Computed tomography evaluation of risk factors for an undesirable buccal split during sagittal split ramus osteotomy.

Sagittal split ramus osteotomy (SSRO) sometimes induces an irregular split pattern referred to as a bad split. We investigated the risk factors for bad splits in the buccal plate of the ramus during SSRO. Ramus morphology and bad splits in the buccal plate of the ramus were assessed using preoperative and postoperative computed tomography images. Of the 53 rami analyzed, 45 had a successful split, and 8 had a bad split in the buccal plate. Horizontal images at the height of the mandibular foramen showed that there were significant differences in the ratio of the forward thickness to the backward thickness of the ramus between patients with a successful split and those with a bad split. In addition, the distal region of the cortical bone tended to be thicker and the curve of the lateral region of the cortical bone tended to be smaller in the bad split group than in the good split group. These results indicated that a ramus shape in which the width becomes thinner towards the back frequently induces bad splits in the buccal plate of the ramus during SSRO, and more attention should be paid to patients who have rami of these shapes in future surgeries.

Genes related to cell wall metabolisms are targeted by miRNAs in immature tomato fruits under drought stress.

The metabolism of tomato fruits changes when plants experience drought stress. In this study, we investigated changes in microRNA (miRNA) abundance and detected 32 miRNAs whose expression changes in fruit. The candidate target genes for each miRNA were predicted from the differentially expressed genes identified by transcriptome analysis at the same fruit maturation stage. The predicted targeted genes were related to cell wall metabolisms, response to pathogens, and plant hormones. Among these, we focused on cell wall metabolism-related genes and performed a dual luciferase assay to assess the targeting of their mRNAs by their predicted miRNA. As a result, sly-miR10532 and sly-miR7981e suppress the expression of mRNAs of galacturonosyltransferase-10 like encoding the main enzyme of pectin biosynthesis, while sly-miR171b-5p targets ß-1,3-glucosidase mRNAs involved in glucan degradation. These results will allow the systematic characterization of miRNA and their target genes in the tomato fruit under drought stress conditions.

Transcriptomic and Metabolomic Analyses Provide Insights into the Upregulation of Fatty Acid and Phospholipid Metabolism in Tomato Fruit under Drought Stress.

Transcriptome and metabolome analysis in tomato (Solanum lycopersicum) fruits cultivated under drought conditions showed that drought stress promoted fatty acid synthesis and increased the content of fatty acids in fruits. The accumulation of some phospholipids composed of palmitic acid and oleic acid also was significantly increased, especially in seeds. Moreover, inositol, which is a component of cell membranes and cell walls, was increased through the activity of the myoinositol monophosphatase 1-mediated pathway. In mature fruits, the levels of metabolic regulators such as ß-alanine and 4-aminobutyric acid were elevated. These results showed that these compounds are drought-responsive and enhance drought tolerance and subsequently they could enhance the nutritional value and health benefits of tomato fruit.

Yeast-based reporter assay system for identifying the requirements of intramembrane proteolysis by signal peptide peptidase of Arabidopsis thaliana.

Signal peptide peptidase (SPP) is an aspartic protease with two active sites, YD and GXGD, in the transmembrane domain. SPP cleaves signal peptides, and the released fragments play key roles in the immune system, embryo development and protein turnover in cells. Despite SPP having an important function, a general system to identify the requirements of intramembrane proteolysis by SPP has not been developed because proteolysis occurs in the membrane. In this study, we first established a reporter assay system in yeast to verify the cleavage activity of the Arabidopsis thaliana SPP (AtSPP). Next, we screened candidate substrates of AtSPP from A. thaliana pollen and roots. In the pollen, 13 signal peptides with 'pollen' and 'cell wall' as gene ontology terms were selected. In the roots, mutants overexpressing AtSPP were constructed, and gene expression changes were compared with the wild-type. Nine signal peptides expressed in the roots were selected. Then we used the candidate substrates in our reporter assay system to determine the requirements for proteolysis by AtSPP. Fifteen of 22 signal peptides were cleaved by AtSPP. The absence of the positively charged amino acids, His and Lys on the C terminus of the signal sequence, was observed in cleaved substrates. Moreover, mutation of a helix breaker-to-Leu substitution in the intramembrane region in substrates prevented cleavage by AtSPP. These results indicated that substrates of AtSPP required the helix breaker structure to be cleaved.

