Abstract
Nicotinic acid adenine dinucleotide phosphate (NAADP) is a newly discovered second messenger that gates two pore channels 1 (TPC1) and 2 (TPC2) to elicit endo-lysosomal (EL) Ca2+ release. NAADP-induced lysosomal Ca2+ release may be amplified by the endoplasmic reticulum (ER) through the Ca2+-induced Ca2+ release (CICR) mechanism. NAADP-induced intracellular Ca2+ signals were shown to modulate a growing number of functions in the cardiovascular system, but their occurrence and role in cardiac mesenchymal stromal cells (C-MSCs) is still unknown. Herein, we found that exogenous delivery of NAADP-AM induced a robust Ca2+ signal that was abolished by disrupting the lysosomal Ca2+ store with Gly-Phe β-naphthylamide, nigericin, and bafilomycin A1, and blocking TPC1 and TPC2, that are both expressed at protein level in C-MSCs. Furthermore, NAADP-induced EL Ca2+ release resulted in the Ca2+-dependent recruitment of ER-embedded InsP3Rs and SOCE activation. Transmission electron microscopy revealed clearly visible membrane contact sites between lysosome and ER membranes, which are predicted to provide the sub-cellular framework for lysosomal Ca2+ to recruit ER-embedded InsP3Rs through CICR. NAADP-induced EL Ca2+ mobilization via EL TPC was found to trigger the intracellular Ca2+ signals whereby Fetal Bovine Serum (FBS) induces C-MSC proliferation. Furthermore, NAADP-evoked Ca2+ release was required to mediate FBS-induced extracellular signal-regulated kinase (ERK), but not Akt, phosphorylation in C-MSCs. These finding support the notion that NAADP-induced TPC activation could be targeted to boost proliferation in C-MSCs and pave the way for future studies assessing whether aberrant NAADP signaling in C-MSCs could be involved in cardiac disorders.
Highlights
Nicotinic acid adenine dinucleotide phosphate (NAADP) has emerged as a the most powerful Ca2+-releasing second messenger in mammalian cells (Galione, 2015; Patel, 2015)
In order to assess whether they are endowed with a NAADPsensitive Ca2+ store, cardiac mesenchymal stromal cells (C-MSCs) were loaded with Fura-2/AM (2 μM), a Ca2+ sensitive fluorophore, as shown elsewhere (Maione et al, 2020a)
NAADP is emerging as a crucial regulator of intracellular Ca2+ signalling and Ca2+-dependent processes in the cardiovascular system (Macgregor et al, 2007; Collins et al, 2011; Fameli et al, 2017; Moccia et al, 2021a)
Summary
Nicotinic acid adenine dinucleotide phosphate (NAADP) has emerged as a the most powerful (already in the nanomolar concentration range) Ca2+-releasing second messenger in mammalian cells (Galione, 2015; Patel, 2015). NAADP elicits an increase in intracellular Ca2+ concentration ([Ca2+]i) by gating a novel family of intracellular Ca2+-releasing channels, known as two-pore channels (TPCs), which present two isoforms in mammals (i.e., TPC1 and TPC2) and mobilize endo-lysosomal (EL) Ca2+ into the cytosol (Patel, 2015; Galione, 2019; Jin et al, 2020). NAADP has been recognized as the trigger of the cellular Ca2+ response to extracellular stimuli in multiple tissues (Galione, 2015; Patel, 2015), including the cardiovascular system (Fameli et al, 2017; Moccia et al, 2021a; Negri et al, 2021b). NAADP gates TPC2 to promote the Ca2+-dependent recruitment of RyR3 and global cytosolic Ca2+ waves in pulmonary artery VSMCs stimulated with either endothelin-1 (Kinnear et al, 2004; Jiang et al, 2013) or angiotensin II (Lee et al, 2015). Aberrant NAADP signalling in cardiac myocytes may result in arrhythmia (Nebel et al, 2013) and ischemia-reperfusion injury (Davidson et al, 2015), whereas it could lead to pulmonary artery hypertension in VSMCs (Jiang et al, 2018; Hu et al, 2021)
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