RESUMO
The human fetal immune system begins to develop early during gestation; however, factors responsible for fetal immune-priming remain elusive. We explored potential exposure to microbial agents in utero and their contribution toward activation of memory T cells in fetal tissues. We profiled microbes across fetal organs using 16S rRNA gene sequencing and detected low but consistent microbial signal in fetal gut, skin, placenta, and lungs in the 2nd trimester of gestation. We identified several live bacterial strains including Staphylococcus and Lactobacillus in fetal tissues, which induced in vitro activation of memory T cells in fetal mesenteric lymph node, supporting the role of microbial exposure in fetal immune-priming. Finally, using SEM and RNA-ISH, we visualized discrete localization of bacteria-like structures and eubacterial-RNA within 14th weeks fetal gut lumen. These findings indicate selective presence of live microbes in fetal organs during the 2nd trimester of gestation and have broader implications toward the establishment of immune competency and priming before birth.
Assuntos
Bactérias/metabolismo , Desenvolvimento Embrionário , Feto/citologia , Feto/microbiologia , Leucócitos/citologia , Adulto , Bactérias/genética , Bactérias/ultraestrutura , Proliferação de Células , Células Dendríticas/metabolismo , Feminino , Feto/ultraestrutura , Trato Gastrointestinal/embriologia , Trato Gastrointestinal/ultraestrutura , Humanos , Memória Imunológica , Ativação Linfocitária/imunologia , Viabilidade Microbiana , Gravidez , Segundo Trimestre da Gravidez , RNA Bacteriano/genética , RNA Ribossômico 16S/genética , Reprodutibilidade dos Testes , Linfócitos T/citologiaRESUMO
We employed scRNA sequencing to extensively characterize the cellular landscape of human liver from development to disease. Analysis of â¼212,000 cells representing human fetal, hepatocellular carcinoma (HCC), and mouse liver revealed remarkable fetal-like reprogramming of the tumor microenvironment. Specifically, the HCC ecosystem displayed features reminiscent of fetal development, including re-emergence of fetal-associated endothelial cells (PLVAP/VEGFR2) and fetal-like (FOLR2) tumor-associated macrophages. In a cross-species comparative analysis, we discovered remarkable similarity between mouse embryonic, fetal-liver, and tumor macrophages. Spatial transcriptomics further revealed a shared onco-fetal ecosystem between fetal liver and HCC. Furthermore, gene regulatory analysis, spatial transcriptomics, and in vitro functional assays implicated VEGF and NOTCH signaling in maintaining onco-fetal ecosystem. Taken together, we report a shared immunosuppressive onco-fetal ecosystem in fetal liver and HCC. Our results unravel a previously unexplored onco-fetal reprogramming of the tumor ecosystem, provide novel targets for therapeutic interventions in HCC, and open avenues for identifying similar paradigms in other cancers and disease.
