Xie Xiang-Zhu, Huang Xin, Zhao Shui-Ping, Yu Bi-Lian, Zhong Qiao-Qing, Cao Jian
Department of Geriatric Cardiology, Chinese PLA General Hospital, Beijing 100853, China.
Chin Med J (Engl). 2016 May 5;129(9):1108-12. doi: 10.4103/0366-6999.180519.
Adipocytes behave like a rich source of pro-inflammatory cytokines including monocyte chemoattractant protein-1 (MCP-1). Oxidized low-density lipoprotein (oxLDL) participates in the local chronic inflammatory response, and high-density lipoprotein could counterbalance the proinflammatory function of oxLDL, but the underlying mechanism is not completely understood. This study aimed to evaluate the effect of apolipoprotein A-I mimetic peptide L-4F on the secretion and expression of MCP-1 in fully differentiated 3T3-L1 adipocytes induced by oxLDL and to elucidate the possible mechanisms.
Fully differentiated 3T3-L1 adipocytes were incubated in the medium containing various concentration of L-4F (0-50 μg/ml) with oxLDL (50 μg/ml) stimulated, with/without protein kinase A (PKA) inhibitor H-89 (10 μmol/L) preincubated. The concentrations of MCP-1 in the supernatant, the mRNA expression of MCP-1, the levels of CCAAT/enhancer binding protein α (C/EBPα), and CCAAT/enhancer binding protein β (C/EBPβ) were evaluated. The monocyte chemotaxis assay was performed by micropore filter method using a modified Boyden chamber.
OxLDL stimulation induced a significant increase of MCP-1 expression and secretion in 3T3-L1 adipocytes, which were inhibited by L-4F preincubation in a dose-dependent manner. PKA inhibitor H-89 markedly reduced the oxLDL-induced MCP-1 expression, but no further decrease was observed when H-89 was used in combination with L-4F (50 μg/ml) (P > 0.05). OxLDL stimulation showed no significant effect on C/EBPα protein level but increased C/EBPβ protein level in a time-dependent manner. H-89 and L-4F both attenuated C/EBPβ protein level in oxLDL-induced 3T3-L1 adipocytes.
OxLDL induces C/EBPβ protein synthesis in a time-dependent manner and enhances MCP-1 secretion and expression in 3T3-L1 adipocytes. L-4F dose-dependently counterbalances the pro-inflammatory effect of oxLDL, and cyclic AMP/PKA-C/EBPβ signaling pathway may participate in it.
脂肪细胞表现得像是包括单核细胞趋化蛋白-1（MCP-1）在内的促炎细胞因子的丰富来源。氧化型低密度脂蛋白（oxLDL）参与局部慢性炎症反应，而高密度脂蛋白可以抵消oxLDL的促炎功能，但其潜在机制尚未完全阐明。本研究旨在评估载脂蛋白A-I模拟肽L-4F对oxLDL诱导的完全分化的3T3-L1脂肪细胞中MCP-1分泌和表达的影响，并阐明其可能的机制。
将完全分化的3T3-L1脂肪细胞在含有不同浓度L-4F（0-50μg/ml）的培养基中孵育，同时用oxLDL（50μg/ml）刺激，预先孵育或不预先孵育蛋白激酶A（PKA）抑制剂H-89（10μmol/L）。评估上清液中MCP-1的浓度、MCP-1的mRNA表达、CCAAT/增强子结合蛋白α（C/EBPα）和CCAAT/增强子结合蛋白β（C/EBPβ）的水平。单核细胞趋化试验采用改良的Boyden小室微孔滤膜法进行。
oxLDL刺激导致3T3-L1脂肪细胞中MCP-1表达和分泌显著增加，而预先孵育L-4F可剂量依赖性地抑制这种增加。PKA抑制剂H-89显著降低了oxLDL诱导的MCP-1表达，但当H-89与L-4F（50μg/ml）联合使用时未观察到进一步降低（P>0.05）。oxLDL刺激对C/EBPα蛋白水平无显著影响，但以时间依赖性方式增加了C/EBPβ蛋白水平。H-89和L-4F均减弱了oxLDL诱导的3T3-L1脂肪细胞中C/EBPβ蛋白水平。
oxLDL以时间依赖性方式诱导C/EBPβ蛋白合成，并增强3T3-L1脂肪细胞中MCP-1的分泌和表达。L-4F剂量依赖性地抵消oxLDL的促炎作用，且环磷酸腺苷/PKA-C/EBPβ信号通路可能参与其中。
