http://espace.library.uq.edu.au/ The University of Queensland en Fez http://blogs.law.harvard.edu/tech/rss http://espace.library.uq.edu.au/view/UQ:171865 Dendritic cell (DC) differentiation is abnormal in type 1 diabetes mellitus (T1DM). However, the nature of the relationship between this abnormality and disease pathogenesis is unknown. We studied the LPS response in monocytes and monocyte-derived DCs isolated from T1DM patients and from non-T1DM controls. In T1DM patients, late LPS-mediated nuclear DNA binding by RelA, p50, c-Rel, and RelB was impaired as compared with type 2 DM, rheumatoid arthritis, and healthy subjects, associated with impaired DC CD40 and MHC class I induction but normal cytokine production. In TIDM monocytes, RelA and RelB were constitutively activated, and the src homology 2 domain-containing protein tyrosine phosphatase (SHP-1), a negative regulator of NF-{kappa}B, was overexpressed. Addition of sodium stibogluconate, a SHP-1 inhibitor, to DCs differentiating from monocyte precursors restored their capacity to respond to LPS in ~60% of patients. The monocyte and DC NF-{kappa}B response to LPS is thus a novel phenotypic and likely pathogenetic marker for human T1DM. SHP-1 is at least one NF-{kappa}B regulatory mechanism which might be induced as a result of abnormal inflammatory signaling responses in T1DM monocytes. 2009-03-25T14:50:01Z Mollah, Zia U.A. og Pai, Saparna og Craig Moore og O'Sullivan, Brendan J. og Harrison, Matthew J. og Peng, Judy og Phillips, Karen og Prins, Johannes B. og Cardinal, John og Thomas, Ranjeny http://espace.library.uq.edu.au/view/UQ:35470 The high affinity receptor for human granulocyte-macrophage colony-stimulating factor (GM-CSF) consists of a cytokine-specific alpha-subunit (hGMR alpha) and a common signal-transducing beta-subunit (hpc) that is shared with the interleukin-3 and -5 receptors, We have previously identified a constitutively active extracellular point mutant of hpc, I374N, that can confer factor independence on murine FDC-P1 cells but not BAF-B03 or CTLL-2 cells (Jenkins, B. J., D'Andrea, R. J., and Gonda, T. J. (1995) EMBO J. 14, 4276-4287), This restricted activity suggested the involvement of cell type-specific signaling molecules in the activation of this mutant. We report here that one such molecule is the mouse GMR alpha (mGMR alpha) subunit, since introduction of mGMR alpha, but not hGMR alpha, into BAF-B03 or CTLL-2 cells expressing the I374N mutant conferred factor independence, Experiments utilizing mouse/human chimeric GMR alpha subunits indicated that the species specificity lies in the extracellular domain of GMRa. Importantly, the requirement for mGMR alpha correlated with the ability of I374N (but not wild-type hpc) to constitutively associate with mGMRa. Expression of I374N in human factor-dependent UT7 cells also led to factor-independent proliferation, with concomitant up-regulation of hGMR alpha surface expression. Taken together, these findings suggest a critical role for association with GMR alpha in the constitutive activity of I374N. 2007-08-13T11:00:53Z Jenkins, BJ og Le, F og Gonda, TJ http://espace.library.uq.edu.au/view/UQ:35097 2007-08-13T10:41:33Z McCormack, MP og Jenkins, BJ og Gonda, TJ http://espace.library.uq.edu.au/view/UQ:34947 2007-08-13T10:30:59Z McCormack, MP og Gonda, TJ http://espace.library.uq.edu.au/view/UQ:57457 2007-08-13T16:39:13Z Gonda, TJ og DAndrea, RJ http://espace.library.uq.edu.au/view/UQ:36011 2007-08-13T11:28:13Z DAndrea, RJ og Mulhern, T og Jones, K og Butcher, C og Blake, T og Yandell, C og Booker, G og Vadas, M og Lopez, A og Bagley, C og Gonda, T http://espace.library.uq.edu.au/view/UQ:76387 Human papillomavirus-like particles (HPV-VLP) are a candidate vaccine for prevention of HPV infection, and also are a candidate for an immunogenic delivery system for incorporated antigen. VLP activate in vitro generated dendritic cells (DC) but not Langerhans cells (LC); however, the mechanism of this activation is unknown. We have shown that uptake and activation of DC by VLP involves proteoglycan receptors and can be inhibited by heparin. Heparin has been shown to activate DC by signalling through Toll-like receptor 4 (TLR4) and nuclear factor (NF)-kappaB. The pathway of DC activation by VLP was further investigated in the present study. Exposure to VLP induced costimulatory molecule expression, RelB translocation and IL-10 production by DC but not by LC. The lack of LC activation was reversible when TGF-beta was removed from the LC medium. VLP-induced induction of costimulatory molecule expression, RelB activation and cytokine secretion by DC was blocked by inhibition of NF-kappaB activation, heparin or TLR4 mAb. The data provide evidence that HPV-VLP signal DC through a pathway involving proteoglycan receptors, TLR4 and NF-kappaB, and shed light on the mechanism by which VLP stimulate immunity in the absence of adjuvants in vivo. LC may resist activation in normal epithelium abundant in TGF-beta, but not in situations in which TGF-beta concentrations are reduced. 2007-08-15T06:26:38Z Yan, M. og Peng, J. C. og Abdul Jabbar, I. og Liu, X. og Filguueira, L. og Frazer, I. H. og Thomas, R. http://espace.library.uq.edu.au/view/UQ:165352 Human papillomavinis-like particles (HPV-VLP) are a candidate vaccine for prevention of HPV infection, asid also are a candidate for an immunogenic delivery system for ineorporated antigen. VLP activate in vitro generated dendritic cells (DC) but not Langerhans cells (LC); however, the mechanism of this activation is unknown. We have shown that uptake and activation of DC by VLP involves proteoglycan receptors and can be inhibited by heparin. Heparin has been shown to activate DC by signalling through Toll-like receptor 4 (TLR4) and nuclear faetor (NF)-KB. The pathway of DC aetivation by VLP was further investigated in the present study. Exposure to VLP induced eostimulatory tnoleeiile expression. RelB transioeation and lL-IO production by DC but not hy LC. The lack of LC activation was reversible when TGF-p was removed from the LC medium. VLP-indueed induction of costimulatory molecule expression. RelB aetivation and eytokine secretion by DC was bloeked by inhihition of NF-icB activation, heparin or TLR4 mAb. The data provide evidence that HPV-VLP signal DC through a pathway involving proteoglyean receptors. TLR4 and NF-KB, and shed light on the mechanism by which VLP stimulate immunity in the absence of adjuvants in vivo. LC may resist activation in nonnal epithelium abundant in TCiF-p. but not in situations in whieh TGF-[3 concentrations are redueed. 