43 results on '"Piper KP"'
Search Results
2. Effects of elexacaftor/tezacaftor/ivacaftor on liver fibrosis markers in adults with cystic fibrosis.
- Author
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Tewkesbury DH, Scott J, Barry PJ, Bright-Thomas RJ, Hanley KP, Athwal V, and Jones AM
- Subjects
- Humans, Male, Female, Adult, Drug Combinations, Chloride Channel Agonists therapeutic use, Pyrroles therapeutic use, Quinolines therapeutic use, Pyrrolidines therapeutic use, Cystic Fibrosis drug therapy, Cystic Fibrosis complications, Benzodioxoles therapeutic use, Liver Cirrhosis drug therapy, Liver Cirrhosis etiology, Liver Cirrhosis diagnosis, Indoles therapeutic use, Aminophenols therapeutic use, Biomarkers blood, Quinolones therapeutic use, Pyridines therapeutic use, Pyrazoles therapeutic use
- Abstract
Background: There are limited studies to date on the effects of elexacaftor/tezacaftor/ivacaftor (E/T/I) on markers of liver fibrosis in adults with cystic fibrosis (CF). This study aims to analyse changes in makers of liver fibrosis before and after initiation of E/T/I in CF adults., Methods: Outcome measures of liver fibrosis, including liver stiffness measurement (LSM) using FibroScan, AST-to-platelet-ratio index (APRI) and gamma-GT-to-platelet-ratio (GPR) were available in 74 CF adults following initiation of E/T/I. This was compared to historical data collected in 2018 prior to UK availability of E/T/I., Results: The median duration of E/T/I therapy at the time liver fibrosis markers were repeated was 21 (IQR: 17-25) months. There was an increase in APRI from historical measurement to follow-up but no change in LSM or GPR. There were no differences in change in fibrosis markers according to CF liver disease (CFLD) status, although those with a raised LSM at baseline (>6.8 kPa) (n = 14) had a significant reduction in LSM from historical measurement to follow-up versus those with a normal historical value (-3.3 kPa vs 0.25 kPa, p < 0.01)., Conclusions: Apart from APRI, we found no changes in liver fibrosis outcomes after initiation of E/T/I in adults with CF. Those with a historical diagnosis of CFLD had no significant worsening or improvement of liver fibrosis markers. We did observe a reduction in LSM in those with liver nodularity, with an initial highest result suggesting a potential positive treatment effect of E/T/I in this category of those with severe CFLD., Competing Interests: Declaration of Competing Interest DT, JS, RBT, KPH and AJ have no conflicts of interest to disclose. PB has accepted speaking fees and honoraria from Vertex pharmaceuticals and advisory board fees from INSMED. VA has accepted speaking fees and honoraria from Vertex pharmaceuticals and Roche diagnostics., (Copyright © 2023. Published by Elsevier B.V.)
- Published
- 2024
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3. Aetiology of Significant Liver Test Abnormalities in a Single-Centre Cohort of People with Cystic Fibrosis Exposed to Elexacaftor/Tezacaftor/Ivacaftor, Utilizing the Updated RUCAM.
- Author
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Tewkesbury D, Jones AM, Bright-Thomas R, Cratchley A, Hanley KP, Wyatt J, Athwal V, and Barry PJ
- Subjects
- Adult, Humans, Cystic Fibrosis Transmembrane Conductance Regulator genetics, Aminophenols adverse effects, Benzodioxoles adverse effects, Transaminases therapeutic use, Necrosis chemically induced, Mutation, Cystic Fibrosis drug therapy, Liver Diseases, Chemical and Drug Induced Liver Injury etiology
- Abstract
Background: The cystic fibrosis (CF) transmembrane conductance regulator (CFTR) modulator elexacaftor/tezacaftor/ivacaftor (E/T/I) has been associated with substantial multisystem benefits for people with CF eligible for therapy. In a minority, tolerance has been limited by hepatic toxicity. It is unknown whether there may be particular risk factors for significant drug-induced elevation in transaminases., Objective: We aimed to determine the cause of raised transaminases following the introduction of E/T/I, and whether E/T/I can safely be continued in some individuals with elevated transaminases., Methods: At a large, single, adult CF centre, individuals with transaminases >3 × the upper limit of normal (ULN) since commencing E/T/I underwent clinical assessment to exclude known causes of raised transaminases. Where an alternative cause could not be identified, individuals were discussed with hepatology to advise on further investigations to establish aetiology in addition to calculation of the updated Roussel Uclaf Causality Assessment Method (RUCAM) score to assess causality grading of drug-induced liver injury (DILI) due to E/T/I, and to guide management of ongoing CFTR modulator therapy., Results: Of 337 adults taking E/T/I for a median of 27 months, 19 (5.6%) had transaminases >3 × ULN. In 12 individuals, there was clear evidence of an aetiology unrelated to E/T/I (RUCAM scores -2 to 1 [excluded-unlikely]). Of the remaining cases, two had RUCAM scores in the 'possible' range and one had a RUCAM score in the 'probable' range. Liver biopsy was performed in four individuals, showing hepatic steatosis in one individual, normal histology in one individual, and hepatocyte necrosis suggestive of DILI in two individuals. E/T/I was suspended in those with hepatocyte necrosis, with one permanent discontinuation due to synthetic dysfunction. One individual with hepatocyte necrosis on histology was successfully re-established on E/T/I therapy., Conclusions: Alternative causes were identified in the majority of patients with clinically significant increases in transaminases following E/T/I, highlighting the importance of thorough investigation. Multidisciplinary assessment involving an experienced hepatologist is crucial in cases of diagnostic uncertainty or suggestion of significant DILI, as discontinuation of therapy can have significant consequences for individuals., (© 2023. The Author(s), under exclusive licence to Springer Nature Switzerland AG.)
- Published
- 2023
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4. Antagonic effect of ghrelin and LEAP-2 on hepatic stellate cell activation and liver fibrosis in obesity-associated nonalcoholic fatty liver disease.
- Author
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Ezquerro S, Tuero C, Becerril S, Valentí V, Moncada R, Landecho MF, Catalán V, Gómez-Ambrosi J, Mocha F, Silva C, Hanley KP, Escalada J, Frühbeck G, and Rodríguez A
- Subjects
- Humans, Transforming Growth Factor beta1 adverse effects, Transforming Growth Factor beta1 metabolism, Hepatic Stellate Cells metabolism, Hepatic Stellate Cells pathology, Phosphatidylinositol 3-Kinases metabolism, Phosphatidylinositol 3-Kinases pharmacology, Ghrelin adverse effects, Receptors, Ghrelin, Liver Cirrhosis etiology, Liver metabolism, Non-alcoholic Fatty Liver Disease metabolism, Obesity, Morbid
- Abstract
Background: Growing evidence suggests the key role of ghrelin in the onset and progression of nonalcoholic fatty liver disease (NAFLD). The potential participation of ghrelin and the ghrelin receptor antagonist, LEAP-2, in the onset of liver fibrosis in patients with severe obesity and NAFLD through the regulation of TGF-β1-induced hepatic stellate cell (HSC) activation was investigated., Methods: Circulating (n = 179) and hepatic expression (n = 95) of ghrelin and LEAP-2 were measured in patients with severe obesity and available liver pathology analysis undergoing Roux-en-Y gastric bypass (RYGB). The effect of ghrelin isoforms and LEAP-2 on TGF-β1-induced HSC activation, fibrogenic response, and contractile properties was evaluated in vitro in human LX-2 cells., Results: Plasma and hepatic ghrelin were negatively associated, while LEAP-2 exhibited a positive association with liver fibrosis in patients with obesity and NAFLD. Six months after RYGB, hepatic function was improved and, although acylated ghrelin and LEAP-2 concentrations remained unchanged, both hormones were inversely related to post-surgical levels of profibrogenic factors TGF-β1 and TIMP-1. Acylated ghrelin treatment reversed TGF-β1-induced myofibroblast-like phenotype, collagen contractile properties, and the upregulation of factors involved in HSC activation and fibrogenesis via PI3K/Akt/mTOR pathway. Moreover, acylated ghrelin inhibited the mild HSC activation induced by LEAP-2., Conclusions: Ghrelin is an anti-fibrogenic factor blocking HSC activation induced by the most potent fibrogenic cytokine, TGF-β1, and LEAP-2. The imbalance between acylated ghrelin and ghrelin receptor antagonist LEAP-2 might contribute to maintain liver fibrosis in patients with obesity and NAFLD., (© The Author(s) 2023. Published by Oxford University Press on behalf of European Society of Endocrinology.)
- Published
- 2023
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5. The prevalence and burden of Rome IV bowel disorders of gut brain interaction in patients with non-alcoholic fatty liver disease: a cross-sectional study.
- Author
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Purssell H, Bennett L, Street O, Hanley KP, Hanley N, Vasant DH, and Athwal VS
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- Humans, Female, Middle Aged, Male, Cross-Sectional Studies, Quality of Life psychology, Prevalence, Activities of Daily Living, Rome, Surveys and Questionnaires, Brain, Liver Cirrhosis complications, Liver Cirrhosis epidemiology, Irritable Bowel Syndrome complications, Irritable Bowel Syndrome epidemiology, Irritable Bowel Syndrome diagnosis, Non-alcoholic Fatty Liver Disease complications, Non-alcoholic Fatty Liver Disease epidemiology, Brain Diseases complications
- Abstract
Rome IV bowel disorders of gut brain interaction (DGBI) and non-alcoholic fatty liver disease (NAFLD) are highly prevalent entities with overlapping pathophysiology and risk factors. We aimed to evaluate the prevalence and burden of Rome IV irritable bowel syndrome (IBS) in patients with NAFLD. Patients diagnosed with NAFLD were recruited from a specialist liver clinic. All participants completed assessments to determine liver fibrosis severity, including liver stiffness measurement (LSM), completed the Rome IV diagnostic questionnaire for bowel disorders of gut brain interaction, the IBS symptom severity score (IBS-SSS), and the EQ-5D-5L to measure of quality-of-life (QoL). 142 patients with NAFLD (71 (50%) female, mean age 53.5 (SD ± 14.9), BMI 35.2 (SD ± 8.1) kg/M
2 ) were recruited. 79 (55.6%) patients met criteria for a Rome IV bowel DGBI, including 50 patients (35.2%) who met the criteria for IBS (mean IBS-SSS 277.2 (SD ± 131.5)). There was no difference in liver fibrosis scores between those with and without Rome IV IBS (FIB-4 scores p = 0.14, LSM p = 0.68). Patients with NAFLD and Rome IV IBS had significantly worse QoL scores (EQ-VAS p = 0.005 and EQ-5D-5L index p = 0.0007), impairment of usual activities of daily living (p = 0.012) and were more likely to report anxiety or depression (p = 0.038). Rome IV bowel DGBI such as IBS are highly prevalent in patients with NAFLD attending liver clinics and are associated with impaired QoL and psychosocial distress., (© 2023. The Author(s).)- Published
- 2023
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6. Rapid Generation of Pulmonary Organoids from Induced Pluripotent Stem Cells by Co-Culturing Endodermal and Mesodermal Progenitors for Pulmonary Disease Modelling.
