• E. coli O157 persistence in the environment.

      Brown, Michael R. W.; Smith, Anthony W.; Barker, John; Humphrey, Thomas J.; Dixon, Bernard (Society for General Microbiology, 2002)
    • Effect of carrier particle shape on dry powder inhaler performance.

      Kaialy, Waseem; Alhalaweh, Amjad; Velaga, Sitaram P; Nokhodchi, Ali (Elsevier, 2011-12-12)
      The aim of this study was to characterise the aerosolisation properties of salbutamol sulphate (SS) from dry powder inhaler (DPI) formulations containing different carrier products. The difference in the elongation ratio (ER) of the different carriers was highlighted. Different set of carriers, namely commercial mannitol (CM), commercial lactose (CL), cooling crystallised mannitol (CCM), acetone crystallised mannitol (ACM) and ethanol crystallised mannitol (ECM) were used and inspected in terms of size, shape, density, crystal form, flowability, and in vitro aerosolisation performance using Multi Stage Liquid Impinger (MSLI) and Aerolizer inhaler device. Solid-state and morphological characterization showed that CM product was in pure β-form having particles with smaller ER (CM: ER=1.62 ± 0.04) whereas ACM and ECM mannitol particles were in pure α form with higher ER (ACM: ER=4.83 ± 0.18, ECM: ER=5.89 ± 0.19). CCM product crystallised as mixtures of β-form and δ-form and showed the largest variability in terms of particle shape, size, and DPI performance. Linear relationships were established showing that carrier products with higher ER have smaller bulk density (D(b)), smaller tap density (D(t)), higher porosity (P), and poorer flow properties. In vitro aerosolisation assessments showed that the higher the ER of the carrier particles the greater the amounts of SS delivered to lower airway regions indicating enhanced DPI performance. Yet, DPI performance enhancement by increasing carrier ER reached a "limit" as increasing carrier ER from 4.83±0.18 (ACM) to 5.89±0.19 (ECM) did not significantly alter fine particle fraction (FPF) of SS. Also, carrier particles with higher ER were disadvantageous in terms of higher amounts of SS remained in inhaler device (drug loss) and deposited on throat. Linear relationship was established (r(2)=0.87) showing that the higher the carrier ER the lower the drug emission (EM) upon inhalation. Moreover, poorer flowability for carrier products with higher ER is disadvantageous in terms of DPI formulation dose metering and processing on handling scale. In conclusion, despite that using carrier particles with higher ER can considerably increase the amounts of drug delivered to lower airway regions; this enhancement is restricted to certain point. Also, other limitations should be taken into account including higher drug loss and poorer flowability.
    • Effect of material deprivation on Epstein-Barr virus infection in Hodgkin's disease in the West Midlands.

      Flavell, Joanne R.; Constandinou, C.; Lowe, D.; Scott, K.; Newey, C.; Evans, D.; Dutton, A.; Simmons, S.; Smith, Richard; Crocker, John; et al. (nature.com, 1999)
      We have used Townsend scores from postcode data to compare levels of material deprivation and Epstein-Barr virus (EBV)-positivity for 223 patients diagnosed with Hodgkin's disease (HD) in the period 1981-1997. The presence of EBV in HD tumours was determined using in situ hybridization to target the abundantly expressed EBV early RNAs. EBV was detected in the malignant Hodgkin and Reed-Sternberg cells in 47/223 HD cases (21%). There was found to be a tendency for higher Townsend scores (indicative of higher levels of material deprivation) in EBV-positive HD patients, but this association was not statistically significant. When various subgroups of patients from the study were examined separately the indication of higher Townsend scores in EBV-positive patients was found to be more marked for patients with mixed cellularity disease (P = 0.09) and for females (P = 0.03). The results of this study suggest that differences in the level of material deprivation are important in determining the likelihood of EBV-positive HD in the UK, particularly for certain subgroups of patients. It is not known what specific socioeconomic factors are responsible for these differences, although alterations in the timing or rate of primary EBV infection, or decline in the level of EBV-specific immunity, may be important. (Cancer Research UK)
    • Elucidation of the mechanisms underlying the angiogenic effects of ginsenoside Rg(1) in vivo and in vitro.

