IGFBP-4 Proteolysis by PAPP-A in a Primary Culture of Rat Neonatal Cardiomyocytes under Normal and Hypertrophic Conditions.

Cardiovascular diseases (CVD) are among the leading causes of death and disability worldwide. Pregnancy-associated plasma protein-A (PAPP-A) is a matrix metalloprotease localized on the cell surface. One of the substrates that PAPP-A cleaves is the insulin-like growth factor binding protein-4 (IGFBP-4), a member of the family of proteins that bind insulin-like growth factor (IGF). Proteolysis of IGFBP-4 by PAPP-A occurs at a specific site resulting in formation of two proteolytic fragments - N-terminal IGFBP-4 (NT-IGFBP-4) and C-terminal IGFBP-4 (CT-IGFBP-4), and leads to the release of IGF activating various cellular processes including migration, proliferation, and cell growth. Increased levels of the proteolytic IGFBP-4 fragments correlate with the development of CVD complications and increased risk of death in patients with the coronary heart disease, acute coronary syndrome, and heart failure. However, there is no direct evidence that PAPP-A specifically cleaves IGFBP-4 in the cardiac tissue under normal and pathological conditions. In the present study, using a primary culture of rat neonatal cardiomyocytes as a model, we have demonstrated that: 1) proteolysis of IGFBP-4 by PAPP-A occurs in the conditioned medium of cardiomyocytes, 2) PAPP-A-specific IGFBP-4 proteolysis is increased when cardiomyocytes are transformed to a hypertrophic state. Thus, it can be assumed that the enhancement of IGFBP-4 cleavage by PAPP-A and hypertrophic changes in cardiomyocytes accompanying CVD are interrelated, and PAPP-A appears to be one of the activators of the IGF-dependent processes in normal and hypertrophic-state cardiomyocytes.

Detection of Neprilysin-Derived BNP Fragments in the Circulation: Possible Insights for Targeted Neprilysin Inhibition Therapy for Heart Failure.

BACKGROUND: Entresto™ is a new heart failure (HF) therapy that includes the neprilysin (NEP) inhibitor sacubitril. One of the NEP substrates is B-type natriuretic peptide (BNP); its augmentation by NEP inhibition is considered as a possible mechanism for the positive effects of Entresto. We hypothesized that the circulating products of BNP proteolysis by NEP might reflect NEP impact on the metabolism of active BNP. We suggest that NEP-based BNP cleavage at position 17-18 results in BNP ring opening and formation of a novel epitope with C-terminal Arg-17 (BNP-neo17 form). In this study, we use a specific immunoassay to explore BNP-neo17 in a rat model and HF patient plasma. METHODS: We injected BNP into rats, with or without NEP inhibition with sacubitril. BNP-neo17 in plasma samples at different time points was measured with a specific immunoassay with neglectable cross-reactivity to intact forms. BNP-neo17 and total BNP were measured in EDTA plasma samples of HF patients. RESULTS: BNP-neo17 generation in rat circulation was prevented by NEP inhibition. The maximum 13.2-fold difference in BNP-neo17 concentrations with and without sacubitril was observed at 2 min after injection. BNP-neo17 concentrations in 32 HF patient EDTA plasma samples ranged from 0 to 37 pg/mL (median, 5.4; interquartile range, 0-9.1). BNP-neo17/total BNP had no correlation with total BNP concentration (with r = -0.175, P = 0.680) and showed variability among individuals. CONCLUSIONS: BNP-neo17 formation is NEP dependent. Considering that BNP-neo17 is generated from the active form of BNP by NEP, we speculate that BNP-neo17 may reflect both the NEP activity and natriuretic potential and serve for HF therapy guidance.