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1.
Clin Exp Allergy ; 54(1): 21-33, 2024 01.
Artigo em Inglês | MEDLINE | ID: mdl-38177093

RESUMO

BACKGROUND: Vancomycin, a glycopeptide antibiotic used for Gram-positive bacterial infections, has been linked with drug reaction with eosinophilia and systemic symptoms (DRESS) in HLA-A*32:01-expressing individuals. This is associated with activation of T lymphocytes, for which glycolysis has been isolated as a fuel pathway following antigenic stimulation. However, the metabolic processes that underpin drug-reactive T-cell activation are currently undefined and may shed light on the energetic conditions needed for the elicitation of drug hypersensitivity or tolerogenic pathways. Here, we sought to characterise the immunological and metabolic pathways involved in drug-specific T-cell activation within the context of DRESS pathogenesis using vancomycin as model compound and drug-reactive T-cell clones (TCCs) generated from healthy donors and vancomycin-hypersensitive patients. METHODS: CD4+ and CD8+ vancomycin-responsive TCCs were generated by serial dilution. The Seahorse XFe96 Analyzer was used to measure the extracellular acidification rate (ECAR) as an indicator of glycolytic function. Additionally, T-cell proliferation and cytokine release (IFN-γ) assay were utilised to correlate the bioenergetic characteristics of T-cell activation with in vitro assays. RESULTS: Model T-cell stimulants induced non-specific T-cell activation, characterised by immediate augmentation of ECAR and rate of ATP production (JATPglyc). There was a dose-dependent and drug-specific glycolytic shift when vancomycin-reactive TCCs were exposed to the drug. Vancomycin-reactive TCCs did not exhibit T-cell cross-reactivity with structurally similar compounds within proliferative and cytokine readouts. However, cross-reactivity was observed when analysing energetic responses; TCCs with prior specificity for vancomycin were also found to exhibit glycolytic switching after exposure to teicoplanin. Glycolytic activation of TCC was HLA restricted, as exposure to HLA blockade attenuated the glycolytic induction. CONCLUSION: These studies describe the glycolytic shift of CD4+ and CD8+ T cells following vancomycin exposure. Since similar glycolytic switching is observed with teicoplanin, which did not activate T cells, it is possible the master switch for T-cell activation is located upstream of metabolic signalling.


Assuntos
Teicoplanina , Vancomicina , Humanos , Vancomicina/efeitos adversos , Linfócitos T CD8-Positivos , Ativação Linfocitária , Citocinas , Glicólise
2.
Arch Toxicol ; 95(4): 1335-1347, 2021 04.
Artigo em Inglês | MEDLINE | ID: mdl-33585966

RESUMO

Mitochondrial DNA (mtDNA) is highly polymorphic and encodes 13 proteins which are critical to the production of ATP via oxidative phosphorylation. As mtDNA is maternally inherited and undergoes negligible recombination, acquired mutations have subdivided the human population into several discrete haplogroups. Mitochondrial haplogroup has been found to significantly alter mitochondrial function and impact susceptibility to adverse drug reactions. Despite these findings, there are currently limited models to assess the effect of mtDNA variation upon susceptibility to adverse drug reactions. Platelets offer a potential personalised model of this variation, as their anucleate nature offers a source of mtDNA without interference from the nuclear genome. This study, therefore, aimed to determine the effect of mtDNA variation upon mitochondrial function and drug-induced mitochondrial dysfunction in a platelet model. The mtDNA haplogroup of 383 healthy volunteers was determined using next-generation mtDNA sequencing (Illumina MiSeq). Subsequently, 30 of these volunteers from mitochondrial haplogroups H, J, T and U were recalled to donate fresh, whole blood from which platelets were isolated. Platelet mitochondrial function was tested at basal state and upon treatment with compounds associated with both mitochondrial dysfunction and adverse drug reactions, flutamide, 2-hydroxyflutamide and tolcapone (10-250 µM) using extracellular flux analysis. This study has demonstrated that freshly-isolated platelets are a practical, primary cell model, which is amenable to the study of drug-induced mitochondrial dysfunction. Specifically, platelets from donors of haplogroup J have been found to have increased susceptibility to the inhibition of complex I-driven respiration by 2-hydroxyflutamide. At a time when individual susceptibility to adverse drug reactions is not fully understood, this study provides evidence that inter-individual variation in mitochondrial genotype could be a factor in determining sensitivity to mitochondrial toxicants associated with costly adverse drug reactions.


