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Activated Wnt signaling induces myofibroblast differentiation of mesenchymal stem cells, contributing to pulmonary fibrosis.

Sun Z, Wang C, Shi C, Sun F, Xu X, Qian W, Nie S, Han X - Int. J. Mol. Med. (2014)

Bottom Line: Histological examination data demonstrated that the engraftment of MSCs did not attenuate lung injury and pulmonary fibrosis.The in vitro study results demonstrated that activation of the Wnt/β-catenin signaling stimulated MSCs to express myofibroblast markers; however, this process was attenuated by Wnt antagonist DKK1.Therefore, the results demonstrated that the aberrant activation of Wnt signaling induces the myofibroblast differentiation of engrafted MSCs, thus contributing to pulmonary fibrosis following lung injury.

View Article: PubMed Central - PubMed

Affiliation: Immunology and Reproductive Biology Laboratory, Medical School of Nanjing University, Nanjing, Jiangsu 210093, P.R. China.

ABSTRACT
Acute lung injury may lead to fibrogenesis. However, no treatment is currently available. This study was conducted to determine the effects of bone marrow-derived mesenchymal stem cells (MSCs) in a model of HCl-induced acute lung injury in Sprague-Dawley (SD) rats. Stromal cell-derived factor (SDF)-1 and its receptor CXC chemokine receptor (CXCR)4 have been shown to participate in mobilizing MSCs. Adenovirus carrying the CXCR4 gene was used to transfect MSCs in order to increase the engraftment numbers of MSCs at injured sites. Histological examination data demonstrated that the engraftment of MSCs did not attenuate lung injury and pulmonary fibrosis. The results showed that engraftment of MSCs almost differentiated into myofibroblasts, but rarely differentiated into lung epithelial cells. Additionally, it was demonstrated that activated canonical Wnt/β-catenin signaling in injured lung tissue regulated the myofibroblast differentiation of MSCs in vivo. The in vitro study results demonstrated that activation of the Wnt/β-catenin signaling stimulated MSCs to express myofibroblast markers; however, this process was attenuated by Wnt antagonist DKK1. Therefore, the results demonstrated that the aberrant activation of Wnt signaling induces the myofibroblast differentiation of engrafted MSCs, thus contributing to pulmonary fibrosis following lung injury.

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Wnt signaling regulates the differentiation of mesenchymal stem cells (MSCs). (A) The Wnt signaling pathway was highly activated after lung injury. The expression of Wnt signaling components β-catenin and MMP-2 were increased in each group. The activated Wnt signaling may determine the differentiation of MSCs in vivo. (B and C) MSCs were cultured until they reached confluence and then treated with Wnt3α and DKK1 for 14 days. Activation of Wnt signaling induces MSCs to differentiate into myofibroblasts which was inhibited by treatment with DKK1. (B) Immunofluorescent analysis of β-catenin, vimentin and α-SMA expression in MSCs. Scale bar, 50 μm. (C) Expression of β-catenin, vimentin and α-SMA was evaluated in whole cell lysates by western blotting. β-actin was used as the control.
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f7-ijmm-33-05-1097: Wnt signaling regulates the differentiation of mesenchymal stem cells (MSCs). (A) The Wnt signaling pathway was highly activated after lung injury. The expression of Wnt signaling components β-catenin and MMP-2 were increased in each group. The activated Wnt signaling may determine the differentiation of MSCs in vivo. (B and C) MSCs were cultured until they reached confluence and then treated with Wnt3α and DKK1 for 14 days. Activation of Wnt signaling induces MSCs to differentiate into myofibroblasts which was inhibited by treatment with DKK1. (B) Immunofluorescent analysis of β-catenin, vimentin and α-SMA expression in MSCs. Scale bar, 50 μm. (C) Expression of β-catenin, vimentin and α-SMA was evaluated in whole cell lysates by western blotting. β-actin was used as the control.

