慢病毒介导Lipin1表达对糖尿病大鼠周围神经病变的影响

发布时间：2018-06-29 16:51

本文选题：脂素 + 糖尿病　；参考：《山东大学》2015年硕士论文

【摘要】：研究背景：糖尿病周围神经病变(diabetic peripheral neuropathy, DPN)是糖尿病(diabetic mellitus, DM)最常见的慢性并发症之一,其患病率达50%,是造成糖尿病足和截肢的主要原因,严重影响患者的生活质量,目前临床上尚无有效的治疗方法。目前对DPN发病机制的研究多集中在多元醇通路的代谢异常、糖基化终末产物(AGEs)的积聚、氧化应激、神经营养因子等。脂素(Lipin)由脂素基因(LPIN)表达产生,是一种新发现的具有双向调控功能的一个家族,可调控身体脂肪代谢,主要包括Lipin 1、Lipin2和Lipin3。脂素Lipin1作为一种新的脂肪因子,在机体糖脂代谢中起着重要作用。Lipin1主要表达于白色脂肪组织及肌肉,同时也表达于其他组织如周围神经、肝脏及肾脏。有研究发现Lipin1缺乏是造成周围神经病变的原因之一,主要机制在于Lipin1缺乏导致构成神经外膜的脂肪垫以及雪旺细胞中的磷脂酸磷酸酯酶活性缺乏,髓磷脂合成减少,神经传导速度减慢。研究目的：本研究通过构建糖尿病周围神经病变大鼠模型,将携带Lipin1的慢病毒载体转染糖尿病大鼠,观察糖尿病大鼠坐骨神经的神经电生理及病理形态学变化,探讨Lipin1对糖尿病大鼠周围神经早期病变的影响。方法：36只雄性Wistar大鼠高糖高脂饲料喂养8周后给予小剂量(30 mg/kg)链脲佐菌素(STZ)腹腔注射,成功诱导其成为2型糖尿病(type2 diabetes mellitus, T2DM)大鼠模型,随机分为空载体组、Lipin1过表达组,另选择12只健康大鼠作为正常对照组。Lipin1过表达(LV-Lipin1)组、空载体(LV-contro1)组分别通过尾静脉注射lipin1慢病毒载体(5x107TU)和空慢病毒载体(5×107 TU),每月1次,连续2次。8周后对照观察各组大鼠血糖(BG)、体质量(BW)；应用甲苯胺蓝染色,通过光镜及透射电镜观察坐骨神经病理形态学改变；应用免疫组化法测定Lipin1表达。数据均采用应用SPSS 19.0统计软件,计量资料采用x±s,多组间数据比较采用单因素方差分析,两两比较采用SNK法,P0.05为差异有统汁学意义。结果：1.体重和血糖至干预结束时,正常组体质量呈生理性正常加重,血糖正常。空载体组大鼠血糖高于正常组,其差异有统计学意义(P0.05)。Lipin1过表达组血糖、体质量较空载体组无显著差异(P0.05)。2.Lipin1在大鼠坐骨神经中的表达糖尿病大鼠成模2月时,免疫组化观察坐骨神经Lipin1表达,空载体组较正常对照组坐骨神经纵切面Lipin1表达量明显减少；Lipin1过表达组较空载体组Lipin1表达量明显增多。3.神经传导速度糖尿病大鼠造模后4w即可出现神经传导速度减慢,Lipin1转染后8周,Lipin1过表达组神经传导速度较空载体组明显改善(P0.05)。4.对糖尿病大鼠病理改变的影响糖尿病大鼠造模后8w,空载体组较正常对照组的神经纤维直径、密度、面积均减少,Lipin1过表达组以上病变较空载体组显著减轻(P0.05)。结论：1.实验性糖尿病大鼠成模后4w发现周围神经病变,随病程延长神经损伤逐渐加重。2.糖尿病周围神经病变大鼠坐骨神经中Lipin1表达降低,上调Lipin1表达后糖尿病大鼠坐骨神经神经传导速度改善,病理学观察中亦可见到神经髓鞘结构和轴突形态的改善。
[Abstract]:Background: diabetic peripheral neuropathy (DPN) is one of the most common chronic complications of diabetes (diabetic mellitus, DM). Its prevalence rate is 50%. It is the main cause of diabetic foot and amputation, which seriously affects the living quality of patients. At present, there is no effective treatment in clinical. The research on the pathogenesis of DPN mainly focuses on the metabolic abnormality of the polyol pathway, the accumulation of glycosylated end products (AGEs), oxidative stress, neurotrophic factor, and so on. Lipoprotein (Lipin) is produced by the expression of the lipoprotein gene (LPIN). It is a newly discovered family with bi-directional regulation function, which can regulate body fat metabolism, mainly including Lipin 1, Lipi N2 and Lipin3. lipoprotein Lipin1, as a new fat factor, play an important role in the metabolism of glycolipid,.Lipin1 is mainly expressed in white adipose tissue and muscles, but also in other tissues such as peripheral nerve, liver and kidney. It is found that the lack of Lipin1 is one of the causes of peripheral neuropathy, and the main mechanism is Li Pin1 deficiency leads to the lipid pads that constitute the outer membrane of the nerve and the deficiency of phosphatidate phosphatidase activity in the Schwann cells, the decrease of the synthesis of myelin and the slow speed of nerve conduction. The effect of Lipin1 on the early pathological changes of peripheral