Assuntos
Plaquetas/efeitos dos fármacos , DNA Mitocondrial/efeitos dos fármacos , Flutamida/análogos & derivados , Tolcapona/toxicidade , Adolescente , Adulto , DNA Mitocondrial/genética , Feminino , Flutamida/toxicidade , Genótipo , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Masculino , Pessoa de Meia-Idade , Adulto Jovem
3.
Biochem Soc Trans ; 48(3): 787-797, 2020 06 30.
Artigo em Inglês | MEDLINE | ID: mdl-32453388

RESUMO

The mitochondrion is an essential organelle responsible for generating cellular energy. Additionally, mitochondria are a source of inter-individual variation as they contain their own genome. Evidence has revealed that mitochondrial DNA (mtDNA) variation can confer differences in mitochondrial function and importantly, these differences may be a factor underlying the idiosyncrasies associated with unpredictable drug-induced toxicities. Thus far, preclinical and clinical data are limited but have revealed evidence in support of an association between mitochondrial haplogroup and susceptibility to specific adverse drug reactions. In particular, clinical studies have reported associations between mitochondrial haplogroup and antiretroviral therapy, chemotherapy and antibiotic-induced toxicity, although study limitations and conflicting findings mean that the importance of mtDNA variation to toxicity remains unclear. Several studies have used transmitochondrial cybrid cells as personalised models with which to study the impact of mitochondrial genetic variation. Cybrids allow the effects of mtDNA to be assessed against a stable nuclear background and thus the in vitro elucidation of the fundamental mechanistic basis of such differences. Overall, the current evidence supports the tenet that mitochondrial genetics represent an exciting area within the field of personalised medicine and drug toxicity. However, further research effort is required to confirm its importance. In particular, efforts should focus upon translational research to connect preclinical and clinical data that can inform whether mitochondrial genetics can be useful to identify at risk individuals or inform risk assessment during drug development.


Assuntos
Efeitos Colaterais e Reações Adversas Relacionados a Medicamentos/genética , Genótipo , Mitocôndrias/genética , Animais , Antibacterianos/toxicidade , Antirretrovirais/toxicidade , Antineoplásicos/toxicidade , Núcleo Celular , DNA Mitocondrial/genética , Desenvolvimento de Medicamentos , Variação Genética , Haplótipos , Humanos , Polimorfismo de Nucleotídeo Único
4.
Toxicol Appl Pharmacol ; 403: 115163, 2020 09 15.
Artigo em Inglês | MEDLINE | ID: mdl-32730777

RESUMO

During its clinical development fialuridine caused liver toxicity and the death of five patients. This case remains relevant due to the continued development of mechanistically-related compounds against a back-drop of simple in vitro models which remain limited for the preclinical detection of such delayed toxicity. Here, proteomic investigation of a differentiated, HepaRG, and proliferating, HepG2 cell model was utilised to confirm the presence of the hENT1 transporter, thymidine kinase-1 and -2 (TK1, TK2) and thymidylate kinase, all essential in order to reproduce the cellular activation and disposition of fialuridine in the clinic. Acute metabolic modification assays could only identify mitochondrial toxicity in HepaRG cells following extended dosing, 2 weeks. Toxic effects were observed around 10 µM, which is within a range of 10-15 X approximate Cmax. HepaRG cell death was accompanied by a significant decrease in mitochondrial DNA content, indicative of inhibition of mitochondrial replication, and a subsequent reduction in mitochondrial respiration and the activity of mitochondrial respiratory complexes, not replicated in HepG2 cells. The structural epimer of fialuridine, included as a pharmacological negative control, was shown to have no cytotoxic effects in HepaRG cells up to 4 weeks. Overall, these comparative studies demonstrate the HepaRG model has translational relevance for fialuridine toxicity and therefore may have potential in investigating the inhibition of mitochondrial replication over prolonged exposure for other toxicants.


Assuntos
Antivirais/farmacologia , Arabinofuranosiluracila/análogos & derivados , Hepatócitos/efeitos dos fármacos , Mitocôndrias/efeitos dos fármacos , Arabinofuranosiluracila/farmacologia , Linhagem Celular Tumoral , Replicação do DNA/efeitos dos fármacos , DNA Mitocondrial/fisiologia , Relação Dose-Resposta a Droga , Humanos , Mitocôndrias/fisiologia
5.
Cytotherapy ; 19(4): 555-569, 2017 04.
Artigo em Inglês | MEDLINE | ID: mdl-28214127

