123 results on '"Clark GC"'
Search Results
2. Isolation and characterization of a novel gene induced by 2,3,7,8-tetrachlorodibenzo-p-dioxin in rat liver
- Author
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Selmin, O, Lucier, GW, Clark, GC, Tritscher, AM, Vanden Heuvel, JP, Gastel, JA, Walker, NJ, Sutter, TR, and Bell, DA
- Published
- 1997
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3. Biopterin metabolism and nitric oxide recoupling in cancer.
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Clark GC, Lai A, Agarwal A, Liu Z, and Wang XY
- Abstract
Tetrahydrobiopterin is a cofactor necessary for the activity of several enzymes, the most studied of which is nitric oxide synthase. The role of this cofactor-enzyme relationship in vascular biology is well established. Recently, tetrahydrobiopterin metabolism has received increasing attention in the field of cancer immunology and immunotherapy due to its involvement in the cytotoxic T cell response. Past research has demonstrated that when the availability of BH4 is low, as it is in chronic inflammatory conditions and tumors, electron transfer in the active site of nitric oxide synthase becomes uncoupled from the oxidation of arginine. This results in the production of radical species that are capable of a direct attack on tetrahydrobiopterin, further depleting its local availability. This feedforward loop may act like a molecular switch, reinforcing low tetrahydrobiopterin levels leading to altered NO signaling, restrained immune effector activity, and perpetual vascular inflammation within the tumor microenvironment. In this review, we discuss the evidence for this underappreciated mechanism in different aspects of tumor progression and therapeutic responses. Furthermore, we discuss the preclinical evidence supporting a clinical role for tetrahydrobiopterin supplementation to enhance immunotherapy and radiotherapy for solid tumors and the potential safety concerns., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Clark, Lai, Agarwal, Liu and Wang.)
- Published
- 2024
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4. RREACT: A mobile multidisciplinary response to overdose.
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Ulintz AJ, McCloskey RJ, Hammond GC, Parrish M, Toliver I, Sharafutdinova A, and Lyons MS
- Abstract
Opioid overdose is a leading cause of death in the United States, and engaging with patients following overdose to provide harm reduction and recovery resources can prove difficult. Quick response models use mobile, multidisciplinary teams to establish a time-sensitive connection between individuals who overdosed and harm reduction and recovery resources that improve outcomes. These quick response models are consistent with the broader field of mobile-integrated health programs that are growing in number and acceptability, though the literature base is sparse and programs vary. We describe the 5-year reach, effectiveness, adoption, implementation and maintenance (RE-AIM) framework of the Rapid Response Emergency Addiction and Crisis Team (RREACT), a fire/emergency medical services-led, multidisciplinary (firefighter/paramedic, law enforcement officer, social worker) mobile outreach team. RREACT provides harm reduction, linkage/transportation to care and wrap-around services to individuals following a nonfatal opioid overdose that resulted in an emergency response in Columbus, Franklin County, Ohio, United States. Between 2018 and 2022, RREACT made 22,157 outreach attempts to 11,739 unique patients. RREACT recorded 3,194 direct patient contacts during this time, resulting in 1,200 linkages to care: 799 direct transports to opioid use disorder treatment and 401 warm handoffs to community treatment agencies. Furthermore, RREACT's staffing increased from 4 full-time equivalent staff in 2018 to 15.5 in 2022 and was supported by the surrounding community through 287 community outreach events and the development of an alumni program. These preliminary results further support the deployment of multidisciplinary mobile outreach teams to increase access to harm reduction and recovery resources following opioid overdose., Competing Interests: CONFLICT OF INTEREST DISCLOSURE Mighty Crow Media, LLC, employs an author of this article and serves as the grant evaluator for RREACT’s Bureau of Justice Assistance COSSAP grant. Grant funding does not support time or other costs associated with this publication. Alexander Ulintz was supported by AHRQ T32 grant (T32 HS029590).
- Published
- 2024
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5. From "crisis to recovery": A complete insight into the mechanisms of chlorine injury in the lung.
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Clark GC, Elfsmark L, Armstrong S, Essex-Lopresti A, Gustafsson Å, Ryan Y, Moore K, Paszkiewicz K, Green AC, Hiscox JA, David J, and Jonasson S
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- Mice, Humans, Animals, Proteomics, Lung, Bronchoalveolar Lavage Fluid, Chlorine toxicity, Proteome
- Abstract
Chlorine (Cl
2 ) gas is a toxic industrial chemical (TIC) that poses a hazard to human health following accidental and/or intentional (e.g. terrorist) release. By using a murine model of sub-lethal Cl2 exposure we have examined the airway hyper responsiveness, cellular infiltrates, transcriptomic and proteomic responses of the lung. In the "crisis" phase at 2 h and 6 h there is a significant decreases in leukocytes within bronchoalveolar lavage fluid accompanied by an upregulation within the proteome of immune pathways ultimately resulting in neutrophil influx at 24 h. A flip towards "repair" in the transcriptome and proteome occurs at 24 h, neutrophil influx and an associated drop in the lung function persisting until 14 d post-exposure and subsequent "recovery" after 28 days. Collectively, this research provides new insights into the mechanisms of damage, early global responses and processes of repair induced in the lung following the inhalation of Cl2 ., (Crown Copyright © 2022. Published by Elsevier Inc. All rights reserved.)- Published
- 2023
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6. A prototype lateral flow assay for detection of orthopoxviruses.
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Ulaeto DO, Lonsdale SG, Laidlaw SM, Clark GC, Horby P, and Carroll MW
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- Biological Assay, DNA, Viral, Humans, Polymerase Chain Reaction, Orthopoxvirus
- Abstract
Competing Interests: MWC received funding from the US FDA to provide materials for this research. PH received the tecoviramat drug from Siga Technologies for an expanded access protocol for monkeypox treatment in Central African Republic. All other authors declare no competing interests.
- Published
- 2022
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7. Radiation induces ESCRT pathway dependent CD44v3 + extracellular vesicle production stimulating pro-tumor fibroblast activity in breast cancer.
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Clark GC, Hampton JD, Koblinski JE, Quinn B, Mahmoodi S, Metcalf O, Guo C, Peterson E, Fisher PB, Farrell NP, Wang XY, and Mikkelsen RB
- Abstract
Despite recent advances in radiotherapeutic strategies, acquired resistance remains a major obstacle, leading to tumor recurrence for many patients. Once thought to be a strictly cancer cell intrinsic property, it is becoming increasingly clear that treatment-resistance is driven in part by complex interactions between cancer cells and non-transformed cells of the tumor microenvironment. Herein, we report that radiotherapy induces the production of extracellular vesicles by breast cancer cells capable of stimulating tumor-supporting fibroblast activity, facilitating tumor survival and promoting cancer stem-like cell expansion. This pro-tumor activity was associated with fibroblast production of the paracrine signaling factor IL-6 and was dependent on the expression of the heparan sulfate proteoglycan CD44v3 on the vesicle surface. Enzymatic removal or pharmaceutical inhibition of its heparan sulfate side chains disrupted this tumor-fibroblast crosstalk. Additionally, we show that the radiation-induced production of CD44v3
+ vesicles is effectively silenced by blocking the ESCRT pathway using a soluble pharmacological inhibitor of MDA-9/Syntenin/SDCBP PDZ1 domain activity, PDZ1i. This population of vesicles was also detected in the sera of human patients undergoing radiotherapy, therefore representing a potential biomarker for radiation therapy and providing an opportunity for clinical intervention to improve treatment outcomes., Competing Interests: PBF is a scientific co-founder and has equity in InterLeukin Combinatorial Therapies, Inc. (ILCT). VCU also has equity in ILCT. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest, (Copyright © 2022 Clark, Hampton, Koblinski, Quinn, Mahmoodi, Metcalf, Guo, Peterson, Fisher, Farrell, Wang and Mikkelsen.)- Published
- 2022
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8. RIPpore: A Novel Host-Derived Method for the Identification of Ricin Intoxication through Oxford Nanopore Direct RNA Sequencing.
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Ryan Y, Harrison A, Trivett H, Hartley C, David J, Clark GC, and Hiscox JA
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- Adenine metabolism, RNA metabolism, Ribosomes metabolism, Sequence Analysis, RNA, Nanopores, Ricin metabolism, Ricin toxicity
- Abstract
Ricin is a toxin which enters cells and depurinates an adenine base in the sarcin-ricin loop in the large ribosomal subunit, leading to the inhibition of protein translation and cell death. We postulated that this depurination event could be detected using Oxford Nanopore Technologies (ONT) direct RNA sequencing, detecting a change in charge in the ricin loop. In this study, A549 cells were exposed to ricin for 2-24 h in order to induce depurination. In addition, a novel software tool was developed termed RIPpore that could quantify the adenine modification of ribosomal RNA induced by ricin upon respiratory epithelial cells. We provided demonstrable evidence for the first time that this base change detected is specific to RIP activity using a neutralising antibody against ricin. We believe this represents the first detection of depurination in RNA achieved using ONT sequencers. Collectively, this work highlights the potential for ONT and direct RNA sequencing to detect and quantify depurination events caused by ribosome-inactivating proteins such as ricin. RIPpore could have utility in the evaluation of new treatments and/or in the diagnosis of exposure to ricin.
- Published
- 2022
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9. Does mental health screening and assessment in child welfare improve mental health service receipt, child safety, and permanence for children in out-of-home care? An evaluation of the Gateway CALL demonstration.
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Bunger AC, Maguire-Jack K, Yoon S, Mooney D, West KY, Hammond GC, and Kranich C
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- Child, Child Protective Services, Child Welfare psychology, Humans, Mental Health, United States, Home Care Services, Mental Health Services
- Abstract
Background: Unmet mental health service needs among children in out-of-home care are sometimes attributed to poor assessments and referrals in child welfare. The Gateway CALL project implemented mental health screening, diagnostic assessment, and referral to treatment practices., Objective: We examined the effectiveness of Gateway CALL for improving children's mental health service receipt, safety, and permanency outcomes., Participants and Setting: Participants included 538 children (birth to 18 years) in out-of-home placements through a county-based child welfare agency over a 17-month period., Methods: We compared the mental health service receipt, safety, and permanency outcomes for 175 children who received Gateway CALL with 175 children who received "services as usual" identified through propensity score matching. Participant demographics, safety, and permanency outcomes were drawn from child welfare administrative records, and mental health service visits and diagnoses were drawn from Medicaid billing records., Results: Gateway CALL appeared to increase the number of mental health service visits children received (z = 2.14, p = 0.032), although not the likelihood of receiving services. In terms of child safety, children in Gateway CALL had a greater number of screened-in calls after the intervention than those in the comparison group [t(348) = -1.92, p = 0.03]; there were no differences in substantiations. There were also no observed effects on permanency., Conclusions: Despite systematic efforts to identify, assess, and refer children to mental health services through the Gateway CALL intervention, substantial unmet mental health service needs among children persisted. Results have implications for designing interventions that promote cross-system service access., (Copyright © 2021 The Author(s). Published by Elsevier Ltd.. All rights reserved.)
