Interleukin (IL)-4 induces production of cytokine-induced neutrophil chemoattractants (CINCs) and intercellular adhesion molecule (ICAM)-1 in lungs of asthmatic rats / 华中科技大学学报（医学）（英德文版）

The present study aimed to examine the effect of interleukin (IL)-4 on neutrophil chemotaxis in airway inflammation in asthmatic rats and the possible mechanism. Male Wistar rats were intranasally instilled with recombinant rat (rr) IL-4 (rrIL-4) at different doses [2, 4 or 8 μg/animal, dissolved in 200 μL normal saline (NS)] or rrIL-4 at 4 μg/animal (dissolved in 200 μL NS). NS (200 μL) and LPS (6 mg/kg/animal, dissolved in 200 μL NS) were intranasally given respectively in the negative and positive control groups. Moreover, the asthmatic lung inflammation was induced in rats which were then intranasally treated with rrIL-4 (4 μg/animal) or LPS (6 mg/kg/animal). The normal rats treated with different doses of rrIL-4 and those asthmatic rats were sacrificed 6 h later. And animals instilled with rrIL-4 at 4 μg were sacrificed 6, 12 or 24 h later. The bronchoalveolar lavage fluid (BALF) and lungs were harvested for detection of leukocyte counts by Wright-Giemsa staining and lung histopathology by haematoxylin-eosin (HE) staining. The levels of cytokine-induced neutrophil chemoattractant (CINC)-1 and intercellular adhesion molecule (ICAM)-1 in BALF were determined by ELISA. Real-time PCR was used to measure the mRNA expression of CINCs (CINC-1, CINC-2α, CINC-2β, CINC-3) and ICAM-1 in lung tissues. The results showed that the intranasal instillation of IL-4 did not induce a recruitment of neutrophils in BALF in rats. However, IL-4 could increase the CINC-1 level in BALF in a dose-dependent manner at 6 h. But the mRNA expression levels of CINC-1, CINC-2α, CINC-2β, CINC-3 were not significantly increased in lungs of IL-4-treated rats relative to NS negative control group. Moreover, IL-4 was found to augment the mRNA expression of ICAM-1 in lungs and the ICAM-1 level in BALF at 6 h. However, the increase in CINC-1 and ICAM-1 levels in BALF of IL-4-treated asthmatic rats was not significantly different from that in untreated asthmatic rats. These findings indicate that IL-4 does not directly recruit neutrophils in the rat lungs, but it may contribute to airway neutrophilia through up-regulation of CINC-1 and ICAM-1.

Interleukin (IL)-4 induces production of cytokine-induced neutrophil chemoattractants (CINCs) and intercellular adhesion molecule (ICAM)-1 in lungs of asthmatic rats / 华中科技大学学报（医学）（英德文版）

The present study aimed to examine the effect of interleukin (IL)-4 on neutrophil chemotaxis in airway inflammation in asthmatic rats and the possible mechanism. Male Wistar rats were intranasally instilled with recombinant rat (rr) IL-4 (rrIL-4) at different doses [2, 4 or 8 μg/animal, dissolved in 200 μL normal saline (NS)] or rrIL-4 at 4 μg/animal (dissolved in 200 μL NS). NS (200 μL) and LPS (6 mg/kg/animal, dissolved in 200 μL NS) were intranasally given respectively in the negative and positive control groups. Moreover, the asthmatic lung inflammation was induced in rats which were then intranasally treated with rrIL-4 (4 μg/animal) or LPS (6 mg/kg/animal). The normal rats treated with different doses of rrIL-4 and those asthmatic rats were sacrificed 6 h later. And animals instilled with rrIL-4 at 4 μg were sacrificed 6, 12 or 24 h later. The bronchoalveolar lavage fluid (BALF) and lungs were harvested for detection of leukocyte counts by Wright-Giemsa staining and lung histopathology by haematoxylin-eosin (HE) staining. The levels of cytokine-induced neutrophil chemoattractant (CINC)-1 and intercellular adhesion molecule (ICAM)-1 in BALF were determined by ELISA. Real-time PCR was used to measure the mRNA expression of CINCs (CINC-1, CINC-2α, CINC-2β, CINC-3) and ICAM-1 in lung tissues. The results showed that the intranasal instillation of IL-4 did not induce a recruitment of neutrophils in BALF in rats. However, IL-4 could increase the CINC-1 level in BALF in a dose-dependent manner at 6 h. But the mRNA expression levels of CINC-1, CINC-2α, CINC-2β, CINC-3 were not significantly increased in lungs of IL-4-treated rats relative to NS negative control group. Moreover, IL-4 was found to augment the mRNA expression of ICAM-1 in lungs and the ICAM-1 level in BALF at 6 h. However, the increase in CINC-1 and ICAM-1 levels in BALF of IL-4-treated asthmatic rats was not significantly different from that in untreated asthmatic rats. These findings indicate that IL-4 does not directly recruit neutrophils in the rat lungs, but it may contribute to airway neutrophilia through up-regulation of CINC-1 and ICAM-1.