- Author
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Mitchell A, Yu C, Zhao X, Pearmain L, Shah R, Hanley KP, Felton T, and Wang T
- Abstract
Differentiation of induced pluripotent stem cells to a range of target cell types is ubiquitous in monolayer culture. To further improve the phenotype of the cells produced, 3D organoid culture is becoming increasingly prevalent. Mature organoids typically require the involvement of cells from multiple germ layers. The aim of this study was to produce pulmonary organoids from defined endodermal and mesodermal progenitors. Endodermal and mesodermal progenitors were differentiated from iPSCs and then combined in 3D Matrigel hydrogels and differentiated for a further 14 days to produce pulmonary organoids. The organoids expressed a range of pulmonary cell markers such as SPA, SPB, SPC, AQP5 and T1α. Furthermore, the organoids expressed ACE2 capable of binding SARS-CoV-2 spike proteins, demonstrating the physiological relevance of the organoids produced. This study presented a rapid production of pulmonary organoids using a multi-germ-layer approach that could be used for studying respiratory-related human conditions., Competing Interests: The authors declare no conflict of interest.
- Published
- 2023
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7. Residual Lung Abnormalities after COVID-19 Hospitalization: Interim Analysis of the UKILD Post-COVID-19 Study.
- Author
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Stewart I, Jacob J, George PM, Molyneaux PL, Porter JC, Allen RJ, Aslani S, Baillie JK, Barratt SL, Beirne P, Bianchi SM, Blaikley JF, Chalmers JD, Chambers RC, Chadhuri N, Coleman C, Collier G, Denneny EK, Docherty A, Elneima O, Evans RA, Fabbri L, Gibbons MA, Gleeson FV, Gooptu B, Greening NJ, Guio BG, Hall IP, Hanley NA, Harris V, Harrison EM, Heightman M, Hillman TE, Horsley A, Houchen-Wolloff L, Jarrold I, Johnson SR, Jones MG, Khan F, Lawson R, Leavy O, Lone N, Marks M, McAuley H, Mehta P, Parekh D, Hanley KP, Platé M, Pearl J, Poinasamy K, Quint JK, Raman B, Richardson M, Rivera-Ortega P, Saunders L, Saunders R, Semple MG, Sereno M, Shikotra A, Simpson AJ, Singapuri A, Smith DJF, Spears M, Spencer LG, Stanel S, Thickett DR, Thompson AAR, Thorpe M, Walsh SLF, Walker S, Weatherley ND, Weeks ME, Wild JM, Wootton DG, Brightling CE, Ho LP, Wain LV, and Jenkins GR
- Subjects
- Humans, SARS-CoV-2, Bayes Theorem, Lung diagnostic imaging, Hospitalization, COVID-19 epidemiology, Lung Diseases, Interstitial
- Abstract
Rationale: Shared symptoms and genetic architecture between coronavirus disease (COVID-19) and lung fibrosis suggest severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection may lead to progressive lung damage. Objectives: The UK Interstitial Lung Disease Consortium (UKILD) post-COVID-19 study interim analysis was planned to estimate the prevalence of residual lung abnormalities in people hospitalized with COVID-19 on the basis of risk strata. Methods: The PHOSP-COVID-19 (Post-Hospitalization COVID-19) study was used to capture routine and research follow-up within 240 days from discharge. Thoracic computed tomography linked by PHOSP-COVID-19 identifiers was scored for the percentage of residual lung abnormalities (ground-glass opacities and reticulations). Risk factors in linked computed tomography were estimated with Bayesian binomial regression, and risk strata were generated. Numbers within strata were used to estimate posthospitalization prevalence using Bayesian binomial distributions. Sensitivity analysis was restricted to participants with protocol-driven research follow-up. Measurements and Main Results: The interim cohort comprised 3,700 people. Of 209 subjects with linked computed tomography (median, 119 d; interquartile range, 83-155), 166 people (79.4%) had more than 10% involvement of residual lung abnormalities. Risk factors included abnormal chest X-ray (risk ratio [RR], 1.21; 95% credible interval [CrI], 1.05-1.40), percent predicted Dl
CO less than 80% (RR, 1.25; 95% CrI, 1.00-1.56), and severe admission requiring ventilation support (RR, 1.27; 95% CrI, 1.07-1.55). In the remaining 3,491 people, moderate to very high risk of residual lung abnormalities was classified at 7.8%, and posthospitalization prevalence was estimated at 8.5% (95% CrI, 7.6-9.5), rising to 11.7% (95% CrI, 10.3-13.1) in the sensitivity analysis. Conclusions: Residual lung abnormalities were estimated in up to 11% of people discharged after COVID-19-related hospitalization. Health services should monitor at-risk individuals to elucidate long-term functional implications.- Published
- 2023
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8. Activating natural product synthesis using CRISPR interference and activation systems in Streptomyces.
- Author
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Ameruoso A, Villegas Kcam MC, Cohen KP, and Chappell J
- Subjects
- Anti-Bacterial Agents, CRISPR-Cas Systems genetics, Multigene Family, Biological Products, Streptomyces genetics
- Abstract
The rise of antibiotic-resistant bacteria represents a major threat to global health, creating an urgent need to discover new antibiotics. Natural products derived from the genus Streptomyces represent a rich and diverse repertoire of chemical molecules from which new antibiotics are likely to be found. However, a major challenge is that the biosynthetic gene clusters (BGCs) responsible for natural product synthesis are often poorly expressed under laboratory culturing conditions, thus preventing the isolation and screening of novel chemicals. To address this, we describe a novel approach to activate silent BGCs through rewiring endogenous regulation using synthetic gene regulators based upon CRISPR-Cas. First, we refine CRISPR interference (CRISPRi) and create CRISPR activation (CRISPRa) systems that allow for highly programmable and effective gene repression and activation in Streptomyces. We then harness these tools to activate a silent BGC by perturbing its endogenous regulatory network. Together, this work advances the synthetic regulatory toolbox for Streptomyces and facilitates the programmable activation of silent BGCs for novel chemical discovery., (© The Author(s) 2022. Published by Oxford University Press on behalf of Nucleic Acids Research.)
- Published
- 2022
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9. DNA and modified vaccinia Ankara prime-boost vaccination generates strong CD8 + T cell responses against minor histocompatibility antigen HA-1.
- Author
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Eldershaw SA, Pearce H, Inman CF, Piper KP, Abbotts B, Stephens C, Nicol S, Croft W, Powell R, Begum J, Taylor G, Nunnick J, Walsh D, Sirovica M, Saddique S, Nagra S, Ferguson P, Moss P, and Malladi R
- Subjects
- Adult, Aged, Allografts, Cytotoxicity, Immunologic, Epitopes immunology, Gene Rearrangement, beta-Chain T-Cell Antigen Receptor, HLA-A2 Antigen immunology, Hematopoietic Stem Cell Transplantation, Humans, Immunogenicity, Vaccine, Immunologic Memory, Male, Middle Aged, Peptides immunology, Receptors, Antigen, T-Cell, alpha-beta immunology, Vaccines, Attenuated, Vaccines, DNA immunology, Viral Vaccines immunology, Antigens, Neoplasm immunology, Graft vs Leukemia Effect immunology, Minor Histocompatibility Antigens immunology, Oligopeptides immunology, T-Lymphocytes, Cytotoxic immunology, Vaccination, Vaccines, DNA therapeutic use, Vaccinia virus immunology, Viral Vaccines therapeutic use
- Abstract
Allogeneic immune responses underlie the graft-versus-leukaemia effect of stem cell transplantation, but disease relapse occurs in many patients. Minor histocompatibility antigen (mHAg) peptides mediate alloreactive T cell responses and induce graft-versus-leukaemia responses when expressed on patient haematopoietic tissue. We vaccinated nine HA-1-negative donors against HA-1 with a 'prime-boost' protocol of either two or three DNA 'priming' vaccinations prior to 'boost' with modified vaccinia Ankara (MVA). HA-1-specific CD8
+ T cell responses were observed in seven donors with magnitude up to 1·5% of total CD8+ T cell repertoire. HA-1-specific responses peaked two weeks post-MVA challenge and were measurable in most donors after 12 months. HA-1-specific T cells demonstrated strong cytotoxic activity and lysed target cells with endogenous HA-1 protein expression. The pattern of T cell receptor (TCR) usage by HA-1-specific T cells revealed strong conservation of T cell receptor beta variable 7-9 (TRBV7-9) usage between donors. These findings describe one of the strongest primary peptide-specific CD8+ T cell responses yet recorded to a DNA-MVA prime-boost regimen and this may reflect the strong immunogenicity of mHAg peptides. Prime-boost vaccination in donors or patients may prove of substantial benefit in boosting graft-versus-leukaemia responses., (© 2021 British Society for Haematology and John Wiley & Sons Ltd.)- Published
- 2021
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10. Dynamic changes in the epigenomic landscape regulate human organogenesis and link to developmental disorders.
- Author
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Gerrard DT, Berry AA, Jennings RE, Birket MJ, Zarrineh P, Garstang MG, Withey SL, Short P, Jiménez-Gancedo S, Firbas PN, Donaldson I, Sharrocks AD, Hanley KP, Hurles ME, Gomez-Skarmeta JL, Bobola N, and Hanley NA
- Subjects
- Animals, Animals, Genetically Modified, Databases, Genetic, Enhancer Elements, Genetic, Gene Expression Regulation, Developmental, Histone Code genetics, Humans, Models, Genetic, Mutation, Organogenesis physiology, Promoter Regions, Genetic, Tissue Distribution, Transcription Factors metabolism, Zebrafish embryology, Zebrafish genetics, Developmental Disabilities genetics, Epigenesis, Genetic, Organogenesis genetics
- Abstract
How the genome activates or silences transcriptional programmes governs organ formation. Little is known in human embryos undermining our ability to benchmark the fidelity of stem cell differentiation or cell programming, or interpret the pathogenicity of noncoding variation. Here, we study histone modifications across thirteen tissues during human organogenesis. We integrate the data with transcription to build an overview of how the human genome differentially regulates alternative organ fates including by repression. Promoters from nearly 20,000 genes partition into discrete states. Key developmental gene sets are actively repressed outside of the appropriate organ without obvious bivalency. Candidate enhancers, functional in zebrafish, allow imputation of tissue-specific and shared patterns of transcription factor binding. Overlaying more than 700 noncoding mutations from patients with developmental disorders allows correlation to unanticipated target genes. Taken together, the data provide a comprehensive genomic framework for investigating normal and abnormal human development.
