Tachtsis Bill, Whitfield Jamie, Hawley John A, Hoffman Nolan J
Exercise and Nutrition Research Program, Mary MacKillop Institute for Health Research, Australian Catholic University, Melbourne, VIC, Australia.
Front Physiol. 2020 Jun 3;11:563. doi: 10.3389/fphys.2020.00563. eCollection 2020.
Accumulation of excess saturated free fatty acids such as palmitate (PAL) in skeletal muscle leads to reductions in mitochondrial integrity, cell viability and differentiation. Omega-3 polyunsaturated fatty acids (n-3 PUFAs) such as eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) counteract PAL-induced lipid accumulation. EPA and DHA, as well as the n-3 PUFA docosapentaenoic acid (DPA), may therefore mitigate PAL-induced lipotoxicity to promote skeletal muscle cell survival and differentiation. C2C12 myoblasts were treated with 50 μM EPA, DPA, or DHA in the absence or presence of 500 μM PAL for 16 h either prior to myoblast analysis or induction of differentiation. Myoblast viability and markers of apoptosis, endoplasmic reticulum (ER) stress and myotube differentiation capacity were investigated using fluorescence microscopy and immunoblotting. High-resolution respirometry was used to assess mitochondrial function and membrane integrity. PAL induced cell death via apoptosis and increased protein content of ER stress markers BiP and CHOP. EPA, DPA, and DHA co-treatment maintained cell viability, prevented PAL-induced apoptosis and attenuated PAL-induced increases in BiP, whereas only DPA prevented increases in CHOP. PAL subsequently reduced protein content of the differentiation marker myogenin and inhibited myotube formation, and all n-3 PUFAs promoted myotube formation in the presence of PAL. Furthermore, DPA prevented PAL-induced release of cytochrome c and maintained mitochondrial integrity. These findings demonstrate the n-3 PUFAs EPA, DPA and DHA elicit similar protective effects against PAL-induced impairments in muscle cell viability and differentiation. Mechanistically, the protective effects of DPA against PAL lipotoxicity are attributable in part to its ability to maintain mitochondrial respiratory capacity via mitigating PAL-induced loss of mitochondrial membrane integrity.
骨骼肌中过量饱和游离脂肪酸（如棕榈酸酯，PAL）的积累会导致线粒体完整性、细胞活力和分化能力下降。ω-3多不饱和脂肪酸（n-3 PUFAs），如二十碳五烯酸（EPA）和二十二碳六烯酸（DHA），可抵消PAL诱导的脂质积累。因此，EPA、DHA以及n-3多不饱和脂肪酸二十二碳五烯酸（DPA）可能减轻PAL诱导的脂毒性，从而促进骨骼肌细胞的存活和分化。在进行成肌细胞分析或诱导分化之前，将C2C12成肌细胞在不存在或存在500 μM PAL的情况下，用50 μM EPA、DPA或DHA处理16小时。使用荧光显微镜和免疫印迹法研究成肌细胞活力、凋亡标志物、内质网（ER）应激和肌管分化能力。采用高分辨率呼吸测定法评估线粒体功能和膜完整性。PAL通过凋亡诱导细胞死亡，并增加ER应激标志物BiP和CHOP的蛋白质含量。EPA、DPA和DHA联合处理可维持细胞活力，防止PAL诱导的凋亡，并减弱PAL诱导的BiP增加，而只有DPA可防止CHOP增加。PAL随后降低了分化标志物肌细胞生成素的蛋白质含量，并抑制了肌管形成，而在存在PAL的情况下，所有n-3多不饱和脂肪酸均促进了肌管形成。此外，DPA可防止PAL诱导的细胞色素c释放，并维持线粒体完整性。这些发现表明，n-3多不饱和脂肪酸EPA、DPA和DHA对PAL诱导的肌肉细胞活力和分化损伤具有相似的保护作用。从机制上讲，DPA对PAL脂毒性的保护作用部分归因于其通过减轻PAL诱导的线粒体膜完整性丧失来维持线粒体呼吸能力的能力。
