F­actin cytoskeleton reorganization is associated with hepatic stellate cell activation.

The activation of hepatic stellate cells (HSCs) is involved in the development of hepatic fibrosis. Previous studies have indicated that the acquisition of certain properties by activated HSCs is highly dependent on the reorganization of the actin cytoskeleton. However, direct evidence showing that the reorganization of the actin cytoskeleton is responsible for HSC activation is lacking. The aim of the present study was to investigate the role of cytoskeletal reorganization during HSC activation and to clarify the underlying mechanism. HSC-T6 cells were treated either with the F-actin stabilizer jasplakinolide (Jas) or the depolymerizer cytochalasin D (Cyto D). The actin cytoskeleton was evaluated via assessment of stress fiber formation. Furthermore, the activation properties of HSCs, including proliferation, adhesion, migration and the expression of α-smooth muscle actin (α-SMA) and collagen 1, were investigated in vitro. The results showed that Jas and Cyto D affected the actin distribution in HSC-T6 cells. Treatment with Jas resulted in thick actin bundles and a patchy appearance in the cytoplasm in HSC-T6 cells. In parallel, polymerization of actin microfilaments induced by Jas upregulated the expression of α-SMA and collagen 1, and also enhanced the migration and adhesion properties of HSC-T6 cells. Furthermore, the activation of HSC-T6 cells induced by the reorganization of the actin cytoskeleton was associated with the p38 mitogen-activated protein kinase (p38 MAPK) pathway. In conclusion, the present study suggests that the reorganization of the F-actin cytoskeleton is associated with HSC activation and that the p38 MAPK pathway is involved in this process. The inhibition of F-actin reorganization may thus be a potential key factor or molecular target for the control of liver fibrosis or cirrhosis.

[Effects of endothelial progenitor cell-conditioned medium on the proliferation, adhesion and migration of vascular smooth muscle cells].

OBJECTIVE: To investigate the effects of endothelia progenitor cells conditioned medium (EPC-CM) on the migration, adhesion and proliferation of vascular smooth muscle cells (VSMCs). METHODS: Mononuclear cells were isolated from rat bone marrow by density gradient centrifugation,plated on dishes precoated with 5% fibronectin, and then cultured with complete M199 medium (including 15% fetal calf serum, 10 microg/L VEGF and 5 microg/L bFGF). EPC-CM was collected and used to incubate VSMCs isolated from rat arteriae aorta. After 24 h, VSMCs proliferation, adhesion and migration were assayed with CCK-8, adhesion test and modified Boyden chamber assay, respectively. RESULTS: The proliferation, adhesion and migration of VSMCs were obviously decreased when the cells were cultured with EPC-CM. CONCLUSION: EPC-CM could inhibit VSMC functions, which would be one of the mechanisms against atherosclerosis by EPCs.

[Effects of extracellular matrix on biological characteristics of late endothelial progenitor cells].

The present study was designed to investigate the effects of various extracellular matrix (ECM) proteins on the biological characteristics of late endothelial progenitor cells (EPCs). Density gradient centrifugation-isolated rat bone marrow mononuclear cells were cultured in complete M199 medium, which contained 15% fetal calf serum, 10 µg/L vascular endothelial growth factor (VEGF) and 5 µg/L basic fibroblast growth factor (bFGF). EPCs were plated on substrates containing fibronectin (Fn), laminin (Ln) or rat tail tendon collagen (Col), and the corresponding cells were defined as Fn, Ln and Col groups. The 3rd generation EPCs, namely late EPCs, were harvested. The proliferation, adhesion, migration and the ability of forming tubes were assayed using CCK-8, adhesion test, wound healing assay and Matrigel, respectively. The mRNA expressions of endothelial cell differentiation markers, vWF and CD31, were analyzed by real time RT-PCR. The apoptosis was assayed by flow cytometry (FCM). The results showed that cell proliferation ability of Fn and Col groups were higher than that of Ln group; Fn group showed increased adhesion compared to Col and Ln groups (P < 0.01); The migration ability of Fn and Col groups were higher than that of Ln group. Moreover, Fn group showed increased tube formation abilities compared to Col and Ln groups (P < 0.05). Although 24-hour free-serum-induced apoptosis in Ln group was the highest, there was no difference of auto-apoptosis among the three groups. Furthermore, the mRNA expressions of vWF and CD31 exhibited no difference among the three groups. These results suggest the ECM affects the biological functions of late EPCs, which would have a high probability of providing new directions that lead to the development of artificial heart and blood vessels.