糖皮质激素对角膜移植术后大鼠角膜TLR2表达的影响

The effects of steriod on expression of TLR2 in rat corneas after penetrating keratoplasty

目的检测大鼠角膜组织TLR2mRNA的表达,研究糖皮质激素对穿透角膜移植术后角膜TLR2表达的影响及对术后排斥反应的作用。方法实验动物分正常对照组(N)、同种同体角膜移植组(A)、同种异体角膜移植组(B)和同种异体角膜移植激素(典必殊滴眼液点术眼,每日2次)抗排斥组(C),A、B、C3组大鼠给予穿透角膜移植术,术后裂隙灯下观察角膜透明度、新生血管并用排斥反应指数评价;分别于术后第5、7、9天取材,行组织病理学检查和实时荧光定量PCR检测,动态观察角膜组织中TLR2mRNA的表达。结果术后随时间变化角膜植片均出现不同程度的水肿、混浊、新生血管生长。B组大鼠平均在穿透角膜移植术后7d发生排斥反应并获病理学支持;A组和C组大鼠术后排斥反应指数计分未达到诊断标准。术后3组手术干预组角膜TLR2mRNA的表达均增加,同种异体角膜移植组角膜TLR2mRNA的表达较同种同体角膜移植组显著增加,激素抗排斥组角膜TLR2mRNA表达明显降低(P<0.05)。结论糖皮质激素可能通过抑制角膜TLR2的表达及其介导的信号转导,抑制排斥反应,对移植物起保护作用。

Objective Our previous study demonstrated that toll-like receptor 2（TLR2） can distinguish extraneous antigen and prevent the immunological response. This study was designed to detect the expression of toll-like receptor 2 （TLR2） mRNA in cornea and investigate the effect of steroid on TLR2 expression in rats cornea following allograft penetrating keratoplasty. Methods The penetrating keratoplasty models were established in SPF rats with the 108 SD rats as receiptors and 36 SPF Wistar rats as donors,and other 6 SPF SD rats worked as normal controls. The receiptor rats were divided randomly into autografl group, allograft group and steriod group. The clarity and neovascularization of corneas of experiment rats were examined under the slit- lamp microscope and the rejection index was calculated based on Holland criteria. Corneal histopathological examination was carried out by hemotoxylin and eosin staining under the light microscope, and real time-PCR was employed for the detect of TLR2 mRNA in the corneas at the fifth,seventh and ninth day after operation. The experimental animals were obtained from the Animal Experimental Center of Southern Medical University and the procedure followed the Statement of Association for Research in Vision and Ophthalmology. Results The rejection occurred in 7 days after operation in allografl group, and only mild edema,opacity and neovascularization of corneas were found at different degrees in 9 days after operation in autograft group and steriod group. Severe corneal edema, a lots of inflammatory cells infiltration and new vessels in stroma were seen in allograft group,and mild inflammatory response was found in autografl group and steriod group. Normal corneal structure was exhibited in normal control group under the light microscope. The fold differences of TLR2 mRNA expression in cornea after amplification was significantly different among three groups and different time points （ Fsro.p = 39.46, P = 0. 00 ; Ftime = 35.38, P = 0. 00 ; Fi ion = 45.66 ,P = 0.00 ） ,and the evident enhance of TLR2 mRNA expression was revealed in allograft group compared with autograft group（P 〈 0. 05 ） and declined in steriod group （ P 〈 0.05 ）. Conclusion Steriod may restrain the acute allograft rejection by down-regulating the expression of TLR2 in corneas and its signals transaction. This result suggests that steriod offer a protection from rejection of cornea after penetrating keratoplasty.