Background/Purpose:  Systemic sclerosis (SSc) is an autoimmune connective tissue disease characterized by activation of the immune system, vascular dysfunction and tissue fibrosis. Vascular dysfunction in SSc is one of the most prominent features of the disease that manifest by Raynaud’s phenomenon in the early stages and by proliferative vasculopathy that affect all involved organs and progress throughout the disease course. Endothelin 1 (ET-1) is overexpressed in SSc as illustrated by elevated circulating levels and increased tissue expression in involved organs. We previously reported increased proliferative capacity of SSc- vascular smooth muscle cells (vSMCs) in association with resistance to apoptosis in comparison to control cells. In this study, we sought to examine the role of ET-1 and its signaling pathways in the generation of the activated vSMCs phenotype.

Methods:  We isolated vSMCs from 4mm punch skin biopsies from 3 patients with SSc and 3 healthy controls. We evaluated the expression levels of EDN1, EDNRA and EDNRB, which encodes for ET-1, Endothelin Receptor A (ERA) and ERB, respectively, in vSMCs from SSc patients and controls. We treated vSMCs with ET-1, in the presence or absence of selective ETA antagonist (BQ123), selective ETB antagonist (BQ788) or dual ETA+B antagonist (PD145065). Under these conditions, we examined the effects of ET-1 on cell proliferation using BrdU assay, viability by MTT assay and apoptosis by Tunel assay.

Results:  SSc-vSMCs overexpressed EDN1 compared to control vSMCs (120 fold). Furthermore, and compared to control-vSMCs,  EDRA expression levels were significantly increased in SSc-vSMCs while EDNRB expression levels were lower than control values. At baseline, SSc-vSMCs exhibited 1.8-fold increase in cell proliferation compared to control vSMCs (P= 0.0031). Upon treating control-vSMCs with 10nM ET-1, we demonstrate increased vSMCs proliferation by 1.6 folds (P= 0.03) in association with decreased cell apoptosis by 36%. The addition of selective ETB antagonist (BQ788) or dual ETA+B antagonist (PD145065) reduced vSMCs proliferation by 72% and 69%, respectively (P<0.05), but we did not find significant effect for selective ETA antagonist (PD156707) on cell proliferation (P>0.05). Moreover, BQ788 and PD145065 increased apoptosis of vSMCs. We did not see this effect using the selective ETA antagonist.

Conclusion:  We provide an experimental evidence for increased proliferation, viability and decreased apoptosis of vSMCs by ET-1 in vitro, and we demonstrate that this effect is mediated by ETB receptors. Selective ETB and non-selective ETA+B receptor antagonists have a theoretical advantage over selective ETA antagonists in reducing the effect of ET-1 on proliferation of cells in tunica media, and that ET-1 have an important role in the development of vascular remodeling in SSc.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/the-role-of-endothelin-1-in-activation-of-vascular-smooth-muscle-cells-in-systemic-sclerosis-increased-cell-proliferation-and-resistance-to-apoptosis-mediated-by-endotehlin-b-receptors/