Hirudo Suppress The Expressions Of Gelatinase A & B Via Inhibiting Thrombin Receptor PAR-1 Activation And Fibrin Deposition In Murine Lupus Nephritis

Coagulation and fibrinolytic processes are involved in the pathogenesis of lupus nephritis The presence of thrombin activation and fibrin deposition in the glomeruli may be associated with more severe diseases We have found that hirudo reduce fibrin deposition and ameliorate renal lesions in primary glomerulonephntis In the present study we investigated its effect on inhibiting disease progression in MRL lpr/lpr mice MRL lpr/lpr mice were admimstered daily with hirudo (3g/kgld orally) at the age of 8 weeks After treatment for 20 weeks, a progressive reduction in positive proteinuria number/total mice (40% vs 80%, proteinuria over 300mgIdl as positive) and a prolonged survival rate (70% vs 50%) were found when compared with the control group Histological analysis of kidney tissues indicated that hirudo could inhibit the mesangial proliferation and renal sclerosis that was evident in murine lupus nephntis No significant differences were observed in the tubular and interstitial lesions Using SDS-PAGE gelatin zymography, we have identified increased expressions of both latent and activated form enzymes of MMP-2 and MMP-9 in unne and kidney extraction Immunohistochemical staining showed both MMP-2 and MMP-9 were obviously up-regulated within glomerulus in control group To determine the net gelatinase activities in murine lupus kidney, film in situ zymography was used, we found a markedly increased gelatinase activities in munne lupus nephritis, consistent with the immunohistochemical staining Hirudo suppressed 4 the expression of both latent and activated form of MMP-2 and MMP-9 We compared thrombin receptor PAR-i expression in glomerulus using immunohistochemistry and PAR-i mRNA expression by RT-PCR analysis among the groups With the development of murine lupus nephntis, we observed an increase in thrombin receptor mRNA, while thrombin receptor protein expression was strikingly down regulated, suggesting its continuous activation and degradation Hirudo inhibited PAR-i activation significantly correlated with the reduced fibrin deposition To investigate the effects of covering fibnn on the expression of MMP-2 and MMP-9 in cultured glomerular mesangial cells (GMC) RT-PCR and gelatin zymography were used to detect the changes of MMP-2 and MMP-9 in GMC at levels of mRNA expressions and protein activities The relationship between plasminogen activator/plasmin and the activation of MMP-2 and -9 were also investigated Fibnn treatment induced a dramatic expression of activated MMP-2 and MMP-9 in GMC in a dose and time dependent manner Fibrin zymography showed fibnn also induced the expression of tissue type plasminogen activator (tPA) as well as urokinase type plasminogen activator (uPA) Inhibition of the activities of plasmin by aprotimn can totally block the conversion from pro-MMP to active MMP These in vitro and in vivo results suggested that PAR-i activation and fibnn deposition might play an important role in regulating the MMP-2/-9 expressions in munne lupus nephritis, contributing to the remodeling of extracellular matrix Hirudo delays the development of glomerulonephntis and improves survival in MRL lpr/lpr mice at least by suppressing the expressions of MMP-2/-9 via inhibiting thrombin receptor PAR-i activation and fibrin deposition...