ABSTRACT
<p><b>OBJECTIVE</b>To study effect of electrical stimulations of different intensities on mitochondrial oxidative stress in C2C12 myotubes and explore the molecular mechanisms.</p><p><b>METHODS</b>After 7 days of differentiation, C2C12 myotubes were subjected to electrical stimulations (15 V, 3Hz, 30 ms) for 60, 120, or 180 min, and the morphological changes of muscular tubes were observed under inverted microscope. The levels of MDA and SOD activity of the cells were detected, and flow cytometry was used to detect mitochondrial reactive oxygen species (ROS) and membrane potential. Western blotting was used to detect the expression of PGC1, AMPK-Ser485, AMPK-Thr172, and AMPK in the cells.</p><p><b>RESULTS</b>No significant changes occurred in the morphology of C2C12 myotubes in response to electrical stimulations. Electrical stimulation for 60 min resulted in significantly increased levels of MDA, AMPK-Ser485 and AMPK-Thr172 in the cells (P<0.05); simulations of the cells for 120 and 180 min caused significantly increased MDA, ROS, mitochondrial ROS, AMPK-Ser485 and PGC1 along with marked reduction of mitochondrial membrane potential (P<0.05).</p><p><b>CONCLUSION</b>Electrical stimulation significantly activates oxidative stress, and a longer stimulation time causes stronger mitochondrial oxidation. AMPK-Thr172 regulates oxidative stress induced by stimulations for a moderate time length, while AMPK-Ser485 and PGC1 function to modulate oxidative stress following prolonged stimulations.</p>
ABSTRACT
<p><b>OBJECTIVE</b>To investigate the role of Rictor/mTORC2 in the formation of blood testis barrier (BTB), testicular development, and spermatogenesis.</p><p><b>METHODS</b>Amh Cre positive mice homozygous for rictor loxP with Sertoli cell specific deletion of rictor were obtained by cross breeding Amh Cre mice with rictor loxP mice. The histology of the reproductive organs, seminiferous tubules and epididymis of the transgenic mice was observed with HE staining. The cell subgroups of the germ cells in the seminiferous tubule were detected by flow cytometry with propidium iodide labeling. The expression levels of Ki 67 and separase were detected with immunofluorescence assay, and the expression levels of BTB associated proteins were detected with immunofluorescence and Western blotting.</p><p><b>RESULTS</b>Compared with the control (Amh Cre, rictoror rictor) mice, the mice with Sertoli cell specific rictor deletion showed significantly decreased testicular weight and epididymis weight (P<0.05), significantly increased diploid cells (P<0.01), and decreased haploid cells (P<0.01) but comparable tetraploid cells and similar expression levels of Ki 67 and separase. The mice with rictor knockout also showed aberrant localization of BTB associated proteins, which were scattered over the whole seminiferous epithelium, but the expression levels of the protein remained stable.</p><p><b>CONCLUSION</b>Rictor in testicular Sertoli cells is essential for maintaining BTB integrity and function and ensuring normal spermatogenesis in mice.</p>