| ObjectiveWith the development of cataract surgery, cataract surgery complications is becoming less, such as, vitreous into the anterior chamber, pseudophakic bullous keratopathy, glaucoma, iris damage, hyphema and lens decentration. But Posterier Capsuler Opacification has became more impormentoOver this years, The studies of preventing PCO now concentrated in inhibiting the proliferation and migration of the remained lens epithelial cells (LECs). Currently a hot research focused on drugs that suppress the proliferation aspects of lens epithelial cells. The outcome of the former method is unsatisfactory. The key point is the inhibition of the LECs proliferation by pharmaceutical ways at present. Decades, we have studied some drugs, such as, anti-metabolism drugs, natural herbs, tissue plasminogen activator, anti-inflammatory drugs, cytotoxic druys and immunotoxin. this druys have not be used in clinical becaused of their strongly side effects. In recent years, the study focuses on inhibition of lens epithelial cell proliferation and migration by impacting proliferation of bFGF in mang ways.LY294002 is phosphatidylinositol 3-kinase (PI3K) channel blocker, PI3K is a key signaling molecule in many life activities and is involved in regulating cell division, differentiation, apoptosis and other activities. Studies have shown that it is involved in proliferation of LEC. serine/theonine kinase is PI3K's direct downstream traget proteins, AKT is a multifunctional regulator and is involved in the regulation of cell survival and apoptosis.Variety growth factors may result a high-affinity site that can combinate with PI3K's p85 subunit by the growth factor receptor tyrosine (Y740/Y751) phosphorylation, then induce activation of PKB. This activation process can be blocked by PI3K specific inhibitors wortmannin and LY294002. The main mechanism is that wortmannin and LY294002 can combinate with PI3K's the p110 subunit by non-competitive irreversible ways, then blocks the downstream activation of AKT. and can inhibite the proliferation and migration of the lens epithelial cells(LEC). Phosphatidylinositol 3-kinase (PI3K) channel inhibitor, is expected to be the ideal drug to prevent PCO because of its less toxic side-effects.This experiment by adding different concentrations of LY294002 in vitro cultured rabbit lens epithelial cells to interfere with its proliferation. Inhibitory rate of cells was observed by MTT and flow cytometry analysis of cell cycle. In order to facilitate observation the extent of inhibition that LY294002 acts on rabbit lens epithelial cells.Materials and methodsRabbit lens epithelial cells were divided experimental groups and contrast groups, contrast groups were divided blank group and bFGF-induced group, experimental groups were divided only with LY294002 groups and combined bFGF-induced groups. Blank control group is consisted by serum-free DMEM medium, bFGF-induced group is consisted by blank control group and (10ng/mL). experimental groups were divided into two groups:â‘ Five only with LY294002 groups (10-8mol/L,10-7mol/L,10-6mol/L,10-5mol/L,10-4mol/L)â‘¡Five combined bFGF-induced groups (10-8mol/L,10-7mol/L,10-6mol/L,10-5mol/L,10-4mol/L) +bFGF(10ng/mL). The 3rd generation of rabbit LECs that grow well were passaged in culture plates and culture bottle. After the normal culture, we joined in different concentrations of grugs according to these groups. The rabbit LECs were cultured for 48 hours, then MTT tested inhibition rate and flow cytometry cell cycle.Results1. MTT tests LY294002 on inhibition of LEC proliferationApplication of LY294002 in a separate group:When the LY294002 group compered with blank control group, result showed 10-8mol/L concentrations of LY294002 effects on rabbit LECs after 48h is not significant inhibition (P>0.01),10-4 mol/L,10-5 mol/L,10-6mol/L,10-7mol/L concentration of LY294002 can significantly inhibite the proliferation of rabbit LECs(P<0.01). The combined treatment group compared with the positive control group, there was a slight inhibition of rabbit LEC, when 10-8mol/L concentration of LY294002 effects on rabbit LECs after 48h(P=0.08).10-4 mol/L,10-5 mol/L,10-6mol/L,10-7mol/L concentration of LY294002 can significantly inhibite the proliferation of rabbit LECs(P<0.01). When the combined treatment group compared with the control group, the difference was statistically significant(P<0.01).2. LY294002 on rabbit LEC of the cell cycleWhen the LY294002 effects on rabbit LECs after 48h, the experimental group compared with control group, G0/G1 phase cells increased, while the S phase and G2 /M phase cells decreased.Analysis of Cells in GO phase showed:Application of LY294002 in a separate group, when the LY294002 group compered with blank control group, the result showed 10-8mol/L concentrations of LY294002 effects on rabbit LECs after 48h is not significant inhibition (P>0.01).10-4 mol/L,10-5 mol/L,10-6mol/L,10-7mol/L concentration of LY294002 can significantly inhibite the proliferation of rabbit LECs(P<0.01). The combined treatment group compared with the positive control group, there was not significant inhibition of rabbit LEC, when 10-8mol/L concentration of LY294002 effects on rabbit LECs after 48h(P>0.01).10-4 mol/L, 10-5 mol/L,10-6mol/L,10-7mol/L concentration of LY294002 can significantly inhibite the proliferation of rabbit LECs(P<0.01). When the combined treatment group compared with the control group, the difference was statistically significant(P<0.01). The results of S phase and M phase are generally same with the G0/G1phase.Conclusion1. LY294002 can inhibit the rabbit lens epithelial cell proliferation.2. bFGF can promote rabbit lens epithelial cell proliferation, the number of cells in the proliferative phase was incread after combined with bFGF.3. The promoting cell proliferation of bFGF can be blocked, when the cincentrations of LY294002 is high, and the number of cells in the proliferative phase signficantly reduced, at the same time, the number of cells in quiescent phase is significantly increased.
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