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Background: Despite the better prognosis given by surgical resection and chemotherapy in low-grade glioma (LGG), progressive transformation is still a huge concern. In this case, the S100A gene family, being capable of regulating inflammatory responses, can promote tumor development. Methods: The analysis was carried out via ONCOMINE, GEPIA, cBioPortal, String, GeneMANIA, WebGestalt, LinkedOmics, TIMER, CGGA, R 4.0.2 and immunohistochemistry. Results: S100A2, S100A6, S100A10, S100A11, and S100A16 were up-regulated and S100A1 and S100A13 were down-regulated in LGG compared to normal tissues. S100A3, S100A4, S100A8, and S100A9 expression was up-regulated during the progression of glioma grade. In addition, genetic variation of the S100A family was high in LGG, and the S100A family genes mostly function through IL-17 signaling pathway, S100 binding protein, and inflammatory responses. The TIMER database also revealed a relationship between gene expression and immune cell infiltration. High expression of S100A2, S100A3, S100A4, S100A6, S100A8, S100A9, S100A10, S100A11, S100A13, and S100A16 was significantly associated with poor prognosis in LGG patients. S100A family genes S100A2, S100A3, S100A6, S100A10, and S100A11 may be prognosis-related genes in LGG, and were significantly associated with IDH mutation and 1p19q codeletion. The immunohistochemical staining results also confirmed that S100A2, S100A3, S100A6, S100A10, and S100A11 expression was upregulated in LGG. Conclusion: The S100A family plays a vital role in LGG pathogenesis, presumably facilitating LGG progression via modulating inflammatory state and immune cell infiltration.

The type VI secretion system (T6SS), a multifunctional protein secretion device, plays very important roles in bacterial killing and/or virulence to eukaryotic cells. Although T6SS genes have been found in many Xanthomonas species, the biological function of T6SSs has not been elucidated in most xanthomonads. In this study, we identified two phylogenetically distinct T6SS clusters, T6SS1 and T6SS2, in a newly sequenced Chinese strain GX01 of Xanthomonas oryzea pv. oryzicola (Xoc) which causes bacterial leaf streak (BLS) of rice (Oryza sativa L.). Mutational assays demonstrated that T6SS1 and T6SS2 are not required for the virulence of Xoc GX01 on rice. Nevertheless, we found that T6SS2, but not T6SS1, played an important role in bacterial killing. Transcription and secretion analysis revealed that hcp2 gene is actively expressed and that Hcp2 protein is secreted via T6SS. Moreover, several candidate T6SS effectors were predicted by bioinformatics analysis that might play a role in the antibacterial activity of Xoc. This is the first report to investigate the type VI secretion system in Xanthomonas oryzae. We speculate that Xoc T6SS2 might play an important role in inter-bacterial competition, allowing this plant pathogen to gain niche advantage by killing other bacteria.

Background A growing body of evidence suggests that stem cell-derived exosomal microRNAs (miRNAs) could be a promising cardioprotective therapy in the context of hypoxic conditions. The present study aims to explore how miRNA-144 (miR-144), a miRNA contained in bone marrow mesenchymal stem cell (MSC)-derived exosomes, exerts a cardioprotective effect on cardiomyocyte apoptosis in the context of hypoxic conditions and identify the underlying mechanisms. Methods MSCs were cultured using the whole bone marrow adherent method. MSC-derived exosomes were isolated using the total exosome isolation reagent and confirmed by nanoparticle trafficking analysis as well as western blotting using TSG101 and CD63 as markers. The hypoxic growth conditions for the H9C2 cells were established using the AnaeroPack method. Treatment conditions tested included H9C2 cells pre-incubated with exosomes, transfected with miR-144 mimics or inhibitor, or treated with the PTEN inhibitor SF1670, all under hypoxic growth conditions. Cell apoptosis was determined by flow cytometry using 7-ADD and Annexin V together. The expression levels of the miRNAs were detected by real-time PCR, and the expression levels of AKT/p-AKT, Bcl-2, caspase-3, HIF-1α, PTEN, and Rac-1 were measured by both real-time PCR and western blotting. Results Exosomes were readily internalized by H9C2 cells after co-incubation for 12 h. Exosome-mediated protection of H9C2 cells from apoptosis was accompanied by increasing levels of p-AKT. MiR-144 was found to be highly enriched in MSC-derived exosomes. Transfection of cells with a miR-144 inhibitor weakened exosome-mediated protection from apoptosis. Furthermore, treatment of cells grown in hypoxic conditions with miR-144 mimics resulted in decreased PTEN expression, increased p-AKT expression, and prevented H9C2 cell apoptosis, whereas treatment with a miR-144 inhibitor resulted in increased PTEN expression, decreased p-AKT expression, and enhanced H9C2 cell apoptosis in hypoxic conditions. We also validated that PTEN was a target of miR-144 by using luciferase reporter assay. Additionally, cells treated with SF1670, a PTEN-specific inhibitor, resulted in increased p-AKT expression and decreased H9C2 cell apoptosis. Conclusions These findings demonstrate that MSC-derived exosomes inhibit cell apoptotic injury in hypoxic conditions by delivering miR-144 to cells, where it targets the PTEN/AKT pathway. MSC-derived exosomes could be a promising therapeutic vehicle to facilitate delivery of miRNA therapies to ameliorate ischemic conditions.