- Published
- 2020
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11. Human notochordal cell transcriptome unveils potential regulators of cell function in the developing intervertebral disc.
- Author
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Rodrigues-Pinto R, Ward L, Humphreys M, Zeef LAH, Berry A, Hanley KP, Hanley N, Richardson SM, and Hoyland JA
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- Biomarkers, CD24 Antigen genetics, CD24 Antigen metabolism, Computational Biology methods, Gene Expression Profiling, Gene Expression Regulation, Developmental, Humans, Reproducibility of Results, Intervertebral Disc cytology, Intervertebral Disc embryology, Notochord cytology, Stem Cells cytology, Stem Cells metabolism, Transcriptome
- Abstract
The adult nucleus pulposus originates from the embryonic notochord, but loss of notochordal cells with skeletal maturity in humans is thought to contribute to the onset of intervertebral disc degeneration. Thus, defining the phenotype of human embryonic/fetal notochordal cells is essential for understanding their roles and for development of novel therapies. However, a detailed transcriptomic profiling of human notochordal cells has never been achieved. In this study, the notochord-specific marker CD24 was used to specifically label and isolate (using FACS) notochordal cells from human embryonic and fetal spines (7.5-14 weeks post-conception). Microarray analysis and qPCR validation identified CD24, STMN2, RTN1, PRPH, CXCL12, IGF1, MAP1B, ISL1, CLDN1 and THBS2 as notochord-specific markers. Expression of these markers was confirmed in nucleus pulposus cells from aged and degenerate discs. Ingenuity pathway analysis revealed molecules involved in inhibition of vascularisation (WISP2, Noggin and EDN2) and inflammation (IL1-RN) to be master regulators of notochordal genes. Importantly, this study has, for the first time, defined the human notochordal cell transcriptome and suggests inhibition of inflammation and vascularisation may be key roles for notochordal cells during intervertebral disc development. The molecules and pathways identified in this study have potential for use in developing strategies to retard/prevent disc degeneration, or regenerate tissue.
- Published
- 2018
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12. Altered Phenotype of β-Cells and Other Pancreatic Cell Lineages in Patients With Diffuse Congenital Hyperinsulinism in Infancy Caused by Mutations in the ATP-Sensitive K-Channel.
- Author
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Salisbury RJ, Han B, Jennings RE, Berry AA, Stevens A, Mohamed Z, Sugden SA, De Krijger R, Cross SE, Johnson PP, Newbould M, Cosgrove KE, Hanley KP, Banerjee I, Dunne MJ, and Hanley NA
- Subjects
- Case-Control Studies, Cell Lineage, Cell Proliferation, Child, Child, Preschool, Congenital Hyperinsulinism metabolism, Cyclin-Dependent Kinase 6 metabolism, Cyclin-Dependent Kinase Inhibitor p27 metabolism, Fetus cytology, Glucagon-Secreting Cells cytology, Homeobox Protein Nkx-2.2, Homeodomain Proteins metabolism, Humans, Infant, Infant, Newborn, Insulin metabolism, Insulin-Secreting Cells cytology, Mutation, Nuclear Proteins, Paired Box Transcription Factors metabolism, Potassium Channels, Inwardly Rectifying genetics, Somatostatin-Secreting Cells cytology, Sulfonylurea Receptors genetics, Transcription Factors metabolism, Zebrafish Proteins, Congenital Hyperinsulinism genetics, Glucagon-Secreting Cells metabolism, Insulin-Secreting Cells metabolism, Somatostatin-Secreting Cells metabolism
- Abstract
Diffuse congenital hyperinsulinism in infancy (CHI-D) arises from mutations inactivating the KATP channel; however, the phenotype is difficult to explain from electrophysiology alone. Here we studied wider abnormalities in the β-cell and other pancreatic lineages. Islets were disorganized in CHI-D compared with controls. PAX4 and ARX expression was decreased. A tendency toward increased NKX2.2 expression was consistent with its detection in two-thirds of CHI-D δ-cell nuclei, similar to the fetal pancreas, and implied immature δ-cell function. CHI-D δ-cells also comprised 10% of cells displaying nucleomegaly. In CHI-D, increased proliferation was most elevated in duct (5- to 11-fold) and acinar (7- to 47-fold) lineages. Increased β-cell proliferation observed in some cases was offset by an increase in apoptosis; this is in keeping with no difference in INSULIN expression or surface area stained for insulin between CHI-D and control pancreas. However, nuclear localization of CDK6 and P27 was markedly enhanced in CHI-D β-cells compared with cytoplasmic localization in control cells. These combined data support normal β-cell mass in CHI-D, but with G1/S molecules positioned in favor of cell cycle progression. New molecular abnormalities in δ-cells and marked proliferative increases in other pancreatic lineages indicate CHI-D is not solely a β-cell disorder., (© 2015 by the American Diabetes Association. Readers may use this article as long as the work is properly cited, the use is educational and not for profit, and the work is not altered.)
- Published
- 2015
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13. Proteome-wide analyses of human hepatocytes during differentiation and dedifferentiation.
- Author
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Rowe C, Gerrard DT, Jenkins R, Berry A, Durkin K, Sundstrom L, Goldring CE, Park BK, Kitteringham NR, Hanley KP, and Hanley NA
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- Alcohol Dehydrogenase metabolism, Cells, Cultured, Cytochrome P-450 Enzyme System metabolism, Glutathione Transferase metabolism, Hep G2 Cells, Humans, Liver cytology, Liver embryology, Liver metabolism, Liver Neoplasms metabolism, Liver Neoplasms pathology, Cell Differentiation genetics, Hepatocytes metabolism, Hepatocytes pathology, Proteomics methods
- Abstract
Unlabelled: Failure to predict hepatotoxic drugs in preclinical testing makes it imperative to develop better liver models with a stable phenotype in culture. Stem cell-derived models offer promise, with differentiated hepatocyte-like cells currently considered to be "fetal-like" in their maturity. However, this judgment is based on limited biomarkers or transcripts and lacks the required proteomic datasets that directly compare fetal and adult hepatocytes. Here, we quantitatively compare the proteomes of human fetal liver, adult hepatocytes, and the HepG2 cell line. In addition, we investigate the proteome changes in human fetal and adult hepatocytes when cultured in a new air-liquid interface format compared to conventional submerged extracellular matrix sandwich culture. From albumin and urea secretion, and luciferase-based cytochrome P450 activity, adult hepatocytes were viable in either culture model over 2 weeks. The function of fetal cells was better maintained in the air-liquid interface system. Strikingly, the proteome was qualitatively similar across all samples but hierarchical clustering showed that each sample type had a distinct quantitative profile. HepG2 cells more closely resembled fetal than adult hepatocytes. Furthermore, clustering showed that primary adult hepatocytes cultured at the air-liquid interface retained a proteome that more closely mimicked their fresh counterparts than conventional culture, which acquired myofibroblast features. Principal component analysis extended these findings and identified a simple set of proteins, including cytochrome P450 2A6, glutathione S transferase P, and alcohol dehydrogenases as specialized indicators of hepatocyte differentiation., Conclusion: Our quantitative datasets are the first that directly compare multiple human liver cells, define a model for enhanced maintenance of the hepatocyte proteome in culture, and provide a new protein "toolkit" for determining human hepatocyte maturity in cultured cells., (Copyright © 2013 by the American Association for the Study of Liver Diseases.)
- Published
- 2013
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14. Osteopontin is a novel downstream target of SOX9 with diagnostic implications for progression of liver fibrosis in humans.
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Pritchett J, Harvey E, Athwal V, Berry A, Rowe C, Oakley F, Moles A, Mann DA, Bobola N, Sharrocks AD, Thomson BJ, Zaitoun AM, Irving WL, Guha IN, Hanley NA, and Hanley KP
- Subjects
- Animals, Disease Progression, Humans, Male, Osteopontin biosynthesis, Rats, Rats, Sprague-Dawley, SOX9 Transcription Factor biosynthesis, Liver Cirrhosis diagnosis, Liver Cirrhosis etiology, Osteopontin physiology, SOX9 Transcription Factor physiology
- Abstract
Unlabelled: Osteopontin (OPN) is an important component of the extracellular matrix (ECM), which promotes liver fibrosis and has been described as a biomarker for its severity. Previously, we have demonstrated that Sex-determining region Y-box 9 (SOX9) is ectopically expressed during activation of hepatic stellate cells (HSC) when it is responsible for the production of type 1 collagen, which causes scar formation in liver fibrosis. Here, we demonstrate that SOX9 regulates OPN. During normal development and in the mature liver, SOX9 and OPN are coexpressed in the biliary duct. In rodent and human models of fibrosis, both proteins were increased and colocalized to fibrotic regions in vivo and in culture-activated HSCs. SOX9 bound a conserved upstream region of the OPN gene, and abrogation of Sox9 in HSCs significantly decreased OPN production. Hedgehog (Hh) signaling has previously been shown to regulate OPN expression directly by glioblastoma (GLI) 1. Our data indicate that in models of liver fibrosis, Hh signaling more likely acts through SOX9 to modulate OPN. In contrast to Gli2 and Gli3, Gli1 is sparse in HSCs and is not increased upon activation. Furthermore, reduction of GLI2, but not GLI3, decreased the expression of both SOX9 and OPN, whereas overexpressing SOX9 or constitutively active GLI2 could rescue the antagonistic effects of cyclopamine on OPN expression., Conclusion: These data reinforce SOX9, downstream of Hh signaling, as a core factor mediating the expression of ECM components involved in liver fibrosis. Understanding the role and regulation of SOX9 during liver fibrosis will provide insight into its potential modulation as an antifibrotic therapy or as a means of identifying potential ECM targets, similar to OPN, as biomarkers of fibrosis., (Copyright © 2012 American Association for the Study of Liver Diseases.)
