Heat stress (HS) results in health problems in animals. This study was conducted to investigate the effect and the underlying mechanism of HS on the proliferation and differentiation process of 3T3-L1 preadipocytes. 3T3-L1 preadipocytes were treated at 37 °C or 41.5 °C. HS up-regulated the mRNA and protein expression level of heat shock protein 70 (HSP70). Furthermore, the proliferation of 3T3-L1 preadipocytes were significantly inhibited after HS treatment for 2 days. A large number of accumulated lipid droplets were observed under the microscope after HS treatment for 8 days. Notably, the result of oil red O staining showed that the number of lipid droplets increased significantly and the differentiation ability of the cells was enhanced after HS. Moreover, after 2 and 8 d of differentiation, HS increased the transcription levels of fat synthesis genes including peroxisome proliferators activated receptor γ (PPARγ), fatty acid binding protein 2 (AP2), fatty acid synthase (FAS) and CCAAT enhancer binding protein α (CEBPα) genes, while decreasing the transcription levels of lipid decomposition genes including ATGL and HSL genes. In addition, HS reduced the expression of AMPK and PGC-1α, as well as the dephosphorylation of AMPK. 5-Aminoimidazole-4-carboxamide ribonucleotide (AICAR) can eliminate HS induced lipogenesis by activating AMPK. These results indicated that HS inhibited the proliferation of 3T3-L1 preadipocytes and promoted lipid accumulation by inhibiting the AMPK-PGC-1α signaling pathway in 3T3-L1 preadipocytes. This work lays a theoretical foundation for improving the effect of HS on meat quality of livestock and provides a new direction for the prevention of obesity caused by HS.

热应激 (HS) 会导致动物出现健康问题。本研究旨在探讨HS对3T3-L1前脂肪细胞增殖和分化过程的影响及其潜在机制。 3T3-L1前脂肪细胞在37°C或41.5°C下处理。 HS 上调热休克蛋白 70 (HSP70) 的 mRNA 和蛋白表达水平。此外，HS处理2天后，3T3-L1前脂肪细胞的增殖受到显着抑制。 HS处理8天后，在显微镜下观察到大量积累的脂滴。值得注意的是，油红O染色结果显示，HS后脂滴数量显着增加，细胞分化能力增强。此外，分化2天和8天后，HS增加了脂肪合成基因的转录水平，包括过氧化物酶体增殖物激活受体γ（PPARγ）、脂肪酸结合蛋白2（AP2）、脂肪酸合酶（FAS）和CCAAT增强子结合蛋白α （CEBPα）基因，同时降低脂质分解基因（包括 ATGL 和 HSL 基因）的转录水平。此外，HS还降低了AMPK和PGC-1α的表达，以及AMPK的去磷酸化。 5-氨基咪唑-4-甲酰胺核糖核苷酸 (AICAR) 可以通过激活 AMPK 消除 HS 诱导的脂肪生成。这些结果表明，HS通过抑制3T3-L1前脂肪细胞中的AMPK-PGC-1α信号通路来抑制3T3-L1前脂肪细胞的增殖并促进脂质积累。该工作为提高HS对牲畜肉品质的影响奠定了理论基础，为预防HS引起的肥胖提供了新的方向。