Investigation Of Epithelial-mesenchymal Transition In The Relationship Between Lens Epithelial Cells And Cataracts

Objective:Epithelial-mesenchymal transition (EMT) is a dynamic process during which the epithelial cells acquire mesenchymal, fibroblasts-like properties, show reduced intercellular adhesion and increased motility. Recent studies indicate that EMT of the lens epithelial cells plays an important role in the pathogenesis of anterior subcapsular cataract (ASC) and posterior capsular opacification (PCO). Epidemiology reveals the onset of cataract among diabetes sufferers is 9.6 years earlier compared with normal people. Up to now, phacoemusification with intraocular lens implantation is the best way to treat cataract. However, PCO which is a fairly common post-operative complication of cataract often leads to a decline of the patients' vision again. Some recent studies indicate that members of the TGF-β superfamily play an important role in lens fiber trans-differentiation, and inappropriate TGF- β superfamily of the lens epithelial cells may result in an EMT which bears morphological and molecular characters similar to the process of cataract formation.Methods:1. Rabbit lens epithelial cells (RELCs) that were cultured with normal culture medium (4.5g/L glucose) and with HG(9, 18, 36g/L glucose) were maintained in a humidified atmosphere containing 5% CO2. At the end of this treatment, TGF-β2, α-SMA and PCNA were examined by immunochemistry and absorbency of RLECs were tested by MTT assay.2. Preparing recombined pRNAiH1.TGF-β2^shRNA and cells model and rabbit model for the next transfecting experiments.3. pRNAiH1.TGF- β 2^shRNA transfected RELCs that were treated with 18g/L glucose, the expressions of TGF- β 2 and a-SMA were tested by Real-time PCR. pRNAiHl.TGF-β 2^shRNA being injected into rabbit anterior chamber, the expression of TGF- β 2 of RLECs was tested by Real-time PCR.Results:1. RLECs nucleus were oval or round and borderlines of cells were clear. After the treatment of HG, RLECs took on a elongated aspect like fibroblast cells and formed reticulation or vesicle in some cells.2. TGF-P2 positive strain expression in the 4.5 and 9g/L glucose was less than that in the 18 and 36g/L glucose and this difference was statistical significant(p<0.05). a-SMA positive strain expression was enhanced with the glucose in the culture medium substrate increasing. The expression in the 4.5 and 9g/L glucose was less than that in the 18 and 36g/L glucose and this difference was statistical significant(p<0.05).3. RLECs transfected by pRNAiHl.TGF-3 2shRNA appeared that the TGF-p2 and a-SMA positive strain expression declined(p<0.05) and the result in vivo was also that pRNAiHl.TGF- P 2shRNA could reduce the expression of TGF- P 2.Conclusions :1. HG can make RLECs appear EMT in vitro and increase the expression of TGF- P 2. So we conclude that the reason of earlier arising of diabetes patients' cataracts is possible that higher glucose in the aqueous humor in diabetes patients causes lens epithelial cells to appear EMT.2. TGF-P is the most important cell factor to regulate EMT that is a dynamic process in which cells change from the epithelial state differentiating into a mesenchymal phenotype. In this study, we prepared pRNAiHl.TGF-P 2shRNA, restrained TGF-P2 expression of RLECs and consequently made a more research in the process of EMT in the formation of cataracts (ASC, PCO and so on).3. Our results testifies that restraining TGF-02 expression can decrease the a-SMAexpression and regulate the process of EMT in lens epithelial cells, so we suppose that interfering targeted gene TGF-02 could be a feasible therapy to cataracts related to EMT.