2009-02-27T13:27:19Z Yan, Mengyong og Peng, Judy og Jabbar, Ibtissam og Lui, Xiaosong og Filgueira, Luis og Frazer, Ian og Thomas, Ranjeny http://espace.library.uq.edu.au/view/UQ:44529 Natural killer (NK) cells are innate effector lymphocytes necessary for defence against stressed, microbe-infected, or malignant cells. NK cells kill target cells by either of two major mechanisms that require direct contact between NK cells and target cells. In the first pathway, cytoplasmic granule toxins, predominantly a membrane-disrupting protein known as perforin, and a family of structurally related serine C, proteases (granzymes) with various substrate specificities, are secreted by exocytosis and together induce apoptosis of the target cell. The granule-exocytosis pathway potently activates cell-death mechanisms that operate through the activation of apoptotic cysteine proteases (caspases), but can also cause cell death in the absence of activated caspases. The second pathway involves the engagement of death receptors (e.g. Fas/CD95) on target cells by their cognate ligands (e.g. FasL on NK cells, resulting in classical caspase-dependent apoptosis. The comparative role of these pathways in the pathophysiology of many diseases is being dissected by analyses of gene-targeted mice that lack these molecules, and humans who have genetic mutations affecting these pathways. We are also now learning that the effector function of NK cells is controlled by interactions involving specific NK cell receptors and their cognate ligands, either on target cells, or other cells of the immune system. This review will discuss the functional importance of NK cell cytotoxicity and the receptor/ligand interactions that control these processes. (C) 2004 Elsevier Ltd. All rights reserved. 2007-08-13T15:00:47Z Smyth, MJ og Cretney, E og Kelly, JM og Westwood, JA og Street, SEA og Yagita, H og Takeda, K og van Dommelen, SLH og Degli-Espostic, MA og Hayakawa, Y http://espace.library.uq.edu.au/view/UQ:164052 In contrast to cutaneous melanoma, there is no evidence that BRAF mutations are involved in the activation of the mitogen-activated protein kinase (MAPK) pathway in uveal melanoma, although there is increasing evidence that this pathway is activated frequently in the latter tumours. In this study, we performed mutation analysis of the RAS and BRAF genes in a panel of 11 uveal melanoma cell lines and 19 primary uveal melanoma tumours. In addition, Western blot and immunohistochemical analyses were performed on downstream members of the MAPK pathway in order to assess the contribution of each of these components. No mutations were found in any of the three RAS gene family members and only one cell line carried a BRAF mutation (V599E). Despite this, mitogenactivated protein kinase/extracellular signal-regulated kinase kinase (MEK), ERK and ELK were constitutively activated in all samples. These data suggest that activation of the MAPK pathway is commonly involved in the development of uveal melanoma, but occurs through a mechanism different to that of cutaneous melanoma. 2009-02-11T18:24:53Z Zuidervaart, W. og van Nieuwpoort, F. og Stark, M. og Dijkman, R. og Packer, L. og Borgstein, A. -M. og Pavey, S. og van der Velden, P. og Out, C. og Jager, M. J. og Hayward, N. K. og Gruis, N. A. http://espace.library.uq.edu.au/view/UQ:123139 New lines of treatment targeting cytokines have been successfully developed recently and are now widely used in therapy. They are based on passive administration of cytokine inhibitors either soluble receptors or mAbs and the major example is TNFalpha in rheumatoid arthritis (RA). Since a few years, our group has developed a novel alternative approach targeting cytokines by using active immunization against biologically inactive but immunogenic cytokine derivatives. In the present work, we present a new aspect of this research, based on immunization against specific cytokine peptides chosen by molecular modelling. We could elicit a significant humoral response against four TNFalpha peptides by active immunization, and show that the Abs generated cross-reacted with the native cytokine with good titers as determined by ELISA. Interestingly, during coimmunization experiments with couples of peptides, one showed a clear immunodominant effect over the other. Overall, we could not show the neutralization of TNFalpha biological activity in vitro by the immunized sera, but it seems that it is not a prerequisite to observe clinical efficacy. Indeed, using the LPS/galactosamine-induced shock, we could demonstrate that one of the four peptides tested conferred a clinical protection. These results validate the feasibility and efficacy of active immunization against cytokine peptides, and confirm that active immunization against cytokines could represent in the future an alternative to passive immunization in many diseases. (C) 2004 Elsevier Ltd. All rights reserved. 2008-01-25T16:20:02Z Capini, CJ og Bertin-Maghit, SM og Bessis, N og Haumont, PM og Bernier, EM og Muel, EG og Laborie, MA og Autin, L og Paturance, S og Chomilier, J og Boissier, MC og Briand, JP og Muller, S og Cavaillon, JM og Therwath, A og Zagury, JF http://espace.library.uq.edu.au/view/UQ:65351 One of the major regulators of mitosis in somatic cells is cdc25B. cdc25B is tightly regulated at multiple levels. The final activation step involves the regulated binding of 14-3-3 proteins. Previous studies have demonstrated that Ser-323 is a primary 14-3-3 binding site in cdc25B, which influences its activity and cellular localization. 14-3-3 binding to this site appeared to interact with the N-terminal domain of cdc25B to regulate its activity. The presence of consensus 14-3-3 binding sites in the N-terminal domain suggested that the interaction is through direct binding of the 14-3-3 dimer to sites in the N-terminal domain. We have identified Ser-151 and Ser-230 in the N-terminal domain as functional 14-3-3 binding sites utilized by cdc25B in vivo. These low affinity sites cooperate to bind the 14-3-3 dimer bound to the high affinity Ser-323 site, thus forming an intramolecular bridge that constrains cdc25B structure to prevent access of the catalytic site. Loss of 14-3-3 binding to either N-terminal site relaxes cdc25B structure sufficiently to permit access to the catalytic site, and the nuclear export sequence located in the N-terminal domain. Mutation of the Ser-323 site was functionally equivalent to the mutation of all three sites, resulting in the complete loss of 14-3-3 binding, increased access of the catalytic site, and access to nuclear localization sequence. 