| Background and Objectives:Diabetic nephropathy(DN) is one of the main primary diseases resulting in the end-stage renal disease (ESRD). The degree of renal interstitium lesions which characterized as extracellular matrix accumulating determines its long-term prognosis. Tubular epithelial cells can undergo epithelial-mesenchymal transition(EMT), thus promts renal intertsitial fibrosis.β-catenin,a multitasking molecule, plays a dual function in epithelial cells. One as an integral structural component of cadherin-based adherens junctions, the other as a key effector for signal transduction. The later one is related with its metabolism.:cytoplasmic P-catenin is constantly degraded by the action of the axin complex, which is composed of many kinds of proteins, some have enzymatic activity. β-catenin is phosphorylated by CK1and GSK3sequentially, resulting in its recognition by β-trcp, and subsequent ubiquitination and proteasomal degradation.β-catenin phosphorylatin determines this degradation,so non-phosphorylated β-catenin is the active and stable form of β-catenin, it can’t be recognized, so accumulates in the cytoplasm and travels to the nucleus,combines with TCF/LEF and activates target genes expression. Many studies show β-catenin is activated in the EMT in DN, they often observe the changes of location of β-catenin by immunohistochemistry assay,or detect the changes of protein expression of total/nuclear β-catenin by western blot, few detect the non-phosphorylated (3-catenin derectly,no such study was reported in DN.Cordyceps sinensis is one of famous traditional chinese medicine,which is widely used in the therapy of kidney disease,and shows certain effect for DN. Recent studies suggest its renal protection effect may involve the mechanism of inhibiting EMT. It can ameliorate the high glucose-induced phenotype changes in tubular epithelial cells, reduce urinary protein excretion and renal interstitial collagen deposition, alleviate the renal injury, but the concrete details behide this is still unknown.This reserch aims at detecting non-phosphorylated (3-catenin expression in high glucose environment cultured HK-2cells, exploring the relationship between β-catenin activation and EMT. Furthermore, observe the changes after cordyceps sinensis’intervention, trying to find possible new mechanism for its renal protection effect. Methods:HK-2cells were divided into the following six groups:normal group, high glucose group, mannitol group, high glucose+5μg/ml cordyceps sinensis group (HG+C5), high glucose+10μug/ml cordyceps sinensis group (HG+C10), high glucose+20μg/ml cordyceps sinensis group (HG+C20). After cultured for48hours, extract total cell protein, and detect the E-cadherin, a-SMA, non-phosphorylated β-catenin expression by Western blot.Results:Compared to the normal group, the expression of E-cadherin decreased(P<0.01), the expression of a-SMA and non-phosphorylated β-catenin increased in the high glucose group (P<0.01). After the cordyceps sinensis intervention, compared to High glucose group, the expression of E-cadherin in both HG+C10group and HG+C20group are up-regulated(P<0.01), the expression of a-SMA in all three intervention groups are down-regulated (P<0.01); the non-phosphorylated P-catenin in all three intervention groups are down-regulated, and these decreases are concentration-dependent on cordyceps sinensis.Conclusions:E-cadherin expression decreased and a-SMA expression increased in high glucose environment cultured HK-2cells, indicates that high glucose can induce Tubular epithelial cells undergoing Epithelial-mesenchymal transition(EMT). Non-phosphorylated P-catenin expression(the active β-catenin)is up-regulated at the same time, suggests it as a possible promoting factor for EMT. Cordyceps sinensis intervention can up-regulate E-cadherin expression and down-regulate α-SMA expression in high glucose environment cultured HK-2cells, this effect of inhibiting EMT may involve the mechanism of preveting the activation of β-catenin. |
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