Regulation of connective tissue growth factor activity in cultured rat mesangial cells and its expression in experimental diabetic glomerulosclerosis.
Connective tissue growth factor (CTGF) is a peptide secreted by cultured endothelial cells and fibroblasts when stimulated by transforming growth factor-beta (TGF-beta), and is overexpressed during fibrotic processes in coronary arteries and in skin. To determine whether CTGF is implicated in the pathogenesis of diabetic glomerulosclerosis, cultured rat mesangial cells (MC) as well as kidney cortex and microdissected glomeruli were examined from obese, diabetic db/db mice and their normal counterparts. Exposure of MC to recombinant human CTGF significantly increased fibronectin and collagen type I production. Furthermore, unstimulated MC expressed low levels of CTGF message and secreted minimal amounts of CTGF protein (36 to 38 kD) into the media. However, sodium heparin treatment resulted in a greater than fourfold increase in media-associated CTGF, suggesting that the majority of CTGF produced was cell- or matrix-bound. Exposure of MC to TGF-beta, increased glucose concentrations, or cyclic mechanical strain, all causal factors in diabetic glomerulosclerosis, markedly induced the expression of CTGF transcripts, while recombinant human CTGF was able to autoinduce its own expression. TGF-, and high glucose, but not mechanical strain, stimulated the concomitant secretion of CTGF protein, the former also inducing abundant quantities of a small molecular weight form of CTGF (18 kD) containing the heparin-binding domain. The induction of CTGF protein by a high glucose concentration was mediated by TGF-beta, since a TGF-beta-neutralizing antibody blocked this stimulation. In vivo studies using quantitative reverse transcription-PCR demonstrated that although CTGF transcripts were low in the glomeruli of control mice, expression was increased 28-fold after approximately 3.5 mo of diabetes. This change occurred early in the course of diabetic nephropathy when mesangial expansion was mild, and interstitial disease and proteinuria were absent. A substantially reduced elevation of CTGF mRNA (twofold) observed in whole kidney cortices indicated that the primary alteration of CTGF expression was in the glomerulus. These results suggest that CTGF upregulation is an important factor in the pathogenesis of mesangial matrix accumulation and progressive glomerulosclerosis, acting downstream of TGF-beta.
Riser
Bruce L
BL
Department of Internal Medicine, Division of Nephrology, Henry Ford Hospital, Detroit, Michigan.
Denichilo
Mark
M
FibroGen, South San Francisco, California.
Cortes
Pedro
P
Department of Internal Medicine, Division of Nephrology, Henry Ford Hospital, Detroit, Michigan.
Baker
Cathryn
C
FibroGen, South San Francisco, California.
Grondin
Janet M
JM
Department of Internal Medicine, Division of Nephrology, Henry Ford Hospital, Detroit, Michigan.
Yee
Jerry
J
Department of Internal Medicine, Division of Nephrology, Henry Ford Hospital, Detroit, Michigan.
Narins
Robert G
RG
Department of Internal Medicine, Division of Nephrology, Henry Ford Hospital, Detroit, Michigan.
D000818:Animals; D002478:Cells, Cultured; D055513:Connective Tissue Growth Factor; D003921:Diabetes Mellitus, Experimental / Q000378:metabolism*; D003928:Diabetic Nephropathies / Q000378:metabolism* / Q000473:pathology; D004195:Disease Models, Animal; D005109:Extracellular Matrix / Q000235:genetics; D005353:Fibronectins / Q000378:metabolism; D015870:Gene Expression; D005920:Glomerular Mesangium / Q000378:metabolism* / Q000473:pathology; D005947:Glucose / Q000378:metabolism; D006133:Growth Substances / Q000235:genetics / Q000378:metabolism*; D017874:Immediate-Early Proteins; D036341:Intercellular Signaling Peptides and Proteins; D007668:Kidney / Q000378:metabolism; D008297:Male; D051379:Mice; D008820:Mice, Obese; D012333:RNA, Messenger / Q000032:analysis; D051381:Rats; D011916:Rats, Inbred F344; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013314:Stress, Mechanical; D016212:Transforming Growth Factor beta / Q000378:metabolism; D015854:Up-Regulation
C526317:CCN2 protein, human; C526318:CCN2 protein, mouse; C526316:CCN2 protein, rat; D005353:Fibronectins; D006133:Growth Substances; D017874:Immediate-Early Proteins; D036341:Intercellular Signaling Peptides and Proteins; D012333:RNA, Messenger; D016212:Transforming Growth Factor beta; D055513:Connective Tissue Growth Factor; D005947:Glucose