In vitro proliferation of rat Leydig cells / 中华男科学杂志

ABSTRACT

<p><b>OBJECTIVE</b>To investigate the feasibility of in vitro proliferation of rat Leydig cells by modifying the cell culture system.</p><p><b>METHODS</b>Leydig cells were isolated from three-week-old rats by a procedure combining collagenase dispersion, stainless steel mesh infiltration and differential adhesion. The isolated cells were cultured in DMEM/F12 and modified media for stem cell proliferation, and the proliferation of the cultured cells was evaluated by cell counting and MTP test. The expression of 3beta-HSD in the cultured cells was detected by immunohistochemistry and flow cytometry, and testosterone productivity in the isolated Leydig cells with or without hCG stimulation was determined at 2 hours and 4 days after cell isolation.</p><p><b>RESULTS</b>The Leydig cells cultured in the modified media proliferated actively, with a doubling time of (2.26 +/- .31) days, as compared with (16.32 +/- 2.14) days for those cultured in the traditional media (P <0.05). The 3beta-HSD positive rate in the cultured cells was (554.3 +/- 7.1)% after 2 hours and (93.6 +/- 4.6)% after 4 days of culture. All the proliferated cells exhibited testosterone productivity, and their testosterone secretion was significantly upregulated by hCG stimulation (P <0.05).</p><p><b>CONCLUSION</b>Leydig cells isolated by differential adhesion proliferate actively in the modified culture media.</p>