- Published
- 2021
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10. Fundamental Vibrational Frequencies and Spectroscopic Constants of Substituted Cyclopropenylidene (c-C 3 HX, X = F, Cl, CN).
- Author
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Westbrook BR, Patel DJ, Dallas JD, Swartzfager GC, Lee TJ, and Fortenberry RC
- Abstract
The recent detection of ethynyl-functionalized cyclopropenylidene (c-C
3 HC2 H) has initiated the search for other functional forms of cyclopropenylidene (c-C3 H2 ) in space. There is existing gas-phase rotational spectroscopic data for cyano-cyclopropenylidene (c-C3 HCN), but the present work provides the first anharmonic vibrational spectral data for that molecule, as well as the first full set of both rotational and vibrational spectroscopic data for fluoro- and chloro-cyclopropenylidenes (c-C3 HF and c-C3 HCl). All three molecules have fundamental vibrational frequencies with substantial infrared intensities. Namely, c-C3 HCN has a moderately intense fundamental frequency at 1244.4 cm-1 , while c-C3 HF has two large intensity modes at 1765.4 and 1125.3 cm-1 and c-C3 HCl again has two large intensity modes at 1692.0 and 1062.5 cm-1 . All of these frequencies are well within the spectral range covered by the high-resolution EXES instrument on NASA's Stratospheric Observatory for Infrared Astronomy (SOFIA). Further, all three molecules have dipole moments of around 3.0 D in line with c-C3 H2 , enabling them to be observed by pure rotational spectroscopy, as well. Thus, the rovibrational spectral data presented herein should assist with future laboratory studies of functionalized cyclopropenylidenes and may lead to their interstellar or circumstellar detection.- Published
- 2021
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11. In Vitro Priming of Human T Cells by Dendritic Cells Provides a Screening Tool for Candidate Vaccines for Burkholderia pseudomallei .
- Author
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Reddi D, Durant L, Bernardo D, Noble A, English NR, Hendy P, Clark GC, Prior JL, Williamson ED, and Knight SC
- Abstract
Murine dendritic cells, when pulsed with heat-killed Burkholderia pseudomallei and used to immunise naïve mice, have previously been shown to induce protective immunity in vivo . We have now demonstrated the in vitro priming of naïve human T cells against heat-killed B. pseudomallei , by co-culture with syngeneic B. pseudomallei- pulsed dendritic cells. Additionally, we have enriched the DC fraction such that a study of the differential response induced by pulsed DCs of either myeloid or plasmacytoid lineage in syngeneic human T cells was achievable. Whilst both mDCs and pDCs were activated by pulsing, the mDCs contributed the major response to B. pseudomallei with the expression of the migration marker CCR7 and a significantly greater secretion of the proinflammatory TNFα and IL1β. When these DC factions were combined and used to prime syngeneic T cells, a significant proliferation was observed in the CD4
+ fraction. Here, we have achieved human T cell priming in vitro with unadjuvanted B. pseudomallei , the causative organism of melioidosis, for which there is currently no approved vaccine. We propose that the approach we have taken could be used to screen for the human cellular response to candidate vaccines and formulations, in order to enhance the cell-mediated immunity required to protect against this intracellular pathogen and potentially more broadly against other, difficult-to-treat intracellular pathogens. To date, the polysaccharide capsule of B. pseudomallei , fused to a standard carrier protein, e.g., Crm, looks a likely vaccine candidate. Dendritic cells (DCs), providing, as they do, the first line of defence to infection, process and present microbial products to the immune system to direct downstream immune responses. Here, we have sought to use DCs ex vivo to identify immunogenic products from heat-killed B. pseudomallei . Using practical volumes of fresh human donor blood, we show that heat-killed B. pseudomallei activated and stimulated the expression of pro-inflammatory cytokines TNF-α, IL-1β and IL-6 from both myeloid and plasmacytoid DCs. Furthermore, B. pseudomallei- pulsed DCs cultured with naïve syngeneic T cells ex vivo , induced the activation and proliferation of the CD4+ T-cell population, which was identified by cell surface marker staining using flow cytometry. Thus, both DC subsets are important for driving primary T helper cell responses to B. pseudomallei in healthy individuals and have the potential to be used to identify immunogenic components of B. pseudomallei for future therapies and vaccines.- Published
- 2021
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12. Establishment of a Novel Oral Murine Model of Ricin Intoxication and Efficacy Assessment of Ovine Ricin Antitoxins.
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Whitfield SJ, Padgen DB, Knight S, Gwyther RJ, Holley JL, Clark GC, and Green AC
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- Administration, Oral, Animals, Antitoxins isolation & purification, Ricinus communis toxicity, Chemical Warfare Agents isolation & purification, Chemical Warfare Agents toxicity, Dose-Response Relationship, Drug, Female, Gastrointestinal Tract pathology, Mice, Mice, Inbred BALB C, Ricin isolation & purification, Sheep, Sheep, Domestic, Treatment Outcome, Antitoxins administration & dosage, Disease Models, Animal, Gastrointestinal Tract drug effects, Ricin administration & dosage, Ricin toxicity
- Abstract
Ricin, produced from the castor beans of Ricinus communis , is a cytotoxin that exerts its action by inactivating ribosomes and causing cell death. Accidental (e.g., ingestion of castor beans) and/or intentional (e.g., suicide) exposure to ricin through the oral route is an area of concern from a public health perspective and no current licensed medical interventions exist to protect from the action of the toxin. Therefore, we examined the oral toxicity of ricin in Balb/C mice and developed a robust food deprivation model of ricin oral intoxication that has enabled the assessment of potential antitoxin treatments. A lethal oral dose was identified and mice were found to succumb to the toxin within 48 h of exposure. We then examined whether a despeciated ovine F(ab')
2 antibody fragment, that had previously been demonstrated to protect mice from exposure to aerosolised ricin, could also protect against oral intoxication. Mice were challenged orally with an LD99 of ricin, and 89 and 44% of mice exposed to this otherwise lethal exposure survived after receiving either the parent anti-ricin IgG or F(ab')2 , respectively. Combined with our previous work, these results further highlight the benefit of ovine-derived polyclonal antibody antitoxin in providing post-exposure protection against ricin intoxication.- Published
- 2020
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13. Erratum: Blume, C., et al. Modulation of Human Airway Barrier Functions during Burkholderia thailandensis and Francisella tularensis Infection Running Title: Airway Barrier Functions during Bacterial Infections. Pathogens 2016, 5 , 53.
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Blume C, David J, Bell RE, Laver JR, Read RC, Clark GC, Davies DE, and Swindle EJ
- Abstract
There is an error in the title [...].
- Published
- 2020
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14. Single and mixture per- and polyfluoroalkyl substances accumulate in developing Northern leopard frog brains and produce complex neurotransmission alterations.
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Foguth RM, Hoskins TD, Clark GC, Nelson M, Flynn RW, de Perre C, Hoverman JT, Lee LS, Sepúlveda MS, and Cannon JR
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- Animals, Environmental Exposure analysis, Larva metabolism, Rana pipiens metabolism, Synaptic Transmission physiology, Water Pollutants, Chemical metabolism, Alkanesulfonic Acids pharmacology, Brain drug effects, Caprylates pharmacology, Fluorocarbons pharmacology, Synaptic Transmission drug effects
- Abstract
Per- and polyfluoroalkyl substances (PFAS) are present in water and >99% of human serum. They are found in brains of wildlife; however, little is known about effects on the developing brain. To determine the effects of PFAS on brain and cardiac innervation, we conducted an outdoor mesocosm experiment with Northern leopard frog larvae (Rana pipiens) exposed to control, 10 ppb perfluorooctane sulfonate (PFOS), or a PFAS mixture totaling 10 ppb that mimicked aqueous film forming foam-impacted surface water (4 ppb PFOS, 3 ppb perfluorohexane sulfonate, 1.25 ppb perfluorooctanoate, 1.25 ppb perfluorohexanoate, and 0.5 ppb perfluoro-n-pentanoate). Water was spiked with PFAS and 25 larvae (Gosner stage (GS) 25) added to each mesocosm (n = 4 mesocosms per treatment). After 30 days, we harvested eight brains per mesocosm and remaining larvae developed to GS 46 (i.e. metamorphosis) before brains and hearts were collected. Weight, length, GS, and time to metamorphosis were recorded. Brain concentrations of all five PFAS were quantified using LC/MS/MS. Dopamine and metabolites, serotonin and its metabolite, norepinephrine, γ-aminobutyric acid, and glutamate were quantified using High Performance Liquid Chromatography with electrochemical detection while acetylcholine and acetylcholinesterase activity were quantified with the Invitrogen Amplex Red Acetylcholine Assay. PFOS accumulated in the brain time- and dose-dependently. After 30 days, the mixture decreased serotonin while both PFAS treatments decreased glutamate. Interestingly, acetylcholine increased in PFAS treatments at GS 46. This research shows that developmental environmentally relevant exposure to PFAS changes neurotransmitters, especially acetylcholine., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 Elsevier Inc. All rights reserved.)
- Published
- 2020
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15. Common Host Responses in Murine Aerosol Models of Infection Caused by Highly Virulent Gram-Negative Bacteria from the Genera Burkholderia, Francisella and Yersinia.