The dynamic pattern of volatiles during Pinot Noir winemaking in monsoon climate with yeast extracellular extract (EE) treatment was analyzed. EE from selected Pichia fermentans and Rhodotorula mucilaginosa strains, and almond β-glucosidase were added after 12-h alcohol fermentation, and the volatiles were determined every 24 h by GC–MS. After 6-month storage, wine aroma was evaluated instrumentally as well as by well-trained panelists. Results showed that enzyme treatments improved the contents of both varietal and fermentative volatiles. The levels of C6 compounds, terpenes, and higher alcohols increased constantly during alcohol fermentation, whereas acetates, short and medium chain fatty acid ethyl esters, phenylethyls, and fatty acids increased first, followed by gradual decrease. EE treatment retarded the decrease of fruity ester content in wine. Mathematical regression between wine aroma and volatiles showed that the relatively higher contents of acetates, ethyl esters, and C13-norisoprenoids in 6-month EE-treated wine were responsible for the improvement in floral aroma intensity.

Background: Glioblastoma (GBM) has a high degree of malignancy, aggressiveness and recurrence rate. However, there are limited options available for the treatment of GBM, and they often result in poor prognosis and unsatisfactory outcomes. Materials and methods: In order to identify potential core genes in GBM that may provide new therapeutic insights, we analyzed three gene chips (GSE2223, GSE4290 and GSE50161) screened from the GEO database. Differentially expressed genes (DEG) from the tissues of GBM and normal brain were screened using GEO2R. To determine the functional annotation and pathway of DEG, Gene Ontology (GO) and KEGG pathway enrichment analysis were conducted using DAVID database. Protein interactions of DEG were visualized using PPI network on Cytoscape software. Next, 10 Hub nodes were screened from the differentially expressed network using MCC algorithm on CytoHubba software and subsequently identified as Hub genes. Finally, the relationship between Hub genes and the prognosis of GBM patients was described using GEPIA2 survival analysis web tool. Results: A total of 37 up-regulated and 187 down-regulated genes were identified through microarray analysis. Amongst the 10 Hub genes selected, SV2B appeared to be the only gene associated with poor prognosis in glioblastoma based on the survival analysis. Conclusion: Our study suggests that high expression of SV2B is associated with poor prognosis in GBM patients. Whether SV2B can be used as a new therapeutic target for GBM requires further validation.

Aim Inflammation within the perivascular adipose tissue (PVAT) in obesity plays an important role in cardiovascular disorders. C-reactive protein (CRP) level in obesity patients is significantly increased and associated with the occurrence and progression of cardiovascular disease. We tested the hypothesis CRP derived from PVAT in obesity contributes to vascular remodeling after injury. Methods A high-fat diet (HFD) significantly increased CRP expression in PVAT. We transplanted thoracic aortic PVAT from wild-type (WT) or transgenic CRP-expressing (CRPTG) mice to the injured femoral artery in WT mice. Results At 4 weeks after femoral artery injury, the neointimal/media ratio was increased significantly in WT mice that received PVAT from CRPTG mice compared with that in WT mice that received WT PVAT. Transplanted CRPTG PVAT also significantly accelerated adventitial macrophage infiltration and vasa vasorum proliferation. It was revealed greater macrophage infiltration in CRPTG adipose tissue than in WT adipose tissue and CRP significantly increased the adhesion rate of monocytes through receptor Fcγ RI. Proteome profiling showed CRP over-expression promoted the expression of chemokine (C-X-C motif) ligand 7 (CXCL7) in adipose tissue, transwell assay showed CRP increased monocyte migration indirectly via the induction of CXCL7 expression in adipocytes. Conclusion CRP derived from PVAT was significantly increased in HFD mice and promoted neointimal hyperplasia after vascular injury.

Cabernet Gernischt (CG) is a famous Chinese wine grape cultivar, the red wine of which is known for its green trait, especially when produced from grapes cultivated in regions with monsoon climate. To modify CG wine aroma, three enzyme preparations (H. uvarum extracellular enzyme, AR2000, and pectinase) were introduced in different winemaking stages with Saccharomyces cerevisiae. Free and bound aroma compounds in young wines were detected using headspace solid-phase micro-extraction and gas chromatography-mass spectrometry, and aroma characteristics were quantified by trained panelists. Results showed that simultaneous inoculation of enzymes and yeasts improved wine aroma. Partial least-squares regression revealed that the green trait was due mainly to varietal compounds, especially C6 compounds, and could be partly weakened by fermentative compounds. Moreover, H. uvarum enzyme treatments enriched the acid fruit note of CG wine by enhancing the synergistic effect of varietal volatiles and certain fermentative compounds, such as esters and phenylethyls.

The major aroma components in grapes and wine include free volatile compounds and glycosidic nonvolatile compounds. The latter group of compounds is more than 10 times abundant of the former, and constitutes a big aroma reserve in grapes and wine. This review summarizes the research results obtained recently for the identification of aroma glycosides in grapes and wine, including grape glycoside structures, differences in aroma glycosides among grape varieties, hydrolysis mechanisms, and the factors that influence them. It also presents the analytical techniques used to identify the glycosidic aroma precursors. The operational strategies, challenges, and improvements of each step encountered in the analysis of glycosidic aroma precursors are described. This review intends to provide a convenient reference for researchers interested in the methods used for the determination of the aroma glucosides composition and the recognition of their chemical structures.

