While cigarette smoke is known to induce eryptosis, the impact of alternative nicotine delivery systems on erythrocyte viability remains largely unknown. In this study, isolated human erythrocytes were exposed to increasing concentrations (40–80%) of heating tobacco product extract (HTPE) or electronic cigarette extract (e-CigE) for 24 h, and eryptotic markers and effectors were assessed by flow cytometry, microscopy and immunoblotting. HTPE induced a concentration-dependent increase in phosphatidylserine exposure and cell shrinkage whereas e-CigE did not affect any eryptotic parameter. HTPE -treated cells exhibited echinocytic morphology and disrupted Band 3–cytoskeleton interactions, accompanied by enhanced phosphorylation of Band 3 and adducin. Calpain inhibition markedly reduced HTPE -induced phosphatidylserine externalization, while caspase-3 inhibition had no effect. HTPE exposure caused a strong, PGE 2 -dependent calcium influx essential for eryptosis, as both calcium chelation and cyclooxygenase inhibition prevented phosphatidylserine exposure. Moreover, HTPE promoted PKCα membrane translocation, and PKC inhibition with staurosporine completely suppressed eryptosis and cytoskeletal phosphorylation. No reactive oxygen species accumulation or Fas/FADD/caspase-8 recruitment was detected. These findings demonstrate that HTPE, but not e-CigE, triggers eryptosis through a calcium- and PKC-dependent mechanism involving PGE 2 -mediated calcium entry, cytoskeletal destabilisation and calpain activity, independently of oxidative stress or caspase activation. Overall, our results provide mechanistic evidence that HTPE exposure compromises erythrocyte integrity and caution against considering tobacco heating products as inherently safe alternatives to conventional cigarettes. • HTP aerosol induces eryptosis in human red blood cells • HTPE triggers eryptosis via a calcium- and PKC-dependent mechanism • HTPE -induced eryptosis involves cytoskeletal proteins remodeling. • Cytoskeletal destabilization involves PKC-dependent phosphorylation of band 3 and adducin • HTP aerosol and cigarette smoke induce eryptosis through distinct signaling pathways.
Massaro et al. (Sun,) studied this question.