The isolation and measurement of luteinizing hormone-releasing hormone (LHRH) from the rat testis.
The isolation and measurement of luteinizing hormone-releasing hormone (LHRH) from the rat testis.
复制标题
大鼠睾丸中黄体生成素释放激素（LHRH）的分离和测定。
DOI：
10.1016/0303-7207(85)90104-2
复制
发表时间：
1985
影响因子：
4.1
通讯作者：
deKretser,DM
中科院分区：
文献类型：
--
作者：
Hedger,MP;Robertson,DM;Browne,CA;deKretser,DM
虽然LHRH已被牵连在大鼠Leydig细胞功能的直接控制，LHRH尚未在大鼠睾丸中检测到。采用酸提、ODS-硅胶柱层析、氯仿/乙醇提取、乙醚萃取、Sephadex G-15凝胶过滤和反相高效液相色谱法从大鼠睾丸中分离LHRH。LHRH活性通过体外LHRH生物测定系统评估，该系统采用单层培养的大鼠垂体前叶细胞和几种LHRH放射免疫测定法。通过在提取前将外源性LHRH添加到一些冻干睾丸样品中来监测LHRH回收率。在不存在外源性LHRH的情况下，在睾丸提取物的凝胶过滤曲线的LHRH区域中未检测到LHRH生物活性;然而，使用最具特异性的LHRH RIA程序，在所有提取物中均发现LHRH免疫活性，其在RP-HPLC后与LHRH共层析。基于从含有外源性LHRH的提取物中测定的31.0%的平均LHRH回收率，确定提取物中内源性LHRH免疫活性的水平相当于5.6 pg LHRH/g干重或1.0 pg LHRH/睾丸。这些结果表明，成年大鼠睾丸中的LHRH水平明显低于下丘脑。基于这些发现，可以设计一个简化的分馏/测定程序，具有足够的灵敏度，睾丸LHRH的定量作为澄清其在性腺功能中的生理作用的一种手段。
Although LHRH has been implicated in the direct control of rat Leydig cell function, LHRH has not been previously detected in the rat testis. An optimized fractionation procedure, which involved acid extraction, bulk fractionation on ODS-silica, extraction in chloroform/ethanol, ether extraction, gel filtration on Sephadex G-15 and RP-HPLC, was employed to isolate LHRH from lyophilized adult rat testes. LHRH activity was assessed by an in vitro LHRH bioassay system employing rat anterior pituitary cells in monolayer culture and several radioimmunoassays for LHRH. LHRH recoveries were monitored by the addition of exogenous LHRH to some samples of lyophilized testes prior to extraction. LHRH bioactivity was non-detectable in the LHRH region of the gel filtration profile of testis extracts in the absence of exogenous LHRH; however, using the most specific LHRH RIA procedure, LHRH immunoactivity which co-chromatographed with LHRH following RP-HPLC was found in all extracts. Based on an average LHRH recovery of 31.0%, as determined from the extracts containing exogenous LHRH, the level of endogenous LHRH immunoactivity in the extracts was determined to be the equivalent to 5.6 pg LHRH/g dry weight or 1.0 pg LHRH/testis. These results indicate that the levels of LHRH in the adult rat testis are considerably less than those of the hypothalamus. Based on these findings a simplified fractionation/assay procedure with sufficient sensitivity can be devised for the quantitation of testicular LHRH as a means of clarifying its physiological role in gonadal function.