INTRACELLULAR-LOCALIZATION OF THE TESTICULAR AND SPERM-SPECIFIC LACTATE-DEHYDROGENASE ISOZYME C-4 IN MICE
INTRACELLULAR-LOCALIZATION OF THE TESTICULAR AND SPERM-SPECIFIC LACTATE-DEHYDROGENASE ISOZYME C-4 IN MICE
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DOI：
10.1095/biolreprod53.1.84
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发表时间：
1995-07-01
影响因子：
3.6
通讯作者：
BLANCO, A
中科院分区：
文献类型：
--
作者：
BURGOS, C;MALDONADO, C;BLANCO, A
提出的精子特异性乳酸脱氢酶（EC 1.1.1.27）同工酶C-4（LDH C-4）的双重细胞内分布是基于间接证据。为了获得直接的证据，在小鼠的LDH同工酶的本地化，postembedding免疫细胞化学在超微结构水平上进行睾丸，附睾精子，和分离的睾丸线粒体。免疫金技术应用于薄切片孵育的第一部分纯化的特异性抗LDH C-4兔IgG，和免疫反应位点检测与胶体金吸附到抗兔IgG。在精巢中，免疫组化染色见于精母细胞和精子细胞的细胞质以及分化中的精子的主段和中段。附睾精子也表现出重标记的胶体金在他们的中间和主要部分，但免疫染色弱的特殊类型的线粒体存在于精母细胞，精子细胞和精子（精子型线粒体，STM）。STM的分离与原位分离相比产生了一些形态学变化，包括线粒体基质中LDH C-4标记的增强，来自精母细胞和非生精细胞的另一种类型的线粒体（非STM）未被免疫染色并作为背景对照。表明LDH C-4在精母细胞、精子细胞和精子的胞质溶胶中以及在精子型线粒体的基质中的双重定位。
The proposed dual intracellular distribution of the sperm-specific lactate dehydrogenase (EC 1.1.1.27) isozyme C-4 (LDH C-4) has been based on indirect evidence. In order to obtain direct evidence on the localization of this LDH isozyme in mice, postembedding immunocytochemistry at ultrastructural level was performed on testes, epididymal spermatozoa, and isolated testicular mitochondria. The immunogold technique was applied to thin sections incubated first in partially purified specific anti-LDH C-4 rabbit IgG, and immunoreactive sites were detected with colloidal gold adsorbed to anti-rabbit IgG. In the testis, immunostaining was found in the cytoplasm of spermatocytes and spermatids and in the principal and middle pieces of differentiating spermatozoa. Spermatozoa from epididymis also exhibited heavy labeling of colloidal gold in their middle and principal pieces, but the immunostaining was weak in the special type of mitochondria present in spermatocytes, spermatids, and spermatozoa (sperm-type mitochondria, STM). The isolation of STM produced several morphological changes in comparison with those in situ, including an enhancement of the LDH C-4 labeling in the mitochondrial matrix, The other type of mitochondria (non-STM) from spermatocytes and nonspermatogenic cells were not immunostained and served as background control.The results presented here confirm previous findings, gathered by indirect methods, indicating a dual localization of LDH C-4 in the cytosol of spermatocytes, spermatids, and spermatozoa, as well as in the matrix of sperm-type mitochondria.