Background/Purpose:

SSc patients frequently suffer from primary cardiac involvement. The main histological feature is fibrosis, but the mechanisms responsible for the heart failure remain unclear. It is known that cellular progenitors, including stromal cells, differentiate into myofibroblasts and autophagy may favor fibrosis through enhanced differentiation of cardiac stromal cells. Here we describe the role of Fos-related antigen 2 (fra-2)/autophagy crosstalk in TGFβ-dependent myocardial fibrosis in SSc.

Methods:

Immunohistochemistry (IHC) and immunofluorescence (IF) were performed on endomyocardial biopsies (EMBs) from SSc patients and hearts from fra-2^tg SSc mouse model and WT mice. Myocardial stromal Ter119^–CD45^–CD31^– (Lin^–) gp38⁺ stromal cells were sorted for in vitro culture. The cellular phenotype was assessed by qPCR, IF, stress fiber staining, SIRCOL and contraction assay on sorted cells. The antisense oligonucleotide GapmeR was used to knock-down fra-2.

Results:

Fibrosis, collagen deposition and αSMA⁺ myofibroblasts were increased in the myocardium of SSc patients (n=10). Fra-2 and autophagy markers such as LC3B, Beclin-1 and Atg5 were expressed in fibrotic cardiac tissues. In parallel, the myocardium of fra-2^tg mice showed higher expression of the profibrotic markers αSMA, vimentin and collagen I compared to WT mice (n=5), as well the expression of LC3B and Beclin-1 in fibrotic regions. Among cardiac stromal cells (Lin^–), the frequency of gp38⁺ cells was significantly higher in fra2^tg myocardium compared to WT mice (n=8, p=0.02). Lin^–/gp38⁺ cells co-expressed αSMA, vimentin and collagen I together with LC3B and Beclin-1 (n=3). Following TGFβ stimulation, WT Lin^–/gp38⁺ cells entered fibroblast-to-myofibroblast transition characterized by increased mRNA and protein levels of αSMA, collagen I, fibronectin (n=3-6), formation of αSMA⁺ fibers and stress fibers (n=3), increased cell proliferation (n=5; p=0.04), contraction capability (n=5; p<0.05) and collagen secretion (n=5; p=0.04). TGFβ stimulation of WT Lin^–/gp38⁺ cells induced the expression of LC3B, Beclin-1 and Atg5 at mRNA and protein level (n=3-5). Accordingly, TGFβ inhibition caused the downregulation of these markers (n=3). In contrast to WT cells, fra-2^tg Lin^–/gp38⁺ cells showed the presence of αSMA⁺ fibers, stress fibers and an increased contraction capability even without TGFβ stimulation.
Finally, fra-2 silencing resulted in decreased Lin^–/gp38⁺ cell differentiation and autophagy activity as following: levels of αSMA and collagen I (n=5; p=0.007), secreted collagens (n=5; p<0.05) and αSMA fibers (n=3) were significantly downregulated in addition to a significant decrease of mRNA and protein expression of LC3B, Beclin-1 and Atg5 (n=3).

Conclusion:

The TGFβ/fra-2 axis regulates the autophagy process, leading in turn to stromal-to-myofibroblast transition. Therefore, targeting autophagy might ameliorate the fibrotic process during heart remodeling in SSc.

« Back to 2017 ACR/ARHP Annual Meeting

ACR Meeting Abstracts - https://acrabstracts.org/abstract/cardiac-remodeling-in-systemic-sclerosis-tgf-%ce%b2fra2-dependent-autophagy-as-a-novel-target-for-heart-fibrosis/