Qinying Sun

Slit2-N inhibits PDGF-induced migration in rat airway smooth muscle cells: WASP and Arp2/3 involved.

BACKGROUND Slit2 has been reported to be implicated in many kinds of cell migration. However little is known about the effect of Slit2 on airway smooth muscle cell migration. This study was to detect the expression of Slit2 in rat airway smooth muscle (RASM) cells stimulated by platelet-derived growth factor (PDGF) and characterized the effect of Slit2-N on PDGF-induced migration of RASM cells in vitro. METHODS mRNAs of Slit-Robo in RASM cells were examined by RT-PCR, and the effect of exogenous Slit2-N at different doses on PDGF-induced migration of RASM cells were examined by transwell and scrape-wound assays. Actin filaments (F-actin) were stained with rhodamine-conjugated phalloidin and the levels of protein expression were detected by western blot. RESULTS RASM cells were identified to express Slit2, Slit3, Robo1, Robo2 and Robo4 in vitro. Slit2-N caused a time- and dose-dependent inhibition of cell proliferation, while had no significantly effect on cell apoptosis. Slit2-N pretreatment attenuated the elongated morphologic characteristics, reduced lamellipodia formation, inhibited actin rearrangement and cell migration induced by PDGF. PDGF-induced increase of WASP and Arp2/3 proteins were dramatically inhibited by 100 ng/ml Slit2-N. CONCLUSION Slit2-N inhibits RASM cells migration at least partly through attenuating the expressions of WASP and Arp2/3, inhibiting actin rearrangement in vitro. The results contribute to provide new insights into the pathogenesis of airway remodeling in asthma and may be helpful for development of effective treatments.

5-Aza-2′-deoxycytidine inhibited PDGF-induced rat airway smooth muscle cell phenotypic switching

Airway smooth muscle (ASM) cell phenotypic switching played an important role in airway remodeling in asthma. In vitro platelet-derived growth factor (PDGF) induced ASM cell phenotypic switching from a mature to pro-remodeling phenotype, but the mechanism remained incompletely understood. This study was to explore the effect of DNA methyltransferase inhibitor 5-Aza-2′-deoxycytidine (Aza-CdR) on PDGF-induced rat ASM cell phenotypic switching and biological behaviors. Rat airway smooth muscle (RASM) cells were obtained by primary explant techniques. Western blot, 3-dimensional gel contraction, transwell and wound healing assay, and MTT were applied to detect cell phenotypic switching, contractility, migration and proliferation, respectively. Cytoskeleton rearrangement was observed by immunofluorescence. Results showed Aza-CdR inhibited PDGF-induced down-regulation of contractile markers in RASM cells and increased cell contractility. Aza-CdR inhibited PDGF-induced RASM cell migration by abrogating cell morphology change and cytoskeletal reorganization and attenuated the effect of PDGF on proliferating cell nuclear antigen expression and cell cycle progression, ultimately cell proliferation. PDGF-induced DNA methyltransferase 1 (DNMT1) expression was mediated by activation of PI3K/Akt and ERK signaling in RASM cells. Selective depletion of DNMT1 protein by Aza-CdR inhibited PDGF-induced RASM cell phenotypic switching, revealing DNMT1-mediated DNA methylation was implicated in asthmatic ASM remodeling. We proposed for the first time that DNMT1 played a key role in PDGF-induced RASM cell phenotypic switching and Aza-CdR is promising in intervening ASM remodeling in asthma. Although study of abnormal DNA methylation in PDGF-stimulated ASM cells is in its infancy, this work contributes to providing new insights into the mechanism of ASM remodeling and may be helpful for developing effective treatments for airway remodeling in asthma.

