The relationship between high glucose-induced secretion of IGFs and PKC or oxidative stress in mesangial cells

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(1)³¤¥Èó. * .* a. (2004) 44 4 Korean J Vet Res(2004) 44(4) : 497~505. .FTBOHJBM^

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(4). *ÎL >~ ÒL. *§L >~ ÒL , Ún*W \² (²Òß: 2004j 11ú 8¢). 1. The relationship between high glucose-induced secretion of IGFs and PKC or oxidative stress in mesangial cells Soo-hyun Park, Jung-sun Heo, Chang-won Kang1 and Ho-jae Han* Department of Veterinary Physiology, College of Veterinary Medicine, Chonnam National University, Gwangju 500-757, Korea 1 Bio-safety Research Institute, College of Veterinary Medicine, Chonbuk National University, Jeonju 561-756, Korea (Accepted= November 8, 2004) Abstract : The proliferation of mesangial cells has been associated with the development of diabetic nephropathy. The cell proliferation has been regulated by diverse growth factors. Among them, insulin like growth factors(IGFs) are also involved in the pathogenesis of diabetic nephropathy. However, it is not yet known about the effect of high glucose on IGF-I and IGF-II secretion and the relationship between high glucose-induced secretion of IGFs and PKC or oxidative stress in the mesangial cells. Thus, we examined the mechanisms by which high glucose regulates secretion of IGFs in mesangial cells. High glucose(25 mM) increased IGF-I and IGF-II secretion. High glucose-induced increase of IGF-I and IGF-II secretion were blocked by taurine(2×10−3 M), N-acetyl cystein(NAC, 10−5 M), or GSH(10−5 M) (antioxidants), suggesting the role of oxidative stress. High glucose-induced secretion of IGF-I and IGF-II were blocked by H-7, staurosporine, and bisindolylmaleimide I(protein kinase C inhibitors). On the other hand, high glucose also increased lipid peroxide (LPO) formation in a dose dependent manner. In addition, high glucoseinduced stimulation of LPO formation was blocked by PKC inhibitors. These results suggest that PKC is responsible for the increase of oxidative stress in the action of high glucose-induced secretion of IGF-I and IGF-II in mesangial cells. In conclusion, high glucose stimulates IGF-I and IGF-II secretion via PKCoxidative stress signal pathways in mesangial cells. Key words : Insulin-like growth factors, high glucose, PKC, oxidative stress, mesangial cell, kidney. * . ^

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(72) 502. ;>*Áî;FÁ;?öÁ^Ò. Fig. 6. Effects of antioxidants (A) and PKC inhibitors (B) on high glucose-induced increase of lipid peroxide formation. Mesangial cells were incubated with NAC (10−5 M), GSH (10−5 M), taurine (2 mM), staurosporine (10−8 M), H-7 (10−4 M), or bisindolylmaleimide I (10−6 M) for 30 min prior to the treatment of 25 mM glucose or were incubated with 25 mM glucose alone or together with hydrogen peroxide (10−5 M) or TPA (100 ng/ml ) for 72 hr. Values are means ±S.E. of 9 separate experiments performed on 3 different cultures. *p<0.05 vs. control, **p <0.05 vs. 25 mM glucose.. ¶ ÿö Ã& º Öf ¢~~& [18]. ~ æò ·Ï ÚÆ ^*ã¢ Û~ Úæºæö &Bº C&ææ p~ . ÖzW ÊÞ.Êº ÷W Ã~ B÷ &7 çê¢ < ®b [27], STZö ~ FêB ÷ ÑÞ 5 ÷ ~¶~ ãÖö WÖ²ö V ç ¶ç ¢Ú¾º ©b > ® [44]. . þöBê

