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Due to their aggressive behavior, local recurrence and distant metastasis, survival rate of advanced stage of the patients with head and neck squamous cell carcinoma (HNSCC) is very poor. Currently available epidermal growth factor receptor (EGFR)-targeted therapies are not considered curative for HNSCC. Therefore, novel approaches for identification of therapeutic targets in HNSCC are needed. All members of the miRNA-29 family (miR-29a, miR-29b, and miR-29c) were downregulated in HNSCC tissues by analysis of RNA-sequencing based microRNA (miRNA) expression signature. Ectopic expression of mature miRNAs demonstrated that the miR-29 family inhibited cancer cell migration and invasion by HNSCC cell lines. Comprehensive gene expression studies and in silico database analyses were revealed that integrin β1 (ITGB1) was regulated by the miR-29 family in HNSCC cells. Overexpression of ITGB1 was confirmed in HNSCC specimens, and high expression of ITGB1 significantly predicted poor survival in patients with HNSCC (p = 0.00463). Knockdown of ITGB1 significantly inhibited cancer cell migration and invasion through regulating downstream of ITGB1-mediated oncogenic signalling. In conclusion, regulation of the antitumor miR-29 family affected integrin-mediated oncogenic signalling to modulate HNSCC pathogenesis; these molecules may be novel therapeutic targets for HNSCC.

For patients with head and neck squamous cell carcinoma (HNSCC), survival rates have not improved due to local recurrence and distant metastasis. Current targeted molecular therapies do not substantially benefit HNSCC patients. Therefore, it is necessary to use advanced genomic approaches to elucidate the molecular mechanisms underlying the aggressiveness of HNSCC cells. Analysis of our microRNA (miRNA) expression signature by RNA sequencing showed that the miR-199 family (miR-199a-5p, miR-199a-3p, miR-199b-5p and miR-199b-3p) was significantly reduced in cancer tissues. Ectopic expression of mature miRNA demonstrated that all members of the miR-199 family inhibited cancer cell migration and invasion by HNSCC cell lines (SAS and HSC3). These findings suggested that both passenger strands and guide strands of miRNA are involved in cancer pathogenesis. In silico database and genome-wide gene expression analyses revealed that the gene coding for integrin α3 (ITGA3) was regulated by all members of the miR-199 family in HNSCC cells. Knockdown of ITGA3 significantly inhibited cancer cell migration and invasion by HNSCC cells. Moreover, overexpression of ITGA3 was confirmed in HNSCC specimens, and high expression of ITGA3 predicted poorer survival of the patients (P = 0.0048). Our data revealed that both strands of pre-miR-199a (miR-199a-5p and miR-199a-3p) and pre-miR-199b (miR-199b-5p and miR-199b-3p) acted as anti-tumor miRNA in HNSCC cells. Importantly, the involvement of passenger strand miRNA in the regulation of cellular processes is a novel concept in RNA research. Novel miRNA-based approaches for HNSCC can be used to identify potential targets for the development of new therapeutic strategies.

