Viability of meiotic prophase spermatocytes of rats is facilitated in primary culture of dispersed testicular cells on collagen gel by supplementing epinephrine or norepinephrine: Evidence that meiotic prophase spermatocytes complete meiotic divisions in vitro-猫眼课题宝

通过补充肾上腺素或去甲肾上腺素，在胶原凝胶上分散睾丸细胞的原代培养中促进大鼠减数分裂前期精母细胞的活力：减数分裂前期精母细胞在体外完成减数分裂的证据

DOI：

发表时间：

1989

期刊：

In Vitro Cellular & Developmental Biology

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中科院分区：

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作者：

Y. Nagao

本实验采用含10%胎牛血清的Ham’s F12-Leibovitz’s L15培养基（F12-L15培养基）1∶1混合液，在1型胶原凝胶上培养14日龄大鼠睾丸细胞。生精细胞的活力，促进补充大鼠肾上腺提取物到培养基中。用分子筛高效液相色谱法从肾上腺的酸性提取物中分离纯化了有效物质（存活因子），并鉴定为肾上腺素和去甲肾上腺素。肾上腺素和去甲肾上腺素在10 ng/ml的半饱和剂量下均能促进生精细胞的存活。通过补充生存因子（肾上腺素或去甲肾上腺素），可以在胶原凝胶上培养2周的生精细胞进行Giemsa染色和DNA流式细胞术。结果表明：（1）14日龄大鼠的生精细胞中不含生精细胞（LC细胞）; B）培养2 ~ 7 d，减数分裂前期精母细胞（4c-细胞）与减数分裂前细胞（2c-细胞）的比例增加。在第7天，超过90%的存活细胞是减数分裂前期精母细胞。c）在第10天，精子细胞（LC-细胞）首次出现。精子细胞在体外培养中首次出现的时间与体内的时间一致。这些结果表明，肾上腺素和去甲肾上腺素都促进了减数分裂前期精母细胞的活力，部分减数分裂前期精母细胞在睾丸细胞培养中完成了减数分裂。

SummaryDispersed testicular cells prepared from 14-d-old rats were cultured on type 1 collagen gels using a medium composed of a 1∶1 mixture of Ham’s F12 medium and Leibovitz’s L15 medium (F12-L15 medium) containing 10% (vol/vol) fetal bovine serum. The viability of the spermatogenic cells was facilitated by supplementing a rat adrenal extract into the medium. The effective substance(s) (the survival factor) was purified from acid extracts of adrenals by molecular sieve high performance liquid chromatography and identified as epinephrine and norepinephrine. Both epinephrine and norepinephrine promoted the survival of the spermatogenic cells with a half saturating dose of 10 ng/ml. The spermatogenic cells, which could be cultured for 2 wk on a collagen gel by supplementing with the survival factor (epinephrine or norepinephrine), were subjected to Giemsa staining and to DNA flow cytometry. The following results were obtained: a) The spermatogenic cells from 14-d-old rats did not contain spermiogenic cells (lc-cells). b) During a culture period of 2 to 7 d the ratio of meiotic prophase spermatocytes (4c-cells) to premeiotic cells (2c-cells) increased. On Day 7, more than 90% of the surviving cells were meiotic prophase spermatocytes. c) On Day 10, spermatids (lc-cells) appeared for the first time. The time of the first appearance of spermatids in the culture was consistent with that in vivo. These results suggest that both epinephrine and norepinephrine facilitated the viability of meiotic prophase spermatocytes and that a part of the meiotic prophase spermatocytes completed the meiotic divisions in the testicular cell culture.