- Published
- 2012
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15. Chronic lymphocytic leukaemia cells drive the global CD4+ T cell repertoire towards a regulatory phenotype and leads to the accumulation of CD4+ forkhead box P3+ T cells.
- Author
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Piper KP, Karanth M, McLarnon A, Kalk E, Khan N, Murray J, Pratt G, and Moss PA
- Subjects
- Adult, Aged, Aged, 80 and over, CD4-Positive T-Lymphocytes metabolism, Cells, Cultured, Coculture Techniques, Flow Cytometry, Humans, Immunomodulation, Interleukin-2 Receptor alpha Subunit biosynthesis, Interleukin-7 Receptor alpha Subunit biosynthesis, Leukemia, Lymphocytic, Chronic, B-Cell drug therapy, Leukemia, Lymphocytic, Chronic, B-Cell pathology, Lymphocyte Count, Middle Aged, T-Lymphocyte Subsets metabolism, T-Lymphocytes, Regulatory metabolism, T-Lymphocytes, Regulatory pathology, Tumor Necrosis Factor Receptor Superfamily, Member 7 biosynthesis, Vidarabine analogs & derivatives, Vidarabine therapeutic use, CD4-Positive T-Lymphocytes immunology, Forkhead Transcription Factors biosynthesis, Leukemia, Lymphocytic, Chronic, B-Cell immunology, T-Lymphocyte Subsets immunology, T-Lymphocytes, Regulatory immunology
- Abstract
Advanced chronic lymphocytic leukaemia (CLL) is associated with profound immunodeficiency, including changes in T regulatory cells (T(regs)). We determined the pattern of expression of forkhead box P3 (FoxP3), CD25, CD27 and CD127 and showed that the frequency of CD4+ FoxP3+ T cells was increased in CLL patients (12% versus 8% in controls). This increase was seen only in advanced disease, with selective expansion of FoxP3-expressing cells in the CD4+ CD25(low) population, whereas the number of CD4+ CD25(high) FoxP3+ cells was unchanged. CD4+ CD25(low) cells showed reduced expression of CD127 and increased CD27, and this regulatory phenotype was also seen on all CD4 T cells subsets in CLL patients, irrespective of CD25 or FoxP3 expression. Incubation of CD4+ T cells with primary CLL tumours led to a sixfold increase in the expression of FoxP3 in CD4+ CD25- T cells. Patients undergoing treatment with fludarabine demonstrated a transient increase in the percentage of CD4+ FoxP3+ T cells, but this reduced to normal levels post-treatment. This work demonstrates that patients with CLL exhibit a systemic T cell dysregulation leading to the accumulation of CD4+ FoxP3+ T cells. This appears to be driven by interaction with malignant cells, and increased understanding of the mechanisms that are involved could provide novel avenues for treatment., (© 2011 The Authors. Clinical and Experimental Immunology © 2011 British Society for Immunology.)
- Published
- 2011
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16. Understanding the role of SOX9 in acquired diseases: lessons from development.
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Pritchett J, Athwal V, Roberts N, Hanley NA, and Hanley KP
- Subjects
- Animals, Humans, Disease genetics, Gene Expression Regulation, Developmental, SOX9 Transcription Factor genetics, SOX9 Transcription Factor metabolism
- Abstract
The transcription factor SOX9 is crucial for multiple aspects of development. Mutations in SOX9 cause campomelic dysplasia, a haploinsufficiency disorder concordant with the expression profile of SOX9 during embryogenesis. The mechanistic understanding of development has revealed roles for SOX9 in regulating cartilage extracellular matrix (ECM) production and cell proliferation, among others. More recently, it transpires that SOX9 becomes expressed and induces destructive ECM components in organ fibrosis and related disorders. Although commonly absent from the parent cell type, SOX9 is expressed in a wide range of cancers, where it regulates cell proliferation. These data have potential diagnostic, prognostic and therapeutic relevance, suggesting that disease mechanisms might result from re-expressing this developmental transcription factor in ectopic locations., (Copyright © 2010 Elsevier Ltd. All rights reserved.)
- Published
- 2011
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17. CD8(+) T-cell immunity against cancer-testis antigens develops following allogeneic stem cell transplantation and reveals a potential mechanism for the graft-versus-leukemia effect.
- Author
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McLarnon A, Piper KP, Goodyear OC, Arrazi JM, Mahendra P, Cook M, Clark F, Pratt G, Craddock C, and Moss PA
- Subjects
- Adult, Female, Humans, Immunity, Cellular, Male, Middle Aged, Multiple Myeloma immunology, Multiple Myeloma therapy, Testis immunology, Young Adult, Antigens, Neoplasm immunology, CD8-Positive T-Lymphocytes immunology, Graft vs Leukemia Effect immunology, Hematopoietic Stem Cell Transplantation methods, Leukemia, Myeloid, Acute immunology, Leukemia, Myeloid, Acute therapy
- Abstract
Background: Allogeneic stem cell transplantation is associated with a powerful 'graft-versus-leukemia' effect that is generally considered to result from an alloreactive T-cell immune response. However, disease remission can also be observed after syngeneic transplantation and we investigated whether a T-cell immune response to cancer-testis antigens can be detected in patients in the post-transplant period., Design and Methods: The T-cell immune response against cancer-testis antigens was studied in a cohort of 41 patients who underwent allogeneic stem cell transplantation for the management of acute myeloid leukemia or multiple myeloma. The cytokine secretion assay was combined with magnetic selection to allow detection of an interferon-gamma-secreting T-cell response to a panel of cancer-testis antigen peptides., Results: A cancer-testis antigen-specific CD8(+) T-cell immune response was observed in the peripheral blood of five patients with an average magnitude of 0.045% of the CD8(+) T-cell repertoire. Four of these patients had undergone reduced intensity conditioning transplantation with alemtuzumab for the treatment of acute myeloid leukemia and three remain in long-term remission. T-cell immunity was focused against peptides derived from MAGE proteins and was markedly increased within the bone marrow., Conclusions: Functional cancer-testis antigen-specific CD8(+) T-cell immune responses develop in the early period following reduced intensity allogeneic stem cell transplantation and are preferentially localized to bone marrow. These immune responses are likely to contribute to the cellular basis of the graft-versus-leukemia effect.
- Published
- 2010
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18. Inactivation of Six2 in mouse identifies a novel genetic mechanism controlling development and growth of the cranial base.
- Author
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He G, Tavella S, Hanley KP, Self M, Oliver G, Grifone R, Hanley N, Ward C, and Bobola N
- Subjects
- Animals, Animals, Newborn, Bone and Bones, Cartilage growth & development, Cartilage metabolism, Cell Differentiation, Chondrogenesis, Homeodomain Proteins metabolism, Humans, Mice, Mice, Knockout, Mice, Transgenic, Nerve Tissue Proteins, Osteogenesis physiology, Proteins metabolism, Skull Base growth & development
- Abstract
The cranial base is essential for integrated craniofacial development and growth. It develops as a cartilaginous template that is replaced by bone through the process of endochondral ossification. Here, we describe a novel and specific role for the homeoprotein Six2 in the growth and elongation of the cranial base. Six2-null newborn mice display premature fusion of the bones in the cranial base. Chondrocyte differentiation is abnormal in the Six2-null cranial base, with reduced proliferation and increased terminal differentiation. Gain-of-function experiments indicate that Six2 promotes cartilage development and growth in other body areas and appears therefore to control general regulators of chondrocyte differentiation. Our data indicate that the main factors restricting Six2 function to the cranial base are tissue-specific transcription of the gene and compensatory effects of other Six family members. The comparable expression during human embryogenesis and the high protein conservation from mouse to human implicate SIX2 loss-of-function as a potential congenital cause of anterior cranial base defects in humans., (Copyright 2010 Elsevier Inc. All rights reserved.)
- Published
- 2010
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19. Transcriptional regulation of the Alström syndrome gene ALMS1 by members of the RFX family and Sp1.
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Purvis TL, Hearn T, Spalluto C, Knorz VJ, Hanley KP, Sanchez-Elsner T, Hanley NA, and Wilson DI
- Subjects
- Adult, Carrier Proteins metabolism, Cell Cycle Proteins, Cells, Cultured, Chromosome Mapping, DNA-Binding Proteins metabolism, Gene Expression Regulation, Humans, Mutation, RNA Interference, Regulatory Factor X Transcription Factors, Transcription Factors metabolism, Transcription, Genetic, Alstrom Syndrome genetics, DNA-Binding Proteins physiology, Promoter Regions, Genetic, Proteins genetics, Sp1 Transcription Factor physiology, Transcription Factors physiology, Transcription Initiation Site
- Abstract
Mutations in the human gene ALMS1 cause Alström syndrome, a disorder characterised by neurosensory degeneration, metabolic defects and cardiomyopathy. ALMS1 encodes a centrosomal protein implicated in the assembly and maintenance of primary cilia. Expression of ALMS1 varies between tissues and recent data suggest that its transcription is modulated during adipogenesis and growth arrest. However the ALMS1 promoter has not been defined. This study focused on identifying and characterising the ALMS1 proximal promoter, initially by using 5' RACE to map transcription start sites. Luciferase reporter assay and EMSA data strongly suggest that ALMS1 transcription is regulated by the ubiquitous factor Sp1. In addition, reporter assay, EMSA, chromatin immunoprecipitation and RNA interference data indicate that ALMS1 transcription is regulated by regulatory factor X (RFX) proteins. These transcription factors are cell-type restricted in their expression profile and known to regulate genes of the ciliogenic pathway. We show binding of RFX proteins to an evolutionarily conserved X-box in the ALMS1 proximal promoter and present evidence that these proteins are responsible for ALMS1 transcription during growth arrest induced by low serum conditions. In summary, this work provides the first data on transcription factors regulating general and context-specific transcription of the disease-associated gene ALMS1., (Copyright 2010 Elsevier B.V. All rights reserved.)
- Published
- 2010
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20. Generating hepatic cell lineages from pluripotent stem cells for drug toxicity screening.
