Characterization of connective tissue growth factor expression in primary cultures of human tubular epithelial cells: modulation by hypoxia.
Tubular epithelial cells secrete connective tissue growth factor (CTGF, CCN2), which contributes to tubulointerstitial fibrosis. However, the molecular regulation of CTGF in human primary tubular epithelial cells (hPTECs) is not well defined. Therefore, CTGF expression was characterized in hPTECs isolated from healthy parts of tumor nephrectomies, with special emphasis on the regulation by transforming growth factor-beta (TGF-beta) and hypoxia, essential factors in the development of fibrosis. CTGF synthesis was strongly dependent on cell density. High CTGF levels were detected in sparse cells, whereas CTGF expression was reduced in confluent cells. Concomitantly, stimulation of CTGF by TGF-beta or the histone deacetylase inhibitor trichostatin was prevented in dense cells. Exposure of hPTECs to low oxygen tension (1% O2) or the hypoxia mimetic dimethyl-oxalylglycine for 24 h reduced CTGF gene expression in most of the 17 preparations analyzed. Preincubation of the cells under hypoxic conditions significantly reduced TGF-beta-mediated upregulation of CTGF. In line with these data, CTGF mRNA was only induced in interstitial cells, but not in tubular cells in kidneys of mice exposed to hypoxia. Longer exposure to hypoxia or TGF-beta (up to 72 h) did not induce hPTECs to adopt a mesenchymal phenotype characterized by upregulation of alpha-smooth muscle actin, downregulation of E-cadherin, or increased sensitivity of the cells in terms of CTGF expression. Sensitivity was restored by inhibition of DNA methylation. Taken together, our data provide evidence that exposure to hypoxia decreased CTGF gene expression. Furthermore, hypoxia per se was not sufficient to induce a mesenchymal phenotype in primary tubular epithelial cells.
Kroening
Sven
S
Department of Nephrology and Hypertension, University Hospital of Erlangen-Nuremberg, Erlangen, Germany.
Neubauer
Emily
E
Wullich
Bernd
B
Aten
Jan
J
Goppelt-Struebe
Margarete
M
D000600:Amino Acids, Dicarboxylic / Q000494:pharmacology; D018929:Cell Culture Techniques; D015687:Cell Hypoxia; D049109:Cell Proliferation; D002478:Cells, Cultured; D055513:Connective Tissue Growth Factor / Q000235:genetics / Q000378:metabolism*; D019175:DNA Methylation; D015254:DNA Modification Methylases / Q000037:antagonists & inhibitors / Q000378:metabolism; D003857:Deoxyuridine / Q000031:analogs & derivatives / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004847:Epithelial Cells / Q000187:drug effects / Q000378:metabolism*; D056572:Histone Deacetylase Inhibitors / Q000494:pharmacology; D006801:Humans; D006877:Hydroxamic Acids / Q000494:pharmacology; D007684:Kidney Tubules / Q000187:drug effects / Q000378:metabolism*; D010100:Oxygen / Q000378:metabolism; D010641:Phenotype; D012333:RNA, Messenger / Q000378:metabolism; D013997:Time Factors; D016212:Transforming Growth Factor beta / Q000378:metabolism; D015854:Up-Regulation
D000600:Amino Acids, Dicarboxylic; C526317:CCN2 protein, human; D004791:Enzyme Inhibitors; D056572:Histone Deacetylase Inhibitors; D006877:Hydroxamic Acids; D012333:RNA, Messenger; D016212:Transforming Growth Factor beta; D055513:Connective Tissue Growth Factor; C012589:trichostatin A; D015254:DNA Modification Methylases; D010100:Oxygen; C040947:oxalylglycine; D003857:Deoxyuridine