Overexpression of response gene to complement 32 (RGC32) promotes cell invasion and induces epithelial–mesenchymal transition in lung cancer cells via the NF-κB signaling pathway

Response gene to complement 32 (RGC32) is a novel cellular protein that has been reported to be expressed aberrantly in multiple types of human tumors. However, the role of RGC32 in cancer is still controversial, and the molecular mechanisms by which RGC32 contributes to the development of cancer remain largely unknown. In the present study, we constructed a recombinant expression vector pCDNA3.1-RGC32 and transfected it into human lung cancer A549 cells. Stable transformanted cells were identified by real-time PCR and Western blot analysis. Functional analysis showed that forced overexpression of RGC32 increased invasive and migration capacities of lung cancer cells in vitro, and induced the acquisition of epithelial–mesenchymal transition (EMT) phenotype, as demonstrated by the spindle-like morphology, downregulation of E-cadherin, and upregulation of Vimentin, Fibronectin, Snail and Slug. Also, overexpression of RGC32 increased expression and activities of matrix metalloproteinase (MMP)-2 and MMP-9 in A549 cells. Furthermore, the downregulation of E-cadherin induced by RGC32 was remarkably attenuated by nuclear factor-κB (NF-κB) inhibitor BAY 11-7028 and small interfering RNA targeting NF-κB p65, suggesting a role of the NF-κB signaling pathway in RGC32-induced EMT. Taken together, our data suggest that RGC32 promotes cell migration and invasion and induces EMT in lung cancer cells via the NF-κB signaling pathway.

A Clinical Study of Tracheobronchopathia Osteochondroplastica: Findings from a Large Chinese Cohort

Background and Study Aims Tracheobronchopathia osteochondroplastica (TO) is an uncommon disease of the tracheobronchial system that leads to narrowing of the airway lumen from cartilaginous and/or osseous submucosal nodules. The aim of this study is to perform a detailed review of this rare disease in a large cohort of patients with TO proven by fiberoptic bronchoscopy from China. Patients and Methods Retrospective chart review was performed on 41,600 patients who underwent bronchoscopy in the Department of Respiratory Medicine of Changhai Hospital between January 2005 and December 2012. Cases of TO were identified based on characteristic features during bronchoscopic examination. Results 22 cases of bronchoscopic TO were identified. Among whom one-half were male and the mean age was 47.45±10.91 years old. The most frequent symptoms at presentation were chronic cough (n = 14) and increased sputum production (n = 10). Radiographic abnormalities were observed in 3/18 patients and findings on computed tomography consistent with TO such as beaded intraluminal calcifications and/or increased luminal thickenings were observed in 18/22 patients. Patients were classified into the following categories based on the severity of bronchoscopic findings: Stage I (n = 2), Stage II (n = 6) and Stage III (n = 14). The result that bronchoscopic improvement was observed in 2 patients administered with inhaled corticosteroids suggested that resolution of this disease is possible. Conclusions TO is a benign disease with slow progression, which could be roughly divided into 3 stages on the basis of the characteristic endoscopic features and histopathologic findings. Chronic inflammation was thought to be more important than the other existing plausible hypotheses in the course of TO. Inhaled corticosteroids might have some impact on patients at Stage I/II.

Nogo-B protects mice against lipopolysaccharide-induced acute lung injury

Nogo-B, a member of the reticulon 4 protein family, plays a critical role in tissue repair and acute inflammation. Its role in acute lung injury (ALI) remains unclear. Here, we assessed the function of Nogo-B during tissue injury in a lipopolysaccharide (LPS)-induced ALI mouse model. We found that pulmonary Nogo-B was significantly repressed after LPS instillation in C57BL/6 mice. Over-expression of pulmonary Nogo-B using an adenovirus vector carrying the Nogo-B-RFP-3flag gene (Ad-Nogo-B) significantly prolonged the survival of mice challenged with a lethal dose of LPS. The Ad-Nogo-B-treated mice also had less severe lung injury, less alveolar protein exudation, and a higher number of macrophages but less neutrophil infiltration compared with Ad-RFP-treated mice. Interestingly, microarray analysis showed that the Ad-Nogo-B-treated mice had different gene expression profiles compared with the controls and the prominent expression of genes related to wound healing and the humoral immune response after LPS induction. Of the 49 differently expressed genes, we found that the expression of PTX3 was significantly up-regulated following Nogo-B over-expression as observed in lung tissues and RAW264.7 cells. In conclusion, Nogo-B plays a protective role against LPS-induced ALI, and this effect might be exerted through the modulation of alveolar macrophage recruitment and PTX3 production.