Ping Chen

Atorvastatin reduces lipopolysaccharide-induced expression of cyclooxygenase-2 in human pulmonary epithelial cells

ObjectiveTo explore the effects of atorvastatin on expression of cyclooxygenase-2 (COX-2) in human pulmonary epithelial cells (A549).MethodsA549 cells were incubated in DMEM medium containing lipopolysaccharide (LPS) in the presence or absence of atorvastatin. After incubation, the medium was collected and the amount of prostaglandin E2 (PGE2) was measured by enzyme-linked immunosorbent assay (ELISA). The cells were harvested, and COX-2 mRNA and protein were analyzed by RT-PCR and western-blot respectively.ResultsLPS increased the expression of COX-2 mRNA and production of PGE2 in a dose- and time-dependent manner in A549. Induction of COX-2 mRNA and protein by LPS were inhibited by atorvastatin in a dose-dependent manner. Atorvastatin also significantly decreased LPS-induced production of PGE2. There was a positive correlation between reduced of COX-2 mRNA and decreased of PGE2 (r = 0.947, P < 0.05).ConclusionAtorvastatin down-regulates LPS-induced expression of the COX-2 and consequently inhibits production of PGE2 in cultured A549 cells.

Endothelin-1 increases expression of cyclooxygenase-2 and production of interlukin-8 in hunan pulmonary epithelial cells

Inducible cyclooxygenase (COX-2) and inflammatory cytokines play important roles in inflammatory processes of chronic obstructive pulmonary disease (COPD). Endothelin-1 (ET-1) might be also involved in the pathophysilogical processes in COPD. In the present study, we determined whether ET-1 could regulate the expression of COX-2 and alter the production of interleukin-8 (IL-8) in human pulmonary epithelial cells (A549). Induced sputum samples were collected from 13 stable COPD patients and 14 healthy subjects. The COX-2 protein, ET-1, PGE(2) and IL-8 in these sputum samples were analyzed. A549 cells were incubated with ET-1 in the presence or absence of celecoxib, a selective COX-2 inhibitor. The expression of COX-2 protein in the cell and the amounts of PGE(2) and IL-8 in the medium were measured. The levels of COX-2 protein, ET-1, PGE(2) and IL-8 were significantly increased in induced sputum from COPD patients when compared to healthy subjects. ET-1 increased the expression of COX-2 protein, as well as the production of PGE(2) in A549 cells. Increased production of PGE(2) was inhibited by celecoxib. ET-1 also increased the production of IL-8. Interestingly, ET-1-induced production of IL-8 was also inhibited by celecoxib. These findings indicate that ET-1 plays important roles in regulating COX-2 expression and production of IL-8 in A549 cells. ET-1 mediated production of IL-8 is likely through a COX-2-dependent mechanism.

Involvement of PKC, p38 MAPK and AP-2 in IL-1β-induced expression of cyclooxygenase-2 in human pulmonary epithelial cells

Objective:u2003 The aim of this study was to identify the signal molecules involved in IL‐1β‐induced expression of cyclooxygenase (COX)‐2 in human pulmonary epithelial (A549) cells.

Expression and methylation of mitochondrial transcription factor a in chronic obstructive pulmonary disease patients with lung cancer

Background Apoptosis plays a central role in the pathogenesis of chronic obstructive pulmonary disease (COPD), and this process can be regulated by mitochondrial transcription factor A (mtTFA). Epigenetics is involved in the regulation and modification of the genes involved in lung cancer and COPD. In this study, we determined the expression of mtTFA and its methylation levels in the COPD patients with lung cancer. Methods Twenty-one squamous cell lung cancer patients, 11 with COPD and 10 without COPD, undergoing pneumonectomy were enrolled. The apoptotic index (AI) of pulmonary vascular endothelial cells was analyzed by transferase-mediated deoxyuridine triphosphate-biotin nick end labeling assay. The expression of mtTFA mRNA and protein was measured using PCR, immunohistochemistry and Western-blot. Methylation of the mtTFA promoter was detected using bisulfite sequencing PCR. Results Compared to the non-COPD group, the AI was higher, and expression of mtTFA mRNA and protein was lower in the COPD group (P<0.001). Expression of the mtTFA protein was positively correlated with FEV1/Pre (ru200a=u200a0.892, P<0.001), and negatively correlated with AI (ru200a=u200a−0.749, P<0.001) and smoke index (ru200a=u200a−0.763, P<0.001). Percentage of mtTFA promoter methylation in the COPD patients was significantly higher compared to the non-COPD patients (P<0.05). Conclusion These results suggest that the expression of mtTFA mRNA and protein is down-regulated in the lung tissue from the COPD patients with squamous cell lung cancer, and the level of mtTFA protein is related to apoptosis of pulmonary vascular endothelial cells. Aberrant mtTFA methylation may also play an important role in the pathogenesis of COPD.