      Yue, Patrick Y. K.; Wong, Daisy Y. L.; Ha, Wai-Yan; Fung, M.C.; Mak, Nai Ki; Yeung, H.W.; Leung, Hei Wun; Chan, Kelvin C.; Liu, Liang; Fan, T. P. David; et al. (Springer Verlag, 2005)
      The major active constituents of ginseng are ginsenosides, and Rg(1) is a predominant compound of the total extract. Recent studies have demonstrated that Rg(1) can promote angiogenesis in vivo and in vitro. In this study, we used a DNA microarray technology to elucidate the mechanisms of action of Rg(1). We report that Rg(1) induces the proliferation of HUVECs, monitored using [(3)H]-thymidine incorporation and Trypan blue exclusion assays. Furthermore, Rg(1) (150-600 nM) also showed an enhanced tube forming inducing effect on the HUVEC. Rg(1) was also demonstrated to promote angiogenesis in an in vivo Matrigel plug assay, and increase endothelial sprouting in the ex vivo rat aorta ring assay. Differential gene expression profile of HUVEC following treatment with Rg(1) revealed the expression of genes related to cell adhesion, migration and cytoskeleton, including RhoA, RhoB, IQGAP1, CALM2, Vav2 and LAMA4. Our results suggest that Rg(1) can promote angiogenesis in multiple models, and this effect is partly due to the modulation of genes that are involved in the cytoskeletal dynamics, cell-cell adhesion and migration.
    • Engineered mannitol ternary additives improve dispersion of lactose-salbutamol sulphate dry powder inhalations.

      Kaialy, Waseem; Nokhodchi, Ali (Springer, 2013-07)
      The aim of this study was to evaluate the influence of novel engineered fine mannitol particles (4.7%, w/w) on the performance of lactose-salbutamol sulphate dry powder inhaler (DPI) formulations to obtain promising aerosolisation properties. The results showed that the more elongated the fine mannitol particles, the weaker the drug-carrier adhesion, the better the drug content homogeneity, the higher the amount of drug expected to be delivered to the lower airways and the higher the total DPI formulation desirability. Linear relationships were established showing that mannitol particles with a more elongated shape generated powders with broader size distributions and that were less uniform in shape. The weaker the drug-carrier adhesion, the higher the fine particle fraction of the drug is upon aerosolisation. It is believed that more elongated fine mannitol particles reduce the number of drug-carrier and drug-drug physical contact points and increase the ability of the drug particles to travel into the lower airways. Additionally, a lower drug-carrier contact area, lower drug-carrier press-on forces and easier drug-carrier detachment are suggested in the case of formulations containing more elongated fine mannitol particles. Ternary 'drug-coarse carrier-elongated fine ternary component' DPI formulations were more favourable than both 'drug-coarse carrier' and 'drug-elongated coarse carrier' binary formulations. This study provides a comprehensive approach for formulators to overcome the undesirable properties of dry powder inhalers, as both improved aerosolisation performance and reasonable flow characteristics were obtained using only a small amount of elongated engineered fine mannitol particles.
    • Epidermal growth factor receptor kinase domain mutations in esophageal and pancreatic adenocarcinomas.

      Kwak, Eunice L.; Jankowski, Janusz; Thayer, Sarah P.; Lauwers, Gregory Y.; Brannigan, Brian W.; Harris, Patricia L.; Okimoto, Ross A.; Haserlat, Sara M.; Driscoll, David R.; Ferry, David R.; et al. (American Association for Cancer Research, 2006)
      PURPOSE: Specific activating mutations within the epidermal growth factor receptor (EGFR) identify a subset of non-small cell lung cancers with dramatic sensitivity to the specific tyrosine kinase inhibitors (TKI), gefitinib and erlotinib. Despite the abundant expression of EGFR protein in a broad range of epithelial cancers, EGFR mutations have not been reported in a substantial fraction of other cancers. Given recent reports of TKI-responsive cases of esophageal and pancreatic cancer, this study was designed to determine the prevalence of EGFR mutations in these gastrointestinal cancers. EXPERIMENTAL DESIGN: We sequenced exons 18 to 21 of EGFR from 21 cases of Barrett's esophagus, 5 cases of high-grade esophageal dysplasia, 17 cases of esophageal adenocarcinoma, and 55 cases of pancreatic adenocarcinoma. Subsets of esophageal (n = 7) and pancreatic cancer cases (n = 5) were obtained from patients who were subsequently treated with gefitinib or erlotinib-capecitabine, respectively. RESULTS: Mutations of EGFR were identified in two esophageal cancers (11.7%), three cases of Barrett's esophagus (14.2%), and two pancreatic cancers (3.6%). The mutations consisted of the recurrent missense L858R and in-frame deletion delE746-A750, previously characterized as activating EGFR mutations in non-small cell lung cancer. We also identified the TKI drug resistance-associated EGFR T790M mutation in an untreated case of Barrett's esophagus and the corresponding adenocarcinoma. CONCLUSION: The presence of activating mutations within EGFR in both esophageal and pancreatic adenocarcinomas defines a previously unrecognized subset of gastrointestinal tumors in which EGFR signaling may play an important biological role. EGFR mutations in premalignant lesions of Barrett's esophagus also point to these as an early event in transformation of the esophageal epithelium. The role of genotype-directed TKI therapy should be tested in prospective clinical trials.
    • Evaluation of community pharmacists' recommendations to standardized patient scenarios.

