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1.
Article de Chinois | WPRIM | ID: wpr-252706

RÉSUMÉ

<p><b>AIM</b>To observe the effect of FAK-related non-Kinase (FRNK) plasmid on hepatic stellate cell (HSC) proliferation stimulated by fibronectin (FN).</p><p><b>METHODS</b>FRNK plasmid was transfected into HSC with transient liposomal transfection. The proteins of FRNK, FAK and p-FAK(Tyr397) were assayed by Western blotting analysis. The proliferation of HSC was evaluated by improved MTT assay, and cell cycle pattern was determined by flow cytometry (FCM).</p><p><b>RESULTS</b>(1) The expression of FRNK protein increased after FRNK transfected HSC, and it was at 48 h that the expression of FRNK protein was the highest (P < 0.01). The protein level of FAK was no significant difference between before FRNK plasmid transfection and after transfection (P > 0.05). The expression of p-FAK(Tyr397) protein was down-regulated after FRNK had been transfected in HSC, (P < 0.01). (2) The HSC proliferation inhibition rates at 12 h, 24 h and 48 h after FRNK transfection were 20.07%, 26.16%, 29.77%, respectively (P < 0.01). (3) Compared with the non-FRNK plasmid group, the FRNK-transfected HSCs almost arrested in G0/G1 phase (71.4 +/- 2.81 vs 48.9 +/- 1.66, P < 0.01).</p><p><b>CONCLUSION</b>After FRNK were transfected successfully in HSCs using lipofectamine, the phosphorylation of FAK was inhibited. The HSC proliferation was restrained in a time-dependent manner and the HSC was arrested in G0/G1 phase.</p>


Sujet(s)
Humains , Lignée cellulaire , Prolifération cellulaire , Fibronectines , Cellules étoilées du foie , Biologie cellulaire , Phosphorylation , Plasmides , Génétique , Protein-tyrosine kinases , Génétique , Transfection
2.
Zhonghua ganzangbing zazhi ; Zhonghua ganzangbing zazhi;(12): 849-853, 2008.
Article de Chinois | WPRIM | ID: wpr-250102

RÉSUMÉ

<p><b>OBJECTIVES</b>To investigate the effects of FAK-related non-kinase (FRNK) on the apoptosis of hepatic stellate cells (HSC) in vitro and on the extracellular signal-regulated kinase (ERK) signal transduction pathway.</p><p><b>METHODS</b>HSC were stimulated by fibronectin (FN), and then they were transfected with FAK-related non-kinase (FRNK) plasmids mediated by cationic liposome. The apoptosis of FRNK-induced HSC was examined by annexin-V/propidium iodide double-labeled flow cytometry (FCM), gel electrophoresis and transmission electron microscopy. Levels of FRNK, FAK, p-FAK (Tyr397), ERK1 and p-ERK in HSC were assayed by Western blot on the protein level, and by RT-PCR on the mRNA level.</p><p><b>RESULTS</b>The expression of FRNK was enhanced after FRNK plasmids were transiently transfected into the HSC. The apoptotic rate of the HSC exposed to FRNK plasmids for 48 h was higher than that in the non-FRNK plasmid group (25.37%+/-1.92% vs 9.28%+/-1.05%, P less than 0.01), and was accompanied by a significantly higher activity of caspase-3 both in the protein and in the mRNA levels [(264.17+/-12.60 vs 185.82+/-9.69), P less than 0.01; (4.19+/-0.48 vs 1.07+/-0.27), P less than 0.01]. After exposure of HSC to FRNK plasmids, compared with the non-FRNK plasmid group, the expressions of p-FAK, ERK1 and p-ERK in protein and mRNA levels were lower; on the contrary, compared with the control group, the expressions of p-FAK, ERK1 and p-ERK in the FN group were higher.</p><p><b>CONCLUSION</b>The expression of FRNK was enhanced and the phosphorylation of FAK was inhibited after FRNK was transiently transfected into HSC in vitro. FRNK induces apoptosis of HSC. FAK-ERK signal transduction pathway perhaps is involved in the process.</p>


Sujet(s)
Humains , Apoptose , Lignée cellulaire , Prolifération cellulaire , Extracellular Signal-Regulated MAP Kinases , Métabolisme , Cellules étoilées du foie , Métabolisme , Anatomopathologie , Protein-tyrosine kinases , Génétique , Métabolisme , ARN messager , Génétique , Transduction du signal
3.
Zhonghua ganzangbing zazhi ; Zhonghua ganzangbing zazhi;(12): 757-761, 2008.
Article de Chinois | WPRIM | ID: wpr-279683

RÉSUMÉ

<p><b>OBJECTIVES</b>To investigate the effects of FAK-related non-kinase (FRNK) on expressions of type I collagen and matrix metalloproteinase-2 (MMP-2) mRNA and tissue inhibitor of metalloproteinase-2 (TIMP-2) mRNA in rat hepatic stellate cells (HSC).</p><p><b>METHODS</b>Using in vitro cell culture technique, FRNK plasmids were transfected into HSC mediated by cationic liposome. Type I collagen synthesis capability in HSC was examined by 3H-Pro incorporation assay. The levels of FRNK in HSC were assayed by Western blot, and the expressions of MMP-2 and TIMP-2 were assayed by RT-PCR on mRNA levels.</p><p><b>RESULTS</b>The exposure of HSC to FRNK caused the expression of FRNK protein to be up-regulated, and the FRNK protein contents reached the highest point at 48 h after the transfection, P less than 0.05. The expressions of MMP-2 mRNA were up-regulated by FRNK; the expressions of TIMP-2 mRNA were down-regulated by FRNK; the ratios of MMP-2 mRNA/TIMP-2 mRNA were enhanced by FRNK.</p><p><b>CONCLUSION</b>After FRNK was transfected, the capability of type I collagen synthesis in HSC was inhibited, which may be related to the up-regulation of MMP-2 mRNA/TIMP-2 mRNA.</p>


Sujet(s)
Animaux , Rats , Cellules cultivées , Collagène de type I , Métabolisme , Cellules étoilées du foie , Biologie cellulaire , Métabolisme , Matrix metalloproteinase 2 , Métabolisme , Plasmides , Protein-tyrosine kinases , Génétique , ARN messager , Génétique , Inhibiteur tissulaire de métalloprotéinase-2 , Métabolisme , Transfection
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