BACKGROUND: Epidermal growth factor receptor (EGFR) gene mutations are the most common driver mutations in non-small cell lung cancer (NSCLC). To prolong the survival of the patients, EGFR tyrosine kinase inhibitors (TKIs) resistance in NSCLC is a major challenge that needs to be addressed urgently, and this study focuses on investigating the mechanism of cigarette smoke (CS) induced Gefitinib resistance in NSCLC. METHODS: PC-9 and A549 cells were cultured in vitro and treated with 1 µmol/L Gefitinib for 4 h and 10% cigarette smoke extract (CSE) for 48 h. Western blot was used to detect Sirtuin 3 (Sirt3) and superoxide dismutase 2 (SOD2) protein expressions; DCFH-DA probe was used to detect intracellular reactive oxygen species (ROS); CCK-8 kit was used to detect cell activity, and EdU was used to detect cell proliferation ability. Sirt3 overexpression plasmid (OV-Sirt3) was transfected in PC-9 and A549 cells and treated with 1 µmol/L Gefitinib for 4 h and 10% CSE for 48 h after N-acetylcysteine (NAC) action. The expressions of Sirt3 and SOD2 were detected by Western blot; the ROS level in the cells was detected by DCFH-DA probe, and the cell activity was detected by CCK-8. RESULTS: CSE induced an increase in the 50% inhibitory concentration (IC50) of both PC-9 and A549 cells to Gefitinib (P<0.01) and enhanced the proliferation of PC-9 and A549 cells, suggesting that CS induced Gefitinib resistance in NSCLC. ROS was involved in CSE-induced Gefitinib resistance (P<0.05). CSE induced low expressions of Sirt3 and SOD2 (P<0.01), and Sirt3/SOD2 was associated with poor prognosis in lung cancer patients (P<0.05). OV-Sirt3 in PC-9 and A549 cells reversed CSE-induced Gefitinib resistance (P<0.05) and significantly reduced ROS production. NAC reversed CSE-induced Gefitinib resistance in PC-9 and A549 cells (P<0.05). CONCLUSIONS The ROS/Sirt3/SOD2 pathway is involved in CS-induced Gefitinib resistance in NSCLC.
Humans ; Gefitinib/therapeutic use* ; Carcinoma, Non-Small-Cell Lung/metabolism* ; Sirtuin 3/therapeutic use* ; Lung Neoplasms/metabolism* ; Reactive Oxygen Species/therapeutic use* ; Antineoplastic Agents/therapeutic use* ; Cigarette Smoking ; Sincalide/therapeutic use* ; ErbB Receptors/metabolism* ; Drug Resistance, Neoplasm/genetics* ; Cell Line, Tumor

Animals ; Bronchoalveolar Lavage Fluid ; Cigarette Smoking ; Interleukin-17/genetics* ; Lung ; MAP Kinase Signaling System ; Mice ; Mice, Inbred C57BL ; Pulmonary Disease, Chronic Obstructive

Objective: To screen differentially expressed miRNAs in the testis of male rats exposed to cigarette smoke (CS) and identify the early molecular markers of CS-induced apoptosis of testicular cells. METHODS: We randomly divided 200 SPF male SD rats into blank control and low-dose (10 non-filter cigarettes/d), medium-dose (20 non-filter cigarettes/d) and high-dose (30 non-filter cigarettes/d) CS exposure groups. After 2, 4, 6, 8 and 12 weeks of CS exposure, we observed the histopathological changes of the testis by HE staining, detected the apoptosis of the testicular cells by TUNEL, and determined the expressions of caspase-3 and caspase-9 in the testis tissue by immunohistochemistry, RT-PCR and Western blot. Based on the laboratory results, we selected 4 testicular samples from the 12-week high-dose group and another 4 from the control for miRNA microarray-based screening, bioinformatics analysis, and verification of differentially expressed miRNAs in all the animals by RT-PCR. RESULTS: Compared with the controls, the CS-exposed rats showed dose- and time-dependent increase in the atrophy of the testis and significantly increased number of apoptotic testis cells from the 6th week of exposure (P < 0.05), with dramatically up-regulated expressions of caspase-3 (P < 0.01) and caspase-9 protein and mRNA (P < 0.05) in the testis tissue. Microarray-based screening and RT-PCR revealed 5 differentially expressed miRNAs in the testis of the CS-exposed rats, of which miR-138-5p, miR-181d-5p, miR-19a-3p and miR-3588 were down-regulated, and miR-155-5p up-regulated, and the target genes of the differentially expressed miRNAs positively regulated the apoptosis of the testicular cells. CONCLUSIONS The differentially expressed miRNAs miR-155-5p, miR-138-5p, miR-181d-5p, miR-19a-3p and miR-3588 regulate CS-induced apoptosis of testicular cells, and may become biomarkers for early diagnosis and prognosis of CS-induced spermatogenesis obstruction.《.
Animals ; Male ; MicroRNAs/genetics* ; RNA, Messenger ; Rats ; Rats, Sprague-Dawley ; Smoking ; Testis