RESUMO

BACKGROUND AIMS: Tracking cells during regenerative cytotherapy is crucial for monitoring their safety and efficacy. Macrophages are an emerging cell-based regenerative therapy for liver disease and can be readily labeled for medical imaging. A reliable, clinically applicable cell-tracking agent would be a powerful tool to study cell biodistribution. METHODS: Using a recently described chemical design, we set out to functionalize, optimize and characterize a new set of superparamagnetic iron oxide nanoparticles (SPIONs) to efficiently label macrophages for magnetic resonance imaging-based cell tracking in vivo. RESULTS: A series of cell health and iron uptake assays determined that positively charged SPIONs (+16.8 mV) could safely label macrophages more efficiently than the formerly approved ferumoxide (-6.7 mV; Endorem) and at least 10 times more efficiently than the clinically approved SPION ferumoxytol (-24.2 mV; Rienso). An optimal labeling time of 4 h at 25 µg/mL was demonstrated to label macrophages of mouse and human origin without any adverse effects on cell viability whilst providing substantial iron uptake (>5 pg Fe/cell) that was retained for 7 days in vitro. SPION labeling caused no significant reduction in phagocytic activity and a shift toward a reversible M1-like phenotype in bone marrow-derived macrophages (BMDMs). Finally, we show that SPION-labeled BMDMs delivered via the hepatic portal vein to mice are localized in the hepatic parenchyma resulting in a 50% drop in T2* in the liver. Engraftment of exogenous cells was confirmed via immunohistochemistry up to 3 weeks posttransplantation. DISCUSSION: A positively charged dextran-coated SPION is a promising tool to noninvasively track hepatic macrophage localization for therapeutic monitoring.


Assuntos
Rastreamento de Células/métodos , Dextranos/química , Ferro/metabolismo , Macrófagos/citologia , Macrófagos/metabolismo , Imageamento por Ressonância Magnética/métodos , Nanopartículas de Magnetita/química , Animais , Células da Medula Óssea/citologia , Células da Medula Óssea/metabolismo , Transplante de Medula Óssea/métodos , Sobrevivência Celular , Células Cultivadas , Dextranos/farmacocinética , Óxido Ferroso-Férrico/química , Óxido Ferroso-Férrico/farmacocinética , Humanos , Cirrose Hepática/terapia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Distribuição Tecidual
6.
Arch Toxicol ; 91(1): 439-452, 2017 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-27039104

RESUMO

The application of primary human hepatocytes following isolation from human tissue is well accepted to be compromised by the process of dedifferentiation. This phenomenon reduces many unique hepatocyte functions, limiting their use in drug disposition and toxicity assessment. The aetiology of dedifferentiation has not been well defined, and further understanding of the process would allow the development of novel strategies for sustaining the hepatocyte phenotype in culture or for improving protocols for maturation of hepatocytes generated from stem cells. We have therefore carried out the first proteomic comparison of primary human hepatocyte differentiation. Cells were cultured for 0, 24, 72 and 168 h as a monolayer in order to permit unrestricted hepatocyte dedifferentiation, so as to reveal the causative signalling pathways and factors in this process, by pathway analysis. A total of 3430 proteins were identified with a false detection rate of <1 %, of which 1117 were quantified at every time point. Increasing numbers of significantly differentially expressed proteins compared with the freshly isolated cells were observed at 24 h (40 proteins), 72 h (118 proteins) and 168 h (272 proteins) (p < 0.05). In particular, cytochromes P450 and mitochondrial proteins underwent major changes, confirmed by functional studies and investigated by pathway analysis. We report the key factors and pathways which underlie the loss of hepatic phenotype in vitro, particularly those driving the large-scale and selective remodelling of the mitochondrial and metabolic proteomes. In summary, these findings expand the current understanding of dedifferentiation should facilitate further development of simple and complex hepatic culture systems.


Assuntos
Regulação da Expressão Gênica no Desenvolvimento , Hepatócitos/metabolismo , Farmacologia/métodos , Proteoma/metabolismo , Toxicologia/métodos , Desdiferenciação Celular/efeitos dos fármacos , Células Cultivadas , Complexo I de Transporte de Elétrons/antagonistas & inibidores , Complexo I de Transporte de Elétrons/metabolismo , Perfilação da Expressão Gênica , Regulação da Expressão Gênica no Desenvolvimento/efeitos dos fármacos , Hepatócitos/citologia , Hepatócitos/efeitos dos fármacos , Humanos , Cinética , Mitocôndrias Hepáticas/efeitos dos fármacos , Mitocôndrias Hepáticas/enzimologia , Mitocôndrias Hepáticas/metabolismo , Estabilidade Proteica/efeitos dos fármacos , Proteoma/genética , Reprodutibilidade dos Testes , Rotenona/farmacologia , Desacopladores/farmacologia
7.
Health Commun ; 31(6): 782-5, 2016.
Artigo em Inglês | MEDLINE | ID: mdl-26580230

RESUMO

Climate change is one of the most serious and pervasive challenges facing us today. Our changing climate has implications not only for the ecosystems upon which we depend, but also for human health. Health communication scholars are well-positioned to aid in the mitigation of and response to climate change and its health effects. To help theorists, researchers, and practitioners engage in these efforts, this primer explains relevant issues and vocabulary associated with climate change and its impacts on health. First, this primer provides an overview of climate change, its causes and consequences, and its impacts on health. Then, the primer describes ways to decrease impacts and identifies roles for health communication scholars in efforts to address climate change and its health effects.


