关键词: 超微机制, 压力, 仿生培养, 角膜内皮细胞

Abstract:

 Objective To investigate the micro-mechanism of high pressure damage on corneal endothelial cells. Design Experimental study. Participants Rabbit corneal endothelial cells. Methods The first generation of rabbit corneal endothelial cells in vitro culture (RCEC) were divided into five groups: group A: 30 mmHg pressure,  group B: the fluctuated pressure, pressure would be set to 15 mmHg-25 mmHg-20 mmHg-10 mmHg, each pressure level for six hours, group C: 50 mmHg pressure , group D: the normal pressure group, group E: without pressure group. The original generation cells were identified by immunohistochemical; cell activity was detected by Flow cytometry. The protein of BcL-2 and P53 of cell was detected with Western blotting test. The expression of Fas/FasL was detected with RT-PCR. The expression in cytoplasm Cytc was detected by Immuno-fluorescence. Main outcome Measures The expression about BcL-2 and P53 and Fas/FasL and Cytc. Results NSE antibody of the primary corneal endothelial cells was positive, and no corneal epithelial cells and corneal stromal cells were found. Corneal endothelial cell activity was lower in high pressures groups by Flow cytometry and most of damaged cells were apoptosis. In the group of 30 mmHg, 50 mmHg and the fluctuated pressure groups, the protein P53 relative expression (OPTDIP53/OPTDIactin) were 0.253±0.014, 0.670±0.019, 0.474±0.016, respectively (F=1210, P=0.000). The expression of Fas/FasL of each group was negative by RT-PCR analysis in each time period. The expression of Cytc in cytoplasm was positive in the high pressure groups with immunofluorescence. Conclusion The damage of the corneal endothelial cell under high pressure environments was caused by apoptosis, while the start of apoptosis pathway was the Cytc being released from mitochondria into the cell cytoplasm, which was endogenous way of apoptosis. (Ophthalmology, 2018, 27: 36-41)

Key words: micro-mechanism, pressure, bionic culture, corneal endothelial cells