Metalloproteinase generation by human glomerular epithelial cells.
In the present study cultured human glomerular epithelial cells (HGEC) were used to examine the potential role for these cells in the turnover of the glomerular basement membrane (GBM) through the secretion of matrix metalloproteinases. The cells were shown by substrate gel electrophoresis to secrete gelatinase activity of molecular weights 72 kDa and 92 kDa. The gelatinolytic activity was inhibited by EDTA (10 mM), and by both TIMP-I and TIMP-II, but was not inhibited by PMSF (2.5 mM), indicating that the enzymes belonged to the metalloproteinase family. The identity of the enzymes was confirmed by the use of specific antisera to gelatinase A and gelatinase B. In addition, reverse transcription polymerase chain reaction (RT PCR) amplification of HGEC mRNA using specific primers to the two enzymes yielded single bands of amplified DNA and served to verify the identity of the enzymes. In supplementary experiments using both specific antiserum and PCR primers it was shown that HGEC express message and secrete both the specific metalloproteinase inhibitors TIMP-I and TIMP-II. These results indicate that the synthesis and secretion of degradative enzymes and their controlling inhibitors by HGEC have the potential to be involved in the turnover of the extracellular matrix.
Knowlden
J
J
Institute of Nephrology, University of Wales College of Medicine, Cardiff Royal Infirmary, United Kingdom.
Martin
J
J
Davies
M
M
Williams
J D
JD
D001483:Base Sequence; D002478:Cells, Cultured; D004847:Epithelial Cells; D004848:Epithelium / Q000378:metabolism; D005455:Fluorescent Antibody Technique; D005780:Gelatin / Q000378:metabolism; D018093:Gelatinases / Q000378:metabolism; D006023:Glycoproteins / Q000378:metabolism; D006801:Humans; D007678:Kidney Glomerulus / Q000166:cytology / Q000378:metabolism*; D008666:Metalloendopeptidases / Q000037:antagonists & inhibitors / Q000096:biosynthesis*; D015335:Molecular Probes / Q000235:genetics; D008969:Molecular Sequence Data; D011480:Protease Inhibitors / Q000378:metabolism; D011506:Proteins / Q000378:metabolism; D019716:Tissue Inhibitor of Metalloproteinase-2; D019714:Tissue Inhibitor of Metalloproteinases
D006023:Glycoproteins; D015335:Molecular Probes; D011480:Protease Inhibitors; D011506:Proteins; D019714:Tissue Inhibitor of Metalloproteinases; D019716:Tissue Inhibitor of Metalloproteinase-2; D005780:Gelatin; D018093:Gelatinases; D008666:Metalloendopeptidases