Doxorubicin (Dox) is commonly used for the treatment of malignant tumors, including colon cancer. However, the development of P-glycoprotein (P-gp)-mediated multidrug resistance (MDR) in tumor chemotherapy has seriously reduced the therapeutic efficacy of Dox. Natural product curcumin (Cur) was demonstrated to have a variety of pharmacological effects, such as anti-tumor, anti-oxidation and anti-aging activities. Here, we examined the MDR reversal capability of Cur in drug sensitive-(SW620) and resistant-(SW620/Ad300) colon cancer cells, and elucidated the underlying molecular mechanisms at the metabolic level. It was found that Cur reversed P-gp-mediated resistance in SW620/Ad300 cells by enhancing the Dox-induced cytotoxicity and apoptosis. Further mechanistic studies indicated that Cur inhibited the ATP-dependent transport activity of P-gp, thereby increasing the intra-celluar accumulation of Dox in drug-resistant cells. Metabolomics analysis based on UPLC-MS/MS showed that the MDR phenomenon in SW620/Ad300 cells was closely correlated with the upregulation of spermine and spermidine synthesis and D-glutamine metabolism. Cur significantly inhibited the biosynthesis of spermine and spermidine by decreasing the expression of ornithine decarboxylase (ODC) and suppressed D-glutamine metabolism, which in turn decreased the anti-oxidative stress ability and P-gp transport activity of SW620/Ad300 cells, eventually reversed MDR. These findings indicated the MDR reversal activity and the related mechanism of action of Cur, suggesting that Cur could be a promising MDR reversal agent for cancer treatment.

The aim of this work was to rapidly screen indigenous yeasts with high levels of β-glucosidase activity and assess the potential of glycosidase extracts for aroma enhancement in winemaking. A semiquantitative colorimetric assay was applied using 96-well plates to screen yeasts from 3 different regions of China. Isolates with high β-glucosidase activity were confirmed by the commonly used pNP assay. Among 493 non-Saccharomyces isolates belonging to 8 generas, 3 isolates were selected for their high levels of β-glucosidase activity and were identified as Hanseniaspora uvarum, Pichia membranifaciens, and Rhodotorula mucilaginosa by sequence analysis of the 26S rDNA D1/D2 domain. β-Glucosidase in the glycosidase extract from H. uvarum strain showed the highest activity in winemaking conditions among the selected isolates. For aroma enhancement in winemaking, the glycosidase extract from H. uvarum strain exhibited catalytic specificity for aromatic glycosides of C13 -norisoprenoids and some terpenes, enhancing fresh floral, sweet, berry, and nutty aroma characteristics in wine.

UNLABELLED The aim of the work was to evaluate the application potential of a glycosidase extract of one indigenous non-Saccharomyces strain in wine aroma enhancement. The isolate was selected from a local winemaking region in China for its high β-glucosidase level and was identified as Rhodotorula mucilaginosa. The tolerance of the glycosidase extract to the typical winemaking conditions was assessed using the activity of its β-glucosidase. After that, the hydrolysis capacity of R. mucilaginosa glycosidase for liberation of grape aroma glycosides was characterized in comparison to commercial enzyme preparations. Results of this work revealed that glycosidase extract from R. mucilaginosa proved to be active in the presence of 0-20% (w/v) glucose, 0-20% (v/v) ethanol and at pH 3·0-5·0. In the hydrolysis of aroma precursors, enzymes obtained from different origins possessed various levels of specificity and activity, showing high origin dependence (α = 0·05). Compared to commercial enzymes, the indigenous R. mucilaginosa glycosidase extract presented better catalytic preference for the 'fruity and floral' glycosides of benzenic compounds and C13 -norisoprenoids, but less sensitivity to the glycosides of C6 compounds and volatile phenols. SIGNIFICANCE AND IMPACT OF THE STUDY This work presents a novel extracellular glycosidase preparation from an indigenous Rhodotorula mucilaginosa strain selected from a local winemaking region in China. This enzyme extract exhibits strong tolerance towards winemaking conditions. It shows hydrolysis specificity for glycosides of benzenic compounds and C13 -norisoprenoids, proving a potential candidate for improving floral and fruity aroma characteristics of wine.

To investigate the effect of alternol on pancreatic cancer cells.Pancreatic cancer cells PANC-1 and BxPC3 were treated with various concentrations of alternol for 24, 48 and 72 h. Cell proliferation was measured by cell counting. Cell cycle distribution and mitochondrial membrane potential were determined by flow cytometry. Apoptosis was determined by a TdT-mediated dUTP nick end labeling assay and Hoechst staining. Expression of caspase 3, Bcl-2, p53 and p21 was measured by western blotting.Alternol showed dose- and time-dependent inhibition of the proliferation of PANC-1 and BxPC3 cells in vitro. Alternol induced apoptosis and cell cycle arrest at S phase and decreased mitochondrial membrane potential. Alternol activated caspase 3, upregulated p53 and p21 expression, and downregulated Bcl-2 expression in a dose-dependent manner.Our results suggested that alternol is a candidate for treatment of pancreatic cancer.

