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CCN5 attenuates profibrotic phenotypes of fibroblasts through the Smad6-CCN2 pathway: Potential role in epidural fibrosis.

Xu H, Liu C, Sun Z, Guo X, Zhang Y, Liu M, Li P - Int. J. Mol. Med. (2015)

Bottom Line: CCN5 upregulation also reduced an increase in collagen type I, α1 (COL1A1) and total collagen concentrations.Additionally, CCN5 over-expression decreased CCN2 expression and increased Smad6 phosphorylation.Mechanism analysis revealed that blocking Smad6 signaling significantly ameliorated the inhibitory effect of CCN5 on the CCN2 levels, accompanied by the reduction in cell proliferation and collagen production.

View Article: PubMed Central - PubMed

Affiliation: Department of Orthopaedics, The Third Affiliated Hospital (Shaanxi Provincial People's Hospital), Medical College of Xi'an Jiaotong University, Xi'an, Shaanxi 710061, P.R. China.

ABSTRACT
Epidural fibrosis is characterized by the development of dense and thick scar tissue adjacent to the dural mater and ranked as the major contributor for post-operative pain recurrence after laminectomy or discectomy. Recently, CCN5 exhibited an inhibitory effect on connective tissue growth factor (CTGF)/CCN2 (a critical regulator for fibrotic disease)‑mediated fibrogenesis. However, its function in epidural fibrosis and the underlying mechanisms involved remain to be determined. In this study, an obvious downregulation of CCN5 was observed in scar tissues from laminectomized rats, concomitant with a marked upregulation of CCN2, suggesting a potential negative regulatory role of CCN5 in fibrogenesis. Furthermore, CCN5 overexpression notably mitigated transforming growth factor‑β1-enhanced fibroblast viability and proliferation. Of note, CCN5 upregulation inhibited the switch of fibroblasts into myofibroblasts as its overexpression abrogated the expression of the myofibroblast marker, α-smooth muscle actin (α-SMA). CCN5 upregulation also reduced an increase in collagen type I, α1 (COL1A1) and total collagen concentrations. Additionally, CCN5 over-expression decreased CCN2 expression and increased Smad6 phosphorylation. Mechanism analysis revealed that blocking Smad6 signaling significantly ameliorated the inhibitory effect of CCN5 on the CCN2 levels, accompanied by the reduction in cell proliferation and collagen production. These results confirm that CCN5 exerts an anti-fibrotic function by regulating the Smad6-CCN2 pathway, thereby indicating a potential approach for ameliorating epidural fibrosis after laminectomy.

No MeSH data available.


Related in: MedlinePlus

CCN5 transfection enhances CCN5 expression in primary fibroblasts. Primary fibroblasts were obtained from the tail skin of rats and cultured in DMEM medium. Lentivirus plasmid pWPT-GFP was introduced to construct the recombinant pWPT-CCN5 plasmids, following packaging with vectors of pCMV-VSV-G and pCMV-dR8.91. The corresponding transfection effect of CCN5 mRNA (A) and protein levels (B) was assessed individually with RT-PCR and western blotting. *P<0.05.
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f2-ijmm-36-01-0123: CCN5 transfection enhances CCN5 expression in primary fibroblasts. Primary fibroblasts were obtained from the tail skin of rats and cultured in DMEM medium. Lentivirus plasmid pWPT-GFP was introduced to construct the recombinant pWPT-CCN5 plasmids, following packaging with vectors of pCMV-VSV-G and pCMV-dR8.91. The corresponding transfection effect of CCN5 mRNA (A) and protein levels (B) was assessed individually with RT-PCR and western blotting. *P<0.05.

Mentions: Fibroblasts are essential for epidural fibrosis due to their function in fibrotic matrix formation during scar formation (3,4). To investigate the effect of CCN5 on scar formation, the recombinant lentiviral vector-carrying CCN5 (LV-CCN5) was constructed and transfected into the isolated primary fibroblasts. As shown in Fig. 2A, an ~4.1-fold increase in CCN5 mRNA levels was observed when the cells were transfected with LV-CCN5. Furthermore, western blot analysis confirmed the obvious upregulation of CCN5 protein following LV-CCN5 transfection, compared with the control and vector-treated groups (Fig. 2B), indicating that a stable overexpression system of CCN5 had been successfully constructed.