      Rutter, Paul M.; Horsley, Emma; Brown, David T. (Harvey Whitney Books Company, 2004)
      BACKGROUND: Recent trends in the global nonprescription drug market show a massive increase in medicine availability to the general public. This places greater responsibility on community pharmacists to ensure that patients exercise self-care appropriately. This small study examined pharmacist performance in facilitating patient self-care. OBJECTIVE: To determine whether an appropriate course of action was taken by UK community pharmacists practicing in one geographic area when presented with 1 of 2 case scenarios: headache or abdominal pain. METHODS: A covert researcher, posing as a patient, spoke with the pharmacist on duty at each of 30 pharmacies, asking for advice on one of the scenarios. The pharmacist was interviewed and their responses were examined by an expert panel. RESULTS: Thirty pharmacies were visited resulting in 28 consultations, 14 each for both scenarios. Pharmacists performed better when counseling the "patient" with abdominal pain than headache. The majority of questions asked were categorized as being relevant by the panel (66% for headache, 89% for abdominal pain), although in both scenarios, the expected outcome of referral was observed only in 7 of the consultations for headache and in 8 cases for abdominal pain. Questioning centered on quantification and clarification of the presenting problem and rarely on questions related to history taking. CONCLUSIONS: Performance levels of the community pharmacists varied considerably. For all pharmacists to perform well, greater emphasis should be placed on eliciting information from the patient to ensure that the appropriate course of action is taken.
    • Experimental and Numerical Investigation of the Effect of Pellet Size on the Adsorption Characteristics of Activated Carbon/Ethanol

      Elsayed, A.; Mahmoud, S.; Al-Dadah, R.; Bowen, J.; Kaialy, Waseem (2014)
      Low temperature adsorption cooling is an attractive heat powered cooling technology suitable for various applications where waste heat is available. The use of activated carbon as adsorbent with ethanol offers potential for low temperature cooling applications like the food retail industry. Activated carbons are commercially available in the form of powders, granules and pellets. Although powder materials have the advantage of good adsorption kinetics but they are difficult to integrate in adsorption beds. Pellets and granules come at various shapes and sizes and can be effectively accommodated in adsorption beds but offer slower kinetics compared to the powder form. This work experimentally and numerically investigates the effect of pellet size on the ethanol adsorption characteristics of Norit RX3 activated carbon. Dynamic vapour sorption (DVS) testing was used for measuring the adsorption isotherms and kinetics for a range of pellet lengths ranging from 3mm to 12mm. COMSOL Multiphysics was used to simulate the adsorption effect taking into account the diffusion process. Results showed that increasing the pellet dimension in terms of diameter and length reduces the adsorption kinetics.
    • Expression profiling in ependymoma reveals differences between benign and anaplastic ependymoma