Epidemiological data indicate an increasing incidence of asthma in the obese individuals recent decades, while very little is known about the possible association between them. Here, we compared the roles of adipocyte-derived factors, including leptin, adiponectin and resistin on proliferation, wound repair and apoptosis in human bronchial epithelial cells (HBECs) which play an important role in the pathogenesis of asthma. The results showed that exogenous globular adiponectin (gAd) promoted proliferation, cell-cycle and wound repair of HBECs. This effect may be relevant to Ca(2+)/calmodulin signal pathway. Besides, gAd inhibited apoptosis induced by ozone and release of lactate dehydrogenase (LDH) of HBECs via regulated adipoR1 and reactive oxygen species. No effects of leptin or resistin on proliferation, wound repair and apoptosis of HBECs were detectable. These data indicate that airway epithelium is the direct target of gAd which plays an important role in protecting HBECs from mechanical or oxidant injuries and may have therapeutic implications in the treatment of asthma.

If " Four faces surrounded by lotuses and three by willows, mountain scenery as the whole city and half covered by lake." is the wonderful portraiture of the Spring city Jinan, then "Every family has springs and willow " is the real description of Hibiscus Street and its surrounding houses. Hibiscus Street is located in the heart of the old city, the residential architecture has been preserved relatively intact. Though Hibiscus Street located in the north, it has characteristics of the southern residential, that is why it has important research values.

Cystic fibrosis transmembrane conductance regulator (CFTR) is an apical membrane chloride channel critical to the regulation of fluid, chloride, and bicarbonate transport in epithelia and other cell types. The most common cause of cystic fibrosis (CF) is the abnormal trafficking of CFTR mutants. Therefore, understanding the cellular machineries that transit CFTR from the endoplasmic reticulum to the cell surface is important. Vasoactive intestinal polypeptide (VIP) plays an important role in CFTR-dependent chloride transport. The present study was designed to observe the affection of VIP on the trafficking of CFTR, and channel gating in human bronchial epithelium cells (HBEC). Confocal microscopy revealed CFTR immunofluorescence extending from the apical membrane deeply into the cell cytoplasm. After VIP treatment, apical extension of CFTR immunofluorescence into the cell was reduced and the peak intensity of CFTR fluorescence shifted towards the apical membrane. Western blot showed VIP increased cell surface and total CFTR. Compared with the augmented level of total CFTR, the surface CFTR increased more markedly. Immunoprecipitation founded that the mature form of CFTR had a marked increase in HBEC treated with VIP. VIP led to a threefold increase in Cl− efflux in HBEC. Glibenclamide-sensitive and DIDS-insensitive CFTR Cl− currents were consistently observed after stimulation with VIP (10−8 mol/L). The augmentation of CFTR Cl− currents enhanced by VIP (10−8 mol/L) was reversed, at least in part, by the protein kinase A (PKA) inhibitor, H-89 and the protein kinase C (PKC) inhibitor, H-7, suggesting PKA and PKC participate in the VIP-promoted CFTR Cl− currents. J. Cell. Biochem. 112: 902–908, 2011. © 2010 Wiley-Liss, Inc.

The shedding of airway epithelial cells and loss of epithelial functional homeostasis are major pathological characteristics of asthma; however, the mechanism underlying these pathologies remains obscure. Our previous work showed that there were three variation sites in 5' flanking region of integrin beta4 in asthma patients, which was correlated with decreased expression of integrin beta4 in peripheral leukocytes. Integrin beta4 is an important structural adhesion molecule on airway epithelia to keep the structural adhesion of epithelial cells. In this work, we further demonstrated that integrin beta4 expression was downregulated in airway epithelia of asthma patients. To probe the relationship between imbalanced expression of integrin beta4 and dysfunction of the airway epithelial cells in asthma, integrin beta4 was silenced in human bronchial epithelium cells (16HBE14O) by integrin beta4 small-interfering RNA lentivirus vector. Upon silencing of integrin beta4, 16HBE14O cells showed reduced proliferation and wound repair. Most cells were shown to be arrested in G1 phase after integrin beta4 silencing, and increased apoptosis was induced in the integrin beta4-silenced cells. In summary, our results provided compelling evidence that integrin beta4 was involved in the structural integrity and functional homeostasis of airway epithelial cells. It is likely that downregulation of integrin beta4 on asthma airway epithelia contributes to the structural disruption and dysfunction of airway epithelial cells.