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Baxter MA, Rowe C, Alder J, Harrison S, Hanley KP, Park BK, Kitteringham NR, Goldring CE, and Hanley NA
- Subjects
- Animals, Drug Evaluation, Preclinical, Drug Industry, Humans, Cell Culture Techniques methods, Cell Lineage, Liver cytology, Liver drug effects, Pluripotent Stem Cells cytology
- Abstract
Hepatotoxicity is an enormous and increasing problem for the pharmaceutical industry. Early detection of problems during the drug discovery pathway is advantageous to minimize costs and improve patient safety. However, current cellular models are sub-optimal. This review addresses the potential use of pluripotent stem cells in the generation of hepatic cell lineages. It begins by highlighting the scale of the problem faced by the pharmaceutical industry, the precise nature of drug-induced liver injury and where in the drug discovery pathway the need for additional cell models arises. Current research is discussed, mainly for generating hepatocyte-like cells rather than other liver cell-types. In addition, an effort is made to identify where some of the major barriers remain in translating what is currently hypothesis-driven laboratory research into meaningful platform technologies for the pharmaceutical industry., (Copyright 2010 Elsevier B.V. All rights reserved.)
- Published
- 2010
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21. Patients with Wegener's granulomatosis demonstrate a relative deficiency and functional impairment of T-regulatory cells.
- Author
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Morgan MD, Day CJ, Piper KP, Khan N, Harper L, Moss PA, and Savage CO
- Subjects
- Adult, Aged, Aged, 80 and over, Antibodies, Antineutrophil Cytoplasmic blood, Antibodies, Antineutrophil Cytoplasmic immunology, Cell Separation, Female, Flow Cytometry, Forkhead Transcription Factors immunology, Humans, Male, Middle Aged, Young Adult, Granulomatosis with Polyangiitis immunology, T-Lymphocyte Subsets immunology, T-Lymphocytes, Regulatory immunology
- Abstract
An increased proportion of CD4(+) CD25(+) T cells has been reported in Wegener's granulomatosis (WG) and may represent an accumulation of regulatory T cells (Treg). CD25 is also expressed on recently activated effector T cells. We have determined the relative proportion of these subsets in a large patient cohort. The fraction of Treg in peripheral blood mononuclear cells from patients and healthy controls was determined by assessment of Foxp3 expression on CD4(+) CD25(+) T cells. The functional activity of Treg was determined by their ability to suppress proliferation and cytokine production in response to proteinase-3. Although WG patients demonstrated an increased fraction of CD4(+) CD25(+) T cells, the percentage of Foxp3-positive cells was decreased. In addition, the percentage of Treg was inversely related to the rate of disease relapse. CD4(+) CD25(hi) T cells were able to suppress T-cell proliferation to proteinase-3 in healthy controls and anti-neutrophil cytoplasm antibody (ANCA)- negative patients (at time of sampling) but not in ANCA-positive patients. In patients with active disease, an increased proportion of CD4(+) Foxp3(+) cells was associated with a more rapid disease remission. Patients with WG demonstrate abnormalities in the number and function of Treg and this is most pronounced in those with most active disease. This information is of value in understanding the pathogenesis and potential treatment of this disease.
- Published
- 2010
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22. Fetal microchimerism: the cellular and immunological legacy of pregnancy.
- Author
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Lissauer DM, Piper KP, Moss PA, and Kilby MD
- Subjects
- Animals, Antigen Presentation immunology, Antigen Presentation physiology, Autoimmune Diseases pathology, Autoimmune Diseases therapy, Autoimmunity immunology, Cell Differentiation immunology, Cell Lineage immunology, Cells immunology, Female, Hematopoietic Stem Cell Transplantation methods, Humans, Immunologic Memory physiology, Maternal-Fetal Exchange immunology, Pregnancy, T-Lymphocytes immunology, T-Lymphocytes physiology, Transplantation Tolerance immunology, Chimerism embryology, Immunologic Memory immunology, Pregnancy Complications immunology
- Abstract
During pregnancy there is transplacental traffic of fetal cells into the maternal circulation. Remarkably, cells of fetal origin can then persist for decades in the mother and are detectable in the circulation and in a wide range of tissues. Maternal CD8 T cell responses directed against fetal antigens can also be detected following pregnancy. However, the impact that the persistence of allogenic cells of fetal origin and the maternal immune response towards them has on the mother's health remains unclear and is the subject of considerable investigation. The potentially harmful effects of fetal microchimerism include an association with autoimmune disease and recurrent miscarriage. Beneficial effects that have been explored include the contribution of persistent fetal cells to maternal tissue repair. A link between fetal microchimerism and cancer has also been proposed, with some results supporting a protective role and others, conversely, suggesting a role in tumour development. The phenomenon of fetal microchimerism thus provokes many questions and promises to offer further insights not only into the biology of pregnancy but fields such as autoimmunity, transplantation biology and oncology.
- Published
- 2009
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23. Early reconstitution of effector memory CD4+ CMV-specific T cells protects against CMV reactivation following allogeneic SCT.
- Author
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Pourgheysari B, Piper KP, McLarnon A, Arrazi J, Bruton R, Clark F, Cook M, Mahendra P, Craddock C, and Moss PA
- Subjects
- Adult, Aged, Antibodies, Monoclonal immunology, CD8-Positive T-Lymphocytes immunology, Female, Humans, Male, Middle Aged, Transplantation, Homologous, Young Adult, CD4-Positive T-Lymphocytes immunology, Cytomegalovirus immunology, Cytomegalovirus Infections immunology, Hematopoietic Stem Cell Transplantation, Immunologic Memory immunology
- Abstract
Reactivation of CMV is a common complication following allogeneic haematopoietic SCT and is associated with significant morbidity and mortality. The relative importance of the CD4+ and CD8+ components of the CMV-specific immune response in protection from reactivation is unclear. The CMV-specific CD4+ and CD8+ immune response was measured at serial time points in 32 patients following allogeneic HSCT. Intracellular cytokine staining following CMV lysate stimulation and HLA-peptide tetramers were used to determine CMV-specific CD4+ and CD8+ responses, respectively. A deficient CMV-specific CD4+ T-cell immune response within the first 30-50 days post transplant was associated with high risk of viral reactivation. Patients with combined impairment of the CD4+ and CD8+ immune response within the first 100 days were susceptible to late viral reactivation. The frequency of CMV-specific CD4+ T cells correlated with CMV-specific CD8+ T cells, comprising 10% of the whole T-cell repertoire. Early CMV-specific CD4+ T-cell reconstitution was dominated by effector memory cells with normal levels of IL-2 resuming 6 months following transplantation. In summary, both CD4 and CD8 CMV-specific immune reconstitution is required for protection from recurrent activation. Measurement of the magnitude of the CMV-specific CD4+ immune response is useful in managing viral reactivation following HSCT.
- Published
- 2009
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24. Secondary anchor polymorphism in the HA-1 minor histocompatibility antigen critically affects MHC stability and TCR recognition.
- Author
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Nicholls S, Piper KP, Mohammed F, Dafforn TR, Tenzer S, Salim M, Mahendra P, Craddock C, van Endert P, Schild H, Cobbold M, Engelhard VH, Moss PA, and Willcox BE
- Subjects
- ATP Binding Cassette Transporter, Subfamily B, Member 2, ATP-Binding Cassette Transporters metabolism, Arginine metabolism, Cell Separation, Circular Dichroism, Crystallography, X-Ray, Epitopes chemistry, Epitopes immunology, HLA-A2 Antigen chemistry, Humans, Leukemia, Myelogenous, Chronic, BCR-ABL Positive immunology, Proteasome Endopeptidase Complex metabolism, Protein Binding, Protein Stability, Protein Structure, Secondary, Protein Transport, Receptors, Antigen, T-Cell chemistry, Surface Plasmon Resonance, T-Lymphocytes, Cytotoxic immunology, Tissue Donors, Minor Histocompatibility Antigens chemistry, Minor Histocompatibility Antigens genetics, Polymorphism, Genetic, Receptors, Antigen, T-Cell immunology
- Abstract
T cell recognition of minor histocompatibility antigens (mHags) underlies allogeneic immune responses that mediate graft-versus-host disease and the graft-versus-leukemia effect following stem cell transplantation. Many mHags derive from single amino acid polymorphisms in MHC-restricted epitopes, but our understanding of the molecular mechanisms governing mHag immunogenicity and recognition is incomplete. Here we examined antigenic presentation and T-cell recognition of HA-1, a prototypic autosomal mHag derived from single nucleotide dimorphism (HA-1(H) versus HA-1(R)) in the HMHA1 gene. The HA-1(H) peptide is restricted by HLA-A2 and is immunogenic in HA-1(R/R) into HA-1(H) transplants, while HA-1(R) has been suggested to be a "null allele" in terms of T cell reactivity. We found that proteasomal cleavage and TAP transport of the 2 peptides is similar and that both variants can bind to MHC. However, the His>Arg change substantially decreases the stability and affinity of HLA-A2 association, consistent with the reduced immunogenicity of the HA-1(R) variant. To understand these findings, we determined the structure of an HLA-A2-HA-1(H) complex to 1.3A resolution. Whereas His-3 is accommodated comfortably in the D pocket, incorporation of the lengthy Arg-3 is predicted to require local conformational changes. Moreover, a soluble TCR generated from HA-1(H)-specific T-cells bound HA-1(H) peptide with moderate affinity but failed to bind HA-1(R), indicating complete discrimination of HA-1 variants at the level of TCR/MHC interaction. Our results define the molecular mechanisms governing immunogenicity of HA-1, and highlight how single amino acid polymorphisms in mHags can critically affect both MHC association and TCR recognition.
- Published
- 2009
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25. Weighing up beta-cell mass in mice and humans: self-renewal, progenitors or stem cells?
- Author
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Hanley NA, Hanley KP, Miettinen PJ, and Otonkoski T
- Subjects
- Animals, Cell Proliferation, Cell Size, Humans, Mice, Pancreas growth & development, Species Specificity, Insulin-Secreting Cells cytology, Stem Cells cytology
- Abstract
Understanding how beta-cells maintain themselves in the adult pancreas is important for prioritizing strategies aimed at ameliorating or ideally curing different forms of diabetes. There has been much debate over whether beta-cell proliferation, as a means of self-renewal, predominates over the existence and differentiation of a pancreatic stem cell or progenitor cell population. This article describes the two opposing positions based largely on research in laboratory rodents and its extrapolation to humans.
- Published
- 2008
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26. The diabetic phenotype in HNF4A mutation carriers is moderated by the expression of HNF4A isoforms from the P1 promoter during fetal development.