BACKGROUND: Lung cancer is a malignant tumor disease with high morbidity and high mortality. The non-small cell lung cancer (NSCLC) is the most common type, among them, lung squamous cell carcinoma own special pathological type and specific treatment, is a subtype of non-small cell lung cancer and can be divided into peripheral type and central type according to clinical phenotype. This study explores the differences in gene levels and their potential values based on clinical differences between central and peripheral in lung squamous cell carcinoma. METHODS: The lung squamous cell carcinoma dataset was collected from The Cancer Genome Atlas (TCGA) database, clinical information and the corresponding gene expression profiles were downloaded. Then we further sort and analyze all these data. RESULTS In clinical characteristics analysis, result showed that central lung squamous cell carcinoma was more likely to metastasis with lymph node than peripheral lung squamous cell carcinoma (46.2%, 67/145 vs 28.9%, 26/90; P=0.019), while there were no significant differences in gender, age, tumor size, distant metastasis, tumor node metastasis (TNM) stage, and EGFR mutation. Gene expression analysis showed 1,031 differentially expressed genes between central and peripheral lung squamous cell carcinoma, of which 629 genes were up-regulated and 402 genes were down-regulated (peripheral vs central). Further enrichment analysis showed differentially expressed genes were mainly riched in 6 signaling pathways. Among them, the neuroactive ligand-receptor interaction pathway was the main enrichment pathway of differentially expressed genes, and other differential expressed genes were mainly involved in lipid metabolism and glucose metabolism. The analysis of interaction network showed that hepatocyte nuclear factor 1 homeobox A (HNF1A) and cytochrome p450 family, Cytochrome P450 3A4 (CYP3A4) own widely effect in up-regulated genes, while ALB and APOA1 at the key positions of the network in down-regulated genes were CONCLUSIONS: Central and peripheral lung squamous cell carcinoma showed clinical phenotype difference not only reflected in the incidence of lymph node metastasis, but also in gene expression profiles. Among them, HNF1A, CYP3A4, ALB, APOA1 at the key position of the differential gene interaction network and maybe as regulatory factors in the phenotypic difference.
Aged ; Carcinoma, Squamous Cell ; genetics ; Databases, Genetic ; Female ; Gene Expression Profiling ; Gene Regulatory Networks ; Humans ; Kaplan-Meier Estimate ; Lung Neoplasms ; genetics ; Male ; Middle Aged ; Smoking ; genetics

BackgroundThe hedgehog signaling system (HHS) plays an important role in the regulation of cell proliferation and differentiation during the embryonic phases. However, little is known about the involvement of HHS in the malignant transformation of cells. This study aimed to detect the role of HHS in the malignant transformation of human bronchial epithelial (16HBE) cells.

MethodsIn this study, two microfluidic chips were designed to investigate cigarette smoke extract (CSE)-induced malignant transformation of cells. Chip A contained a concentration gradient generator, while chip B had four cell chambers with a central channel. The 16HBE cells cultured in chip A were used to determine the optimal concentration of CSE for inducing malignant transformation. The 16HBE cells in chip B were cultured with 12.25% CSE (Group A), 12.25% CSE + 5 μmol/L cyclopamine (Group B), or normal complete medium as control for 8 months (Group C), to establish the in vitro lung inflammatory-cancer transformation model. The transformed cells were inoculated into 20 nude mice as cells alone (Group 1) or cells with cyclopamine (Group 2) for tumorigenesis testing. Expression of HHS proteins was detected by Western blot. Data were expressed as mean ± standard deviation. The t-test was used for paired samples, and the difference among groups was analyzed using a one-way analysis of variance.

ResultsThe optimal concentration of CSE was 12.25%. Expression of HHS proteins increased during the process of malignant transformation (Group B vs. Group A, F = 7.65, P < 0.05). After CSE exposure for 8 months, there were significant changes in cellular morphology, which allowed the transformed cells to grow into tumors in 40 days after being inoculated into nude mice. Cyclopamine could effectively depress the expression of HHS proteins (Group C vs. Group B, F = 6.47, P < 0.05) and prevent tumor growth in nude mice (Group 2 vs. Group 1, t = 31.59, P < 0.01).