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Clark GC, Essex-Lopresti A, Moore KA, Williamson ED, Lukaszewski R, Paszkiewicz K, and David J
- Abstract
Highly virulent bacterial pathogens cause acute infections which are exceptionally difficult to treat with conventional antibiotic therapies alone. Understanding the chain of events that are triggered during an infection of a host has the potential to lead to new therapeutic strategies. For the first time, the transcriptomic responses within the lungs of Balb/C mice have been compared during an acute infection with the intracellular pathogens Burkholderia pseudomallei , Francisella tularensis and Yersinia pestis . Temporal changes were determined using RNAseq and a bioinformatics pipeline; expression of protein was also studied from the same sample. Collectively it was found that early transcriptomic responses within the infected host were associated with the (a) slowing down of critical cellular functions, (b) production of circulatory system components, (c) lung tissue integrity, and (d) intracellular regulatory processes. One common molecule was identified, Errfi1 (ErbB receptor feedback inhibitor 1); upregulated in response to all three pathogens and a potential novel marker of acute infection. Based upon the pro-inflammatory responses observed, we sought to synchronise each infection and report that 24 h p.i. of B. pseudomallei infection closely aligned with 48 h p.i. of infection with F. tularensis and Y. pestis . Post-transcriptional modulation of RANTES expression occurred across all pathogens, suggesting that these infections directly or indirectly modulate cell trafficking through chemokine expression/detection. Collectively, this unbiased NGS approach has provided an in-depth characterisation of the host transcriptome following infection with these highly virulent pathogens ultimately aiding in the development of host-directed therapies as adjuncts or alternatives to antibiotic treatment.
- Published
- 2019
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16. CD200R deletion promotes a neutrophil niche for Francisella tularensis and increases infectious burden and mortality.
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Casulli J, Fife ME, Houston SA, Rossi S, Dow J, Williamson ED, Clark GC, Hussell T, D'Elia RV, and Travis MA
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- Animals, Cells, Cultured, Disease Models, Animal, Female, Francisella tularensis immunology, Humans, Immunoglobulin Fc Fragments, Lung immunology, Lung microbiology, Lung pathology, Macrophages immunology, Macrophages microbiology, Membrane Glycoproteins genetics, Membrane Glycoproteins metabolism, Mice, Mice, Inbred C57BL, Mice, Knockout, Neutrophils metabolism, Neutrophils microbiology, Primary Cell Culture, Reactive Oxygen Species immunology, Reactive Oxygen Species metabolism, Tularemia microbiology, Francisella tularensis pathogenicity, Host-Pathogen Interactions immunology, Membrane Glycoproteins immunology, Neutrophils immunology, Tularemia immunology
- Abstract
Pulmonary immune control is crucial for protection against pathogens. Here we identify a pathway that promotes host responses during pulmonary bacterial infection; the expression of CD200 receptor (CD200R), which is known to dampen pulmonary immune responses, promotes effective clearance of the lethal intracellular bacterium Francisella tularensis. We show that depletion of CD200R in mice increases in vitro and in vivo infectious burden. In vivo, CD200R deficiency leads to enhanced bacterial burden in neutrophils, suggesting CD200R normally limits the neutrophil niche for infection. Indeed, depletion of this neutrophil niche in CD200R
-/- mice restores F. tularensis infection to levels seen in wild-type mice. Mechanistically, CD200R-deficient neutrophils display significantly reduced reactive oxygen species production (ROS), suggesting that CD200R-mediated ROS production in neutrophils is necessary for limiting F. tularensis colonisation and proliferation. Overall, our data show that CD200R promotes the antimicrobial properties of neutrophils and may represent a novel antibacterial therapeutic target.- Published
- 2019
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17. Friends or Foes? Emerging Impacts of Biological Toxins.
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Clark GC, Casewell NR, Elliott CT, Harvey AL, Jamieson AG, Strong PN, and Turner AD
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- Animals, Humans, Toxins, Biological chemistry, Toxins, Biological adverse effects, Toxins, Biological therapeutic use
- Abstract
Toxins are substances produced from biological sources (e.g., animal, plants, microorganisms) that have deleterious effects on a living organism. Despite the obvious health concerns of being exposed to toxins, they are having substantial positive impacts in a number of industrial sectors. Several toxin-derived products are approved for clinical, veterinary, or agrochemical uses. This review sets out the case for toxins as 'friends' that are providing the basis of novel medicines, insecticides, and even nucleic acid sequencing technologies. We also discuss emerging toxins ('foes') that are becoming increasingly prevalent in a range of contexts through climate change and the globalisation of food supply chains and that ultimately pose a risk to health., (Crown Copyright © 2019. Published by Elsevier Ltd. All rights reserved.)
- Published
- 2019
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18. Mitogen-activated protein kinases (MAPKs) are modulated during in vitro and in vivo infection with the intracellular bacterium Burkholderia pseudomallei.
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D'Elia RV, Saint RJ, Newstead SL, Clark GC, and Atkins HS
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- Animals, Benzamides pharmacology, Burkholderia pseudomallei immunology, Burkholderia pseudomallei metabolism, Cell Line, Diphenylamine analogs & derivatives, Diphenylamine pharmacology, Enzyme Activation, Extracellular Signal-Regulated MAP Kinases antagonists & inhibitors, Female, Macrophages microbiology, Melioidosis immunology, Melioidosis microbiology, Mice, Mice, Inbred BALB C, Burkholderia pseudomallei growth & development, Chemokine CCL2 biosynthesis, Extracellular Signal-Regulated MAP Kinases metabolism, Melioidosis pathology, Tumor Necrosis Factor-alpha biosynthesis, p38 Mitogen-Activated Protein Kinases metabolism
- Abstract
Burkholderia pseudomallei is a Gram-negative intracellular bacterium that causes the disease melioidosis. The disease can be fatal if left untreated or when antibiotic therapy is delayed and total clearance of the pathogen from the host is often not accomplished with current therapies. Thus, new therapeutic approaches for the treatment of infections caused by B. pseudomallei are required. To better understand host responses to B. pseudomallei infection, the activation of key proteins involved in the TLR inflammatory cascade was measured by western blotting. Activation of the mitogen-activated protein kinases (MAPKs) p38 and ERK were both significantly altered during both in vitro and in vivo infection. In considering an approach for therapy of B. pseudomallei infection the inhibition of ERK was achieved in vitro using the inhibitor PD0325901, along with decreased TNF-α production. However, the reduction in phosphorylated ERK and TNF-α release did not correspond with decreased bacterial replication or enhance clearance from infected macrophages. Despite this apparent lack of effect on the intracellular growth of B. pseudomallei in vitro, it is not clear what effect inhibition of ERK activation might have on outcome of disease in vivo. It may be that decreasing the levels of TNF-α in vivo could aid in reducing the overactive immune response that is known to ensue following B. pseudomallei infection, thereby increasing host survival.
- Published
- 2017
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19. Production, Characterisation and Testing of an Ovine Antitoxin against Ricin; Efficacy, Potency and Mechanisms of Action.
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Whitfield SJC, Griffiths GD, Jenner DC, Gwyther RJ, Stahl FM, Cork LJ, Holley JL, Green AC, and Clark GC
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- Animals, Antibodies, Neutralizing analysis, Chlorocebus aethiops, Female, Mice, Inbred BALB C, Ricin pharmacokinetics, Ricin toxicity, Sheep, Vero Cells, Antitoxins immunology, Ricin immunology
- Abstract
Ricin is a type II ribosome-inactivating toxin that catalytically inactivates ribosomes ultimately leading to cell death. The toxicity of ricin along with the prevalence of castor beans (its natural source) has led to its increased notoriety and incidences of nefarious use. Despite these concerns, there are no licensed therapies available for treating ricin intoxication. Here, we describe the development of a F(ab')₂ polyclonal ovine antitoxin against ricin and demonstrate the efficacy of a single, post-exposure, administration in an in vivo murine model of intoxication against aerosolised ricin. We found that a single dose of antitoxin afforded a wide window of opportunity for effective treatment with 100% protection observed in mice challenged with aerosolised ricin when given 24 h after exposure to the toxin and 75% protection when given at 30 h. Treated mice had reduced weight loss and clinical signs of intoxication compared to the untreated control group. Finally, using imaging flow cytometry, it was found that both cellular uptake and intracellular trafficking of ricin toxin to the Golgi apparatus was reduced in the presence of the antitoxin suggesting both actions can contribute to the therapeutic mechanism of a polyclonal antitoxin. Collectively, the research highlights the significant potential of the ovine F(ab')₂ antitoxin as a treatment for ricin intoxication., Competing Interests: The authors declare no conflict of interest.
- Published
- 2017
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20. Morphology and the gradient of a symmetric potential predict gait transitions of dogs.
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Wilshin S, Haynes GC, Porteous J, Koditschek D, Revzen S, and Spence AJ
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- Animals, Biomechanical Phenomena, Dogs, Forelimb physiology, Hindlimb physiology, Gait physiology
- Abstract
Gaits and gait transitions play a central role in the movement of animals. Symmetry is thought to govern the structure of the nervous system, and constrain the limb motions of quadrupeds. We quantify the symmetry of dog gaits with respect to combinations of bilateral, fore-aft, and spatio-temporal symmetry groups. We tested the ability of symmetries to model motion capture data of dogs walking, trotting and transitioning between those gaits. Fully symmetric models performed comparably to asymmetric with only a [Formula: see text] increase in the residual sum of squares and only one-quarter of the parameters. This required adding a spatio-temporal shift representing a lag between fore and hind limbs. Without this shift, the symmetric model residual sum of squares was [Formula: see text] larger. This shift is related to (linear regression, [Formula: see text], [Formula: see text]) dog morphology. That this symmetry is respected throughout the gaits and transitions indicates that it generalizes outside a single gait. We propose that relative phasing of limb motions can be described by an interaction potential with a symmetric structure. This approach can be extended to the study of interaction of neurodynamic and kinematic variables, providing a system-level model that couples neuronal central pattern generator networks and mechanical models.
- Published
- 2017
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21. Longitudinal quasi-static stability predicts changes in dog gait on rough terrain.
- Author
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Wilshin S, Reeve MA, Haynes GC, Revzen S, Koditschek DE, and Spence AJ
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- Animals, Biomechanical Phenomena, Extremities, Female, Male, Models, Theoretical, Walking physiology, Dogs physiology, Gait
- Abstract
Legged animals utilize gait selection to move effectively and must recover from environmental perturbations. We show that on rough terrain, domestic dogs, Canis lupus familiaris , spend more time in longitudinal quasi-statically stable patterns of movement. Here, longitudinal refers to the rostro-caudal axis. We used an existing model in the literature to quantify the longitudinal quasi-static stability of gaits neighbouring the walk, and found that trot-like gaits are more stable. We thus hypothesized that when perturbed, the rate of return to a stable gait would depend on the direction of perturbation, such that perturbations towards less quasi-statically stable patterns of movement would be more rapid than those towards more stable patterns of movement. The net result of this would be greater time spent in longitudinally quasi-statically stable patterns of movement. Limb movement patterns in which diagonal limbs were more synchronized (those more like a trot) have higher longitudinal quasi-static stability. We therefore predicted that as dogs explored possible limb configurations on rough terrain at walking speeds, the walk would shift towards trot. We gathered experimental data quantifying dog gait when perturbed by rough terrain and confirmed this prediction using GPS and inertial sensors ( n =6, P <0.05). By formulating gaits as trajectories on the n -torus we are able to make tractable the analysis of gait similarity. These methods can be applied in a comparative study of gait control which will inform the ultimate role of the constraints and costs impacting locomotion, and have applications in diagnostic procedures for gait abnormalities, and in the development of agile legged robots., Competing Interests: Competing interestsThe authors declare no competing or financial interests., (© 2017. Published by The Company of Biologists Ltd.)
