Study on inhibition of acute rejection in rat limb allografts by inducible costimulator pathway blockade / 中国地方病学杂志

ABSTRACT

Objective To observe the effect of inducible costimulator(ICOS) costimulation pathway blockade in rat limb allografts acute rejection by RNA interference. Methods Twenty-seven cases of modified hind llmb allotransplantation were performed from Wistar to SD rats. The rats were divided into 3 gronps(each n=9) the rejection group not given a special disposal; the control group, consisting of SD rats that received injection of pSilencer 4.1 and Sofast complex by vein post transplantation; and the interference group that received injection of pSilencer 4.1-ICOSshRNA and Sofast complex. On the eighth day posttransplantation, 3 rats were killed to study the pathological changes in each group. The expressions of ICOS gene in vivo were detected by flow cytometry and RT-PCR. The mixed lymphocyte reaction (MLR) was performed and eytokines in blood were measured by ELISA. The rest rats were used to record limb survival time. Results The mean survival time in rats of the rejection and the control groups were(11.34±1.21) and (11.14±1.32) days respectively. In the interference group, the mean survival time of limb allografts was (16.85±1.73) days(P<0.05). The rats in the rejection and the control groups experienced moderate to serious acute rejections with skin epidermal necrosis, a large quantity of lymphocyte infdtration, muscle cell necrosis and interstitial edema, while the pathological changes in rats of the interference group were mild. The splenocyte ICOS mRNA expression level in the interference group(18.75%) was significantly lower than that of the rejection group(100%) and the control group(98.51%). ICOS cell surface expression level as judged by the fluorescence intensity was 45.59±12.87 in the interference group, 103.72±21.76 in the rejection group, and 93.47±29.55 in the control group(F=6.89, P<0.05). In stimulation assays, a one-way mixed lymphocyte reaction stimulation index(SI), with spleen cells from Wistar and Lewis rats, respectively, the rejection group (5.26±0.42,5.18±0.29) and the control group (5.37±0.27,4.93±0.44) had significantly greater reactions than the interference group(2.37±0.35, 4.87±0.36), respectivily(F=7.29, P<0.05; F=6.19, P0.05). In the IFN-γ and IL-4 expression assays, reactions of the interference group (230.17±38.47,160.32±59.13) were lower than those of the rejection group(490.73±51.48,230.67±45.21) and the control group(480.15±43.96, 240.53± 47.36), (F=7.23,6.75, all P<0.01). Conclusions In vivo transfection of pSilencer 4.1-ICOS shRNA interference plasmid can effectively block T-cell co-stimulation pathway, suppress acute rejection, and prolong limb allografts survival.