      Suarez-Merino, Blanca; Hubank, Mike; Hayward, Richard; Harkness, William; Thompson, Dominic; Phipps, Kim; Revesz, Tamas; Darling, John L.; Thomas, David G.; Warr, Tracy (Society for Neuro-Oncology and Duke University Press, 2003)
      Ependymomas arise from the ependymal cells lining the ventricular system of the CNS and account for approximately 10% of paediatric brain tumours. Approximately 70% of ependymomas are histologically benign and correspond to WHO grade II, whilst the remainder are anaplastic (WHO grade III). The 5-year survival rates in children are 34–45%, with local recurrence being the major source of therapeutic failure. Anaplasia does not appear to be associated with worse prognosis, and at present there are no molecular or genetic markers which can be used as predictors of outcome. Indeed, the genetic events that contribute to the pathogenesis of ependymoma are essentially unknown. We have used human oligonucleotide arrays to generate gene expression profiles in 10 ependymoma samples from patients with different histopathological/clinical parameters in order to identify new prognostic markers. Our sample group is composed of 7 ependymoma (WHO grade II) and 3 anaplastic ependymoma (WHO grade III). Three patients were <3 years of age at first presentation. Five tumours have chromosomal aberrations identified by comparative genomic hybridisation (CGH). Our preliminary data show that overexpression of specific functional categories of genes is dependant on histology when compared to normal controls. Cell cycle and adhesion related genes, oncogenes, and genes involved in apoptosis were mainly overexpressed in anaplastic tumours. Benign tumours, however, overexpressed mainly growth factor related genes. Some common candidates emerged for all tumours; Wee1+, a cell cycle related gene that regulates entry into mitosis, was up to six fold overexpressed in tumours. The oncogene c-myc, which maps to an amplicon at 8q24 detected by CGH in a subset of ependymomas, was also overexpressed in some tumours and may be an interesting candidate in their development. To our knowledge, none of these genes have been associated previously with this class of brain tumours. Further analysis of differential gene expression profiles using large series of tumours will help in the identification of molecular markers. This information, when linked to clinical and pathology data, could also help in the classification of these tumours and the choice of therapy.
    • Framing stress and associated behaviours at work: an ethnography study in the United Kingdom

      Hampton, Paul; Chinyio, Ezekiel; Riva, Silvia (Emerald Publishing Group, 2019-02-01)
      Aim: The purpose is to understand more precisely the culture and interpersonal behaviours associated with stress. Methods: The research was conducted using a qualitative approach through an ethnographic methodology in relation to three companies. The greater part of the data collection period was structured into observations that ranged between 2 and 4 hours per day, 1 to 3 days per week, for a period of 6 months. A total of 10 sites were explored; and on each site, the observations involved activities by 5 to 20 people. Findings: The results showed the pivotal importance of interpersonal relationships in coping with the uncertainty of working conditions, the coordination of team-work, and managing responsibilities and power interactions. It was found that the impact of stress is multifaceted, affecting the physical status, interpersonal relationships, work performance, and emotional wellbeing of construction workers. The workers who were studied emphasised five sources of support that help moderate work-related stress: additional tools such as communication systems and software, a facilitated access to professional help (e.g. psychological services), organisational changes in leadership, provision of resources for the wellbeing of personnel (e.g. job training) and better teamwork. Practical implications: The study underlines the importance of dedicated services for stress management and specific training-related abilities devoted to reinforcing positive person-organization dynamics. In particular, the abilities should relate to managing the impact of stress in terms of physique, interpersonal relationships, work performance, and emotional well-being. Originality/value: This is one of the first studies to adopt a psychological perspective for understanding construction scenarios and phenomena and was conducted by a qualified psychologist.
    • Fungal gene sequences make excellent models for teaching data mining

      Hooley, Paul; Burns, Alan T. H.; Whitehead, Michael P. (Elsevier Science Direct, 2004)
      A brief introductory exercise in the use of on-line databases to examine fungal genes and their products is described. Fungal genes make particularly good teaching models owing to their relatively simple eukaryotic structure and wide range of homologues in higher organisms including humans. An evaluation of students' reactions to the exercise is included.
    • Fungal osmotolerance.

      Hooley, Paul; Fincham, Daron A.; Whitehead, Michael P.; Clipson, Nicholas J. W. (Elsevier Science Direct, 2003)
      Abstract not available. Article Outline as follows: I. Introduction. II. Physiological Mechanisms of Osmotolerance. III. Control of Osmotic Responses at the Molecular Level. A. Osmotic Adjustment in Yeasts: 1. Saccharomyces cerevisiae. a. Regulation of osmotic responses at the molecular level. b. Transcriptional responses. 2. Debaryomyces hansenii. 3. Other Yeasts. B. Oscp>smotic Adjustment in Filamentous Fungal Species: 1. Genes with “Osmotic” Phenotypes. 2. Molecular Approaches. 3. A. nidulans as a Model Expression System. 4. Life Cycle Stage Sensitivity. 5. Regulation of Osmotic Responses in A. nidulans. a. Introduction to transcription factors. IV. Conclusions. A. Comparisons of Yeast and Filamentous Models. B. Sporulation and Development. C. Origins of Fungal Stress Response Genes. D. Specificity of Stress Responses. E. Concluding Remarks. Acknowledgements. References.
    • Gain of 1q and loss of 22 are the most common changes detected by comparative genomic hybridisation in paediatric ependymoma.