2007-08-15T01:25:11Z Giles, N. og Forrest, A. og Gabrielli, B. http://espace.library.uq.edu.au/view/UQ:163645 Adiponectin is a recently described adipokine that has been recognized as a key regulator of insulin sensitivity and tissue inflammation. It is produced by adipose tissue (white and brown) and circulates in the blood at very high concentrations. It has direct actions in liver, skeletal muscle and the vasculature, with prominent roles to improve hepatic insulin sensitivity, increase fuel oxidation [via up-regulation of adenosine monophosphate-activated protein kinase (AMPK) activity] and decrease vascular inflammation. Adiponectin exists in the circulation as varying molecular weight forms, produced by multimerization. Recent data indicate that the high-molecular weight (HMW) complexes have the predominant action in the liver. In contrast to other adipokines, adiponectin secretion and circulating levels are inversely proportional to body fat content. Levels are further reduced in subjects with diabetes and coronary artery disease. Adiponectin antagonizes many effects of tumour necrosis factor-α(TNF-α) and this, in turn, suppresses adiponectin production. Furthermore, adiponectin secretion from adipocytes is enhanced by thiazolidinediones (which also act to antagonize TNF-α effects). Thus, adiponectin may be the common mechanism by which TNF-α promotes, and the thiazolidinediones suppress, insulin resistance and inflammation. Two adiponectin receptors, termed AdipoR1 and AdipoR2, have been identified and these are ubiquitously expressed. AdipoR1 is most highly expressed in skeletal muscle and has a prominent action to activate AMPK, and hence promote lipid oxidation. AdipoR2 is most highly expressed in liver, where it enhances insulin sensitivity and reduces steatosis via activation of AMPK and increased peroxisome-proliferator-activated receptor α ligand activity. T-cadherin, which is expressed in endothelium and smooth muscle, has been identified as an adiponectin-binding protein with preference for HMW adiponectin multimers. Given the low levels of adiponectin in subjects with the metabolic syndrome, and the beneficial effect of the adipokine in animal studies, there is exciting potential for adiponectin replacement therapy in insulin resistance and related disorders 2009-02-10T13:26:49Z Whitehead, J. P. og Richards, A. A. og Hickman, I. J. og Macdonald , G. A. og Prins, J. B. http://espace.library.uq.edu.au/view/UQ:81816 Adiponectin is a recently described adipokine that has been recognized as a key regulator of insulin sensitivity and tissue inflammation. It is produced by adipose tissue (white and brown) and circulates in the blood at very high concentrations. It has direct actions in liver, skeletal muscle and the vasculature, with prominent roles to improve hepatic insulin sensitivity, increase fuel oxidation [via up-regulation of adenosine monophosphate-activated protein kinase (AMPK) activity] and decrease vascular inflammation. Adiponectin exists in the circulation as varying molecular weight forms, produced by multimerization. Recent data indicate that the high-molecular weight (HMW) complexes have the predominant action in the liver. In contrast to other adipokines, adiponectin secretion and circulating levels are inversely proportional to body fat content. Levels are further reduced in subjects with diabetes and coronary artery disease. Adiponectin antagonizes many effects of tumour necrosis factor-alpha(TNF-alpha) and this, in turn, suppresses adiponectin production. Furthermore, adiponectin secretion from adipocytes is enhanced by thiazolidinediones (which also act to antagonize TNF-alpha effects). Thus, adiponectin may be the common mechanism by which TNF-alpha promotes, and the thiazolidinediones suppress, insulin resistance and inflammation. Two adiponectin receptors, termed AdipoR1 and AdipoR2, have been identified and these are ubiquitously expressed. AdipoR1 is most highly expressed in skeletal muscle and has a prominent action to activate AMPK, and hence promote lipid oxidation. AdipoR2 is most highly expressed in liver, where it enhances insulin sensitivity and reduces steatosis via activation of AMPK and increased peroxisome-proliferator-activated receptor alpha ligand activity. T-cadherin, which is expressed in endothelium and smooth muscle, has been identified as an adiponectin-binding protein with preference for HMW adiponectin multimers. Given the low levels of adiponectin in subjects with the metabolic syndrome, and the beneficial effect of the adipokine in animal studies, there is exciting potential for adiponectin replacement therapy in insulin resistance and related disorders. 2007-08-15T09:58:48Z Whitehead, J. P. og Richards, A. A. og Hickman, I. J. og Macdonald, G. A. og Prins, J. B. http://espace.library.uq.edu.au/view/UQ:82165 Adiponectin is a secreted, multimeric protein with insulin-sensitizing, antiatherogenic, and antiinflammatory properties. Serum adiponectin consists of trimer, hexamer, and larger high-molecular-weight (HMW) multimers, and these HMW multimers appear to be the more bioactive forms. Multimer composition of adiponectin appears to be regulated; however, the molecular mechanisms involved are unknown. We hypothesize that regulation of adiponectin multimerization and secretion occurs via changes in posttranslational modifications (PTMs). Although a structural role for intertrimer disulfide bonds in the formation of hexamers and HMW multimers is established, the role of other PTMs is unknown. PTMs identified in murine and bovine adiponectin include hydroxylation of multiple conserved proline and lysine residues and glycosylation of hydroxylysines. By mass spectrometry, we confirmed the presence of these PTMs in human adiponectin and identified three additional hydroxylations on Pro71, Pro76, and Pro95. We also investigated the role of the five modified lysines in multimer formation and secretion of recombinant human adiponectin expressed in mammalian cell lines. Mutation of modified lysines in the collagenous domain prevented formation of HMW multimers, whereas a pharmacological inhibitor of prolyl- and lysyl-hydroxylases, 2,2'-dipyridyl, inhibited formation of hexamers and HMW multimers. Bacterially expressed human adiponectin displayed a complete lack of differentially modified isoforms and failed to form bona fide trimers and larger multimers. Finally, glucose-induced increases in HMW multimer production from human adipose explants correlated with changes in the two-dimensional electrophoresis profile of adiponectin isoforms. Collectively, these data suggest that adiponectin multimer composition is affected by changes in PTM in response to physiological factors. 