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Vangl2 participates in the primary ciliary assembly under low fluid shear stress in hUVECs

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Abstract

The pattern of blood fluid shear stress (FSS) is considered the main factor that affects ciliogenesis in human umbilical vein endothelial cells (hUVECs), the underlying mechanism is unclear. Microfluidic chamber experiments were carried out to load hUVECs with low fluid shear stress (LSS, 0.1 dynes/cm2) or high fluid shear stress (HSS, 15 dynes/cm2). Van Gogh2 (Vangl2), a core protein in the planar cell polarity (PCP) pathway, was silenced and overexpressed in hUVECs. Immunofluorescence analysis showed that primary cilia assemble under LSS while disassembling under HSS. Vangl2 expression was consistent with cilia assembly, and its localization showed a polar distribution under LSS. Furthermore, the average number of ciliated cells and primary cilia length were increased in the Vangl2 overexpressing cell lines (the OE group) but decreased in the Vangl2 silenced cell lines (the SH group). When these cells were loaded with different FSS, more ciliated cells with longest primary cilia were observed in the LSS loaded OE group compared with those in the other groups. Immunoprecipitation showed that the interaction between Bardet-Biedl syndrome 8 (BBS8) and Vangl2 was enhanced following LSS loading compared to that under HSS. However, the interactions between phosphorylated dishevelled segment polarity protein 2 (pDvl2), kinesin family member 2a (Kif2a), and polo-like kinase 1 (Plk1) and Vangl2 were restrained following LSS loading. Overall, the results indicated that Vangl2 played a significant role during LSS-induced primary cilia assembly by recruiting BBS to promote the apical docking of basal bodies and by restraining Dvl2 phosphorylation from reducing primary cilia disassembly.

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Primary cilia length among ciliated cells was quantified from 22 Images and 3 independent experiments using ImageJ. (d) The percentage of ciliated cells was manually counted in 9 different images for each group. At least 200 cells were counted from each image. The results are presented as average ± SD, n = 5, *P < 0.05, **P < 0.01, vs. BC groups; ##P < 0.01, vs. the OE groups, t-test for independent samples; &&P < 0.01, vs. the SH-HSS group, multi-factor ANOVA analyze the effect of dual factors

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Acknowledgements

The authors thank Prof. Zhi Zhang and Zhuo Ming for their technical assistance.

Funding

This work was supported by a grant from the National Natural Science Foundation of China (No. 31360278).

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  1. Xin Sheng and Shuanglin Gao contributed equally to this work.

Authors and Affiliations

  1. Department of Biochemistry, Zunyi Medical University, Zunyi, 563000, People’s Republic of China

    Xin Sheng, Shuanglin Gao, Xiadan Xie, Junhua Wang & Yan He

  2. Laboratory of Basic Medical Morphology, Zunyi Medical University, Zunyi, 563000, People’s Republic of China

    Yan Sheng

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Contributions

XS conceived and coordinated the study, designed, performed, and analyzed the experiments, wrote the paper. SLG carried out the data collection, YS analyzed data, XDX revised the paper, WJH contributed to the reagents and materials, and HY provided analysis tools. All authors reviewed the results and approved the final version of the manuscript.

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Correspondence to Xin Sheng.

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Sheng, X., Gao, S., Sheng, Y. et al. Vangl2 participates in the primary ciliary assembly under low fluid shear stress in hUVECs. Cell Tissue Res 387, 95–109 (2022). https://doi.org/10.1007/s00441-021-03546-0

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