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Abstract #45945 Published in IGR 13-2
Inhibition effect of P27 tumor suppressor gene transfection on human Tenon's fibroblast cells
Sun L; Luo W; Li J-MChinese Ophthalmic Research 2010; 28: 1119-1123
Background: The failure rate of glaucoma filtrating surgery is 15%-30% during 2-year duration and the main reason is proliferation of fibroblast and scarring of filtering tunnel. Clinical researches found some adverse effects after administration of anti-scarring drugs. Objective: Present study was to discuss the cationic liposome-mediated P27 tumor suppressor gene in cultured human Tenon's fibroblast inhibition in vitro. Methods: Human Tenon's capsule tissue was collected during the strabismus correct surgery, and adherent expiants culture method was used to cultivate the fibroblast in DMEM + 10% FCS. With liposome-mediated method, the constructed pEGFP-N1 P27 eukaryotic expression plasmid was transfected in vitro into cultured human Tenon's fibroblasts, and recombinant plasmid pEGFP-N1 was transfected in the same way as the empty vector transfection group. Only cultured fibroblasts were used as non-transfection group. Expression of P27 in human Tenon's fibroblasts was detected by Western blot. The cell cycle after transfection was observed by flow cytometry. The proliferation activity of the fibroblasts following the pEGFP-N1 P27 transfection was assessed by methylthiazlyl tetrazolium (MTT) assay. Results: The transfected fibroblasts cells showed the green fluorescence under the inverted fluorescence microscope. The fibroblasts transfected by pEGFP-N1 P27 gene presented with the green fluorescence. Western blot assay showed that P27 gene was expressed in the fibroblasts at the protein level. The percentage of the fibroblasts in G(0)-G(1) phase and S phase was 73.16% and 26.84% respectively in pECFP-N1 P27 transfection group, 63.29% and 58.16% in pEGFP-N1 transfection group, 36.71% and 41.84% in non-transfection group. After transfection of pEGFP-N1 P27, the A(490) value of the fibroblasts was significantly declined in comparison with pEGFP-N1 group (0.079(plus or minus)0.054 vs 0.127(plus or minus)0.106) and non-transfection group (0.079(plus or minus)0.054 vs 0.180(plus or minus)0.007) (P = 0.000;P =0.011). Conclusion: The P27 gene transfection may inhibit the proliferation of human Tenon's fibroblasts in vitro.
L. Sun. Department of Ophthalmology, Dalian Medical University, Dalian 116027, China.
Classification:
12.8.10 Woundhealing antifibrosis (Part of: 12 Surgical treatment > 12.8 Filtering surgery)
11.14 Investigational drugs; pharmacological experiments (Part of: 11 Medical treatment)