Draft Genome Sequence of Zygosaccharomyces mellis CA-7, Isolated from Honey.

In this study, we report the draft genome sequence of Zygosaccharomyces mellis CA-7, isolated from purchased honey imported from Canada. The 10.19-Mb genome contains 4,963 gene models. To our knowledge, this annotated genome sequence is the first from the species Z. mellis and will contribute to a better understanding of the osmotolerance of microorganisms in high-sugar products.

Oryza sativa Brittle Culm 1-like 6 modulates ß-glucan levels in the endosperm cell wall.

The endosperm cell wall affects post-harvest grain quality by affecting the mechanical fragility and water absorption of the grain. Therefore, understanding the mechanism underlying endosperm cell wall synthesis is important for determining the growth and quality of cereals. However, the molecular machinery mediating endosperm cell wall biosynthesis is not well understood. In this study, we investigated the role of Oryza sativa Brittle Culm 1-like 6 (OsBC1L6), a member of the COBRA-like protein family, in cellulose synthesis in rice. OsBC1L6 mRNA was expressed in ripening seeds during endosperm enlargement. When OsBC1L6-RFP was expressed in Arabidopsis cell cultures, this fusion protein was transported to the plasma membrane. To investigate the target molecules of OsBC1L6, we analyzed the binding interactions of OsBC1L6 with cellohexaose and the analogs using surface plasmon resonance, determining that cellohexaose bound to OsBC1L6. The ß-glucan contents were significantly reduced in OsBC1L6-RNAi calli and OsBC1L6-deficient seeds from a Tos insertion mutant, compared to their wild-type counterparts. These findings suggest that OsBC1L6 modulates ß-glucan synthesis during endosperm cell wall formation by interacting with cellulose moieties on the plasma membrane during seed ripening.

Gene Expression Profiling in Short-Term Imbibition of Wheat: Tools for Dissecting of Pasting Properties of Imbibed Wheat Seeds.

Germination of wheat maximizes phytochemical content and antioxidant activity while altering chemical composition, gluten content, and pasting properties. This study investigated the effect of short-term imbibition on gene expression profiles and the physical and functional characteristics of wheat. Changes in gene expression profiles of wheat during short-term imbibition (0, 16, and 24 hr) were evaluated by DNA microarray analysis. Gene Ontology (GO) analysis was carried out to categorize the function of genes with altered expression. Genes related to cellulose and cell wall synthesis were upregulated by imbibition for 16 hr, whereas those associated with polysaccharide catabolism and nucleosome assembly were upregulated in the subsequent 8 hr. The genes related to proteases and gluten were expressed in dry seeds but disappeared after 16 hr of imbibition. Genes encoding α-amylase were not expressed in dry seeds whereas those encoding ß-amylase were expressed in dry seeds and downregulated by imbibition. According to quantitative real-time PCR and enzymatic activity assay, α-Amylase expression increased by imbibition and reached a maximum 24 hr after imbibition, with a corresponding increase in enzymatic activity. Pasting properties of flour made from wheat seeds imbibed for different times were decreased when seeds were imbibed for over 16 hr, by examination with Rapid Visco Analyzer. Gluten content did not significantly change until 24-hr imbibition, although expression of genes encoding gliadin and glutenin disappeared by 16-hr imbibition. The data indicated that it was possible to use 16-hr imbibed wheat, with up to the 50% w/w replacement of nonimbibed wheat.

Expression of protein arginine methyltransferase-5 in oral squamous cell carcinoma and its significance in epithelial-to-mesenchymal transition.

