大鼠角膜碱、酸、热烧伤后ICAM-1、VCAM-1、CD44及E-selectin mRNA的表达

Expression of ICAM-1,VCAM-1,CD44 and E-selectin mRNA after corneal alkali burn,acid burn and thermal burn in rat corneas

目的利用RT-PCR法检测大鼠角膜烧伤后角膜局部ICAM-1、VCAM-1、CD44及E-selectin的mRNA表达情况,探讨黏附分子在角膜烧伤中的变化,以进一步完善角膜烧伤的免疫机制。方法制备Wistar大鼠角膜碱、酸、热烧伤模型。提取大鼠烧伤后3d和2周角膜RNA,反转录扩增ICAM-1、VCAM-1、CD44及E-selectin,根据DNA条带的光密度值确定DNA含量。观察烧伤后3d和2周以上各指标的变化。结果角膜烧伤后3d,与正常对照组(1.658±0.722)相比,3个烧伤组ICAM-1mR-NA表达量增加(5.699±0.934、3.182±0.830、3.403±0.916),差异均有显著统计学意义(均为P<0.01);2周时,3个烧伤组ICAM-1mRNA较3d时明显减少,其中碱烧伤组伤后3d和2周ICAM-1mRNA的表达量均较酸和热烧伤组明显增多,差异均有显著统计学意义(均为P<0.01)。同正常对照组(2.172±0.368)相比,烧伤后2周时,碱烧伤组VCAM-1mRNA表达量(8.591±1.021)增多,差异有极显著统计学意义(P<0.001),而酸和热烧伤组差异均无显著统计学意义。同正常对照组(1.402±0.439)相比,伤后3d时3个烧伤组CD44mRNA表达量无显著变化,2周时碱烧伤组CD44mRNA表达量(5.587±0.753)增加,与对照组比较,差异有显著统计学意义(P<0.01),酸和热烧伤组CD44mRNA表达量增加,但差异无统计学意义。3d时角膜各烧伤组E-selectin的表达量低于正常对照组,差异均有显著统计学意义(均为P<0.01);2周时各烧伤组E-selectinmRNA的表达量明显增加,高于正常对照组,差异均有显著统计学意义(均为P<0.01),3个烧伤组间差异无显著统计学意义。结论在角膜烧伤的急性期,ICAM-1介导中性粒细胞与血管内皮细胞的黏附,表达增加。CD44及其配体在角膜烧伤特别是角膜碱烧伤的愈合过程中发挥重要作用。作为介导白细胞与血管内皮细胞最初黏附过程的黏附分子,E-selectin在角膜烧伤的病理发生中的作用机制有待于进一步研究。

Objective To investigate the changes of adhesion molecules in corneal burn to complete the immunologic mechanisms of corneal burn by detecting the expression of ICAM-1,VCAM-1,CD44 and E-selectin mRNA after alkali burn,acid burn and thermal burn in the rat corneas by RT-PCR.Methods Wistar rat models of alkali burn,acid burn and thermal burn were made.Corneal RNA was extracted at 3 days and 2 weeks after corneal burn.ICAM-1,VCAM-1,CD44 and E-selectin were amplified with reverse transcription.DNA content was determined by optical density of DNA strap.Changes of each index were observed at 3 days and 2 weeks after corneal burn.Results The expression of ICAM-1 mRNA were 5.699±0.934,3.182±0.830 and 3.403±0.916 at 3 days after corneal alkali burn,acid burn and thermal burn higher than 1.658±0.722 in normal control group,there were significant differences（all P0.01）.The expression of ICAM-1 mRNA at 2 weeks after corneal burn was fewer than those at 3 days after corneal burn（P0.01）.At 3 days and 2 weeks after corneal burn,expression of ICAM-1 mRNA in alkali burn group was obviously higher than those in acid burn group and thermal burn group after corneal burn,there were significant differences（all P0.01）.At 2 weeks after corneal burn,the expression of VCAM-1 mRNA was 8.591±1.021 in alkali burn group higher than 2.172±0.368 in normal control group,and there was significant difference（P0.001）.While no obvious difference was found in expression of VCAM-1 mRNA for acid burn group and thermal group compared with normal control group.At 3 days after corneal burn,there was no obvious difference in expression of CD44 mRNA of 3 corneal burn groups compared with 1.402±0.439 in normal control group.At 2 weeks after corneal burn,expression of CD44 mRNA was 5.587±0.753 in alkali burn group higher than that in normal control group,there was significant difference（P0.01）;Expression of CD44 mRNA increased at 2 weeks after corneal burn in acid burn group and thermal burn group,but no difference was found.At 3 days after corneal burn,expression of E-selectin mRNA groups in 3 corneal burn groups were fewer than that in control group,there were significant differences（all P0.01）,At 2 weeks after corneal burn,expression of E-selectin mRNA increased in 3 corneal burn groups,and were higher than that in normal control group,there were significant differences（all P0.01）,while no difference was found among 3 corneal burn groups.Conclusions At acute stage after corneal burn,ICAM-1 can mediate adherence of neutrophilic granulocytes and vascular endothelial cells,and the expression increases.CD44 and its ligands play important roles in healing of corneal burn,especially for corneal alkali burn.As adhesion molecule to mediate initiatory adherence of leucocytes and vascular endothelial cells,E-selectin takes part in pathology of corneal burn,and more studies of its mechanisms are needed.