2007-08-15T10:11:40Z Richards, Ayanthi A. og Stephens, Tim og Charlton, Hayley K. og Jones, Alun og Macdonald, Graeme A. og Prins, Johannes B. og Whitehead, Jonathan P. http://espace.library.uq.edu.au/view/UQ:58604 2007-08-14T15:15:29Z Niesler, C. og Prins, J. B. og Orahilly, S. og Siddle, K. og Montague, C. T. http://espace.library.uq.edu.au/view/UQ:67964 2007-08-14T10:52:15Z Hutley, L. og Newell, F. S. og Suchting, S. J. og Prins, J.B. http://espace.library.uq.edu.au/view/UQ:55929 Because CD4(+) T cells play a key role in aiding cellular immune responses, we wanted to assess whether increasing numbers of gene-engineered antigen-restricted CD4(+) T cells could enhance an antitumor response mediated by similarly gene-engineered CD8(+) T cells. In this study, we have used retroviral transduction to generate erbB2-reactive mouse T-cell populations composed of various proportions of CD4(+) and CD8(+) cells and then determined the antitumor reactivity of these mixtures. Gene-modified CD4(+) and CD8(+) T cells were shown to specifically secrete Tc1 (T cytotoxic-1) or Tc2 cytokines, proliferate, and lyse erbB2(+) tumor targets following antigen ligation in vitro. In adoptive transfer experiments using severe combined immunodeficient (scid) mice, we demonstrated that injection of equivalent numbers of antigen-specific engineered CD8(+) and CD4(+) T cells led to significant improvement in survival of mice bearing established lung metastases compared with transfer of unfractionated (largely CD8(+)) engineered T cells. Transferred CD4(+) T cells had to be antigen-specific (not just activated) and secrete interferon gamma (IFN-gamma) to potentiate the antitumor effect. Importantly, antitumor responses in these mice correlated with localization and persistence of gene-engineered T cells at the tumor site. Strikingly, mice that survived primary tumor challenge could reject a subsequent re-challenge. Overall, this study has highlighted the therapeutic potential of using combined transfer of antigen-specific gene-modified CD8(+) and CD4(+) T cells to significantly enhance T-cell adoptive transfer strategies for cancer therapy. 2007-08-13T15:38:43Z Moeller, M og Haynes, NM og Kershaw, MH og Jackson, JT og Teng, MWL og Street, SE og Cerutti, L og Jane, SM og Trapani, JA og Smyth, MJ og Darcy, PK http://espace.library.uq.edu.au/view/UQ:172110 We have previously shown in the rat slow-twitch soleus muscle that adrenaline greatly potentiates insulin-stimulated protein kinase B (PKB) phosphorylation without having an effect alone. However, insulin signalling capacity through the PKB pathway is higher in soleus than in fast-twitch muscles, whereas adrenaline activates phosphorylase more strongly in epitrochlearis. Therefore, the aim of the present study was to investigate the interaction between adrenaline and insulin signalling in the fast-twitch epitrochlearis muscle. Insulin increased insulin receptor substrate-1 (IRS-1)-associated phosphoinositide (PI) 3-kinase activity threefold, and adrenaline did not influence basal or insulin-stimulated PI 3-kinase activity. Insulin but not adrenaline increased PKB activity and phosphorylation of Ser473 and Thr308. It is interesting to note that adrenaline potentiated insulin-stimulated PKB activity and PKB Ser473 and Thr308 phosphorylation. These effects were mimicked by dibutyryl-cyclic adenosine monophosphate (db-cAMP). Adrenaline and db-cAMP increased glycogen synthase kinase (GSK)-3β Ser9 phosphorylation independently of PKB activation and enhanced insulin-stimulated GSK-3β Ser9 phosphorylation. Although adrenaline increased GSK-3 phosphorylation (inhibiting activity), phosphorylation of its target sites on glycogen synthase was increased, and adrenaline blocked insulin-stimulated glycogen synthase dephosphorylation of Ser641 and Ser645,649,653,657, glycogen synthase activation and glycogen synthesis. Insulin-stimulated glucose transport was not influenced by adrenaline despite the increased PKB activation. In conclusion, as in the slow-twitch soleus muscle, adrenaline potentiates insulin-stimulated PKB activation in the fast-twitch glycolytic epitrochlearis muscle without increasing IRS-1-associated PI 3-kinase activity. Furthermore, adrenaline induces phosphorylation of a pool of GSK-3 that is not involved in the regulation of glycogen metabolism. These results indicate that the combination of adrenaline and insulin may activate novel signalling molecules rather than just summing up their effects on linear pathways. 2009-03-26T13:57:32Z Jensen, Jørgen og Gronning-Wang, Line M. og Jebens, Einar og Whitehead, Jonathan P. og Zorec, Robert og Shepherd, Peter R. http://espace.library.uq.edu.au/view/UQ:79503 2007-08-15T08:23:46Z Frazer, Ian H. og Cox, J. Thomas og Mayeaux, Edward John og Franco, Eduardo L. og Moscicki, Anna-Barbara og Palefsky, Joel M. og Ferris, Daron G. og Ferenczy, Alex S. og Villa, Luisa L. http://espace.library.uq.edu.au/view/UQ:57742 Cytokines are secreted proteins that regulate important cellular responses such as proliferation and differentiation(1). Key events in cytokine signal transduction are well defined: cytokines induce receptor aggregation, leading to activation of members of the JAK family of cytoplasmic tyrosine kinases. In turn, members af the STAT family of transcription factors are phosphorylated, dimerize and increase the transcription of genes with STAT recognition sites in their promoters(1-4). Less is known of how cytokine signal transduction is switched off. We have cloned a complementary DNA encoding a protein SOCS-1, containing an SH2-domain, by its ability to inhibit the macrophage differentiation of M1 cells in response to interleukin-6. Expression of SOCS-1 inhibited both interleukin-6-induced receptor phosphorylation and STAT activation. We have also cloned two-relatives of SOCS-1, named SOCS-2 and SOCS-3, which together with the previously described CIS (ref. 5) form a new family of proteins. Transcription of all four SOCS genes is increased rapidly in response to interleukin-6, in vitro and in vivo, suggesting they may act in a classic negative feedback loop to regulate cytokine signal transduction. 