Protein arginine methyltransferases (PRMT) 5, a member of type II arginine methyltransferases, catalyzes the symmetrical dimethylation of arginine residues on histone and non-histone substrates. Although the overexpression of PRMT5 has been reported in various cancers, its role in oral squamous cell carcinoma (OSCC) has not been elucidated. In the present study, we immunohistochemically examined the expression of PRMT5 in surgically resected oral epithelial dysplasia (OED, n = 8), oral intraepithelial neoplasia (OIN)/carcinoma in situ (CIS) (n = 11) and OSCC (n = 52) with or without contiguous OED lesions. In the normal epithelium, PRMT5 was weakly expressed in the cytoplasm of basal layer cells. In OED, OIN/CIS, and OSCC, its expression consistently and uniformly increased in the cytoplasm of dysplastic and cancer cells. Moreover, nuclear and cytoplasmic localization was detected in the invasive front of cancer cells, particularly in cases showing poor differentiation or aggressive invasion patterns. The concomitant nuclear and cytoplasmic expression of PRMT5 correlated with the loss of E-cadherin and cytokeratin 17, and the upregulation of vimentin, features that are both indicative of epithelial-to-mesenchymal transition. PRMT5 may play a role from early oncogenesis through to the progression of OSCC, particularly in the aggressive mode of stromal invasion.

Establishment of highly metastatic KRAS mutant lung cancer cell sublines in long-term three-dimensional low attachment cultures.

Decreased cell-substratum adhesion is crucially involved in metastasis. Previous studies demonstrated that lung cancer with floating cell clusters in histology is more likely to develop metastasis. In the present study, we investigated whether cancer cells in long-term, three-dimensional low attachment cultures acquire high metastatic potential; these cells were then used to examine the mechanisms underlying metastasis. Two KRAS-mutated adenocarcinoma cell lines (A549 and H441) were cultured and selected on ultra-low attachment culture dishes, and the resulting cells were defined as FL (for floating) sublines. Cancer cells were inoculated into NOD/SCID mice via an intracardiac injection, and metastasis was evaluated using luciferase-based imaging and histopathology. In vitro cell growth (in attachment or suspension cultures), migration, and invasion were assayed. A whole genomic analysis was performed to identify key molecular alterations in FL sublines. Upon detachment on low-binding dishes, parental cells initially formed rounded spheroids with limited growth activity. However, over time in cultures, cells gradually formed smaller spheroids that grew slowly, and, after 3-4 months, we obtained FL sublines that regained prominent growth potential in suspension cultures. On ordinary dishes, FL cells reattached and exhibited a more spindle-shaped morphology than parental cells. No marked differences were observed in cell growth with attachment, migration, or invasion between FL sublines and parental cell lines; however, FL cells exhibited markedly increased growth potential under suspended conditions in vitro and stronger metastatic abilities in vivo. A genomic analysis identified epithelial-mesenchymal transition (EMT) and c-Myc amplification in A549-FL and H441-FL cells, respectively, as candidate mechanisms for metastasis. The growth potential of FL cells was markedly inhibited by lentiviral ZEB1 knockdown in A549-FL cells and by the inhibition of c-Myc through lentiviral knockdown or the pharmacological inhibitor JQ1 in H441-FL cells. Long-term three-dimensional low attachment cultures may become a useful method for investigating the mechanisms underlying metastasis mediated by decreased cell-substratum adhesion.

Bacillus cereus Response to a Proanthocyanidin Trimer, a Transcriptional and Functional Analysis.

Proanthocyanidins are abundant in peanut skin, and in this study, the antibacterial effects of a peanut skin extract (PSE) against food-borne bacteria were investigated to find its minimum inhibitory concentration. Food-borne gram-positive bacteria, and in particular Bacillus cereus, was more sensitive to PSE. In particular, the inhibitory activity of epicatechin-(4ߠ→ 6)-epicatechin-(2ߠ→ O→7, 4ߠ→ 8)-catechin (EEC), a proanthocyanidin trimer from peanut skin, against B. cereus was stronger than that of procyanidin A1, a proanthocyanidin dimer. DNA microarray analysis of B. cereus treated with EEC was carried out, with a finding that 597 genes were significantly up-regulated. Analysis of the up-regulated genes suggested that EEC disrupted the normal condition of the cell membrane and wall of B. cereus and alter its usual nutritional metabolism. Moreover, treatment of B. cereus with EEC inhibited glucose uptake, suggesting that EEC affects the cell-surface adsorption.

Recent Advances in the Management of Chronic Hepatitis B Including Suppression of Hepatocellular Carcinoma by Entecavir and Interferon.

