Abstract Background During atherosclerosis, S100A4 was demonstrated to play crucial roles in smooth muscle cells (SMCs) phenotypic transition, to which it confers pro-inflammatory properties. Using a S100A4 neutralizing antibody in an ApoE deficient mouse model, we previously showed a significant reduction in plaque burden. Nevertheless, this approach did not allow us to discriminate between the effect of different S100A4-expressing cells, or to establish a direct link with disease development. Purpose Our research aims at understanding the roles of S100A4 in atherosclerosis, and how various S100A4-expressing cells impact the disease. Methods First, we engineered a double-knockout mouse model (ApoE-/-; S100A4 -/-) fed on a 12-week high cholesterol diet to address S100A4 functions in vivo. Male and female mice were selected for the study and body weight was measured at three-week intervals over the course of the regimen. At the study endpoint, complete lipid measurements were established upon blood samples collection. Plaque burden was quantified by en face Oil Red O staining of the whole aorta, and by immunofluorescence for CD68, α-SMA and S100A4 on six 5 µm-OCT aortic root sections per animal to assess both plaque volume and cellular composition. Second, to study the cellular crosstalk between S100A4-mediated SMCs and macrophages, we utilized a transwell co-culture system. Human aortic SMCs, placed on the upper compartment, were treated with oligomeric S100A4, PDGF-BB, or combinations of both, while monocytes isolated from human buffy coats, were cultivated on the lower compartment. Polarization of macrophages were then detected by flow cytometry analysis, after staining for CD14, CD16, HLA-DR, CD86, CD163 and CD206. Results We first showed that despite no significant changes in lipid profiles, double-knockout mice gained significantly more weight on average compared to control mice (ApoE-/-; S100A4-/- 6.446 g, n=29 vs ApoE-/-; S100A4 +/+ 4.289 g, n=28, p=0.0001). Plaque burden was significantly diminished along the aorta in double-knockout mice when compared to controls (10.450%, n=23 vs 15.219%, n=21, respectively, p=0.003). At the level of the aortic roots, we observed a prominent S100A4 sex-dependent effect in double-knockout female mice. Although these mice displayed larger atherosclerotic lesions (172.6 x103 µm, n=10, vs 107,7x103 µm, n=17, p=0.05), they exhibited smaller necrotic cores (16,30% vs 24.06%, p=0.05), suggesting a role of S100A4 in plaque stabilization. In vitro studies proposed that S100A4-treated SMCs might modulate macrophage phenotypes. Indeed, our preliminary results (n= 3 donors) showed an upregulation of CD163 expression, suggesting a shift towards an anti-inflammatory profile. Conclusions Our research reveals that loss of S100A4 in ApoE−/− mice markedly impacts both the size and cellular composition of atherosclerotic plaques, positioning S100A4 as a critical regulator of vascular inflammation.For image description, please refer to the figure legend and surrounding text.
Sarhdaoui et al. (Fri,) studied this question.