ConclusionsThe activity of HHS is upregulated during the CSE-induced malignant transformation of 16HBE cells. Cyclopamine can effectively depress expression of HHS proteins in vitro and prevent tumor growth of the transformed cells in vivo.

Animals ; Cell Transformation, Neoplastic ; genetics ; metabolism ; Gene Expression Regulation, Neoplastic ; genetics ; physiology ; Hedgehog Proteins ; genetics ; metabolism ; Lab-On-A-Chip Devices ; Mice ; Mice, Inbred BALB C ; Mice, Nude ; Microfluidics ; Signal Transduction ; genetics ; physiology ; Smoke ; Smoking ; adverse effects

OBJECTIVE: We aimed to study the association between rs7525173, rs2236518, rs2493264 single nucleotide polymorphism (SNP) in the PRDM16 gene, smoking, alcohol exposures, and nonsyndromic cleft lip with or without cleft palate (NSCL/P). METHODS: A total of 157 case-parent trios were selected, and SNPs were genotyped by using ligase detection reaction (LDR) and direct sequencing methods. Transmission disequilibrium test (TDT) and linkage disequilibrium (LD) tests were con-ducted to analyze the data. A total of 1 710 patients with orofacial clefts and 956 healthy newborns were enrolled in the epidemiological survey. The smoking and drinking exposures of parents during early pregnancy were analyzed. RESULTS: The C allele at rs2236518 was over-transmitted for NSCPO (P<0.05). Statistical differences were observed among three factors, namely, maternal smoking, maternal passive smoking, and maternal drinking (P<0.05). CONCLUSIONS The rs2236518 at PRDM16 gene, maternal smoking, maternal passive smoking, and maternal drinking were closely related to the occurrence of NSCL/P.
Alcohol Drinking ; adverse effects ; Case-Control Studies ; Cleft Lip ; genetics ; Cleft Palate ; genetics ; DNA-Binding Proteins ; Environmental Exposure ; Female ; Genotype ; Humans ; Infant, Newborn ; Mothers ; Polymorphism, Single Nucleotide ; Pregnancy ; Smoking ; adverse effects ; Transcription Factors

Objective: To evaluate the association of family history with risk of major coronary events (MCE) and ischemic heart disease (IHD). Methods: After excluding participants with heart disease, stroke or cancer at baseline survey, a total of 485 784 participants from the China Kadoorie Biobank, who had no missing data on critical variables, were included in the analysis. Cox regression analysis was used to estimate the hazard ratios (HR) and 95% CI. Subgroup analyses were performed according to the baseline characteristics. Results: During a median of 7.2 years of follow-up, we documented 3 934 incident cases of MCE and 24 537 cases of IHD. In multivariable-adjusted models, family history was significantly associated with risk of MCE and IHD. The adjusted HRs (95%CI) were 1.41 (1.19-1.65) and 1.25 (1.18-1.33), respectively. History of disease among siblings was more strongly associated with early-onset MCE than parental history (HR=2.97, 95%CI: 1.80-4.88). Moreover, the association of family history with MCE and IHD was stronger in persons who were overweight or obesive, and the association between family history and MEC was stronger in smokers. Conclusion: This large-scale, prospective study indicated that family history was an independent risk factor for MCE and IHD in China. The intervention targeting major known lifestyle risk factors and the management of chronic diseases should be strengthened for Chinese population, especially for the individuals with family history were at high risk.
Asian People/statistics & numerical data* ; China/epidemiology* ; Coronary Disease/genetics* ; Humans ; Incidence ; Myocardial Ischemia/genetics* ; Overweight/ethnology* ; Proportional Hazards Models ; Prospective Studies ; Risk Assessment ; Risk Factors ; Smoking/ethnology*