Integrin beta 4 (ITGB4) is a structural adhesion molecule which engages in maintaining the integrity of airway epithelial cells. Its specific cytomembrane structural feature strongly indicates that ITGB4 may engage in many signaling pathways and physiologic processes. However, in addition to adhesion, the specific biologic significance of ITGB4 in airway epithelial cells is almost unknown. In this article, we investigated the expression and functional properties of ITGB4 in airway epithelial cells in vivo and in vitro. Human bronchial epithelial cell line (16HBE14O-cells) and primary rat tracheal epithelial cells (RTE cells) were used to determine ITGB4 expression under ozone tress or mechanical damage, respectively. An ovalbumin (OVA)-challenged asthma model was used to investigate ITGB4 expression after antigen exposure in vivo. In addition, an ITGB4 overexpression vector and ITGB4 silence virus vector were constructed and transfected into RTE cells. Then, wound repair ability and anti-oxidation capacity was evaluated. Our results demonstrated that, on the edge of mechanically wounded cell areas, ITGB4 expression was increased after mechanical injury. After ozone stress, upregulation expression of ITGB4 was also detected. In the OVA-challenged asthma model, ITGB4 expression was decreased on airway epithelial cells accompanying with structural disruption and damage of anti-oxidation capacity. Besides, our study revealed that upregulation of ITGB4 promotes wound repair ability and anti-oxidative ability, while such abilities were blocked when ITGB4 was silenced. Taken together, these results showed that ITGB4 was a new interesting molecule involved in the regulation of wound repair and anti-oxidation processes for airway epithelial cells.

The cytotoxic effects and mechanism of action of cisplatin and the mTOR inhibitor rapamycin on Hep-2 laryngeal cancer cells were investigated. Hep-2 cells were cultured in the presence of different concentrations of rapamycin, cisplatin, or the two combined. Inhibition of cell growth, apoptosis, and AKT, mTOR, S6K, and ERCC1 protein levels were assessed. All combinations of rapamycin and cisplatin resulted in synergistic inhibition of cell growth (as indicated by q values determined using Jin's formula > 1.15). Rapamycin inhibited Hep-2 cell growth, induced G1 arrest, and when combined with cisplatin, enhanced apoptosis. p-mTOR and S6K expressions were significantly downregulated by rapamycin. ERCC1 expression was significantly upregulated with cisplatin treatment. Combined cisplatin and rapamycin treatment resulted in significant downregulated p-mTOR and S6K expression, but no change in ERCC1 expression. Rapamycin and cisplatin act in a synergistic manner, increasing the cytotoxic effect on Hep-2 cells. Rapamycin may facilitate increased Hep-2 cell apoptosis with cisplatin via inhibiting downstream expression of proteins in the AKT-mTOR signaling pathway.

Background: Parental studies are often necessary subsequent to the identification of a chromosome abnormality. The recommended studies are based on assumptions about how chromosome rearrangements occur. One such assumption is that deletion size is stable through generations. Results: We have identified a family where a small subtelomeric deletion in a phenotypically and cytogenetically normal mother expanded nearly 10-fold into a clinically consequential and cytogenetically visible deletion in her affected daughter. Conclusion: Traditional parental follow-up studies would have not identified this expansion, but would have rather classified the deletion in the daughter as either de novo or benign. Only by sizing the deletion by array comparative genomic hybridisation in both the mother and the daughter was the expansion recognised. Previous assumptions about chromosome behaviour suggest that this phenomenon may have been easily missed in other cases of chromosomal deletions. Therefore, this case illustrates the need for more comprehensive analyses of parental chromosome structure when characterising an abnormality in a child.

The complete mitochondrial genome sequence of a funnel-web weaving spider, Agelena silvatica (Agelenidae), was obtained using long PCR and conventional PCR methods. The circular genome is 14,776 bp in length. It contains 13 protein-coding genes, 22 transfer RNAs, 2 ribosomal RNAs, and a control region. The A + T content (74.46%) of the all base composition is higher than G + C content (25.54%). Bayesian and maximum likelihood phylogenetic analyses demonstrate that Agelena silvatica is the sister clade of the water spider, Argyroneta aquatica.

DJ-1, a novel mitogen-dependent oncogene, has an important role in the progression of human malignancies, whereas tumor suppressor phosphatase and tensin homolog (PTEN) is known to control a variety of processes associated with cell survival, proliferation and invasion. DJ-1 overexpression was reported to be negatively correlated with PTEN expression in tumor tissues of patients with laryngeal squamous cell carcinoma (LSCC). In the present study, the effect of DJ-1 on PTEN in laryngeal cancer cells was investigated by transfecting DJ-1-specific small interfering (si)RNA into Hep-2 and SNU-899 cells. Cell survival and cell proliferative and invasive capacity were then evaluated. The results showed that siRNA targeting of DJ-1 effectively upregulated PTEN expression, resulting in enhanced cell death as well as decreased proliferation and invasion of Hep-2 and SNU-899 cells. The results of the present study indicated, for the first time, to the best of our knowledge, that DJ-1-induced PTEN downregulation is associated with proliferative and invasive activity of laryngeal cancer cells. The DJ-1 gene may have an important role in the tumorigenesis of LSCC.

CD4+ regulatory T cells (Tregs) mediate immune tolerance in laryngeal squamous cell carcinoma (LSCC). However, Tregs are functionally heterogeneous. Recently, we reported that three distinct Treg subsets (resting Tregs, activated Tregs and cytokine-secreting CD45RA-Foxp3lowCD4+ T cells) vary in the peripheral circulation of patients with head and neck squamous cell carcinoma (HNSCC); however, the potential implication of these Treg subsets in LSCC immunity is unclear. Here, we report that activated Tregs and cytokine‑secreting CD45RA-Foxp3lowCD4+ T cells were increased in LSCC patients compared with healthy donors (HD) (p

BACKGROUND. Cytogenetic, molecular cytogenetic, and molecular studies of prostate cancer have revealed an enormous amount of data regarding chromosomal loci that are aberrant in prostate tumors. METHODS. These data have been compared and condensed in this review to determine which chromosomes and chromosome sites have been most frequently reported. RESULTS. Loss of the Y chromosome, gain of 7, 8, and X, and interstitial deletions on 6q, 7q, 8p, 10q, 13q, 16q, 17q, and 18q are the most prevalent. CONCLUSIONS. A potential model for genetic control of tumor progression is presented, as are data regarding the evaluation of a new series of tumors.