      Ward, Samantha; Harding, Brian; Wilkins, Peter; Harkness, William; Hayward, Richard; Darling, John L.; Thomas, David G.; Warr, Tracy (Wiley Interscience, 2001)
      Ependymomas are the third most common brain tumour in the paediatric population. Although cytogenetic and molecular analyses have pinpointed deletions of chromosomes 6q, 17, and 22 in a subset of tumours, definitive patterns of genetic aberrations have not been determined. In the present study, we analysed 40 ependymomas from paediatric patients for genomic loss or gain using comparative genomic hybridisation (CGH). Eighteen of the tumours (45%) had no detectable regions of imbalance. In the remaining cases, the most common copy number aberrations were loss of 22 (25% of tumours) and gain of 1q (20%). Three regions of high copy number amplification were noted at 1q24-31 (three cases), 8q21-23 (two cases), and 9p (one case). Although there was no association with the loss or gain of any chromosome arm or with benign versus anaplastic histologic characteristics, the incidence of gain of 7q and 9p and loss of 17 and 22 was significantly higher in recurrent versus primary tumours. This study has identified a number of chromosomal regions that may contain candidate genes involved in the development of different subgroups of ependymoma.
    • Gene encoding the collagen type I and thrombospondin receptor CD36 is located on chromosome 7q11.2.

      Fernández-Ruiz, Elena; Armesilla, Angel Luis; Sánchez-Madrid, Francisco; Vega, Miguel A. (Elsevier BV, 1993)
      The human CD36 is a member of a gene family of structurally related glycoproteins and functions as a receptor for collagen type I and thrombospondin. CD36 also binds to red blood cells infected with the human malaria parasite Plasmodium falciparum. In the present study, the CD36 gene was assigned to chromosome 7 by using the polymerase chain reaction with DNA from human-hamster somatic cell hybrids. Furthermore, the use of a CD36 genomic probe has allowed the localization of the CD36 locus to the 7q11.2 band by fluorescence in situ hybridization coupled with GTG-banding.
    • Generating monoclonal antibody probes and techniques for characterizing and localizing reactivity to antigenic determinants

      Nelson, Paul N. (Oxford: Oxford University Press, 2001)
      This book: Epitope Mapping covers all the major methods for the identification and definition of epitopes. The Pepscan assay is used to define B cell epitopes and makes use of synthetic peptides but can only be used if the amino acid sequence is known. It can be adapted for the delineation of both helper T cells and cytotoxic T cells. The identification of combined B and T cell epitopes can also be achieved using synthetic peptides. There are other methodologies for analysing for cytotxic T cell epitopes such as the purification of antigens presented by MHC class I molecules and expression cloning. Site directed mutagenesis is also a powerful tool in epitope mapping and can be used to evaluate the role of single amino acids in immune complex formation. Protein footprinting makes use of monoclonal antibodies produced by hybridoma technology and relies on the fact that the epitope is protected from cleavage when bound as an antibody-antigen complex. It is only useful for small antigens. Other monoclonal antibody assays such as enzyme linked immunosorbent assay and haemaglutination and slot-blotting may also be used in epitope mapping. Random phage display libraries bring together the genetic and amino acid peptide sequence and can be screened with antibody and the resulting peptide DNA sequenced to confirm the amino acid sequence of a specific eptiope. Investigation of carbohydrates can also be useful to eptitope mapping as deglycosylation can lead to loss of antigenic activity. Epitopes are important to the pharmaceutical industry and wherever appropriate, pharmaceutical applications of the methods described are included. For each method there is a description of the technology, protocols, trouble-shooting, and advice on when to use the method. This book will therefore be invaluable to any researcher involved in epitope mapping.
    • Generation and characterization of monoclonal antibodies to the neural crest.

