Calcium-sensing receptor signalling and neutrophil dysfunction: implications in critical illness

Background: Critically ill (CI) patients often develop neutrophil dysfunction that contributes to increased susceptibility to infections such as hospital-acquired pneumonia. The upregulation of calcium-sensing receptor (CaSR) can drive chronic inflammatory lung disease, but whether CaSR contributes to the neutrophil dysfunction in CI patients is yet unknown. Objective: To investigate signalling of CaSR in neutrophils from healthy and CI donors. Methods and Results: Human circulating blood neutrophils were obtained from healthy volunteers and CI patients. Immunofluorescence confirmed CaSR expression by neutrophils. Treatment of neutrophils with spermine, a polyamine and CaSR agonist involved in infection and inflammation, resulted in improved bacterial clearance. However, spermine led to concentration-dependent neutrophils aggregation and death with EC₅₀=314 μM (healthy) and EC₅₀=156 μM (CI patients). Moreover, spermine induced increases in intracellular calcium concentrations (Oregon Green BAPTA-1 AM fluorescence), activation of transmembrane currents (patch-clamp) and reactive oxygen species (ROS; DCFDA kit) production by neutrophils in a concentration-dependent manner. A CaSR inhibitor (NPS2143, 1 µM) improved neutrophil viability, reversed polyamine-induced enhancement of bacterial killing and ROS production. Conclusions: Spermine activated CaSR in circulating neutrophils to enhance bacterial clearance by inducing increased intracellular calcium, ROS production, and cell death, suggesting therapeutic avenues for allosteric CaSR modulators to regulate neutrophil functions.