One of the most common chromosomal abnormalities in prostate cancer involves loss of 10q22-qter. Rarely, a smaller deletion, involving 10q24-q25, has been observed, suggesting the presence of a tumor suppressor gene at this site. We previously demonstrated that the MXI1 gene maps to 10q24-q25 and is mutated in some tumors with cytogenetically detectable deletions of this locus. MXI1 encodes a basic-helix-loop-helix protein that suppresses the transcriptional activity of the MYC oncoprotein by competing for the common dimerization partner, MAX, and binding to identical DNA sites. Because more than 90% of prostate tumors contain no cytogenetic abnormality of 10q, the relevance of MXI1 loss and/or mutation to the vast majority of cases remains unclear. We prospectively evaluated prostate tumors for loss of MXI1 by fluorescence in situ hybridization (FISH) and cytogenetic techniques. Twenty-one of 40 tumors (53%) demonstrated loss of a single MXI1 allele as determined by FISH. Ten cases with cytogenetically normal 10qs, but with FISH-documented deletion of MXI1, were examined at the molecular level, and eight mutations were identified, albeit at low frequency. Five of the mutant proteins were unable to bind DNA in association with MAX. We conclude that MXI1 gene loss in prostate cancer is common and most frequently involves a cytogenetically undetectable deletion.

Hepatocellular carcinoma (HCC) is the third most common cause of cancer-related deaths worldwide. Such deaths are due, in large part, to its propensity to metastasize. We have examined the effect of alternol on human HCC cells and the underlying molecular mechanism. Therapeutic effects of alternol on cancer cell migration and invasion were analyzed with Boyden chamber and wound healing assays. Effects of alternol on the levels of various proteins involved in cancer cell migration and invasion were determined with gelatin zymography, immunofluorescence, and Western blotting. As shown, treatment with alternol has resulted in a concentration-dependent inhibition of cell migration and invasion of HepG2 cells. The inhibition of HCC invasion by alternol was associated with the suppression of MMP-9 expression and reversal of epithelial-to-mesenchymal transition (EMT). The above results indicated that alternol has the ability to inhibit the migration and invasion of human HCC cells by reversing the process of EMT, suggesting that alternol may be developed as an alternative drug for the treatment of HCC.

Purpose: The breast-ovarian cancer susceptibility gene, BRCA1, has been implicated by both epidemiologic and genetic studies to be involved in prostate cancer. We wished to test the frequency of BRCA1 deletion and that of other markers in the region of proximal 17q in prostate tumor cells.Materials and Methods: We used a dual-color fluorescence in situ hybridization (FISH) assay using P1 phage probes for the BRCA1 gene and 3 flanking sites at 17q12-21, as well as a chromosome 17 centromere-specific alpha-satellite probe, to detect deletions in single-cell suspensions and touch preparations from 23 primary clinical stage B prostate tumors and adjacent nontumor prostate tissues. Lymphoblastoid cells and prostate cells from a normal donor were used to determine control loss values.Results: Significant loss (p less than 0.05) of at least 1 of the P1s was detected in 16 of 23 (70 percent) cases, and in 4 of those cases all markers were lost, consistent with whole chromosome loss. Of the 12 cases with s...

Neurofibromatosis type 1 (NFl) is a common genetic disorder characterized primarily by the development of multiple neurofibromas and pigmentary changes. The recent identification of contiguous gene deletions in NF1 a previously unrecognized molecular basis for this disorder, raises important questions regarding deletion frequency in the patient population and the role that contiguous genes may play in the physical manifestations of NF1 patients. To facilitate the identification of patients with large NFl deletions, we have isolated clones carrying large genomic segments from the NF1 locus and tested their efficacy as probes for fluorescence in situ hybridization (FISH). Clone P1-9 spans approximately 65 kb of the NFl gene, including exons 2–11, and clone P1-12 carries ∼55 kb of NFl intron 27B. FISH studies performed with P1-9, P1-12, and a set of overlapping 1F10 cosmid clones mapping telomeric to the NF1 locus identified large deletions in two new neurofibromatosis type 1 patients who, like previously characterized deletion patients, had mildly dysmorphic facial features and large numbers of cutaneous neurofibromas.

It is known that ozone stress can induce airway hyperresponsiveness (AHR). The underlying cellular and molecular mechanisms are not fully understood. We constructed a successive ozone-stressed rat model and showed that AHR caused by ozone stress presented as increased lung resistance (R(L)) to inhaled histamine but not baseline R(L). Meanwhile, structural disruption and decreased expression of integrin beta4 on airway epithelia were observed. Further regression analysis revealed a significant negative correlation between increases in R(L) to histamine (at 0.32 mg/ml) and mRNA expression of integrin beta4. Moreover, when integrin beta4 on human bronchial epithelial cells was knocked down, we found that reactive oxygen species was increased and apoptosis rates were higher. Overall, this study suggests that downregulation of integrin beta4 is important for the development ozone stress-induced AHR, presumably because it causes increased oxidative damage and epithelial apoptosis.