- Published
- 2017
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22. Current and future developments in the treatment of virus-induced hypercytokinemia.
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Wong JP, Viswanathan S, Wang M, Sun LQ, Clark GC, and D'Elia RV
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- Animals, Antiviral Agents chemistry, Birds immunology, Birds virology, Cytokines blood, Cytokines immunology, Dengue blood, Dengue complications, Dengue pathology, Hemorrhagic Fever, Ebola blood, Hemorrhagic Fever, Ebola complications, Hemorrhagic Fever, Ebola pathology, Humans, Inflammation complications, Inflammation immunology, Inflammation pathology, Influenza in Birds blood, Influenza in Birds complications, Influenza in Birds pathology, Antiviral Agents pharmacology, Antiviral Agents therapeutic use, Cytokines antagonists & inhibitors, Dengue immunology, Hemorrhagic Fever, Ebola immunology, Inflammation prevention & control, Influenza in Birds immunology
- Abstract
Emerging pathogenic viruses such as Ebola and Middle Eastern Respiratory Syndrome coronavirus (MERS-CoV) can cause acute infections through the evasion of the host's antiviral immune responses and by inducing the upregulation of inflammatory cytokines. This immune dysregulation, termed a cytokine storm or hypercytokinemia, is potentially fatal and is a significant underlying factor in increased mortality of infected patients. The prevalence of global outbreaks in recent years has offered opportunities to study the progression of various viral infections and have provided an improved understanding of hypercytokinemia associated with these diseases. However, despite this increased knowledge and the study of the infections caused by a range of emerging viruses, the therapeutic options still remain limited. This review aims to explore alternative experimental strategies for treating hypercytokinemia induced by the Ebola, avian influenza and Dengue viruses; outlining their modes of action, summarizing their preclinical assessments and potential clinical applications.
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- 2017
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23. Mitogen-activated protein kinases (MAPKs) are modulated during Francisella tularensis infection, but inhibition of extracellular-signal-regulated kinases (ERKs) is of limited therapeutic benefit.
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Saint RJ, D'Elia RV, Bryant C, Clark GC, and Atkins HS
- Subjects
- Animals, Bacterial Load, Benzamides administration & dosage, Cell Line, Cytokines metabolism, Diphenylamine administration & dosage, Diphenylamine analogs & derivatives, Disease Models, Animal, Female, Gene Expression Profiling, Liver microbiology, Liver pathology, Macrophages, Alveolar enzymology, Macrophages, Alveolar parasitology, Mice, Inbred BALB C, Protein Kinase Inhibitors administration & dosage, Spleen microbiology, Spleen pathology, Treatment Outcome, Extracellular Signal-Regulated MAP Kinases biosynthesis, Host-Pathogen Interactions, Tularemia pathology
- Abstract
Francisella tularensis is a Gram-negative intracellular bacterium that causes the disease tularemia. The disease can be fatal if left untreated and there is currently no licenced vaccine available; the identification of new therapeutic targets is therefore required. Toll-like receptors represent an interesting target for therapeutic modulation due to their essential role in generating immune responses. In this study, we analysed the in vitro expression of the key mitogen-activated protein kinases (MAPKs) p38, JNK and ERK in murine alveolar macrophages during infection with F. tularensis. The phosphorylation profile of ERK highlighted its potential as a target for therapeutic modulation and subsequently the effect of ERK manipulation was measured in a lethal intranasal F. tularensis in vivo model of infection. The selective ERK1/2 inhibitor PD0325901 was administered orally to mice either pre- or post-challenge with F. tularensis strain LVS. Both treatment regimens selectively reduced ERK expression, but only the pre-exposure treatment produced decreased bacterial burden in the spleen and liver, which correlated with a significant reduction in the pro-inflammatory cytokines IFN-γ, MCP-1, IL-6, and TNF-α. However, no overall improvements in survival were observed for treated animals in this study. ERK may represent a useful therapeutic target where selective dampening of the immune response (to control the damaging pathology seen during infection) is combined with antibiotic treatment required to eradicate bacterial infection. This combination treatment strategy has been shown to be effective in other models of tularemia., Competing Interests: Compliance with ethical standards Funding This work was funded by the UK Ministry of Defence. Conflict of Interest To the authors knowledge there is no conflict of interest arising from the publication of this manuscript. Ethical approval All procedures performed in studies involving animals were in accordance with the ethical standards of the institution or practice at which the studies were conducted. Informed consent Not applicable.
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- 2016
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24. Modulation of Human Airway Barrier Functions during Burkholderia thailandensis and Francisella tularensis Infection Running Title: Airway Barrier Functions during Bacterial Infections.
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Blume C, David J, Bell RE, Laver JR, Read RC, Clark GC, Davies DE, and Swindle EJ
- Abstract
The bronchial epithelium provides protection against pathogens from the inhaled environment through the formation of a highly-regulated barrier. In order to understand the pulmonary diseases melioidosis and tularemia caused by Burkholderia thailandensis and Fransicella tularensis, respectively, the barrier function of the human bronchial epithelium were analysed. Polarised 16HBE14o- or differentiated primary human bronchial epithelial cells (BECs) were exposed to increasing multiplicities of infection (MOI) of B. thailandensis or F. tularensis Live Vaccine Strain and barrier responses monitored over 24-72 h. Challenge of polarized BECs with either bacterial species caused an MOI- and time-dependent increase in ionic permeability, disruption of tight junctions, and bacterial passage from the apical to the basolateral compartment. B. thailandensis was found to be more invasive than F. tularensis. Both bacterial species induced an MOI-dependent increase in TNF-α release. An increase in ionic permeability and TNF-α release was induced by B. thailandensis in differentiated BECs. Pretreatment of polarised BECs with the corticosteroid fluticasone propionate reduced bacterial-dependent increases in ionic permeability, bacterial passage, and TNF-α release. TNF blocking antibody Enbrel(®) reduced bacterial passage only. BEC barrier properties are disrupted during respiratory bacterial infections and targeting with corticosteroids or anti-TNF compounds may represent a therapeutic option., Competing Interests: The authors declare no conflict of interest.
- Published
- 2016
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25. miR-9 Acts as an OncomiR in Prostate Cancer through Multiple Pathways That Drive Tumour Progression and Metastasis.
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Seashols-Williams SJ, Budd W, Clark GC, Wu Q, Daniel R, Dragoescu E, and Zehner ZE
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- Animals, Antigens, CD, Cadherins genetics, Cadherins metabolism, Cell Line, Tumor, Cell Movement genetics, Cell Proliferation, Disease Models, Animal, Disease Progression, Gene Expression Profiling, High-Throughput Nucleotide Sequencing, Humans, Male, Mice, Models, Biological, Neoplasm Metastasis, Prostatic Neoplasms metabolism, RNA, Messenger genetics, RNA, Messenger metabolism, Suppressor of Cytokine Signaling Proteins genetics, Suppressor of Cytokine Signaling Proteins metabolism, Gene Expression Regulation, Neoplastic, MicroRNAs genetics, Oncogenes, Prostatic Neoplasms genetics, Prostatic Neoplasms pathology
- Abstract
Identification of dysregulated microRNAs (miRNAs) in prostate cancer is critical not only for diagnosis, but also differentiation between the aggressive and indolent forms of the disease. miR-9 was identified as an oncomiR through both miRNA panel RT-qPCR as well as high-throughput sequencing analysis of the human P69 prostate cell line as compared to its highly tumorigenic and metastatic subline M12, and found to be consistently upregulated in other prostate cell lines including DU-145 and PC3. While miR-9 has been characterized as dysregulated either as an oncomiR or tumour suppressor in a variety of other cancers including breast, ovarian, and nasopharyngeal carcinomas, it has not been previously evaluated and proven as an oncomiR in prostate cancer. miR-9 was confirmed an oncomiR when found to be overexpressed in tumour tissue as compared to adjacent benign glandular epithelium through laser-capture microdissection of radical prostatectomy biopsies. Inhibition of miR-9 resulted in reduced migratory and invasive potential of the M12 cell line, and reduced tumour growth and metastases in male athymic nude mice. Analysis showed that miR-9 targets e-cadherin and suppressor of cytokine signalling 5 (SOCS5), but not NF-ĸB mRNA. Expression of these proteins was shown to be affected by modulation in expression of miR-9.
- Published
- 2016
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26. Mechanisms of Disease: Host-Pathogen Interactions between Burkholderia Species and Lung Epithelial Cells.
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David J, Bell RE, and Clark GC
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- Cell Line, Humans, Models, Biological, Burkholderia mallei physiology, Burkholderia pseudomallei physiology, Epithelial Cells microbiology, Epithelial Cells physiology, Host-Pathogen Interactions, Lung microbiology, Lung pathology
- Abstract
Members of the Burkholderia species can cause a range of severe, often fatal, respiratory diseases. A variety of in vitro models of infection have been developed in an attempt to elucidate the mechanism by which Burkholderia spp. gain entry to and interact with the body. The majority of studies have tended to focus on the interaction of bacteria with phagocytic cells with a paucity of information available with regard to the lung epithelium. However, the lung epithelium is becoming more widely recognized as an important player in innate immunity and the early response to infections. Here we review the complex relationship between Burkholderia species and epithelial cells with an emphasis on the most pathogenic species, Burkholderia pseudomallei and Burkholderia mallei. The current gaps in knowledge in our understanding are highlighted along with the epithelial host-pathogen interactions that offer potential opportunities for therapeutic intervention.
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- 2015
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27. Dual Action of miR-125b As a Tumor Suppressor and OncomiR-22 Promotes Prostate Cancer Tumorigenesis.