Assuntos
Carcinoma Hepatocelular/patologia , Células Endoteliais/metabolismo , Microambiente Tumoral/genética , Adulto , Animais , Carcinoma Hepatocelular/genética , Linhagem Celular , Modelos Animais de Doenças , Células Endoteliais/patologia , Feminino , Receptor 2 de Folato/metabolismo , Perfilação da Expressão Gênica/métodos , Humanos , Fígado/patologia , Neoplasias Hepáticas/genética , Macrófagos/metabolismo , Masculino , Proteínas de Membrana/metabolismo , Camundongos , Receptores Notch/genética , Receptores Notch/metabolismo , Transdução de Sinais/genética , Transcriptoma/genética , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/metabolismoRESUMO
The COVID-19 pandemic has devastated the world, despite all efforts in infection control and treatment/vaccine development. Hospitals are currently overcrowded, with health statuses of patients often being hard to gauge. Therefore, methods for determining infection severity need to be developed so that high-risk patients can be prioritized, resources can be efficiently distributed, and fatalities can be prevented. Electrochemical prognostic biosensing of various biomarkers may hold promise in solving these problems as they are low-cost and provide timely results. Therefore, we have reviewed the literature and extracted the most promising biomarkers along with their most favourable electrochemical sensors. The biomarkers discussed in this paper are C-reactive protein (CRP), interleukins (ILs), tumour necrosis factor alpha (TNFα), interferons (IFNs), glutamate, breath pH, lymphocytes, platelets, neutrophils and D-dimer. Metabolic syndrome is also discussed as comorbidity for COVID-19 patients, as it increases infection severity and raises chances of becoming infected. Cannabinoids, especially cannabidiol (CBD), are discussed as a potential adjunct therapy for COVID-19 as their medicinal properties may be desirable in minimizing the neurodegenerative or severe inflammatory damage caused by severe COVID-19 infection. Currently, hospitals are struggling to provide adequate care; thus, point-of-care electrochemical sensor development needs to be prioritized to provide an approximate prognosis for hospital patients. During and following the immediate aftermath of the pandemic, electrochemical sensors can also be integrated into wearable and portable devices to help patients monitor recovery while returning to their daily lives. Beyond the COVID-19 pandemic, these sensors will also prove useful for monitoring inflammation-based diseases such as cancer and cardiovascular disease.
RESUMO
OBJECTIVE: Evaluate performance-based outcomes and return-to-sport rate in National Hockey League (NHL) athletes. DESIGN: Retrospective cohort study. SETTING: Public records. No direct patient care was provided. PARTICIPANTS: National Hockey League athletes who underwent different orthopedic procedures were identified using public records. Three hundred thirty-seven athletes met inclusion criteria. INDEPENDENT VARIABLES: Common orthopedic surgical procedures in NHL athletes. MAIN OUTCOME MEASURES: Return-to-play and preoperative and postoperative performance measures were collected to calculate a position-specific performance score. Short-term and medium-term outcomes were defined as 1 and 2 to 3 seasons after surgery, respectively. RESULTS: Three hundred seven athletes (92.6%) successfully returned to play. The number of games played during the first season after surgery compared with baseline was significantly decreased for hip arthroscopy (HA), noninstability shoulder arthroscopy (ie, shoulder arthroscopy procedure to address pathology other than shoulder instability), knee arthroscopy, and sports hernia repair (P = 0.002, 0.009, 0.03, and 0.01, respectively). The number of games played for seasons 2 and 3 after surgery was significantly decreased for both HA and noninstability shoulder arthroscopy (P = 0.01 and 0.001, respectively). Short-term postoperative performance scores were significantly decreased for HA, noninstability shoulder arthroscopy, and anterior cruciate ligament reconstruction (P = 0.00004, 0.02, and 0.02, respectively) while medium-term scores were significantly decreased for HA only (P = 0.009). CONCLUSIONS: National Hockey League athletes return to play at a high rate after common orthopedic surgeries. However, certain procedures portend poorer performance scores and game participation than others. In particular, HA and noninstability shoulder arthroscopy have the greatest negative effect on NHL careers after surgery.
Assuntos
Desempenho Atlético , Hóquei , Procedimentos Ortopédicos , Volta ao Esporte , Reconstrução do Ligamento Cruzado Anterior , Artroscopia , Atletas , Herniorrafia , Articulação do Quadril/cirurgia , Humanos , Articulação do Joelho/cirurgia , Estudos Retrospectivos , Articulação do Ombro/cirurgiaRESUMO
Correction for 'Glutamate sensing in biofluids: recent advances and research challenges of electrochemical sensors' by Jessica Schultz et al., Analyst, 2020, 145, 321-347. DOI: 10.10.1039/C9AN01609K.
