抗可诱导共刺激分子抗体对大鼠慢性移植肾肾病的影响

Induction of immune tolerance by the blockade of ICOS/B7RP-1 costimulation pathway in chronic allograft nephropathy of rats

目的探讨应用抗可诱导共刺激分子（ICOS）抗体阻断ICOS-B7RP-1共刺激通路对大鼠慢性移植肾肾病（CAN）的影响及其可能机制。方法实验分为3组：（1）同系对照组，供、受者均为Wistar大鼠，进行肾移植；（2）同种对照组，以SD大鼠为供者，Wistar大鼠为受者进行肾移植，术后灌胃给予环孢素A（CsA），用10d；（3）实验组，在同种对照组的基础上，于使用CsA后经腹腔注射抗ICOS单克隆抗体，2mg／kg，2次／周，共用3个月。各组分别于术后4、8和12周，检测受者血清肌酐；观察移植肾的病理学变化；用免疫组织化学法检测移植肾和受者脾脏中ICOS的表达；检测移植肾组织中ICOS mRNA的表达。另取存活6个月的实验组和同系对照组大鼠，检测其外周血中CD4^＋CD25^＋T淋巴细胞的比例。结果同系对照组术后各个时间点的血清肌酐变化不大；同种对照组血清肌酐明显升高，12周时达（310±18）／μmol／L；实验组血清肌酐也呈上升趋势，但上升幅度明显低于同种对照组，至12周时为（122±13）μmol／L。术后12周时，同种对照组移植肾肾小球硬化率和Banff评分（总分）分别为（54．75±3．06）％和8．28±1．41，实验组为（15．05±1．64）％和6．13±1．00，两组比较，差异有统计学意义（P〈0．01）。同系对照组受者脾脏边缘区可见ICOS表达，白髓区少见ICOS表达，该组移植肾内未见ICOS明显表达；在同种对照组，移植后各时间点移植肾和受者脾脏中都可见ICOS明显表达；实验组移植肾和受者脾脏内ICOS的表达明显下降，且主要表达于脾脏的白髓区，边缘区很少表达。术后4、8和12周时，实验组各个时间点ICOSmRNA的表达明显低于同种对照组。实验组存活〉6个月者外周血中CD4^＋CD25^＋T淋巴细胞的比例平均为13．5％，同系对照组平均为9．1％。结论应用抗ICOS抗体阻断ICOS-BTRP-1共刺激通路可在一定程度上抑制CAN的进展，其机制可能与移植肾组织ICOS的表达受到抑制及ICOS在受者脾脏内的重新分布有关，其中也可能有CD4^＋CD25^＋调节性T淋巴细胞的参与。

Objective To study the efficacy and mechanisms of blockade of ICOS/B7RP-1 pathway to induce immune tolerance in treatment of rat chronic allograft nephropathy. Methods Orthotopic kidney transplantation was performed following the procedure of Kamada with our modification. All of experimental rats were divided into three groups： group A,isograft group; B, control group （CAN group）; C, anti-ICOS antibody group （intraperitoneal injection of anti-ICOS antibody,2 mg/kg twice weekly）. The pathological changes were observed at the 4th, 8th and 12th week post-transplantation by HE,PAS and Masson. When the transplanted rat survival time exceeded 6 months, CD25^ ＋/CD4^＋ T cells in peripheral blood were tested by using fluorescence activated cell sorter analysis （FACS）. The immunohistochemistry and real-time quantitative PCR were used to detect localization and expression of ICOS in transplant kidney and spleen of the recipients. Results The pathological changes of CAN in control group were significant, and the score according to Banff Standards in control group was higher than in other groups. Anti-ICOS treatment led to a significant decrease of chronic rejection lesions. The population of CD25^ ＋/CD4^＋ T cells in peripheral blood in group C （13. 5 %） was higher than that in group A （9. 1%） （P〈0. 05）. The expression of ICOS in transplant kidney and spleen of the recipients in group C was down-regulated as compared with group B （P 〈 0. 01 ）. Conclusion Blockade of ICOS-B7RP 1 with anti-ICOS-antibody to induce immune tolerance may attenuate remarkably the development of CAN, and the mechanisms may be the redistribution of ICOS in the spleen of recipients and a high population of CD25^＋/CD4^＋T cells in peripheral blood.