Background/Purpose: Systemic sclerosis (SSc) is a multisystem autoimmune disease characterized by initial vascular injuries and resultant fibrosis of skin and certain internal organs. Although there have been no established treatments against SSc, recent studies have demonstrated that bosentan, a dual endothelin receptor antagonist, reverses nailfold capillary changes in SSc patients. However, the molecular mechanism explaining this observation still remains unknown. Therefore, we herein investigated the mechanism underlying the reversal effect of bosentan on SSc vasculopathy by using endothelial cell (EC)-specific Fli1 knockout mice (Fli1 ECKO mice), which mimic the functional and morphological abnormalities characteristic of SSc vasculopathy.

Methods: Eight-week-old Fli1 ECKO mice were administered intra-peritoneally with bosentan for 4 weeks. The expression levels of VE-Cadherin, PECAM1, a-SMA, and Fli1 were evaluated by immunohistochemistry. Vascular structure in the back skin was visualized by intravenously injected FITC-dextran. Vascular permeability was evaluated by the degree of Evans blue dye extravasation. The effects of bosentan on c-Abl, PKC-d, and Fli1 in human dermal microvascular ECs (HDMECs) were evaluated by immunoblotting.

Results: The structural and functional abnormalities, including stenosis of arterioles, dilation of capillaries, and increased vascular permeability, of small dermal blood vessels were improved by the administration of bosentan in Fli1 ECKO mice. Bosentan reversed the abnormal expression of various genes regulating EC-pericyte interaction, EC-EC interaction, and degradation of vascular basement membranes in dermal microvascular ECs isolated from Fli1 ECKO mice. In HDMECs, bosentan suppressed the activity of c-Abl and PKC-d, resulting in decreased phosphorylation levels of Fli1 at threonine 312. Consistent with the evidence that, when unphosphorylated, the DNA binding ability and the protein stability of Fli1 are increased, Fli1 protein levels were elevated along with the increase of its DNA binding, while Fli1 mRNA levels were not affected, after the exposure to bosentan in those cells.

Conclusion: Bosentan reverses the functional and morphological abnormalities of Fli1 ECKO mice at least partially by increasing the transcriptional activity and the protein levels of Fli1 through post-translational modification without affecting its mRNA expression. These results suggest that the efficacy of bosentan on SSc vasculopathy, in which Fli1 deficiency due to the epigenetic mechanism is deeply involved, is partly attributable to its reversal effect on Fli1 transcriptional activity and expression in SSc endothelial cells.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/bosentan-improves-vascular-abnormalities-in-endothelial-cell-specific-fli1-knockout-mice-by-increasing-the-dna-binding-ability-of-fli1-a-possible-mechanism-explaining-the-effect-of-bosentan-on/