Effects Of KIM-1 On Autophagy Induced By High Glucose In Human Tubular Epithelial Cells

Background and aimsDiabetic nephropathy(DN) is one of the most serious complications of diabetes. Its main characteristic is the formation of progressive renal interstitial fibrosis(RIF), which is the primary reason for leading to the end- stage renal disease(ESRD). Therefore, it is of great significance to seek for the new targets of therapy for the prevention and treatment of diabetic nephropathy. Autophagy is one of the way to balance the metabolism of cellular protein and maintain the stability of introcellular environmental. Reactive oxygen species(ROS)、 endoplasmic reticulum stress and hypoxia are supposed to be the pathogenesis of diabetic nephropathy,and the activator of autophagy. Recent studies suggest that the increasing level of cell autophagy can promote the progress of RIF. Kidney injury molecular 1(KIM- 1) is one kind of transmembrane glycoprotein that expressed in the injuried proximal renal tubular epithelial cells. It is not only an important indicator for the extent of renal damage in acute kidney injury(AKI) and chronic kidney diseases(CKD), but also the participator in the process of injury and repairment in renal tubular epithelial cell. In the progression of AKI, KIM- 1 can transform the renal tubular epithelial cells into "semiprofessional" phagocytes and enhance the ability of cell autophagy. At the same time, in the progression of CKD,Which was caused by variety of protopathy(including diabetic nephropathy), there is a positive correlation between the extent of RIF and the level of expression of KIM- 1. Whether KIM– 1could regulate cell autophagy, and then participate in the progress of diabetic nephropathy is still not clear. This experiment is designed to consider the human tubular epithelial cells as the research object,and the high glucose environment as the stimulating factors. Observing the effects of KIM-l induced the expression of LC3II(microtubuleassociated protein 1 light chain 3II) and the formation of autophagosome when the cells were cultured at the different time of high glucose condition,clarifing the relationship between KIM-l and autophagy, and exploring the possible mechanism of KIM-1involved in the progression of diabetic nephropathy. So as to providing the new ideas and targets for the prevention and treatment of diabetic nephropathy. MethodsHK2 were cultured in vitro and divided into five groups as follows:(1) Normal control group(D-glucose 5.6 mmol/L);(2) Hypertonic group(D-glucose 5.6 mmol/L+D-mannitol 24.4 mmol/L);(3) High glucose group(D-glucose 30 mmol/L);(4) KIM-1 si RNA group;(5) Control si RNA group.The corresponding indicators were measured at different time.Western blotting was used to detect the protein expression of KIM-1 and autophagy protein microtubule-associated protein 1 light chain 3II(LC3II);Real Time-PCR was used to detect m RNA expression of KIM-1 and LC3II;The autophagosomes formed in human tubular epithelial cells were observed by transmission electron microscope(TEM). ResultsCompared with the control group,the protein and m RNA expression of KIM-1 and LC3 II in the high glucose group were increased in a time-dependent manner(P<0.05);the number of autophagosomes in the high glucose group were increased in a time-dependent manner(P< 0.05).Compared with the high glucose group,the protein and m RNA expressions of KIM-1 and LC3 II in KIM-1 si RNA group were decreased(P<0.05);the number of autophagosomes in the high glucose group were reduced(P<0.05). Conclusions1、The expression of KIM1 and LC3II and the number of autophagosome are increased in renal tubular cells when cultured in high glucose environment;2、Down-regulating the expression of KIM-1 can inhibit the expression of LC3 II and the formation of autophagosomes, which suggests that KIM-1 may be involved in the progress of DN by regulating the autophagy in human tubular epithelial cells.