Plasminogen activator inhibitor-1 reduces cardiac fibrosis and promotes M2 macrophage polarization in inflammatory cardiomyopathy

Fig. 1. Cardiac PAI-1 expression is elevated in inflammatory-dilated cardiomyopathy with increasing lymphocytic infiltration. a Quantification of PAI-1 levels in endomyocardial biopsies of patients diagnosed for dilated cardiomyopathy (DCM) or inflammatory dilated cardiomyopathy (DCMi). DCMi patients were clustered into low-grade inflammation (DCMi-low, CD3+ lymphocytes = 14–30 cells/mm2) and high-grade inflammation (DCMi-high, CD3+ lymphocytes > 30 cells/mm2). Data are represented as mean ± SEM (n = 27–88 patients per group). ***p ≤ 0.001 as indicated. b Representative photomicrographs immunostained with anti PAI-1 antibody. Magnification: ×200. Scale bar: 100 µm

Fig. 2. Increased PAI-1 level in DCMi-high patients attenuates fibrosis by inhibiting the activation of cardiac myofibroblasts. a Representative photomicrographs from cardiac sections of patients with DCM, DCMi low-grade inflammation (DCMi-low) and DCMi high-grade inflammation (DCMi-high) stained with Trichrome stain AB solution. Fibrous connective tissue is identified in blue. Magnification: ×400. Scale bar: 20 µm. b Quantification of fibrotic area (n = 6–9 patients per group). c Representative photomicrographs from cardiac sections immunostained with anti α-smooth muscle actin (α-SMA) antibody. α-SMA+ myofibroblasts are identified in brown. Magnification: ×600. Scale bar: 40 µm. d Quantification of α-SMA+ cells (n = 6–12 patients per group). Data are represented as mean ± SEM. *p ≤ 0.05 vs. DCM and DCMi-low or as indicated

Fig. 3. Increased expression of TGF-β in inflammatory dilative cardiomyopathy is attenuated by elevated PAI-1 levels. a, b Relative TGF-β mRNA (a) and protein (b) levels in endomyocardial biopsies of patients diagnosed for dilative cardiomyopathy (DCM) or inflammatory dilative cardiomyopathy (DCMi). DCMi group were clustered into low-grade inflammation (DCMi-low, CD3 + lymphocytes = 14–30/mm2) and high-grade inflammation (DCMi-high, CD3 + lymphocytes > 30/mm2). Data are represented as mean ± SEM (n = 7–12 patients per group). *p ≤ 0.05 as indicated. c Representative photomicrographs immunostained with anti-TGF-β antibody. Magnification: ×400. Scale bar: 20 µm

Fig. 4. Cardiac PAI-1 expression is associated with increased prevalence of M2 macrophages. a, c Representative photomicrographs from cardiac sections of patients with DCM, DCMi low-grade inflammation (DCMi-low) and DCMi-high grade inflammation (DCMi-high), double immunostained against CD14/CD68 (green/red) and CD16/CD68 (green/red) (a), or CD86/CD68 (green/red) and CD163/CD68 (green/red) (c), respectively. Double positive cells are depicted in yellow and indicate presence of M1 (CD68+ CD14+ and CD68+ CD86+) or M2 (CD68+ CD16+ and CD68+ CD163+) macrophages. Magnification: ×400. Scale bar: 50 µm. b, d M1 to M2 ratio was calculated from the total number of M1 (CD68+ CD14+ (b) or CD68+ CD86+ (d) double positive cells) and M2 (CD68+ CD16+ (b) or CD68+ CD163+ (d) double positive cells) macrophages (n = 6–11 patients per group). Data are represented as mean ± SEM. *p ≤ 0.05, **p ≤ 0.01