      Shakil, T.; Richardson, M. K.; Waldron, E.E.; Conde, Gillian; Wood, S.; Bland, Y.; Reynolds, Gary; Murray, Paul G.; Nelson, Paul N. (New Rochelle (NY): Mary Ann Liebert, Inc., 2001)
      The generation of monoclonal antibodies (MAbs) specific for quail neural crest may provide valuable tools for studying the differentiation of embryonic precursor cells. Unfortunately, relatively few antibodies are available because of the difficulty in obtaining sufficient cells for in vivo immunization strategies. We have overcome this problem by using intrasplenic immunization with formaldehyde-fixed cells harvested from neural crest cultures. In addition, booster injections of cultured whole-embryo cells were administered intraperitoneally. Following two fusions, a total of 18 hybridomas were generated with antibody reactivity to the cytoplasm of neural crest cells. Furthermore, 32 were reactive against both somite (a noncrest mesodermal control) and crest cultures, whilst 15 were not reactive. Out of those hybridomas reactive with neural crest, six designated 160D, 164D, OE, 12E, 120E and 124E were further characterized. Interestingly MAb supernatants OE, 12E, 120E, and 124E exhibited reactivity against some but not all neural crest cells suggesting that they might recognise subpopulations. Immunoglobulin isotyping of supernatants revealed that 4 (160D, 164D, OE, and 120E) were IgM and 2 (12E and 124E) were IgG(2b). On assessing their reactivity against human tissue sections, all six hybridoma supernatants cross-reacted with neuroendocrine cells within appendix, colon and rectum. These MAbs could provide novel reagents for the understanding of neural crest development.
    • Genomic and transcriptomic characterisation of undifferentiated pleomorphic sarcoma of bone

      Ali, Naser M.; Niada, Stefania; Brini, Anna T.; Morris, Mark R.; Kurusamy, Sathishkumar; Alholle, Abdullah; Huen, David; Antonescu, Cristina R.; Tirode, Franck; Sumathi, Vaiyapuri; et al. (Wiley, 2018-10-03)
      Undifferentiated pleomorphic sarcoma of bone (UPSb), is a rare primary bone sarcoma that lacks a specific line of differentiation. There is very little information about the genetic alterations leading to tumourigenesis or malignant transformation. Distinguishing between UPSb and other malignant bone sarcomas, including dedifferentiated chondrosarcoma and osteosarcoma, can be challenging due to overlapping features. To explore the genomic and transcriptomic landscape of UPSb tumours, whole-exome sequencing (WES) and RNA Sequencing (RNA-Seq) were performed on UPSb tumours. All tumours lacked hotspot mutations in IDH1/2 132 or 172 codons, thereby excluding the diagnosis of dedifferentiated chondrosarcoma. Recurrent somatic mutations in TP53 were identified in 4/14 samples (29%). Moreover, recurrent mutations in histone chromatin remodelling genes, including H3F3A, ATRX and DOT1L, were identified in 5/14 samples (36%), highlighting the potential role of deregulated chromatin remodelling pathways in UPSb tumourigenesis. The majority of recurrent mutations in chromatin remodelling genes identified here are reported in COSMIC, including the H3F3A G35 and K36 hotspot residues. Copy number alteration analysis identified gains and losses in genes that have been previously altered in UPSb or UPS of soft tissue. Eight somatic gene fusions were identified by RNA-Seq, two of which, CLTC-VMP1 and FARP1-STK24, were reported previously in multiple cancers. Five gene fusions were genomically characterised. Hierarchical clustering analysis, using RNA-Seq data, distinctly clustered UPSb tumours from osteosarcoma and other sarcomas, thus molecularly distinguishing UPSb from other sarcomas. RNA-Seq expression profiling analysis and quantitative RT-PCR showed an elevated expression in FGF23 which can be a potential molecular biomarker in UPSb. To our knowledge, this study represents the first comprehensive WES and RNA-Seq analysis of UPSb tumours revealing novel protein-coding recurrent gene mutations, gene fusions and identifying a potential UPSb molecular biomarker, thereby broadening the understanding of the pathogenic mechanisms and highlighting the possibility of developing novel targeted therapeutics.
    • Germline Mutations in the CDKN2B Tumor Suppressor Gene Predispose to Renal Cell Carcinoma.