Purpose: The University of Utah Comparative Genomic Hybridization Microarray Laboratory was one of the first US laboratories to offer comparative genomic hybridisation (CGH) microarray testing using a commercial platform in a clinical setting. Results for 1076 patients (1598 chips) are presented. Methods: The Spectral Genomics/PerkinElmer Constitutional Chip TM (targeted array), SpectralChip 2600 TM (whole genome array) and a “Combo” chip (both arrays run simultaneously) were the tests offered. Abnormal results were confirmed by an alternative method, most often fluorescence in situ hybridisation. Results: In 669 cases with known normal cytogenetics, an abnormal detection rate of 10.8% was observed, (5.3%, 12.2% and 14.1% for the Constitutional Chip TM , SpectralChip 2600 TM and Combo assay, respectively). Known copy number variants and single clone abnormalities are not included in these rates. Single clone abnormalities are reported separately. For 1076 total cases, we report an average abnormal rate of 16.9% (8.7%, 23.7% and 18.6% for the three assays). This rate includes characterisation of some abnormalities previously identified by cytogenetics. Conclusions: CGH microarray provides a likely aetiology for the clinical phenotype in many cytogenetically normal cases, and a whole genome array generally identifies copy number changes more effectively than a targeted chip alone.

Prostate cancer is the most common cancer in males in the United States, yet the etiology of this disease is still poorly understood. In previous work from our laboratory, one or more deleted regions were found in prostate tumors distal to the breast and ovarian cancer susceptibility gene (BRCA1) on chromosome 17. This suggested that genes at 17q21 may play a pivotal role in prostate cancer progression, and there may be new tumor suppressor genes at this locus. We now present a physical map built with P1, P1 artificial chromosome, and bacterial artificial chromosome clones encompassing a DNA sequence anchored by multiple STS markers. The analysis of prostate tumors indicated an 85-kb novel commonly deleted interval flanked by D17S1184-D17S183-D17S1203-D17S1860, which is at least 470 kb distal to the BRCA1 gene. Fifty-four of 126 prostrate cancer cases (43%) showed a deletion by a direct FISH technique using P1 probes in this region. Searching with clone end sequences in the sequence database BLAST, the deleted clone covered genomic DNA sequence that contained upstream binding factor (UBF), EPB3 genes, SHCL1, ASB-4-like sequence, and acidic protein-like sequence. PCR for the ESTs confirmed that these genes or ESTs are within the deletion region. Our results will be helpful for finding candidate tumor suppressor genes in prostate cancer.

There is a clear genetic component to prostate cancer susceptibility. Regions reported to be linked to prostate cancer include 1q24-25 (HPC-1), 1q42.2-43, and Xq27-28. There is limited genetic information on familial prostate tumors. We used the Utah Population Database to identify familial prostate cancer cases and selected 35 cases from high-risk families. Tissue blocks containing discernable tumor were available from 19 cases; 13 of these yielded adequate specimens for analysis. Six cases came from families with linkage to HPC-1, 3 were known to have linkage to Xq27-28, and 4 had no linkage to a known locus; 7 cases were analyzed from patients who showed no known linkage (sporadic tumors) as controls. These paraffin-embedded tumors were laser microdissected, degenerate oligonucleotide (DOP)-amplified, and labeled for fluorescence detection by comparative genomic hybridization (CGH). Loss of 7q, 10q, and 16q and gain of 8q were common abnormalities present in both familial and sporadic tumors. Distinctive abnormalities included loss of 3p12-3p22 in 3 of 6 HPC-7-linked cases and in 2 of 3 X-linked cases and gain of 6q11-6q21 in 2 each of HPC-1 and X-linked tumors. In conclusion, laser microdissection, DOP-PCR, and CGH is a feasible method for analysis of paraffin-embedded prostate tumors. This study provides preliminary data suggesting that familial prostate cancer harbors some unique genetic changes when compared with sporadic prostate tumors.

Aneusomy of chromosome 7 and loss at 7q (especially 7q31.1) have been reported in prostate cancer. To further investigate abnormalities of 7q and the relationship with whole chromosome 7 changes, we have conducted a dual-color fluorescence in situ hybridization (FISH) analysis on isolated nuclei from 28 primary prostate cancers. A pericentromeric probe for chromosome 7, five newly isolated sequence-specific bacterial artificial chromosome (BAC) probes from 7q31.1, and one BAC for the epidermal growth factor receptor (EGFR) gene at 7p12 were used in dual color hybridizations. Pericentromeric probes for chromosomes X and 4 were also used as controls. Sixteen (57.1%) of the 28 tumors showed clonal aberrations. Nine of them were trisomy 7 and four were hypertetrasomy for chromosome 7. Deletions at 7q31.1 were found in two of the high grade tumors. With the exception of these two cases, all other cases showed concordant results using all probes. These findings confirm previous studies that aneusomy of 7 is associated with prostate cancer progression, and there may be a tumor suppressor gene (TSG) at 7q31.1 which is associated with tumor progression. In addition, our study indicates: (1) the deletion pattern of individual nuclei infers that deletions at 7q31.1 precede reduplications of chromosome 7; and (2) the amplification of EGFR was not detected at the DNA level, suggesting that activation of this oncogene may play a minor role in prostate cancer.

