Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2014 Dec 2;9(12):e112695.
doi: 10.1371/journal.pone.0112695. eCollection 2014.

BMP4 increases the expression of TRPC and basal [Ca2+]i via the p38MAPK and ERK1/2 pathways independent of BMPRII in PASMCs

Yi Zhang  1 , Yingfeng Wang  2 , Kai Yang  3 , Lichun Tian  2 , Xin Fu  2 , Yan Wang  2 , Yueqian Sun  4 , Qian Jiang  2 , Wenju Lu  5 , Jian Wang  6
Affiliations

BMP4 increases the expression of TRPC and basal [Ca2+]i via the p38MAPK and ERK1/2 pathways independent of BMPRII in PASMCs

Yi Zhang et al. PLoS One. .

Abstract

Multiple abnormalities of bone morphogenetic protein (BMPs) signaling are implicated in the process of pulmonary arterial hypertension (PAH). BMP4 plays an important role during the process of pulmonary arterial remodeling and mutant of the principle BMP4 receptor, BMP receptors II (BMPRII), is found to associate with the development of PAH. However, the likely mechanism defining the contribution of BMPRII to BMP4 mediated signaling in pulmonary arterial smooth muscle cells (PASMCs) remains comprehensively unclear. We previously found that enhanced store operated calcium entry (SOCE) and basal intracellular calcium concentration [Ca2+]i were induced by BMP4 via upregulation of TRPC1, 4 and 6 expression in PASMCs, and that BMP4 modulated TRPC channel expression through activating p38MAPK and ERK1/2 signaling pathways. In this study, BMPRII siRNA was used to knockdown BMPRII expression to investigate whether BMP4 upregulates the expression of TRPC and activating Smad1/5/8, ERK1/2 and p38MAPK pathway via BMPRII in distal PASMCs. Our results showed that knockdown of BMPRII: 1) attenuated BMP4 induced activation of P-Smad1/5/8, without altering BMP4 induced P-p38MAPK and P-ERK1/2 activation in PASMCs; 2) did not attenuate the BMP4-induced TRPC1, 4 and 6 expression; 3) did not affect BMP4-enhanced SOCE and basal [Ca2+]i. Thus, we concluded that BMP4 activated Smad1/5/8 pathway is BMPRII-dependent, while the BMP4 - ERK/p-P38 - TRPC - SOCE signaling axis are likely mediated through other receptor rather than BMPRII.

PubMed Disclaimer

Conflict of interest statement

Competing Interests: The authors have declared that no competing interests exist.