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Budd WT, Seashols-Williams SJ, Clark GC, Weaver D, Calvert V, Petricoin E, Dragoescu EA, O'Hanlon K, and Zehner ZE
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- Animals, Biomarkers, Tumor metabolism, Cell Line, Tumor, Extracellular Signal-Regulated MAP Kinases metabolism, Humans, Male, Mice, Nude, Mitogen-Activated Protein Kinase Kinases metabolism, Oncogene Protein v-akt metabolism, Phosphatidylinositol 3-Kinases metabolism, Prostate cytology, Prostatic Neoplasms pathology, Proteomics, Carcinogenesis metabolism, Genes, Tumor Suppressor, MicroRNAs metabolism, Prostatic Neoplasms metabolism
- Abstract
MicroRNAs (miRs) are a novel class of small RNA molecules, the dysregulation of which can contribute to cancer. A combinatorial approach was used to identify miRs that promote prostate cancer progression in a unique set of prostate cancer cell lines, which originate from the parental p69 cell line and extend to a highly tumorigenic/metastatic M12 subline. Together, these cell lines are thought to mimic prostate cancer progression in vivo. Previous network analysis and miR arrays suggested that the loss of hsa-miR-125b together with the overexpression of hsa-miR-22 could contribute to prostate tumorigenesis. The dysregulation of these two miRs was confirmed in human prostate tumor samples as compared to adjacent benign glandular epithelium collected through laser capture microdissection from radical prostatectomies. In fact, alterations in hsa-miR-125b expression appeared to be an early event in tumorigenesis. Reverse phase microarray proteomic analysis revealed ErbB2/3 and downstream members of the PI3K/AKT and MAPK/ERK pathways as well as PTEN to be protein targets differentially expressed in the M12 tumor cell compared to its parental p69 cell. Relevant luciferase+3'-UTR expression studies confirmed a direct interaction between hsa-miR-125b and ErbB2 and between hsa-miR-22 and PTEN. Restoration of hsa-miR-125b or inhibition of hsa-miR-22 expression via an antagomiR resulted in an alteration of M12 tumor cell behavior in vitro. Thus, the dual action of hsa-miR-125b as a tumor suppressor and hsa-miR-22 as an oncomiR contributed to prostate tumorigenesis by modulations in PI3K/AKT and MAPK/ERK signaling pathways, key pathways known to influence prostate cancer progression.
- Published
- 2015
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28. Immune profiling of the progression of a BALB/c mouse aerosol infection by Burkholderia pseudomallei and the therapeutic implications of targeting HMGB1.
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Laws TR, Clark GC, and D'Elia RV
- Subjects
- Aerosols, Animals, Cytokines, HMGB1 Protein genetics, Interferon-gamma therapeutic use, Liver immunology, Melioidosis metabolism, Mice, Mice, Inbred BALB C, Signal Transduction, Spleen microbiology, Burkholderia pseudomallei, HMGB1 Protein metabolism, Melioidosis immunology
- Abstract
Introduction: The role of damage-associated molecular pattern HMGB1 signalling in a murine BALB/c model of severe respiratory melioidosis (Burkholderia pseudomallei infection) was explored in this study., Methods: Time course experiments were performed., Results: It was established that HMGB1 was released in concert with increasing weight of organs and increasing concentration of liver enzymes in the blood a short time after cytokine release. Differences in the cytokine response between organs were observed, where the lungs contained higher concentrations of chemokines and interleukin 17, while the spleen produced more interferon-gamma, which is essential in the host defence against B. pseudomallei. This is evidence as to why the disease is seemingly more severe in the respiratory form. The effect of depleting HMGB1 using an antibody was also evaluated. It was found that this treatment significantly reduced bacterial load in the liver, spleen, and, to a greater degree, in the lungs. Cytokine quantification indicated that this reduction in bacterial load is likely due to the treatment reducing the release of a variety of pro-inflammatory cytokines., Conclusion: It is concluded that anti-HMGB1 treatment would be effective alongside other therapeutics, where it would reduce the characteristic over-inflammation associated with late stage infection., (Crown Copyright © 2015. Published by Elsevier Ltd. All rights reserved.)
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- 2015
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29. Delayed presence of alternatively activated macrophages during a Francisella tularensis infection.
- Author
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D'Elia RV, Laws TR, Núñez A, Taylor C, and Clark GC
- Subjects
- Animals, Disease Models, Animal, Francisella tularensis physiology, Humans, Male, Mice, Mice, Inbred BALB C, Nitric Oxide Synthase Type II genetics, Nitric Oxide Synthase Type II immunology, Tularemia enzymology, Tularemia genetics, Tularemia microbiology, Francisella tularensis immunology, Macrophages immunology, Tularemia immunology
- Abstract
Francisella tularensis is an intracellular bacterium that has the ability to multiply within the macrophage. The phenotype of a macrophage can determine whether the infection is cleared or the host succumbs to disease. Previously published data has suggested that F. tularensis LVS actively induces the alternative phenotype as a survival mechanism. In these studies we demonstrate that this is not the case for the more virulent strain of F. tularensis SCHU-S4. During an intranasal mouse model of infection, immuno-histochemistry identified that iNOS positive ("classical") macrophages are present at 72 h post-infection and remain high (supported by CCL-5 release) in numbers. In contrast, arginase/FIZZ-1 positive ("alternative") cells appear later and in low numbers during the development of the disease tularemia., (Copyright © 2014. Published by Elsevier Ltd.)
- Published
- 2015
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30. "FoxP3 Hunting" during infection with Francisella tularensis.
- Author
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D'Elia RV, Laws TR, Núñez A, and Clark GC
- Subjects
- Animals, Forkhead Transcription Factors analysis, Liver immunology, Lung immunology, Male, Mice, Mice, Inbred BALB C, Spleen immunology, Forkhead Transcription Factors physiology, T-Lymphocytes, Regulatory immunology, Tularemia immunology
- Abstract
Francisella tularensis is a Gram-negative intracellular bacterium that can cause acute disease in mouse models of infection when administered via the inhalational route. The immune response to a pulmonary infection is typified by an initial lack of pro-inflammatory cytokines, followed by hypercytokinemia prior to host death. It remains unclear what causes this delay in the host immune response. In this study we determine the presence of FoxP3 regulatory T cells in the lung, liver and spleen following intranasal infection with F. tularensis SCHU S4. In the lung, the site of initial infection, there is an increase in FoxP3+ cells during the first few days of infection and a notable absence of these cells at the point of cytokine storm and death (day 4 post-infection). This coincides with a decrease in the anti-inflammatory cytokine TGF-β and increases of chemokines MIP-1α, MIP-1β and RANTES. In our model, we also observed an overall decrease in the number of regulatory T cells in the spleen, which was not as evident in the liver. Overall, this data suggests that early on in an acute F. tularensis SCHUS4 infection regulatory T cells contribute to a dampening of the pro-inflammatory response, allowing for bacterial replication and spread.
- Published
- 2014
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31. Targeting the "Rising DAMP" during a Francisella tularensis Infection.
- Author
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D'Elia RV, Laws TR, Carter A, Lukaszewski R, and Clark GC
- Abstract
Antibiotic efficacy is greatly enhanced the earlier it is administered following infection with a bacterial pathogen. However, in a clinical setting antibiotic treatment usually commences following the onset of symptoms, which in some cases (e.g., biothreat agents) may be too late. In a BALB/c murine intranasal model of infection for Francisella tularensis SCHU S4 infection, we demonstrate during a time course experiment that proinflammatory cytokines and the damage-associated molecular pattern HMGB1 were not significantly elevated above naive levels in tissue or sera until 72 h postinfection. HMGB1 was identified as a potential therapeutic target that could extend the window of opportunity for the treatment of tularemia with antibiotics. Antibodies to HMGB1 were administered in conjunction with a delayed/suboptimal levofloxacin treatment of F. tularensis We found in the intranasal model of infection that treatment with anti-HMGB1 antibody, compared to an isotype IgY control antibody, conferred a significant survival benefit and decreased bacterial loads in the spleen and liver but not the lung (primary loci of infection) 4 days into infection. We also observed an increase in the production of gamma interferon in all tested organs. These data demonstrate that treatment with anti-HMGB1 antibody is beneficial in enhancing the effectiveness of current antibiotics in treating tularemia. Strategies of this type, involving antibiotics in combination with immunomodulatory drugs, are likely to be essential for the development of a postexposure therapeutic for intracellular pathogens., (Copyright © 2013, American Society for Microbiology. All Rights Reserved.)
- Published
- 2013
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32. Scaling of Haversian canal surface area to secondary osteon bone volume in ribs and limb bones.
- Author
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Skedros JG, Knight AN, Clark GC, Crowder CM, Dominguez VM, Qiu S, Mulhern DM, Donahue SW, Busse B, Hulsey BI, Zedda M, and Sorenson SM
- Subjects
- Adult, Aged, Animals, Anthropology, Physical, Anthropometry, Bone Remodeling, Female, Humans, Linear Models, Male, Middle Aged, Bone and Bones anatomy & histology, Haversian System anatomy & histology
- Abstract
Studies of secondary osteons in ribs have provided a great deal of what is known about remodeling dynamics. Compared with limb bones, ribs are metabolically more active and sensitive to hormonal changes, and receive frequent low-strain loading. Optimization for calcium exchange in rib osteons might be achieved without incurring a significant reduction in safety factor by disproportionally increasing central canal size with increased osteon size (positive allometry). By contrast, greater mechanical loads on limb bones might favor reducing deleterious consequences of intracortical porosity by decreasing osteon canal size with increased osteon size (negative allometry). Evidence of this metabolic/mechanical dichotomy between ribs and limb bones was sought by examining relationships between Haversian canal surface area (BS, osteon Haversian canal perimeter, HC.Pm) and bone volume (BV, osteonal wall area, B.Ar) in a broad size range of mature (quiescent) osteons from adult human limb bones and ribs (modern and medieval) and various adult and subadult non-human limb bones and ribs. Reduced major axis (RMA) and least-squares (LS) regressions of HC.Pm/B.Ar data show that rib and limb osteons cannot be distinguished by dimensional allometry of these parameters. Although four of the five rib groups showed positive allometry in terms of the RMA slopes, nearly 50% of the adult limb bone groups also showed positive allometry when negative allometry was expected. Consequently, our results fail to provide clear evidence that BS/BV scaling reflects a rib versus limb bone dichotomy whereby calcium exchange might be preferentially enhanced in rib osteons., (Copyright © 2013 Wiley Periodicals, Inc.)
