Details

Autor(en) / Beteiligte

• MORCOS, Michael
• SAYED, Ahmed A. R
• TRITSCHLER, Hans
• KASPER, Michael
• SCHWENGER, Vedat
• HAMANN, Andreas
• DUGI, Klaus A
• SCHMIDT, Anne-Marie
• STERN, David
• ZIEGLER, Reinhard
• HAERING, Hans U
• ANDRASSY, Martin
• BIERHAUS, Angelika
• VAN DER WOUDE, Fokko
• NAWROTH, Peter P
• YARD, Benito
• WALDHERR, Rüdiger
• MERZ, Wolfgang
• KLOETING, Ingrid
• SCHLEICHER, Erwin
• MENTZ, Stefani
• ABD EL BAKI, Randa F

Titel

Activation of Tubular Epithelial Cells in Diabetic Nephropathy

Ist Teil von

• Diabetes (New York, N.Y.), 2002-12, Vol.51 (12), p.3532-3544

Ort / Verlag

Alexandria, VA: American Diabetes Association

Erscheinungsjahr

2002

Link zum Volltext

Quelle

MEDLINE

Beschreibungen/Notizen

• Activation of Tubular Epithelial Cells in Diabetic Nephropathy Michael Morcos 1 , Ahmed A.R. Sayed 1 , Angelika Bierhaus 1 , Benito Yard 2 , Rüdiger Waldherr 3 , Wolfgang Merz 4 , Ingrid Kloeting 5 , Erwin Schleicher 6 , Stefani Mentz 1 , Randa F. Abd el Baki 1 , Hans Tritschler 7 , Michael Kasper 8 , Vedat Schwenger 1 , Andreas Hamann 1 , Klaus A. Dugi 1 , Anne-Marie Schmidt 9 , David Stern 9 , Reinhard Ziegler 1 , Hans U. Haering 6 , Martin Andrassy 1 , Fokko van der Woude 2 and Peter P. Nawroth 1 1 Department of Internal Medicine 1 and Department of Nephrology, University of Heidelberg, Heidelberg, Germany 2 Department of Nephrologie, University Hospital of Mannheim, Mannheim, Germany 3 Gemeinschaftspraxis für Pathologie, Heidelberg, Germany 4 Biochemiezentrum, University of Heidelberg, Germany 5 Department of Laboratory Animal Science, Institute of Pathophysiology, Faculty of Medicine, University Greifswald, Greifswald, Germany 6 Department of Medicine, University of Tuebingen, Tuebingen, Germany 7 Asta Medica, Frankfurt, Germany 8 Department of Anatomy, Department of Pathology and Institute of Food Chemistry, Technical University Dresden, Dresden, Germany 9 College of Surgeons, Columbia University, New York, New York Abstract Previous studies have shown that renal function in type 2 diabetes correlates better with tubular changes than with glomerular pathology. Since advanced glycation end products (AGEs; AGE-albumin) and in particular carboxymethyllysine (CML) are known to play a central role in diabetic nephropathy, we studied the activation of nuclear factor κB (NF-κB) in tubular epithelial cells in vivo and in vitro by AGE-albumin and CML. Urine samples from healthy control subjects ( n = 50) and type 2 diabetic patients ( n = 100) were collected and tested for excretion of CML and the presence of proximal tubular epithelial cells (pTECs). CML excretion was significantly higher in diabetic patients than in healthy control subjects ( P < 0.0001) and correlated with the degree of albuminuria ( r = 0.7, P < 0.0001), while there was no correlation between CML excretion and HbA 1c ( r = 0.03, P = 0.76). Urine sediments from 20 of 100 patients contained pTECs, evidenced by cytokeratin 18 positivity, while healthy control subjects ( n = 50) showed none ( P < 0.0001). Activated NF-κB could be detected in the nuclear region of excreted pTECs in 8 of 20 patients with pTECs in the urine sediment (40%). Five of eight NF-κBp65 antigen-positive cells stained positive for interleukin-6 (IL-6) antigen (62%), while only one of the NF-κB-negative cells showed IL-6 positivity. pTECs in the urine sediment correlated positively with albuminuria ( r = 0.57, P < 0.0001) and CML excretion ( r = 0.55, P < 0.0001). Immunohistochemistry in diabetic rat kidneys and a human diabetic kidney confirmed strong expression of NF-κB in tubular cells. To further prove an AGE/CML-induced NF-κB activation in pTECs, NF-κB activation was studied in cultured human pTECs by electrophoretic mobility shift assays (EMSAs) and Western blot. Stimulation of NF-κB binding activity was dose dependent and was one-half maximal at 250 nmol/l AGE-albumin or CML and time dependent at a maximum of activation after 4 days. Functional relevance of the observed NF-κB activation was demonstrated in pTECs transfected with a NF-κB-driven luciferase reporter plasmid and was associated with an increased release of IL-6 into the supernatant. The AGE- and CML-dependent activation of NF-κBp65 and NF-κB-dependent IL-6 expression could be inhibited using the soluble form of the receptor for AGEs (RAGE) (soluble RAGE [sRAGE]), RAGE-specific antibody, or the antioxidant thioctic acid. In addition transcriptional activity and IL-6 release from transfected cells could be inhibited by overexpression of the NF-κB-specific inhibitor κBα. The findings that excreted pTECs demonstrate activated NF-κB and IL-6 antigen and that AGE-albumin and CML lead to a perpetuated activation of NF-κB in vitro infer that a perpetuated increase in proinflammtory gene products, such as IL-6, plays a role in damaging the renal tubule. Footnotes Address correspondence and reprint requests to Michael Morcos, MD, Department of Internal Medicine 1, University of Heidelberg, Bergheimerstr 58, 69115 Heidelberg, Germany. E-mail: michael_morcos{at}med.uni-heidelberg.de . Received for publication 25 February 2002 and accepted in revised form 13 September 2002. M.M. and A.A.R.S. contributed equally to this study. AGE, advanced glycation end product; CC, contingency coefficient; CML, carboxymethyllysine; ELISA, enzyme-linked immunosorbent assay; EMSA, electrophoretic mobility shift assay; IκB, inhibitory κB; LPS, lipopolysaccharide; NF-κB, nuclear factor κB; pTEC, proximal tubular epithelial cell; RAGE, receptor for AGEs; sRAGE, soluble RAGE; TA, thioctic acid. DIABETES