CAJ | 학술논문

目的观察OX40/OX40L共刺激通路在调节性T细胞(Treg)诱导移植耐受中的作用.方法建立小鼠胰岛移植模型,分3组:IgG组、抗-OX40L单抗(RM134L)+抗-CD154单抗( MR1)组、联合RM134L+ MR1及抗-CD25单抗(PC61)组.存活超过150 d小鼠切除移植侧肾脏,在对侧行同或不同主要组织相容性复合体( MHC)的2次胰岛移植.观察各组平均存活时间(MST).检测效应性T细胞(Teff)、野生型和CD154敲除小鼠Treg OX40表达.检测同基因小鼠胰岛、胰岛移植不加或加RM134L+ MR1治疗的移植物内Foxp3表达.Treg与Teff加或不加OX40激动剂(OX86)以不同比例共培养,检测Teff增殖抑制情况.结果 RM134L+ MR1组MST为＞150 d(n =8),较IgG组(19d,n=5)和联合PC61组(23 d,n=4)明显延长(P＜0.05).同或不同MHC的2次胰岛移植MST分别为＞60 d(n=3)和17 d(n =3) (P ＜0.05).OX40在Teff不表达,在野生型和CD154敲除小鼠Treg表达.3组小鼠胰岛移植物内Foxp3相对表达量分别为8、21、123 AU(P ＜0.05).Teff与Treg以1∶2和1∶4共培养,β-液体闪烁计数仪测定Teff的核素每分钟计数(CPM)分别为63×103和41×103,与对照组(140×103)比较增殖受抑制(P＜0.05),加OX86时CPM值分别为128×103和135×103 (P ＞0.05).结论 CD4+ CD25+ Treg在诱导移植耐受中发挥重要作用,OX40在其中发挥负性调节作用.
목적 관찰OX40/OX40L공자격통로재조절성T세포(Treg)유도이식내수중적작용.방법 건립소서이도이식모형,분3조:IgG조、항-OX40L단항(RM134L)+항-CD154단항( MR1)조、연합RM134L+ MR1급항-CD25단항(PC61)조.존활초과150 d소서절제이식측신장,재대측행동혹불동주요조직상용성복합체( MHC)적2차이도이식.관찰각조평균존활시간(MST).검측효응성T세포(Teff)、야생형화CD154고제소서Treg OX40표체.검측동기인소서이도、이도이식불가혹가RM134L+ MR1치료적이식물내Foxp3표체.Treg여Teff가혹불가OX40격동제(OX86)이불동비례공배양,검측Teff증식억제정황.결과 RM134L+ MR1조MST위＞150 d(n =8),교IgG조(19d,n=5)화연합PC61조(23 d,n=4)명현연장(P＜0.05).동혹불동MHC적2차이도이식MST분별위＞60 d(n=3)화17 d(n =3) (P ＜0.05).OX40재Teff불표체,재야생형화CD154고제소서Treg표체.3조소서이도이식물내Foxp3상대표체량분별위8、21、123 AU(P ＜0.05).Teff여Treg이1∶2화1∶4공배양,β-액체섬삭계수의측정Teff적핵소매분종계수(CPM)분별위63×103화41×103,여대조조(140×103)비교증식수억제(P＜0.05),가OX86시CPM치분별위128×103화135×103 (P ＞0.05).결론 CD4+ CD25+ Treg재유도이식내수중발휘중요작용,OX40재기중발휘부성조절작용.
Objective To observe the role of OX40/OX40L costimulation pathway in the CD4+CD25 + regulatory T (Treg) cells-induced tolerance of islet transplantation.Methods Diabetes mellitus was induced in C57BL/6 mice as recipients,and islets from DBA/2 mice were transplanted into the C57BL/6 mice.The recipients were divided into three groups (n =8):treated with IgG as controls,antiOX40L mAb (RM134L) + anti-CD154 mAb (MR1),and RM134L + MR1 + anti-CD25 mAb.The mean survival time (MST) of allograft was observed.The expression of OX40 in T effector cells,Treg cells of wide type and CD154 knock-out mice was detected.The expression of Foxp3 gene in allograft in the absence and absence of anti-OX40L mAb and anti-CD154 mAb was detected.T effector cells and Treg cells were co-cultured at different ratio,and the cell proliferation was assayed.Results The MST in RM134L +MR1 group was ＞ 150 days (n =8),significantly longer than in IgG group ( MST + 19 days,n =5 ) and RM134L + MR1 + anti-CD25 mAb (23 days,n =4) (P ＜0.05).Anti-OX40L mAb and anti-CD154 mAb treatment could successfully induce tolerance,and also induce donor specific tolerance,but the tolerance was blocked by anti-CD25 mAb treatment (P ＜ 0.05 ).OX40 was expressed on the Treg cells surface,whereas the T effector cells had no OX40 expression.The expression level of Foxp3 gene in the allograft treated with anti-OX40L mAb and anti-CD154 mAb was 123 AU,significantly higher than without treatment (21 AU) and syngenic control ( 8 AU) ( P ＜ 0.05 ).After Treg cells were co-cultured with T effector cells at different ratio ( 1∶2 and 1∶ 4),the mean count per minute (CPM) of T effector cells was 63 x 103and 41 × 103 respectively (P ＜0.05),and after addition of OX40,the CPM was 128 × 103 and 135 × 103respectively ( P ＞ 0.05 ).Conclusion CD4 + CD25 + Treg cells were critical to tolerance induction in mice islet transplant recipients through targeting OX40/OX40L costimulation pathway.