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Establishment of culture model of seminiferous tubules treated by ethane dimethanesulfonate in vitro

ZHANG Lei1,3, ZHANG Qi-hao2,3, SU Zhi-jian2,3, HUANG Ya-dong2,3   

  1. 1.Guangdong Provincial Institute of Biological Products and Materia Medica， Guangzhou 510440， China； 2.Department of Cell Biology, Jinan University, Guangzhou 510632, China; 3.Biopharmaceutical Research and Development Center, Jinan University， Guangzhou 510632, China
  • Online:2015-03-28 Published:2015-03-26
  • Supported by:

    National Natural Science Foundation of China, 81070477, 31000663, 31271607； Fundamental Research Funds for the Central Universities of China, 21611378； Guangdong Province Universities and Colleges Pearl River Scholar Funded Scheme 2012

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Abstract:

Objective To establish the culture model of seminiferous tubules treated by ethane dimethanesulfonate (EDS) in vitro. Methods Male SD rats were intraperitioneally injected with EDS 7 d before experiment. Seminiferous tubules of rats were removed and cultured in vitro, and divided into the basic treatment group [treated by insulin-transferin-selenium (ITS), i.e. the ITS group] and luteinizing hormone (LH) treatment group (treated by ITS and LH, i.e. the ITS+ LH group). EdU staining was adopted to detect the proliferation of stem Leydig cells. Radioimmunoassay and 3β-HSD staining were employed to detect the secretion of testosterone and differentiation of stem Leydig cells. Seminiferous tubules were isolated from testicles of adult male mouse and treated by EDS of different concentrations. Then seminiferous tubules were treated by ITS (ITS group) and ITS+LH (ITS+ LH group). The secretion of testosterone of culture supernatant was detected by the radioimmunoassay. Results Compared with the ITS group, the proliferation of stem Leydig cells of seminiferous tubules of the ITS+LH group was more significant. The secretion of testosterone was consistent with the development and differentiation of Leydig cells. The production of testosterone of the ITS+LH group was large and the duration was long after being treated by LH, which showed that the culture model of seminiferous tubules was sensitive to LH. EDS of 1.72 mmol/L effectively killed mature Leydig cells of seminiferous tubules. Testosterone was produced after the model was cultured and the testosterone level of culture supernatant of seminiferous tubules of the ITS+LH group was significantly higher than that of the ITS group (P<0.01), which indicated that EDS could kill mature Leydig cells and the model was also sensitive to LH. Conclusion The development and differentiation processes of stem Leydig cells of the male rat and mouse models and the culture model of seminiferous tubules in vitro are similar. The culture model is suitable for studying the development and differentiation of Leydig cells.

Key words: seminiferous tubules, Leydig cells, ethane dimethanesulphonate, luteinizing hormone