Characterization of a plasminogen activator and its inhibitor in human mesangial cells.
In the course of some pathological and experimental nephropathies, intraglomerular fibrin deposits develop, possibly as a consequence of inefficient fibrinolysis. In vitro human glomeruli exhibit fibrinolytic activity due to the synthesis of plasminogen activators (PAs) such as, tissue-type PA (t-PA) and urokinase-type PA (u-PA). Immunofluorescence studies have previously shown that t-PA is localized in the capillary tufts and u-PA in the visceral epithelial cells. We have now investigated the fibrinolytic activity of cultured human mesangial cells. Inhibitory activity towards u-PA or t-PA but not plasmin was found in both conditioned medium and cellular extracts. Analysis of the conditioned medium by zymography revealed a single band of PA-activity (Mr: 110 to 120 kDa). Immunoneutralization with anti-t-PA and anti-plasminogen activator inhibitor (PAI-1) IgG but not anti-u-PA or anti-PAI-2 removed this band. Reverse fibrin autography demonstrated the presence of PAI-1 in both cellular extracts and in conditioned medium. Western Blot analysis showed that two bands (50 kD and 120 kD) were recognized by the anti-PAI-1 antibody. By ELISA t-PA and PAI-1 antigens were found to increase progressively with time in the culture medium but not in cellular extracts. Both t-PA and PAI-1, but not u-PA and PAI-2, were also detected by immunofluorescence studies. Thus human glomerular mesangial cells synthesize and secrete t-PA and PAI-1 in vitro. PAI-1 is produced in excess, therefore t-PA is only found in the form of a complex with PAI-1.
Lacave
R
R
INSERM U 64, Hôpital Tenon, Paris, France.
Rondeau
E
E
Ochi
S
S
Delarue
F
F
Schleuning
W D
WD
Sraer
J D
JD
D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D005455:Fluorescent Antibody Technique; D005920:Glomerular Mesangium / Q000032:analysis*; D006023:Glycoproteins / Q000302:isolation & purification*; D006801:Humans; D015151:Immunoblotting; D066298:In Vitro Techniques; D010960:Plasminogen Activators / Q000037:antagonists & inhibitors*; D015849:Plasminogen Inactivators; D010959:Tissue Plasminogen Activator / Q000302:isolation & purification*
D006023:Glycoproteins; D015849:Plasminogen Inactivators; D010960:Plasminogen Activators; D010959:Tissue Plasminogen Activator