Assuntos
Mudança Climática , Comunicação em Saúde , Transtornos de Estresse por Calor/epidemiologia , Humanos
8.
Health Commun ; 31(12): 1447-59, 2016 12.
Artigo em Inglês | MEDLINE | ID: mdl-27054822

RESUMO

How we cope with the many stressors that we encounter throughout our lives has implications for our well-being. By affecting how individuals appraise stressful events, communication can prolong or ameliorate physiological and emotional responses to stress. This study investigated the short-term effects of hope-inducing and rumination-inducing messages on heart rate, state anxiety, and emotions after a standardized, social-evaluative stressor. Continuous heart rate was monitored for 127 college students (64 female, 63 male) throughout an experiment that included a performance stressor and messages designed to (a) cause feelings of hope, (b) evoke rumination, or (c) be a distraction (control). Heart rate varied by message, such that heart rate was lowest in the hope evocation condition. State anxiety was lower in the hope evocation and distraction control conditions than in the rumination condition. The rumination condition led to greater anger, greater guilt, and less happiness than did the other conditions. This study advances our knowledge about potential ways that communication messages can counter the psychological and biological effects of stressful life events. Overall, the study provides preliminary evidence that hope evocation messages may be a form of supportive communication and can ameliorate stress.


Assuntos
Ansiedade/psicologia , Comunicação , Emoções , Frequência Cardíaca/fisiologia , Esperança , Estresse Psicológico , Pensamento , Adolescente , Adulto , Feminino , Humanos , Masculino , Análise e Desempenho de Tarefas , Adulto Jovem
9.
Forensic Sci Med Pathol ; 12(1): 4-13, 2016 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-26782961

RESUMO

PURPOSE: The purpose of this study was to determine the major subcategories and clinicopathologic features of sudden unexpected death in young children in a large retrospective cohort, and to confirm the association of sudden unexplained death in children (abbreviated by us for unexplained deaths as SUDC) with hippocampal pathology and/or febrile seizures. METHODS: We undertook analysis of a retrospective cohort of 151 cases, of which 80% (121/151) were subclassified as SUDC, 11% (16/151) as explained, 7% (10/151) as undetermined, and 3% (4/151) as seizure-related. RESULTS: There were no significant differences between SUDC and explained cases in postnatal, gestational, or postconceptional age, frequency of preterm birth, gender, race, or organ weights. In contrast, 96.7% (117/121) of the SUDC group were discovered during a sleep period compared to 53.3% (8/15) of the explained group (p < 0.001), and 48.8% (59/121) of the SUDC cases had a personal and/or family history of febrile seizures compared to 6.7% (1/15) of the explained group (p < 0.001). Of the explained deaths, 56% (9/16) were subclassified as infection, 31% (5/16) cardiac, 6% (1/16) accidental, and 6% (1/16) metabolic. Two of the three cases specifically tested for cardiac channelopathies at autopsy based upon clinical indications had genetic variants in cardiac genes, one of uncertain significance. Bacterial cultures at autopsy typically revealed organisms interpreted as contaminants. Two of the four seizure-related deaths were witnessed, with two of the brains from these cases showing generalized malformations. Hippocampal anomalies, including a specific combination we termed hippocampal maldevelopment associated with sudden death, were found in almost 50% (40/83) of the SUDC and undetermined cases in which hippocampal sections were available. CONCLUSIONS: This study highlights the key role for the hippocampus, febrile seizures, and sleep in SUDC pathophysiology. It also demonstrates the role of known predisposing conditions such as cardiac channelopathies and infections in causing sudden unexpected death in childhood, and the need for improved ancillary testing and protective strategies in these cases, even when the cause of death is established at autopsy.