// Keiichi Koshizuka 1, 2 , Nijiro Nohata 3 , Toyoyuki Hanazawa 2 , Naoko Kikkawa 2 , Takayuki Arai 1 , Atsushi Okato 1 , Ichiro Fukumoto 1, 2 , Koji Katada 2 , Yoshitaka Okamoto 2 , Naohiko Seki 1 1 Department of Functional Genomics, Chiba University Graduate School of Medicine, Chuo-Ku, Chiba, Japan 2 Department of Otorhinolaryngology/Head and Neck Surgery, Chiba University Graduate School of Medicine, Chiba, Japan 3 Moores Cancer Center, University of California, San Diego, La Jolla, CA, USA Correspondence to: Naohiko Seki, email: naoseki@faculty.chiba-u.jp Keywords: microRNA, miR-150, ITGA3, ITGA6, TNC Received: January 17, 2017 Accepted: March 09, 2017 Published: March 17, 2017 ABSTRACT We adopted into RNA-sequencing technologies to construct the microRNA (miRNA) expression signature of head and neck squamous cell carcinoma (HNSCC). Our signature revealed that a total of 160 miRNAs (44 upregulated and 116 downregulated) were aberrantly expressed in cancer tissues. Expression of miR-150-5p (guide strand miRNA) and miR-150-3p (passenger strand miRNA) were significantly silenced in cancer tissues, suggesting both miRNAs act as antitumor miRNAs in HNSCC cells. Ectopic expression of mature miRNAs, miR-150-5p and miR-150-3p inhibited cancer cell aggressiveness. Low expression of miR-150-5p and miR-150-3p predicted significantly shorter overall survival in patients with HNSCC ( P = 0.0091 and P = 0.0386) by Kaplan–Meier survival curves analyses. We identified that integrin α3 ( ITGA3 ), integrin α6 ( ITGA6 ), and tenascin C ( TNC ) were coordinately regulated by these miRNAs in HNSCC cells. Knockdown assays using siRNAs showed that ITGA3, ITGA6 and TNC acted as cancer promoting genes in HNSCC cells. Moreover, ITGA3, ITGA6 , and TNC alterations were associated with significantly poorer overall survival ( P = 0.0177, P = 0.0237, and P = 0.026, respectively). Dual strands of pre-150 ( miR-150-5p and miR-150-3p ) functioned as antitumor miRNAs based on the miRNA expression signature of HNSCC. Identification of antitumor miR-150 -mediated RNA networks may provide novel insights into pathogenesis of HNSCC.

Analysis of the microRNA (miRNA) expression signature of head and neck squamous cell carcinoma (HNSCC) based on RNA sequencing showed that dual strands of pre‑miR‑145 (miR‑145‑5p, guide strand; and miR‑145‑3p, passenger strand) were significantly reduced in cancer tissues. In miRNA biogenesis, passenger strands of miRNAs are degraded and have no biological activities in cells. The aims of this study were to investigate the functional significance of the passenger strand of miR‑145 and to identify miR‑145‑3p‑regulated oncogenic genes in HNSCC cells. Expression levels of miR‑145‑5p and miR‑145‑3p were significantly downregulated in HNSCC tissues and cell lines (SAS and HSC3 cells). Ectopic expression of miR‑145‑3p inhibited cancer cell proliferation, migration and invasion, similar to miR‑145‑5p, in HNSCC cells. Myosin 1B (MYO1B) was directly regulated by miR‑145‑3p, and knockdown of MYO1B by siRNA inhibited cancer cell aggressiveness. Overexpression of MYO1B was confirmed in HNSCC clinical specimens by analysis of protein and mRNA levels. Interestingly, high expression of MYO1B was associated with poor prognosis in patients with HNSCC by analysis of The Cancer Genome Atlas database (p=0.00452). Our data demonstrated that the passenger strand of miR‑145 acted as an antitumor miRNA through targeting MYO1B in HNSCC cells. The involvement of dual strands of pre‑miR‑145 (miR‑145‑5p and miR‑145‑3p) in the regulation of HNSCC pathogenesis is a novel concept in present RNA research.

Objective Our recent studies have revealed that both strands of pre-miRNAs, the guide strand and the passenger strand, are involved in cancer pathogenesis. Analyses of miRNA expression signatures by RNA sequencing in head and neck squamous cell carcinoma (HNSCC) showed that both of the strands of pre-miR-150 (miR-150-5p and miR-150-3p) were significantly downregulated, and that these miRNAs acted as antitumor miRNAs in HNSCC cells. The aim of this study was to identify oncogenic genes in HNSCC cells that were regulated by miR-150-5p and miR-150-3p. Methods Genome-wide gene expression studies, in silico analyses and dual-luciferase reporter assays were carried out to predict miR-150-5p and miR-150-3p regulation in HNSCC cells. Knockdown assay was applied to investigate the functional significance of the target gene. Overall patient survival as a function of target gene expression was estimated by The Cancer Genome Atlas (TCGA) database. Results A total of 19 genes were putative targets of both miR-150-5p and miR-150-3p regulation. Among them, SPOCK1 (SPARC/osteonectin, cwcv and kazal-like domains proteoglycan 1) was directly regulated by both miRNAs in HNSCC cells. Knockdown studies using si-SPOCK1 showed that expression of SPOCK1 enhanced HNSCC cell aggressiveness. Overexpression of SPOCK1/SPOCK1 was confirmed in HNSCC clinical specimens. Interestingly, analysis of a large number of patients in the TCGA database (n = 248) demonstrated that patients with high SPOCK1 expression had significantly shorter survival than did those with low SPOCK1 expression (P = 0.0003). Moreover, 15 pathways were identified as SPOCK1-mediated downstream pathways. Conclusion Downregulation of both strands of pre-miR-150 (miR-150-5p and miR-150-3p) and overexpression of SPOCK1 contribute to the aggressive nature of HNSCC. The involvement of passenger strand miRNA in the regulation of HNSCC pathogenesis is a novel concept in RNA research.

