Li-Yun Miao

Discoidin domain receptor 1 is associated with poor prognosis of non-small cell lung cancer and promotes cell invasion via epithelial-to-mesenchymal transition

Discoidin domain receptors (DDRs) are a novel class of receptor tyrosine kinases that respond to several collagens and facilitate cell adhesion. DDR1 is highly expressed in a variety of human cancers, and it is clear that DDR1 is primarily expressed in epithelial cells including lung, colon and brain. Moreover, DDR1 expression can be stimulated by collagen types I, II, III, IV, V, VIII and XI, and aberrant signaling induced by DDR1 dysregulated expression is involved in various steps of tumorigenesis. However, the molecular mechanism underlying the role of DDR1 in cancer development is not well documented. In this study, we found that the expression of DDR1 is upregulated in non-small cell lung cancer (NSCLC) tissues and cells when compared with counterpart normal tissues and cells. Furthermore, collagen I could induce DDR1 expression, and activated DDR1 promoted NSCLC cell migration and invasion, while knockdown of DDR1 by transfection with siRNA resulted in a significant decrease in cell migrativeness and invasiveness. Enhanced DDR1 expression mediated by collagen I could activate MMP-2, N-cadherin and vimentin expression, but reduce E-cadherin expression; however, inhibition of DDR1 expression could suppress MMP-2, N-cadherin and vimentin expression and induce E-cadherin activation. In conclusion, our findings indicated that upregulation of DDR1 induced by collagen I may contribute to the development and progression of NSCLC and this effect may be associated with increased invasiveness, at least in part, via promoting epithelial-to-mesenchymal transition.

SPOCK1 is a novel transforming growth factor-β target gene that regulates lung cancer cell epithelial-mesenchymal transition

Lung cancer is the leading cause of cancer related death worldwide and the prognosis is still poor with 5-year survival of approximately 15%. Metastasis is the leading cause of death by cancer. Recent researches have demonstrated that epithelial-to-mesenchymal transition (EMT) plays a key role in the early process of metastasis of cancer cells. Here, we identified that SPARC/osteonectin, cwcv and kazal-like domains proteoglycan 1 (SPOCK1) is a novel transforming growth factor-β1 (TGF-β) target gene that regulates lung cancer cell EMT. TGF-β has been reported as a major inductor of EMT. We observed that the expression of SPOCK1 in lung cancer tumor tissues is significantly higher than matched normal lung tissues. Moreover, the expression of SPOCK1 was also significantly higher in metastasis tumor tissues than non-metastasis tumor tissues. Levels of SPOCK1 mRNA were increased among patients with shorter disease-free survival times, indicating the potential role of SPOCK1 in lung cancer progression and metastasis. Silencing SPOCK1 expression with endoribonuclease-prepared small interfering RNA (esiRNA) in lung cells inhibits lung cancer cell growth, colony formation and invasion in vitro. Interestingly, ectopic expression of SPOCK1 in epithelial lung cancer cells induced EMT with increased expression of the mesenchymal marker Vimentin and decreased expression of epithelial marker E-cadherin. We also found that the expression of SPOCK1 was increased under treatment of TGF-β, indicating that SPOCK1 is a novel downstream target of TGF-β. Taken together, our study showed that SPOCK1 is a novel metastasis related biomarker in lung cancer and may be new diagnostic and therapeutic target for lung cancer.

Identification of novel driver mutations of the discoidin domain receptor 2 (DDR2) gene in squamous cell lung cancer of Chinese patients