Assuntos
Morte Súbita/etiologia , Acidentes/mortalidade , Canalopatias/mortalidade , Criança , Pré-Escolar , Estudos de Coortes , Feminino , Febre/mortalidade , Patologia Legal , Cardiopatias/congênito , Cardiopatias/mortalidade , Hipocampo/anormalidades , Hipocampo/patologia , Humanos , Lactente , Infecções/mortalidade , Masculino , Doenças Metabólicas/mortalidade , Estudos Retrospectivos , Convulsões Febris/mortalidade , Sono
10.
Forensic Sci Med Pathol ; 12(1): 14-25, 2016 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-26782962

RESUMO

PURPOSE: Sudden unexplained death in childhood (SUDC), while rare, accounts for an important fraction of unexpected deaths in children >1 year of age. Previously we reported an association between febrile seizures, hippocampal maldevelopment, and sudden, unexpected deaths in young children (1-6 years), termed "hippocampal maldevelopment associated with sudden death (HMASD)." Here, we characterize in greater detail the hippocampal pathology in a large cohort of cases (n = 42) of this entity, and attempt to define possible new entities responsible for sudden, unexplained death in young children without HMASD/febrile seizure phenotypes. METHODS: We performed comparative analysis on cases, which we classified in a cohort of 89 sudden and unexpected deaths as HMASD, explained deaths, SUDC with febrile seizure phenotype (SUDC-FS) but without hippocampal pathology, and SUDC (without hippocampal pathology or febrile seizure phenotype). RESULTS: The frequency of each subgroup was: HMASD 48% (40/83); SUDC 27% (22/83); SUDC-FS 18% (15/83); explained 7% (6/83). HMASD was characterized clinically by sudden, sleep-related death, term birth, and discovery in the prone position. Key morphologic features of HMASD were focal granule cell bilamination of the dentate gyrus with or without asymmetry and/or malrotation of the hippocampus, associated with significantly increased frequencies of 11 other developmental abnormalities. We identified no other distinct phenotype in the unexplained categories, except for an association of febrile seizures without hippocampal maldevelopment. CONCLUSIONS: HMASD is a distinct clinicopathologic entity characterized by a likely developmental failure of neuronal migration in the dentate gyrus. Future research is needed to determine the causal role of HMASD in sudden death in early childhood.


Assuntos
Morte Súbita/etiologia , Hipocampo/anormalidades , Hipocampo/patologia , Criança , Pré-Escolar , Estudos de Coortes , Giro Denteado/patologia , Feminino , Patologia Legal , Humanos , Lactente , Masculino , Neurônios/patologia , Decúbito Ventral , Estudos Retrospectivos , Sono , Lobo Temporal/patologia , Nascimento a Termo
11.
Health Commun ; 30(6): 598-611, 2015.
Artigo em Inglês | MEDLINE | ID: mdl-25297455

RESUMO

Hope has the potential to be a powerful motivator for influencing behavior. However, hope and messages that evoke hope (hope appeals) have rarely been the focus of theoretical development or empirical research. As a step toward the effective development and use of hope appeals in persuasive communication, this study conceptualized and operationalized hope appeals in the context of climate change prevention. Then, the study manipulated components of the hope evocation part of a hope appeal. Specifically, the components were designed to address appraisals of the importance, goal congruence, future expectation, and possibility of climate protection, resulting in a 2 (strong/weak importance) × 2 (strong/weak goal congruence) × 2 (strong/weak future expectation) × 2 (strong/weak possibility) between-subjects pretest-posttest factorial design. Two hundred forty-five undergraduate students were randomly assigned to one of the 16 message conditions and completed the study online. The study tested whether the four appraisals predict feelings of hope. It determined whether message components that address importance, goal congruence, future expectation, and possibility affect appraisals, feelings of hope, and persuasion outcomes. Finally, this study tested the effects of feelings of hope on persuasion outcomes. This study takes an important step toward enabling the effective use of hope appeals in persuasive communication.


Assuntos
Esperança , Comunicação Persuasiva , Teoria Psicológica , Adolescente , Adulto , Comportamento , Mudança Climática , Cognição , Emoções , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Motivação , Adulto Jovem
12.
Toxicol Res (Camb) ; 13(1): tfae012, 2024 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-38328743

RESUMO

Background: ß-d-N4-Hydroxycytidine (NHC) is the active metabolite of molnupiravir, a broad-spectrum antiviral approved by the MHRA for COVID-19 treatment. NHC induces lethal mutagenesis of the SARS-CoV-2 virus, undergoing incorporation into the viral genome and arresting viral replication. It has previously been reported that several nucleoside analogues elicit off-target inhibition of mitochondrial DNA (mtDNA) or RNA replication. Although NHC does not exert these effects in HepG2 cells, HepaRG are proven to be advantageous over HepG2 for modelling nucleoside analogue-induced mitochondrial dysfunction. Therefore, the objective of this work was to assess the mitotoxic potential of NHC in HepaRG cells, a model more closely resembling physiological human liver. Methods: Differentiated HepaRG cells were exposed to 1-60 µM NHC for 3-14 days to investigate effects of sub-, supra-, and clinically-relevant exposures (in the UK, molnupiravir for COVID-19 is indicated for 5 days and reported Cmax is 16 µM). Following drug incubation, cell viability, mtDNA copy number, mitochondrial protein expression, and mitochondrial respiration were assessed. Results: NHC induced minor decreases in cell viability at clinically relevant exposures, but did not decrease mitochondrial protein expression. The effects on mtDNA were variable, but typically copy number was increased. At supra-clinical concentrations (60 µM), NHC reduced mitochondrial respiration, but did not appear to induce direct electron transport chain dysfunction. Conclusions: Overall, NHC does not cause direct mitochondrial toxicity in HepaRG cells at clinically relevant concentrations, but may induce minor cellular perturbations. As HepaRG cells have increased physiological relevance, these findings provide additional assurance of the mitochondrial safety profile of NHC.

