Activation of muscle satellite cells in single-fiber cultures.
Satellite stem cell activation is the process by which quiescent precursor cells resident on muscle fibers are recruited to cycle and move. Two processes are reported to affect satellite cell activation. In vivo, nitric oxide (NO) produced by NO synthase in fibers (NOS-Imu) promotes activation. In cell cultures, hepatocyte growth factor (HGF) is the major activating factor isolated from crushed muscle extract (CME). In this study we hypothesized that distinct and possibly related events were mediated by NO and HGF during activation. Intact fibers were cultured in the presence of bromodeoxyuridine (BrdU) to label DNA synthesis over 48 h. Experiments were designed to test the effects of CME, HGF, a NOS substrate L-arginine, and the NOS inhibitor L-NAME on activation, determined as the number of BrdU-positive satellite cells per fiber. Activation was increased significantly by CME, HGF, and L-arginine. L-Arginine increased activation in a dose-response manner. CME-induced activation was reduced significantly by NOS inhibition. Exposure to marcaine (10 min) caused reversible membrane damage without hypercontraction, as shown by characterizing the sarcolemmal integrity. The resulting decrease in satellite cell activation could be overcome by exogenous HGF. Results support the hypothesis that NO is involved in recruiting to cycle those satellite cells resident on fibers. Separate assessments of resident and free muscle cells showed that HGF and NO also participate in mobilizing satellite cells. Since HGF counteracted NOS inhibition and marcaine-induced membrane damage, data suggest that NO may mediate early steps in activation and precede HGF-mediated events.
Anderson
Judy
J
Department of Human Anatomy and Cell Science, University of Manitoba, 730 William Avenue, Winnipeg, Manitoba, Canada R3E 0W3. janders@ms.umanitoba.ca
Pilipowicz
Orest
O
D000818:Animals; D001120:Arginine / Q000494:pharmacology; D018929:Cell Culture Techniques / Q000379:methods; D002457:Cell Extracts / Q000494:pharmacology; D004247:DNA / Q000096:biosynthesis; D004791:Enzyme Inhibitors / Q000494:pharmacology; D017228:Hepatocyte Growth Factor / Q000494:pharmacology; D051379:Mice; D008810:Mice, Inbred C57BL; D018485:Muscle Fibers, Skeletal / Q000166:cytology* / Q000187:drug effects; D009569:Nitric Oxide / Q000502:physiology; D019001:Nitric Oxide Synthase / Q000037:antagonists & inhibitors / Q000378:metabolism; D032496:Satellite Cells, Skeletal Muscle / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*
D002457:Cell Extracts; D004791:Enzyme Inhibitors; D009569:Nitric Oxide; D017228:Hepatocyte Growth Factor; D004247:DNA; D001120:Arginine; D019001:Nitric Oxide Synthase