CCN5 attenuates profibrotic phenotypes of fibroblasts through the Smad6-CCN2 pathway: Potential role in epidural fibrosis.

Xu H, Liu C, Sun Z, Guo X, Zhang Y, Liu M, Li P - Int. J. Mol. Med. (2015)

CCN5 transfection enhances CCN5 expression in primary fibroblasts. Primary fibroblasts were obtained from the tail skin of rats and cultured in DMEM medium. Lentivirus plasmid pWPT-GFP was introduced to construct the recombinant pWPT-CCN5 plasmids, following packaging with vectors of pCMV-VSV-G and pCMV-dR8.91. The corresponding transfection effect of CCN5 mRNA (A) and protein levels (B) was assessed individually with RT-PCR and western blotting. *P<0.05.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4494601&req=5

f2-ijmm-36-01-0123: CCN5 transfection enhances CCN5 expression in primary fibroblasts. Primary fibroblasts were obtained from the tail skin of rats and cultured in DMEM medium. Lentivirus plasmid pWPT-GFP was introduced to construct the recombinant pWPT-CCN5 plasmids, following packaging with vectors of pCMV-VSV-G and pCMV-dR8.91. The corresponding transfection effect of CCN5 mRNA (A) and protein levels (B) was assessed individually with RT-PCR and western blotting. *P<0.05.
Mentions: Fibroblasts are essential for epidural fibrosis due to their function in fibrotic matrix formation during scar formation (3,4). To investigate the effect of CCN5 on scar formation, the recombinant lentiviral vector-carrying CCN5 (LV-CCN5) was constructed and transfected into the isolated primary fibroblasts. As shown in Fig. 2A, an ~4.1-fold increase in CCN5 mRNA levels was observed when the cells were transfected with LV-CCN5. Furthermore, western blot analysis confirmed the obvious upregulation of CCN5 protein following LV-CCN5 transfection, compared with the control and vector-treated groups (Fig. 2B), indicating that a stable overexpression system of CCN5 had been successfully constructed.

Bottom Line: CCN5 upregulation also reduced an increase in collagen type I, α1 (COL1A1) and total collagen concentrations.Additionally, CCN5 over-expression decreased CCN2 expression and increased Smad6 phosphorylation.Mechanism analysis revealed that blocking Smad6 signaling significantly ameliorated the inhibitory effect of CCN5 on the CCN2 levels, accompanied by the reduction in cell proliferation and collagen production.

View Article: PubMed Central - PubMed

Affiliation: Department of Orthopaedics, The Third Affiliated Hospital (Shaanxi Provincial People's Hospital), Medical College of Xi'an Jiaotong University, Xi'an, Shaanxi 710061, P.R. China.

ABSTRACT
Epidural fibrosis is characterized by the development of dense and thick scar tissue adjacent to the dural mater and ranked as the major contributor for post-operative pain recurrence after laminectomy or discectomy. Recently, CCN5 exhibited an inhibitory effect on connective tissue growth factor (CTGF)/CCN2 (a critical regulator for fibrotic disease)‑mediated fibrogenesis. However, its function in epidural fibrosis and the underlying mechanisms involved remain to be determined. In this study, an obvious downregulation of CCN5 was observed in scar tissues from laminectomized rats, concomitant with a marked upregulation of CCN2, suggesting a potential negative regulatory role of CCN5 in fibrogenesis. Furthermore, CCN5 overexpression notably mitigated transforming growth factor‑β1-enhanced fibroblast viability and proliferation. Of note, CCN5 upregulation inhibited the switch of fibroblasts into myofibroblasts as its overexpression abrogated the expression of the myofibroblast marker, α-smooth muscle actin (α-SMA). CCN5 upregulation also reduced an increase in collagen type I, α1 (COL1A1) and total collagen concentrations. Additionally, CCN5 over-expression decreased CCN2 expression and increased Smad6 phosphorylation. Mechanism analysis revealed that blocking Smad6 signaling significantly ameliorated the inhibitory effect of CCN5 on the CCN2 levels, accompanied by the reduction in cell proliferation and collagen production. These results confirm that CCN5 exerts an anti-fibrotic function by regulating the Smad6-CCN2 pathway, thereby indicating a potential approach for ameliorating epidural fibrosis after laminectomy.

No MeSH data available.


Related in: MedlinePlus