13.
Biomedicines ; 11(5)2023 May 19.
Artigo em Inglês | MEDLINE | ID: mdl-37239154

RESUMO

The United States Food and Drug Administration Adverse Event Reporting System (FAERS) logged 27,140 rhabdomyolysis cases from 2004 to 31 March 2020. We used FAERS to identify 14 drugs frequently reported in 6583 rhabdomyolysis cases and to investigate whether mitochondrial toxicity is a common pathway of drug-induced rhabdomyolysis by these drugs. Preliminary screening for mitochondrial toxicity was performed using the acute metabolic switch assay, which is adapted here for use in murine L6 cells. Fenofibrate, risperidone, pregabalin, propofol, and simvastatin lactone drugs were identified as mitotoxic and underwent further investigation, using real-time respirometry (Seahorse Technology) to provide more detail on the mechanism of mitochondrial-induced toxicity. To confirm the human relevance of the findings, fenofibrate and risperidone were evaluated in primary human skeletal muscle-derived cells (HSKMDC), using the acute metabolic switch assay and real-time respirometry, which confirmed this designation, although the toxic effects on the mitochondria were more pronounced in HSKMDC. Overall, these studies demonstrate that the L6 model of acute modification may find utility as an initial, cost-effective screen for identifying potential myotoxicants with relevance to humans and, importantly, that drug-induced mitochondrial dysfunction may be a common mechanism shared by some drugs that induce myotoxicity.

14.
Elife ; 122023 Jun 06.
Artigo em Inglês | MEDLINE | ID: mdl-37278519

RESUMO

Background: Evidence supports an important link between mitochondrial DNA (mtDNA) variation and adverse drug reactions such as idiosyncratic drug-induced liver injury (iDILI). Here, we describe the generation of HepG2-derived transmitochondrial cybrids, to investigate the impact of mtDNA variation on mitochondrial (dys)function and susceptibility to iDILI. This study created 10 cybrid cell lines, each containing distinct mitochondrial genotypes of haplogroup H or haplogroup J backgrounds. Methods: HepG2 cells were depleted of mtDNA to make rho zero cells, before the introduction of known mitochondrial genotypes using platelets from healthy volunteers (n=10), thus generating 10 transmitochondrial cybrid cell lines. The mitochondrial function of each was assessed at basal state and following treatment with compounds associated with iDILI; flutamide, 2-hydroxyflutamide, and tolcapone, and their less toxic counterparts bicalutamide and entacapone utilizing ATP assays and extracellular flux analysis. Results: Whilst only slight variations in basal mitochondrial function were observed between haplogroups H and J, haplogroup-specific responses were observed to the mitotoxic drugs. Haplogroup J showed increased susceptibility to inhibition by flutamide, 2-hydroxyflutamide, and tolcapone, via effects on selected mitochondrial complexes (I and II), and an uncoupling of the respiratory chain. Conclusions: This study demonstrates that HepG2 transmitochondrial cybrids can be created to contain the mitochondrial genotype of any individual of interest. This provides a practical and reproducible system to investigate the cellular consequences of variation in the mitochondrial genome, against a constant nuclear background. Additionally, the results show that inter-individual variation in mitochondrial haplogroup may be a factor in determining sensitivity to mitochondrial toxicants. Funding: This work was supported by the Centre for Drug Safety Science supported by the Medical Research Council, United Kingdom (Grant Number G0700654); and GlaxoSmithKline as part of an MRC-CASE studentship (grant number MR/L006758/1).


