Influence of temperature on spermatogenesis-related genes in spermatogonia cultured in vitro / 中华男科学杂志

ABSTRACT

<p><b>OBJECTIVE</b>To investigate the influence of temperature on the expressions of c-kit and PI3K in spermatogonia cultured in vitro at 32 degrees C and 37 degrees C, and provide basic scientific data for the mechanism of spermatogenic impairment due to body temperature (37 degrees C).</p><p><b>METHODS</b>Isolated spermatogenic cells were cultured in vitro at 32 degrees C and 37 degrees C, and their adherence, proliferation and morphologic changes were observed and recorded under the inverted phase contrast microscope. At 8 days, the spermatogonia were separated by Percoll density gradient centrifugation and the differential adhesion method. The expressions of c-kit and PI3K mRNA and proteins in the separated cells were detected by real time polymerase chain reaction and Western blot, respectively. The c-kit gene was sequenced to identify the occurrence of mutations.</p><p><b>RESULTS</b>Adherence, division and proliferation of the cells were observed in both the 32 degrees C and 37 degrees C groups. The expressions of c-kit and PI3K mRNA and proteins in the spermatogonia were significantly higher in the 32 degrees C group than in the 37 degrees C group (P < 0.05). The 32 degrees C group showed no mutation of c-kit in exon 9, 11 and 13; the 37 degrees C group exhibited no mutation in exon 11 and 13, but possible insertion or deletion mutations in exon 9.</p><p><b>CONCLUSION</b>Culturing in vitro at 37 degrees C could inhibit the expression of proliferation- and differentiation-related genes in spermatogenic cells and lead to the mutation of the c-kit gene.</p>