Effects Of Mechanical Stimulus On Proliferation And The Bfgf Expression Of Keratocytes

Laser in situ keratomileusis (LASIK)is now a widely performed refractive procedure with an estimated over one million annual procedures performed worldwide. The main mechanism of LASIK is to change the central optical curvature of cornea to correct the refractive errors. It involves the use of a microkeratome to create a thin corneal flap followed by excimer laser ablation of the corneal stroma and repositioning of the flap, which makes epithelial cell and Broman’s membrane integrated; LASIK created less corneal hazeCorneal tissue repair after LASIK is a complicated process which involves many factors. Alteration of the mechanical environment is an important one. The corneal tissue will be subjected to greater stress under the same intraocular pressures(IOP) when it becomes thinner after LASIK.The mechanical environment of corneal fibroblasts will be also change,which can influence many aspects of cellular behaviors such as cell growth,proliferation and biosynthesis. Previous studies showed that the basic fibroblast growth factor(bFGF) played an important role in wound healing and tissue repair.Our aim is to investagte the expression of bFGF in corneal tissue in vivo and in vitro and the effects of mechanical stimulation on proliferation of keratocytes after LASIK. It will provide a valuable evidence and gudiance for clinical therapy.The main research contents and conclusions are as follows:1. Rabbit corneas in different mechanics environment were established by LASIK with different residual stroma bed. The expression of bFGF in epithelial layer and stromal layer was observed by immunofluorescence staining, changes on proliferation of keratocytes are observed by HE staining.Our results showed that:bFGF was found in both epithelial layer and stromal layer,the fluorescence intensity of stromal layer was higher than epithelial layer; At one week after LASIK, expression of bFGF in stromal layer increased significantly in the group with residual stroma bed30%of the whole cornea compared to epithelial layer (P<0.05);At one week after LASIK,compared with the normal group, the number of cornea cells decreased at first,then increased at one month after LASIK. Analysis for the same surgery fashion at one week and one month after LASIK showed that the number of cells at one month were obviously more than at one week(P<0.05).2. Experimental animals were killed at one week or one month after LASIK and corneas were harvested.The concentration of bFGF was detected by ELISA. Our results showed that at one week after LASIK, the expression of bFGF increased significantly in the group with residual stroma bed30%of the whole cornea compared to the control group (P<0.05);Then it decreased to the normal level in all groups at one month after surgery.There were no significant differences at different surgery fashion;Analysis for the same surgery fashion at different time point showed that there was significant difference between one week and one month only in30%group (P<0.05).3. Corneal fibroblasts isolated from normal rabbits were subjected to cyclic stretch regimen of5%、10%and15%elongation at0.1Hz respectively.The concentration of bFGF was assayed on aliquots of culture medium by ELISA method after stretch for6h and24h respectively. Our results showed that, when the corneal fibroblasts were subjected to15%elongation for6h, the concentration of bFGF in the culture medium supernatant was significantly elevated compared to the control (P<0.05), then decreased after stretch for24h (P<0.05).Our results indicated that the proliferation of corneal stromal cell was changed after LASIK, the expression of bFGF was mainly concentrate on the stromal layer. The expression of bFGF was up-regulated in eary stage after LASIK in vivo and mechanical stretch expriments in vitro. It will be beneficial to cornea repair after LASIK.