【摘要】 目的 探讨巨噬细胞转录因子阴阳1（YY1）基因对葡萄糖耐量、胰岛素敏感性的影响。方法 通过Cre-Loxp系统，成功获得巨噬细胞YY1基因特异性敲除(YY1^f/f;lyz2-cre)小鼠。高脂喂养小鼠12～13周，检测小鼠葡萄糖耐量（血糖水平）和胰岛素耐量（相对血糖值）。将高脂喂养的YY1基因敲除小鼠(实验组，YY1^f/f;lyz2-cre)与对照组(YY1^f/f)小鼠分别注射0.9%氯化钠溶液和胰岛素（剂量均为0.75 U/kg）,取其附睾脂肪、肌肉、肝脏组织，采用免疫印迹法检测蛋白激酶B（Akt）-糖原合成酶激酶3β（Gsk3β）通路关键蛋白（Akt、Gsk3β）的磷酸化水平。采用ELISA法检测高脂饮食后实验组小鼠与对照组小鼠血清胰岛素、脂联素和瘦素水平。应用实时荧光定量PCR（qRT-PCR）检测正常饮食的两组小鼠骨髓来源巨噬细胞（BMDM）中炎症因子mRNA相对表达量。结果 琼脂糖凝胶电泳见700 bp处显示条带，表明lyz2-cre基因成功表达。实验组小鼠BMDM中，YY1基因mRNA相对表达量显著低于对照组，表明巨噬细胞YY1基因特异性敲除小鼠模型构建成功。高脂喂养下，实验组小鼠注射葡萄糖后60、90 min时的血糖水平均显著低于对照组同时间点（P值均<0.05）,实验组小鼠注射胰岛素后30、60、90 min时的相对血糖值均显著低于对照组同时间点（P值分别<0.05、0.001、0.01）。正常饮食喂养下，两组各时间点间血糖水平和相对血糖值的差异均无统计学意义（P值均>0.05）。实验组小鼠附睾脂肪中胰岛素信号通路关键蛋白Akt及Gsk3β磷酸化水平较对照组均明显上调，肌肉和肝脏组织中无明显变化。实验组小鼠血清瘦素、脂联素水平均显著高于对照组（P值均<0.05）;实验组小鼠血清胰岛素水平较对照组有下降趋势，但差异无统计学意义（P>0.05）。实验组小鼠BMDM中IL-6 mRNA、TNF-α mRNA、IL-1β mRNA相对表达量均显著低于对照组（P值均<0.001）。结论 巨噬细胞转录因子YY1基因敲除，可以通过减轻巨噬细胞炎症，上调脂联素表达，增强脂肪组织胰岛素敏感性，从而改善高脂饮食小鼠的胰岛素抵抗。更多还原

【Abstract】 Objective To explore the effect of macrophageYin Yang 1(YY1) gene on glucose tolerance and insulin sensitivity. Methods We used Cre-LoxP system to breed myeloid-specific YY1 knockout mice. The glucose tolerance(blood glucose level) and insulin tolerance(relative blood glucose level) were measured in mice fed with high fat diet for 12 to 13 weeks. High-fat-diet YY1 knockout mice(experimental group) and control mice(control group) were injected with normal saline and insulin(0.75 U/kg), respectively, and their epididymal white adipose, muscle and liver tissues were collected. The expression levels of key proteins in the protein kinase B(Akt)-glycogen synthase kinase 3β(Gsk3β) pathway were detected by Western blotting. Serum insulin, adiponectin and leptin levels in YY1 knockout mice and control mice fed with high-fat diet were measured by enzyme-linked immunosorbent assay(ELISA). The mRNA expression of inflammatory factors in bone marrow derived macrophage(BMDM) from YY1 knockout mice and control mice with chow diet were detected by quantitative real-time fluorescent PCR(qRT-PCR). Results Agarose gel electrophoresis showed a band at 700 bp, indicating that lyz2-cre gene was successfully expressed. The mRNA relative expression of YY1 gene in BMDM in the experimental group was significantly lower than that in the control group, indicating that BMDM YY1-specific knockout mouse model was successfully constructed. After high-fat feeding, the blood glucose level of the experimental group at 60 and 90 min after glucose injection was significantly lower than that of the control group(both P<0.05), and the relative blood glucose level of the experimental group at 30, 60 and 90 min after insulin injection was significantly lower than that of the control group(P<0.05, 0.001, 0.01). There was no significant difference in the glucose tolerance or insulin tolerance between the two groups after chow diet(all P>0.05). The level of phosphorylation of Akt and Gsk3β in adipose tissue, and serum leptin and adiponectin levels were significantly increased in the experimental group as compared with the control group(all P<0.05). Serum insulin level in the experimental group was slightly decreased compared with the control group, but there was no significant difference in the serum insulin level between the two groups(P>0.05). The mRNA levels of proinflammatory factors in BMDM, including IL-6, TNF-α and IL-1β were significantly lower in the experimental group than those in the control group(all P<0.001). Conclusion YY1 deletion in macrophages can improve glucose tolerance and insulin sensitivity probably through activating AKT-GSK3β insulin pathway, which may be associated with the secretion of adipokines and amelioration of macrophage inflammation.更多还原