Fibronectin turnover in human mesangial cell cultures as affected by Adriamycin |
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Authors: | Mechthild Soose Simone Wenzel Hilmar Stolte |
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Institution: | (1) Institute of Animal Physiology, Justus-Liebig-University Giessen, Wartweg 95, 6300 Giessen, Germany;(2) Department of Nephrology, Medical School Hannover, Hannover, Germany |
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Abstract: | Fibronectin (FN) turnover and turnover changes induced by the anticancer drug Adriamycin (ADR) were measured in human mesangial cells (HMC) in vitro. HMC cultures synthesize cellular FN (2.2+-0.3% of totalprotein synthesis; n = 12) which is secreted and incorporated into a fibrillar extracellular matrix (ECM). A 24 hr incubation of HMC with ADR (0.5–5 g/ml) resulted in an accumulation of FN in the culture medium, with a maximum increase following 5 pglml(7.3+-2.3pg/cell vs. controls: 4.4+-1.9pg/cell; n= 10). Correspondingly, radioactively labeled immunoprecipitable FN was increased in a dosage-dependent manner in the culture medium up to 50% vs. controls. The incorporation of radioactively labeled FN into ECM was significantly increased following 2 g ADR/ml. In accordance, immunofZuorescence staining revealed an expansion ofpericellular FNfibers in cultures exposed to 2 g ADR/ml. Concomitant with the accumulation of extracelhlar FN, radioactively labeled FN in the cells was reduced by 22%. Qualitative characterization of FN patterns revealed a diminished number of degradation products in the culture medium ofADR-treated HMC. These data suggest thatADR interferes with the turnover of FN secreted by HMC in vitro in such a way that FN accumulates extracellularly. This in turn leads to a reduced FN synthesis. These findings are compatible with a loss of urinary FN degradation products accompanying the onset ofproteinuria in ADR-treated rats.Abbreviations ADR
adriamycin
- BSA
bovine serum albumin
- DTT
dithiothreitol
- ECM
extracellular matrix
- EDTA
ethylenediamine tetraacetic acid disodium salt
- ELISA
enzyme-linked immunosorbent assay
- FCS
fetal calf serum
- FITC
fluorescein isothiocyanate
- FN
fibronectin
- HMC
human mesangial cell
- PBS
phosphate buffered saline
- PMSF
phenylmethylsulfonyl fluoride
- SDS
sodium dodecyl sulfate
- SDS-PAGE
SDS-polyacrylamide gel electrophoresis |
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Keywords: | Adriamycin extracellular matrix expansion fibronectin turnover human mesangial cells nephrotoxicity |
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