2007-08-13T16:52:31Z Starr, R og Willson, TA og Viney, EM og Murray, LJL og Rayner, JR og Jenkins, BJ og Gonda, TJ og Alexander, WS og Metcalf, D og Nicola, NA og Hilton, DJ http://espace.library.uq.edu.au/view/UQ:171557 2009-03-24T14:42:36Z Ali Naderi og Hughes-Davies, L. http://espace.library.uq.edu.au/view/UQ:61190 Background. Posttransplant lymphoproliferative disease (PTLD), driven by the presence of Epstein-Barr virus (EBV), is becoming an increasingly important clinical problem after solid organ transplantation. The use of immunosuppressive therapy leads to the inhibition of the cytotoxic T cells that normally control the EBV latently infected B cells. The prognosis for many patients with PTLD is poor, and the optimal treatment strategy is not well defined. Method. This study investigates the use of a histone deacetylase inhibitor, azelaic bishydroxamic acid (ABRA), for its ability to effectively kill EBV-transformed lymphoblastoid cell lines. Results. In vitro treatment of lymphoblastoid cell lines with ABRA showed that they were effectively killed by low doses of the drug (ID50 2-5 mug/ml) within 48 hr. As well as being effective against polyclonal B-cell lines, ABHA was also shown to be toxic to seven of eight clonal Burkitt's lymphoma cell lines, indicating that the drug may also be useful in the treatment of late-occurring clonal PTLD. In addition, ABHA treatment did not induce EBV replication or affect EBV latent gene expression. Conclusion. These studies suggest that ABHA effectively kills both polyclonal and clonal B-cell lines and has potential in the treatment of PTLD. 2007-08-14T17:02:13Z Sculley, T. B. og Buck, M. og Gabrielli, B. og Parsons, P. G. og Krauer, K. G. http://espace.library.uq.edu.au/view/UQ:61244 The contribution of the UV component of sunlight to the development of skin cancer is widely acknowledged, although the molecular mechanisms that are disrupted by UV radiation (UVR) resulting in the loss of normal growth controls of the epidermal stem cell keratinocytes and melanocytes is still poorly understood. alpha-Melanocyte stimulating hormone (alpha-MSH), acting via its receptor MC1, has a key role in skin pigmentation and the melanizing response after exposure to UVR. The cell cycle inhibitor p16/CDKN2A also appears to have an important function in a cell cycle checkpoint response in skin after exposure to UVR. Both of these genes have been identified as risk factors in skin cancer, MC1R variants are associated with increased risk to both melanoma and nonmelanoma skin cancers, and p16/CDKN2A with increased risk of melanoma. Here we demonstrate that the increased expression of p16 after exposure to sub-erythemal doses of UVR is potentiated by alpha-MSH, a ligand for MC1R, and this effect is mimicked by cAMP, the intracellular mediator of alpha-MSH signaling via the MC1 receptor. This link between p16 and MC1R may provide a molecular basis for the increased skin cancer risk associated with MC1R polymorphisms. 2007-08-14T17:04:20Z Pavey, S. og Gabrielli, B. http://espace.library.uq.edu.au/view/UQ:68388 CD4-CD8 ratio is an important diagnostic measure of immune system functioning. In particular, CD4-CD8 ratio predicts the time taken for progression of HIV infection to acquired immune deficiency syndrome (AIDS) and the long-term survival of AIDS patients. To map genes that regulate differences between healthy individuals in CD4-CD8 ratio, we typed 757 highly polymorphic microsatellite markers at an average spacing of similar to5 cM across the genome in 405 pairs of dizygotic twins at ages 12, 14 and 16. We used multipoint variance components linkage analysis to test for linkage between marker loci and CD4-CD8 ratio at each age. We found suggestive evidence of linkage on chromosome 11p in 12-year-old twins (LOD=2.55, P=0.00031) and even stronger evidence of linkage in the same region at age 14 (LOD 3.51, P=0.00003). Possible candidate genes include CD5 and CD6, which encode cell membrane proteins involved in the positive selection of thymocytes. We also found suggestive evidence of linkage at other areas of the genome including regions on chromosomes 1, 3, 4, 5, 6, 12, 13, 15, 17 and 22. 2007-08-15T03:03:59Z Evans, D. M. og Zhu, G. og Duffy, D. L. og Frazer, I. H. og Montgomery, G. W. og Martin, N. G. http://espace.library.uq.edu.au/view/UQ:36281 Granulocyte-macrophage colony stimulating factor (GM-CSF), Interleukin-3 (IL-3) and Interleukin-5 (IL-5) have overlapping, pleiotropic effects on hematopoietic cells, including neutrophils, eosinophils, monocytes and early progenitor cells. The high-affinity receptors for human GM-CSF, IL-3, and IL-5 share a common beta-subunit (h beta(c)), which is essential for signalling and plays a major role in recruiting intracellular signalling molecules. While activation of the cytoplasmic tyrosine kinase JAK2 appears to be the initiating event for signalling, the immediate events that trigger this are still unclear. We have isolated a number of activated mutants of h beta(c), which can be grouped into classes defined by their state of receptor phosphorylation, their requirement for alpha subunit as a cofactor, and their activities in primary cells and cell lines. We discuss these findings with regard to the stoichiometry, activation, and signalling of the normal GM-CSF/IL-3/IL-5 receptor complexes. Specifically, this work has implications for the role of the ligand-specific alpha-subunits in initiating the signalling through the beta-subunit, the role of beta subunit dimerization as a receptor trigger, and the function of receptor tyrosine phosphorylation in generating growth and survival signals. Based on the properties of the activated mutants and the recent structures of erythropoietin receptor (Epo-R) complexes, we propose a model in which (1) activation of h beta(c) can occur via alternative states that differ with respect to stoichiometry and subunit assembly, but which all mediate proliferative responses, and (2) each of the different classes of activated mutants mimics one of these alternative states. (C) 2000 International Society for Experimental Hematology. Published by Elsevier Science Inc. 2007-08-13T11:39:51Z DAndrea, RJ og Gonda, TJ http://espace.library.uq.edu.au/view/UQ:56054 2007-08-13T15:44:14Z Narayan, S. og Fernando, G. og Frazer, I. H. og Leggatt, G. http://espace.library.uq.edu.au/view/UQ:132956 Cell cycle checkpoints respond to a wide range of stresses to prevent compromise to the integrity of the cell. The best studied checkpoints are those induced by genotoxic agents that cause DNA damage. Histone deacetylase inhibitors not only increase the acetylation state of chromatin histones, but they also perturb the cell cycle, causing both Gl and G2 phase arrests, the latter by initiating a checkpoint response. In this chapter we will describe the analysis of the histone deacetylase inhibitor-sensitive G 2 checkpoint using synchronized cell populations. 