CTRP4基因表达上调通过抑制炎症因子调控食欲调节相关蛋白表达

上调中枢补体C1q/肿瘤坏死因子相关蛋白4(complement-C1q/tumor necrosis factor-related protein 4, CTRP4) 可以改变下丘脑食欲调节相关蛋白的表达，抑制小鼠摄食且降低其体重。然而，CTRP4如何调控食欲调节相关蛋白的表达尚不清楚。本研究通过上调小鼠神经母细胞瘤细胞(N2a)中的CTRP4，探讨CTRP4调控食欲调节相关蛋白的潜在作用机制。通过对N2a细胞未做干预、转染绿色荧光蛋白(green fluorescent protein, GFP)重组腺病毒和CTRP4过表达重组腺病毒，将其分为空白对照组(Control组)、阴性对照组(Ad-GFP组)及CTRP4过表达组(Ad-CTRP4组)。干预72 h时，用实时荧光定量PCR(RT-PCR)检测细胞CTRP4 mRNA表达，采用Western 印迹检测细胞CTRP4、Pomc、Npy、p-STAT3/t-STAT3、TNF-α、IL-6、SOCS3在蛋白质水平的表达。结果显示，与对照组相比，Ad-CTRP4组的CTRP4 mRNA水平(26 258.44±10 403.47 vs. 1.81±0.79 vs. 1.00±0.00, P<0.01)及蛋白质水平显著增加(10.44±7.99 vs.0.64±0.62 vs. 1.00±0.75, P<0.01)。Ad-CTRP4组的p-STAT3/t-STAT3(3.38±1.70 vs. 0.86±0.57 vs. 1.00±0.63, P<0.01)和Pomc(1.81±0.19 vs. 1.15±0.18 vs. 1.00±0.22, P<0.01)表达均显著增高；SOCS3(0.69±0.15 vs. 1.00±0.12 vs. 1.00±0.07, P<0.01)，IL-6(0.40±0.19 vs. 1.03±0.17 vs. 1.00±0.16, P<0.01)，TNF.α(0.39±0.27 vs. 1.05±0.46 vs. 1.00±0.29, P<0.05)及Npy (0.55±0.14 vs. 1.21±0.38 vs. 1.00±0.24, P<0.05)表达均显著下降。上述结果提示，在N2a细胞中，上调CTRP4可能通过抑制炎症因子TNF-α和IL-6，降低负性调节因子SOCS3的表达，增加STAT3磷酸化表达水平，从而调控食欲调节相关蛋白的表达。

Upregulation of CTRP4 Regulates Appetite-regulating Proteins Expression by Inhibiting Inflammatory Factors

Upregulation of complement-C1q/tumor necrosis factor-related protein 4 (CTRP4) gene expression could change the expression of the appetite-regulating proteins in the hypothalamus, which lead to inhibit feeding and reduce body weight of mice. However, it is not clear how CTRP4 regulates the expression of the appetite-regulating proteins. To investigate the molecular mechanisms through which CTRP4 regulates appetite-regulating protein expression in mouse neuroblastoma cells (N2a). The N2a cells were divided into blank group (control group), negative group (Ad-GFP group), and CTRP4 overexpression group (Ad-CTRP4 group). After infection with green fluorescent protein (GFP) recombinant adenovirus or CTRP4 overexpressed recombinant adenovirus for 72 hours, RT-PCR was performed to determine the mRNA levels of CTRP4. Western blot was performed to determine the protein levels of CTRP4, TNF-α, SOCS3, p-STAT3/t-STAT3, Pomc and Npy. The results showed that compared with the control group, the levels of CTRP4 mRNA (26258.44±10403.47 vs. 1.81±0.79 vs. 1.00±0.00, P<0.01) and protein (10.44±7.99 vs. 0.64±0.62 vs. 1.00±0.75, P<0.01) were significantly increased in N2A cells of CTRP4 overexpression group. The levels of p-STAT3/t-STAT3 (3.38±1.7 vs. 0.86±0.57 vs. 1.00±0.63, P<0.01) and Pomc (1.81±0.19 vs. 1.15±0.18 vs. 1.00±0.22, P<0.01) were markedly increased in Ad-CTRP4 group. The expression levels of SOCS3 (0.69±0.15 vs. 1.00±0.12 vs. 1.00±0.07, P<0.01), IL-6 (0.40±0.19 vs. 1.03±0.17 vs. 1.00±0.16, P<0.01), TNF-α (0.39±0.27 vs. 1.05±0.46 vs. 1.00±0.29, P<0.05) and Npy (0.55±0.14 vs. 1.21±0.38 vs. 1.00±0.24, P<0.05) were obviously decreased. The findings suggest that increased CTRP4 level in N2a cells could inhibit inflammatory factors such as TNF-α and IL-6, reduce the expression of negative regulator SOCS3, and increase p-STAT3/t-STAT3 level, to regulate appetite-regulating proteins.