Airway smooth muscle STIM1 and Orai1 are upregulated in asthmatic mice and mediate PDGF-activated SOCE, CRAC currents, proliferation, and migration

Abstract

Airway smooth muscle cell (ASMC) remodeling contributes to the structural changes in the airways that are central to the clinical manifestations of asthma. Ca2+ signals play an important role in ASMC remodeling through control of ASMC migration and hypertrophy/proliferation. Upregulation of STIM1 and Orai1 proteins, the molecular components of the store-operated Ca2+ entry (SOCE) pathway, has recently emerged as an important mediator of vascular remodeling. However, the potential upregulation of STIM1 and Orai1 in asthmatic airways remains unknown. An important smooth muscle migratory agonist with major contributions to ASMC remodeling is the platelet-derived growth factor (PDGF). Nevertheless, the Ca2+ entry route activated by PDGF in ASMC remains elusive. Here, we show that STIM1 and Orai1 protein levels are greatly upregulated in ASMC isolated from ovalbumin-challenged asthmatic mice, compared to control mice. Furthermore, we show that PDGF activates a Ca2+ entry pathway in rat primary ASMC that is pharmacologically reminiscent of SOCE. Molecular knockdown of STIM1 and Orai1 proteins inhibited PDGF-activated Ca2+ entry in these cells. Whole-cell patch clamp recordings revealed the activation of Ca2+ release-activated Ca2+ (CRAC) current by PDGF in ASMC. These CRAC currents were abrogated upon either STIM1 or Orai1 knockdown. We show that either STIM1 or Orai1 knockdown significantly inhibited ASMC proliferation and chemotactic migration in response to PDGF. These results implicate STIM1 and Orai1 in PDGF-induced ASMC proliferation and migration and suggest the potential use of STIM1 and Orai1 as targets for ASMC remodeling during asthma.

Airway smooth muscle in airway reactivity and remodeling: what have we learned?

“…In PC12 cells, changes in extracellular pH can influence store-operated Ca 2ϩ influx that is dependent on the SR Ca 2ϩ protein stromal interacting molecule 1 (STIM1) (299). Given the important role of STIM1/Orai1 in human ASM (80,137,225,264,292,357), it is possible that OGR1 could modulate store-operated Ca 2ϩ entry (SOCE) by enhancing STIM1 interactions with Orai1 or other channels that contribute to Ca 2ϩ influx, particularly canonical transient receptor potentials (TRPCs) (80,338). Furthermore, the limited data in human ASM suggest that OGR1 can further result in p42/44 MAPK phosphorylation (269) and IL-6 production (122) and therefore have more complex effects in the setting of disease.…”

Mechanisms of BDNF regulation in asthmatic airway smooth muscle

“…As elevated [Ca 2ϩ ] i is a hallmark of asthmatic ASM, it is possible that higher [Ca 2ϩ ] i also enhances BDNF expression and/or secretion, but this aspect has not been previously explored. Further, elevated [Ca 2ϩ ] i in ASM involves Ca 2ϩ regulatory proteins such as transient receptor potential (TRPC) and Orai1 channels, which have been implicated in asthma (40,48), and initial studies from our group have shown that TRPC3 channel activation is important for ASM BDNF secretion (44). Along similar lines, in fetal ASM cells, cAMP and Epac2 appear to be important in driving BDNF expression and secretion (43).…”

Molecular pharmacology of store-operated CRAC channels

“…18,19 Ca 2+ influx through CRAC channels is essential to activate the transcription factor NFAT that regulates the expression of a variety of cytokines essential for T-cell function, a pathway that is defective in the SCID patients. 20 Since then, the importance of this pathway has been highlighted in a range of functions in many different cell types including various subsets of T cells, 21 B cells, 22,23 mast cells, 24,25 vascular smooth muscle, [26][27][28] endothelium, 29 platelets, 30,31 melanocytes, 32 and skeletal muscle. [33][34][35] Despite the biophysical characterization of CRAC channels, the molecular mechanism that linked store-depletion to channel activation as well as the molecular identity of the channel itself REVIEW REVIEW resting ER Ca 2+ concentration.…”