- Author
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Harries LW, Locke JM, Shields B, Hanley NA, Hanley KP, Steele A, Njølstad PR, Ellard S, and Hattersley AT
- Subjects
- Adult, Aged, Female, Gene Expression Profiling, Humans, Kidney physiology, Kidney physiopathology, Male, Middle Aged, Pancreas physiology, Pancreas physiopathology, Polymerase Chain Reaction, Protein Isoforms genetics, White People, Alternative Splicing, Diabetes Mellitus genetics, Diabetes Mellitus, Type 2 genetics, Fetal Development genetics, Hepatocyte Nuclear Factor 4 genetics, Mutation, Promoter Regions, Genetic
- Abstract
Objective: Mutations in the alternatively spliced HNF4A gene cause maturity-onset diabetes of the young (MODY). We characterized the spatial and developmental expression patterns of HNF4A transcripts in human tissues and investigated their role as potential moderators of the MODY phenotype., Research Design and Methods: We measured the expression of HNF4A isoforms in human adult tissues and gestationally staged fetal pancreas by isoform-specific real-time PCR. The correlation between mutation position and age of diagnosis or age-related penetrance was assessed in a cohort of 190 patients with HNF4A mutations., Results: HNF4A was expressed exclusively from the P2 promoter in adult pancreas, but from 9 weeks until at least 26 weeks after conception, up to 23% of expression in fetal pancreas was of P1 origin. HNF4A4-6 transcripts were not detected in any tissue. In whole pancreas, HNF4A9 expression was greater than in islets isolated from the endocrine pancreas (relative level 22 vs. 7%). Patients with mutations in exons 9 and 10 (absent from HNF4A3, HNF4A6, and HNF4A9 isoforms) developed diabetes later than those with mutations in exons 2-8, where all isoforms were affected (40 vs. 24 years; P = 0.029). Exon 9/10 mutations were also associated with a reduced age-related penetrance (53 vs. 10% without diabetes at age 55 years; P < 0.00001)., Conclusions: We conclude that isoforms derived from the HNF4A P1 promoter are expressed in human fetal, but not adult, pancreas, and that their presence during pancreatic development may moderate the diabetic phenotype in individuals with mutations in the HNF4A gene.
- Published
- 2008
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27. Ectopic SOX9 mediates extracellular matrix deposition characteristic of organ fibrosis.
- Author
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Hanley KP, Oakley F, Sugden S, Wilson DI, Mann DA, and Hanley NA
- Subjects
- Base Sequence, Cell Line, Tumor, Collagen chemistry, Gene Silencing, HeLa Cells, Hepatocytes cytology, Hepatocytes metabolism, High Mobility Group Proteins metabolism, Histone Deacetylase Inhibitors, Humans, Models, Biological, Molecular Sequence Data, SOX9 Transcription Factor, Sequence Homology, Nucleic Acid, CCAAT-Binding Factor metabolism, Extracellular Matrix metabolism, Fibrosis pathology, High Mobility Group Proteins biosynthesis, Transcription Factors biosynthesis, Transcription Factors metabolism
- Abstract
Appropriate temporospatial expression of the transcription factor SOX9 is important for normal development of a wide range of organs. Here, we show that when SOX9 is expressed ectopically, target genes become expressed that are associated with disease. Histone deacetylase inhibitors in clinical trials for cancer therapy induced SOX9 expression via enhanced recruitment of nuclear factor Y (NF-Y) to CCAAT elements in the SOX9 proximal promoter. The effect of histone deacetylase inhibitors could be elicited in cells that normally lack SOX9, such as hepatocytes. In human fetal hepatocytes, this aberrant induction of SOX9 protein caused ectopic expression of COL2A1 and COMP1 that encode extracellular matrix (ECM) components normally associated with chondrogenesis. Previously, ectopic expression of this "chondrogenic" profile has been implicated in vascular calcification. More broadly, inappropriate ECM deposition is a hallmark of fibrosis. We demonstrated that induction of SOX9 expression also occurred during activation of fibrogenic cells from the adult liver when the transcription factor was responsible for expression of the major component of fibrotic ECM, type 1 collagen. These combined data identify new aspects in the regulation of SOX9 expression. They support a role for SOX9 beyond normal development as a transcriptional regulator in the pathology of fibrosis.
- Published
- 2008
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28. Posttransplantation imatinib as a strategy to postpone the requirement for immunotherapy in patients undergoing reduced-intensity allografts for chronic myeloid leukemia.
- Author
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Olavarria E, Siddique S, Griffiths MJ, Avery S, Byrne JL, Piper KP, Lennard AL, Pallan L, Arrazi JM, Perz JB, O'Shea D, Goldman JM, Apperley JF, and Craddock CF
- Subjects
- Adult, Aged, Benzamides, Humans, Imatinib Mesylate, Immunotherapy, Leukemia, Myelogenous, Chronic, BCR-ABL Positive drug therapy, Lymphocyte Transfusion, Middle Aged, Transplantation, Homologous, Treatment Outcome, Hematopoietic Stem Cell Transplantation methods, Leukemia, Myelogenous, Chronic, BCR-ABL Positive therapy, Piperazines administration & dosage, Pyrimidines administration & dosage, Secondary Prevention
- Abstract
Disease relapse is a major cause of treatment failure after reduced-intensity allografts and while donor lymphocyte infusions (DLIs) can be effective salvage therapy they are associated with severe graft-versus-host disease (GVHD) when administered early after transplantation. We have therefore examined whether imatinib mesylate can delay relapse and postpone the requirement for DLI in 22 patients with chronic myeloid leukemia (CML) allografted using a reduced-intensity regimen. Imatinib was commenced on day + 35 and continued until 1 year after transplantation. Posttransplantation imatinib was well tolerated and abolished the risk of relapse during this period. Twenty-one patients completed 11 months of imatinib therapy, 15 of whom subsequently relapsed and received DLI. Ten patients to date have achieved molecular remission after DLI. Adjunctive targeted therapy allows the kinetics of disease relapse after a reduced-intensity allograft to be manipulated and represents a novel strategy by which outcome may be improved in patients who undergo transplantation for CML and other leukemias.
- Published
- 2007
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29. CXCL10-CXCR3 interactions play an important role in the pathogenesis of acute graft-versus-host disease in the skin following allogeneic stem-cell transplantation.
- Author
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Piper KP, Horlock C, Curnow SJ, Arrazi J, Nicholls S, Mahendra P, Craddock C, and Moss PA
- Subjects
- Acute Disease, Adult, Animals, Biopsy, Cell Movement, Disease Models, Animal, Epidermis metabolism, Epidermis pathology, Female, Gene Expression Regulation, Graft vs Host Disease pathology, Humans, Inflammation blood, Inflammation pathology, Keratinocytes metabolism, Keratinocytes pathology, Lymphocyte Count, Male, Mice, Middle Aged, Skin Diseases pathology, T-Lymphocytes metabolism, T-Lymphocytes pathology, Transplantation, Homologous, Chemokine CXCL10 blood, Graft vs Host Disease blood, Receptors, CXCR3 biosynthesis, Skin Diseases blood, Stem Cell Transplantation
- Abstract
Acute graft-versus-host disease (aGVHD) remains a serious complication following allogeneic stem-cell transplantation (SCT), and is mediated by infiltration of alloreactive donor T cells into recipient tissue. Chemokines and their receptors play a central role in controlling the recruitment of T cells into discrete tissue sites, and determine the clinical features of GVHD in murine models. In this study, we have analyzed the serum concentration of molecules that control leukocyte migration in serial samples from 34 patients following allogeneic SCT. The chemokine CXCL10 (IP-10) was significantly elevated (> 2-fold) in serum at the time of aGVHD. Because the ligand for CXCL10 is CXCR3, the number of CXCR3(+) T cells was determined in peripheral blood, but was not increased during episodes of GVHD. To investigate the role of chemokines in the recruitment of T cells to the anatomic site of GVHD, skin biopsies were stained for CXCL10 and CXCR3 expression. CXCL10 expression was observed in the basal keratinocytes of the epidermis in patients with GVHD together with positive staining for CXCR3 on cells in dermal infiltrates. These findings indicate that CXCL10 plays a central role in the pathogenesis of skin aGVHD by the recruitment of CXCR3(+) T cells to the sites of inflammation.
- Published
- 2007
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30. Persistence of fetal cells in the mother: friend or foe?
- Author
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Lissauer D, Piper KP, Moss PA, and Kilby MD
- Subjects
- DNA analysis, Female, Humans, Male, Pregnancy, Chimerism, Fetus cytology, Maternal-Fetal Exchange physiology, Mothers
- Abstract
There is increasing evidence that both circulating cells and free fetal DNA persist in the maternal circulation after delivery of the fetus. In some cases, this has been described many years after the end of the pregnancy. This article reviews the evidence for these cells being present, the potential methodologies used to identify such cells and the potential effects on maternal immunomodulation. Data relating to the potential beneficial and potentially harmful effects are discussed.
- Published
- 2007
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31. Functional HY-specific CD8+ T cells are found in a high proportion of women following pregnancy with a male fetus.
- Author
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Piper KP, McLarnon A, Arrazi J, Horlock C, Ainsworth J, Kilby MD, Martin WL, and Moss PA
- Subjects
- Adolescent, Adult, Amino Acid Sequence, CD8-Positive T-Lymphocytes drug effects, Cell Line, Chimerism, Clone Cells, Female, Humans, Interferon-gamma metabolism, Male, Molecular Sequence Data, Peptides immunology, Peptides pharmacology, CD8-Positive T-Lymphocytes immunology, Fetus immunology, H-Y Antigen analysis, Pregnancy immunology
- Abstract
Recent studies have demonstrated that fetal cells can be detected in the maternal circulation during virtually all human pregnancies. These fetal cells can engraft and may be isolated for many decades after pregnancy, leading to a state that may be maintained by the passage of pregnancy-associated progenitor cells. The clinical consequences of fetal cell microchimerism are unclear but may be potentially detrimental or valuable to the mother. One possibility is the generation of an alloreactive immune response by the mother to antigens expressed by the fetus; for example, the HY protein encoded by the Y chromosome. To test this we have screened a cohort of women with a range of parity histories within 8 yr of their last pregnancy for the presence of an HY-specific CD8+ T-cell response. Fluorescent HLA-peptide (HY) tetramers were used to stain short-term T-cell cultures from these women for analysis by flow cytometry. Responses were detected in 37% of women with a history of pregnancies that produced males, and this value rose to 50% in women with two or more pregnancies that produced males. HY-specific CD8+ T cells also could be detected directly in the peripheral blood of women with a history of at least two pregnancies that produced males. These HY-specific CD8+ T cells produced interferon gamma (IFNG) following peptide stimulation, demonstrating their functional capacity. In conclusion, our data indicate that alloreactive CD8+ T cells are generated frequently following normal pregnancy and retain functional capability for years following pregnancy.
