中华实验外科杂志
中華實驗外科雜誌
중화실험외과잡지
CHINESE JOURNAL OF EXPERIMENTAL SURGERY
2011年
10期
1670-1673
,共4页
树突状细胞%免疫耐受%FasL%转化生长因子-β1%肝移植
樹突狀細胞%免疫耐受%FasL%轉化生長因子-β1%肝移植
수돌상세포%면역내수%FasL%전화생장인자-β1%간이식
Dendritic cells%Immune tolerance%FasL%TGF-β1%Liver transplantation
目的 联合人凋亡相关因子配体(hFasL)及人转化生长因子-β1( hTGF-β1)基因共修饰大鼠骨髓来源的未成熟树突状细胞( imDC),提高imDC诱导大鼠同种异体原位肝移植免疫耐受的能力,以延长受鼠生存期.方法 行以DA大鼠为供体、近交系Lewis大鼠为受体的同种异体原位肝移植140例,随机均分为7组:未给移植受鼠注射DC组(对照组)、mDC组、imDC组、空载体组、FasL组、TGF组及共转染组；分别于移植前5d每天给Lewis大鼠腹腔注射mDC、imDC、空载体转染的imDC、hFasL修饰的imDC、hTGF-β1修饰的imDC、hFasL和hTGF-β1共修饰的imDC各2×106个,于肝移植术后3、7、10d各处死4只,观察外周血肝功能变化[丙氨酸转氨酶(ALT)和总胆红素(TBIL)],肝脏病理改变,肝脏凋亡,外周血清细胞因子,并进行生存分析.结果 肝脏病理示术后7d,7组差异有统计学意义(P＜0.01),共转染组排斥反应均轻于TGF组和FasL组；10d时,7组差异有统计学意义(P＜0.01),共转染组排斥反应轻于TGF组和FasL组,TGF组轻于FasL组.肝功能示共转染组于7d时就已基本恢复正常,FasL组于7d时有所降低,但10 d时反而升高,ALT及TBIL的浓度均高于共转染组(P＜0.01,P＜0.05)；TGF组到10 d时,肝功能已恢复正常,ALT及TBIL的浓度均低于FasL组(P＜0.01,P＜0.05),但与共转染组比较,差异无统计学意义(P＞0.05).酶联免疫吸附试验(ELISA)检测示,术后7d共转染组和TGF组的血清白细胞介素(IL)-1、IL-10及IL-12与FasL组比较,差异均有统计学意义(P＜0.05)；但共转染组和TGF组之间差异无统计学意义(P＞0.05).TUNEL显示FasL组的肝、脾及腹腔淋巴结中淋巴细胞凋亡指数与共转染组比较,差异均无统计学意义(P＞0.05),但多于TGF组,差异均有统计学意义(P＜0.05).FasL组中位生存期20 d与imDC组(23 d)差异无统计学意义(P＞0.05),而TGF组延长受鼠生存时间有限(48 d)；只有共转染组出现长期存活的受鼠(＞90 d),均长于FasL组和TGF(P＜0.01,P=0.01).结论 imDC、hFasL或hTGF-β1单基因修饰imDC均未能诱导同种异体大鼠肝移植长期存活；hFasL 和hTGF-β1共转染的imDC诱导同种异体大鼠肝移植免疫耐受的能力显著增强,显著延长同种异体肝移植大鼠的生存期.