- Published
- 2013
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33. Targeting the "cytokine storm" for therapeutic benefit.
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D'Elia RV, Harrison K, Oyston PC, Lukaszewski RA, and Clark GC
- Subjects
- Humans, Communicable Diseases immunology, Communicable Diseases pathology, Cytokines antagonists & inhibitors, Cytokines immunology, Immunologic Factors therapeutic use
- Abstract
Inflammation is the body's first line of defense against infection or injury, responding to challenges by activating innate and adaptive responses. Microbes have evolved a diverse range of strategies to avoid triggering inflammatory responses. However, some pathogens, such as the influenza virus and the Gram-negative bacterium Francisella tularensis, do trigger life-threatening "cytokine storms" in the host which can result in significant pathology and ultimately death. For these diseases, it has been proposed that downregulating inflammatory immune responses may improve outcome. We review some of the current candidates for treatment of cytokine storms which may prove useful in the clinic in the future and compare them to more traditional therapeutic candidates that target the pathogen rather than the host response.
- Published
- 2013
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34. Endoscopic vein harvest of the lesser saphenous vein in the supine position: a unique approach to an old problem.
- Author
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Brandt CP, Greene GC, Maggart ML, Hall WC, Harville LE, Pollard TR, and Stouffer CW
- Subjects
- Humans, Tissue and Organ Harvesting adverse effects, Treatment Outcome, Coronary Artery Bypass, Endoscopy adverse effects, Patient Positioning, Saphenous Vein transplantation, Supine Position, Tissue and Organ Harvesting methods
- Abstract
Objectives: To obtain a suitable conduit from the lesser (short) saphenous system for use in coronary artery bypass surgery. We wanted to perform this while the patient was in the supine position as to not disrupt the standard operation, and at the same time, utilizing the endoscopic vein harvest technique with its obvious abilities to decrease vein harvest morbidity. We also theorized that through endoscopic techniques instead of the open technique we could harvest greater lengths of conduit, thus providing quality vein segments for additional grafts if needed., Methods: We were able to perform endoscopic vein harvest while in the supine position with one unique centrally located incision that has not been previously described., Results: The lesser saphenous vein harvested in the described technique provided excellent conduit for our patients that were conduit poor. The endoscopic technique allowed increased length of harvested segments, by giving us the ability to travel under the gastrocnemius muscle with minimal morbidity as opposed to the open technique, where the traditional endpoint is the aforementioned muscle. Conduits were harvested successfully from 14 of 16 candidates. No wound infections or healing problems were experienced. Neurovascular integrity was maintained in all patients., Conclusions: Endoscopic vein harvest of the lesser saphenous vein with the patient in the supine position is safe, effective and affords conduits for a unique subset of patients undergoing coronary artery bypass grafting.
- Published
- 2013
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35. Residential family treatment for parents with substance use disorders who are involved with child welfare: two perspectives on program design, collaboration, and sustainability.
- Author
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Hammond GC and McGlone A
- Subjects
- Adult, Child, Cooperative Behavior, Female, Humans, Male, Minnesota, Oregon, Program Evaluation statistics & numerical data, Residential Treatment statistics & numerical data, Child Welfare statistics & numerical data, Family, Parents, Program Evaluation methods, Residential Treatment methods, Substance-Related Disorders therapy
- Abstract
This article discusses the service design, implementation, and evaluation findings of two residential family treatment programs: Wayside House (MN) and OnTrack (OR). Both programs specialize in family-centered services for adults with substance use disorders (SUD) who are involved with child welfare. Information on program design, services offered, and key collaborations are detailed. Implications for program sustainability are provided.
- Published
- 2013
36. Reactive oxygen species and the MEK/ERK pathway are involved in the toxicity of clostridium perfringens α-toxin, a prototype bacterial phospholipase C.
- Author
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Monturiol-Gross L, Flores-Díaz M, Araya-Castillo C, Pineda-Padilla MJ, Clark GC, Titball RW, and Alape-Girón A
- Subjects
- Animals, Antipyrine administration & dosage, Antipyrine analogs & derivatives, Cell Line, Disease Models, Animal, Edaravone, Free Radical Scavengers administration & dosage, Gas Gangrene drug therapy, Gas Gangrene mortality, Gas Gangrene pathology, Mice, Muscle, Skeletal pathology, Survival Analysis, Treatment Outcome, Bacterial Toxins toxicity, Calcium-Binding Proteins toxicity, Clostridium perfringens pathogenicity, MAP Kinase Signaling System, Reactive Oxygen Species toxicity, Type C Phospholipases toxicity
- Abstract
Clostridium perfringens, the most broadly distributed pathogen in nature, produces a prototype phospholipase C, also called α-toxin, which plays a key role in the pathogenesis of gas gangrene. α-Toxin causes plasma membrane disruption at high concentrations, but the role of intracellular mediators in its toxicity at low concentrations is unknown. This work demonstrates that α-toxin causes oxidative stress and activates the MEK/ERK pathway in cultured cells and furthermore provides compelling evidence that O(2)(-.), hydrogen peroxide, and the OH(.) radical are involved in its cytotoxic and myotoxic effects. The data show that antioxidants and MEK1 inhibitors reduce the cytotoxic and myotoxic effects of α-toxin and demonstrate that edaravone, a clinically used hydroxyl radical trap, reduces the myonecrosis and the mortality caused by an experimental infection with C. perfringens in a murine model of gas gangrene. This knowledge provides new insights for the development of novel therapies to reduce tissue damage during clostridial myonecrosis.
- Published
- 2012
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37. Subtype analysis of Blastocystis isolates in Swedish patients.
- Author
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Forsell J, Granlund M, Stensvold CR, Clark CG, and Evengård B
- Subjects
- Adolescent, Adult, Aged, Aged, 80 and over, Animals, Blastocystis isolation & purification, Child, Child, Preschool, Cluster Analysis, Feces parasitology, Female, Genotype, Humans, Infant, Male, Middle Aged, Molecular Epidemiology, Molecular Sequence Data, Prevalence, RNA, Ribosomal genetics, Sequence Analysis, DNA, Sweden epidemiology, Young Adult, Blastocystis classification, Blastocystis genetics, Blastocystis Infections epidemiology, Blastocystis Infections parasitology, Genetic Variation
- Abstract
Blastocystis is a genetically diverse and widespread intestinal parasite of animals and humans with controversial pathogenic potential. At least nine subtypes of Blastocystis have been found in humans. The genetic diversity of Blastocystis was examined in stool samples from 68 patients from the Stockholm area, Sweden. Blastocystis was identified by light microscopy, and subtyped by sequencing the 5'-end of the small subunit ribosomal RNA gene. Five Blastocystis subtypes were identified in the 63 patients whose samples were successfully subtyped: ST1 (15.9%), ST2 (14.3%), ST3 (47.6%), ST4 (20.6%), and ST7 (1.6%). ST3 was more common in males compared to females (P=0.049). Comparative molecular analysis of Blastocystis sequences revealed intra-subtype variations within the identified subtypes with the exception of ST4. Among ST4 sequences in this study, as well as in the majority of human GenBank sequences, a limited genetic diversity was found compared to what was found among the other common subtypes (ST1, ST2 and ST3). The relative prevalence of ST4 in this study was comparable to the overall distribution of ST4 in European cohorts (16.5%). This contrasts with the sparse reports of ST4 in studies from other continents, which may indicate that the distribution of this subtype is geographically heterogeneous.
- Published
- 2012
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38. Pegylation of antimicrobial peptides maintains the active peptide conformation, model membrane interactions, and antimicrobial activity while improving lung tissue biocompatibility following airway delivery.
- Author
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Morris CJ, Beck K, Fox MA, Ulaeto D, Clark GC, and Gumbleton M
- Subjects
- Animals, Anti-Infective Agents chemical synthesis, Cell Line, Chromatography, High Pressure Liquid, Circular Dichroism, Enzyme-Linked Immunosorbent Assay, Male, Mass Spectrometry, Microbial Sensitivity Tests, Peptides chemical synthesis, Rats, Rats, Sprague-Dawley, Spectrometry, Fluorescence, Anti-Infective Agents chemistry, Anti-Infective Agents pharmacology, Lung drug effects, Lung metabolism, Peptides chemistry, Peptides pharmacology, Polyethylene Glycols chemistry
- Abstract
Antimicrobial peptides (AMPs) have therapeutic potential, particularly for localized infections such as those of the lung. Here we show that airway administration of a pegylated AMP minimizes lung tissue toxicity while nevertheless maintaining antimicrobial activity. CaLL, a potent synthetic AMP (KWKLFKKIFKRIVQRIKDFLR) comprising fragments of LL-37 and cecropin A peptides, was N-terminally pegylated (PEG-CaLL). PEG-CaLL derivatives retained significant antimicrobial activity (50% inhibitory concentrations [IC(50)s] 2- to 3-fold higher than those of CaLL) against bacterial lung pathogens even in the presence of lung lining fluid. Circular dichroism and fluorescence spectroscopy confirmed that conformational changes associated with the binding of CaLL to model microbial membranes were not disrupted by pegylation. Pegylation of CaLL reduced AMP-elicited cell toxicity as measured using in vitro lung epithelial primary cell cultures. Further, in a fully intact ex vivo isolated perfused rat lung (IPRL) model, airway-administered PEG-CaLL did not result in disruption of the pulmonary epithelial barrier, whereas CaLL caused an immediate loss of membrane integrity leading to pulmonary edema. All AMPs (CaLL, PEG-CaLL, LL-37, cecropin A) delivered to the lung by airway administration showed limited (<3%) pulmonary absorption in the IPRL with extensive AMP accumulation in lung tissue itself, a characteristic anticipated to be beneficial for the treatment of pulmonary infections. We conclude that pegylation may present a means of improving the lung biocompatibility of AMPs designed for the treatment of pulmonary infections.
- Published
- 2012
- Full Text
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39. Altered carnitine metabolism in dialysis patients with reduced physical function may be due to dysfunctional fatty acid oxidation.
