基于转录组分析母代孕期及哺乳期低蛋白饮食的子代雌性小鼠胰岛功能受损的相关机制  被引量:1

Mechanisms underlying the impairment of islet function in female offspring mice exposed to maternal low-protein diet during pregnancy and lactation:a transcriptomic analysis

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作  者:彭粤跃 严晋华[1] 陈亚兰 刘芷谷 林倍思[1] 凌萍 徐芬[1] 刘子瑜 李晏丽 梁华 许雯[1] Peng Yueyue;Yan Jinhua;Chen Yalan;Liu Zhigu;Lin Beisi;Ling Ping;Xu Fen;Liu Ziyu;Li Yanli;Liang Hua;Xu Wen(Department of Endocrinology and Metabolism,Third Affiliated Hospital of Sun Yat-sen University,Guangdong Provincial Key Laboratory of Diabetology,Guangzhou 510630,China;Guangzhou First People′s Hospital,Guangzhou 510180,China;Department of Endocrinology and Metabolism,Sixth Affiliated Hospital of Sun Yat-sen University,Guangzhou 510655,China;Department of Endocrinology,Second Affiliated Hospital of Guangzhou Medical University,Guangzhou 510260,China;Department of Endocrinology and Metabolism,Shunde Hospital of Southern Medical University(First People′s Hospital of Shunde),Foshan 528300,China)

机构地区:[1]中山大学附属第三医院内分泌与代谢病科,广东省糖尿病防治重点实验室,广州510630 [2]广州市第一人民医院,广州510180 [3]中山大学附属第六医院内分泌与代谢病科,广州510655 [4]广州医科大学附属第二医院内分泌科,广州510260 [5]南方医科大学顺德医院(顺德区第一人民医院)内分泌与代谢病科,佛山528300

出  处:《中华糖尿病杂志》2024年第6期680-690,共11页CHINESE JOURNAL OF DIABETES MELLITUS

基  金:广东省自然科学基金(2022A1515012364);广州市科技计划项目(202102010175,202201020550)。

摘  要:目的基于全转录组测序技术,探讨母鼠孕期和哺乳期低蛋白饮食对子代雌鼠胰岛功能的影响及其可能机制。方法33只C57BL/6J雌鼠受孕当日按照随机数表法分为两组,在整个孕期及哺乳期分别予以对照饮食(蛋白质22%)(13只)和低蛋白饮食(20只)(蛋白质9%)干预。哺乳期结束后,所有子代予以标准饮食。根据母代的喂养方式,将其雌性子代分为对照饮食组(CD组,20只)和低蛋白饮食组(LPD组,20只)。收集子代雌鼠体重和空腹血糖(FPG),采用腹腔内注射葡萄糖耐量实验(IPGTT)及同步血清胰岛素释放实验(IRT)、腹腔内注射胰岛素耐量实验(IPITT)评估子代胰岛功能及其胰岛素敏感性,采用全转录组测序法对两组胰岛组织中差异表达的长链非编码RNA(LncRNA)进行靶基因预测,采用生物信息学分析法对这些靶基因与测序得到的差异信使RNA(mRNA)进行分析,采用实时荧光定量PCR(qRT-PCR)及蛋白免疫印迹法对生信分析结果进行实验验证,对mRNA及LncRNA预测靶基因的共同差异基因乳酸脱氢酶A(Ldha)进行免疫荧光染色并构建LncRNA Gm38850敲减的Min6细胞模型以研究Gm38850对胰岛功能的影响及其与Ldha的调控关系。采用两独立样本t检验或校正t检验进行组间比较。结果与CD组相比,LPD组子代雌鼠在第17周龄时出现糖耐量受损(P<0.05)和胰岛素释放减少(P<0.05),但两组间空腹血糖及IPITT血糖的曲线下面积差异均无统计学意义(P>0.05)。全转录组测序筛选出两组子代雌鼠胰岛细胞间差异表达mRNA共55个(42个上调,13个下调);差异表达LncRNA共4个(均下调)。通路富集分析提示,这些差异表达RNA主要与丙酮酸代谢、白细胞介素-17信号通路、胰高糖素途径、晚期糖基化终产物及其受体(AGE-RAGE)信号通路及缺氧诱导因子-1(HIF-1)信号通路等生物学功能相关。胰腺组织qRT-PCR验证测序差异表达倍数前9的mRNA表达情况,结果与生信分析结果趋势�Objective To investigate how a low-protein diet during pregnancy and lactation affects the islet function in the female offspring mice,and to explore the potential mechanisms using transcriptome sequencing technology.Methods Thirty-three female C57BL/6J mice were randomly divided into two groups according to a random number table method on the day of conception and given a control diet(22%protein)(13 mice)and a low-protein diet(9%protein)(20 mice)throughout pregnancy and lactation,respectively.After weaning,all offspring were given a standard diet.The female offspring were divided into the control diet group(CD group,20 mice)and the low protein diet group(LPD group,20 mice)based on their maternal diet.Body weight and fasting plasma glucose(FPG)data of the female offspring were collected.Intraperitoneal glucose tolerance test(IPGTT),simultaneous serum insulin release test(IRT)and intraperitoneal insulin tolerance test(IPITT)were performed to assess the function of the offspring islets and their insulin sensitivity.Long non-cody RNA(LncRNA)from the offspring primary islets were extracted for transcriptome sequencing.The bioinformatics analysis applied between the targeted genes and the differentially expressed messenger RNA(mRNA).Experimental verification of biosignature analysis was performed using quantitative real-time PCR(qRT-PCR)and Western blotting.To verify the expression of(lactate dehydrogenase A)Ldha,a common differential gene of mRNA and LncRNA predicted target genes,we performed immunofluorescence staining on paraffin sections of pancreatic tissue.Thereafter,LncRNA Gm38850 knockdown Min6 cell model was constructed to study the effect of Gm38850 on islet function and its regulatory relationship with Ldha.Group comparisons were conducted using either independent samples t-test or corrected t-test.Results The LPD group showed impaired glucose tolerance(P<0.05)and reduced insulin release(P<0.05)at week 17 of age compared to the CD group.However,no significant differences were observed in FPG and blood gluco

关 键 词:转录组测序 低蛋白饮食 胰岛功能 

分 类 号:R587.1[医药卫生—内分泌] R-332[医药卫生—内科学]

 

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