Human amniotic membrane isolated from the placenta contained basement membrane components such as type IV collagen, laminin, and 6 and 4 integrins, all of which remained detectable while preserved in glycerin for one week. One month after the n-heptanol removal of the total corneal epithelium and the limbal lamellar keratectomy, all rabbit eyes carried features of limbal deficiency, including conjunctival epithelial ingrowth, vascularization and chronic inflammation. Ten control eyes then received a total keratectomy, and 13 experimental eyes received an additional amniotic membrane transplantation. Three-month follow-ups revealed that all control corneas were revascularized to the center with granuloma and retained a conjunctival phenotype. In contrast, in the experimental groups, 5 corneas became clear with either minimal or no vascularization; the rest had either mild peripheral (5) or total (3) vascularization and more cloudy stroma. Using monoclonal antibodies for epithelial markers and matrix components, we concluded that the success correlated with the return of a cornea-like epithelial phenotype and the preservation of the amniotic membrane, whereas the failure maintained a conjunctival epithelial phenotype and the amniotic membrane was either partially degraded or covered by host fibrovascular stroma. Measures taken to facilitate the former might prove this procedure clinically useful for ocular surface reconstruction.
Amnion/chemistry/*transplantation ; Animals ; Antibodies, Monoclonal ; Basement Membrane/chemistry/pathology ; Cornea/pathology/*surgery ; Corneal Neovascularization/pathology/*prevention & control/surgery ; Epithelium/pathology/surgery ; Extracellular Matrix Proteins/analysis ; Female ; Fluorescent Antibody Technique ; Humans ; Immunophenotyping ; Male ; Rabbits

OBJECTIVETo evaluate the efficacy of epigallocatechin gallate (EGCG) in treatment of corneal alkali burn injury in mice.

METHODSCorneal alkali burn injury was induced by sodium hydroxide method in C57BL/6J mice. The mice with cornea burns were treated intraperitoneally with EGCG solution or phosphate buffer solution (PBS) respectively. The healing of corneal epithelium, the formation of corneal neovascularization (CNV) and the inflammation reaction were assessed by slit -lamp microscopy and histological examination. Expression of vascular endothelial growth factor (VEGF) mRNA and protein in cornea was evaluated by real -time reverse transcription polymerase chain reaction (RT-PCR) and immunohistochemistry, respectively. Myeloperoxidase (MPO) assay was used to quantitatively evaluate the polymorphonuclear neutrophils (PMNs) infiltration in the corneas.

RESULTSThe healing rate of corneal epithelium in EGCG group was significantly higher than that of PBS group at d1, d3 and d7 after treatment (d1: 41.0%±13.0% vs 23.8%±7.6%; d3: 76.6%±7.5% vs 61.2%±6.8%; d7: 87.8%±8.5% vs 74.0%±9.1%; all P <0.05). The CNV scores and the number of CNV in the corneal sections of EGCG group were significantly lower than those of PBS group at d3, d7 and d14 after treatment (CNV score: d3: 1.1±0.5 vs 6.6±1.0; d7: 1.3±0. 3 vs 8.1±1.0; d14: 0.9±0.2 vs 9.2±1.1; CNV number: d3: 1.68±0.61 vs 2.92±0.95; d7: 4.80±1.36 vs 7.92±1.28; d14: 3.64±0.71 vs 5.88±0.76; all P<0.05) . The expression of VEGF protein at d3 (0.19±0.05 vs 0.45±0.08) and d7 (0.42±0.07 vs 0.84±0.09), the expression of VEGF mRNA at d1, d3 and d7 in EGCG group were significantly lower than those in PBS group (all P <0.05). Compared to PBS group, the inflammatory index at d3 (3.2±0.4 vs 3.7±0.5) and d7 (2.3±0.5 vs 4.0±0.0), the number of PMNs in the corneal sections and the MPO values at d3, d7 and d14 in EGCG group were significantly decreased (PMNs: d3: 34.5±15.7 vs 90.0±28.8; d7: 17.1±11.4 vs 54.9±25.9; d14: 12. 8±4.6 vs 39.0±17.9; all P <0.05).

CONCLUSIONIn the murine corneal alkali burn model, intraperitoneal injection of EGCG solution can promote the healing of corneal epithelium, inhibit the formation of CNV and reduce the inflammatory cell infiltration in the corneas.

Alkalies ; Animals ; Burns, Chemical ; drug therapy ; Catechin ; analogs & derivatives ; therapeutic use ; Cornea ; drug effects ; pathology ; Corneal Neovascularization ; prevention & control ; Disease Models, Animal ; Eye Burns ; drug therapy ; Inflammation ; drug therapy ; immunology ; Mice ; Mice, Inbred C57BL ; Neutrophils ; cytology ; RNA, Messenger ; Vascular Endothelial Growth Factor A ; metabolism