      Jafri, Mariam; Wake, Naomi C; Ascher, David B; Pires, Douglas E V; Gentle, Dean; Morris, Mark R.; Rattenberry, Eleanor; Simpson, Michael A; Trembath, Richard C; Weber, Astrid; et al. (American Association for Cancer Research, 2015-07)
      Familial renal cell carcinoma (RCC) is genetically heterogeneous and may be caused by mutations in multiple genes, including VHL, MET, SDHB, FH, FLCN, PTEN, and BAP1. However, most individuals with inherited RCC do not have a detectable germline mutation. To identify novel inherited RCC genes, we undertook exome resequencing studies in a familial RCC kindred and identified a CDKN2B nonsense mutation that segregated with familial RCC status. Targeted resequencing of CDKN2B in individuals (n = 82) with features of inherited RCC then revealed three candidate CDKN2B missense mutations (p.Pro40Thr, p.Ala23Glu, and p.Asp86Asn). In silico analysis of the three-dimensional structures indicated that each missense substitution was likely pathogenic through reduced stability of the mutant or reduced affinity for cyclin-dependent kinases 4 and 6, and in vitro studies demonstrated that each of the mutations impaired CDKN2B-induced suppression of proliferation in an RCC cell line. These findings identify germline CDKN2B mutations as a novel cause of familial RCC.
    • Glucose induces and leptin decreases expression of uncoupling protein-2 mRNA in human islets.

      Brown, James E. P.; Thomas, Steven; Digby, Janet E.; Dunmore, Simon J. (Elsevier BV, 2002)
      Elevated islet uncoupling protein-2 (UCP-2) impairs beta-cell function and UCP-2 may be increased in clinical obesity and diabetes. We investigated the effects of glucose and leptin on UCP-2 expression in isolated human islets. Human islets were incubated for 24 h with glucose (5.5-22 mmol/l)+/-leptin (0-10 nmol/l). Some islet batches were incubated at high (22 mmol/l), and subsequently lower (5.5 mmol/l), glucose to assess reversibility of effects. Leptin effects on insulin release were also measured. Glucose dose-dependently increased UCP-2 expression in all islet batches, maximally by three-fold. This was not fully reversed by subsequently reduced glucose levels. Leptin decreased UCP-2 expression by up to 75%, and maximally inhibited insulin release by 47%, at 22 mmol/l glucose. This is the first report of UCP-2 expression in human islets and provides novel evidence of its role in the loss of beta-cell function in diabetes.
    • Granulocyte-macrophage colony-stimulating factor, phorbol ester, and sodium butyrate induce the CD11c integrin gene promoter activity during myeloid cell differentiation.

      Rubio, M.A.; Lopez-Rodriguez, C.; Nueda, A.; Aller, P.; Armesilla, Angel Luis; Vega, Miguel A.; Corbí, A.L. (American Society of Hematology, 1995)
      To analyze the activity of the CD11c promoter during myeloid differentiation without the limitations of transient expression systems, we have stably transfected the myeloid U937 cell line with the pCD11C361-Luc plasmid, in which the expression of the firefly luciferase cDNA is driven by the CD11c promoter region -361/+43, previously shown to confer myeloid specificity to reporter genes. The stable transfectants (U937-C361) retained the ability to differentiate in response to phorbol-ester (PMA), sodium butyrate (SB), granulocyte-macrophage colony-stimulating factor (GM-CSF), and other differentiating agents. U937-C361 differentiation correlated with increased cellular luciferase levels, showing the inducibility of the CD11c promoter during myeloid differentiation and establishing the U937-C361 cells as a suitable system for studying the myeloid differentiation-inducing capacity of cytokines, growth, factors, and other biological response modifiers. Unexpectedly, the inducibility of the CD11c gene promoter showed distinct kinetics and magnitude on the PMA-, SB-, GM-CSF-triggered differentiation. Moreover, SB synergized with either PMA or GM-CSF in enhancing both the CD11c promoter activity and the cell surface expression of p150,95 on differentiating U937 cells. Furthermore, we showed the existence of a c-Myb-binding site at -85, the importance of the -99/-61 region in the CD11c promoter inducibility during PMA- or SB-triggered differentiation, and the dependency of the GM-CSF and PMA responsiveness of the CD11c promoter on an intact AP-1-binding site located at -60. These results, together with the lack of functional effect of mutations disrupting the Sp1-and Myb-binding sites within the proximal region of the CD11c promoter, indicate that the myeloid differentiation pathways indicated by SB and phorbol esters (or GM-CSF) activate a distinct set of transcription factors and show that the myeloid differentiation-inducibility of the CD11c gene maps to the -99/-53 proximal region of the promoter.