Objectives Evaluate the cytotoxicity and genotoxicity of short- and long-term e-cigarette vapor exposure on a panel of normal epithelial and head and neck squamous cell carcinoma (HNSCC) cell lines. Materials and Methods HaCaT, UMSCC10B, and HN30 were treated with nicotine-containing and nicotine-free vapor extract from two popular e-cigarette brands for periods ranging from 48 hours to 8 weeks. Cytotoxicity was assessed using Annexin V flow cytometric analysis, trypan blue exclusion, and clonogenic assays. Genotoxicity in the form of DNA strand breaks was quantified using the neutral comet assay and γ-H2AX immunostaining. Results E-cigarette-exposed cells showed significantly reduced cell viability and clonogenic survival, along with increased rates of apoptosis and necrosis, regardless of e-cigarette vapor nicotine content. They also exhibited significantly increased comet tail length and accumulation of γ-H2AX foci, demonstrating increased DNA strand breaks. Conclusion E-cigarette vapor, both with and without nicotine, is cytotoxic to epithelial cell lines and is a DNA strand break-inducing agent. Further assessment of the potential carcinogenic effects of e-cigarette vapor is urgently needed.

BackgroundAlcohol consumption is a well-established risk factor for head and neck squamous cell carcinoma (HNSCC); however, the molecular mechanisms by which alcohol promotes HNSCC pathogenesis and progression remain poorly understood. Our study sought to identify microRNAs that are dysregulated in alcohol-associated HNSCC and investigate their contribution to the malignant phenotype.MethodUsing RNA-sequencing data from 136 HNSCC patients, we compared the expression levels of 1,046 microRNAs between drinking and non-drinking cohorts. Dysregulated microRNAs were verified by qRT-PCR in normal oral keratinocytes treated with biologically relevant doses of ethanol and acetaldehyde. The most promising microRNA candidates were investigated for their effects on cellular proliferation and invasion, sensitivity to cisplatin, and expression of cancer stem cell genes. Finally, putative target genes were identified and evaluated in vitro to further establish roles for these miRNAs in alcohol-associated HNSCC.ResultsFrom RNA-sequencing analysis we identified 8 miRNAs to be significantly upregulated in alcohol-associated HNSCCs. qRT-PCR experiments determined that among these candidates, miR-30a and miR-934 were the most highly upregulated in vitro by alcohol and acetaldehyde. Overexpression of miR-30a and miR-934 in normal and HNSCC cell lines produced up to a 2-fold increase in cellular proliferation, as well as induction of the anti-apoptotic gene BCL-2. Upon inhibition of these miRNAs, HNSCC cell lines exhibited increased sensitivity to cisplatin and reduced matrigel invasion. miRNA knockdown also indicated direct targeting of several tumor suppressor genes by miR-30a and miR-934.ConclusionsAlcohol induces the dysregulation of miR-30a and miR-934, which may play crucial roles in HNSCC pathogenesis and progression. Future investigation of the alcohol-mediated pathways effecting these transformations will prove valuable for furthering the understanding and treatment of alcohol-associated HNSCC.Electronic supplementary materialThe online version of this article (doi:10.1186/s12943-015-0452-8) contains supplementary material, which is available to authorized users.

Head and neck squamous cell carcinoma (HNSCC) is an aggressive disease marked by frequent recurrence and metastasis and stagnant survival rates. To enhance molecular knowledge of HNSCC and define a non-coding RNA (ncRNA) landscape of the disease, we profiled the transcriptome-wide dysregulation of long non-coding RNA (lncRNA), microRNA (miRNA), and PIWI-interacting RNA (piRNA) using RNA-sequencing data from 422 HNSCC patients in The Cancer Genome Atlas (TCGA). 307 non-coding transcripts differentially expressed in HNSCC were significantly correlated with patient survival, and associated with mutations in TP53, CDKN2A, CASP8, PRDM9, and FBXW7 and copy number variations in chromosomes 3, 5, 7, and 18. We also observed widespread ncRNA correlation to concurrent TP53 and chromosome 3p loss, a compelling predictor of poor prognosis in HNSCCs. Three selected ncRNAs were additionally associated with tumor stage, HPV status, and other clinical characteristics, and modulation of their expression in vitro reveals differential regulation of genes involved in epithelial-mesenchymal transition and apoptotic response. This comprehensive characterization of the HNSCC non-coding transcriptome introduces new layers of understanding for the disease, and nominates a novel panel of transcripts with potential utility as prognostic markers or therapeutic targets.