BackgroundAlthough many of the recently approved genomically targeted therapies have improved outcomes for patients in non–small-cell lung cancer (NSCLC) with lung adenocarcinoma, little is known about the genomic alterations that drive lung squamous cell cancer (SCC) and development of effective targeted therapies in lung SCC is a promising area to be further investigated. Discoidin domain receptor 2 (DDR2), is a novel receptor tyrosine kinases that respond to several collagens and involved in tissue repair, primary and metastatic cancer progression.MethodsExpression of DDR2 mRNA was analyzed in 54 lung SCC tissues by qRT-PCR. Over-expression approaches were used to investigate the biological functions of DDR2 and its’ mutations in lung SCC cells. Conventional Sanger sequencing was used to investigate the mutations of DDR2 gene in 86 samples. The effect of DDR2 and its’ mutations on proliferation was evaluated by MTT and colony formation assays; cell migration and invasion was evaluated by trasnwell assays. Lung SCC cells stably transfected with pEGFP-DDR2 WT, pEGFP-DDR2-S131C or empty vector were injection into nude mice to study the effect of DDR2 and its’ mutation on tumorigenesis in vivo. Protein and mRNA expression levels of E-cadherin and MMP2 were determined by qRT-PCR and western blot analysis. Differences between groups were tested for significance using Student’s t-test (two-tailed).ResultsIn this study, we found that DDR2 mRNA levels were significantly decreased in 54 lung SCC tissues compared with normal lung tissues. Moreover, there were 3 novel DDR2 mutations (G531V, S131C, T681I) in 4 patients and provide the mutation rate of 4.6% in the 86 patients with lung SCC. The mutation of S131C in DDR2 could promote lung SCC cells proliferation, migration and invasion via inducing MMP-2, but reducing E-cadherin expression.ConclusionsThese data indicated that the novel DDR2 mutation may contribute to the development and progression of lung SCC and this effect may be associated with increased proliferation and invasiveness, at least in part, via regulating E-cadherin expression.

Meta-Analysis of the Adverse Effects of Long-Term Azithromycin Use in Patients with Chronic Lung Diseases

ABSTRACT The adverse effects of azithromycin on the treatment of patients with chronic lung diseases (CLD) were evaluated in the present study. MEDLINE and other databases were searched for relevant articles published until August 2013. Randomized controlled trials that enrolled patients with chronic lung diseases who received long-term azithromycin treatment were selected, and data on microbiological studies and azithromycin-related adverse events were abstracted from articles and analyzed. Six studies were included in the meta-analysis. The risk of bacterial resistance in patients receiving long-term azithromycin treatment was increased 2.7-fold (risk ratio [RR], 2.69 [95% confidence interval {95% CI}, 1.249, 5.211]) compared with the risk in patients receiving placebo treatment. On the other hand, the risk of bacterial colonization decreased in patients receiving azithromycin treatment (RR, 0.551 [95% CI, 0.460, 0.658]). Patients receiving long-term azithromycin therapy were at risk of increased impairment of hearing (RR, 1.168 [95% CI, 1.030, 1.325]). This analysis provides evidence supporting the idea that bacterial resistance can develop with long-term azithromycin treatment. Besides the increasingly recognized anti-inflammatory role of azithromycin used in treating chronic lung diseases, we should be aware of the potential for adverse events with its long-term use.

Loss of long noncoding RNA FOXF1-AS1 regulates epithelial-mesenchymal transition, stemness and metastasis of non-small cell lung cancer cells.

Although recent evidence shows that long noncoding RNAs (lncRNAs) are involved in the regulation of gene expression and cancer progression, the understanding of the role of lncRNAs in lung cancer metastasis is still limited. To identify novel lncRNAs in non-small cell lung cancer (NSCLC), we profile NSCLC tumor and matched normal samples using GeneChip® Human Gene 2.0 ST Array, which provides the most accurate, sensitive, and comprehensive measurement of protein coding and lncRNA transcripts. We identified a panel of key factors dysregulated in lung cancer. Among them, the expression of FOXF1-AS1 was significantly downregulated in lung cancer. Stable overexpression of FOXF1-AS1 inhibits lung cancer cell migration and invasion by regulating EMT. Meanwhile, loss of FOXF1-AS1 mediates stem-like properties of lung cancer cells. Interestingly, we found that FOXF1-AS1 physically associates with PRC2 components EZH2 and loss of FOXF1-AS1 mediates cell migration and stem-like properties require EZH2. Loss of FOXF1-AS1 is also correlated with downregulation of FOXF1 in lung cancer. These results suggested that FOXF1-AS1 might regulate EMT, stemness and metastasis of NSCLC cells via EZH2, indicating it as a therapeutic target for future treatment of NSCLC.