Assuntos
Doença Hepática Induzida por Substâncias e Drogas , Flutamida , Humanos , Flutamida/metabolismo , Flutamida/farmacologia , Tolcapona/metabolismo , Tolcapona/farmacologia , Haplótipos , Mitocôndrias/metabolismo , DNA Mitocondrial/genética , DNA Mitocondrial/metabolismo , Genótipo , Doença Hepática Induzida por Substâncias e Drogas/genética , Doença Hepática Induzida por Substâncias e Drogas/metabolismo
15.
Biomed Pharmacother ; 150: 112999, 2022 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-35461087

RESUMO

SLC2A1 mediates glucose cellular uptake; key to appropriate immune function. Our previous work has shown efavirenz and lopinavir exposure inhibits T cell and macrophage responses, to known agonists, likely via interactions with glucose transporters. Using human cell lines as a model, we assessed glucose uptake and subsequent bioenergetic profiles, linked to immunological responses. Glucose uptake was measured using 2-deoxyglucose as a surrogate for endogenous glucose, using commercially available reagents. mRNA expression of SLC transporters was investigated using qPCR TaqMan™ gene expression assay. Bioenergetic assessment, on THP-1 cells, utilised the Agilent Seahorse XF Mito Stress test. In silico analysis of potential interactions between SLC2A1 and antiretrovirals was investigated using bioinformatic techniques. Efavirenz and lopinavir exposure was associated with significantly lower glucose accumulation, most notably in THP-1 cells (up to 90% lower and 70% lower with efavirenz and lopinavir, respectively). Bioenergetic assessment showed differences in the rate of ATP production (JATP); efavirenz (4 µg/mL), was shown to reduce JATP by 87% whereas lopinavir (10 µg/mL), was shown to increase the overall JATP by 77%. Putative in silico analysis indicated the antiretrovirals, apart from efavirenz, associated with the binding site of highest binding affinity to SLC2A1, similar to that of glucose. Our data suggest a role for efavirenz and lopinavir in the alteration of glucose accumulation with subsequent alteration of bioenergetic profiles, supporting our hypothesis for their inhibitory effect on immune cell activation. Clarification of the implications of this data, for in vivo immunological responses, is now warranted to define possible consequences for these, and similar, therapeutics.


Assuntos
Fármacos Anti-HIV , Infecções por HIV , Trifosfato de Adenosina , Alcinos/uso terapêutico , Fármacos Anti-HIV/farmacologia , Benzoxazinas/farmacologia , Ciclopropanos , Metabolismo Energético , Glucose/uso terapêutico , Transportador de Glucose Tipo 1/genética , Infecções por HIV/tratamento farmacológico , Humanos , Lopinavir/farmacologia , Ritonavir
16.
Front Cardiovasc Med ; 9: 997013, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-36158799

RESUMO

Hyperglycaemia at the time of myocardial infarction has an adverse effect on prognosis irrespective of a prior diagnosis of diabetes, suggesting glucose is the damaging factor. In ex vivo models of ischaemia, we demonstrated that deleterious effects of acutely elevated glucose are PKCα/ß-dependent, and providing PKCα/ß are inhibited, elevated glucose confers cardioprotection. Short pre-treatments with high glucose were used to investigate time-dependent glucose cardiotoxicity, with PKCα/ß inhibition investigated as a potential mechanism to reverse the toxicity. Freshly isolated non-diabetic rat cardiomyocytes were exposed to elevated glucose to investigate the time-dependence toxic effects. High glucose challenge for >7.5 min was cardiotoxic, proarrhythmic and lead to contractile failure, whilst cardiomyocytes exposed to metabolic inhibition following 5-min high glucose, displayed a time-dependent protection lasting ∼15 min. This protection was further enhanced with PKCα/ß inhibition. Cardioprotection was measured as a delay in contractile failure and KATP channel activation, improved contractile and Ca2+ transient recovery and increased cell survival. Finally, the effects of pre-ischaemic treatment with high glucose in a whole-heart coronary ligation protocol, where protection was evident with PKCα/ß inhibition. Selective PKCα/ß inhibition enhances protection suggesting glycaemic control with PKC inhibition as a potential cardioprotective therapeutics in myocardial infarction and elective cardiac surgery.