Objective While tobacco smoking is a well-known risk factor for head and neck squamous cell carcinoma (HNSCC), the molecular mechanisms underlying tobacco-induced HNSCC remain unclear. This study sought to comprehensively identify microRNA (miRNA) alterations and evaluate their clinical relevance in smoking-induced HNSCC pathogenesis and progression. Materials and methods Using small RNA-sequencing data and clinical data from 145 HNSCC patients, we performed a series of differential expression and correlation analyses to identify a panel of tobacco-dysregulated miRNAs associated with key clinical characteristics in HNSCC. We then examined the expression patterns of these miRNAs in normal epithelial cell lines following exposure to cigarette smoke extract. Results Our analyses revealed distinct panels of miRNAs to be dysregulated with smoking status and associated with additional clinical features, including tumor stage, metastasis, anatomic site, and patient survival. The differential expression of key miRNAs, including miR-101, miR-181b, miR-486, and miR-1301, was verified in cigarette-treated epithelial cell lines, suggesting their potential roles in the early development of smoking-related HNSCCs. Conclusion Specific alterations in miRNA expression may be traced to tobacco use and are associated with important HNSCC clinical characteristics. Future studies of these miRNAs may be valuable for furthering the understanding and targeted treatment of smoking-associated HNSCC.

Objective Smoking remains a primary etiological factor in head and neck squamous cell carcinoma (HNSCC). Given that non-coding RNAs (ncRNAs), including PIWI-interacting RNAs (piRNAs), have emerged as mediators of initiation and progression in head and neck malignancies, we undertook a global study of piRNA expression patterns in smoking-associated HNSCC. Materials and Methods Using RNA-sequencing data from 256 current smoker and lifelong nonsmoker samples in The Cancer Genome Atlas (TCGA), we analyzed the differential expression patterns of 27,127 piRNAs across patient cohorts stratified by tobacco use, with HPV16 status and tumor status taken into account. We correlated their expression to clinical characteristics and to smoking-induced alteration of PIWI proteins, the functional counterparts of piRNAs. Finally, we correlated our identified piRNAs and PIWI proteins to known chromosomal aberrations in HNSCC to understand their wider-ranging genomic effects. Results and Conclusion Our analyses implicated a 13-member piRNA panel in smoking-related HNSCC, among which NONHSAT123636 and NONHSAT113708 are associated with tumor stage, NONHSAT067200 with patient survival, and NONHSAT081250 with smoking-altered PIWIL1 protein expression. 6 piRNAs as well as PIWIL1 correlated with genomic alterations common to HNSCC, including TP53 mutation, TP53-3p co-occurrence, and 3q26, 8q24, and 11q13 amplification. Collectively, our findings provide novel insights into the etiology-specific piRNA landscape of smoking-induced HNSCC.

Alcohol consumption and chronic hepatitis B virus (HBV) infection are two well-established risk factors for Hepatocellular carcinoma (HCC); however, there remains a limited understanding of the molecular pathway behind the pathogenesis and progression behind HCC, and how alcohol promotes carcinogenesis in the context of HBV+ HCC. Using next-generation sequencing data from 130 HCC patients and 50 normal liver tissues, we identified a panel of microRNAs that are significantly dysregulated by alcohol consumption in HBV+ patients. In particular, two microRNAs, miR-944 and miR-223-3p, showed remarkable correlation with clinical indication and genomic alterations. We confirmed the dysregulation of these two microRNAs in liver cell lines treated by alcohol and acetaldehyde, and showed that manipulation of miR-223-3p and miR-944 expression induces significant changes in cellular proliferation, sensitivity to doxorubicin, and the expression of both direct-binding and downstream mRNA targets. Together, the results of this study suggest that alcohol consumption in HBV+ HCCs regulates microRNAs that likely play previously uncharacterized roles in the alcohol-associated carcinogenesis of HCC, and future studies of these microRNAs may be valuable for furthering the understanding and treatment of alcohol and HBV-associated HCC.