2008-03-27T09:37:57Z Beamish, Heather og Warrener, Robyn og Gabrielli, Brian G. http://espace.library.uq.edu.au/view/UQ:55601 2007-08-13T15:24:33Z Brown, AL og Wilkinson, C og Waterman, S og Salerno, D og Diakiw, S og Tsykin, A og Goodall, G og Solomon, P og Gonda, TJ og DAndrea, RJ http://espace.library.uq.edu.au/view/UQ:119687 2007-10-17T15:26:32Z Pointon, JJ og Timms, AE og Bradbury, L og Brown, MA og Wordsworth, P http://espace.library.uq.edu.au/view/UQ:73648 2007-08-15T04:45:27Z Pavey, S. og Gabrielli, B. G. http://espace.library.uq.edu.au/view/UQ:163791 Homologous recombination, a major double strand break repair pathway, plays critical roles in maintaining genome stability. Genetic polymorphisms in HR genes have been implicated in cancer risk. We report a novel assay system for evaluating polymorphisms in human homologous recombination genes using a panel of chicken DT40 repair mutants. We established mutant cell lines complemented with either wild-type or variant cDNAs of three human genes, RAD51, XRCC2, and XRCC3, and assessed their sensitivity to cisplatin and mitomycin C. DT40 mutants complemented with RAD51 coding and 5′UTR variants, and with a XRCC3 coding variant showed equivalent sensitivity as those with wild-type cDNAs. Interestingly, Xrcc2−/− DT40 cells complemented with variant XRCC2 (R188H) were more tolerant to cisplatin than those with wild-type XRCC2. Considering that the XRCC2 (R188H) allele reduces risk to epithelial ovarian cancer, the increased XRCC2 activity with the R188H polymorphism may have clinical benefit in preventing cancer risk. 2009-02-10T18:03:59Z Danoy, Patrick og Sonoda, Eiichiro og Lathrop, Mark og Takeda,Shunichi og Matsuda, Fumihiko http://espace.library.uq.edu.au/view/UQ:117258 2007-10-17T13:06:28Z Zhang, Y og Johnson, K og Wordsworth, P og Russell, G og Carr, A og Terkeltaub, RA og Brown, MA http://espace.library.uq.edu.au/view/UQ:115516 2007-10-17T11:31:52Z Lanctot, C og Bessette, M og Gaumond, M og Gingras, R og Godin, E og Moffatt, P og Salois, P og Sellin, K og St-Amant, N og Thomas, G http://espace.library.uq.edu.au/view/UQ:117038 We have developed and validated a semi-automated fluorescent method of genotyping human leucocyte antigen (HLA)-DRB1 alleles, HLA-DRB1*01-16, by multiplex primer extension reactions. This method is based on the extension of a primer that anneals immediately adjacent to the single-nucleotide polymorphism with fluorescent dideoxynucleotide triphosphates (minisequencing), followed by analysis on an ABI Prism 3700 capillary electrophoresis instrument. The validity of the method was confirmed by genotyping 261 individuals using both this method and polymerase chain reaction with sequence-specific primer (PCR-SSP) or sequencing and by demonstrating Mendelian inheritance of HLA-DRB1 alleles in families. Our method provides a rapid means of performing high-throughput HLA-DRB1 genotyping using only two PCR reactions followed by four multiplex primer extension reactions and PCR-SSP for some allele groups. In this article, we describe the method and discuss its advantages and limitations. 2007-10-17T12:56:09Z Jaakkola, E og Herzberg, I og Crane, AM og Pointon, JJ og Laiho, K og Kauppi, M og Kaarela, K og Wordsworth, BP og Tuomilehto, J og Brown, MA http://espace.library.uq.edu.au/view/UQ:135784 Recent studies have reported loss of function mutations in the LEMD3 gene, encoding an inner nuclear membrane protein that influences Smad signaling, as a cause of osteopoikilosis, Buschke-Ollendorff syndrome, and melorheostosis. We investigated LEMD3 in a three-generation family with osteopoikilosis from the Azores, an affected father and daughter from Ireland with osteopoikilosis (the daughter also had melorheostosis), and two other individuals from the UK with isolated melorheostosis. We found a novel C to T substitution at position 2032 bp (cDNA) in exon 8 of LEMD3, resulting in a premature stop codon at amino acid position 678. This mutation co-segregates with the osteopoikilosis phenotype in both the Azorean family and the Irish family. It was not detected in any of the six unaffected family members or in 342 healthy Caucasian individuals. No LEMD3 mutations were detected in the two patients with sporadic melorheostosis. The LEMD3 mutation reported was clearly the cause of osteopoikilosis in the two families but its relationship to melorheostosis in one of the family members is still unclear. Perhaps unsurprisingly in what is a segmental disease, we did not find LEMD3 mutations in peripheral-blood-derived DNA from the two other individuals with sporadic melorheostosis. The nature of the additional genetic and/or environmental influences required for the development of melorheostosis in those with osteopoikilosis requires further investigation. 2008-04-21T12:26:21Z Couto, Ana. R. og Bruges-Armas, Jacome og Peach, Chris A. og Chapmsn, Kay og Brown, Matthew A. og Wordsworth, B. Paul og Zhang, Yun http://espace.library.uq.edu.au/view/UQ:117206 Many viruses have evolved mechanisms to antagonize the interferon (IFN) system, targeting all the major components involved in receptor binding and signaling. Although a number of these vital proteins are homologous to cellular proteins involved in IFN downregulation (e.g., viral IFN regulatory factors [vIRFs]), many share little resemblance to known proteins. To determine the IFN-blocking properties of these proteins, functional assays are required. Here, we present a new and rapid functional screening method, based on the 2fTGH cell line, which is able to determine viral gene products that inhibit the IFN-alpha/Jak-Stat signaling pathway. Expression cloning of viral IFN-blocking genes into 2fTGH and consequent selection with IFN-alpha and 6-thioguanine result in the outgrowth of cells that are no longer responsive to IFN-alpha. We also demonstrate that selection occurs if members of the Jak-Stat signaling pathway are lost. To show the utility of our system, we have used a known suppressor of IFN signaling, the human papillomavirus (HPV) E7 gene. Expression of E7 causes the loss of ability of 2fTGH cells to respond to IFN-alpha treatment because of a functional disruption of the signaling pathway. This approach offers a new strategy for identifying novel viral genes or new functions of already described viral genes that have a role in IFN-alpha signaling inhibition. 