去势小鼠血清和脑组织中抑制素B的含量及分布变化

Expression and distribution of inhibin B in serum and brain of orchidectomized mice

目的分析去势小鼠血清和脑组织中抑制素B(INH-B)水平变化及INH-B亚基在脑组织中的分布。方法将4周龄去势小鼠(睾丸切除,去势组)及正常(对照组)雄性C57BL/6小鼠各12只分别喂养至8周龄(去势后4周)和12周龄(去势后8周)后,收集每只小鼠血清约50μl,并用10%水合氯醛麻醉后开胸进行生理盐水及4%多聚甲醛灌注,然后分离脑组织;仅生理盐水灌注者在4°C条件下收集组织匀浆上清,而继续4%多聚甲醛灌注者将脑组织置于4%多聚甲醛溶液中继续固定保存,石蜡包埋切片。ELISA方法检测两组小鼠血清及脑组织匀浆上清中INH-B含量,免疫荧光方法检测INH-B亚基在脑组织中的定位分布。结果ELISA结果显示,去势组小鼠血清及脑组织匀浆上清中INH-B含量均显著低于对照组(P<0.05),尤其血清中INH-B水平低于检测下限值(<4 pg/ml),但同组小鼠在8周和12周两个时间点的水平变化均无显著性差异(P>0.05)。免疫荧光实验结果显示,去势后小鼠脑组织中INH-Bα亚基和β亚基的信号强度明显减弱。结论去势小鼠体内INH-B维持一个极低水平的状态,提示雄性小鼠INH-B主要来源于睾丸,也可能存在着睾丸外来源。

Objective:To analyze the effect of orchidectomy on concentrations of inhibin B level in serum and brain and distribution of the inhibin B subunits in brain of male mice.Methods:Four-week-old male C57BL/6 mice were divided to orchiectomy group and control group,12 mice for each group.The rats were fed to 8 weeks of age(4 weeks after castration)and 12 weeks of age(8 weeks after castration)respectively.The 50μl of serum was collected from each mouse,and the chest was opened after anesthetized with 10%chloral hydrate and normal saline and 4%paraformaldehyde were perfused,and then the brain tissues were separated.The tissue homogenate supernatant was collected at 4°C for the mice infused with saline only,while the brain tissues were placed in 4%paraformaldehyde solution for continuous preservation for the mice which continued to perfuse with 4%paraformaldehyde followed by paraffin embedding and sectioning.Concentrations of INH-B in the serum and supernatant of brain tissues were detected by Elisa method,and immunofluorescence method was used to detect the location and distribution of INH-B subunits in brain tissue.Results:Concentrations of INH-B in serum and brain of orchidectomized mice were significantly lower than that of control mice(P<0.05).In particular,the INH-B level in the serum was lower than the minimal detection limit(<4 pg/ml),but there was no significant difference in the changes at 8 and 12 weeks in the mice of same group(P>0.05).The results of immunofluorescence experiments showed that the signal intensity ofαandβsubunits of INH-B in the brain tissue of mice was significantly weakened after castration.The results of immunofluorescence showed that the signal intensity ofαandβsubunits of INH-B in the brain of orchidectomized mice decreased significantly.Conclusions:INH-B in orchidectomized mice maintained a very low level,indicating thatαandβsubunits of INH-B mainly originated from the testis in male mice.There may also be an extra-testicular source.