Plasmin plays a key role in the regulation of profibrogenic molecules in hepatic stellate cells.
BACKGROUND
Plasmin role in transforming growth factor-beta (TGF-beta)-responsive gene regulation remains to be elucidated. Also, plasmin action on co-repressor Ski-related novel protein N (SnoN) and differential activation of matrix metalloproteinases (MMPs) are unknown. Thus, the role of plasmin on profibrogenic molecule expression, SnoN transcriptional kinetics and gelatinase activation was investigated.


METHODS
Hepatic stellate cells (HSC) were transduced with adenovirus-mediated human urokinase plasminogen activator (Ad-huPA) (4 x 10(9) viral particles/ml). Overexpression of urokinase plasminogen activator and therefore of plasmin, was blocked by tranexamic acid (TA) in transduced HSC. Gene expression was monitored by reverse transcriptase polymerase chain reaction. HSC-free supernatants were used to evaluate MMP-2 and MMP-9 by zymography. SnoN, TGF-beta and tissue inhibitor of metalloproteinase (TIMP)-1 were analysed by Western blot. Plasmin and SnoN expression kinetics were evaluated in bile duct-ligated (BDL) rats.


RESULTS
Plasmin overexpression in Ad-huPA-transduced HSC significantly decreased gene expression of profibrogenic molecules [alpha1(I)collagen 66%, TIMP-1 59%, alpha-smooth muscle actin 90% and TGF-beta 55%]. Interestingly, both SnoN gene and protein expression increased prominently. Plasmin inhibition by TA upregulated the profibrogenic genes, which respond to TGF-beta-intracellular signalling. In contrast, SnoN mRNA and protein dropped importantly. Plasmin-activated MMP-9 and MMP-2 in HSC supernatants. Taken together, these findings indicate that MMP-9 activation is totally plasmin dependent. SnoN levels significantly decreased in cholestatic-BDL rats (82%) as compared with control animals. Interestingly, hepatic plasmin levels dropped 46% in BDL rats as compared with control.


CONCLUSIONS
Plasmin plays a key role in regulating TGF-beta-responding genes. In particular, regulation of TGF-beta-co-repressor (SnoN) is greatly affected, which suggests SnoN as a cardinal player in cholestasis-induced fibrogenesis.
Martínez-Rizo
Abril
A
Department of Molecular Biology and Genomics, CUCS, Institute for Molecular Biology in Medicine and Gene Therapy, University of Guadalajara, and OPD Hospital Civil de Guadalajara, Guadalajara, Jalisco, Mexico.
Bueno-Topete
Miriam
M
González-Cuevas
Jaime
J
Armendáriz-Borunda
Juan
J
D000818:Animals; D002460:Cell Line; D024042:Collagen Type I / Q000378:metabolism; D005341:Fibrinolysin / Q000502:physiology*; D005786:Gene Expression Regulation; D055166:Hepatic Stellate Cells / Q000187:drug effects / Q000378:metabolism* / Q000473:pathology; D006801:Humans; D008106:Liver Cirrhosis, Experimental / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D009419:Nerve Tissue Proteins / Q000235:genetics* / Q000378:metabolism; D051381:Rats; D019715:Tissue Inhibitor of Metalloproteinase-1 / Q000235:genetics / Q000378:metabolism; D019716:Tissue Inhibitor of Metalloproteinase-2 / Q000235:genetics; D014148:Tranexamic Acid / Q000494:pharmacology; D014157:Transcription Factors / Q000235:genetics* / Q000378:metabolism; D014161:Transduction, Genetic; D016212:Transforming Growth Factor beta / Q000235:genetics* / Q000378:metabolism; D014568:Urokinase-Type Plasminogen Activator / Q000235:genetics / Q000378:metabolism
D024042:Collagen Type I; D009419:Nerve Tissue Proteins; C511209:Skil_v1 protein, rat; D019715:Tissue Inhibitor of Metalloproteinase-1; D014157:Transcription Factors; D016212:Transforming Growth Factor beta; D019716:Tissue Inhibitor of Metalloproteinase-2; D014148:Tranexamic Acid; D005341:Fibrinolysin; D014568:Urokinase-Type Plasminogen Activator