中国医药
中國醫藥
중국의약
CHINA MEDICINE
2014年
12期
1842-1847
,共6页
糖尿病%人骨髓间充质干细胞%分泌胰岛素细胞%胰高血糖素样肽1
糖尿病%人骨髓間充質榦細胞%分泌胰島素細胞%胰高血糖素樣肽1
당뇨병%인골수간충질간세포%분비이도소세포%이고혈당소양태1
Diabetes mellitus%Bone marrow mesenchymal stem cells%Insulin-producing cells%Glueagon like peptide-1
目的 探讨并优化人骨髓间充质干细胞(hBMSCs)向分泌胰岛素细胞(IPCs)定向分化的条件,为糖尿病替代疗法寻找新出路.方法 hBMSCs复苏培养后,联合应用胰高血糖素样肽1(GLP-1)、Ex-tentin-4等细胞因子通过三步诱导方案进行体外向IPCs诱导分化.应用反转录-聚合酶链反应检测与β细胞发育和功能相关的基因表达;免疫细胞化学、双硫腙染色证实IPCs的生成;电化学发光法检测细胞胰岛素的分泌量.结果 诱导分化18d后,镜下观察到胰岛样细胞团的生成,双硫腙及免疫细胞化学染色胰岛素均呈现阳性反应;诱导分化的细胞在第18天则可以观察到胰十二指肠同源盒基因1、胰岛素的高表达;诱导分化的细胞接受葡萄糖刺激后能够分泌胰岛素,且胰岛素的分泌量随葡萄糖浓度的提高而增加,高糖胰岛素分泌量[(188.7±1.5)mU/L]与低糖胰岛素分泌量[(60.1±0.9)mU/L]相比,差异有统计学意义(P＜0.05).结论 体外联合应用GLP-l、Extentin-4、尼克酰胺等细胞因子采用三阶段诱导方案可以大大提高胰岛素细胞诱导分化率,并能促进IPCs的成熟和胰岛素的释放.
目的 探討併優化人骨髓間充質榦細胞(hBMSCs)嚮分泌胰島素細胞(IPCs)定嚮分化的條件,為糖尿病替代療法尋找新齣路.方法 hBMSCs複囌培養後,聯閤應用胰高血糖素樣肽1(GLP-1)、Ex-tentin-4等細胞因子通過三步誘導方案進行體外嚮IPCs誘導分化.應用反轉錄-聚閤酶鏈反應檢測與β細胞髮育和功能相關的基因錶達;免疫細胞化學、雙硫腙染色證實IPCs的生成;電化學髮光法檢測細胞胰島素的分泌量.結果 誘導分化18d後,鏡下觀察到胰島樣細胞糰的生成,雙硫腙及免疫細胞化學染色胰島素均呈現暘性反應;誘導分化的細胞在第18天則可以觀察到胰十二指腸同源盒基因1、胰島素的高錶達;誘導分化的細胞接受葡萄糖刺激後能夠分泌胰島素,且胰島素的分泌量隨葡萄糖濃度的提高而增加,高糖胰島素分泌量[(188.7±1.5)mU/L]與低糖胰島素分泌量[(60.1±0.9)mU/L]相比,差異有統計學意義(P＜0.05).結論 體外聯閤應用GLP-l、Extentin-4、尼剋酰胺等細胞因子採用三階段誘導方案可以大大提高胰島素細胞誘導分化率,併能促進IPCs的成熟和胰島素的釋放.
목적 탐토병우화인골수간충질간세포(hBMSCs)향분비이도소세포(IPCs)정향분화적조건,위당뇨병체대요법심조신출로.방법 hBMSCs복소배양후,연합응용이고혈당소양태1(GLP-1)、Ex-tentin-4등세포인자통과삼보유도방안진행체외향IPCs유도분화.응용반전록-취합매련반응검측여β세포발육화공능상관적기인표체;면역세포화학、쌍류종염색증실IPCs적생성;전화학발광법검측세포이도소적분비량.결과 유도분화18d후,경하관찰도이도양세포단적생성,쌍류종급면역세포화학염색이도소균정현양성반응;유도분화적세포재제18천칙가이관찰도이십이지장동원합기인1、이도소적고표체;유도분화적세포접수포도당자격후능구분비이도소,차이도소적분비량수포도당농도적제고이증가,고당이도소분비량[(188.7±1.5)mU/L]여저당이도소분비량[(60.1±0.9)mU/L]상비,차이유통계학의의(P＜0.05).결론 체외연합응용GLP-l、Extentin-4、니극선알등세포인자채용삼계단유도방안가이대대제고이도소세포유도분화솔,병능촉진IPCs적성숙화이도소적석방.
Objective To explore an effective and appropriate condition of inducing human bone marrow mesenchymal stem cells(hBMSCs) into insulin-producing cells.Methods hBMSCs were induced to insulin producing cells(IPCs) by glueagon like peptide-1 (GLP-l),extentin-4 and the additions of many cell stimulating growth factors according to a three-stage protocol.The expression of multiple genes related to Panereatic β-cell development and function was detected by RT-PCR.The identity of the IPCs was illustrated by the analysis of morphology,ditizone staining and immunocytochemistry and release of insulin.Results Typical islet-like cell clusters were observed at the end of Protocol (18 days).Ditizone staining and immunocytochemistry for insulin were both positive.These differentiated cells at the end of Protocol expressed PDX-l mRNA and insulin mRNA.Insulin was secreted by these cells in response to different concentration of glucose stimulation.In the situation of the incubation with high glucose solution(16.7 mmol/L),the insulin secretion in the induced group showed significantly increased as compared with that in the control group [(188.7 ± 1.5) mU/L vs(60.1 ± 0.9) mU/L,P ＜ 0.05].Conclusion With combined GLP-l and extentin-4,hBMSCs can improve the rate of insulin-producing cells differentiation,promote IPCs maturation and release of insulin.
