脐带间充质干细胞通过分泌吲哚胺2,3过

译者：韩健校对：马丹审核：张莉芸

摘要：

目的：研究脐带间充质干细胞(UC-MSCs)对原发性干燥综合征(pSS)患者滤泡辅助性T细胞(cTfh)的作用。

方法：使用流式细胞术分析外周血单核细胞(PBMCs)中CXCR5(+)PD-1(+)CD4(+)T细胞的比例。根据细胞间接触原理或双室细胞培养法将PBMCs与UC-MSCs进行共培养。在Tfh细胞处于细胞极化环境下将从PBMCs分离出的幼粒CD4(+)T细胞与UC-MSCs进行共培养。使用流式细胞术计算CXCR5(+)PD-1(+)CD4(+)T细胞、二醋酸盐琥珀酰亚胺酯(CFSE)荧光强度及膜联蛋白V的比例。使用实时PCR及Luminex细胞因子化验分析mRNA表达及上清蛋白水平。使用HPLC计算吲哚胺2,3-过氧化酶(IDO)活性。

结果：pSS患者的cTfh细胞表达增加并且和血清抗La/SSB抗体水平及欧洲风湿病联盟SS疾病活动指数评分呈正相关。在体外实验中，UC-MSCs抑制了cTfh细胞的增值和分化。实时PCR分析表明UC-MSCs与幼粒CD4(+)T细胞共培养时IDOmRNA表达明显增加。然而，在双室细胞培养法中当UC-MSCs与幼粒CD4(+)T细胞共培养时IDOmRNA表达仅有少量增加。此外，HPLC表明在UC-MSCs与幼粒CD4(+)T细胞共培养基的上清液中IDO酶活性增加。IDO抑制剂1-MT的增加一定程度上降低了UC-MSCs对cTfh细胞分化的抑制作用。

结论：UC-MSCs通过分泌IDO抑制cTfh细胞的分化，由活性CD4(+)T细胞分泌的可溶性因子可以促进UC-MSCs分泌IDO。

Abstract：

OBJECTIVE:Theaimofthisstudywastoinvestigatetheeffectofumbilicalcordmesenchymalstemcells(UC-MSCs)oncirculatingTfollicularhelper(cTfh)cellsinprimarySS(pSS)patients.

METHODS:ThepercentageofCXCR5(+)PD-1(+)CD4(+)Tcellsinperipheralbloodmononuclearcells(PBMCs)wasanalysedbyflowcytometry.PBMCswereco-culturedwithUC-MSCsbycell-to-cellcontactorinatrans-wellsystem.NaiveCD4(+)TcellswereisolatedfromPBMCsandthenco-culturedwithUC-MSCsunderTfhcell-polarizingconditions.ThepercentageofCXCR5(+)PD-1(+)CD4(+)Tcells,carboxyfluoresceinsuccinimidylester(CFSE)fluorescenceintensityandannexinVweredeterminedbyflowcytometricanalysis.Real-timePCRandLuminexcytokineassaywereperformedtodetectmRNAexpressionandsupernatantproteinlevels.Theactivityofindoleamine2,3-dioxygenase(IDO)wasmeasuredbyHPLC.

RESULTS:IncreasedfrequencyofcTfhcellswasfoundinpSSpatientsandwaspositivelycorrelatedwithserumanti-La/SSBlevelsandtheEuropeanLeagueAgainstRheumatismSSDiseaseActivityIndexscore.Invitro,UC-MSCssuppressedthedifferentiationandproliferationofcTfhcells.Real-timePCRanalysisshowedsignificantlyhigherIDOmRNAexpressiononUC-MSCswhenco-culturedwithnaiveCD4(+)TcellsunderTfhcell-polarizingconditionsinpSSpatients.However,IDOmRNAexpressiononUC-MSCswasonlyalittlehigherwhenUC-MSCswereco-culturedwithnaiveCD4(+)Tcellsinatrans-wellsystem.Inaddition,HPLCshowedincreasedIDOenzymicactivityinthesupernatantofUC-MSCsco-culturedwithnaiveCD4(+)TcellsunderTfhcell-polarizingconditionsinpSS.TheadditionoftheIDOinhibitor1-MTpartlyreversedthesuppressiveeffectofUC-MSCsonthedifferentiationofcTfhcells.

CONCLUSION:TheseresultssuggestaninhibitoryeffectofUC-MSCsonthedifferentiationofcTfhcellsviathesecretionofIDO,andsolublefactorssecretedbyactivatedCD4(+)TcellsmightcontributetoIDOsecretionbyUC-MSCs.

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