Bortezomib inhibits hypoxia-induced increase of Orai-1 expression in pulmonary arterial smooth muscle cell

Abstract

Objective: In this study, a primary culture system for the rat distal pulmonary arterial smooth muscle cell (PASMC) was established to observe the effect of Bortezomib a treatment on the basal intracellular calcium concentration ([Ca(2+) ](i)), store operated calcium entry (SOCE) and Orai-1 expression in rat PASMC. Methods: We employed the primary culture method for the rat distal PASMC including the enzymatically dissociation of PASMC from the freshly isolated distal pulmonary artery and the culture of PASMC. The In Cyte system was used to measure the basal [Ca(2+) ](i) and SOCE after substantial treatment.Orai-1 protein expression in rat pulmonary artery smooth muscle was detected by Western blot. Results: Compared with Hypoxia group, the basal [Ca(2+) ](i) were significantly reduced in Hypoxia+ BTZ group(P<0.01). The basal [Ca(2+) ](i) A340/A380 ratio of Normoxia group was(1.07±0.02). The basal [Ca(2+) ](i) of Hypoxia group was(1.49±0.03); The Hypoxia+ BTZ group was(1.17±0.03). Compared with Hypoxia group, the store operated calcium entry were significantly reduced in Hypoxia+ BTZ group(P<0.01). The SOCE A340/A380 ratio of Normoxia group was(0.56±0.02). The SOCE of Hypoxia group was(0.84±0.02); The Hypoxia+ BTZ group was(0.66±0.02). The level of Orail-1 protein in pulmonary artery smooth muscle of Hypoxia group was (181.5±12.7)% higher than control group which was(100±0)%, (P<0.05). In the Hypoxia+ BTZ group Orai-1 protein expression was recovered(146.7±15.1)%, (P<0.05). Conclusion: Bortezomib inhibit chronically hypoxic enhancement of Orail-1 protein expression, basal [Ca(2+) ](i) and SOCE in rat distal pulmonary arterial smooth muscle cells.