- Author
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Murphy WJ, Steiber A, Connery GC, Carder J, Spry L, and Hoppel C
- Subjects
- Aged, Anemia drug therapy, Anemia etiology, Carnitine blood, Cross-Sectional Studies, Erythrocytes pathology, Erythropoietin therapeutic use, Female, Follow-Up Studies, Humans, Lipid Metabolism, Male, Oxidation-Reduction, Prognosis, Activities of Daily Living, Carnitine analogs & derivatives, Carnitine metabolism, Fatty Acids metabolism, Quality of Life, Renal Dialysis adverse effects
- Abstract
Background: It has been reported that hemodialysis patients have elevated plasma acylcarnitine concentrations, which correlates to reduced red blood cell integrity. It has also been reported that the supplementation of L-carnitine for these patients improves anemia, glucose metabolism and muscle function, but the mechanism of these relationships remains unknown. We hypothesized that the cause of increased plasma acylcarnitines is incomplete fatty acid oxidation and the underlying disturbance of metabolism reduces muscle function, resulting in decreased ability to function and quality of life, and glucose availability, resulting in decreased red blood cell integrity and worsened anemia., Methods: This analysis was conducted on baseline data from a clinical trial of carnitine supplementation in hemodialysis patients with reduced physical function and free carnitine levels. Partial correlations controlling for age, gender, hemoglobin and subjective global assessment score for each acylcarnitine species and outcome were computed using SPSS version 17.0 and a significance level of P < 0.05. To measure the impact of acylcarnitine acyl chain length on these relationships, the correlation coefficients were categorized by chain length and linear regressions were computed for each outcome measure., Results: Linear regression analysis (n = 58) revealed significant negative relationships between chain length and Short Form-36 physical composite score, sit-to-stand count and 6-min walk distance (r(2) = 0.635, 0.332 and 0.347, respectively) and a significant positive relationship with erythropoietin dose (r(2) = 0.181)., Conclusion: Our data revealed that longer acyl chain length significantly predicts poorer physical function and worsened anemia, and this data supports our proposed mechanism, which may lead to increased understanding of altered carnitine metabolism in hemodialysis patients.
- Published
- 2012
- Full Text
- View/download PDF
40. Analysis of the effect of osteon diameter on the potential relationship of osteocyte lacuna density and osteon wall thickness.
- Author
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Skedros JG, Clark GC, Sorenson SM, Taylor KW, and Qiu S
- Subjects
- Adult, Animals, Bone and Bones anatomy & histology, Bone and Bones physiology, Deer, Extremities anatomy & histology, Haversian System anatomy & histology, Haversian System physiology, Horses, Humans, Male, Osteocytes physiology, Ribs anatomy & histology, Ribs physiology, Sheep, Young Adult, Bone Remodeling, Bone and Bones cytology, Extremities growth & development, Haversian System cytology, Osteocytes cytology, Ribs cytology
- Abstract
An important hypothesis is that the degree of infilling of secondary osteons (Haversian systems) is controlled by the inhibitory effect of osteocytes on osteoblasts, which might be mediated by sclerostin (a glycoprotein produced by osteocytes). Consequently, this inhibition could be proportional to cell number: relatively greater repression is exerted by progressively greater osteocyte density (increased osteocytes correlate with thinner osteon walls). This hypothesis has been examined, but only weakly supported, in sheep ulnae. We looked for this inverse relationship between osteon wall thickness (On.W.Th) and osteocyte lacuna density (Ot.Lc.N/B.Ar) in small and large osteons in human ribs, calcanei of sheep, deer, elk, and horses, and radii and third metacarpals of horses. Analyses involved: (1) all osteons, (2) smaller osteons, either ≤150 μm diameter or less than or equal to the mean diameter, and (3) larger osteons (>mean diameter). Significant, but weak, correlations between Ot.Lc.N/B.Ar and On.W.Th/On.Dm (On.Dm = osteon diameter) were found when considering all osteons in limb bones (r values -0.16 to -0.40, P < 0.01; resembling previous results in sheep ulnae: r = -0.39, P < 0.0001). In larger osteons, these relationships were either not significant (five/seven bone types) or very weak (two/seven bone types). In ribs, a negative relationship was only found in smaller osteons (r = -0.228, P < 0.01); this inverse relationship in smaller osteons did not occur in elk calcanei. These results do not provide clear or consistent support for the hypothesized inverse relationship. However, correlation analyses may fail to detect osteocyte-based repression of infilling if the signal is spatially nonuniform (e.g., increased near the central canal)., (Copyright © 2011 Wiley-Liss, Inc.)
- Published
- 2011
- Full Text
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41. Comparison of a nontoxic variant of Clostridium perfringens α-toxin with the toxic wild-type strain.
- Author
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Vachieri SG, Clark GC, Alape-Girón A, Flores-Díaz M, Justin N, Naylor CE, Titball RW, and Basak AK
- Subjects
- Animals, Apoptosis drug effects, Apoptosis genetics, Bacterial Toxins genetics, Bacterial Toxins metabolism, Bacterial Toxins toxicity, Calcium-Binding Proteins genetics, Calcium-Binding Proteins metabolism, Calcium-Binding Proteins toxicity, Catalytic Domain genetics, Cattle, Cell Line, Cell Membrane metabolism, Creatine Kinase blood, Crystallization, Endothelial Cells pathology, Host-Pathogen Interactions, Humans, Mice, Mice, Inbred Strains, Mutant Proteins genetics, Mutant Proteins metabolism, Mutant Proteins toxicity, Protein Binding, Protein Conformation, Substrate Specificity genetics, Type C Phospholipases genetics, Type C Phospholipases metabolism, Type C Phospholipases toxicity, Bacterial Toxins chemistry, Calcium-Binding Proteins chemistry, Clostridium perfringens pathogenicity, Endothelial Cells drug effects, Mutant Proteins chemistry, Phospholipids metabolism, Type C Phospholipases chemistry
- Abstract
The α-toxin produced by Clostridium perfringens is one of the best-studied examples of a toxic phospholipase C. In this study, a nontoxic mutant protein from C. perfringens strain NCTC8237 in which Thr74 is substituted by isoleucine (T74I) has been characterized and is compared with the toxic wild-type protein. Thr74 is part of an exposed loop at the proposed membrane-interfacing surface of the toxin. The mutant protein had markedly reduced cytotoxic and myotoxic activities. However, this substitution did not significantly affect the catalytic activity towards water-soluble substrate or the overall three-dimensional structure of the protein. The data support the proposed role of the 70-90 loop in the recognition of membrane phospholipids. These findings also provide key evidence in support of the hypothesis that the hydrolysis of both phosphatidylcholine and sphingomyelin are required for the cytolytic and toxic activity of phospholipases.
- Published
- 2010
- Full Text
- View/download PDF
42. Exploring smoking prevalence, quit attempts, and readiness to quit cigarette use among women in substance abuse treatment.
- Author
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Teater B and Hammond GC
- Subjects
- Adolescent, Adult, Aged, Comorbidity, Female, Humans, Logistic Models, Middle Aged, Models, Psychological, Multivariate Analysis, Ohio epidemiology, Prevalence, Smoking epidemiology, Tobacco Use Disorder epidemiology, Patient Acceptance of Health Care psychology, Smoking Cessation psychology, Smoking Prevention, Substance-Related Disorders epidemiology, Tobacco Use Disorder prevention & control
- Abstract
Client questionnaires from 38 gender-specific substance abuse facilities throughout Ohio were analyzed to explore smoking prevalence, quit attempts, and readiness to quit cigarette use. The analysis revealed 79.7% of women used cigarettes at the time of the survey, 33.5% of current smokers had made at least one quit attempt within the past 12 months, and 55.2% of current smokers reported either contemplating or preparing to make a quit attempt. A multinomial logistic regression revealed that clients who experienced a past quit attempt were more likely to be in the contemplation and preparation stages and clients who smoked 30 out of the past 30 days were least likely to be in the preparation stage. Clients who reported smoking between 10-15 cigarettes a day were more likely to be in the contemplation stage than those who reported smoking <10 cigarettes a day. A three-pronged approach that examines the physiological, emotional, and social components of addiction is recommended.
- Published
- 2010
- Full Text
- View/download PDF
43. Ophthalmic presentation of Wegener's granulomatosis on a background of polymyalgia rheumatica.
- Author
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Clark GC, Moloney G, and Sutton G
- Subjects
- Aged, Cyclophosphamide therapeutic use, Drug Therapy, Combination, Glucocorticoids therapeutic use, Granulomatosis with Polyangiitis drug therapy, Humans, Immunosuppressive Agents therapeutic use, Keratoconjunctivitis drug therapy, Limbus Corneae drug effects, Male, Granulomatosis with Polyangiitis diagnosis, Keratoconjunctivitis diagnosis, Limbus Corneae pathology, Polymyalgia Rheumatica complications
- Abstract
We present a case of Wegener's granulomatosis (WG) in a 79-year-old man with limbitis and granulomatous conjunctivitis, on a background of polymyalgia rheumatica (PMR). The undifferentiated nature of ocular presentations of WG can be diagnostically challenging, especially in cases, such as this, where findings are initially inconclusive and evolve with time. This case highlights the significance of a history of PMR in patients with ocular inflammation. The systemic inflammatory systems of WG, including arthralgias, may mimic other conditions such as PMR. Patients with undifferentiated ocular inflammatory syndromes should be questioned regarding arthralgias, myalgias and stiffness. Such symptoms, or a background of PMR, should raise suspicion of WG.
- Published
- 2010
- Full Text
- View/download PDF
44. Novel peptide therapeutics for treatment of infections.
- Author
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Oyston PCF, Fox MA, Richards SJ, and Clark GC
- Subjects
- Humans, Protein Conformation, Antimicrobial Cationic Peptides therapeutic use, Bacterial Infections drug therapy, Carrier Proteins therapeutic use
- Abstract
As antibiotic resistance increases worldwide, there is an increasing pressure to develop novel classes of antimicrobial compounds to fight infectious disease. Peptide therapeutics represent a novel class of therapeutic agents. Some, such as cationic antimicrobial peptides and peptidoglycan recognition proteins, have been identified from studies of innate immune effector mechanisms, while others are completely novel compounds generated in biological systems. Currently, only selected cationic antimicrobial peptides have been licensed, and only for topical applications. However, research using new approaches to identify novel antimicrobial peptide therapeutics, and new approaches to delivery and improving stability, will result in an increased range of peptide therapeutics available in the clinic for broader applications.