Using fluorescence in situ hybridization (FISH), we have identified seven NF1-related loci, two separate loci on chromosome 2, at bands 2q21 and 2q33-q34, and one locus each on five other chromosomes at bands 14q11.2, 15q11.2, 18p11.2, 21q11.2-q21, and 22q11.2. Application of PCR using NF1 primer pairs and genomic DNA from somatic cell hybrids confirmed the above loci, identified additional loci on chromosomes 12 and 15, and showed that the various loci do not share homology beyond NF1 exon 27b. Sequenced PCR products representing segments corresponding to NF1 exons from these loci demonstrated greater than 95% sequence identity with the NF1 locus. We used sequence differences between bona fide NF1 and NF1-homologous loci to strategically design primer sets to specifically amplify 30 of 36 exons within the 5' end of the NF1 gene. These developments have facilitated mutation analysis at the NF1 locus using genomic DNA as template.

Glioblastoma multiforme (GBM) is the most malignant glial brain tumor in humans. The fact that deleted copies of chromosome 10 are observed frequently in primary GBM tumors supports the hypothesis that one or more tumor suppressor genes located on chromosome 10 occupy crucial growth control checkpoints for glial cells. Deletion mapping in primary GBM tumors using the loss of heterozygosity (LOH) test has implicated the 10q24-10qter region as one possible site for a gene. We report here on the molecular cytogenetic analysis of chromosome 10 abnormalities in a human GBM cell line, JBSA. LOH testing showed that JBSA cells were hemizygous for chromosome 10. Molecular cytogenetic analysis showed that the undeleted homologue was involved in a reciprocal translocation, t(7;10)(p21;q22). The translocation breakpoint on chromosome 10 lay within band q22 between D10S19 and D10S4. The fact that JBSA cells lack one homologue of chromosome 10 and carry a translocation breakpoint on the remaining one, proximal to the smallest region of overlap reported in primary tumor deletions, suggests that 10q22 may be another possible site for a tumor suppressor gene involved in GBM.

A human delayed rectifier K{sup +} channel gene has been localized to the long arm of human chromosome 20q13.2 by fluorescence in situ hybridization of genomic P1 clones from this locus. A polymorphic (GA) microsatellite repeat was identified in one of the P1 clones. The new SSR marker (D20S436) was genotyped in four CEPH pedigrees. Two-point linkage analysis indicated linkage of this marker to a PCR marker, D20S109, with a maximum lod score of 9.32 at {theta} = 0.001. The assignment of this K{sup +} channel gene to 20q13.2 eliminates it as a candidate for the gene associated with benign familial neonatal convulsions (BFNC), which has been localized to 20q13.3. Genetically, the K{sup +} channel gene maps more than 30 cM proximal to the BFNC locus. 16 refs., 1 fig., 1 tab.

Twenty prostate tumor specimens, obtained from radical prostatectomies, and two lymph node metastases were examined by classical and molecular cytogenetic methods. A sample from each tumor was analyzed histologically and used for touch preparations. Adjacent samples were used for preparation of single-cell suspensions before cell culture (DirFISH) and for establishing cell cultures, which were subsequently harvested for classical G-banding analysis. Fluorescence in situ hybridization (FISH) was performed on touch preparations, DirFISH, and cells obtained from tissue culture. Biotinylated pericentromeric probes for chromosomes 7 and 17, in addition to a digoxigenin-labeled X-chromosome probe, were used in a dual-color FISH assay. The results indicated that, in uncultured tumor cells, chromosome 17 was lost in 55% of specimens, chromosome 7 was gained in 16% of specimens, and 9% of specimens showed large tetraploid populations. After cell culture, 23% of specimens showed loss of chromosome 17, no specimens showed gain of chromosome 7, and no tetraploid populations were present. This study suggests that loss of chromosome 17 may play an important role in the development of prostate cancer, and that genetic changes observed after selection in vitro may not represent those in the original tumor.

Xanthine dehydrogenase (XDH, EC 1.1.1.204) oxidizes a variety of purines, pterins, and other heterogenic nitrogen compounds, serving as a rate-limiting enzyme in nucleic acid degradation. The genetic defect of XDH results in hereditary xanthinuria and other disorders in purine metabolism. Based on the cloning and sequencing results of human XDH cDNA in our laboratory, we studied the localization and sublocalization of the XDH gene. A Version 3.0 human-hamster somatic cell hybrid PCRable DNA panel and specific PCR primers derived from human XDH cDNA for amplification were used to assign the XDH gene to human chromosome 2. The fidelity of the PCR product was confirmed by nucleotide sequencing the PCR product. The assignment of the XDH gene to chromosome 2 at band p22 was established by fluorescence in situ hybridization on human metaphase chromosomes using a clone from a pWE 15 cosmid library containing the XDH gene. The results should be useful for further studies of the molecular basis for hereditary xanthinuria and other genetic disorders related to abnormal XDH activity.