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(99) êf PKC-ÖzW ÊÞ.Ê ^* ã ¢ Û~ IGF-I 5 IGF-II ªj·Ïj Ã&Êº © b .. 7.. 8.. 9.. 10.. 11.. ^^ò 1. Abou-Seif, M. A. and Youssef, A. A. Oxidative stress and male IGF-1, gonadotropin and related hormones in diabetic patients. Clin. Chem. Lab. Med. 2001, 39, 618623. 2. Agardh, C. D., Stenram, U., Torffvit, O. and Agardh, E. R. Effects of inhibition of glycation and oxidative stress on the development of diabetic nephropathy in rats. J. Diabetes Complications. 2002, 16, 395-400. 3. Babazono, T., Kapor-Drezgic, J., Dlugosz, J. A. and Whiteside, C. Altered expression and subcellular localization of diacylglycerol-sensitive protein kinase C isoforms in diabetic rat glomerular cells. Diabetes. 1998, 47, 668-676. 4. Berfield, A. K., Spicer, D. and Abrass, C. K. Insulinlike growth factor I (IGF-I) induces unique effects in the cytoskeleton of cultured rat glomerular mesangial cells. J. Histochem. Cytochem. 1997, 45, 583-593. 5. Binoux, M. The IGF system in metabolism regulation. Diabetes Metab. 1995, 21, 330-337. 6. Bowsher, R. R., Lee, W. H., Apathy, J. M., O'Brien, P. J., Ferguson, A. L. and Henry, D. P. Measurement of insulin-like growth factor-II in physiological fluids. 12.. 13.. 14.. 15.. 16.. 17.. 503. and tissues. I. An improved extraction procedure and radioimmunoassay for human and rat fluids. Endocrinology. 1991, 128, 805-814. Catherwood, M. A., Powell, L. A., Anderson, P., McMaster, D., Sharpe, P. C. and Trimble, E. R. Glucose-induced oxidative stress in mesangial cells. Kidney Int. 2002, 61, 599-608. Craven, P. A., Davidson, C. M. and DeRubertis, F. R. Increase in diacylglycerol mass in isolated glomeruli by glucose from de novo synthesis of glycerolipids. Diabetes. 1990, 39, 667-674. Daughaday, W. H. and Rotwein, P. Insulin-like growth factors I and II. Peptide messenger ribonucleic acid and gene structure serum, and tissue concentrations. Endocr. Rev. 1989, 10, 68-91. De La Puente, A., Goya, L., Ramos, S., Martin, M. A., Alvarez, C., Escriva, F. and Pascual-Leone, A. M. Effects of experimental diabetes on renal IGF/ IGFBP system during neonatal period in the rat. Am. J. Physiol. Renal. Physiol. 2000, 279, F1067-F1076. Derubertis, F. R. and Craven, P. A. Activation of protein kinase C in glomerular cells in diabetes. Mechanisms and potential links to the pathogenesis of diabetic glomerulopathy. Diabetes. 1994, 43, 1-8. Elliot, S. J., Striker, L. J., Hattori, M., Yang, C. W., He, C. J., Peten, E. P. and Striker, G. E. Mesangial cells from diabetic NOD mice constitutively secrete increased amounts of insulin-like growth factor-I. Endocrinology. 1993, 133, 1783-1788. Feld, S. and Hirschberg, R. Growth hormone, the insulin-like growth factor system, and the kidney. Endocr Rev. 1996, 17, 423-480. Flyvbjerg, A., Landau, D., Domene, H., Hernandez, L., Gronbaek, H. and LeRoith, D. The role of growth hormone, insulin-like growth factors (IGFs), and IGFbinding proteins in experimental diabetic kidney disease. Metabolism. 1995, 44, 67-71. Gooch, J. L., Tang, Y., Ricono, J. M. and Abboud, H. E. Insulin-like growth factor-I induces renal cell hypertrophy via a calcineurin-dependent mechanism. J. Biol. Chem. 2001, 276, 42492-42500. Gronbaek, H., Nielsen, B., Frystyk, J., Flyvbjerg, A. and Orskov, H. Effect of lanreotide on local kidney IGF-I and renal growth in experimental diabetes in the rat. Exp. Nephrol. 1996, 4, 295-303. Ha, H., Yu, M. R., Choi, Y. J, Kitamura, M. and Lee, H. B. Role of high glucose-induced nuclear factor-.