Hepatocellular carcinoma (HCC) is one of the leading causes of cancer-related deaths because of frequent late detection and poor therapeutic outcomes, necessitating the need to identify effective biomarkers for early diagnosis and new therapeutic targets for effective treatment. Long noncoding RNAs (lncRNAs) have emerged as promising molecular markers for diagnosis and treatment. Through analysis of patient samples from The Cancer Genome Atlas database, we identified putative lncRNAs dysregulated in HCC and by its risk factors, hepatitis infection and alcohol consumption. We identified 184 lncRNAs dysregulated in HCC tumors versus paired normal samples, 53 lncRNAs dysregulated in alcohol-drinking patients with hepatitis B, and 5, 456 lncRNAs dysregulated in patients with hepatitis infection. A panel of these candidate lncRNAs’ expressions correlated significantly with patient survival, clinical variables, and known genomic alteration in HCC. Two most significantly dysregulated lncRNAs in our computational analysis, lnc-CFP-1:1 and lnc-CD164L2-1:1, were validated in vitro to be dysregulated by alcohol. Our findings suggest that lncRNAs dysregulated by different etiologies of HCC serve as potential disease markers and can be further investigated to develop personalized prevention, diagnosis, and treatment strategies.

Background: We have previously reported the 1-year outcomes of arthroscopic suprapectoral biceps tenodesis (ASPBT) versus open subpectoral biceps tenodesis (OSPBT) for the management of long head of the biceps tendon (LHBT) pathology. While patients had similar 1-year biceps muscle strength and pain, longer-term functional outcomes are unknown. Purpose: To directly compare clinical outcomes of ASPBT versus OSPBT with interference screw fixation, distal to the bony bicipital groove, at a minimum of 2 years’ follow-up. Study Design: Randomized controlled trial; Level of evidence, 1. Methods: A total of 85 patients undergoing biceps tenodesis (BT) for LHBT disease were randomized into the ASPBT or OSPBT group. Both techniques utilized polyether ether ketone inference screws for tenodesis fixation. Patients completed American Shoulder and Elbow Surgeons (ASES), Constant subjective, and Single Assessment Numeric Evaluation (SANE) questionnaires preoperatively and again at 6 months, 12 months, and at the final follow-up at a minimum of 24 months. Results: A total of 73 patients (37/42 randomized to ASPBT [88%]; 36/42 randomized to OSPBT [86%]) with a mean age of 50.4 ± 10.3 years and a mean body mass index of 29 ± 7.9 were included in clinical outcome analyses. The mean final follow-up was 2.9 years (ASPBT, 3 years; OSPBT, 2.8 years [range 2-5.2 years]). Comparison of demographic characteristics and intraoperative findings showed no significant differences in age, sex, concomitant procedures, and rotator cuff disease. No statistically significant differences in the ASES ( P = .25), Constant subjective ( P = .52), and SANE scores ( P = .61) were found at the final follow-up. Clinical outcomes scores showed no significant improvement from a mean of 12.6 months to the final follow-up at 34.5 months (ASPBT: ASES, P = .43; Constant, P = .25; SANE, P = .45 vs OSPBT: ASES, P = .65; Constant, P = .78; SANE, P = .70). No patients required revision of BT in either group. Conclusion: This study reported a minimum of 2-year follow-up of patients undergoing ASPBT or OSPBT, utilizing the same interference screw technique, for the management of LHBT pathology in the setting of concomitant shoulder procedures. There were no significant differences in patient-reported outcomes and complication rates found at any time point. Registration: NCT02192073 (ClinicalTrials.gov identifier).