Elevated Platelets Enhance Cancer Cell Migration, Promote Hematogenous Metastasis and Associate with a Poor Prognosis in Advanced Non-small Cell Lung Cancer Cases

Although correlations between platelets and lung cancer has been recognized, effects on non-small cell lung cancer (NSCLC) metastasis remain to be determined in detail. In the present study, wound healing assays revealed a role of platelets in NSCLC cell migration . Thus the mean migration rate of lung adenocarcinoma A549 cells was significantly elevated after co-culture with platelets (81.7±0.45% vs 41.0±3.50%, P<0.01). Expression of GAPDH was examined by reverse transcription-polymerase chain reaction to study the effect of platelets on NSCLC cell proliferation. The result showed that the proliferation of A549 and SPC-A1 cells was not affected. Mouse models were established by transfusing A549 cells and SPC-A1 cells into mice lateral tail veins. We found tumor metastasis nodules in lungs to be increased significantly after co-transfusion with platelets (in A549, 4.33±0.33 vs 0.33±0.33, P=0.01; in SPC-A1, 2.67±0.33 vs 0.00±0.00, P=0.01). In addition, consecutive inoperable patients with newly diagnosed NSCLC (TNM stage III or IV) between January 2009 and December 2011 were retrospectively reviewed. Using the Kaplan-Meier method, NSCLC patients with a high platelet counts demonstrated a significantly shorter progression free survival compared with those with a low platelet count (>200x109/L, 3 months versus ≤200x109/L, 5 months, P=0.001). An elevated platelet count was also identified as an independent prognostic factor by Cox regression analysis for prgression free survival (adjusted hazard ratio: 1.69; 95% CI: 1.16, 2.46; P=0.006). This study suggested that platelets might contribute to the hematogenous metastatic process by promoting cancer cell migration, which eventually affects the prognosis of NSCLC.

The overexpression of glypican-5 promotes cancer cell migration and is associated with shorter overall survival in non-small cell lung cancer

Although the correlation between glypican-5 (GPC5) and lung cancer is well known, the effect of GPC5 expression on non-small cell lung cancer (NSCLC) survival remains to be determined. In the present study, GPC5 expression in A549, H3255, and SPC-A1 NSCLC cell lines was evaluated by reverse transcription-polymerase chain reaction (RT-PCR) and western blot analysis. GPC5 mRNA and protein expression levels were found to be higher in A549 and H3255 cells compared with SPC-A1 cells. The role of GPC5 in NSCLC cell migration was evaluated in vitro by shRNA-mediated knockdown or the overexpression of GPC5 through scratch and transwell assays. The mean migration rates of cancer cells transfected with pRNAT-shRNA-GPC5-1 were reduced compared with the controls in A549 (P<0.001) and H3255 (P=0.001), while the migration rate of SPC-A1 with GPC5 overexpression was higher than that of the control (P=0.001). The downregulation of GPC5 impeded the transmigration of A549 and H3255 while the upregulation of GPC5 expression promoted the transmembrane invasion of SPC-A1. Furthermore, a panel of formalin-fixed paraffin-embedded NSCLC tissues from 127 patients undergoing curative resection (stages I, II and III) between January, 2003 and December, 2008 were obtained in order to investigate the correlation between GPC5 expression and clinicopathological factors using immunohistochemical methods. The results demonstrated that high GPC5 expression levels in NSCLC were associated with respiratory symptoms in lung cancer diagnosis, poor differentiation, vascular invasion, regional lymph node metastasis and a higher TNM stage. Using the Kaplan-Meier method, NSCLC patients with high levels of GPC5 expression demonstrated a significantly shorter overall survival time compared with those with low GPC5 expression levels (median postsurgical survival time: 14.0 months vs. 59.0 months, P=0.001). GPC5 expression was also identified as an independent prognostic factor by Cox regression analysis [adjusted hazard ratio: 2.18; 95% confidence interval (CI): 1.35–3.52; P=0.001]. This study suggested that increased levels of GPC5 expression are a poor prognostic marker for NSCLC.

Clinical and Radiological Profile of Acute Fibrinous and Organizing Pneumonia: A Retrospective Study