Mentions: Following transplantation, MSCs did not attenuate lung injury and pulmonary fibrosis as we expected, however, MSCs were involved in pulmonary fibrogenesis. We suspect that certain factors or signaling pathway affected the differentiation process of MSCs. It is necessary to explore the regulation mechanisms of MSC differentiation in vivo. In a previous study, we demonstrated that Wnt/β-catenin signaling regulates the MSC differentiation in vitro (24). In a co-culture system, we demonstrated that activation of Wnt signaling prevented the epithelial differentiation of MSCs, but inhibition of Wnt signaling promoted MSCs to differentiate into epithelial-like cells when MSCs were co-cultured with epithelial cells. Wnt/β-catenin signaling is also involved in regulating lung tissue remodeling, fibrosis or destruction and lung diseases. Therefore, Wnt/β-catenin signaling may regulate the differentiation of MSCs in lung tissue. In this study, we detected the expression of β-catenin and MMP-2, which are the essential components of canonical Wnt signaling. Western blot analysis revealed that the protein expression of β-catenin and MMP-2 was increased significantly in the ALI and ALI+MSC-CXCR4/GFP groups compared with the control group, which demonstrated that Wnt signaling is highly activated (Fig. 7A). It indicated that the abnormal activation of Wnt/β-catenin signaling may induce engraftment of MSCs to differentiate into myofibroblasts or fibroblasts and prevent the epithelial differentiation of MSCs, resulting in tissue repair failure and severe pulmonary fibrosis. The functional role of canonical Wnt signaling in MSC differentiation in vitro was investigated. Immunofluorescent staining and western blotting results revealed that treatment of MSCs for 14 days with Wnt3α (100 ng/ml) resulted in an increase in the protein expression of β-catenin, fibroblast marker vimentin and myofibroblast marker α-SMA (Fig. 7B and C). Conversely, DKK1 decreased the expression levels of β-catenin, vimentin and α-SMA in MSCs. The activation of Wnt/β-catenin signaling therefore induced by Wnt3α ameliorated the possibility of MSCs to differentiate into myofibroblasts. By contrast, the inhibition of Wnt/β-catenin signaling caused by DKK1 prevented myofibroblast differentiation of MSCs. It indicated that inhibition of the Wnt/β-catenin signaling pathway following MSC transplantation suggests a positive unique therapeutic approach for lung injury or pulmonary fibrosis.


Activated Wnt signaling induces myofibroblast differentiation of mesenchymal stem cells, contributing to pulmonary fibrosis.

Sun Z, Wang C, Shi C, Sun F, Xu X, Qian W, Nie S, Han X - Int. J. Mol. Med. (2014)

Wnt signaling regulates the differentiation of mesenchymal stem cells (MSCs). (A) The Wnt signaling pathway was highly activated after lung injury. The expression of Wnt signaling components β-catenin and MMP-2 were increased in each group. The activated Wnt signaling may determine the differentiation of MSCs in vivo. (B and C) MSCs were cultured until they reached confluence and then treated with Wnt3α and DKK1 for 14 days. Activation of Wnt signaling induces MSCs to differentiate into myofibroblasts which was inhibited by treatment with DKK1. (B) Immunofluorescent analysis of β-catenin, vimentin and α-SMA expression in MSCs. Scale bar, 50 μm. (C) Expression of β-catenin, vimentin and α-SMA was evaluated in whole cell lysates by western blotting. β-actin was used as the control.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4020487&req=5