Objective: To explore the SNP effects of patatin-like phospholipase domain which containing 3 (PNPLA3), transmembrane 6 superfamily member 2 (TM6SF2) gene, environmental effects of smoking, alcohol drinking and interaction between gene-gene, gene-environment and drinking-smoking on hepatitis B virus-associated hepatocellular carcinoma (HBV-HCC). Methods: We collected anticoagulant peripheral blood from patients of HBV-HCC, chronic hepatitis B (CHB), liver cirrhosis (LC) and from healthy controls to detect the single nucleotide polymorphism (SNP) of patatin-like phospholipase domain containing 3 (PNPLA3) gene loci rs738409 and transmembrane 6 superfamily member 2 (TM6SF2) gene loci rs58542926, using the flight mass spectrometry method. The optimal assignment value of gene polymorphisms was defined by using the online SNP stats. Hardy-Weinberg (H-W) balance was tested for SNP. Effects of the genetic and environmental factors to HBV-HCC were analyzed by using the multiple classification logistic regression method. The gene-gene, gene-smoking and alcohol drinking interaction effects were investigated by Fork-Life analysis and binary logistic regression methods. Results: The frequency distribution of CHB group rs738409 loci seemed not in conformity with the H-W balance (χ(2)=11.980, P<0.005). Two loci frequency distributions in the other groups were all in accordandce with the H-W balance. After adjusting for influences on age and sex and comparing to the healthy group, the rs58542926 mutation appeared as OR=1.659, 95%CI: 1.026-2.684, P=0.039, in the HBV-HCC group. When comparing to CHB group, the HBV-HCC group presented that drinking as OR=1.680, 95%CI: 1.121-2.519, P=0.012. When comparing to the LC group, the ORs of drinking and smoking were 1.539 (1.071-2.213) and 1.453 (1.005-2.099) respectively, in the HBV-HCC group. When comparing to the CHB+LC group, interactions between the HBV-HCC group were found rs738409 and rs58542926 on additive model OR=1.548 (U=1.885, P=0.029) and OR=1.658 (P=0.024) on logistic regression model while drinking was rs738409 on interaction additive model with OR=1.811(U=1.965, P=0.024). As for drinking and mutation of rs738409, the multiplication model of logistic regression showed no statistically significant differences. Interaction between smoking and drinking appeared as OR=1.756 (P<0.001) in the logistics regression multiplication model. Conclusions: Factors as mutation of TM6SF2, smoking and drinking all appeared as risk factors for HBV-HCC. Mutations of both PNPLA3 and TM6SF2, together with smoking and drinking all served as risk factors for HBV-HCC. However, the mutation of single PNPLA3 appeared as a protective factor on HBV-HCC.
Alcohol Drinking/adverse effects* ; Carcinoma, Hepatocellular/virology* ; Case-Control Studies ; Epistasis, Genetic ; Gene-Environment Interaction ; Genetic Predisposition to Disease ; Genotype ; Hepatitis B virus ; Hepatitis B, Chronic ; Humans ; Lipase/genetics* ; Liver Cirrhosis, Alcoholic/complications* ; Liver Neoplasms/virology* ; Membrane Proteins/genetics* ; Polymorphism, Single Nucleotide ; Smoking/adverse effects*

Asthma exacerbation (AE) usually denotes worsening of asthma symptoms that requires intense management to prevent further deterioration. AE has been reported to correlate with clinical and demographic factors, such as race, gender, and treatment compliance as well as environmental factors, such as viral infection, smoking, and air pollution. In addition, recent observations suggest that there are likely to be genetic factors specific to AE. Understanding genetic factors specific to AE is essential to develop therapy tailored for exacerbation-prone asthma. Here, we summarize the results of studies involving genetic risk factors for AE. To simplify and enhance understanding, we reviewed the studies according to the following categories: hypothesis-driven approaches, hypothesis-free approaches, gene-environment interactions, and pharmacogenetics.
Air Pollution ; Asthma* ; Compliance ; Continental Population Groups ; Demography ; Gene-Environment Interaction ; Genetics ; Genome-Wide Association Study ; Humans ; Pharmacogenetics ; Risk Factors ; Smoke ; Smoking

Recently, genetic variants in the WNT signaling pathway have been reported to affect the survival outcome of Caucasian patients with early stage non-small cell lung cancer (NSCLC). We therefore attempted to determine whether these same WNT signaling pathway gene variants had similar impacts on the survival outcome of NSCLC patients in a Korean population. A total of 761 patients with stages I-IIIA NSCLC were enrolled in this study. Eight variants of WNT pathway genes were genotyped and their association with overall survival and disease-free survival were analyzed. None of the eight variants were significantly associated with overall survival or disease-free survival. There were no differences in survival outcome after stratifying the subjects according to age, gender, smoking status, and histological type. These results suggest that genetic variants in the WNT signaling pathway may not affect the survival outcome of NSCLC in a Korean population.
Aged ; Asian Continental Ancestry Group/*genetics ; Carcinoma, Non-Small-Cell Lung/*genetics/mortality/pathology ; Demography ; Disease-Free Survival ; Female ; Genotype ; Humans ; Kaplan-Meier Estimate ; Lung Neoplasms/*genetics/mortality/pathology ; Male ; Middle Aged ; *Polymorphism, Single Nucleotide ; Republic of Korea ; Smoking ; Wnt Signaling Pathway/*genetics