Growth Regulation of High Glucose and Advanced Glycosylation End-product on MDCK, a Distal Tubular Cell Line - an Exploration on Transforming Growth Factor-b and its Receptors

• Main Authors: Yang, Yu Lin, 楊堉麟
• Other Authors: Chuang Lea Yea
• Format: Others
• Language: zh-TW
• Published: 1998
• Online Access: http://ndltd.ncl.edu.tw/handle/45255080018713693753

博士 === 高雄醫學院 === 醫學研究所 === 86 === Diabetes mellitus and renal disease is the 5th and 7th leading cause of death in Taiwan. Hence, they consume a major portion of medical resources in our country. Furthermore, diabetic nephropathy is a major cause of diabetic morbidity and mortality. Unfortunately, diabetic patients will almost always enter end-stage renal disease (ESRD) once they develop proteinuria. Therefore, the study for the pathogenesis of diabetic nephropathy has become a major topic in biomedical research.Hyperglycemia and advanced glycation end-product (AGE) are two of the essential factors in diabetic nephropathy. The pathology of diabetic nephropathy is characterized by cellular hyperplasia, hypertrophy and the expansion of extracellular matrix which result in renal fibrosis and ESRD. These processes are intimately associated with cytokines/growth factors, esp. transforming growth factor-b (TGF-b). The distal nephron is also important in diabetic nephropathy, although most studies regarding diabetic nephropathy were focused on glomerulopathy and occasionally proximal tubule. Therefore, we performed a series of studies in high glucose and AGE-cultured distal tubule-like MDCK cells. We found that, unlike other renal cells, high glucose did not increase TGF-b production, but it did increase the responsiveness of MDCK cells to TGF-b, which includes: inhibition of cellular mitogenesis, induction of cellular hypertrophy, increase of cell cycle-regulatory retinoblastoma protein (pRb) dephosphorylation and inhibition of cdc2 kinase activity. Affinity-labeling experiments showed that high glucose may increase TGF-b responsiveness by increasing type I and II TGF-b receptor protein expressions. This is the first demonstration that distal tubule is unique in that it responds to high glucose by increasing TGF-b (which may be derived from paracrine sources) responsiveness but not the production of endogenous TGF-b.Regarding the roles of intracellular signal transduction pathways in diabetic nephropathy, protein kinase C (PKC) had been shown to be important. However, the roles of various PKC isoenzymes in diabetic tubulopathy is still not known. Thus, we showed that high glucose induced PKC activation, PKCi and PKCe activation, cytosolic translocation of PKCi(l) and membrane translocation of PKCe. As for the roles of various transcription factors, only AP-1 had been suggested to be involved in diabetic glomerulopathy. Thus, we first showed that high glucose induced activation of transcription factors AP-1 and NF-kB in MDCK cells concomitantly with the induction of type II TGF-b receptor mRNA. We speculate that the above changes in the signal transduction pathways may be involved in the induction of type II TGF-b receptor mRNA, although this speculation awaits further confirmation.Regarding the effects of AGE, we found that, unlike high glucose, AGE inhibited cellular mitogenesis while inducing cellular hypertrophy in the MDCK cells. Moreover, AGE induced the production of bioactive TGF-b in these cells. Importantly, neutralizing anti-TGF-b1 antibody reversed the above AGE-induced effects. Therefore, endogenous TGF-b1 may mediate the above AGE- induced effects in the MDCK cells.Our experiments showed that distal tubular cells behave differently from glomerular and proximal tubular cells in high glucose and AGE cultures. Whereas AGE induced bioactive TGF-b, high glucose only induced the expression of type I and II TGF-b receptors. We conclude that the complex interaction between high glucose, AGE, TGF-b, TGF-b receptors and cell cycle-regulatory proteins (pRb and cdc2) may play important roles in diabetic nephropathy. In addition, PKCi( l), PKCe, AP-1 and NF-kB may mediate some of the above high glucose and AGE-induced effects.