SOX40L: an important inflammatory mediator in adult bronchial asthma

<p><b>INTRODUCTION</b>The role of soluble OX40 ligand (sOX40L) in adult bronchial asthma is unclear. This study aims to determine the serum concentrations of sOX40L in adult patients with bronchial asthma, and discussed its relationship with pulmonary function.</p><p><b>MATERIALS AND METHODS</b>We measured the pulmonary function using the spirometer and detected the serum concentrations of sOX40L by enzyme linked immunosorbent assay (ELISA) in 19 healthy persons in the control group, 58 acute asthmatic adult patients who were grouped according to their disease severity: 18 mild grade, 24 moderate grade, 16 severe grade, and 24 persons in a stable asthmatic group.</p><p><b>RESULTS</b>The serum concentrations of sOX40L in asthmatic adult patients (6.80 ± 4.95 ng/L) were distinctly higher than those in the control group (3.98 ± 2.83 ng/L, P <0.05), and they were negatively correlated with pulmonary function indexes (FEV1%, FVC%, FEV1/FVC) (r = -0.754, P <0.01, r = -0.557, P <0.01, r = -0.457, P <0.01, respectively). Moreover, the serum concentrations of sOX40L showed obvious differences among control, mild, moderate, and severe groups (3.98 ± 2.83, 4.87 ± 1.89, 6.97 ± 5.91, 8.71 ± 5.18 ng/L, respectively; P <0.01). The concentrations of sOX40L decreased to the same extent as the control group after therapeutic treatments were provided to the asthmatic adult patients.</p><p><b>CONCLUSION</b>The concentrations of sOX40L were found to be high in adult asthmatic patients and were associated with the severity of the disease. Therefore, sOX40L could be a potential inflammatory mediator in the pathogenesis of asthma.</p>

Cyclin D1 is involved in human pulmonary artery smooth muscle cells proliferation and migration induced by cigarette smoke extract / 生理学报

The present study was aimed to investigate the role of cyclin D1 in human pulmonary artery smooth muscle cells (HPASMCs) proliferation and migration induced by cigarette smoke extract (CSE). The eukaryotic expression vector of antisense cyclin D1 gene (pIRES2-EGFP-ascyclin D1) was recombinated. The recombinant and empty vector were separately transfected into normal HPASMCs using liposome. Then the cells were treated with or without 5% CSE. The cells were randomly divided into six groups: control group, vector group, antisense cyclin D1 group, 5% CSE group, vector+5% CSE group and antisense cyclin D1+5% CSE group. The expressions of cyclin D1 mRNA and protein were detected by real-time fluorescence RT-PCR and Western blot, respectively. The proliferation of HPASMCs was examined by cell cycle analysis, MTT assay and proliferation cell nuclear antigen (PCNA) immunocytochemical staining. The migration of HPASMCs was measured by Transwell cell test. The results showed that the eukaryotic expression vector of antisense cyclin D1 gene was constructed and transfected into HPASMCs successfully. The cyclin D1 mRNA and protein levels in antisense cyclin D1 group were significantly lower than those in control group (P<0.05). In 5% CSE group, the cyclin D1 mRNA and protein levels were elevated significantly compared with those in control group (P<0.05), and the indicators of cell and migration in antisense cyclin D1+5% CSE group were remarkably lower than those in 5% CSE group (P<0.05). These results suggest that CSE could promote HPASMCs proliferation and migration through up-regulation of cyclin D1 expression. PIRES2-EGFP-ascyclin D1 could attenuate CSE-induced proliferation and migration of HPASMCs by suppressing the expression of cyclin D1, which implicates that cyclin D1 might be involved in the process of HPASMCs proliferation and migration stimulated by CSE.

Cigarette smoke extract promotes human pulmonary artery smooth muscle cells proliferation through protein kinase C alpha-dependent induction of cyclin D1 / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>Exposure to cigarette smoke stimulates the proliferation of human pulmonary artery smooth muscle cells (HPASMCs) in vivo and in vitro. However, the molecular mechanism remains unclear. This study aimed at investigating the role of signaling pathways involving protein kinase C alpha (PKCα) and cyclin D1 in the cigarette smoke extract (CSE)-induced HPASMCs proliferation.</p><p><b>METHODS</b>Synchronized HPASMCs were treated with different concentrations of CSE. Cell proliferation was evaluated by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and cell counting. Cell cycle was analyzed by flow cytometry with propidium iodide staining. Activation of PKCα was measured by detecting the expression of PKCα protein in the cytosolic and membrane fractions using Western blotting analysis. Small interfering RNA (siRNA) was used to knockdown PKCα and cyclin D1. The cyclin D1 mRNA was assessed by real-time RT-PCR. The PKCα and cyclin D1 protein levels were detected by Western blotting.</p><p><b>RESULTS</b>Low concentrations of CSE (1% - 10%) stimulated proliferation of HPASMCs, with its maximal effect at 5%. CSE (5%) led to PKCα activation. Inhibition of PKCα activity using Gö 6976 or siRNA-mediated knockdown of PKCα significantly attenuated CSE-induced cell proliferation and G1/S transition. Cyclin D1, one of key regulators of G1/S transition, was found to be upregulated by 5% CSE at both the mRNA and protein levels. CSE-stimulated cell proliferation and G1/S transition was abolished by cyclin D1 siRNA. Moreover, Gö 6976 or PKCα siRNA significantly suppressed CSE-induced upregulation of cyclin D1 at both the mRNA and protein levels.</p><p><b>CONCLUSION</b>PKCα-cyclin D1 pathway at least partially mediates the CSE-induced proliferation in HPASMCs.</p>