Galectin-3 binding protein is upregulated in heart failure with preserved ejection fraction and is modulated by endothelial nitric oxide synthase deficiency
R Madonna, M C Cufaro, S Gagliardi, R Morganti, F Di Ferdinando, A Marzoppi, A Avagimyan, S Ghelardoni, P Del BoccioAbstract
Abstract
Heart failure with preserved ejection fraction (HFpEF) is a cardiometabolic syndrome characterized by systemic inflammation and endothelial dysfunction, in which impaired endothelial nitric oxide synthase (eNOS) signaling plays a central role. This study aimed to identify HFpEF-associated circulating proteins and to explore their mechanistic link with endothelial dysfunction under metabolic stress. Given the inflammatory and metabolic nature of HFpEF, we set up an in vitro model of insulin resistance and hyperinsulinemia in murine aortic endothelial cells, wild-type and knock-out for eNOS with the aim of examining the effect of metabolic stress on LGALS3BP expression and whether eNOS deletion impacts its expression.
Methods
A total of 109 HFpEF patients and 50 control subjects underwent clinical, laboratory, and echocardiographic assessment. Wild-type (WT) and eNOS knockout (KO) murine aortic endothelial cells (MAECs) were treated for 24 h with basal glucose (5.5 mM), high glucose (HG, 30.5 mM ), high insulin (HI, 10 nM), and the combination of high glucose and high insulin (HG+HI). Protein lysates were collected and analyzed by Western blot.
Results
HFpEF patients showed a high burden of cardiometabolic comorbidities and significantly increased NT-proBNP (2851.2 ± 1565.5 vs 156.0 ± 85.2 pg/mL) and C-reactive protein levels (2.9 ± 4.5 vs 0.31 ± 0.33 mg/dL). Echocardiography revealed elevated filling pressures (E/e′ 16.5 ± 3.8 vs 7.0 ± 1.9), a higher prevalence of high-probability pulmonary hypertension, and impaired right ventricular–pulmonary artery coupling. Proteomic profiling identified galectin-3–binding protein (LGALS3BP) as selectively increased in plasma of HFpEF patients, with significantly higher circulating levels compared with controls (8.65 ± 0.66 vs 2.36 ± 0.26 ng/mL, p < 0.001). Moreover, proteomics analysis revealed that LGALS3BP was indirectly involved in the significant activation of nitric oxide synthase 2 (NOS2) as upstream regulator in HFpEF patients (Figure 1 A-D). In vitro, metabolic stress induced LGALS3BP upregulation and markers for endothelial disfunction (decreased p-eNOS/eNOS expression), insulin-resistance (decreased pAKT/AKT expression) and endothelial activation (increased VCAM-1 expression) in murine endothelial cells, with a more pronounced effect in eNOS⁻/⁻ cells by confirming proteomic signature (Figure 1 E). Notably, eNOS deficiency was associated with the expression of a truncated LGALS3BP isoform (Figure 1F), accompanied by increased markers of endothelial senescence and autophagy (Figure 1G).
Conclusions
LGALS3BP is selectively elevated in HFpEF and mechanistically linked to eNOS-dependent endothelial dysfunction, identifying a novel pathway connecting metabolic stress, endothelial maladaptation, and HFpEF pathophysiology.Figure 1