Desmin is present in proliferating rat muscle satellite cells but not in bovine muscle satellite cells

Abstract

The presence of desmin was characterized in cultured rat and bovine satellite cells and its potential usefulness as a marker for identifying satellite cells in vitro was evaluated. In primary cultures, positive immunohistochemical staining for desmin and skeletal muscle myosin was observed in rat and bovine myotubes. A small number of mononucleated cells (20% of rat satellite cells and 5% of bovine satellite cells) were myosin‐positive, indicative of post‐mitotic differentiated myocytes. In bovine satellite cell cultures 13% of the mononucleated cells were desmin‐positive, while 84% of the mononucleated cells in rat satellite cell cultures were desmin‐positive. Rat satellite cell mass cultures and bovine satellite cell clonal density cultures were pulsed with ³H‐thymidine, and autoradiographic data revealed that >94% of dividing rat cells were desmin‐positive, suggesting that desmin is synthesized in proliferating rat satellite cells. However, no desmin was seen in cells that incorporated labeled thymidine in bovine satellite cell clones. Analysis of clonal density cultures revealed that only 14% of the mononucleated cells in bovine satellite cell colonies were desmin‐positive, whereas 98% of the cells in rat satellite cell colonies were desmin‐positive. Fibroblast colonies from both species were desmin‐negative. In order to further examine the relationship between satellite cell differentiation and desmin expression, 5‐bromo‐2′‐deoxyuridine (BrdU) was added to culture medium at the time of plating to inhibit differentiation. Fusion was inhibited in rat and bovine cultures, and cells continued to divide. Very few desmin‐positive cells were found in bovine cultures, but greater than 90% of the cells in rat cultures stained positive for desmin. The presence of desmin and sarcomeric myosin was also evaluated in regenerating rat tibialis anterior five days after bupivicaine injection. In regenerating areas of the muscle many desmin‐positive cells were present, and only a few cells stained positive for skeletal muscle myosin. Application of desmin staining to rat satellite cell growth assays indicated that rat satellite cells cultured in serum‐containing medium were contaminated with fibroblasts at levels that ranged from approximately 5% in 24 hr cultures to 15% in mature cultures. In defined medium 4 day cultures contain approximately 95% to 98% desmin‐positive satellite cells. The effects of combinations of insulin‐like growth factor I (IGF‐I), basic fibroblast growth factor (bFGF), and transforming growth factor beta (TGF‐β) on rat satellite cell proliferation and differentiation were assessed by desmin staining, and results were found to be consistent with results obtained previously using conventional cell staining and counting techniques (Allen and Boxhorn, 1989). Our experiments indicate that the pattern of desmin expression in satellite cells differs between rat and bovine and that desmin can be a useful marker for cultured rat satellite cells.