To investigate the expression of vascular endothelial growth factor (VEGF) on human pterygium fibroblast angiogenesis induced by vasculogenic mimicry (VM) effect.

6 cases of primary pterygium were collected from Zengcheng District People′s Hospital of Guangzhou province (aged from 41 to 58). Cut fresh tissue of human pterygium, and in vitro adherent cell culture. The human pterygium fibroblasts were identified by immunocytochemical staining of Vimentin and cytokeratin. Groups were divided into control group and different concentrations of VEGF (1 ng/ml, 10 ng/ml and 100 ng/ml) group, take good growth of the 3 to 5 generations of pterygium fibroblasts were cultured. Cell counting kit-8 method was used to detect the proliferation ability of each component of the fibroblast cells, and the cell culture and PAS staining were used to observe the VM of each group, Western blotting was used to detect the expression of matrix metalloproteinase (MMP)-2 and hypoxia inducible factor -1α(HIF-1α) protein in the cells of each group of rats.Each absorbance(OD value), MMP-2, and HIF-1 alpha described by the mean and standard deviation (±s). Multiple groups were compared using one-way analysis of variance, when the difference was statistically significant, two further comparisons between the two. Bivariate correlation analysis using Spearman′s rank correlation analysis.

Human pterygium fibroblasts identification results showed that the cell smear after hematoxylin eosin staining under the microscope in the display cell fusiform, triangle, fan-shaped or irregular shape. The results of immunocytochemical staining showed that the cytoplasm of the cells was positive for Vimentin, and the expression of CK was negative. The results of cell counting kit was used to detect cell proliferation showed that the OD value of control group and 1 ng/ml group, 10 ng/ml group and 100 ng/ml group cells were 0.57±0.023, 0.59±0.012, 0.77±0.024 and 1.014±0.035, differences in control group and 1 ng/ml group was not statistically significant (t=1.21, P>0.05). the OD value of 10 ng/ml group and 100 ng/ml group were higher than that of the control group and 1 ng/ml group (t=14.78, 12.34; P<0.05); the OD value of 100 ng/ml group is the highest, control group, 1 ng/ml and 10 ng/ml groups were significantly different (t=20.12, 19.45, 13.41; P<0.05). Three dimensional cell culture and PAS staining showed that 10 ng/ml group and the 100 ng/ml group had VM structure in the control group and the 1 ng/ml group, no VM appeared in the control group and group. 10 ng/ml group and the density of the 100 ng/ml were respectively 4.40±1.14 and 12.40±2.07, the difference was statistically significant (t=-7.899, P<0.05). The 1 ng/ml group, 10 ng/ml group and 100 ng/ml group cell MMP-2 values were 0.26±0.038, 0.29±0.013, 0.39±0.013 and 0.73±0.014 in the control group; and 1 ng/ml group, 10 ng/ml group and 100 ng/ml group cell HIF-1 values were 0.087±0.0086 and 0.087±0.0031, 0.15±0.0016 and 0.37±0.026 in control. The difference between MMP-2 and HIF-1 alpha gray value of 1 ng/ml group and control group had no statistical significance (t=1.21, 0.01; P>0.05); MMP-2 and HIF-1 alpha gray values were significantly higher than those in control group and 1 ng/ml group (t=7.92, 6.85, 17.64, 17.82; P<0.05); 100 ng/ml group of MMP-2 HIF-1 alpha and gray value were significantly higher than the other 3 groups (t=28.48, 26.45, 18.72, 25.31, 24.78, 15.45; P<0.05). Pterygium fibroblast cells in vasculogenic mimicry density and MMP-2 and HIF-1 expression was positively correlated, and closely correlated ( r=0.509, 0.503; P<0.05).

VEGF can promote cells proliferation and induce VM in human pterygium fibroblasts cells, also enhance the protein expression of MMP-2 and HIF-1α. VEGF may be the molecular mechanism of angiogenesis mimicry and synergistic enhancement MMP-2 and HIF-1α protein expression, thus influence the progress of pterygium.