平成15年1月から平成19年12月までの5年間に当科で治療（原則手術）を施行した耳下腺腫瘍100例のうち，良性は82.0%，悪性は18.0%であった。良性では多形腺腫（54.9%），ワルチン腫瘍（30.5%）と，これらで良性の約85%を占めた。悪性の組織型は多彩であった。耳下腺腫瘍全体でみても，悪性を1グループとして加えるとこれら3グループで88%を占めた。さらに基底細胞腺腫を加えて96%とし，臨床所見などをふまえた上で画像診断を中心にして，術前（病理組織）診断フローチャートを作成した。画像診断ではMRI（造影）と核医学検査（特に99mTcO4）が有用であった。67Ga-citrateは有用とは言えなかった。手術適応に迷う症例や腫瘍摘出後の再建法選択などで準備を要するものについては，さらに超音波ガイド下のFNAや，どうしても必要な場合に限り，手術前2週間以内の針生検を加え診断をさらに絞っている。

Sleep breathing patterns are altered by nasal obstruction and respiratory events. This study aimed to describe the relationships between specific sleep oral flow (OF) patterns, nasal airway obstruction, and respiratory events.Nasal flow and OF were measured simultaneously by polysomnography in 85 adults during sleep. OF was measured 2 cm in front of the lips using a pressure sensor.OF could be classified into three patterns: postrespiratory event OF (postevent OF), during-respiratory event OF (during-event OF), and spontaneous arousal-related OF (SpAr-related OF). Postevent OFs begin at the end of airflow reduction, are preceded by respiratory arousal, and are accompanied by postapneic hyperventilation; during-event OFs occur during nasal flow reduction; and SpAr-related OFs to OF begin during stable breathing, and are preceded by spontaneous arousal but are rarely accompanied by apnea/hypopnea. Multivariate regression showed that nasal obstruction was predictive of SpAr-related OF. The relative frequency of SpAr-related OF events was negatively correlated with the apnea-hypopnea index. The fraction of SpAr-related OF duration relative to total OF duration was significantly greater in patients with nasal obstruction than in those without.SpAr-related OF was associated with nasal obstruction, but not respiratory events. This pattern thus functions as a "nasal obstruction bypass", mainly in normal subjects and patients with mild sleep disordered breathing (SDB). By contrast, the other two types were related to respiratory events and were typical patterns seen in patients with moderate and severe SDB.

Environmental challenge chambers (ECC) have been used to expose people to pollen allergens within a stable atmosphere and to examine the efficacy of treatment. Although pollinosis is one of the typical IgE-mediated type I allergic diseases, allergic inflammation is thought to contribute to the fundamental pathogenesis and prophylactic treatment may reduce exacerbations of pollinosis. The purpose of this study was to compare the efficacy of prophylactic treatment with nasal steroid (mometasone furoate nasal spray) or an antihistamine (fexofenadine) in the control of cedar pollinosis using the ECC. In a randomized, double-blind two-way crossover study, 48 patients received nasal steroid or antihistamine for 7 consecutive days (days 1-7). On day 8, patients were exposed to cedar pollen (8000 grains/m(3)) in the ECC for 3 hours. Nasal symptoms induced by pollen exposure were assessed. Total nasal symptom scores (TNSSs) during the exposure in the ECC were not significantly different between the antihistamine and the nasal steroid groups. Nasal symptoms induced by pollen exposure using the ECC persisted for up to 3 days. TNSSs after pollen exposure on days 8-11 were significantly lower in the nasal steroid group compared with the antihistamine group. Prophylactic treatment with nasal steroid is more effective than antihistamine against pollinosis, particularly in the late phase. Clinical trial registration JAPIC CTI 101182 (www.clinicaltrials.jp/user/ctiMain_e.jsp).