(100) 504. 18.. 19.. 20.. 21.. 22.. 23.. 24.. 25.. ;>*Áî;FÁ;?öÁ^Ò kappaB activation in monocyte chemoattractant protein1 expression by mesangial cells. J. Am. Soc. Nephrol. 2002, 13, 894-902. Heo, Y. R., Kang, C. W. and Cha, Y. S. L-Carnitine changes the levels of insulin-like growth factors (IGFs) and IGF binding proteins in streptozotocin-induced diabetic rat. J. Nutr. Sci. Vitaminol. 2001, 47, 329-334. Hoog, A., Sandberg-Nordqvist, A. C., Abdel-Halim, S. M,, Carlsson-Skwirut, C., Guenifi, A., Tally, M., Ostenson, C. G., Falkmer, S., Sara, V. R., Efendic, S., Schalling, M. and Grimelius, L. Increased amounts of a high molecular weight insulin-like growth factor II (IGF-II) peptide and IGF-II messenger ribonucleic acid in pancreatic islets of diabetic GotoKakizaki rats. Endocrinology. 1996, 137, 2415-2423. Inoguchi, T., Li, P., Umeda, F., Yu, H. Y., Kakimoto, M., Imamura, M., Aoki, T., Etoh, T., Hashimoto, T., Naruse, M., Sano, H., Utsumi, H. and Nawata, H. High glucose level and free fatty acid stimulate reactive oxygen species production through protein kinase Cdependent activation of NAD(P)H oxidase in cultured vascular cells. Diabetes. 2000, 49, 1939-1945. Iori, E., Marescotti, M. C., Vedovato, M., Ceolotto, G., Avogaro, A., Tiengo, A., Del Prato, S. and Trevisan, R. In situ protein Kinase C activity is increased in cultured fibroblasts from Type 1 diabetic patients with nephropathy. Diabetologia. 2003, 46, 524530. Kimura, M., Ishizawa, M., Miura, A., Itaya, S., Kanoh, Y., Yasuda, K., Uno, Y., Morita, H. and Ishizuka, T. Platelet protein kinase C isoform content in type 2 diabetes complicated with retinopathy and nephropathy. Platelets. 2001, 12, 138-143. Koya, D., Jirousek, M. R., Lin, Y. W., Ishii, H., Kuboki, K. and King, G. L. Characterization of protein kinase C beta isoform activation on the gene expression of transforming growth factor-beta, extracellular matrix components, and prostanoids in the glomeruli of diabetic rats. J. Clin. Invest. 1997, 100, 115-126. Kumar, A., Hawkins, K. S., Hannan, M. A. and Ganz, M. B. Activation of PKC-beta(I) in glomerular mesangial cells is associated with specific NF-kappaB subunit translocation. Am. J. Physiol. Renal. Physiol. 2001, 281, F613-F619. Kurzawa, R., Glabowski, W., Baczkowski, T. and Brelik, P. Evaluation of mouse preimplantation embryos. 26.. 27.. 28.. 29.. 30.. 31.. 32.. 33.. 34.. 35.. 36.. exposed to oxidative stress cultured with insulin-like growth factor I and II, epidermal growth factor, insulin, transferrin, and selenium. Reprod. Biol. 2002, 2, 143162. Lee, C. Y. and Henricks, D. M. Comparisions of various acidic treatments of bovine serum on insulinlike growth factor-I immunoreactive and binding activity. J. Endocrinol, 1990, 127, 139-148. Lee, H. B., Yu, M. R., Yang, Y., Jiang, Z. and Ha, H. Reactive oxygen species-regulated signaling pathways in diabetic nephropathy. J. Am. Soc. Nephrol. 2003, 14, S241-S245. Mathews, L. S., Norstedt, G. and Palmiter, R. D. Regulation of insulin-like growth factor I gene expression by growth hormone. Proc. Natl. Acad. Sci. USA. 1986, 83, 9343-9347. Miyatake, N., Shikata, K., Wada, J., Sugimoto, H., Takahashi, S. and Makino, H. Differential distribution of insulin-like growth factor-1 and insulin-like growth factor binding proteins in experimental diabetic rat kidney. Nephron. 1999, 81, 317-323. Nishikawa, T., Edelstein, D., Du, X. L., Yamagishi, S., Matsumura, T., Kaneda, Y., Yorek, M. A. Beebe, D., Oates, P. J., Hammes, H. P., Giardino, I. and Brownlee, M. Normalizing mitochondrial superoxide production blocks three pathways of hyperglycaemic damage. Nature. 2000, 404, 787-790. Obrosova, I. G., Fathallah, L., Liu, E. and NouroozZadeh, J. Early oxidative stress in the diabetic kidney: effect of DL-alpha-lipoic acid. Free. Radic. Biol. Med. 2003, 34, 186-195. Ohkawa, H., Ohishi, N. and Yagi, K. Assay for lipid peroxides in animal tissues by thiobarbituric acid reaction. Anal. Biochem. 1979, 95, 351-358. Rossert, J., Terraz-Durasnel, C. and Brideau, G. Growth factors, cytokines, and renal fibrosis during the course of diabetic nephropathy. Diabetes. Metab. 2000, 26, S16-S24. Schleicher, E. D. and Olgemoller, B. Glomerular changes in diabetes mellitus. Eur. J. Clin. Chem. Clin. Biochem. 1992, 30, 635-640. Schwander, J. C., Hauri, C., Zapf, J. and Froesch, E. R. Synthesis and secretion of insulin-like growth factor and its binding protein by the perfused rat liver: dependence on growth hormone status. Endocrinology. 1983, 113, 297-305. Sheetz, M. J. and King, G. L. Molecular understanding.