- Published
- 2009
- Full Text
- View/download PDF
45. Ultrastructure of the retinal synapses in cubozoans.
- Author
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Gray GC, Martin VJ, and Satterlie RA
- Subjects
- Animals, Microscopy, Electron, Transmission, Cubozoa ultrastructure, Retina ultrastructure, Synapses ultrastructure
- Abstract
Cubomedusae (box jellyfish) are well known for strong directional swimming, rapid responses to visual stimuli, and complex lensed eyes comparable to those of more advanced multicellular animals. They possess a total of 24 eyes that are of four morphologically different types, yet little is known about the neural organization of their eyes. The eyes are located on ganglion-like structures called rhopalia. Each of the four rhopalia contains an upper and a lower lensed eye (with a cornea, lens, and retina), two pit ocelli, and two slit ocelli. Transmission electron microscopy was used to examine the synaptic morphology of the eyes and pacemaker region of four species of cubozoans (Tamoya haplonema, Carybdea marsupialis, Tripedalia cystophora, and Chiropsalmus quadrumanus). Invaginated synapses were found in all four species, but only in the upper and lower lensed eyes. Density measurements indicated that the invaginated synapses were located close to the basal region of photoreceptor cells, and size differences of invaginated synapses were observed between the upper and lower lensed eyes, as well as between species. Four additional types of chemical synapses-clear unidirectional, dense-core unidirectional, clear bidirectional, and clear and dense-core bidirectional-were also observed in the rhopalia. The invaginated synapses of the lensed eyes may be useful as markers to help sort out the neural circuitry in the retinal region of these complex cubomedusan eyes.
- Published
- 2009
- Full Text
- View/download PDF
46. Determination of relative assay response factors for toxic chlorinated and brominated dioxins/furans using an enzyme immunoassay (EIA) and a chemically-activated luciferase gene expression cell bioassay (CALUX).
- Author
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Samara F, Gullett BK, Harrison RO, Chu A, and Clark GC
- Subjects
- Biological Assay methods, Cross Reactions, Gene Expression, Humans, Hydrocarbons, Brominated analysis, Hydrocarbons, Brominated toxicity, Hydrocarbons, Chlorinated analysis, Hydrocarbons, Chlorinated toxicity, Immunoenzyme Techniques methods, Luciferases metabolism, Dioxins analysis, Dioxins toxicity, Ecotoxicology methods, Furans analysis, Furans toxicity
- Abstract
Determination of toxic activity requires knowledge of both the concentration and toxicity to evaluate the risk for adverse human health and environmental effects. A chemically-activated luciferase gene expression cell bioassay system (CALUX) and an antibody-based method enzyme immunoassay (EIA) were used to detect the dioxin-like response of several polybrominated, polychlorinated, and polybrominated/chlorinated dibenzo-p-dioxins/furans (PBDDs/Fs, PCDDs/Fs, and PBCDDs/Fs, respectively). It has been suggested that the biological activity of the brominated and mixed bromo/chloro compounds is similar to their chlorinated analogues (measured by binding to the Ah receptor). PBDD/F, PCDD/F, and PBCDD/F laboratory standards exhibited biological activity ranging over three orders of magnitude. The highest relative potency (REP) values from CALUX analysis, when compared to 2,3,7,8-TCDD, were 2,3,7,8-TBDD at 0.99 (+/-0.07), 1,2,3,7,8-PeCDD at 0.69, and 2-Br-3,7,8-TriCDD at 0.72 (+/-0.02). Cross-reactivities were calculated using EIA for several PBDDs/Fs and PBCDDs. The highest percent cross-reactivity was found for 2,3,7,8-TBDD at 138 (+/-34%), and 2,3,7-TriBDD at 84 (+/-36%).
- Published
- 2009
- Full Text
- View/download PDF
47. Recombinant Bacillus subtilis expressing the Clostridium perfringens alpha toxoid is a candidate orally delivered vaccine against necrotic enteritis.
- Author
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Hoang TH, Hong HA, Clark GC, Titball RW, and Cutting SM
- Subjects
- Administration, Oral, Animals, Antibodies, Bacterial blood, Antibodies, Bacterial immunology, Antigens, Bacterial immunology, Bacillus subtilis genetics, Bacterial Toxins genetics, Bacterial Vaccines administration & dosage, Calcium-Binding Proteins genetics, Enzyme-Linked Immunosorbent Assay, Female, Mice, Mice, Inbred BALB C, Type C Phospholipases genetics, Bacillus subtilis immunology, Bacterial Toxins immunology, Bacterial Vaccines immunology, Calcium-Binding Proteins immunology, Gas Gangrene prevention & control, Poultry Diseases prevention & control, Type C Phospholipases immunology
- Abstract
Recombinant Bacillus subtilis endospores have been used to vaccinate against tetanus and anthrax. In this work, we have developed spores that could be used to vaccinate against Clostridium perfringens alpha toxin and that could be used to protect against gas gangrene in humans and necrotic enteritis in poultry. The primary active agent in both cases is alpha toxin. A carboxy-terminal segment of the alpha toxin gene (cpa) fused to the glutathione-S-transferase (GST) gene was cloned in B. subtilis such that the encoded GST-Cpa(247-370) polypeptide had been expressed in the following three different ways: expression in the vegetative cell, expression on the surface of the spore coat (fused to the CotB spore coat protein), and a combined approach of spore coat expression coupled with expression in the vegetative cell. Mice immunized orally or nasally with three doses of recombinant spores that carried GST-Cpa(247-370) on the spore surface showed the most striking responses. This included seroconversion with anti-Cpa(247-370)-specific immunoglobulin G (IgG) responses in their sera, a Th2 bias, and secretory IgA responses in saliva, feces, and lung samples. Neutralizing IgG antibodies to alpha toxin were detected using in vitro and in vivo assays, and a toxin challenge established protection. Mice immunized nasally or orally with recombinant spores were protected against a challenge with 12 median lethal doses of alpha toxin. Existing use of spores as competitive exclusion agents in animal feeds supports their use as a potentially economical and heat-stable vaccine for the poultry industry.
- Published
- 2008
- Full Text
- View/download PDF
48. AH receptor agonist activity in human blood measured with a cell-based bioassay: evidence for naturally occurring AH receptor ligands in vivo.
- Author
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Connor KT, Harris MA, Edwards MR, Budinsky RA, Clark GC, Chu AC, Finley BL, and Rowlands JC
- Subjects
- Adult, Biological Assay methods, Cytochrome P-450 CYP1A1, Dibenzofurans, Polychlorinated, Diet methods, Female, Humans, Indoles administration & dosage, Ligands, Lipids blood, Male, Receptors, Aryl Hydrocarbon agonists, Receptors, Aryl Hydrocarbon genetics, Vegetables, Benzofurans blood, Polychlorinated Biphenyls blood, Polychlorinated Dibenzodioxins analogs & derivatives, Polychlorinated Dibenzodioxins blood, Receptors, Aryl Hydrocarbon blood
- Abstract
In the present study, an aryl hydrocarbon receptor (AHR)-driven reporter gene bioassay was used to measure the activity, measured as an induction equivalent (IEQ) as compared to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), or IEQ concentration in human blood samples from 10 volunteers under different dietary regimens. Blood concentrations of polychlorinated dibenzo-p-dioxins/furans (PCDD/Fs) and polychlorinated biphenyls (PCBs), as determined by analytical chemistry (HR-GC/MS), and expressed as toxic equivalents (TEQs) with the use of TCDD equivalency factors (TEFs), were within a range that has been reported in the general US population, ranging from 0.022 to 0.119 ppt (whole blood basis). However, the human blood IEQ measured directly via bioassay ranged from 13.4 to 218 ppt (whole blood basis). These order of magnitude greater IEQs compared to the TEQs for dioxins, furans, and certain PCBs suggests that human blood contains a relatively high level of AHR agonists able to activate the CYP1A1 dioxin response element (DRE)-linked reporter gene bioassay and that this AHR activity is not accounted for by PCDDs/Fs and dioxin-like PCBs based on standard HR-GC/MS and TEF analysis. When study participants switched from a "baseline" to a high-vegetable diet, increases in bioassay IEQ were observed that were statistically significant (P<0.05). In addition, IEQ activity was elevated above levels observed following dietary intervention in two subjects given indole-3-carbinol (I3C) supplements. We conclude that a substantial portion of the IEQ activity occurred as a result of the increased intake of natural AHR agonists (NAHRAs) present in many fruits, vegetables. and herbs. Our findings also suggest that dietary NAHRAs constitute a substantial daily dietary intake of AHR-active compounds, and these NAHRAs could influence AHR status in humans and play a role in a basal level of AHR activation.
- Published
- 2008
- Full Text
- View/download PDF
49. Clinical and economic implications of incretin-based therapies.
- Author
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Briggs GC
- Subjects
- Blood Glucose analysis, Female, Humans, Male, Middle Aged, Treatment Outcome, Diabetes Mellitus, Type 2 drug therapy, Diabetes Mellitus, Type 2 economics, Hypoglycemic Agents therapeutic use, Incretins therapeutic use
- Published
- 2007
50. Analysis of protection afforded by a Clostridium perfringens alpha-toxoid against heterologous clostridial phospholipases C.
- Author
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Neeson BN, Clark GC, Atkins HS, Lingard B, and Titball RW
- Subjects
- Animals, Bacterial Vaccines immunology, Cattle, Clostridium Infections enzymology, Clostridium Infections immunology, Clostridium Infections microbiology, Cross Reactions, Female, Mice, Mice, Inbred BALB C, Recombinant Proteins biosynthesis, Recombinant Proteins genetics, Recombinant Proteins isolation & purification, Recombinant Proteins pharmacology, Type C Phospholipases immunology, Type C Phospholipases metabolism, Bacterial Toxins immunology, Bacterial Vaccines pharmacology, Calcium-Binding Proteins immunology, Clostridium Infections prevention & control, Clostridium bifermentans enzymology, Clostridium perfringens immunology, Type C Phospholipases antagonists & inhibitors
- Abstract
The major virulence determinant in clostridial myonecrosis caused by Clostridium perfringens is a phospholipase C (PLC), the alpha-toxin. Previously, mice have been protected against challenge with heterologous alpha-toxin or Clostridium perfringens spores by immunisation with the C-domain (known as Cpa(247-370) or alpha-toxoid) of the alpha-toxin. In this study, we have determined the ability of the alpha-toxoid to protect against the lethal effects of a divergent C. perfringens alpha-toxin and against the PLCs of C. absonum or C. bifermentans, species which have been isolated from cases of clostridial myonecrosis. Protection against the C. perfringens alpha-toxin variant, the C. absonum alpha-toxin or the C. bifermentans PLC was elicited by immunisation with the alpha-toxoid in vivo.
- Published
- 2007
- Full Text
- View/download PDF
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