Head and neck squamous cell carcinoma (HNSCC) is usually found at a late stage and distant metastasis occurs at high frequency; therefore, novel prognostic markers are needed. This study was aimed to identify novel tumor markers in HNSCC. We identified 65 proteins which were significantly increased or decreased in the tumors by 2D-DIGE using 12 HNSCC and adjacent non-cancer tissues. Western blotting and immunohistochemical analysis confirmed that the expression of plectin was significantly increased in most cancer tissues as compared with non-cancer tissues. Strikingly, the suppression of endogenous plectin using siRNA inhibited the proliferation, migration and invasion of HNSCC cells and down-regulated Erk 1/2 kinase. Furthermore, immunohistochemistry using paraffin-embedded tissues from 62 patients showed not only that the frequency of recurrence was correlated with the plectin expression but that the prognosis of patients with a high plectin was extremely poor. Moreover, the survival rate of patients with a high plectin was significantly lower than that of patients with low E-cadherin levels, which is known to correlate with the poor prognosis of HNSCC. Our findings suggest that plectin promotes the migration and invasion of HNSCC cells through activation of Erk 1/2 kinase and is a potential prognostic biomarker of HNSCC.

As to X-ray examination of the naviculare and tibiale externum bones, the authors worked out a way to project them obliquely as follows; A new X-ray projection technique for taking shots anteversio oblique at an angle of 28 degrees to the footbottom. The reason for determining the angle at 28 degrees is that it corresponds to the degree at which the space between the tibiale externum and naviculare can be most clearly projected. It was the average degree out of 21 samples taken. Using this oblique projection technique, the space along these two bones does not overlap and its shadow is projected sharply.The figures of the naviculare and the tibiale externum of 540 adults (25-74 years old) on roentogenograph by means of such oblique projection are now classified as the Oneda-Katada classification in 6 divisions (A', A, B, C, D and E types) and 2 subdivisions (+S1, or +S2 and L, M, or S types) (cf. Figs. 8 and 9 and Tables 2, 3 and 4), referred to the dorso-plantar X-ray projection figures.Further, using this oblique projection method, the authors classified the both bones of 1, 159 growing children from age 6-19 years old (cf. Tables 5, 6 and 7). It was concluded that the frequency of occurrensce of each type was about one or two years earlier for the female than for the male. Due to the frequency of occurrence of each type in each age group, the growth of the naviculare and the tibiale externum is assumed by the authors to be as Fig. 11.It must by assumed therefore, that in some cases growth ceases in each type and that the bones remain unchanged.

Background We previously reported that periplakin (PPL) is downregulated in human esophageal cancer tissues compared to the adjacent non-cancer epithelium. Thus PPL could be a useful marker for detection of early esophageal cancer and evaluation of tumor progression, but largely remains unknown in this field. To investigate PPL involvement in carcinogenesis, tumor progression, cellular movement or attachment activity, siRNAs against PPL were transfected into pharyngeal squamous cancer cell lines and their effects on cellular behaviours were examined. Results PPL knockdown appeared to decrease tumor cell growth together with G2/M phase accumulation in cells attached to a culture dish. However, the extent of cell growth suppression, evaluated by the number of cells attached to the culture dish, was too distinctive to be explained only by cell cycle delay. Importantly, PPL knockdown suppressed cellular movement and attachment to the culture dish accompanied by decreased pAktSer473 phosphorylation. Additionally, LY294002, a PI3K inhibitor that dephosphorylates pAktSer473, significantly suppressed D562 cell migration. Thus PPL potentially engages in cellular movement al least partly via the PI3K/Akt axis. Conclusions PPL knockdown is related to reduced cellular movement and attachment activity in association with PI3K/Akt axis suppression, rather than malignant progression in pharyngeal cancer cells.