Jia Dan, Zheng Weijiang, Jiang Honglin
Department of Animal and Poultry Sciences, Virginia Tech, Blacksburg, VA, United States.
Department of Animal and Poultry Sciences, Virginia Tech, Blacksburg, VA, United States; College of Animal Sciences, Nanjing Agricultural University, Nanjing, China.
Growth Horm IGF Res. 2018 Jun;40:9-16. doi: 10.1016/j.ghir.2018.03.006. Epub 2018 Mar 28.
The C3H10T1/2 cells are considered mesenchymal stem cells (MSCs) because they can be induced to become the progenitor cells for myocytes, adipocytes, osteoblasts, and chondrocytes by the DNA methyltransferase inhibitor 5'-azacytidine. In this study, we determined the effect of growth hormone (GH) on the myogenic and adipogenic lineage commitment in C3H10T1/2 cells. The C3H10T1/2 cells were treated with recombinant bovine GH in the presence or absence of 5'-azacytidine for 4 days. The myogenic commitment in C3H10T1/2 cells was assessed by immunostaining them for MyoD, the marker for myoblasts, and by determining their capacity to differentiate into the multinucleated myotubes. The adipogenic commitment in C3H10T1/2 cells was assessed by determining their ability to differentiate into adipocytes. Myotubes and adipocyteswere identified by immunocytochemistry and Oil Red O staining, respectively. C3H10T1/2 cells treated with 5'-azacytidine and GH for 4 days contained a greater percentage of MyoD-positive cells than those treated with 5'-axacytidine alone (P < 0.05). The former generated more myotubes than the latter upon induced myoblast differentiation (P < 0.05). However, C3H10T1/2 cells treated with GH alone did not form any myotubes. C3H10T1/2 cells treated with 5'-azacytidine formed adipocytes upon adipocyte differentiation induction, whereas C3H10T1/2 cells treated with GH alone did not form any adipocytes. C3H10T1/2 cells treated with both 5'-azacytidine and GH formed fewer adipocytes than those treated with 5'-azacytidine alone (P < 0.05). Both GHR and IGF-I mRNA expression in C3H10T1/2 cells were increased by 5'-azacytidine (P < 0.05), but neither was affected by GH. Overall, this study showed that GH enhanced 5'-azacytidine-induced commitment in C3H10T1/2 cells to myoblasts but inhibited 5'-azacytidine-induced commitment to preadipocytes. These results support the possibility that GH stimulates skeletal muscle growth and inhibits adipose tissue growth in part by stimulating the myogenic commitment and inhibiting the adipogenic commitment, respectively, in mesenchymal stem cells.
C3H10T1/2细胞被认为是间充质干细胞（MSCs），因为它们可被DNA甲基转移酶抑制剂5'-氮杂胞苷诱导成为肌细胞、脂肪细胞、成骨细胞和软骨细胞的祖细胞。在本研究中，我们确定了生长激素（GH）对C3H10T1/2细胞成肌和成脂谱系定向分化的影响。在存在或不存在5'-氮杂胞苷的情况下，用重组牛GH处理C3H10T1/2细胞4天。通过对成肌细胞标志物MyoD进行免疫染色以及测定它们分化为多核肌管的能力，来评估C3H10T1/2细胞的成肌定向分化。通过测定C3H10T1/2细胞分化为脂肪细胞的能力，来评估其成脂定向分化。分别通过免疫细胞化学和油红O染色鉴定肌管和脂肪细胞。用5'-氮杂胞苷和GH处理4天的C3H10T1/2细胞中，MyoD阳性细胞的百分比高于仅用5'-氮杂胞苷处理的细胞（P<0.05）。在诱导成肌细胞分化后，前者产生的肌管比后者更多（P<0.05）。然而，单独用GH处理的C3H10T1/2细胞未形成任何肌管。用5'-氮杂胞苷处理的C3H10T1/2细胞在诱导脂肪细胞分化时形成了脂肪细胞，而单独用GH处理的C3H10T1/2细胞未形成任何脂肪细胞。用5'-氮杂胞苷和GH共同处理的C3H10T1/2细胞形成的脂肪细胞比仅用5'-氮杂胞苷处理的细胞少（P<0.05）。5'-氮杂胞苷可增加C3H10T1/2细胞中GHR和IGF-I mRNA的表达（P<0.05），但两者均不受GH影响。总体而言，本研究表明，GH增强了5'-氮杂胞苷诱导的C3H10T1/2细胞向成肌细胞的定向分化，但抑制了5'-氮杂胞苷诱导的向脂肪前体细胞的定向分化。这些结果支持了GH可能分别通过刺激间充质干细胞的成肌定向分化和抑制成脂定向分化，来刺激骨骼肌生长和抑制脂肪组织生长的可能性。