nerve in diabetic rats. Methods: 36 male Wistar rats were fed with high glucose and high fat diet for 8 weeks, and a small dose (30 mg/kg) of streptozotocin (STZ) was intraperitoneally injected, and it was successfully induced to become type 2 diabetes mellitus (type2 diabetes mel). Litus, T2DM) rat model, randomly divided into empty carrier group, Lipin1 overexpression group, and 12 healthy rats as normal control group.Lipin1 overexpression (LV-Lipin1) group, LV-contro1 group via tail vein injection of lipin1 lentivirus carrier (5x107TU) and empty lentivirus vector (5 x 107 TU), 1 times a month, 2 consecutive.8 weeks after the control view The blood glucose (BG) and body mass (BW) were observed in each group. The pathological changes of the sciatic nerve were observed with toluidine blue, and the expression of Lipin1 was measured by the light microscopy and transmission electron microscopy. The data were used to use the SPSS 19 statistical software, the measurement data were x s, and the data of multiple groups were compared with the single factor analysis of variance, 22 The SNK method was used to compare the significance of P0.05. Results: when 1. body weight and blood sugar were at the end of the intervention, the normal body mass showed normal weight and normal blood sugar. The blood glucose in the no-load group was higher than that of the normal group. The difference was statistically significant (P0.05).Lipin1 overexpression group blood glucose, and there was no significant difference between the body mass and the empty carrier group (P0.05). The expression of Lipin1 in the sciatic nerve was observed by.2.Lipin1 in the rat model of sciatic nerve in the rat's sciatic nerve in February. The expression of Lipin1 expression in the longitudinal section of the sciatic nerve was significantly reduced in the no-load group than in the normal control group, and the expression of Lipin1 in the Lipin1 overexpression group was significantly increased by the.3. nerve conduction velocity diabetic rat model. The nerve conduction velocity decreased after 4W, 8 weeks after Lipin1 transfection, the nerve conduction velocity in the Lipin1 overexpression group was significantly improved (P0.05).4. on the pathological changes of diabetic rats, and 8W in the diabetic rats. The diameter, density, and area of Lipin1 overexpression group were lower than those in the normal control group. In 1. experimental diabetic rats, 4W found peripheral neuropathy in 1. experimental diabetic rats and gradually increased the Lipin1 expression in the sciatic nerve of rats with.2. diabetic peripheral neuropathy with the duration of the disease, and the increase of the nerve conduction velocity of the sciatic nerve in diabetic rats after the up-regulation of Lipin1. The myelin sheath and axonal morphology were also observed in pathological observation.
【学位授予单位】：山东大学
【学位级别】：硕士
【学位授予年份】：2015
【分类号】：R587.2

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