2007-10-17T13:04:08Z Clarke, Daniel T. W. og Irving, Aaron T. og Lambley, Eleanore H. og Payne, Elizabeth og McMillan, Nigel A.J. http://espace.library.uq.edu.au/view/UQ:35096 2007-08-13T10:41:31Z Stomski, F og Dattore, M og Guthridge, M og Woodcock, J og Bagley, C og Le, F og Gonda, T og Lopez, A og Andrews, R og Berndt, M http://espace.library.uq.edu.au/view/UQ:118540 Inflammatory arthropathies such as rheumatoid arthritis, ankylosing spondylitis, and psoriatic arthritis are extremely common in the community, with a prevalence of up to 5%, and they cause substantial morbidity. The development of anti-TNF agents for use initially in rheumatoid arthritis, and subsequently more broadly in inflammatory arthritis, represents the biggest advance in management of these conditions since the introduction of corticosteroid agents, and is a major vindication of public funded arthritis research. However, there are limitations of even these highly effective agents. A significant minority of patients with inflammatory arthritis do not respond to these anti-TNF agents, they are associated with substantial risk of toxicity, require parenteral administration, and are extremely expensive. New antibody treatments in development can be divided into anti-cytokine agents, cell-targeted therapies, co-stimulation inhibitors, and treatments aimed at preventing joint erosion consequent on inflammation. This review discusses the state of the art in the development of these agents for management of this common group of diseases. 2007-10-17T14:10:52Z Brown, MA http://espace.library.uq.edu.au/view/UQ:35205 2007-08-13T10:46:51Z Thomas, R http://espace.library.uq.edu.au/view/UQ:151465 Although the importance of CD4+ T-cell help for generation of an effective CD8+ effector cytotoxic T cell (CTL) response is well established, the role of T-cell help in the activation of memory T cells to become fully functional tumor killer cells is undefined. Using synthetic peptide immunizations corresponding to the major CTLs and T-helper epitopes of ovalbumin, adoptive transfers of ovalbumin-specific memory CTLs (mCTLs), and ovalbumin as the tumor-specific antigen in a mouse tumor model, we have determined that T help is essential for the activation of mCTLs to kill tumors. Our data show that T-helper cells specific for the tumor-associated antigen are required for the reactivation of mCTLs by antigen presented indirectly from tumor. In contrast, effector CTLs do not need T help to kill tumors. These results have implications for induction of tumor immunotherapy by immunization. 2008-06-19T15:20:46Z Gao, Feng Guang og Khammanivong, Vithagna og Liu, Wen Jun og Leggatt, Graham R. og Frazer, Ian H. og Fernando, Germain J. P. http://espace.library.uq.edu.au/view/UQ:65355 Antigen-specific suppression of a previously primed immune response is a major challenge for immunotherapy of autoimmune disease. ReIB activation is required for myeloid DC differentiation. Here, we show that antigen-exposed DCs in which ReIB function is inhibited lack cell surface CD40, prevent priming of immunity, and suppress previously primed immune responses. DCs generated from CD40-deficient mice similarly confer suppression. Regulatory CD4(+) T cells induced by the DCs transfer antigen-specific Infectious tolerance to primed recipients in an interleukin10-dependent fashion. Thus CD40, regulated by ReIB activity, determines the consequences of antigen presentation by myeloid DCs. These observations have significance for autoimmune immunotherapy and suggest a mechanism by which peripheral tolerance might be constitutively maintained by RelB(-) CD40(-) DCs. 2007-08-15T01:25:20Z Martin, E. og O'Sullivan, B. og Low, P. og Thomas, R. http://espace.library.uq.edu.au/view/UQ:39606 Purpose Antigen-specific suppression of a previously primed immune response is a major challenge for immunotherapy of autoimmune disease. We have shown that NF-κB inactivation in dendritic cells (modified DC) converts them into cells that tolerize rather than immunize to specific antigen [1]. Antigen-exposed modified DC prevent priming of immunity, and they suppress previously primed immune responses. Regulatory CD4+ T cells, which can transfer antigen-specific tolerance in an IL-10-dependent fashion, mediate the tolerance. We hypothesized that modified DC exposed to arthritogenic antigen would suppress clinical arthritis after disease onset. Methods Antigen-induced arthritis was induced in C57/Bl6 mice by priming to methylated bovine serum albumin (mBSA) antigen followed by challenge injection of mBSA to one knee. Knee swelling was apparent within 2 days, with peak clinical signs apparent at 5 days. Mice were treated with antigen-exposed modified DC between 2 and 6 days after mBSA challenge to the knee joint. Results Clinical arthritis was suppressed in each group receiving mBSA-exposed modified DC within 4 days compared with mice that received either no DC or keyhole limpet hemocyanin-exposed modified DC. Clinical improvement was associated with mBSA-specific tolerance in mice receiving mBSA-exposed modified DC. Tolerance induction was not impaired by concomitant administration of anti-tumor necrosis factor alpha monoclonal antibody. Subsequent rechallenge with intra-articular IL-1 induced flare of arthritis in all groups, which could be effectively suppressed by a second administration of mBSA-exposed modified DC. Conclusions The data indicate that modified DC induce antigen-specific immune suppression in this model of inflammatory arthritis, even after full clinical expression of the disease. These observations have important implications for antigen-specific therapy of autoimmunity. 2007-08-13T13:48:58Z Martin, E. og O'Sullivan, B. J. og Thomas, R. http://espace.library.uq.edu.au/view/UQ:41105 Purpose Antigen-specific suppression of a previously primed immune response is a major challenge for immunotherapy of autoimmune disease. We have shown that NF-κB inactivation in dendritic cells (modified DC) converts them into cells that tolerize rather than immunize to specific antigen [1]. Antigen-exposed modified DC prevent priming of immunity, and they suppress previously primed immune responses. Regulatory CD4+ T cells, which can transfer antigen-specific tolerance in an IL-10-dependent fashion, mediate the tolerance. We hypothesized that modified DC exposed to arthritogenic antigen would suppress clinical arthritis after disease onset. Methods Antigen-induced arthritis was induced in C57/Bl6 mice by priming to methylated bovine serum albumin (mBSA) antigen followed by challenge injection of mBSA to one knee. Knee swelling was apparent within 2 days, with peak clinical signs apparent at 5 days. Mice were treated with antigen-exposed modified DC between 2 and 6 days after mBSA challenge to the knee joint. Results Clinical arthritis was suppressed in each group receiving mBSA-exposed modified DC within 4 days compared with mice that received either no DC or keyhole limpet hemocyanin-exposed modified DC. Clinical improvement was associated with mBSA-specific tolerance in mice receiving mBSA-exposed modified DC. Tolerance induction was not impaired by concomitant administration of anti-tumor necrosis factor alpha monoclonal antibody. Subsequent rechallenge with intra-articular IL-1 induced flare of arthritis in all groups, which could be effectively suppressed by a second administration of mBSA-exposed modified DC. Conclusions The data indicate that modified DC induce antigen-specific immune suppression in this model of inflammatory arthritis, even after full clinical expression of the disease. These observations have important implications for antigen-specific therapy of autoimmunity. 