Figures

Figure 1
Figure 1. Specific siRNA effectively knockdown BMPRII protein without affecting BMPRIa protein in PASMCs.
Images showing the blots of BMPRII protein for 48 h or 108 h (A) and BMPRIa protein relative to α-actin as detected by Western blotting in PASMCs treated with BMPRII small interference siRNA or non-targeting (NT) control siRNA for 48 h (C). Bar values are means ± SEM (n  =  4 for each group). *P <0.01 vs. respective siNT control (B and D).
Figure 2
Figure 2. BMP4 induced activation of P-Smad1/5/8, P-p38MAPK and P-ERK1/2 in rat distal PASMC.
(A) represents the blots of P-Smad1/5/8, P-p38MAPK, P-ERK1/2, t-Smad1/5/8, t-p38MAPK and t-ERK1/2 upon BMP4 treatment (50 ng/ml) in dose and time dependent manner. (B) Bar values are means ± SEM (n  =  3 for each group). (C) represents the blots of BMPRII upon BMP4 treatment for 5–30 min. (D) Bar values are means ± SEM (n  =  3 for each group).*P <0.01 vs. respective vehicle control.
Figure 3
Figure 3. Knockdown of BMPRII attenuated BMP4 induced phosphorylation of P-Smad1/5/8, but not P-p38MAPK and P-ERK1/2 in PASMC.
(A), (B) and (C) show blots (left) and normalized bar graphs (right) of P-Smad1/5/8, P-p38MAPK, P-ERK1/2, t-Smad1/5/8, t- p38MAPK and t-ERK1/2, respectively. Bar values were means ± SEM (n  =  4 for each group). *P <0.01 vs. respective siNT control.
Figure 4
Figure 4. Knockdown of BMPRII did not affect BMP4 induced TRPC expression in PASMCs.
(A) bar graphs showing the real-time PCR results indicating the mRNA expression of TRPC1, TRPC4 and TRPC6. *P <0.05 vs. respective vehicle control (A). (B) blots represent the protein expression of TRPC1, 4 and 6 after treatments of BMPRII and BMP4 and (C) represent the normalized bar graphs of TRPC1, TRPC4 and TRPC6 protein expression. Bar values are means ± SEM; n  =  3 in each group. §P <0.05 vs. respective siNT control.
Figure 5
Figure 5. Knockdown of BMPRII did not affect BMP4 elevated basal [Ca2+]i and SOCE in PASMCs.
(A) bar graph showing the changes in basal [Ca2+]i in distal PASMCs treated with 50 ng/ml BMP4 (n  =  4 from 120 cells) for 60 h. Bar values are means ± SEM. *P <0.05 vs. siNT control (n  =  4 from 120 cells). (B) represting time course of quenching of Fura-2 AM fluorescence at 360 nm by 200 µM Mn2+ after perfusion with Ca2+ -free KRBS (0 Ca2+) containing Nifedipine (5 µM) and CPA (10 µM) in distal PASMCs treated with 50 ng/ml BMP4 (n = 4 experiments in 110 cells) or vehicle for 60 h, normalized to fluorescence at time 0 (F/F0). (C) Bar graph shows that Mn2+ quenching, expressed as percent decrease in fluorescence at time 10 min from time 0, was greater in BMP4-treated distal PASMCs compared with that in control cells (*P<0.05). Bar values are means ± SEM.
Figure 6
Figure 6. Expression profile of BMP receptors after BMP4 treatment in PASMCs.
Representative blots (A) and normalized bar graphs (B) of BMPRII, ActRIIa, ActRIIb, ALK2, BMPRIa and BMPRIb protein as detected by Western blotting in PASMCs treated with 50 ng/ml BMP4 for 60 h. Bar graph shows mean protein expression for BMPRII, ActRIIa, ActRIIb, ALK2, BMPRIa and BMPRIb protein relative to α-actin (n  =  4 in each group; *P <0.05 vs. vehicle control). Bar values are as means ± SEM.
See this image and copyright information in PMC

Similar articles

See all similar articles

Cited by

See all "Cited by" articles

References

    1. Eickelberg O, Morty RE (2007) Transforming growth factor beta/bone morphogenic protein signaling in pulmonary arterial hypertension: remodeling revisited. Trends Cardiovasc Med 17:263–269. - PubMed
    1. Young KA, Ivester C, West J, Carr M, Rodman DM (2006) BMP signaling controls PASMC KV channel expression in vitro and in vivo. Am J Physiol Lung Cell Mol Physiol 290:L841–848. - PubMed
    1. Takeda M, Otsuka F, Nakamura K, Inagaki K, Suzuki J, et al. (2004) Characterization of the bone morphogenetic protein (BMP) system in human pulmonary arterial smooth muscle cells isolated from a sporadic case of primary pulmonary hypertension: roles of BMP type IB receptor (activin receptor-like kinase-6) in the mitotic action. Endocrinology 145:4344–4354. - PubMed
    1. Hruska KA, Mathew S, Saab G (2005) Bone morphogenetic proteins in vascular calcification. Circ Res 97:105–114. - PubMed
    1. Gutierrez FR, Moran CJ, Ludbrook PA, McKnight RC, Weldon CS (1980) Pulmonary arterial calcification with reversible pulmonary hypertension. AJR Am J Roentgenol 135:177–178. - PubMed

Publication types

MeSH terms