袖状胃切除术影响肥胖小鼠迷走神经传入神经元激素受体的表达

Effect of sleeve gastrectomy on expression of hormonal receptors on vagus afferent neurons in obese mice

目的探讨袖状胃切除术(SG)对迷走神经传入神经元(VANs)食欲相关激素受体表达水平的影响。方法采用10周龄健康的小鼠给予高脂饮食喂养, 获得12只饮食诱导肥胖(DIO)小鼠, 采用随机数字表法随机分配至SG组和假手术配对组, 每组6只。观察两组小鼠体质量和空腹血糖水平变化。6周后, 无菌操作取出颈部迷走神经节, 体外原代培养迷走神经元细胞, 利用神经元特异标志物MAP2鉴定神经元的纯度, 并用免疫组织化学和Western blot方法检测迷走神经元中的抑制食欲激素受体CCK1-R、LepR和GhR的表达情况。计数资料采用t检验。结果术后6周, SG小鼠的体重和血糖水平明显低于对照组, 差异有统计学意义[(31.6±3.4) g比(49.8±2.7) g, t=-8.400, P<0.01;(5.4±0.24) mmol/L比(9.5±0.5) mmol/L, t=-14.406, P<0.01]。体外培养的迷走神经元形态饱满, 形成密集交错的神经元网络。神经元特异标志物鉴定后, 纯度达到(97.01+0.23)%。细胞传代后生长情况及增殖能力良好。SG后迷走神经VANs的CCK1-R与LepR表达均高于假手术组(CCK1-R, 1.92±0.16比0.81±0.12, t=10.626, P<0.001;LepR, 1.72±0.28比0.89±0.09, t=5.395, P<0.05), 而GhR表达低于假手术组(0.77±0.11比1.07±0.11, t=-5.180, P<0.001)。结论 SG术后DIO小鼠VANs抑制食欲激素受体表达增加, 而促食欲激素受体表达减少, 提示SG可以改变VANs表型。

Objective To establish an in vitro primary culture method for vagus neurons in high-fat diet-induced obesity(DIO)mice,and explore the effects of sleeve gastrectomy(SG)on the expression of apetite-related hormone receptors in vagal neurons.Methods The 10-week-old healthy mice were fed on high-fat diet to obtain 12 DIO mice and randomly divided into SG group and sham group according to table of random numbers.Body weight and fasting sugar level during perioperative period were observed.At 6th week after surgery,the cervical vagus ganglions were sampled,and the vagus neurons were cultured in vitro.The neuron-specific marker MAP2 was used to identify the purity of the neurons.Immunohistochemistry and Western blotting were used to detect the expression of receptors of appetite suppressing hormone in the vagus neurons,such as CCK1-R,LepR and GhR.T test was used for counting data.Results At 6th week after surgery,compared with sham group,the body weight and blood glucose level in the SG group decreased significantly,and the difference was statistically significant[(31.6±3.4)g vs.(49.8±2.7)g,t=-8.400,P<0.01;(5.4±0.24)mmol/L vs.(9.5±0.5)mmol/L,t=-14.406,P<0.01].The vagus neurons cultured in vitro were full in shape and formed a densely interlaced neuron network.Identification of neuron-specific markers showed the purity was(97.01+0.23)%.The growth and proliferation ability of the neurons remained good after passage.The expression of CCK1-R and LepR in the vagus nerve VANs after SG was higher than that in the sham group(for CCK1-R,1.92±0.16 vs.0.81±0.12,t=10.626,P<0.001;for LepR,1.72±0.28 vs.0.89±0.09,t=5.395,P<0.05),while the expression of GhR was lower than that in the sham group(0.77±0.11 vs.1.07±0.11,t=-5.180,P<0.01).Conclusion SG can increase the expression of receptors of appetite suppressing hormone in VANs of DIO mice,and reduce the expression of receptors of appetite stimulating hormone,which indicates SG can change the phenotype of VANs.