At present, for adults with chronic hepatitis B virus (HBV) infection, two new analogues, entecavir (ETV) and tenofovir, are recommended as the first-line therapy by the EASL (European Association for the Study of the Liver), AASLD (American Association for the Study of Liver Diseases), and APASL (Asian Pacific Association for the Study of the Liver) guidelines. The use of pegylated interferon-α (PEG IFN-α) is recommended as the first-line therapy instead of standard IFN-α according to the above 3 guidelines. In this paper, the aim was to assess: (1) the long-term efficacy and safety as well as the resistance to ETV and tenofovir disoproxil fumarate (TDF); (2) the efficacy of PEG IFN-α; (3) the role of combination therapy with IFN plus two analogues, such as lamivudine and ETV; (4) the efficacy and safety of two analogues with cirrhosis, and (5) suppression of hepatocellular carcinoma (HCC) by ETV and IFN treatment. The results are as follows: (1) both ETV and TDF showed long-term efficacy and safety; (2) PEG IFN-α resulted in a greater decline in HBV DNA levels and a higher rate of HBeAg seroconversion; (3) combination therapy with IFN plus two analogues did not elevate the rate of sustained responses; (4) both ETV and TDF showed efficacy and safety with cirrhosis (ETV especially displayed efficacy and safety with decompensated cirrhosis), and (5) suppression of HCC was observed by ETV and IFN.

Frequent loss of the expression of multiple subunits of the SWI/SNF complex in large cell carcinoma and pleomorphic carcinoma of the lung.

The switch/sucrose non-fermenting (SWI/SNF) complex has recently emerged as a novel tumor suppressor in various human cancers. In the present study, we analyzed the expression of multiple SWI/SNF subunits in primary non-small cell lung cancer (NSCLC). A total of 133 NSCLC, consisting of 25 squamous cell carcinomas (SCC), 70 adenocarcinomas (AD), 16 large cell carcinomas (LC), and 22 pleomorphic carcinomas (PL), were immunohistochemically examined for the expression of BRG1, BRM, BAF47, ARID1A, and ARID1B. The frequency at which reductions in the expression of BRG1 were observed was significantly higher in the LC-PL group (13/38, 34.2%) than in the SCC-AD group (7/95, 7.4%). Similarly, the frequency at which reductions in the expression of BRM were observed was significantly higher in the LC-PL group (17/38, 44.7%) than in the SCC-AD group (14/95, 14.7%). The loss of the expression of ARID1A, ARID1B, and BAF47 was observed only in a fraction of NSCLC cases. Furthermore, the frequency at which the concurrent loss of multiple subunits of the SWI/SNF complex was observed was significantly higher in the LC-PL group (10/38, 26.3%) than in the SCC-AD group (8/95, 8.4%). Collectively, these results indicate that the loss of the SWI/SNF complex was related to dedifferentiation in NSCLC.

Development of an in vitro model for the analysis of bovine endometrium using simple techniques.

This study aimed to develop an in vitro model for the analysis of the bovine endometrium. Immunofluorescent staining revealed that the hetero-spheroids and the cultured explants showed almost similar structure in the localization of bovine endometrial epithelial cells and endometrial stromal cells, except the glandular-like structure of the epithelial cells inside the explants. Gelatin zymography revealed that the hetero-spheroids did not express matrix metalloproteinases (MMPs) after 4 days of culture, but strong MMP expressions were observed in the cultured explants until 7 days of culture. Additionally, expression of progesterone receptor (PR), estrogen receptor alpha (ERα), type I interferon receptor 1 (IFNAR1) and 2 (IFNAR2) messenger RNA was observed both in the homo- and hetero-spheroids. The expression of oxytocin receptor (OTR) mRNA in E2 and E2+P4 (1,3,5(10)-Estratrien-3, 17ß-diol + 4-Pregnen-3, 20-dinone) treated groups were significantly (P < 0.05) higher than that of the control group of spheroids. In case of cultured explants, the expression of PR and OTR mRNA were significantly (P < 0.05) higher in E2 treated groups compared to the control groups. Hepatocyte growth factor (HGF) mRNA expression was also higher in P4 treated groups at 10 days in culture (P < 0.05). In a nutshell, the in vitro model developed in this study for the analysis of the endometrium may provide a new platform for extensive research on bovine endometrial function.

