Hypoxia-induced mitogenic factor/FIZZ1 induces intracellular calcium release through the PLC-IP₃ pathway

Hypoxia-induced mitogenic factor (HIMF), also known as "found in inflammatory zone 1" (FIZZ1) or resistin-like molecule-α (RELMα), is a profound vasoconstrictor of the pulmonary circulation and a strong mitogenic factor in pulmonary vascular smooth muscle. To further understand the mechanism of these contractile and mitogenic responses, we examined the effect of HIMF on intracellular Ca²⁺ in human pulmonary artery smooth muscle cells (SMC). Ca²⁺ imaging in fluo 4-loaded human pulmonary artery SMC revealed that recombinant murine HIMF increased intracellular Ca²⁺ concentration ([Ca²⁺]_i) in a sustained and oscillatory manner. This increase occurred independent of extracellular Ca²⁺ influx. Pretreatment of human pulmonary artery SMC with U-73122, a specific inhibitor of phosphatidylinositolphospholipase C (PLC) completely prevented the HIMF-induced Ca²⁺ signal. The [Ca ²⁺]_i increase was also abolished by pretreatment with 2-aminoethoxydiphenyl borate (2-APB), an inositol 1,4,5-trisphosphate (IP ₃) receptor antagonist. Ryanodine pretreatment did not affect initiation of [Ca²⁺]_i activation or internal release but reduced [Ca²⁺]_i at the plateau phase. Pretreatment with the Gα_ispecific inhibitor pertussis toxin and the Gα_s-specific inhibitor NF-449 did not block the Ca²⁺ signal. Knockdown of Gα_q/11 expression did not prevent Ca ²⁺ release, but the pattern of Ca²⁺ release changed from the sustained oscillatory transients with prolonged plateau to a series of short [Ca²⁺]_i transients that return to baseline. However, pretreatment with the tyrosine kinase inhibitor genistein completely inhibited the internal Ca²⁺ release. These results demonstrate that HIMF can stimulate intracellular Ca²⁺ release in human pulmonary artery SMC through the PLC signaling pathway in an IP₃- and tyrosine phosphorylation-dependent manner and that Gα_q/11 protein-coupled receptor and ryanodine receptor contribute to the increase of [Ca²⁺]_i.