- Published
- 2007
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32. Haptoglobin levels are associated with haptoglobin genotype and alpha+ -Thalassemia in a malaria-endemic area.
- Author
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Imrie H, Fowkes FJ, Michon P, Tavul L, Hume JC, Piper KP, Reeder JC, and Day KP
- Subjects
- Adolescent, Age Factors, Animals, Child, Child, Preschool, Cohort Studies, Cross-Sectional Studies, Female, Gene Frequency, Genotype, Hemolysis genetics, Humans, Infant, Linear Models, Malaria diagnosis, Malaria genetics, Male, Papua New Guinea epidemiology, Parasitemia diagnosis, Parasitemia epidemiology, Parasitemia genetics, Polymorphism, Genetic, Splenomegaly epidemiology, Endemic Diseases, Haptoglobins analysis, Haptoglobins genetics, Malaria epidemiology, alpha-Thalassemia genetics
- Abstract
Haptoglobin (Hp) is an acute phase protein that removes free hemoglobin (Hb) released during hemolysis. Hp has also been shown to be toxic for malaria parasites. alpha(+)-Thalassemia is a hemoglobinopathy that results in subclinical hemolytic anemia. alpha(+)-Thassemia homozygosity confers protection against severe malarial disease by an as yet unidentified mechanism. Hp levels were measured in a serial cross-sectional survey of children in Madang Province, Papua New Guinea (PNG). Hp levels were related to age, Hp genotype, Hb levels, parasitemia, splenomegaly, and alpha(+)-thalassemia genotype. Surprisingly, children who were homozygous for alpha(+) -thalassemia had significantly higher levels of Hp than did heterozygotes, after controlling for relevant confounders. We suggest that this is the result of either reduced mean cell Hb associated with alpha(+) -thalassemia homozygosity or an elevated IL-6-dependent acute phase response.
- Published
- 2006
33. Evaluating human embryonic germ cells: concord and conflict as pluripotent stem cells.
- Author
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Turnpenny L, Spalluto CM, Perrett RM, O'Shea M, Hanley KP, Cameron IT, Wilson DI, and Hanley NA
- Subjects
- Animals, Cell Culture Techniques methods, Cell Differentiation, Cell Movement, Cell Proliferation, Cell- and Tissue-Based Therapy methods, Culture Media chemistry, Embryo, Mammalian physiology, Gene Expression Profiling, Gene Expression Regulation, Developmental, Germ Cells chemistry, Growth Substances physiology, Humans, Mice, Embryo, Mammalian cytology, Germ Cells physiology, Pluripotent Stem Cells physiology
- Abstract
The realization of cell replacement therapy derived from human pluripotent stem cells requires full knowledge of the starting cell types as well as their differentiated progeny. Alongside embryonic stem cells, embryonic germ cells (EGCs) are an alternative source of pluripotent stem cell. Since 1998, four groups have described the derivation of human EGCs. This review analyzes the progress on derivation, culture, and differentiation, drawing comparison with other pluripotent stem cell populations.
- Published
- 2006
- Full Text
- View/download PDF
34. Human embryonic germ cells for future neuronal replacement therapy.
- Author
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Turnpenny L, Cameron IT, Spalluto CM, Hanley KP, Wilson DI, and Hanley NA
- Subjects
- Animals, Gonads cytology, Gonads embryology, Humans, Brain Diseases therapy, Germ Cells cytology, Neurons cytology, Pluripotent Stem Cells cytology, Stem Cell Transplantation methods
- Abstract
Stem cell therapy offers exciting potential for ambitious cellular replacement to treat human (h) disease, such as Parkinson's disease, Alzheimer's disease or even replacement of the cell death that follows thromboembolic stroke. The realisation of these treatments requires cellular resources possessing three essential characteristics: (i) self-renewal, (ii) the ability to differentiate to physiologically normal cell types and (iii) lack of tumourigenicity. Here, we describe work on human embryonic germ cells (hEGCs), a population of cells alongside human embryonic stem cells (hESCs) with the potential to address these issues.
- Published
- 2005
- Full Text
- View/download PDF
35. Association between Balint training and physician empathy and work satisfaction.
- Author
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Cataldo KP, Peeden K, Geesey ME, and Dickerson L
- Subjects
- Adult, Family Practice, Female, Humans, Male, Middle Aged, South Carolina, Surveys and Questionnaires, Empathy, Job Satisfaction, Physician-Patient Relations, Physicians psychology
- Abstract
Background and Objectives: Balint training is used in many family medicine residency programs to encourage self-reflection and exploration of the physician-patient encounter. There is limited objective research, however, on outcomes of Balint training. This study's purpose was to determine whether Balint training is associated with differences in physician empathy and work satisfaction., Methods: The Jefferson Scale of Physician Empathy and a validated survey on physician work satisfaction were mailed to 182 graduates of the Medical University of South Carolina (MUSC) Family Medicine Residency Program (113 Balint attendees and 69 nonattendees). The Dillman method of survey design was followed., Results: The two groups were not statistically different in demographic measures. Balint attendees had a similar mean empathy score (119.4) as nonattendees (116.7). There was also no statistically significant difference in overall work satisfaction or satisfaction with financial compensation. Balint attendees were, however, more likely to say they would choose the same specialty if they could choose again than did nonattendees (86.1% versus 55%)., Conclusions: There was no association found between Balint training and physician empathy, financial satisfaction, or overall work satisfaction. However, physicians who completed Balint training at the MUSC Family Medicine Residency Program seemed more satisfied with their choice of family medicine as a specialty.
- Published
- 2005
36. Low antibody responses to variant surface antigens of Plasmodium falciparum are associated with severe malaria and increased susceptibility to malaria attacks in Gabonese children.
- Author
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Tebo AE, Kremsner PG, Piper KP, and Luty AJ
- Subjects
- Animals, Case-Control Studies, Child, Child, Preschool, Cohort Studies, Disease Susceptibility, Erythrocytes parasitology, Gabon, Humans, Immunoglobulin G blood, Infant, Longitudinal Studies, Malaria, Falciparum immunology, Malaria, Falciparum parasitology, Severity of Illness Index, Antibodies, Protozoan blood, Antigens, Protozoan immunology, Antigens, Surface immunology, Malaria, Falciparum physiopathology, Plasmodium falciparum immunology
- Abstract
We measured the levels of IgG antibodies with specificity for the variant surface antigens (VSA) of Plasmodium falciparum in plasma samples from a cohort of Gabonese children participating in a longitudinal case-control malaria study. Children with mild malaria had significantly higher anti-VSA IgG responses than their matched counterparts with severe malaria, most markedly during convalescence and when they were healthy. Over the course of the study, almost twice as many children who presented initially with mild rather than severe malaria developed antibodies recognizing the VSA expressed by each of a panel of three isolates, and those with the highest anti-VSA IgG responses had the lowest malaria attack rates. The results suggest that the clinical outcome of P. falciparum infection in young African children depends on their ability to both develop and maintain a broad profile of anti-VSA IgG antibodies, and that this ability is diminished in children who have experienced a severe malaria attack.
- Published
- 2002
- Full Text
- View/download PDF
37. Malaria transmission and naturally acquired immunity to PfEMP-1.
- Author
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Piper KP, Hayward RE, Cox MJ, and Day KP
- Subjects
- Adolescent, Adult, Aging immunology, Animals, Child, Child, Preschool, Cross-Sectional Studies, Humans, Immunoglobulin G blood, Malaria, Falciparum parasitology, Middle Aged, Parasitemia immunology, Parasitemia parasitology, Plasmodium falciparum growth & development, Antibodies, Protozoan blood, Malaria, Falciparum immunology, Malaria, Falciparum transmission, Plasmodium falciparum immunology, Protozoan Proteins immunology
- Abstract
Why there are so few gametocytes (the transmission stage of malaria) in the blood of humans infected with Plasmodium spp. is intriguing. This may be due either to reproductive restraint by the parasite or to unidentified gametocyte-specific immune-mediated clearance mechanisms. We propose another mechanism, a cross-stage immunity to Plasmodium falciparum erythrocyte membrane protein 1 (PfEMP-1). This molecule is expressed on the surface of the erythrocyte infected with either trophozoite or early gametocyte parasites. Immunoglobulin G antibodies to PfEMP-1, expressed on both life cycle stages, were measured in residents from an area where malaria is endemic, Papua New Guinea. Anti-PfEMP-1 prevalence increased with age, mirroring the decline in both the prevalence and the density of asexual and transmission stages in erythrocytes. These data led us to propose that immunity to PfEMP-1 may influence malaria transmission by regulation of the production of gametocytes. This regulation may be achieved in two ways: (i) by controlling asexual proliferation and density and (ii) by affecting gametocyte maturation.
- Published
- 1999
- Full Text
- View/download PDF
38. Virulence and transmission success of the malarial parasite Plasmodium falciparum.
- Author
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Hayward RE, Tiwari B, Piper KP, Baruch DI, and Day KP
- Subjects
- Animals, Antigens, Protozoan genetics, Cell Adhesion, Erythrocyte Membrane parasitology, Genes, Protozoan, Humans, Malaria, Falciparum parasitology, Plasmodium falciparum genetics, Protozoan Proteins genetics, RNA, Messenger genetics, Reverse Transcriptase Polymerase Chain Reaction, Transcription, Genetic, Virulence, Malaria, Falciparum transmission, Plasmodium falciparum pathogenicity
- Abstract
Virulence of Plasmodium falciparum is associated with the expression of variant surface antigens designated PfEMP1 (P. falciparum erythrocyte membrane protein 1) that are encoded by a family of var genes. Data presented show that the transmission stages of P. falciparum also express PfEMP1 variants. Virulence in this host-parasite system can be considered a variable outcome of optimizing the production of sexual transmission stages from the population of disease-inducing asexual stages. Immunity to PfEMP1 will contribute to the regulation of this trade-off by controlling the parasite population with potential to produce mature transmission stages.