RESUMO
Glutamate is a nonessential amino acid and a putative neurotransmitter. When its consumption exceeds its synthesis, it becomes necessary to monitor its levels. Hence, a low-cost, sensitive and real-time monitoring of glutamate to quantify pain and detect neurodegenerative diseases is imperative to improve pharmacotherapy and early diagnosis for health care. While enzymatic electrochemical sensors are promising to address issues in lab-based detection techniques, non-enzymatic sensors are better due to their higher stability and lower cost. In this review, we aim to discuss the recent advances and remaining challenges of sensing glutamate in biofluids. First, we discuss the metabolic routes of glutamate, followed by its transmission processes to the biofluids. Second, we identify the connection of glutamate to pathologies as a potential biomarker. Third, we emphasize electrochemical sensors instantaneously detect glutamate in biofluids in real-time, quantifying pain and monitoring neurodegenerative diseases. The literature shows the concentration of glutamate in biofluids, such as plasma, cerebral spinal fluid, urine, and saliva are in the range of 5-100 µM, 0.5-2 µM, 8.5 (3.3-18.4) µM mM-1 creatinine, and 0.232 ± 0.177 µM respectively. While these concentration levels are sometimes lower than the detection limit of electrochemical sensors, functionalization of the nanomaterials currently being used such as NiO and Co3O4 with carbon nanotubes or beta-cyclodextrin may improve the sensing performance. Another key challenge in the research is to develop relationships between glutamate and biofluids. Finally, we have to advance electrochemical sensors that are compatible to detect glutamate in physiological conditions for long durations of time.
Assuntos
Técnicas Eletroquímicas/métodos , Ácido Glutâmico/análise , Animais , Biomarcadores/análise , Técnicas Biossensoriais/métodos , Técnicas Eletroquímicas/instrumentação , Eletrodos , Humanos , Neoplasias/diagnóstico , Doenças Neurodegenerativas/diagnóstico , Dor/diagnósticoRESUMO
Cancers in the bone produce a number of severe symptoms including pain that compromises patient functional status, quality of life, and survival. The source of this pain is multifaceted and includes factors secreted from tumor cells. Malignant cells release the neurotransmitter and cell-signaling molecule glutamate via the oxidative stress-related cystine/glutamate antiporter, system xC-, which reciprocally imports cystine for synthesis of glutathione and the cystine/cysteine redox cycle. Pharmacological inhibition of system xC- has shown success in reducing and delaying the onset of cancer pain-related behavior in mouse models. This investigation describes the development of a stable siRNA-induced knockdown of the functional trans-membrane system xC- subunit xCT ( SLC7A11) in the human breast cancer cell line MDA-MB-231. Clones were verified for xCT knockdown at the transcript, protein, and functional levels. RNAseq was performed on a representative clone to comprehensively examine the transcriptional cellular signature in response to xCT knockdown, identifying multiple differentially regulated factors relevant to cancer pain including nerve growth factor, interleukin-1, and colony-stimulating factor-1. Mice were inoculated intrafemorally and recordings of pain-related behaviors including weight bearing, mechanical withdrawal, and limb use were performed. Animals implanted with xCT knockdown cancer cells displayed a delay until the onset of nociceptive behaviors relative to control cells. These results add to the body of evidence suggesting that a reduction in glutamate release from cancers in bone by inhibition of the system xC- transporter may decrease the severe and intractable pain associated with bone metastases.
