Pan Shifeng, Chen Yongfang, Zhang Lin, Liu Zhuang, Xu Xingyu, Xing Hua
College of Veterinary Medicine, Yangzhou University, Yangzhou, Jiangsu, 225009, China.
Jiangsu Co-Innovation Center for the Prevention and Control of Important Animal Infectious Disease and Zoonoses, Yangzhou, Jiangsu, 225009, China.
Anim Biosci. 2022 May;35(5):763-777. doi: 10.5713/ab.21.0371. Epub 2021 Nov 1.
Excessive lipid accumulation in adipocytes results in prevalence of obesity and metabolic syndrome. Curcumin (CUR), a naturally phenolic active ingredient, has been shown to have lipid-lowering effects. However, its underlying mechanisms have remained largely unknown. Therefore, the study aims to determine the effect of CUR on cellular lipid accumulation in porcine subcutaneous preadipocytes (PSPA) and to clarify novel mechanisms.
The PSPA were cultured and treated with or without CUR. Both cell counting Kit-8 and lactate dehydrogenase release assays were used to examine cytotoxicity. Intracellular lipid contents were measured by oil-red-o staining extraction and triglyceride quantification. Apoptosis was determined by flow cytometry and the terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate-nick end labelling assay. Adipogenic and apoptosis genes were analyzed by quantitative polymerase chain reaction and Western blot.
The CUR dose-dependently reduced the proliferation and lipid accumulation of PSPA. Noncytotoxic doses of CUR (10 to 20 μM) significantly inhibited extracellular signal-regulated kinase 1/2 (ERK1/2) phosphorylation and expression of adipogenic genes peroxisome proliferation-activity receptor-γ (PPAR-γ), CCAAT/enhancer binding protein-α, sterol regulatory element-binding protein-1c, adipocyte protein-2, glucose transporter-4 as well as key lipogenic enzymes fatty acid synthase and acetyl-CoA carboxylase, while ERK1/2 activation significantly reversed CUR-reduced lipid accumulation by increasing PPAR-γ. Furthermore, compared with differentiation induced media treated cells, higher dose of CUR (30 μM) significantly decreased the expression of AKT and B-cell lymphoma-2 (BCL2), while increased the expression of BCL-2-associated X (BAX) and the BAX/BCL-2 expression ratio, suggesting triggered apoptosis by inactivating AKT and increasing BAX/BCL-2 ratio and Caspase-3 expression. Moreover, AKT activation significantly rescued CUR inhibiting lipid accumulation via repressing apoptosis.
These results demonstrate that CUR is capable of suppressing differentiation by inhibiting ERK1/2-PPAR-γ signaling pathway and triggering apoptosis via decreasing AKT and subsequently increasing BAX/BCL-2 ratio and Caspase-3, suggesting that CUR provides an important method for the reduction of porcine body fat, as well as the prevention and treatment of human obesity.
脂肪细胞中脂质过度积累导致肥胖和代谢综合征的普遍发生。姜黄素（CUR）是一种天然酚类活性成分，已被证明具有降脂作用。然而，其潜在机制在很大程度上仍不清楚。因此，本研究旨在确定CUR对猪皮下前脂肪细胞（PSPA）中细胞脂质积累的影响，并阐明新的机制。
培养PSPA并分别用或不用CUR处理。使用细胞计数试剂盒8和乳酸脱氢酶释放试验检测细胞毒性。通过油红O染色提取和甘油三酯定量测定细胞内脂质含量。通过流式细胞术和末端脱氧核苷酸转移酶介导的脱氧尿苷三磷酸缺口末端标记试验确定细胞凋亡。通过定量聚合酶链反应和蛋白质免疫印迹分析脂肪生成和凋亡相关基因。
CUR剂量依赖性地降低了PSPA的增殖和脂质积累。非细胞毒性剂量的CUR（10至20μM）显著抑制细胞外信号调节激酶1/2（ERK1/2）磷酸化以及脂肪生成基因过氧化物酶体增殖物激活受体γ（PPAR-γ）、CCAAT/增强子结合蛋白α、固醇调节元件结合蛋白1c、脂肪细胞蛋白2、葡萄糖转运蛋白4以及关键脂肪生成酶脂肪酸合酶和乙酰辅酶A羧化酶的表达，而ERK1/2激活通过增加PPAR-γ显著逆转了CUR降低的脂质积累。此外，与分化诱导培养基处理的细胞相比，高剂量的CUR（30μM）显著降低了AKT和B细胞淋巴瘤-2（BCL2）的表达，同时增加了Bcl-2相关X蛋白（BAX）的表达以及BAX/BCL-2表达比率，表明通过使AKT失活、增加BAX/BCL-2比率和半胱天冬酶-3表达触发细胞凋亡。此外，AKT激活通过抑制细胞凋亡显著挽救了CUR对脂质积累的抑制作用。
这些结果表明，CUR能够通过抑制ERK1/2-PPAR-γ信号通路抑制分化，并通过降低AKT水平、随后增加BAX/BCL-2比率和半胱天冬酶-3触发细胞凋亡，表明CUR为降低猪体脂以及预防和治疗人类肥胖提供了一种重要方法。