目的 聯閤人凋亡相關因子配體(hFasL)及人轉化生長因子-β1( hTGF-β1)基因共脩飾大鼠骨髓來源的未成熟樹突狀細胞( imDC),提高imDC誘導大鼠同種異體原位肝移植免疫耐受的能力,以延長受鼠生存期.方法 行以DA大鼠為供體、近交繫Lewis大鼠為受體的同種異體原位肝移植140例,隨機均分為7組:未給移植受鼠註射DC組(對照組)、mDC組、imDC組、空載體組、FasL組、TGF組及共轉染組；分彆于移植前5d每天給Lewis大鼠腹腔註射mDC、imDC、空載體轉染的imDC、hFasL脩飾的imDC、hTGF-β1脩飾的imDC、hFasL和hTGF-β1共脩飾的imDC各2×106箇,于肝移植術後3、7、10d各處死4隻,觀察外週血肝功能變化[丙氨痠轉氨酶(ALT)和總膽紅素(TBIL)],肝髒病理改變,肝髒凋亡,外週血清細胞因子,併進行生存分析.結果 肝髒病理示術後7d,7組差異有統計學意義(P＜0.01),共轉染組排斥反應均輕于TGF組和FasL組；10d時,7組差異有統計學意義(P＜0.01),共轉染組排斥反應輕于TGF組和FasL組,TGF組輕于FasL組.肝功能示共轉染組于7d時就已基本恢複正常,FasL組于7d時有所降低,但10 d時反而升高,ALT及TBIL的濃度均高于共轉染組(P＜0.01,P＜0.05)；TGF組到10 d時,肝功能已恢複正常,ALT及TBIL的濃度均低于FasL組(P＜0.01,P＜0.05),但與共轉染組比較,差異無統計學意義(P＞0.05).酶聯免疫吸附試驗(ELISA)檢測示,術後7d共轉染組和TGF組的血清白細胞介素(IL)-1、IL-10及IL-12與FasL組比較,差異均有統計學意義(P＜0.05)；但共轉染組和TGF組之間差異無統計學意義(P＞0.05).TUNEL顯示FasL組的肝、脾及腹腔淋巴結中淋巴細胞凋亡指數與共轉染組比較,差異均無統計學意義(P＞0.05),但多于TGF組,差異均有統計學意義(P＜0.05).FasL組中位生存期20 d與imDC組(23 d)差異無統計學意義(P＞0.05),而TGF組延長受鼠生存時間有限(48 d)；隻有共轉染組齣現長期存活的受鼠(＞90 d),均長于FasL組和TGF(P＜0.01,P=0.01).結論 imDC、hFasL或hTGF-β1單基因脩飾imDC均未能誘導同種異體大鼠肝移植長期存活；hFasL 和hTGF-β1共轉染的imDC誘導同種異體大鼠肝移植免疫耐受的能力顯著增彊,顯著延長同種異體肝移植大鼠的生存期.
목적 연합인조망상관인자배체(hFasL)급인전화생장인자-β1( hTGF-β1)기인공수식대서골수래원적미성숙수돌상세포( imDC),제고imDC유도대서동충이체원위간이식면역내수적능력,이연장수서생존기.방법 행이DA대서위공체、근교계Lewis대서위수체적동충이체원위간이식140례,수궤균분위7조:미급이식수서주사DC조(대조조)、mDC조、imDC조、공재체조、FasL조、TGF조급공전염조；분별우이식전5d매천급Lewis대서복강주사mDC、imDC、공재체전염적imDC、hFasL수식적imDC、hTGF-β1수식적imDC、hFasL화hTGF-β1공수식적imDC각2×106개,우간이식술후3、7、10d각처사4지,관찰외주혈간공능변화[병안산전안매(ALT)화총담홍소(TBIL)],간장병리개변,간장조망,외주혈청세포인자,병진행생존분석.결과 간장병리시술후7d,7조차이유통계학의의(P＜0.01),공전염조배척반응균경우TGF조화FasL조；10d시,7조차이유통계학의의(P＜0.01),공전염조배척반응경우TGF조화FasL조,TGF조경우FasL조.간공능시공전염조우7d시취이기본회복정상,FasL조우7d시유소강저,단10 d시반이승고,ALT급TBIL적농도균고우공전염조(P＜0.01,P＜0.05)；TGF조도10 d시,간공능이회복정상,ALT급TBIL적농도균저우FasL조(P＜0.01,P＜0.05),단여공전염조비교,차이무통계학의의(P＞0.05).매련면역흡부시험(ELISA)검측시,술후7d공전염조화TGF조적혈청백세포개소(IL)-1、IL-10급IL-12여FasL조비교,차이균유통계학의의(P＜0.05)；단공전염조화TGF조지간차이무통계학의의(P＞0.05).TUNEL현시FasL조적간、비급복강림파결중림파세포조망지수여공전염조비교,차이균무통계학의의(P＞0.05),단다우TGF조,차이균유통계학의의(P＜0.05).FasL조중위생존기20 d여imDC조(23 d)차이무통계학의의(P＞0.05),이TGF조연장수서생존시간유한(48 d)；지유공전염조출현장기존활적수서(＞90 d),균장우FasL조화TGF(P＜0.01,P=0.01).결론 imDC、hFasL혹hTGF-β1단기인수식imDC균미능유도동충이체대서간이식장기존활；hFasL 화hTGF-β1공전염적imDC유도동충이체대서간이식면역내수적능력현저증강,현저연장동충이체간이식대서적생존기.
