Session Title: Systemic Sclerosis, Fibrosing Syndromes, and Raynaud's II
Session Type: Abstract Submissions (ACR)
Background/Purpose: Systemic Sclerosis (SSc) is an autoimmune disorder characterized by tissue fibrosis and vasculopathy. This latter comprises both neointima proliferation and defective angio and vasculogenesis. Although some of the clinical manifestations of the vasculopathy often precede the onset of fibrosis, there is scanty of data investigating the molecular mechanisms eventually linking the profibrotic activation of fibroblasts (FBs) and the poor angio/vasculogenesis in Scleroderma. Recently, a proteomic analysis of the secretome of SSc dermal FBs (SSc FBs), identified among other proteins a consistent increased secretion of Pigment Epithelium Derived Factor (PEDF) associated with the profibrotic phenotype (AJP, 2010). PEDF has been described as produced by retinal-pigmented epithelium and melanocytes, and is a major endogenous inhibitor of intraocular angiogenesis. Here we aimed to validate the increased expression of PEDF in SSc and test the hypothesis that PEDF might play an important role in establishing or perpetuating SSc vasculopathy.
Methods: PEDF expression was investigated in the involved skin and FBs of 4 SSc patients in the early phase of the diffuse form of the disease and 4 healthy controls (HC) by immunohistochemistry (IHC) and real time-PCR analysis. Functional effects of PEDF on angio/vasculogenesis were examined by Matrigel assays. Organotypic co-culture assays were performed seeding HUVECs or microvascular endothelial cells (MVECs), on monolayers of either primary healthy FBs (HC FBs) or SSc FBs or HC FBS silenced for Caveolin-1 (Cav-1). Endothelial cells were evidenced by IHC staining for CD31 in organotypic co-culture assays. Vascular tubule number, length and junctions were identified and analyzed by Angiosys software (TCS CellWorks).
Results: Both Healthy and Scleroderma skin biopsies showed high PEDF protein expression on melanocytes, as expected. Nevertheless, in SSc skin 52% (+/-5.9) of the FBs showed a strong expression of PEDF whereas only 13% (+/-0.68) of the FBs in HC skin were positive (p<0.0006). Double IHC studies indicated that FBs positive for PEDF showed a decreased expression of Cav-1 in both HC and SSc skin. In vitro studies confirmed that SSc FBs showed on average a 5-fold increased in PEDF expression compared to HC FBs (p<0.05). Functional studies confirmed that recombinant PEDF protein had a direct inhibitory effect on vasculogenesis, suppressing both tubule length and number. Consistently, organotypic co-culture assays indicated that SSc FBs or HC FBs silenced for Cav-1 inhibited tubulogenesis both on MVECs or HUVECs, respectively.
Conclusion: PEDF expression is increased in SSc biopsies and SSc FBs. PEDF expression is associated with decreased Cav-1 expression in vivo and it is induced by silencing Cav-1 in vitro. Functionally, PEDF can suppress vasculogenesis both in Matrigel and co-culture assays. This suggests that the decreased expression of Cav-1 observed in SSc FBs may contribute to the vasculopathy of Scleroderma. Further studies unraveling the mechanisms of the antiangiogenic effect of PEDF may shed light in understanding the molecular events linking the profibrotic phenotype and SSc vasculopathy.
F. Del Galdo,
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ACR Meeting Abstracts - https://acrabstracts.org/abstract/pigment-epithelium-derived-factor-secreted-by-activated-fibroblasts-can-contribute-to-impaired-angio-and-vasculogenesis-in-scleroderma/