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Herz

, Volume 44, Issue 1, pp 60–68 | Cite as

Ginsenoside-Rb3 inhibits endothelial–mesenchymal transition of cardiac microvascular endothelial cells

  • L. Yang
  • Q. LiuEmail author
  • Y. Yu
  • H. Xu
  • S. Chen
  • S. Shi
Original articles
  • 123 Downloads

Abstract

Background

We investigated the effect of Ginsenoside-Rb3 (Rb3) on the endothelial-to-mesenchymal transition (EMT) of cardiac microvascular endothelial cells (CMVECs) following coxsackievirus B3 (CVB3) infection.

Methods

CMVECs were infected with 100 TCID50 CVB3 (CVB3 group) or treated with Rb3 (Rb3 group); stably cultured CMVECs were used as control. Cells treated with the Pyk2 inhibitor TAE226 and PI3K inhibitor LY294002 were used for additional experiments. Cell viability was assessed with the Cell Counting Kit-8 (CCK8). Expression of CD31 and α‑smooth muscle actin (α-SMA) was evaluated by immunofluorescence (IF) and Western blotting (WB). Expression of Pyk2, PI3K, and AKT was assessed by real-time polymerase chain reaction (RT-PCR) and WB.

Results

Cell morphology, including cell pyknosis, and viability were significantly impaired by CVB3 infection (p < 0.05). However, the morphology of the Rb3 group was unaffected. The CCK8 assay showed that viability in the Rb3 group was increased compared with the CVB3 group (p < 0.05). Expression of CD31 decreased and α‑SMA increased in the CVB3 group compared with the control group (p < 0.05), but CD31 increased while α‑SMA decreased in the Rb3 group compared with the CVB3 group (p < 0.05). IF staining showed the same trends. The levels of Pyk2, PI3K, AKT, and CD31 were up-regulated in the Rb3 group compared with the CVB3 group, whereas α‑SMA decreased (p < 0.05). In the Pyk2-inhibitor group, PI3K, AKT, and CD31 expression was down-regulated while α‑SMA expression increased in comparison with the Rb3 group (p < 0.05). In the PI3K-inhibitor group, the levels of AKT and CD31 decreased while α‑SMA increased (p < 0.05), although the level of Pyk2 expression showed no obvious change in comparison with the Rb3 group.

Conclusion

Rb3 inhibited EMT in CMVECs following CVB3 infection via the Pyk2–PI3K–AKT signaling pathway.

Keywords

Ginsenosides Endomyocardial fibrosis Myocarditis Coxsackievirus infections Signal pathways 

Ginsenosid-Rb3 inhibiert die endothelial-mesenchymale Transition kardialer mikrovaskulärer Endothelzellen

Zusammenfassung

Hintergrund

Die Autoren untersuchten die Wirkung von Ginsenosid-Rb3 (Rb3) auf die endothelial-mesenchymale Transition (EMT) bei kardialen mikrovaskulären Endothelzellen (CMVEC) nach einer Infektion mit Coxsackie-Virus B3 (CVB3).

Methoden

Die CMVEC wurden mit 100 TCID50 CVB3 (CVB3-Gruppe) infiziert oder mit Rb3 (Rb3-Gruppe) behandelt; stabil kultivierte CMVEC wurden als Kontrollen verwendet. Mit dem Pyk2-Inhibitor TAE226 und dem PI3K-Inhibitor LY294002 behandelte Zellen wurden für weitere Versuche eingesetzt. Die Zellviabilität wurde mit dem Cell Counting Kit-8 (CCK8) bestimmt. Durch Immunfluoreszenz (IF) und Westernblot (WB) wurde die Expression von CD31 und glattmuskulärem α‑Aktin (α-SMA) ermittelt. Die Expression von Pyk2, PI3K und AKT wurde mittels Echtzeitpolymerasekettenreaktion (RT-PCR) und WB erfasst.

Ergebnisse

Die Zellmorphologie, einschließlich Zellpyknose, und die Zellviabilität waren durch die CVB3-Infektion signifikant beeinträchtigt (p < 0,05). Jedoch war die Morphologie in der Rb3-Gruppe nicht beeinflusst. Das CCK8-Assay zeigte, dass die Viabilität in der Rb3-Gruppe im Vergleich zur CVB3-Gruppe höher war (p < 0,05). In der CVB3-Gruppe nahm im Vergleich zur Kontrollgruppe die Expression von CD31 ab und die Expression von α‑SMA zu (p < 0,05), jedoch stieg in der Rb3-Gruppe im Vergleich zur CVB3-Gruppe CD31 an, während α‑SMA abfiel (p < 0,05). Die IF-Färbung zeigte die gleichen Tendenzen. Die Spiegel von Pyk2, PI3K, AKT und CD31 waren in der Rb3-Gruppe im Vergleich zur CVB3-Gruppe hochreguliert, während es bei α‑SMA zu einer Abnahme kam (p < 0,05). In der Pyk2-Inhibitor-Gruppe war die Expression von PI3K, AKT und CD31 herunterreguliert, während die α‑SMA-Expression im Vergleich zur Rb3-Gruppe zunahm (p < 0,05). In der PI3K-Inhibitor-Gruppe nahmen die Spiegel von AKT und CD31 ab, während α‑SMA anstieg (p < 0,05), obwohl die Werte für die Pyk2-Expression keine offensichtlichen Veränderungen gegenüber der Rb3-Gruppe aufwiesen.

Schlussfolgerung

Rb3 inhibierte die EMT in CMVEC nach CVB3-Infektion via Pyk2-PI3K-AKT-Signalweg.

Schlüsselwörter

Ginsenoside Endomyokardfibrose Myokarditis Coxsackie-Virus-Infektionen Signalwege 

Notes

Acknowledgements

This work was supported by research grants from the Natural Science Fund of ZheJiang Province in China (LY13H270013).

Compliance with ethical guidelines

Conflict of interest

L. Yang, Q. Liu, Y. Yu, H. Xu, S. Chen, and S. Shi declare that they have no competing interests.

This article does not contain any studies with human participants or animals performed by any of the authors.

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Copyright information

© Springer Medizin Verlag GmbH 2017

Authors and Affiliations

  • L. Yang
    • 1
  • Q. Liu
    • 1
    Email author
  • Y. Yu
    • 1
  • H. Xu
    • 1
  • S. Chen
    • 1
  • S. Shi
    • 2
  1. 1.The First Affiliated Hospital of Zhejiang Traditional Chinese Medicine UniversityHangzhouChina
  2. 2.College of PharmacyZhejiang Traditional Chinese Medicine UniversityHangzhouChina

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