Objective To improve immature dendritic cell ( imDC)' s tolerance capability which induces rat allogeneic orthotopic liver transplantation and prolong the survival time of allograft,this research co-modifying imDC with FasL and transforming growth factor-β1 ( TGF-[1 ) gene.Methods One hundred and forty cases of rat allogeneic orthotopic liver transplantation were accomplished in the strain combination of DA to Lewis rats,which were divided equally into 7 groups,including non- injection group ( control group),mDC group,imDC group,empty vector group,FasL group,TGF group and cotransfection group.Then 2 × 106 cells were respectively intraperitoneally injected to Lewis rats with distinct cells,such as mDC,imDC,empty vector-transfected imDC,hFasL-modified imDC,hTGF-β1-transformed imDC and comodified imDC with FasL and TGF-β1 gene.Four Lewis rats from each group were randomly sacrificed on days 3,7 and10 after transplantation.Hepatic function [ alanine aminotransferase (ALT) and total bilirubin (TBIL) ] and cytokines alteration were evaluated in peripheral blood.The pathological changes of the liver graft were observed.Survival analysis were performed for remnant rats,except rats dead for technical factor.Results At day 7,the difference was statistically significant ( P ＜ 0.01 ) between seven groups,and the AR in hepatic graft in cotransfection group were all lighter than those in TGF group and FasL group.At day 10,there was statistical significance between seven groups (P ＜0.01 ),the AR in hepatic graft in cotransfection group were all lighter than those in TGF group and FasL group,further more the AR in hepatic graft in TGF group was lighter than that in FasL group.At day 7,hepatic function of cotransfection group had reverted to normal and the hepatic function of FasL group took a turn for the better,but got worse at day 10;the concentration of ALT and TBIL of FasL group were obviously higher than those of cotransfection group at day 10 (P ＜0.01 and P ＜0.05).At day 10,Liver function of TGF group returned to normal,and ALT and TBIL concentrations of TGF group were significantly lower than those of FasL group ( P ＜ 0.01 and P ＜0.05 ).But compared to those of cotransfection group,the difference was not statistically significant ( P ＞0.05 ).The outcome of enzyme linked immunosorbent assay (ELISA) notified that at day 7,the serum levels of interleukin ( IL)-I,IL-10 and IL-12 in FasL group compared to those in cotransfection group and TGF group,the differences were all statistically significant ( P ＜ 0.05 ),on the contraty,the difference was not statistically significant ( P ＞ 0.05 ) between cotransfection group ane TGF group.The MST of rat recepients in FasL group was 20 days and compared to that in imDC group (23 d) with no statistically significance of difference (P ＞ 0.05),and the ability of TGF Group was limited in extending the rat receptors'survival time (48 d).There were rat recepients with long-term survival ( ＞ 90 d) only in cotransfection group,in which the survival of rat recepients was significantly prolonged than that in FasL group (P ＜0.01 ) and TGF group (P ＜0.01 ).Conclusion lmDC,hFasL or hTGF-β1 single gene-modified imDC all didn't induced long-term allograft survival of receptors in allogeneic rat liver transplantation.The ability of imDC with hFasL and hTGF-β1 co-transfection was significantly enhanced by hFasL and hTGF-β1 in inducing allogeneic rat liver transplantation immunology tolerance,which could significantly extended survival of recipients in allogeneic rat liver transplantation.
