Original Article
Construction of a cell model of β2-adrenoceptor downregulation
Abstract
Objective: To establish a cell model of β2-adrenergic receptor (β2AR) downregulation of murine airway smooth muscle induced by salbutamol to elucidate the molecular and biological mechanisms of β2AR downregulation.
Methods: Airway smooth muscle cells (ASMCs) derived from Balb/c mice were primary cultured. Passage 2-5 cells were characterized by cell morphology and indirect immunofluorescence. More than 95% pure cells at passage 3 or 4 were randomly divided into two groups: control and salbutamol-treated groups. β2AR mRNA and protein expression levels were then detected by RT-PCR and western blot analyses.
Results: Primary cultured cells demonstrated a typical “peak and valley”-like growth characteristic. Smooth muscle α-actin filaments paralleled the cell longitudinal axis in the cytoplasm. The origin of the ASMCs was validated and consistent with their morphology and biological characteristics. β2AR mRNA expression in the salbutamol-treated group was lower than that in the control group (P<0.05), and β2AR protein expression was also markedly lower than that in the control group (P<0.05).
Conclusions: We successfully established a cell model of β2AR downregulation in ASMCs, which may provide the foundation for further study of the mechanism of β2AR downregulation in asthmatic patients.
Methods: Airway smooth muscle cells (ASMCs) derived from Balb/c mice were primary cultured. Passage 2-5 cells were characterized by cell morphology and indirect immunofluorescence. More than 95% pure cells at passage 3 or 4 were randomly divided into two groups: control and salbutamol-treated groups. β2AR mRNA and protein expression levels were then detected by RT-PCR and western blot analyses.
Results: Primary cultured cells demonstrated a typical “peak and valley”-like growth characteristic. Smooth muscle α-actin filaments paralleled the cell longitudinal axis in the cytoplasm. The origin of the ASMCs was validated and consistent with their morphology and biological characteristics. β2AR mRNA expression in the salbutamol-treated group was lower than that in the control group (P<0.05), and β2AR protein expression was also markedly lower than that in the control group (P<0.05).
Conclusions: We successfully established a cell model of β2AR downregulation in ASMCs, which may provide the foundation for further study of the mechanism of β2AR downregulation in asthmatic patients.
