羊膜移植及丝裂霉素对碱烧伤大鼠角膜缘基质微环境的影响

Influence of alkaline burned rat corneal limbus microenvironment by transplantation of amniotic membrane and mitomycin C

目的观察羊膜移植及丝裂霉素C(MMC)对碱烧伤大鼠角膜缘基质微环境的影响。方法采用SD大鼠角膜缘碱烧伤模型。将大鼠随机分为5个组:A组为正常组,B组为羊膜移植组,C组为羊膜移植联合MMC组,D组、E组为未干预组(D组为碱烧伤1周组,E组为碱烧伤2周组)。通过苏木精-伊红染色观察病理学改变,免疫组织化学技术观察髓过氧化物酶(MPO)、肝细胞生长因子(HGF)免疫染色阳性细胞数的变化。结果干预组(B组、C组)较未干预组(D组、E组)角膜上皮完整、基质细胞浸润减轻、新生血管减少,组织病理学染色证实角膜缘及其周边上皮细胞为多层结构,角膜新生血管减少及基质中炎性细胞浸润减轻。免疫组织化学结果显示MPO及HGF含量明显减少,B组、C组之间MPO含量差别不明显,C组中的HGF含量明显减少,上皮细胞层数轻微减少。结论羊膜移植和羊膜移植联合MMC可以改善角膜缘碱烧伤大鼠模型中的角膜缘基质微环境,MMC可能通过抑制成纤维细胞分泌HGF而影响上皮细胞的增生和移行。

Objective Destruction of corneal limbus microenvironment may induce limbal stem cells deficiency. It is important to recovery limbus microenvironment when reestablish ocular surface. Present study was to examine the influence of transplantation of amniotic membranes and mitomycin C on alkaline burned rat corneal limbus microenvironment. Methods The model of alkaline cauterization of corneal limbns was established in 16 Sprague-Dawleg（SD） rats by covering the cornea limbus with filter soaking 1 mol/L NaOH for 15 seconds. The rats were divided into 5 groups randomly. Group A was normal rats（4 rats） ,and transplantation of amniotic membrane was performed after alkaline cauterized in group B, and combination of transplantation of amniotic membrane with mitomycin C was used in group C. Group D and E was alkaline asteriated for 1 week and 2 weeks respectively. The change of pathology was observed by HE dyeing under the light microscope. The myeloperoxidase （MPO） and hepatocyte growth factor （HGF） positive cell numbers were evaluated by immunohistochemistry. Results Integrated corneal epithelium,less vascularity and stromal infiltration in group B and C compared with group D and E. Pathological examination verified that the limbal and peripheral corneal epithelium was composed of muhilayer cells, and the neovascularization and stromal inflammatory cells were decreased in group B and C. The immunohistochemistry showed that the positive cell numbers of MPO and HGF were decreased obviously after transplantation of amniotic membrane and usage of mitomycin C in comparison to the untreated group D and E. However, there was no remarkable difference in MPO positive cell numbers between group B and C（P 〉 0. 05）. Less expression of HGF and layers of epithelial cells were found in group C than B. Conclusion Amniotic membrane and amniotic membrane combined with mitomycin C can inhibit the inflammatory reaction in alkaline burned corneal limbus. Mitomycin C inhibit desmocyte to excrete HGF and therefore influent the proliferation and migration of epithelial cells.