Institute of Animal Sciences, Chinese Academy of Agricultural Sciences, Beijing 100193, China.
State Key Laboratory of Animal Nutrition, Beijing 100193, China.
Poult Sci. 2018 Oct 1;97(10):3691-3697. doi: 10.3382/ps/pey023.
Intramuscular fat is important in improving meat quality; however, the lack of high-purity intramuscular preadipocyte (IMP) in vitro has severely limited the in-depth research on the mutual regulation of myocytes and adipocytes in chicken. In this study, we establish a new method by combining the mature adipocyte ceiling method and the transwell co-culture system. Mature intramuscular adipocyte (MIA) and muscle satellite cell (MSC) were obtained from digested pectoralis major, and MIAs were transformed into IMPs by dedifferentiation with ceiling culture. MSCs were then purified by differential adhesion for 2 h. The results by inverted-microscope observation, MTT assay, Oil Red O staining, and q-PCR revealed that the de-differentiated cells from MIA were identified as the IMPs, and had the same the cellular morphology, the capacity on differentiation, proliferation and passage with the abdominal fat preadipocytes (P > 0.05). The applicability of the obtained IMPs in co-cultured experiment with the MSC revealed that it could meet the requirements of the experimental study. Finally, a co-culture system of IMPs and MSCs was established using a transwell chamber. The co-cultured results indicated that MSCs in the proliferative stage tend to accelerate the differentiation of IMPs to induce more fat content in co-cultured IMPs than in single-culture IMPs (P < 0.05), in the non-proliferative stage, the results tend to show the opposite (P < 0.05). The mRNA levels of related genes significantly changed in accordance with the fat content in cells. The results strongly supported the view that the established co-culture system was effective and feasible. In summary, we successfully found a new method to explore the interaction between myocytes and adipocytes of chicken. Our findings can deepen research on the regulation of chicken myocytes and adipocytes.
肌内脂肪在改善肉质方面很重要；然而，由于缺乏高纯度的肌内前体脂肪细胞（IMP），鸡的肌细胞和脂肪细胞之间的相互调节机制的深入研究受到了严重限制。在本研究中，我们结合成熟脂肪细胞天花板培养法和 Transwell 共培养系统建立了一种新的方法。从消化的胸大肌中获得成熟的肌内脂肪细胞（MIA）和肌肉卫星细胞（MSC），并通过天花板培养使 MIA 去分化转化为 IMP。然后通过差速贴壁法纯化 MSC 2 h。倒置显微镜观察、MTT 检测、油红 O 染色和 q-PCR 结果表明，MIA 去分化细胞被鉴定为 IMP，具有与腹部脂肪前体脂肪细胞相同的细胞形态、分化能力、增殖和传代能力（P>0.05）。所获得的 IMP 用于与 MSC 共培养实验的适用性表明，它可以满足实验研究的要求。最后，使用 Transwell 室建立了 IMP 和 MSC 的共培养系统。共培养结果表明，处于增殖期的 MSC 倾向于加速 IMP 的分化，导致共培养的 IMP 中的脂肪含量比单独培养的 IMP 中的脂肪含量更高（P<0.05），在非增殖期，结果则相反（P<0.05）。相关基因的 mRNA 水平根据细胞中的脂肪含量而显著变化。这些结果有力地支持了所建立的共培养系统是有效和可行的观点。总之，我们成功地找到了一种新的方法来探索鸡的肌细胞和脂肪细胞之间的相互作用。我们的研究结果可以加深对鸡肌细胞和脂肪细胞调节机制的研究。
