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Overexpression of SerpinE2/protease nexin-1 Contribute to Pathological Cardiac Fibrosis via increasing Collagen Deposition

View Article: PubMed Central - PubMed

ABSTRACT

Although increases in cardiovascular load (pressure overload) are known to elicit ventricular remodeling including cardiomyocyte hypertrophy and interstitial fibrosis, the molecular mechanisms of pressure overload or AngII -induced cardiac interstitial fibrosis remain elusive. In this study, serpinE2/protease nexin-1 was over-expressed in a cardiac fibrosis model induced by pressure-overloaded via transverse aortic constriction (TAC) in mouse. Knockdown of serpinE2 attenuates cardiac fibrosis in a mouse model of TAC. At meantime, the results showed that serpinE2 significantly were increased with collagen accumulations induced by AngII or TGF-β stimulation in vitro. Intriguingly, extracellular collagen in myocardial fibroblast was reduced by knockdown of serpinE2 compared with the control in vitro. In stark contrast, the addition of exogenous PN-1 up-regulated the content of collagen in myocardial fibroblast. The MEK1/2- ERK1/2 signaling probably promoted the expression of serpinE2 via transcription factors Elk1 in myocardial fibroblast. In conclusion, stress-induced the ERK1/2 signaling pathway activation up-regulated serpinE2 expression, consequently led accumulation of collagen protein, and contributed to cardiac fibrosis.

No MeSH data available.


Potential involvement of ERK 1/2 signal pathway in regulating serpinE2 and collagen deposition.(a) Collagen mRNA levels measured by qRT-PCR after 24 and 48 h treatment with U0126 (The inhibitor of ERK 1/2 signal pathway). n = 10, *P < 0.05 vs. control. (b) Collagen expression levels detected by Sircol™ Collagen Assay after 24 and 48 h treatment with U0126. n = 6, *P < 0.05 vs. control. (c) After treatment with U0126, the protein and mRNA level of serpinE2 was detected in cardiac fibroblasts. n = 6, *P < 0.05 vs. control. Full-length blots/gels are presented in Supplementary Figure 8. (d) Effect of U0126 on expression of phosphor-ERK1/2 (p-ERK1/2) and total ERK1/2 (t-ERK1/2) in myocardial fibroblasts inducd by AngII or TGF-β. n = 8, *P < 0.05 vs. control, #P < 0.05 vs. AngII or TGF-β. Left, western blot. Right, Elisa assay. Full-length blots/gels are presented in Supplementary Figure 9 and 10. (e) Effect of U0126 on expression of serpinE2 and collagen in myocardial fibroblasts inducd by ANG-II. n = 4, *P < 0.05 vs. control, #P < 0.05 vs. ANG-II. Full-length blots/gels are presented in Supplementary Figure 11 and 12. (f) Effect of U0126 on the collagen content inducd by AngII or TGF-β in supernatants of fibroblasts. n = 6, *P < 0.05 vs. control, #P < 0.05 vs. AngII or TGF-β. (g) Effect of U0126 on expression of serpinE2 inducd by AngII in supernatants of fibroblasts. n = 6, *P < 0.05 vs. control, #P < 0.05 vs. ANG-II.
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f6: Potential involvement of ERK 1/2 signal pathway in regulating serpinE2 and collagen deposition.(a) Collagen mRNA levels measured by qRT-PCR after 24 and 48 h treatment with U0126 (The inhibitor of ERK 1/2 signal pathway). n = 10, *P < 0.05 vs. control. (b) Collagen expression levels detected by Sircol™ Collagen Assay after 24 and 48 h treatment with U0126. n = 6, *P < 0.05 vs. control. (c) After treatment with U0126, the protein and mRNA level of serpinE2 was detected in cardiac fibroblasts. n = 6, *P < 0.05 vs. control. Full-length blots/gels are presented in Supplementary Figure 8. (d) Effect of U0126 on expression of phosphor-ERK1/2 (p-ERK1/2) and total ERK1/2 (t-ERK1/2) in myocardial fibroblasts inducd by AngII or TGF-β. n = 8, *P < 0.05 vs. control, #P < 0.05 vs. AngII or TGF-β. Left, western blot. Right, Elisa assay. Full-length blots/gels are presented in Supplementary Figure 9 and 10. (e) Effect of U0126 on expression of serpinE2 and collagen in myocardial fibroblasts inducd by ANG-II. n = 4, *P < 0.05 vs. control, #P < 0.05 vs. ANG-II. Full-length blots/gels are presented in Supplementary Figure 11 and 12. (f) Effect of U0126 on the collagen content inducd by AngII or TGF-β in supernatants of fibroblasts. n = 6, *P < 0.05 vs. control, #P < 0.05 vs. AngII or TGF-β. (g) Effect of U0126 on expression of serpinE2 inducd by AngII in supernatants of fibroblasts. n = 6, *P < 0.05 vs. control, #P < 0.05 vs. ANG-II.

Mentions: ERK1/2 is one of members of the mitogen-activated protein kinase (MAPK) cascade and regulates cell proliferation and differentiation, which is involved in cardiomyocyte hypertrophic growth and fibrosis24. The corresponding results of collagen mRNA and content of collagen expression were also reduced after treatment with ERK1/2 inhibitor U0126 (Fig. 6a,b). Published evidence indicates that activation of ERK signaling up-regulates the serpinE2 expression in mouse embryonic fibroblasts25. Therefore, ERK1/2 signal pathway in regulating serpinE2 in myocardial fibroblasts is highly expected. To test this hypothesis, the expression of serpinE2 were evaluated and the results showed that, along with the inhibition of ERK1/2 signal pathway by U0126, the protein and mRNA level of serpinE2 were both down-regulated significantly (Fig. 6c).


