Human Bronchial Smooth Muscle Cells

The present study aimed to determine the expression of microRNA-146a (miR-146a) in the plasma of children with asthma, and to investigate the effect of miR-146a on the pr... More

The present study aimed to determine the expression of microRNA-146a (miR-146a) in the plasma of children with asthma, and to investigate the effect of miR-146a on the proliferation and apoptosis of bronchial smooth muscle cells (BSMCs). Reverse transcription-quantitative polymerase chain reaction was used to determine the expression levels of miR-146a mimics and its inhibitor. A Cell Counting kit-8 assay was performed to examine the proliferation of BSMCs. Caspase-3/7 activity was determined using a Caspase-Glo 3/7 kit. To measure the expression levels of proteins associated with apoptosis, western blotting was performed. The target gene of miR-146a was identified using a dual-luciferase reporter assay. The plasma levels of miR-146a in children with asthma were significantly higher compared with those in healthy children. Enhanced miR-146a expression inhibited the proliferation of BSMCs. BSMC apoptosis was promoted by miR-146a. The mechanism underlying the miR-146a-induced promotion of BSMC apoptosis may be its direct targeting of epidermal growth factor receptor (EGFR), which affects downstream signaling pathways. In conclusion, miR-146a expression in asthma inhibits the proliferation and promotes the apoptosis of BSMCs by direct targeting of EGFR. Keywords: microRNA-146a, bronchial smooth muscle cell, proliferation, apoptosis Less

Background: Abnormal proliferation, apoptosis, migration and contraction of airway smooth muscle (ASM) cells in airway remodeling in asthma are basically excessive repair... More

Background: Abnormal proliferation, apoptosis, migration and contraction of airway smooth muscle (ASM) cells in airway remodeling in asthma are basically excessive repair responses to a network of inflammatory mediators such as PDGF, but the mechanisms of such responses remain unclear. Nogo-B, a member of the reticulum family 4(RTN4), is known to play a key role in arteriogenesis and tissue repair. Further studies are needed to elucidate the role of Nogo-B in airway smooth muscle abnormalities. Methods: A mouse model of chronic asthma was established by repeated OVA inhalation and subjected to Nogo-B expression analysis using immunohistochemistry and Western Blotting. Then, primary human bronchial smooth muscle cells (HBSMCs) were cultured in vitro and a siRNA interference was performed to knockdown the expression of Nogo-B in the cells. The effects of Nogo-B inhibition on PDGF-induced HBSMCs proliferation, migration and contraction were evaluated. Finally, a proteomic analysis was conducted to unveil the underlying mechanisms responsible for the function of Nogo-B. Results: Total Nogo-B expression was approximately 3.08-fold lower in chronic asthmatic mice compared to naïve mice, which was obvious in the smooth muscle layer of the airways. Interference of Nogo-B expression by siRNA resulted nearly 96% reduction in mRNA in cultured HBSMCs. In addition, knockdown of Nogo-B using specific siRNA significantly decreased PDGF-induced migration of HBSMCs by 2.3-fold, and increased the cellular contraction by 16% compared to negative controls, but had limited effects on PDGF-induced proliferation. Furthermore, using proteomic analysis, we demonstrate that the expression of actin related protein 2/3 complex subunit 5 (ARPC 2/3) decreased and, myosin regulatory light chain 9 isoform a (MYL-9) increased after Nogo-B knockdown. Conclusions: These data define a novel role for Nogo-B in airway remodeling in chronic asthma. Endogenous Nogo-B, which may exert its effects through ARPC 2/3 and MYL-9, is necessary for the migration and contraction of airway smooth muscle cells. Less

IL-13 is a central mediator of allergic inflammation and secreted by Th2 and bronchial smooth muscle cells (BSMC). However, little is known about the regulation of IL-13 ... More

IL-13 is a central mediator of allergic inflammation and secreted by Th2 and bronchial smooth muscle cells (BSMC). However, little is known about the regulation of IL-13 secretion. To address it, a cDNA library of BSMC was screened for the proteins interacted with IL-13 by yeast two-hybridization. Besides IL-13 receptors, human sulfatase modifying factor 2 (SUMF2) was interacted with IL-13. Furthermore, SUMF2 and IL-13 were co-immunoprecipitated from BSMC, which was independent of IL-13 glycosylation. Interestingly, high levels of SUMF2 were expressed by BSMC, accompanied by significantly higher levels of intracellular IL-13, but lower levels of IL-13 secretion from BSMC. In contrast, little of SUMF2 was detected in lymphocytes, accompanied by lower levels of intracellular IL-13, but significantly higher levels of 12 kDa form of IL-13 secretion. Moreover, knockdown of SUMF2 expression by transfection with SUMF2-specific siRNA did not alter IL-13 mRNA transcription, but significantly reduced intracellular IL-13 levels, associated with increased levels of IL-13 secretion from BSMC. While induction of transient SUMF2 expression in lymphocytes failed to modulate IL-13 mRNA transcription. It significantly increased the contents of 12 kDa form of intracellular IL-13, accompanied by significantly reduced levels of IL-13 in their supernatants. In addition, blockage of N-glycosylation by treatment with tunicamycin eliminated 17 kDa form of intracellular IL-13, but failed to promote IL-13 secretion in BSMC. Collectively, our novel data clearly indicated that SUMF2 interacted with IL-13 and inhibited IL-13 secretion in BSMC and lymphocytes, which was independent of IL-13 glycosylation. Less

Background and objectives: In asthma, airway smooth muscle cell (ASMC) hyperplasia plays an important role in airway remodelling. Increased expression of matrix metallopr... More

Background and objectives: In asthma, airway smooth muscle cell (ASMC) hyperplasia plays an important role in airway remodelling. Increased expression of matrix metalloproteinases-9 (MMP-9), a disintegrin and metalloprotease 33 (ADAM33) in ASMCs are also relevant to asthmatic airway remodelling. 1,25-dihydroxyvitamin D(3) (1,25-(OH)(2)D(3)) has potent antiproliferative properties in vitro in various cell types; however, its role in ASMCs is not well understood. This study investigated the effect of 1,25-(OH)(2)D(3) on passively sensitized human bronchial (airway) smooth muscle cell (HASMC) proliferation and MMP-9 and ADAM33 expressions. Methods: The effect of 1,25-(OH)(2)D(3) on cell proliferation was examined by 3-(4,5-dimethylthiazole-2-yl)-2,5-diphenyltetrazolium-bromide colorimetry assay; cell cycle analysis by flow cytometry; and immunocytochemical staining for proliferating cell nuclear antigen (PCNA). The expression of MMP-9 and ADAM33 in HASMCs was investigated by real-time quantitative PCR and Western Blot analysis. Results: 1,25-(OH)(2)D(3) effectively suppressed passively sensitized HASMC proliferation, proliferating cell nuclear antigen expression and G(1)/S transition in HASMCs passively sensitized with asthmatic serum. Further analysis showed that 1,25-(OH)(2)D(3) significantly down-regulated the expressions of protein for MMP-9 and ADAM33, as well as their mRNA levels in passively sensitized HASMCs. Conclusions: 1,25-(OH)(2)D(3) has direct inhibitory effects on passively sensitized HASMCs in vitro, including inhibition of cell proliferation and expression of MMP-9 and ADAM33, suggesting a possible beneficial role for 1,25-(OH)(2)D(3) in preventing and treating asthmatic airway remodelling. Less