18.
Front Genet ; 12: 698825, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34484295

RESUMO

Background: The triad of drug efficacy, toxicity and resistance underpins the risk-benefit balance of all therapeutics. The application of pharmacogenomics has the potential to improve the risk-benefit balance of a given therapeutic via the stratification of patient populations based on DNA variants. A growth in the understanding of the particulars of the mitochondrial genome, alongside the availability of techniques for its interrogation has resulted in a growing body of literature examining the impact of mitochondrial DNA (mtDNA) variation upon drug response. Objective: To critically evaluate and summarize the available literature, across a defined period, in a systematic fashion in order to map out the current landscape of the subject area and identify how the field may continue to advance. Methods: A systematic review of the literature published between January 2009 and December 2020 was conducted using the PubMed database with the following key inclusion criteria: reference to specific mtDNA polymorphisms or haplogroups, a core objective to examine associations between mtDNA variants and drug response, and research performed using human subjects or human in vitro models. Results: Review of the literature identified 24 articles reporting an investigation of the association between mtDNA variant(s) and drug efficacy, toxicity or resistance that met the key inclusion criteria. This included 10 articles examining mtDNA variations associated with antiretroviral therapy response, 4 articles examining mtDNA variants associated with anticancer agent response and 4 articles examining mtDNA variants associated with antimicrobial agent response. The remaining articles covered a wide breadth of medications and were therefore grouped together and referred to as "other." Conclusions: Investigation of the impact of mtDNA variation upon drug response has been sporadic to-date. Collective assessment of the associations identified in the articles was inconclusive due to heterogeneous methods and outcomes, limited racial/ethnic groups, lack of replication and inadequate statistical power. There remains a high degree of idiosyncrasy in drug response and this area has the potential to explain variation in drug response in a clinical setting, therefore further research is likely to be of clinical benefit.

19.
Toxicol In Vitro ; 72: 105096, 2021 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-33460737

RESUMO

Inhibition of dihydroorotate dehydrogenase (DHODH), the rate-limiting enzymatic step in de novo pyrimidine synthesis, has broad immunosuppressive effects in vivo and shows promise as a therapeutic target for the treatment of malignancies, viral infections and auto-immune diseases. Whilst there are numerous DHODH inhibitors under development, leflunomide and teriflunomide are the only FDA approved compounds on the market, each of which have been issued with black-box warnings for hepatotoxicity. Mitochondrial dysfunction is a putative mechanism by which teriflunomide and leflunomide elicit their hepatotoxic effects, however it is as yet unclear whether this is shared by other nascent DHODH inhibitors. The present study aimed to evaluate the propensity for DHODH inhibitors to mediate mitochondrial dysfunction in two hepatic in vitro models. Initial comparisons of cytotoxicity and ATP content in HepaRG® cells primed for oxidative metabolism, in tandem with mechanistic evaluations by extracellular flux analysis identified multifactorial toxicity and moderate indications of respiratory chain dysfunction or uncoupling. Further investigations using HepG2 cells, a hepatic line with limited capability for phase I xenobiotic metabolism, identified leflunomide and brequinar as positive mitochondrial toxicants. Taken together, biotransformation of some DHODH inhibitor species may play a role in mediating or masking hepatic mitochondrial liabilities.


Assuntos
Antineoplásicos/toxicidade , Imunossupressores/toxicidade , Fígado/efeitos dos fármacos , Mitocôndrias/efeitos dos fármacos , Oxirredutases atuantes sobre Doadores de Grupo CH-CH/antagonistas & inibidores , Trifosfato de Adenosina/metabolismo , Compostos de Bifenilo/toxicidade , Linhagem Celular , Respiração Celular/efeitos dos fármacos , Crotonatos/toxicidade , Ácidos Dicarboxílicos/toxicidade , Di-Hidro-Orotato Desidrogenase , Humanos , Hidroxibutiratos/toxicidade , Leflunomida/toxicidade , Fígado/metabolismo , Mitocôndrias/metabolismo , Modelos Biológicos , Nitrilas/toxicidade , Salicilanilidas/toxicidade , Toluidinas/toxicidade , Triazóis/toxicidade
20.
Health Commun ; 25(3): 276-85, 2010 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-20461613

RESUMO

Geographic information systems (GIS) for cancer control present an innovative approach to health communication for comprehensive cancer control (CCC) planning. The ability to spatially depict multivariate views of cancer incidence, treatment site locations, transportation routes, and even environmental exposures within a map represents opportunities to involve communities in novel ways with cancer control. Communities may be involved strategically and/or as a goal in planning efforts. The experiences and perceptions of a near census of U.S. CCC program managers (N = 49) were examined to gain their insights about the compatibility of GIS mapping for CCC, the target audiences to be reached with maps as a CCC message, and relative advantages of this technology in its diffusion. Analysis includes a quantitative assessment of interviews and qualitative statements to illustrate these issues. Results suggest that GIS use for cancer control has the potential to build community capacity and social capital for communities as a way to reduce the cancer burden.


Assuntos
Redes Comunitárias/organização & administração , Sistemas de Informação Geográfica , Planejamento em Saúde/organização & administração , Neoplasias/epidemiologia , Vigilância de Evento Sentinela , Conscientização , Censos , Educação em Saúde , Conhecimentos, Atitudes e Prática em Saúde , Humanos , Neoplasias/prevenção & controle , Estados Unidos
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