Assuntos
Sistema y+ de Transporte de Aminoácidos/metabolismo , Neoplasias da Mama/complicações , Dor do Câncer/etiologia , Dor do Câncer/genética , Regulação Neoplásica da Expressão Gênica/fisiologia , Sistema y+ de Transporte de Aminoácidos/genética , Animais , Neoplasias da Mama/genética , Neoplasias da Mama/metabolismo , Proteínas de Ligação ao Cálcio/genética , Proteínas de Ligação ao Cálcio/metabolismo , Isótopos de Carbono/farmacocinética , Carnitina O-Palmitoiltransferase/genética , Carnitina O-Palmitoiltransferase/metabolismo , Linhagem Celular Tumoral , Cistina/farmacocinética , Modelos Animais de Doenças , Feminino , Ácido Glutâmico/metabolismo , Glutationa/metabolismo , Humanos , Interleucina-1/metabolismo , Fator Estimulador de Colônias de Macrófagos/genética , Fator Estimulador de Colônias de Macrófagos/metabolismo , Camundongos Endogâmicos BALB C , Proteínas de Neoplasias/genética , Proteínas de Neoplasias/metabolismo , Fator de Crescimento Neural/genética , Fator de Crescimento Neural/metabolismo , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismoRESUMO
Aggressive breast cancer subtypes utilize system xc-, a membrane antiporter, to import cystine for glutathione synthesis and maintenance of redox homeostasis, in turn releasing glutamate as a metabolic pro-nociceptive by-product. Metastatic breast cancers establish themselves at distal sites including bone, where changes in extracellular glutamate levels contribute to cancer-induced bone pain. We previously established that stearically blocking system xc- activity with sulfasalazine delays the onset of nociceptive behaviours and that xCT, the functional antiporter subunit, is positively regulated by signal transducer and activator of transcription 3 (STAT3). In the current investigation, a murine xenograft cancer-induced bone pain model was applied to examine whether pharmacological inhibition of phosphorylated STAT3 (pSTAT3) induces changes in nociception. A high glutamate-releasing, xCT/pSTAT3 over-expressing human breast cancer cell line was selected for injection into the distal epiphysis of the right femur of female nude mice. A 14-day regimen of intraperitoneal injections with either vehicle or the novel STAT3 inhibitor DR-1-55 commenced three weeks after initial intrafemoral bone injection. Nociceptive behaviours were temporally monitored by automated von Frey, dynamic weight bearing and open-field testing for the duration of the study, beginning at the baseline. Prior to sacrifice and at ethical end point, tumour-induced osteolytic lesions were radiographically assessed. Treatment with DR-1-55 significantly delayed the onset and severity of spontaneous and induced nociceptive behaviours, also decreasing human SLC7A11 ( xCT) mRNA levels in tumour-bearing limbs without altering osteolysis. In addition, two pro-inflammatory cytokines released by this cell line, interleukin 6 and interleukin 1ß, were also down-regulated at the mRNA level in response to DR-1-55 treatment in vivo, with lower human interleukin 6 levels detected in the host circulation. This study demonstrates that targeting pSTAT3 may be a viable therapeutic means to manage cancer-induced bone pain, alone or in combination with stearic system xc- blockers.
Assuntos
Osso e Ossos/patologia , Dor do Câncer/metabolismo , Regulação Neoplásica da Expressão Gênica/fisiologia , Nociceptividade/fisiologia , Fator de Transcrição STAT3/metabolismo , Animais , Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Neoplasias Ósseas/complicações , Neoplasias Ósseas/tratamento farmacológico , Neoplasias da Mama/complicações , Dor do Câncer/etiologia , Dor do Câncer/terapia , Carcinoma/complicações , Linhagem Celular Tumoral/patologia , Citocinas/sangue , Citocinas/genética , Modelos Animais de Doenças , Comportamento Exploratório/efeitos dos fármacos , Comportamento Exploratório/fisiologia , Feminino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Hiperalgesia/tratamento farmacológico , Hiperalgesia/etiologia , Masculino , Camundongos , Camundongos Nus , Nociceptividade/efeitos dos fármacos , Fator de Transcrição STAT3/antagonistas & inibidores , Fator de Transcrição STAT3/genética , Suporte de Carga/fisiologia , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Breast cancer cells release the signalling molecule glutamate via the system xC- antiporter, which is upregulated to exchange extracellular cystine for intracellular glutamate to protect against oxidative stress. Here, we demonstrate that this antiporter is functionally influenced by the actions of the neurotrophin nerve growth factor on its cognate receptor tyrosine kinase, TrkA, and that inhibiting this complex may reduce cancer-induced bone pain via its downstream actions on xCT, the functional subunit of system xC-. We have characterized the effects of the selective TrkA inhibitor AG879 on system xC- activity in murine 4T1 and human MDA-MB-231 mammary carcinoma cells, as well as its effects on nociception in our validated immunocompetent mouse model of cancer-induced bone pain, in which BALB/c mice are intrafemorally inoculated with 4T1 murine carcinoma cells. AG879 decreased functional system xC- activity, as measured by cystine uptake and glutamate release, and inhibited nociceptive and physiologically relevant responses in tumour-bearing animals. Cumulatively, these data suggest that the activation of TrkA by nerve growth factor may have functional implications on system xC--mediated cancer pain. System xC--mediated TrkA activation therefore presents a promising target for therapeutic intervention in cancer pain treatment.
