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C-type natriuretic peptide attenuates LPS-induced endothelial activation: involvement of p38, Akt, and NF-κB pathways

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Abstract

Endothelial activation elicited by inflammatory agents is regarded as a key event in the pathogenesis of several vascular inflammatory diseases. In the present study, the inhibitory effects and underlying mechanism of C-type natriuretic peptide (CNP) on LPS-induced endothelial activation were examined in human umbilical vein endothelial cells (HUVECs). The effect of CNP on adhesion molecule expression was assessed using quantitative real-time RT-PCR and western blotting analyses. The nuclear factor-κB (NF-κB), MAPK, and PI3K/Akt signaling pathways in LPS-stimulated HUVECs were investigated using western blotting analyses, and the production of intracellular reactive oxygen species (ROS) was measured using a fluorescence method. Pretreatment with CNP inhibited LPS-induced expression of intercellular adhesion molecule-1, vascular cell adhesion molecule-1, E-selectin, and P-selectin in a concentration-dependent manner. CNP suppressed the phosphorylation of p65 and NF-κB activation in LPS-stimulated cells. Moreover, CNP reduced ERK1/2 and p38 phosphorylation induced by LPS but not JNK. Furthermore, CNP induced Akt phosphorylation and activation of hemeoxygenase-1 (HO-1) expression. CNP significantly inhibited the production of intracellular ROS. These results suggest that CNP effectively attenuated LPS-induced endothelial activation by inhibiting the NF-κB and p38 signaling pathways, eliminating LPS-induced intracellular ROS production, and activating the PI3K/Akt/HO-1 pathway in HUVECs; thereby, demonstrating that CNP may be a potential therapeutic target for the treatment of sepsis and inflammatory vascular diseases.

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Abbreviations

CNP:

C-type natriuretic peptide

LPS:

Lipopolysaccharide

VCAM-1:

Vascular cell adhesion molecule-1

ICAM-1:

Intercellular adhesion molecule-1

ROS:

Reactive oxygen species

HO-1:

Hemeoxygenase-1

HUVECs:

Human umbilical vein endothelial cells

NF-κB:

Nuclear factor-κB

PBS:

Phosphate-buffered saline

AREs:

Antioxidant response elements

EGM-2:

Endothelial cell growth medium-2

DCFH-DA:

2′,7-Dichlorodihydrofluorescein diacetate

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Acknowledgments

This work was supported by grants from the National Natural Science Foundation of China (No. 31271001) and the National High Technology Research and Development Program of China (No. 2012AA021902).

Conflict of interest

The authors declare that they have no competing interests.

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Authors and Affiliations

  1. Institute of Transfusion Medicine, Academy of Military Medical Sciences, No. 27th Taiping Road, Haidian, Beijing, China

    Gan Chen, Jingxiang Zhao, Yujing Yin, Bo Wang, Qingjun Liu, Penglong Li, Lian Zhao & Hong Zhou

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  1. Gan Chen
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  2. Jingxiang Zhao
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  3. Yujing Yin
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  4. Bo Wang
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  5. Qingjun Liu
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  7. Lian Zhao
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  8. Hong Zhou
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Correspondence to Lian Zhao or Hong Zhou.

Additional information

G. Chen and J. Zhao contributed equally to this work.

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726_2014_1816_MOESM1_ESM.tif

Supplementary material 1 (TIFF 345 kb) Figure S1. CNP do not affect the viability of HUVECs. HUVECs were plated in 96-well plates, pretreated with CNP (1 μM) for 120 min, followed by incubation with LPS (1 μg/ml) for 24 h. The cell viability was determined by MTT assay. Data are plotted as the mean ± SD of three independent experiments, performed in triplicate. & P < 0.05 versus the control group

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Chen, G., Zhao, J., Yin, Y. et al. C-type natriuretic peptide attenuates LPS-induced endothelial activation: involvement of p38, Akt, and NF-κB pathways. Amino Acids 46, 2653–2663 (2014). https://doi.org/10.1007/s00726-014-1816-x

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