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(103) êö ~ IGFs ªjf PKC 5 ÖzW ÊÞ.Êf~ NWö \. of hyperglycemia’s adverse effects for diabetic complications. JAMA. 2002, 288, 2579-2588. Spranger, J., Buhnen, J., Jansen, V., Krieg, M., Meyer-Schwickerath, R., Blum, W. F., Schatz, H. and Pfeiffer, A. F. Systemic levels contribute significantly to increased intraocular IGF-I, IGF-II and IGF-BP3 [correction of IFG-BP3] in proliferative diabetic retinopathy. Horm. Metab. Res. 2000, 32, 196200. Steff, M.W., Osterby, W. R., Chavers, B. and Mauer, S. M. Mesangial expansion as a central mechanism for loss of kidney function in diabetic patients. Diabetes. 1998, 38, 1077-1081. Studer, R. K, Craven, P. A. and DeRubertis, F. R. Antioxidant inhibition of protein kinase C-signaled increases in transforming growth factor-beta in mesangial cells. Metabolism. 1997, 46, 918-925. Tack, I., Elliot, S. J., Potier, M., Rivera, A., Striker, G. E. and Striker, L. J. Autocrine activation of the IGF-I signaling pathway in mesangial cells isolated from diabetic NOD mice. Diabetes. 2002, 51, 182-188. Tuttle, K. R. and Anderson, P. W. A novel potential. 42. 43.. 44. 45.. 46.. 505. therapy for diabetic nephropathy and vascular complications: protein kinase C beta inhibition. Am. J. Kidney Dis. 2003, 42, 456-465. Wardle, E. N. How does hyperglycaemia predispose to diabetic nephropathy? QJM. 1996, 89, 943-951. Werner, H., Shen-Orr, Z., Stannard, B., Burguera, B., Roberts, C. T. Jr. and LeRoith, D. Experimental diabetes increases insulin-like growth factor I and II receptor concentration and gene expression in kidney. Diabetes. 1990, 39, 1490-1497. West, I. C. Radicals and oxidative stress in diabetes. Diabet Med. 2000, 17, 171-180. Yakar, S., Wu, Y., Setser, J. and Rosen, C. J. The role of circulating IGF-I: lessons from human and animal models. Endocrine. 2002, 19, 239-248. Zhuang, H. X., Wuarin, L., Fei, Z. J. and Ishii, D. N. Insulin-like growth factor (IGF) gene expression is reduced in neural tissues and liver from rats with noninsulin-dependent diabetes mellitus, and IGF treatment ameliorates diabetic neuropathy. J. Pharmacol. Exp. Ther. 1997, 283, 366-374..

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