Effects of 1, 25-(OH)2D3 on airway remodeling and airway epithelial cell apoptosis in a murine model of asthma

Abstract

To investigate the effects of calcitriol on airway remodeling and airway epithelial cell apoptosis in a murine model of bronchial asthma. Twenty-four SPF female Balb/c mice were randomly allocated into three groups according to a random digits table with 8 mice in each group: the control group, the chronic asthma group and the calcitriol intervention group.(1) The chronic asthma group: the mice were sensitized by intraperitoneal injection of Ovalbumin (OVA, 20 μg) and aluminium hydroxide (50 μl) on days 1 and 14.From day 21, the mice were challenged by inhalation of 1% OVA solution (10 ml, 30 min/time, three times a week for consecutively 8 weeks). (2) The calcitriol intervention group: the mice were sensitized and challenged as above, and were given calcitriol 100 ng through intraperitoneal injection 30 min before every inhalation.(3) The control group: the mice were sensitized and challenged by saline instead of OVA.The mice were sacrificed 24 hours after the last challenge, and the left lung were removed, fixed with paraformaldehyde, embedded with paraffin and sectioned.HE staining, Alcian blue and Periodic acid Schiff (AB-PAS) staining, Masson staining, and α-smooth muscle actin (α-SMA) immunohistochemistry staining were conducted.Bronchial basement membrane perimeter (Pbm), total bronchial wall area and positive areas of AB-PAS staining, Masson staining, and α-SMA staining were determined with image analysis software.The results were standardized with the basement membrane perimeter.Paraffin sections of mice lung tissue were detected with terminal transferase deoxyuridine triphosphate (dUTP) nick end labeling enzyme mediated method (TUNEL) for apoptosis of airway epithelial cells and with immunohistochemistry staining for expression of B cell lymphoma/lewkmia-2 (Bcl-2) in the airway epithelium. The mice in the chronic asthma group and calcitriol intervention group showed characteristic airway inflammation and airway remodeling of asthma.The ratios between the total bronchial wall area and positive areas of AB-PAS staining, Masson staining and α-SMA staining and the basement membrane perimeter in the calcitriol intervention group were (14.12±2.13), (3.72±0.57), (4.31±0.65) and (3.27±0.46) μm2/μm, respectively, all of them were significantly lower than (19.24±1.70), (5.23±0.90), (7.63±1.55) and (5.40±0.69) μm2/μm in the chronic asthma group and higher than (7.79±1.01), (0.05±0.03), (1.37±0.25) and (1.40±0.24) μm2/μm in the control group (all P<0.01). The airway epithelial cell apoptosis index in the calcitriol intervention group was significantly lower than that in the chronic asthma group and higher than the control group [(14.89±1.75)% vs (29.73±5.74)% and (0.45±0.38)%, both P<0.01]. The relative expression of Bcl-2 in the calcitriol intervention group was significantly higher than that in the chronic asthma group and lower than the control group (0.114±0.009 vs 0.091±0.023 and 0.160±0.021, both P<0.05). Calcitriol attenuates airway remodeling and reduces the apoptosis of airway epithelial cells in a murine model of chronic asthma.The mechanism of calcitriol in reducing apoptosis of airway epithelial cells is by regulation of expression of the important molecule Bcl-2 protein in mitochondrial apoptotic pathway.