文献类型：期刊文献

中文题名：诱导大鼠胰腺导管干细胞分化形成胰岛素分泌细胞基因表达谱的研究

英文题名：Research on the gene expression profile of inducing pancreatic duct stem cells in rats to differentiate into insulin-secreting cells

作者：任凯[1];郇月蓉[1];吴江[1];韩梦瑶[1];周光现[1];孙平平[1];效梅[1]

机构：[1]广东海洋大学滨海农业学院动物医学系,湛江524088

年份：2025

卷号：33

期号：6

起止页码：449

中文期刊名：中国糖尿病杂志

外文期刊名：Chinese Journal of Diabetes

收录：北大核心2023、、北大核心

基金：广东省自然科学基金(10152408801000023);国家级大学生创新创业训练计划项目(201810566015);留学回国人员科研启动基金(教外司留[2010]1561号)。

语种：中文

中文关键词：胰腺导管干细胞;胰岛素分泌细胞;诱导分化;转录组分析

外文关键词：Pancreatic ductal stem cells;Insulin-secreting cell;Induced differentiation;Transcriptome analysis

中文摘要：目的 探讨诱导大鼠胰腺导管干细胞(PDSCs)分化为胰岛素分泌细胞(IPCs)的基因表达谱,并筛选参与诱导分化的关键基因。方法 扩增PDSCs分为正常对照(NC)组和诱导(Tre)组。NC组继续进行扩增培养,Tre组采用诱导液培养28 d定向分化PDSCs为IPCs。采用双硫腙(DTZ)染色观察细胞阳性结果。采用免疫荧光染色法检测诱导后细胞Ins和胰腺十二指肠同源盒基因1(PDX1)表达。采用ELISA检测IPCs的Ins释放量,验证诱导后的细胞对葡萄糖刺激的Ins分泌反应性。同时收集诱导0、28 d的细胞进行RNA-seq,分析差异表达基因(DEGs)并进行功能注释,结果发现调节因子X3(RFX3)在PDSCs中过表达,并验证上调RFX3对诱导分化的影响。结果 与NC组比较,Tre组DTZ染色呈阳性,表达PDX1和Ins,葡萄糖刺激下Ins释放量提高。RNA-seq分析鉴定DEGs 4270个,基因本体和京都基因与基因组百科全书数据库功能富集分析结果显示,与Ins反应和分泌正向调节、胰腺内分泌细胞发育和胰腺发育等功能相关的基因,如乙醛脱氢酶2、cAMP反应元件结合蛋白5、真核生物翻译起始因子6、叉头框转录因子O1(FOXO1)、RFX3、无翅型MMTV整合位点家族成员5a、氧连接N-乙酰葡萄糖胺转移酶、G蛋白偶联受体39、SMAD家族成员6和瞬时受体电位M2通道等,细胞周期、TGF-β1信号通路、FOXO信号通路和Wnt信号通路等均参与PDSCs向IPCs分化的调控。上调RFX3在72 h内可抑制TGF-β1表达,促进Ins阳性细胞形成和释放Ins。结论 多个与胰岛β细胞功能相关的基因及信号通路共同调控大鼠PDSCs分化形成IPCs,其中RFX3上调可促进分化过程。

外文摘要：Objective To investigate the gene expression profile in rat pancreatic ductal stem cells(PDSCs)when induced to differentiate into insulin-secreting cells(IPCs),with the goal of identifying key genes involved in this differentiation process.Methods The expanded PDSCs were categorized into a normal control(NC)group and an induced(Tre)group.PDSCs continued expansion culture in NC group,and cultured in induction medium for 28 days to facilitate the differentiation of PDSCs into IPCs in Tre group.Dithizone staining was employed to morphologically assess whether the cells exhibited a reddish-brown coloration,indicating a positive result.The immunofluorescence staining method was used to detect the expression of insulin(Ins)and PDX1 in the cells following induction.Additionally,ELISA was conducted to measure the Ins release from IPCs,thereby verifying the responsiveness of the induced cells to glucose-stimulated Ins secretion.Concurrently,cells were collected on induction days 0 and 28 for RNA sequencing(RNA-seq),and differentially expressed genes(DEGs)were analyzed and functionally annotated.The analysis revealed that regulatory factor X3(RFX3)was overexpressed in PDSCs,and the impact of RFX3 upregulation on differentiation induction was subsequently verified.Results Compared with NC group,DTZ staining was positive,PDX1 and Ins proteins were expressed,and an increased release of Ins in response to sugar stimulation was demonstrated in the Tre group.RNA-seq analysis identified 4270 DEGs,and functional enrichment analysis utilizing the Gene Ontology and Kyoto Encyclopedia of Genes and Genomes databases revealed associations with Ins response,positive regulation of Ins secretion,pancreatic endocrine cell development,and overall pancreatic development.Additionally,functionally related genes such as ALDHA2,CREB5,EIF6,FOXO1,RFX3,WNT5a,OGT,GPR39,SMAD6,and TRPM2 were identified,indicating involvement in the cell cycle,TGF-β1 signaling pathway,FOXO signaling pathway,and Wnt signaling pathway in the regulation of the differentiation of pancreatic ductal stem cells(PDSCs)into insulin-producing cells(IPCs).Furthermore,the upregulation of RFX3 can inhibit the expression of TGF-β1 within 72 hours,thereby promoted the formation and release of Ins from insulin-positive cells.Conclusions Multiple genes and signaling pathways associated with pancreaticβ-cell function collectively regulate the differentiation of rat PDSCs into IPCs.Notably,the upregulation of RFX3 enhances this differentiation process.