Atorvastatin reduces lipopolysaccharide-induced expression of C-reactive protein in human lung epithelial cells

Accumulating evidence suggests that statins possess anti-inflammatory properties and may decrease C-reactive protein (CRP) levels in plasma. However, no studies have as yet addressed whether or not statins regulate the expression of CRP in human lung epithelial cells (A549). In this study, we determined whether atorvastatin modulates the lipopolysaccharide (LPS)-induced expression of CRP in A549 cells. A549 cells were incubated in Dulbeccos modified Eagles medium containing LPS in the absence or presence of various concentrations of atorvastatin. After incubation, the medium was collected and the amount of CRP was measured by an enzyme-linked immunosorbent assay. The cells were harvested and CRP messenger ribonucleic acid (mRNA) was analyzed by reverse transcription polymerase chain reaction. Incubation with LPS induced a significant time- and dose-dependent increase in CRP mRNA expression and CRP production in A549 cells, whereas atorvastatin significantly decreased LPS-induced CRP mRNA expression and CRP production in a dose-dependent manner. The present study revealed that A549 cells are capable of LPS-induced CRP expression, and that atorvastatin down-regulates the LPS-induced expression of CRP in cultured A549 cells. Our results suggest that statins ameliorate lung inflammation by regulating CRP production in human lung epithelial cells.

Changes in the number of CD31 − CD45 − Sca-1 + cells and Shh signaling pathway involvement in the lungs of mice with emphysema and relevant effects of acute adenovirus infection

Background COPD is a leading cause of mortality worldwide, and cigarette smoke is a pivotal risk factor. Adenovirus is a common cause of acute exacerbations of COPD and expedites COPD progression. Lung stem/progenitor cells play an important role in the development of COPD, while the relevant mechanism remains elusive. Here, we investigated the number of lung CD31−CD45−Sca-1+ cells and sonic hedgehog (Shh) signaling pathway expression levels in cigarette smoke extract (CSE)-induced emphysema mice, as well as the relevant effects of acute adenovirus infection (AAI). Materials and methods BALB/c mice were treated with CSE by intraperitoneal injection and/or adenovirus endotracheal instillation at different time points for 28 days. Lung function, lung histomorphology, CD31−CD45−Sca-1+ cell count, and expression levels of major components in the Shh signaling pathway in the lungs were measured. Results CSE intraperitoneal injection and adenovirus endotracheal instillation successfully induced emphysema and AAI in mice, respectively. In the lungs of emphysema mice, both the number of CD31−CD45−Sca-1+ cells and expression levels of Shh signaling pathway molecules were reduced. However, AAI increased the number of inhibited CD31−CD45−Sca-1+ cells and activated the suppression of the Shh signaling pathway. Conclusion Both CD31−CD45−Sca-1+ cell numbers and Shh signaling pathway expression levels were downregulated in the lungs of emphysema mice induced by CSE intraperitoneal injection, which likely contributes to the pathogenesis of emphysema. Additionally, these inhibited lung CD31−CD45−Sca-1+ cells and Shh signaling pathway molecules were upregulated during AAI, indicating that they play a protective role in the epithelial repair process after AAI injury.

Effects of One-Hour Training Course and Spirometry on the Ability of Physicians to Diagnose and Treat Chronic Obstructive Pulmonary Disease

Backgrounds In China, the prevalence of chronic obstructive pulmonary disease (COPD) in persons 40 years of age or older is estimated at 8.2%, but this is likely a substantial underestimate. Methods Eight secondary hospitals which didn’t have spirometries were chosen randomly in Hunan province of central south China. Physician subjects at these hospitals underwent a one-hour training course on the Chinese COPD guidelines. Physicians answered questionnaires assessing their knowledge of the guidelines before and after the training session. The mean correct scores of questionnaires were compared before and after training. Four out of the eight hospitals were given access to spirometry. Eligible patient subjects underwent spirometry testing prior to the physician visit. After seeing the patient, physicians were asked to answer a questionnaire relating to the diagnosis and severity of COPD. Physicians were then given the results of the spirometry, and asked to answer the same questionnaire. Physicians’ responses before and after receiving the spirometry results were compared. Results 225 physicians participated in the training session. 207 questionnaires were completed. Mean scores (out of 100) before and after the training were 53.1 ± 21.7 and 93.3 ± 9.8, respectively. 18 physicians and 307 patient subjects participated in the spirometry intervention. Based on spirometric results, the prevalence of COPD was 38.8%. Physicians correctly identified the presence of COPD without spirometric data in 85 cases (76.6%); this increased to 117 cases (97.4%) once spirometric data were available. Without spirometric data, physicians incorrectly diagnosed COPD in 38 patients; this decreased to 6 patients once spirometric data were available. Spirometric data also improved the ability of physicians to correctly grade COPD severity. Conclusions Simple educational training can substantially improve physicians’ knowledge relating to COPD. Spirometry combined with education improves the ability of physicians to diagnose COPD and to assess its severity.