2007-08-13T14:11:13Z Thomas, R. og Martin, E. og O'Sullivan, B. J. http://espace.library.uq.edu.au/view/UQ:181454 Existing therapies for rheumatoid arthritis and other autoimmune diseases are not Ag specific, which increases the likelihood of systemic toxicity. We show that egg phosphatidylcholine liposomes loaded with Ag (OVA or methylated BSA) and a lipophilic NF-{kappa}B inhibitor (curcumin, quercetin, or Bay11-7082) suppress preexisting immune responses in an Ag-specific manner. We injected loaded liposomes into mice primed with Ag or into mice suffering from Ag-induced inflammatory arthritis. The liposomes targeted APCs in situ, suppressing the cells’ responsiveness to NF-{kappa}B and inducing Ag-specific FoxP3+ regulatory T cells. This regulatory mechanism suppressed effector T cell responses and the clinical signs of full-blown Ag-induced arthritis. Thus, liposomes encapsulate Ags and NF-{kappa}B inhibitors stably and efficiently and could be readily adapted to deliver Ags and inhibitors for Ag-specific suppression of other autoimmune and allergic diseases. 2009-09-03T08:31:17Z Capini, Christelle og Jaturanpinyo, Montree og Chang, Hsin-I og Mutalik, Srinivas og McNally, Alice og Street, Shayna og Steptoe, Raymond og O'Sullivan, Brendan og Davies, Nigel og Thomas, Ranjeny http://espace.library.uq.edu.au/view/UQ:115836 Aim: To determine the frequency of tumour budding and somatic APC mutation in a series of colorectal cancers stratified according to DNA microsatellite instability (MSI) status. Material/Methods: Ninety five colorectal cancers were genotyped for APC mutation in the mutation cluster region (exon 15) and scored for the presence of turnout budding at the invasive margin in haematoxylin and eosin stained sections. A subset was immunostained for beta catenin and p16. Results: The frequency of both somatic APC mutation and tumour budding increased pari passu in cancers stratified as sporadic MSI high (MSI-H), hereditary non-polyposis colorectal cancer (HNPCC), MSI low (MSI-L), and microsatellite stable (MSS). Both budding and APC mutation were significantly less frequent in sporadic MSI-H cancers than in MSI-L or MSS cancers. Tumour buds were characterised by increased immunostaining for both beta catenin and p16. Conclusion: Tumour budding is associated with an adverse prognosis. The lack of budding in MSI-H colorectal cancer may account for the improved prognosis of this subset and may be explained by an. intact WNT signalling pathway and/or inactivated p16(INK4A). 2007-10-17T11:50:15Z Jass, JR og Barker, M og Fraser, L og Walsh, MD og Whitehall, VLJ og Gabrielli, B og Young, J og Leggett, BA http://espace.library.uq.edu.au/view/UQ:56059 2007-08-13T15:44:24Z Zhou, F. og Leggatt, G. og Zhong, J. og Zhao, K. og De Kluyver, R. og Frazer, I. H. http://espace.library.uq.edu.au/view/UQ:133502 We have developed a totally new class of nonporphyrin photodynamic therapeutic agents with a specific focus on two lead candidates azadipyrromethene (ADPM)01 and ADPM06. Confocal laser scanning microscopy imaging showed that these compounds are exclusively localised to the cytosolic compartment, with specific accumulation in the endoplasmic reticulum and to a lesser extent in the mitochondria. Light-induced toxicity assays, carried out over a broad range of human tumour cell lines, displayed EC50 values in the micro-molar range for ADPM01 and nano-molar range for ADPM06, with no discernable activity bias for a specific cell type. Strikingly, the more active agent, ADPM06, even retained significant activity under hypoxic conditions. Both photosensitisers showed low to nondeterminable dark toxicity. Flow cytometric analysis revealed that ADPM01 and ADPM06 were highly effective at inducing apoptosis as a mode of cell death. The photophysical and biological characteristics of these PDT agents suggest that they have potential for the development of new anticancer therapeutics. 2008-03-28T15:28:33Z Gallagher, WM og Allen, LT og O'Shea, C og Kenna, T og Hall, M og Gorman, A og Killoran, J og O'Shea, DF http://espace.library.uq.edu.au/view/UQ:101779 2007-08-23T20:25:02Z Rakhit, D. og Downey, M. O. og Moir, W. S. og Prins, J. B. og Marwick, T. H. http://espace.library.uq.edu.au/view/UQ:72300 The AP-2 transcription factor family is presumed to play an important role in the regulation of the keratinocyte squamous differentiation program; however, limited functional data are available to support this. In the present study, the activity and regulation of AP-2 were examined in differentiating human epidermal keratinocytes. We report that (1) AP-2 transcriptional activity decreases in differentiated keratinocytes but remains unchanged in differentiation-insensitive squamous cell carcinoma cell lines, (2) diminished AP-2 transcriptional activity is associated with a loss of specific DNA-bound AP-2 complexes, and (3) there is an increase in the ability of cytoplasmic extracts, derived from differentiated keratinocytes, to phosphorylate AP-2alpha and AP-2beta when cells differentiate. In contrast, extracts from differentiation-insensitive squamous cell carcinoma cells are unable to phosphorylate AP-2 proteins. Finally, the phosphorylation of recombinant AP-2alpha by cytosolic extracts from differentiated keratinocytes is associated with decreased AP-2 DNA-binding activity. Combined, these data indicate that AP-2 trans-activation and DNA-binding activity decrease as keratinocytes differentiate, and that this decreased activity is associated with an enhanced ability to phosphorylate AP-2alpha and beta. 2007-08-15T04:10:58Z Popa, Claudia og Dahler, Alison L. og Serewko-Auret, Magdalena M. og Wong, Chung F. og Smith, Louise og Barnes, Liam M. og Strutton, Geoff M. og Saunders, Nicholas A.