Overexpression of SerpinE2/protease nexin-1 Contribute to Pathological Cardiac Fibrosis via increasing Collagen Deposition
Potential involvement of ERK 1/2 signal pathway in regulating serpinE2 and collagen deposition.(a) Collagen mRNA levels measured by qRT-PCR after 24 and 48 h treatment with U0126 (The inhibitor of ERK 1/2 signal pathway). n = 10, *P < 0.05 vs. control. (b) Collagen expression levels detected by Sircol™ Collagen Assay after 24 and 48 h treatment with U0126. n = 6, *P < 0.05 vs. control. (c) After treatment with U0126, the protein and mRNA level of serpinE2 was detected in cardiac fibroblasts. n = 6, *P < 0.05 vs. control. Full-length blots/gels are presented in Supplementary Figure 8. (d) Effect of U0126 on expression of phosphor-ERK1/2 (p-ERK1/2) and total ERK1/2 (t-ERK1/2) in myocardial fibroblasts inducd by AngII or TGF-β. n = 8, *P < 0.05 vs. control, #P < 0.05 vs. AngII or TGF-β. Left, western blot. Right, Elisa assay. Full-length blots/gels are presented in Supplementary Figure 9 and 10. (e) Effect of U0126 on expression of serpinE2 and collagen in myocardial fibroblasts inducd by ANG-II. n = 4, *P < 0.05 vs. control, #P < 0.05 vs. ANG-II. Full-length blots/gels are presented in Supplementary Figure 11 and 12. (f) Effect of U0126 on the collagen content inducd by AngII or TGF-β in supernatants of fibroblasts. n = 6, *P < 0.05 vs. control, #P < 0.05 vs. AngII or TGF-β. (g) Effect of U0126 on expression of serpinE2 inducd by AngII in supernatants of fibroblasts. n = 6, *P < 0.05 vs. control, #P < 0.05 vs. ANG-II.
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f6: Potential involvement of ERK 1/2 signal pathway in regulating serpinE2 and collagen deposition.(a) Collagen mRNA levels measured by qRT-PCR after 24 and 48 h treatment with U0126 (The inhibitor of ERK 1/2 signal pathway). n = 10, *P < 0.05 vs. control. (b) Collagen expression levels detected by Sircol™ Collagen Assay after 24 and 48 h treatment with U0126. n = 6, *P < 0.05 vs. control. (c) After treatment with U0126, the protein and mRNA level of serpinE2 was detected in cardiac fibroblasts. n = 6, *P < 0.05 vs. control. Full-length blots/gels are presented in Supplementary Figure 8. (d) Effect of U0126 on expression of phosphor-ERK1/2 (p-ERK1/2) and total ERK1/2 (t-ERK1/2) in myocardial fibroblasts inducd by AngII or TGF-β. n = 8, *P < 0.05 vs. control, #P < 0.05 vs. AngII or TGF-β. Left, western blot. Right, Elisa assay. Full-length blots/gels are presented in Supplementary Figure 9 and 10. (e) Effect of U0126 on expression of serpinE2 and collagen in myocardial fibroblasts inducd by ANG-II. n = 4, *P < 0.05 vs. control, #P < 0.05 vs. ANG-II. Full-length blots/gels are presented in Supplementary Figure 11 and 12. (f) Effect of U0126 on the collagen content inducd by AngII or TGF-β in supernatants of fibroblasts. n = 6, *P < 0.05 vs. control, #P < 0.05 vs. AngII or TGF-β. (g) Effect of U0126 on expression of serpinE2 inducd by AngII in supernatants of fibroblasts. n = 6, *P < 0.05 vs. control, #P < 0.05 vs. ANG-II.
Mentions: ERK1/2 is one of members of the mitogen-activated protein kinase (MAPK) cascade and regulates cell proliferation and differentiation, which is involved in cardiomyocyte hypertrophic growth and fibrosis24. The corresponding results of collagen mRNA and content of collagen expression were also reduced after treatment with ERK1/2 inhibitor U0126 (Fig. 6a,b). Published evidence indicates that activation of ERK signaling up-regulates the serpinE2 expression in mouse embryonic fibroblasts25. Therefore, ERK1/2 signal pathway in regulating serpinE2 in myocardial fibroblasts is highly expected. To test this hypothesis, the expression of serpinE2 were evaluated and the results showed that, along with the inhibition of ERK1/2 signal pathway by U0126, the protein and mRNA level of serpinE2 were both down-regulated significantly (Fig. 6c).

View Article: PubMed Central - PubMed

ABSTRACT

Although increases in cardiovascular load (pressure overload) are known to elicit ventricular remodeling including cardiomyocyte hypertrophy and interstitial fibrosis, the molecular mechanisms of pressure overload or AngII -induced cardiac interstitial fibrosis remain elusive. In this study, serpinE2/protease nexin-1 was over-expressed in a cardiac fibrosis model induced by pressure-overloaded via transverse aortic constriction (TAC) in mouse. Knockdown of serpinE2 attenuates cardiac fibrosis in a mouse model of TAC. At meantime, the results showed that serpinE2 significantly were increased with collagen accumulations induced by AngII or TGF-&beta; stimulation in vitro. Intriguingly, extracellular collagen in myocardial fibroblast was reduced by knockdown of serpinE2 compared with the control in vitro. In stark contrast, the addition of exogenous PN-1 up-regulated the content of collagen in myocardial fibroblast. The MEK1/2- ERK1/2 signaling probably promoted the expression of serpinE2 via transcription factors Elk1 in myocardial fibroblast. In conclusion, stress-induced the ERK1/2 signaling pathway activation up-regulated serpinE2 expression, consequently led accumulation of collagen protein, and contributed to cardiac fibrosis.

No MeSH data available.