The protective effect of PRMT6 overexpression on cigarette smoke extract-induced murine emphysema model

Background Cigarette smoke exposure is the most common risk factor for emphysema, which is one of the major pathologies of COPD. Protein arginine methyltransferase 6 (PRMT6) is a nuclear enzyme that specially catalyzes dimethylation of R2 in histone H3 (H3R2me2a). H3R2me2a prevents trimethylation of H3K4 (H3K4me3), which is located in the transcription start sites of genes in mammalian genomes. We attempted to determine the expression of PRMT6 in human samples, and investigate whether the upregulation of PRMT6 expression can attenuate the development of cigarette smoke extract (CSE)-induced emphysema. Further experiments were performed to elucidate the molecular mechanisms involved. Materials and methods Human lung tissues were obtained from patients undergoing pneumonectomy for benign pulmonary lesions. BALB/c mice were treated with lentiviral vectors intratracheally and injected with CSE three times. The protein expression of PRMT6, H3R2me2a, and H3K4me3 in human and mouse samples, as well as B-cell lymphoma-2 (Bcl-2), Bcl-2-associated X protein (Bax), and endothelial nitric oxide synthase (eNOS) in mice were detected in lung homogenates by Western blotting. The mRNA expression of cyclooxygenase-2, interleukin-6, Bcl-2, Bax, and eNOS in mice was measured by quantitative real-time polymerase chain reaction. Results The expression of PRMT6 was significantly downregulated in the pulmonary parenchyma in smokers with COPD as well as in mice treated with CSE. Overexpression of PRMT6 was detected in the CSE + Lenti-PRMT6 group of mice, which reversed the expression of H3R2me2a and H3K4me3. Inflammation, apoptosis, and oxidative stress levels were severe in the CSE-treated emphysema mice compared with the control group, which was inhibited by the overexpression of PRMT6. Conclusion The overexpression of PRMT6 might inhibit inflammation, apoptosis, and oxidative stress in CSE-induced emphysema mediated by H3R2me2a.

Intratracheal transplantation of endothelial progenitor cells attenuates smoking-induced COPD in mice

Background Endothelial progenitor cells (EPCs) might play a protective role in COPD. The aim of this study was to investigate whether intratracheal allogeneic transplantation of bone-marrow-derived EPCs would attenuate the development of smoking-induced COPD in mice. Methods Isolated mononuclear cells from the bone marrow of C57BL/6J mice were cultured in endothelial cell growth medium-2 for 10 days, yielding EPCs. A murine model of COPD was established by passive 90-day exposure of cigarette smoke. On day 30, EPCs or phosphate-buffered saline alone was administered into the trachea. On day 90, EPCs or 30 μL phosphate-buffered saline alone was administered into the trachea, and on day 120, inflammatory cells, antioxidant activity, apoptosis, matrix metalloproteinase (MMP)-2, and MMP-9 were measured. Results After EPC treatment, the lung function of the mice had improved compared with the untreated mice. Mean linear intercept and destructive index were reduced in the EPCs-treated group compared with the untreated group. In addition, the EPCs-treated mice exhibited less antioxidant activity in bronchoalveolar lavage fluid compared with the untreated mice. Moreover, decreased activities of MMP-2, MMP-9, and TUNEL-positive cells in lung tissues were detected in EPCs-treated mice. Conclusion Intratracheal transplantation of EPCs attenuated the development of pulmonary emphysema and lung function disorder probably by alleviating inflammatory infiltration, decelerating apoptosis, inhibiting proteolytic enzyme activity, and improving antioxidant activity.

Pulmonary Diseases and Epigenetics

Abstract Pulmonary diseases are one of the leading causes of death in the world. Chronic obstructive pulmonary disease (COPD), asthma, interstitial lung diseases, pulmonary arterial hypertension (PAH), lung tuberculosis, and pulmonary embolism, as well as other disorders, are significant public health burdens. Although advances in methods of detection, intervention, and treatment have prolonged life for patients, these complex illnesses continue to have serious effects on a large segment of the global population. Epigenetics is defined as the study of heritable changes in gene expression or cellular phenotype caused by mechanisms that do not alter the nucleotide sequence. There are three main classes of epigenetic marks: DNA methylation, histone modifications, and noncoding RNAs. DNA methylation involves the addition of a methyl group to the 5 position of cytosine by DNA methyltransferases and can be inherited through cell division. Histone modifications are posttranslational processes such as acetylation, methylation, phosphorylation, and ubiquitination of the tails of core histones, each of which plays different roles in gene expression. MicroRNAs are approximately 22 nucleotide-long regulatory RNAs that control gene expression by binding to messenger RNA (mRNA), leading to mRNA degradation or inhibition of protein translation. Increasingly, the evidence suggests that these three important epigenetic mechanisms play significant roles in the pathogenetic processes of lung diseases. Understanding these mechanisms may lead to the development of novel diagnostic and therapeutic approaches. In this chapter the influence of epigenetic variations on the pathophysiology of COPD, asthma, interstitial lung diseases, PAH, lung tuberculosis, and pulmonary embolism is described.