ABSTRACT
Background Corneal neovascularization (CNV) is one of the causes of corneal blindness.Studies showed that zonula occludens-1 (ZO-1) can inhibit pathological angiogenesis through physical barrier formed by tight junction structure.However,whether ZO-1 plays a role in CNV is unclear.Objective The aim of this study was to explore the effect of ZO-1,a tight junction protein on experimental CNV.Methods The CNV models were established in the left eyes of 24 clear male BALB/c mice aged 7-8 weeks by putting NaOH filter paper in the center of corneas for 15 seconds (15 s group) or 40 seconds (40 s group).CNV was examined and evaluated under the slit lamp microscope,and the expression of ZO-1 mRNA in the corneas were detected and compared by reverse transcription PCR (RT-PCR) between the two groups 2 weeks after modeling.In addition,54 models created by the same method were assigned to 3 groups according to randomized number table,0.2％ hyaluronic acid (HA),antiZO-1 neutralizing antibody (10 mg/L) +0.2％ HA and mouse hypoxia inducible factor-1α (HIF-1α) recombinant protein (5 mg/L)+0.2％ HA were topically administrated in the mice three times a day for 1 week after modeling respectively.The corneas were extracted 2 weeks after application of the drugs.Expression of CD31 in the CNV was assayed to calculate the number and the area of CNV by immunohistochemistry.The expression of VEGF mRNA in the corneas was detected by RT-PCR.The percentages of macrophage-specific F4/80 positive cells and neutrophilsspecific Ly-6G positive cells were calculated to evaluate the infiltrations of inflammatory cells in the corneas by flow cytometry.Results In 2 weeks after alkali burn of corneas,the number of severe CNV was more in the 40 s group than that in the 15 s group (x2 =6.032,P=0.049),and the expression level of ZO-1 mRNA was lower in the 40 s group than that in the 15 s group (1.15±0.08 versus 1.53±0.04) (t=4.157,P=0.014).CD31 positive cell number was more and the staining area was larger in the ZO-1 antibody group and HIF-1α positive control group than those in the 0.2％ HA group (cells:t=-129.590,-226.820,both at P=0.000;area:t =-5.310,-8.840,both at P=0.000).The relative expressions level of vascular endothelial growth factor (VEGF) mRNA was 1.33±0.10 and 1.46±0.11 in the ZO-1 antibody group and HIF-1 α positive control group respectively,which were significantly higher than 0.93±0.06 of the 0.2％ HA group (t =-5.820,-7.284,both at P =0.000).The percentages of positive cells in the ZO-1 antibody group and HIF-1α positive control group were significantly increased in comparison with the 0.2％ HA group for F4/80 (t =-16.750,-17.480,both at P =0.000) and for Ly-6G (t =-21.450,-27.680,both at P=0.000).Conclusions Alkali burn induced CNV downregulates the expression of ZO-1 mRNA.Administration of ZO-1 antibody causes the rise of VEGF mRNA in CNV and the infiltration inflammation cells,which suggests that the influence of ZO-1 on CNV is associated with the expression of VEGF.
ABSTRACT
This study was performed to evaluate the effects of conditioned media (CM) from human amniotic epithelial cells (HAECs) on the corneal wound healing process. Eighteen rabbits (36 eyes) were used and randomly assigned to three groups according treatment: CM from HAECs (group 1), vehicle alone (group 2), and saline (group 3). Corneal alkali injuries were induced with 1 N sodium hydroxide. Each reagent used for treatment evaluation was injected into the dorsal bulbar subconjunctiva and the area of the corneal epithelial defect was measured every other day. Two animals from each group were euthanized at a time on days 3, 7, and 15, and the cornea was removed for histological examination. The sum of the epithelial defect areas measured on day 0 to day 6 as well as day 0 to day 14 in group 1 was significantly smaller than those of other groups. Histological examination revealed that the group 1 corneas had less inflammatory cell infiltration and showed more intact epithelial features compared to the other groups. These results suggest that CM from HAECs promote corneal wound healing in rabbits.