- Published
- 1999
- Full Text
- View/download PDF
39. Plasmodium falciparum: analysis of the antibody specificity to the surface of the trophozoite-infected erythrocyte.
- Author
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Piper KP, Roberts DJ, and Day KP
- Subjects
- Adult, Agglutination Tests, Animals, Antibodies, Monoclonal, Antibodies, Protozoan biosynthesis, Antigens, Surface immunology, Child, Cross-Sectional Studies, Erythrocytes immunology, Flow Cytometry, Humans, Immunoglobulin G biosynthesis, Immunoglobulin G immunology, Malaria, Falciparum immunology, Malaria, Falciparum parasitology, Papua New Guinea, Antibodies, Protozoan immunology, Antibody Specificity, Antigens, Protozoan immunology, Erythrocytes parasitology, Plasmodium falciparum immunology
- Abstract
Current opinion supports the view that immunity to the surface of the trophozoite-infected erythrocyte (IE) is to Plasmodium falciparum erythrocyte membrane protein 1 (PfEMP-1). Here we provide further evidence using the mutant cell line 1776/C10 which no longer expresses PfEMP-1 at the IE surface, due to a subtelomeric deletion in chromosome 9. We have measured antibody reactivity to this mutant in comparison to it's intact isogenic parent line 1776, which does express PfEMP-1, using the sensitive technique of flow cytometry. IgG-specific antibodies (subclass IgG1) in the plasma of hyperimmune adults, reacted to 1776 but never to the 1776/C10 mutant. Antibody subclasses were also measured in individual plasma samples to the surface of trophozoite-IE. Predominantly IgG1 antibodies were detected, with a few individual plasma having additional IgG3 antibodies. Previous studies have used the agglutination assay to measure sero-conversion to PfEMP-1. Here we show that both agglutination and flow cytometric methods are comparable, suggesting that agglutination of trophozoite-IE is mediated by IgG antibodies. Comparison of the isogenic cell lines 1776 and 1776/C10 differing in expression of PfEMP-1 provides further evidence that IgG antibodies, in particular of the cytophilic subclasses, mediate recognition of PfEMP-1.
- Published
- 1999
- Full Text
- View/download PDF
40. Eosinophilic interleukin 5 (IL-5) transgenic mice: eosinophil activity and impaired clearance of Schistosoma mansoni.
- Author
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Dent LA, Munro GH, Piper KP, Sanderson CJ, Finlay DA, Dempster RK, Bignold LP, Harkin DG, and Hagan P
- Subjects
- Animals, Chemotaxis, Leukocyte, Eosinophils ultrastructure, Female, Immunization, In Vitro Techniques, Interleukin-5 genetics, Liver parasitology, Male, Mesocestoides immunology, Mice, Mice, Inbred CBA, Mice, Transgenic, Microscopy, Electron, Proteus mirabilis immunology, Schistosoma mansoni immunology, Schistosomiasis mansoni parasitology, Eosinophilia complications, Eosinophilia immunology, Eosinophils immunology, Interleukin-5 physiology, Schistosomiasis mansoni complications, Schistosomiasis mansoni immunology
- Abstract
Eosinophilia is a feature common to many invasive helminth infections and eosinophils are often considered to be effector cells in immunity to helminths. This study examined the possible influence of constitutive eosinophilia on the clearance of Schistosoma mansoni infections in mice. Eosinophils from interleukin-5 transgenic mice exhibit normal ultrastructure and function with regard to phagocytosis and killing of bacteria and responses to chemotactic stimuli. IL-5 transgenics and non-transgenic littermates were immunized once or four (hyperimmunization) times with irradiated cercariae of S. mansoni. Animals were challenged percutaneously with unirradiated cercariae one month after their last exposure to irradiated parasites. One month after challenge transgenic animals, whether unimmunized, vaccinated or hypervaccinated, carried significantly more liver-stage parasites than non-transgenic animals. These results suggest that although eosinophils from IL-5 transgenic mice are functional for a number of key parameters, large numbers of eosinophils and/or high levels of IL-5 may in some way impair clearance of S. mansoni. A re-evaluation of the roles of eosinophils and IL-5 in infections with this and other parasites may therefore be warranted.
- Published
- 1997
- Full Text
- View/download PDF
41. The role of T cells in vaccine immunity in the murine model of schistosomiasis mansoni.
- Author
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Piper KP and McLaren DJ
- Subjects
- Animals, Female, Immunity, Cellular, Lung immunology, Mice, Mice, Inbred CBA, Skin immunology, Schistosoma mansoni immunology, Schistosomiasis mansoni immunology, T-Lymphocytes immunology, Vaccination
- Abstract
Naive CBA/Ca mice and mice vaccinated with gamma-irradiated cercariae of Schistosoma mansoni were challenged percutaneously with normal cercariae and depleted of L3T4+ T helper cells through the administration of a specific monoclonal antibody. Three regimes were utilized to target known phases of parasite migration. The in vivo depletion of L3T4+ cells resulted in a significant reduction in immunity (up to 65%) in vaccinated/challenged mice, provided the monoclonal antibody was targeted towards skin-resident schistosomula. When antibody was targeted towards lung phase challenge larvae, however, there was a significant reduction in worm recovery, but no correspondingly significant reduction in vaccine immunity. In contrast, the administration of monoclonal to naive mice, via all three treatment regimes, had no effect on the primary schistosome worm burden. Histopathological studies complemented these worm recovery data. Skin tissue biopsied from vaccinated/challenged mice treated with monoclonal to L3T4+ T cells rarely showed the inflammatory foci which normally characterize untreated vaccinated/challenged mice. This was true when antibody was given either before challenge, or just after challenge, and correlated with the recorded depression in vaccine immunity. Lung tissue collected from monoclonal-treated vaccinated/challenged mice (for all three treatment regimes) exhibited no changes in morphology compared to that from untreated vaccinated/challenged mice. This was not altogether surprising since in the NIMR vaccine mouse model, the lungs represent a poor site for challenge attrition and appear normal in morphology with the exception of a few, small inflammatory reactions. When the monoclonal was given to naive/infected mice, there was no change in the morphology of the pulmonary tissue, as compared to corresponding untreated cohorts. Immunohistochemical studies revealed that Thy-1+ cells dominated the subdermal inflammatory foci of vaccinated/challenged mice. Of the T cells identified, the T helper subset was the most common, with T suppressor cells being only weakly represented, and in some cases not at all. The proportion of macrophages (Mac-1+) varied between reactions.
- Published
- 1993
- Full Text
- View/download PDF
42. Schistosoma mansoni: larval damage and role of effector cell(s) in the synergy between vaccine immunity and praziquantel treatment.
- Author
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Piper KP, Mott RF, Hockley DJ, and McLaren DJ
- Subjects
- Animals, Disease Models, Animal, Eosinophils immunology, Female, Guinea Pigs, Larva drug effects, Larva immunology, Larva ultrastructure, Leukocyte Count, Mice, Mice, Inbred CBA, Microscopy, Electron, Praziquantel pharmacology, Schistosoma mansoni drug effects, Schistosoma mansoni immunology, Schistosomiasis mansoni drug therapy, Schistosomiasis mansoni parasitology, Silicon Dioxide pharmacology, Whole-Body Irradiation, Praziquantel therapeutic use, Schistosoma mansoni ultrastructure, Schistosomiasis mansoni immunology, Vaccination
- Abstract
A number of authors have demonstrated that the schistosomicidal compound, Praziquantel (Pzq), depends for its action upon the immune status of the host (Sabah et al. 1985; Brindley & Sher, 1987; Doenhoff et al. 1987). We have attempted to define the synergistic interaction between immuno- and chemotherapy further, using the murine irradiated vaccine model of schistosomiasis mansoni. In vaccinated mice, resistance operates in the skin but not the lungs; drug targeted towards lung-stage worms exacerbates lung-phase immunity, however, as depicted by the increased number and size of inflammatory reactions in the pulmonary tissues. Parasites are often found trapped within such foci. In the present investigation, light and ultrastructural studies have been utilized to examine the nature and extent of damage inflicted upon lung-stage larvae recovered from day 6 Pzq-treated vaccinated mice. Such studies have revealed that damage involves muscle disorganization, internal disruption and occasionally, loss of the tegument; in the latter case, cells are often seen attached to the denuded lung worms. To identify the crucial cellular effector cell(s) involved in the synergy between immuno- and chemotherapy, cell depletion studies have been performed in vivo. It would appear from these experiments that eosinophils or lymphocytes rather than neutrophils or macrophages are important effector cells in this synergy. Histological studies argue in favour of eosinophils being the key effector cells.
- Published
- 1991
- Full Text
- View/download PDF
43. Schistosoma mansoni: histological analysis of the synergistic interaction between vaccine immunity and praziquantel therapy in the lungs of mice.
- Author
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Piper KP, Mott RF, and McLaren DJ
- Subjects
- Analysis of Variance, Animals, Drug Administration Schedule, Drug Synergism, Female, Immunity, Leukocyte Count drug effects, Lung drug effects, Lung immunology, Lung parasitology, Mice, Mice, Inbred CBA, Schistosoma mansoni isolation & purification, Schistosomiasis mansoni drug therapy, Schistosomiasis mansoni immunology, Schistosomiasis mansoni pathology, Skin parasitology, Skin pathology, Lung pathology, Praziquantel therapeutic use, Schistosomiasis mansoni prevention & control, Vaccines immunology
- Abstract
Naive CBA mice and mice vaccinated 4 weeks previously with gamma-irradiated cercariae of S. mansoni were challenged percutaneously with normal cercariae and then treated with 500 mg/kg body weight of Praziquantel (Pzq). The drug was administered intradermally on day 1 or intramuscularly on day 6, thus targeting against skin stage or lung stage challenge larvae respectively. The skin site of challenge and/or the lungs were removed at various time points to provide samples for histological examination. As reported elsewhere (Flisser, Delgado & McLaren 1989) the efficacy of Pzq was significantly enhanced in vaccinated mice and was influenced by the treatment regime. Histological analysis revealed that when Pzq was administered I/D on day 1 to vaccinated mice, the inflammatory response to challenge differed in extent but not nature from that seen in vaccinated but untreated cohorts. This correlates with worm recovery data showing no (this study), or only marginal synergy between drug treatment and immunity using this regimen of drug treatment (Flisser et al. 1989). Following the day 6 protocol of drug delivery, however, lungs from treated vaccinated mice exhibited many large inflammatory reactions containing trapped challenge larvae. In contrast, lungs from untreated vaccinated mice had only few foci which were small and rarely contained trapped larvae. These data again correlate well with worm recovery data showing that there is a highly significant synergy between vaccination and drug treatment administered at this time (Flisser et al. 1989; this study). It would seem, therefore, that Pzq exacerbates lung phase immunity in the NIMR vaccine mouse model where skin phase immunity predominates and pulmonary attrition is normally minimal. The results are discussed in the light of published data concerning the effector mechanisms thought to characterize skin and lung phase vaccine resistance in the murine model.
- Published
- 1990
- Full Text
- View/download PDF
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