Microvascular Endothelial Cells License APS Vasculopathy Through YAP1- and CCN2-Mediated Signaling
Abstract
BACKGROUND: Whereas antiphospholipid syndrome (APS) is best known for increasing the risk of macrovascular thrombosis, APS vasculopathy is characterized by the abnormal proliferation of endothelial and smooth muscle cells, leading to occlusion of small blood vessels in the skin, kidneys, and heart, among other organs. The underlying mechanisms remain unclear, and targeted treatment options for patients with APS are lacking. METHODS: To identify and analyze APS microvascular endothelial cells (MVECs), skin biopsies of patients with APS complicated by livedo racemosa were characterized using single-cell RNA sequencing. CCN1 (cellular communication network factor 1) and CCN2 were identified in skin and kidney biopsies by immunofluorescence microscopy and in plasma by ELISA. Healthy dermal MVECs were cultured with APS patient–derived serum or immunoglobulin G (IgG), and relevant signaling pathways were characterized using quantitative PCR, immunoblotting, and immunofluorescence microscopy. The proliferation and migration of vascular smooth muscle cells were determined after exposure to conditioned medium from APS IgG–stimulated MVECs. A mouse model of APS IgG–accelerated neointima formation was developed. Anti-CCN2 monoclonal antibodies were tested in vascular smooth muscle cell functional assays and the mouse model. RESULTS: Increased endothelial cell expression of CCN1 and CCN2 was identified by single-cell RNA sequencing and further confirmed in APS skin by microscopy as well as in APS plasma by ELISA. Exposure of healthy MVECs to patient IgG triggered the upregulation of CCN1 and CCN2 via Toll-like receptor 4– and YAP1 (yes-associated protein 1)–mediated signaling. CCN2, originating from APS IgG–stimulated MVECs led to the proliferation and migration of vascular smooth muscle cells, phenotypes that were inhibited by an anti-CCN2 antibody or depletion of EGFR (epidermal growth factor receptor). We further observed increased expression of CCN2, along with evidence of YAP1 nuclear translocation, in kidney vessels of APS nephropathy biopsies. Finally, CCN2 inhibition with an anti-CCN2 monoclonal antibody significantly reduced neointima thickening and cell proliferation in a mouse model of APS IgG–accelerated neointima formation. CONCLUSIONS: This study revealed activation of YAP1-mediated signaling in APS dermal microvessels and demonstrated that the YAP1 target CCN2 plays a role in facilitating pro-proliferative communication between MVECs and vascular smooth muscle cells. These findings offer insights into the cellular and molecular mechanisms underlying APS vasculopathy, providing potential therapeutic targets for patients.
Article Details
Authors (26)
Hui Shi
Wenying Liang
Departments of Internal Medicine (H.S., W.L., C.K.H., Y. Shen, C.S., S.Y., P.K., L.H., C.E.V., A.T., J.A.M., Y.Z., P.-S.T., J.S.K.), University of Michigan, Ann Arbor, MI.
Zhixia Yang
Claire K. Hoy
Departments of Internal Medicine (H.S., W.L., C.K.H., Y. Shen, C.S., S.Y., P.K., L.H., C.E.V., A.T., J.A.M., Y.Z., P.-S.T., J.S.K.), University of Michigan, Ann Arbor, MI.
Qiuyu Wang
Department of Endocrinology and Metabolism, Zhongshan Hospital, Fudan University, Shanghai, China (Q.W.).
Zetao Ding
Department of Rheumatology and Immunology, Ruijin Hospital, School of Medicine, Shanghai Jiao Tong University, Shanghai, China (H.S., Z.Y., Z.D., H.P., C.Y.).
Haoyu Pan
Interdisciplinary Materials Research Center, School of Materials Science and Engineering
Yue Shi
Department of Chemistry, School of Science
Yiran Shen
Departments of Internal Medicine (H.S., W.L., C.K.H., Y. Shen, C.S., S.Y., P.K., L.H., C.E.V., A.T., J.A.M., Y.Z., P.-S.T., J.S.K.), University of Michigan, Ann Arbor, MI.
Cyrus Sarosh
Departments of Internal Medicine (H.S., W.L., C.K.H., Y. Shen, C.S., S.Y., P.K., L.H., C.E.V., A.T., J.A.M., Y.Z., P.-S.T., J.S.K.), University of Michigan, Ann Arbor, MI.
Srilakshmi Yalavarthi
Departments of Internal Medicine (H.S., W.L., C.K.H., Y. Shen, C.S., S.Y., P.K., L.H., C.E.V., A.T., J.A.M., Y.Z., P.-S.T., J.S.K.), University of Michigan, Ann Arbor, MI.
Pooja Kapoor
Departments of Internal Medicine (H.S., W.L., C.K.H., Y. Shen, C.S., S.Y., P.K., L.H., C.E.V., A.T., J.A.M., Y.Z., P.-S.T., J.S.K.), University of Michigan, Ann Arbor, MI.
Lucas Hudgins
Departments of Internal Medicine (H.S., W.L., C.K.H., Y. Shen, C.S., S.Y., P.K., L.H., C.E.V., A.T., J.A.M., Y.Z., P.-S.T., J.S.K.), University of Michigan, Ann Arbor, MI.
Caroline E. Vance
Departments of Internal Medicine (H.S., W.L., C.K.H., Y. Shen, C.S., S.Y., P.K., L.H., C.E.V., A.T., J.A.M., Y.Z., P.-S.T., J.S.K.), University of Michigan, Ann Arbor, MI.
Rachael Wasikowski
Gabriel Figueroa-Parra
Department of Rheumatology, Mayo Clinic, Rochester, MN (G.F.-P., A.D.-G.).
Ajay Tambralli
Departments of Internal Medicine (H.S., W.L., C.K.H., Y. Shen, C.S., S.Y., P.K., L.H., C.E.V., A.T., J.A.M., Y.Z., P.-S.T., J.S.K.), University of Michigan, Ann Arbor, MI.
Jacqueline A. Madison
Departments of Internal Medicine (H.S., W.L., C.K.H., Y. Shen, C.S., S.Y., P.K., L.H., C.E.V., A.T., J.A.M., Y.Z., P.-S.T., J.S.K.), University of Michigan, Ann Arbor, MI.
Lam C. Tsoi
Johann E. Gudjonsson
Chengde Yang
Alí Duarte-García
Department of Rheumatology, Mayo Clinic, Rochester, MN (G.F.-P., A.D.-G.).
Yu Zuo
Allison C. Billi
Pei-Suen Tsou
Departments of Internal Medicine (H.S., W.L., C.K.H., Y. Shen, C.S., S.Y., P.K., L.H., C.E.V., A.T., J.A.M., Y.Z., P.-S.T., J.S.K.), University of Michigan, Ann Arbor, MI.
Jason S. Knight
Departments of Internal Medicine (H.S., W.L., C.K.H., Y. Shen, C.S., S.Y., P.K., L.H., C.E.V., A.T., J.A.M., Y.Z., P.-S.T., J.S.K.), University of Michigan, Ann Arbor, MI.