记的探针在睾丸间质细胞、睾丸支持细胞、精原细胞、初级精母细胞和生精小管精子细胞中检测到HO-1的强转录信号。在所有细胞类型中，转录本主要以确定的模式集中在细胞核中。热应激（42 ℃，20分钟）没有改变HO-1转录表达的细胞群体模式;然而，它确实引起细胞核模式的扭曲和转录信号的扩散。大鼠的高温处理导致睾丸1.8 kb HO-1 mRNA适度（2- 2.8倍），时间依赖性和持续（1-16 h）的增加。睾丸HO-1表达的正常和热休克模式的免疫组织化学分析显示，热休克后支持细胞和间质细胞染色强烈;在正常组织中，这些细胞群的免疫反应性低。与转录本的分布，高温并没有影响HO-1免疫反应的模式，和蛋白质未检测到在生精细胞在控制或应力条件下。在Leydig细胞中，高温导致HO-1蛋白质染色强度增加8倍以上。与HO-1的选择性表达相一致，热休克后6 h，在总睾丸微粒体中检测到的单个HO-1免疫反应蛋白（类似于32 kDa）的水平显示出适度（1.5倍）的增加。数据是一致的建议，HO-1蛋白在生殖细胞系和Sertoli细胞中的差异分布反映了这些细胞类型中的差异HO-1 mRNA加工。增加可能是必不可少的催化变性血红素蛋白，如细胞色素P450和血红蛋白血红素血红素部分，因此可以防止血红素催化的自由基形成。我们认为热应激条件下支持细胞和间质细胞中HO-1蛋白的诱导可能起到保护生精细胞的作用。
By using in situ hybridization and immunohistochemistry, the distribution patterns of heme oxygenase (HO)-1 (HSP32) transcript and protein were studied, and their response to thermal stress was examined. And, by using an HO-1 cDNA probe and polyclonal antibody, the levels of HO-1 mRNA and protein in normal and heat-shocked testis were quantified, The digoxigenin-labeled probe detected a strong signal for HO-1 transcript in Leydig cells, and in the Sertoli cells, spermatogonia, primary spermatocytes, and spermatids of the seminiferous tubules. In all cell types, the transcript was predominantly concentrated in the nucleus in a defined pattern. Thermal stress (42 degrees C, 20 min) did not change the cell population pattern of HO-1 transcript expression; however, it did cause distortion of the nuclear pattern and diffusion of the transcript signal in cells, Hyperthermic treatment of rats resulted in a modest (2- to 2.8-fold), time-dependent, and sustained (1-16 h) increase in testicular 1.8-kb HO-1 mRNA. Immunohistochemical analysis of normal and heat shock patterns of testicular HO-1 expression showed robust staining of Sertoli and Leydig cells after heat shock; in normal tissue, immunoreactivity was low in these cell populations. As with the transcript distribution, hyperthermia did not affect the pattern of HO-1 immunoreactivity, and the protein was not detected in spermatogenic cells under control or stress conditions. In the Leydig cells, hyperthermia led to a more than 8-fold increase in the intensity of cytoplasmic staining for HO-1 protein. Consistent with the selective expression of HO-1, the level of the single HO-1 immunoreactive protein (similar to 32 kDa) detected in total testis microsomes showed a modest (1.5-fold) increase 6 h after heat shock. Data are consistent with the suggestion that differential distribution of HO-1 protein in the germ cell line and Sertoli cells reflects differential HO-1 mRNA processing in these cell types. The increase may be essential for the catalysis of the heme moiety of denatured hemoproteins such as cytochrome P450 and hemoglobin heme and hence may protect against heme-catalyzed free radical formation. We propose that induction of HO-1 protein in Sertoli and Leydig cells may function to protect the spermatogenic cells under conditions of thermal stress.