f7-ijmm-33-05-1097: Wnt signaling regulates the differentiation of mesenchymal stem cells (MSCs). (A) The Wnt signaling pathway was highly activated after lung injury. The expression of Wnt signaling components β-catenin and MMP-2 were increased in each group. The activated Wnt signaling may determine the differentiation of MSCs in vivo. (B and C) MSCs were cultured until they reached confluence and then treated with Wnt3α and DKK1 for 14 days. Activation of Wnt signaling induces MSCs to differentiate into myofibroblasts which was inhibited by treatment with DKK1. (B) Immunofluorescent analysis of β-catenin, vimentin and α-SMA expression in MSCs. Scale bar, 50 μm. (C) Expression of β-catenin, vimentin and α-SMA was evaluated in whole cell lysates by western blotting. β-actin was used as the control.
Mentions: Following transplantation, MSCs did not attenuate lung injury and pulmonary fibrosis as we expected, however, MSCs were involved in pulmonary fibrogenesis. We suspect that certain factors or signaling pathway affected the differentiation process of MSCs. It is necessary to explore the regulation mechanisms of MSC differentiation in vivo. In a previous study, we demonstrated that Wnt/β-catenin signaling regulates the MSC differentiation in vitro (24). In a co-culture system, we demonstrated that activation of Wnt signaling prevented the epithelial differentiation of MSCs, but inhibition of Wnt signaling promoted MSCs to differentiate into epithelial-like cells when MSCs were co-cultured with epithelial cells. Wnt/β-catenin signaling is also involved in regulating lung tissue remodeling, fibrosis or destruction and lung diseases. Therefore, Wnt/β-catenin signaling may regulate the differentiation of MSCs in lung tissue. In this study, we detected the expression of β-catenin and MMP-2, which are the essential components of canonical Wnt signaling. Western blot analysis revealed that the protein expression of β-catenin and MMP-2 was increased significantly in the ALI and ALI+MSC-CXCR4/GFP groups compared with the control group, which demonstrated that Wnt signaling is highly activated (Fig. 7A). It indicated that the abnormal activation of Wnt/β-catenin signaling may induce engraftment of MSCs to differentiate into myofibroblasts or fibroblasts and prevent the epithelial differentiation of MSCs, resulting in tissue repair failure and severe pulmonary fibrosis. The functional role of canonical Wnt signaling in MSC differentiation in vitro was investigated. Immunofluorescent staining and western blotting results revealed that treatment of MSCs for 14 days with Wnt3α (100 ng/ml) resulted in an increase in the protein expression of β-catenin, fibroblast marker vimentin and myofibroblast marker α-SMA (Fig. 7B and C). Conversely, DKK1 decreased the expression levels of β-catenin, vimentin and α-SMA in MSCs. The activation of Wnt/β-catenin signaling therefore induced by Wnt3α ameliorated the possibility of MSCs to differentiate into myofibroblasts. By contrast, the inhibition of Wnt/β-catenin signaling caused by DKK1 prevented myofibroblast differentiation of MSCs. It indicated that inhibition of the Wnt/β-catenin signaling pathway following MSC transplantation suggests a positive unique therapeutic approach for lung injury or pulmonary fibrosis.

Bottom Line: Histological examination data demonstrated that the engraftment of MSCs did not attenuate lung injury and pulmonary fibrosis.The in vitro study results demonstrated that activation of the Wnt/β-catenin signaling stimulated MSCs to express myofibroblast markers; however, this process was attenuated by Wnt antagonist DKK1.Therefore, the results demonstrated that the aberrant activation of Wnt signaling induces the myofibroblast differentiation of engrafted MSCs, thus contributing to pulmonary fibrosis following lung injury.

View Article: PubMed Central - PubMed

Affiliation: Immunology and Reproductive Biology Laboratory, Medical School of Nanjing University, Nanjing, Jiangsu 210093, P.R. China.

ABSTRACT
Acute lung injury may lead to fibrogenesis. However, no treatment is currently available. This study was conducted to determine the effects of bone marrow-derived mesenchymal stem cells (MSCs) in a model of HCl-induced acute lung injury in Sprague-Dawley (SD) rats. Stromal cell-derived factor (SDF)-1 and its receptor CXC chemokine receptor (CXCR)4 have been shown to participate in mobilizing MSCs. Adenovirus carrying the CXCR4 gene was used to transfect MSCs in order to increase the engraftment numbers of MSCs at injured sites. Histological examination data demonstrated that the engraftment of MSCs did not attenuate lung injury and pulmonary fibrosis. The results showed that engraftment of MSCs almost differentiated into myofibroblasts, but rarely differentiated into lung epithelial cells. Additionally, it was demonstrated that activated canonical Wnt/β-catenin signaling in injured lung tissue regulated the myofibroblast differentiation of MSCs in vivo. The in vitro study results demonstrated that activation of the Wnt/β-catenin signaling stimulated MSCs to express myofibroblast markers; however, this process was attenuated by Wnt antagonist DKK1. Therefore, the results demonstrated that the aberrant activation of Wnt signaling induces the myofibroblast differentiation of engrafted MSCs, thus contributing to pulmonary fibrosis following lung injury.

Show MeSH
Related in: MedlinePlus