Background: Acute fibrinous and organizing pneumonia (AFOP) is a unique pathological entity with intra-alveolar fibrin in the form of “fibrin balls” and organizing pneumonia. It was divided into rare idiopathic interstitial pneumonia according to the classification notified by American Thoracic Society/European Respiratory Society in 2013. As a rare pathological entity, it is still not well known and recognized by clinicians. We reviewed the clinical features of 20 patients with AFOP diagnosed in a teaching hospital. Methods: The medical records of 20 patients with biopsy-proven diagnosis of AFOP were retrospectively reviewed. The patients’ symptoms, duration of the disease, comorbidities, clinical laboratory data, pulmonary function testing, radiographic studies, and the response to treatment were extracted and analyzed. Results: Fever was the most common symptom and was manifested in 90% of AFOP patients. For clinical laboratory findings, systematic inflammatory indicators, including C-reactive protein and erythrocyte sedimentation rate, were significantly higher than normal in AFOP patients. In accordance with this increased indicators, injured liver functions were common in AFOP patients. Inversely, AFOP patients had worse clinical conditions including anemia and hypoalbuminemia. For pulmonary function testing, AFOP patients showed the pattern of restrictive mixed with obstructive ventilation dysfunction. For high-resolution computerized tomography (HRCT) findings, the most common pattern for AFOP patients was lobar consolidation which was very similar to pneumonia. However, unlike pneumonia, AFOP patients responded well to glucocorticoids. Conclusion: Patients with AFOP manifest as acute inflammatory-like clinical laboratory parameters and lobar consolidation on HRCT, but respond well to steroid.

Hypoxia Induced High Expression of Thioredoxin Interacting Protein (TXNIP) in Non-small Cell Lung Cancer and its Prognostic Effect

Although associations between thioredoxin interacting protein (TXNIP) and cancers have been recognized, the effects of TXNIP on non-small cell lung cancer (NSCLC) prognosis remained to be determined in detail. In addition, while hypoxia is a key characteristic of tumor cell growth microenvironment, the effect of hypoxia on TXNIP expression is controversial. In this study, formaldehyde fixed and paraffin embedded (FFPE) samples of 70 NSCLC patients who underwent resection between January 2010 and December 2011 were obtained. Evaluation of TXNIP and hypoxia inducible factor-1α (HIF-1α) protein expression in FFPE samples was made by immunohistochemistry. By Kaplan-Meier method, patients with high TXNIP expression demonstrated a significantly shorter progression free survival (PFS) compared with those with low TXNIP expression (18.0 months, 95%CI: 11.7, 24.3 versus 23.0 months, 95%CI: 17.6, 28.4, P=0.02). High TXNIP expression level was also identified as an independent prognostic factor by Cox regression analysis (adjusted hazard ratio: 2.46; 95%CI: 1.08, 5.56; P=0.03). Furthermore, TXNIP expression was found to be significantly correlated with HIF- 1α expression (Spearman correlation=0.67, P=0.000). To further confirm correlations, we established a tumor cell hypoxic culture model. Expression of TXNIP was up-regulated in all three NSCLC cell lines (A549, SPC-A1, and H1299) under hypoxic conditions. This study suggests that hypoxia induces increased TXNIP expression in NSCLC and high TXNIP expression could be a poor prognostic marker.

Lesion with morphologic feature of organizing pneumonia (OP) in CT-guided lung biopsy samples for diagnosis of bronchiolitis obliterans organizing pneumonia (BOOP): a retrospective study of 134 cases in a single center

BACKGROUND Small biopsy samples are generally considered inconclusive for bronchiolitis obliterans organizing pneumonia (BOOP) diagnosis despite their potential to reveal organizing pneumonia (OP) pathologically, necessitating risky invasive tissue biopsy during surgery for reliable confirmation. OBJECTIVE OP by CT-guided lung biopsy was to evaluate the role in the diagnosis of BOOP. METHODS A retrospective review of 134 cases with the OP feature in the CT-guided lung biopsy samples between 2004 and 2011 at a single center was conducted. Diagnostic accuracy of OP by CT-guided lung biopsy and clinical-radiographic data alone were compared. RESULTS After exclusion of 11 cases due to pathology with others besides OP and 15 cases for loss to follow-up, 108 were included. Of these, 95 cases and 13 cases were classified as BOOP and non-BOOP group, respectively. Among BOOP group, only 30 were initially diagnosed as BOOP according to the typical clinical and radiographic features. The other 65 cases with atypical features were diagnosed as BOOP mainly based on OP by CT-guided lung biopsy. Among non-BOOP group, one was misdiagnosed as BOOP, and others were not BOOP according to clinical and radiographic findings. Thus, OP by CT-guided lung biopsy produced a diagnostic accuracy of 87.96% (95/108), much higher than 31.25% (30/96) observed using clinical and radiographic data alone. Combined, these techniques produced diagnostic accuracy of 98.96% (95/96). CONCLUSIONS OP by CT-guided lung biopsy can be effectively used as the pathological evidence for BOOP diagnosis and reducing unnecessary surgery.