广东医学
廣東醫學
엄동의학
Guangdong Medical Journal
2015年
14期
2128-2131,2132
,共5页
肝细胞%小分子化合物%胰岛素分泌细胞%糖尿病
肝細胞%小分子化閤物%胰島素分泌細胞%糖尿病
간세포%소분자화합물%이도소분비세포%당뇨병
hepatocytes%small molecules%insulin-producing cells%diabetes mellitus
目的:分离原代小鼠肝细胞,在小分子化合物作用下将其诱导分化为胰岛素分泌细胞( IPCs )。方法采用肝脏组织直接剪碎的方法分离、培养小鼠肝细胞,并进行肝细胞糖原染色鉴定。随后将肝细胞分别用不同的诱导剂5-氮杂胞苷(5-AZA)、曲古霉素A(TSA)、维甲酸(RA)、胰岛素-转铁蛋白-硒(ITS)、尼克酰胺( NA )及不同糖浓度的培养液进行诱导,同时以不含诱导剂的培养液培养肝细胞作为阴性对照。结果原代肝细胞在5-AZA、TSA的作用下去分化,RT-qPCR显示成熟肝细胞表面标志物白蛋白明显下降,出现巢蛋白( nestin)表达阳性细胞,进一步在含RA、ITS的低糖培养液培养7 d及含NA的低糖培养液培养7 d,分别获得在基因及蛋白水平都表达的Pdx1及胰岛素阳性细胞。结论小分子化合物(5-AZA、TSA、RA、ITS、NA)及依次使用高浓度和低浓度葡萄糖可以在短时期内(17 d)将肝细胞诱导为胰腺前体细胞(Pdx1阳性细胞),并进一步分化为IPCs。
目的:分離原代小鼠肝細胞,在小分子化閤物作用下將其誘導分化為胰島素分泌細胞( IPCs )。方法採用肝髒組織直接剪碎的方法分離、培養小鼠肝細胞,併進行肝細胞糖原染色鑒定。隨後將肝細胞分彆用不同的誘導劑5-氮雜胞苷(5-AZA)、麯古黴素A(TSA)、維甲痠(RA)、胰島素-轉鐵蛋白-硒(ITS)、尼剋酰胺( NA )及不同糖濃度的培養液進行誘導,同時以不含誘導劑的培養液培養肝細胞作為陰性對照。結果原代肝細胞在5-AZA、TSA的作用下去分化,RT-qPCR顯示成熟肝細胞錶麵標誌物白蛋白明顯下降,齣現巢蛋白( nestin)錶達暘性細胞,進一步在含RA、ITS的低糖培養液培養7 d及含NA的低糖培養液培養7 d,分彆穫得在基因及蛋白水平都錶達的Pdx1及胰島素暘性細胞。結論小分子化閤物(5-AZA、TSA、RA、ITS、NA)及依次使用高濃度和低濃度葡萄糖可以在短時期內(17 d)將肝細胞誘導為胰腺前體細胞(Pdx1暘性細胞),併進一步分化為IPCs。
목적:분리원대소서간세포,재소분자화합물작용하장기유도분화위이도소분비세포( IPCs )。방법채용간장조직직접전쇄적방법분리、배양소서간세포,병진행간세포당원염색감정。수후장간세포분별용불동적유도제5-담잡포감(5-AZA)、곡고매소A(TSA)、유갑산(RA)、이도소-전철단백-서(ITS)、니극선알( NA )급불동당농도적배양액진행유도,동시이불함유도제적배양액배양간세포작위음성대조。결과원대간세포재5-AZA、TSA적작용하거분화,RT-qPCR현시성숙간세포표면표지물백단백명현하강,출현소단백( nestin)표체양성세포,진일보재함RA、ITS적저당배양액배양7 d급함NA적저당배양액배양7 d,분별획득재기인급단백수평도표체적Pdx1급이도소양성세포。결론소분자화합물(5-AZA、TSA、RA、ITS、NA)급의차사용고농도화저농도포도당가이재단시기내(17 d)장간세포유도위이선전체세포(Pdx1양성세포),병진일보분화위IPCs。
Objective To induce differentiation of isolated primary mouse hepatocytes into insulin-producing cells ( IPCs) with small molecules.Methods Mouse hepatocytes were isolated and cultured from minced mice livers be-fore being identified by glycogen staining.They were subsequently treated with the following inducers, 5-aza-2′-de-oxycytidine (5-AZA), richostatin A (TSA), retinoic acid (RA), insulin-transferrin-selenium (ITS), nicotinamide ( NA) and with culture media of different glucose concentration.Hepatocytes cultured without inducers were served as the negative control.Results Primary hepatocytes underwent dedifferentiation in the effect of 5-AZA and TSA.RT-qPCR demonstrated significant decrease of albumin, a surface marker of mature hepatocytes, along with nestin-positive cells. Further culture with low glucose medium containing either RA plus ITS or NA alone for 7 d yielded Pdx1 and insulin posi-tive cells at both gene and protein levels.Conclusion Small molecules (5-AZA, TSA, RA, ITS, NA) and the se-quential use of high and low concentration of glucose can induce hepatocytes into pancreatic precursor cells, and further differentiate into insulin-producing cells in a short period (17 d) .