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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

The function of CCN5 on fibroblast cell proliferation and viability. To assess the effect of CCN5 on cell proliferation and viability in response to the transforming growth factor-β (TGF-β) (10 ng/ml) for 12 h, cells were seeded in 24-well plates and treated with LV-CCN5, or vector transfection. Then, 20 µl MTT reagent was added for 6 h to determine the roles of CCN5 overexpression in cell viability (A). Furthermore, the corresponding effects of CCN5 on cell proliferation were analyzed by addition of [3H]-thymidine (B). *P<0.05.
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f3-ijmm-36-01-0123: The function of CCN5 on fibroblast cell proliferation and viability. To assess the effect of CCN5 on cell proliferation and viability in response to the transforming growth factor-β (TGF-β) (10 ng/ml) for 12 h, cells were seeded in 24-well plates and treated with LV-CCN5, or vector transfection. Then, 20 µl MTT reagent was added for 6 h to determine the roles of CCN5 overexpression in cell viability (A). Furthermore, the corresponding effects of CCN5 on cell proliferation were analyzed by addition of [3H]-thymidine (B). *P<0.05.

Mentions: TGF-β1 is known to be critical for scar-triggered epidural fibrosis following laminectomy due to its important role in the development of fibrosis (16). Based on the stable expression of CCN5 in fibroblasts, cell viability and proliferation in response to TGF-β1 was investigated. An MTT assay was used to assess the effect of CCN5 on cell viability. As shown in Fig. 3A, TGF-β1 stimulation exhibited a 2.5-fold increase in cell viability. However, this increase was markedly attenuated when CCN5 was overexpressed in fibroblasts. Further [3H]-TdR analysis confirmed that TGF-β1 treatment induced cell proliferation and resulted in an ~2.0-fold increase in cell numbers (Fig. 3B). Elevated CCN5 expression decreased TGF-β1-induced cell proliferation. These results suggested that CCN5 exhibits a negative inhibitory function during the TGF-β1-induced process of fibrosis by inhibiting fibroblast cell viability and proliferation.


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)

The function of CCN5 on fibroblast cell proliferation and viability. To assess the effect of CCN5 on cell proliferation and viability in response to the transforming growth factor-β (TGF-β) (10 ng/ml) for 12 h, cells were seeded in 24-well plates and treated with LV-CCN5, or vector transfection. Then, 20 µl MTT reagent was added for 6 h to determine the roles of CCN5 overexpression in cell viability (A). Furthermore, the corresponding effects of CCN5 on cell proliferation were analyzed by addition of [3H]-thymidine (B). *P<0.05.
© Copyright Policy - open-access
Related In: Results  -  Collection

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

f3-ijmm-36-01-0123: The function of CCN5 on fibroblast cell proliferation and viability. To assess the effect of CCN5 on cell proliferation and viability in response to the transforming growth factor-β (TGF-β) (10 ng/ml) for 12 h, cells were seeded in 24-well plates and treated with LV-CCN5, or vector transfection. Then, 20 µl MTT reagent was added for 6 h to determine the roles of CCN5 overexpression in cell viability (A). Furthermore, the corresponding effects of CCN5 on cell proliferation were analyzed by addition of [3H]-thymidine (B). *P<0.05.
Mentions: TGF-β1 is known to be critical for scar-triggered epidural fibrosis following laminectomy due to its important role in the development of fibrosis (16). Based on the stable expression of CCN5 in fibroblasts, cell viability and proliferation in response to TGF-β1 was investigated. An MTT assay was used to assess the effect of CCN5 on cell viability. As shown in Fig. 3A, TGF-β1 stimulation exhibited a 2.5-fold increase in cell viability. However, this increase was markedly attenuated when CCN5 was overexpressed in fibroblasts. Further [3H]-TdR analysis confirmed that TGF-β1 treatment induced cell proliferation and resulted in an ~2.0-fold increase in cell numbers (Fig. 3B). Elevated CCN5 expression decreased TGF-β1-induced cell proliferation. These results suggested that CCN5 exhibits a negative inhibitory function during the TGF-β1-induced process of fibrosis by inhibiting fibroblast cell viability and proliferation.

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