RESUMEN
Airway smooth muscle (ASM) regulation of airway structure and contractility is critical in fetal/neonatal physiology in health and disease. Fetal lungs experience higher Ca2+ environment that may impact extracellular Ca2+ ([Ca2+ ]o ) sensing receptor (CaSR). Well-known in the parathyroid gland, CaSR is also expressed in late embryonic lung mesenchyme. Using cells from 18-22 week human fetal lungs, we tested the hypothesis that CaSR regulates intracellular Ca2+ ([Ca2+ ]i ) in fetal ASM (fASM). Compared with adult ASM, CaSR expression was higher in fASM, while fluorescence Ca2+ imaging showed that [Ca2+ ]i was more sensitive to altered [Ca2+ ]o . The fASM [Ca2+ ]i responses to histamine were also more sensitive to [Ca2+ ]o (0-2 mM) compared with an adult, enhanced by calcimimetic R568 but blunted by calcilytic NPS2143. [Ca2+ ]i was enhanced by endogenous CaSR agonist spermine (again higher sensitivity compared with adult). Inhibition of phospholipase C (U73122; siRNA) or inositol 1,4,5-triphosphate receptor (Xestospongin C) blunted [Ca2+ ]o sensitivity and R568 effects. NPS2143 potentiated U73122 effects. Store-operated Ca2+ entry was potentiated by R568. Traction force microscopy showed responsiveness of fASM cellular contractility to [Ca2+ ]o and NPS2143. Separately, fASM proliferation showed sensitivity to [Ca2+ ]o and NPS2143. These results demonstrate functional CaSR in developing ASM that modulates airway contractility and proliferation.
Asunto(s)
Señalización del Calcio/genética , Pulmón/crecimiento & desarrollo , Mioblastos/metabolismo , Receptores Sensibles al Calcio/genética , Calcio/metabolismo , Proliferación Celular/efectos de los fármacos , Proliferación Celular/genética , Feto , Humanos , Pulmón/embriología , Pulmón/metabolismo , Compuestos Macrocíclicos/farmacología , Músculo Liso/metabolismo , Naftalenos/farmacología , Oxazoles/farmacología , Fosfolipasas de Tipo C/antagonistas & inhibidores , Fosfolipasas de Tipo C/genéticaRESUMEN
Supplemental O2 (hyperoxia), necessary for maintenance of oxygenation in premature infants, contributes to neonatal and pediatric airway diseases including asthma. Airway smooth muscle (ASM) is a key resident cell type, responding to hyperoxia with increased contractility and remodeling [proliferation, extracellular matrix (ECM) production], making the mechanisms underlying hyperoxia effects on ASM significant. Recognizing that fetal lungs experience a higher extracellular Ca2+ ([Ca2+]o) environment, we previously reported that the calcium sensing receptor (CaSR) is expressed and functional in human fetal ASM (fASM). In this study, using fASM cells from 18 to 22 week human fetal lungs, we tested the hypothesis that CaSR contributes to hyperoxia effects on developing ASM. Moderate hyperoxia (50% O2) increased fASM CaSR expression. Fluorescence [Ca2+]i imaging showed hyperoxia increased [Ca2+]i responses to histamine that was more sensitive to altered [Ca2+]o, and promoted IP3 induced intracellular Ca2+ release and store-operated Ca2+ entry: effects blunted by the calcilytic NPS2143. Hyperoxia did not significantly increase mitochondrial calcium which was regulated by CaSR irrespective of oxygen levels. Separately, fASM cell proliferation and ECM deposition (collagens but not fibronectin) showed sensitivity to [Ca2+]o that was enhanced by hyperoxia, but blunted by NPS2143. Effects of hyperoxia involved p42/44 ERK via CaSR and HIF1α. These results demonstrate functional CaSR in developing ASM that contributes to hyperoxia-induced contractility and remodeling that may be relevant to perinatal airway disease.