Influence of clinical experience of the Macintosh laryngoscope on performance with the Pentax-AWS Airway Scope(®), a rigid video-laryngoscope, by paramedics in Japan.

PURPOSE: We sought to establish the clinical utility of the Pentax-AWS Airway Scope(®) (AWS) when used by paramedics to intubate the trachea, and to evaluate whether their performance was influenced by previous clinical experience with the Macintosh laryngoscope (ML). METHODS: Twenty paramedics attempted tracheal intubation using the AWS in five patients each in the operating room. We recorded the success rate, the number of intubation attempts, and the time for intubation and adverse events, and compared these based on the paramedics' previous clinical experience with the ML. Ten paramedics had no prior clinical experience of the ML (group A) and 10 had used it on more than 30 occasions (group B). RESULTS: The intubation success rate was 99 % (99/100). Notably, 96 % (47/49) of intubations were achieved on the first attempt by the inexperienced paramedics in group A, compared with 64 % (32/50) by the experienced paramedics in group B (p = 0.0001). The time to intubation (mean ± SD) was significantly shorter in group A than in group B (37 ± 24 vs. 48 ± 21 s, p = 0.002). There were marked variations in the times taken to intubate, but no apparent improvement as the intubators gained experience between their first and fifth cases. No complications were encountered in either group. CONCLUSION: We found that paramedics could achieve a high tracheal intubation success rate using the AWS independent of previous airway management experience. Better intubation performance with the AWS was observed in paramedics without clinical experience with the ML.

Immunohistochemical analysis of the expression of E-cadherin and ZEB1 in non-small cell lung cancer.

We performed an immunohistochemical analysis of the expression of zinc-finger E-box binding homeobox 1 (ZEB1), a master regulator of epithelial-mesenchymal transition (EMT), and determined its relationship with E-cadherin in 157 non-small cell lung carcinomas (93 adenocarcinomas, 36 squamous cell carcinomas, 18 large cell carcinomas, and 10 pleomorphic carcinomas). Although the expression of E-cadherin was low in the subset of adenocarcinomas (10%) and squamous cell carcinomas (11%), ZEB1 expression was only observed in one case of squamous cell carcinoma and none of the adenocarcinomas. In contrast, the low expression of E-cadherin (50% and 90%, respectively) and the positive expression of ZEB1 (11% and 50%, respectively) were more frequently observed in poorly differentiated carcinomas (large cell carcinomas and pleomorphic carcinomas). Overall, the expression of ZEB1 was inversely correlated with that of E-cadherin. Furthermore, the distribution of ZEB1-positive cancer cells was more restricted than in the area in which the expression of E-cadherin was lost, and the former was detected within the latter. We concluded that the expression of ZEB1 was not necessarily associated with the low expression of E-cadherin in lung adenocarcinomas and squamous cell carcinomas. The expression of ZEB1 correlated with an undifferentiated and/or sarcomatoid morphology that may occur in the late stage of EMT.

Characterization of the quality of imbibed soybean at an early stage of pre-germination for the development of a new protein food item.

This was a pilot study carried out to develop a new protein food item from imbibed soybean before germination. It identified the significance of a short stage after imbibition and before germination, and that vitamin C production was activated in as little as 16 h from the start of imbibition, without any influence on the soy protein quality or sensory acceptability, while longer imbibition caused the imbibed soybean to activate its phytophysiological metabolism for germination. DNA microarray analysis indicated that the genes for carbohydrate metabolism were up-regulated prior to 16 h, and that the expression rates of genes responsible for environmental factors were down-regulated. Thereafter, the expression rates of the genes associated with lipid metabolism and secondary metabolite production were changed. This information should contribute to a better understanding of how to develop a new soy protein item in pre-germination before active physiological processes begin.

Stromal fibroblasts are predictors of disease-related mortality in esophageal squamous cell carcinoma.

The growth, invasiveness and metastasis of human cancers are determined not only by cancer cells, but also by their microenvironment. Activated stromal fibroblasts promote tumor progression by secreting growth factors. In the present study, we focused on interrelations between cancer and fibroblasts, the main component of tumor stroma. We retrospectively analyzed the relations of mortality to clinical, pathological, and α-smooth muscle actin (α-SMA) characteristics in 97 consecutive patients with esophageal squamous cell carcinoma (ESCC). In vitro, we used TE-11, KYSE150 and KYSE220 ESCC cell lines and isolated esophageal stromal fibroblasts, some of which were immortalized. Migration assays were conducted to assess the effects of fibroblasts on cancer-cell migration and 3-dimensional organotypic cultures. In vivo, TE-11 and KYSE220 cells plus immortalized fibroblasts were co-transplanted subcutaneously in Nod/Scid mice to assess the effects of fibroblasts on tumorigenicity. Clinicopathologically, the α-SMA expression of cancer stroma was correlated with venous invasion (p<0.01), nodal involvement (p=0.02), recurrence (p=0.01), and was a predictor of survival in patients with stage I and II ESCC (p=0.04). In vitro, the presence of fibroblasts strongly promoted the migration of TE-11, KYSE150 and KYSE220 cells. On organotypic culture, stromal invasion was observed only in the presence of immortalized fibroblasts. In vivo, tumors developed or grew in a fibroblast­dependent manner after implantation. Our findings provide evidence that stromal fibroblasts and tumor cells interact to promote tumor progression in ESCC. In patients with earlier stage ESCC, α-SMA may be a predictor of mortality. Inhibition of paracrine systems associated with tumor fibroblasts may slow or reverse tumor progression, potentially leading to the development of new targeted therapies.

Peanut-skin polyphenols, procyanidin A1 and epicatechin-(4 ß â†’ 6)-epicatechin-(2 ß â†’ O → 7, 4 ß â†’ 8)-catechin, exert cholesterol micelle-degrading activity in vitro.

We identified epicatechin-(4 ß â†’ 6)-epicatechin-(2 ß â†’ O → 7, 4 ß â†’ 8)-catechin (EEC) in the skin of the peanut (Arachis hypogaea L.). EEC (a trimer) showed more potent cholesterol micelle-degrading activity than procyanidin A1 (a dimer) did in vitro. The hypercholesterolemia suppressing effect of a peanut skin polyphenol on rats fed high-cholesterol diet in our preceding experiments might thus have been due primarily to a micelle degrading effect in the intestine.

Lung cancer with loss of BRG1/BRM, shows epithelial mesenchymal transition phenotype and distinct histologic and genetic features.

BRG1 and BRM, two core catalytic subunits in SWI/SNF chromatin remodeling complexes, have been suggested as tumor suppressors, yet their roles in carcinogenesis are unclear. Here, we present evidence that loss of BRG1 and BRM is involved in the progression of lung adenocarcinomas. Analysis of 15 lung cancer cell lines indicated that BRG1 mutations correlated with loss of BRG1 expression and that loss of BRG1 and BRM expression was frequent in E-cadherin-low and vimentin-high cell lines. Immunohistochemical analysis of 93 primary lung adenocarcinomas showed loss of BRG1 and BRM in 11 (12%) and 16 (17%) cases, respectively. Loss of expression of BRG1 and BRM was frequent in solid predominant adenocarcinomas and tumors with low thyroid transcription factor-1 (TTF-1, master regulator of lung) and low cytokeratin7 and E-cadherin (two markers for bronchial epithelial differentiation). Loss of BRG1 was correlated with the absence of lepidic growth patterns and was mutually exclusive of epidermal growth factor receptor (EGFR) mutations. In contrast, loss of BRM was found concomitant with lepidic growth patterns and EGFR mutations. Finally, we analyzed the publicly available dataset of 442 cases and found that loss of BRG1 and BRM was frequent in E-cadherin-low, TTF-1-low, and vimentin-high cases and correlated with poor prognosis. We conclude that loss of either or both BRG1 and BRM is involved in the progression of lung adenocarcinoma into solid predominant tumors with features of epithelial mesenchymal transition and loss of the bronchial epithelial phenotype. BRG1 loss was specifically involved in the progression of EGFR wild-type, but not EGFR-mutant tumors.