Assuntos
Sistema y+ de Transporte de Aminoácidos/metabolismo , Neoplasias Ósseas/complicações , Dor do Câncer/etiologia , Dor do Câncer/metabolismo , Ácido Glutâmico/metabolismo , Receptor trkA/antagonistas & inibidores , Sistema y+ de Transporte de Aminoácidos/genética , Animais , Comportamento Animal , Contagem de Células , Linhagem Celular Tumoral , Feminino , Fêmur/patologia , Humanos , Camundongos Endogâmicos BALB C , Modelos Biológicos , Fator de Crescimento Neural/farmacologia , Nociceptividade/efeitos dos fármacos , Osteólise/metabolismo , Osteólise/patologia , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Receptor trkA/metabolismo , Tirfostinas/farmacologiaRESUMO
Evidence suggests that there are both nociceptive and neuropathic components of cancer-induced pain. We have observed that changes in intrinsic membrane properties and excitability of normally non-nociceptive Aß sensory neurons are consistent in rat models of peripheral neuropathic pain and cancer-induced pain. This has prompted a comparative investigation of the intracellular electrophysiological characteristics of sensory neurons and of the ultrastructural morphology of the dorsal horn in rat models of neuropathic pain and cancer-induced pain. Neuropathic pain model rats were induced with a polyethylene cuff implanted around a sciatic nerve. Cancer-induced pain model rats were induced with mammary rat metastasis tumour-1 rat breast cancer or MATLyLu rat prostate cancer cells implanted into the distal epiphysis of a femur. Behavioural evidence of nociception was detected using von Frey tactile assessment. Aß-fibre low threshold mechanoreceptor neurons in both cancer-induced pain and neuropathic pain models exhibited slower dynamics of action potential genesis, including a wider action potential duration and lower action potential amplitude compared to those in control animals. Enhanced excitability of Aß-fibre low threshold mechanoreceptor neurons was also observed in cancer-induced pain and neuropathic pain models. Furthermore, both cancer-induced pain and neuropathic pain models showed abundant abnormal axonal sprouting in bundles of myelinated axons in the ipsilateral spinal laminae IV and V. The patterns of changes show consistency between rat models of cancer-induced pain and neuropathic pain. These findings add to the body of evidence that animal models of cancer-induced pain and neuropathic pain share features that may contribute to the peripheral and central sensitization and tactile hypersensitivity in both pain states.
Assuntos
Dor do Câncer/fisiopatologia , Gânglios Espinais/fisiopatologia , Neuralgia/fisiopatologia , Plasticidade Neuronal/fisiologia , Medula Espinal/fisiopatologia , Potenciais de Ação/fisiologia , Animais , Dor do Câncer/patologia , Gânglios Espinais/patologia , Hiperalgesia/fisiopatologia , Neuralgia/patologia , Limiar da Dor/fisiologia , Ratos Sprague-Dawley , Células Receptoras Sensoriais/fisiologiaRESUMO
BACKGROUND: Bone cancer pain is often severe, yet little is known about mechanisms generating this type of chronic pain. While previous studies have identified functional alterations in peripheral sensory neurons that correlate with bone tumours, none has provided direct evidence correlating behavioural nociceptive responses with properties of sensory neurons in an intact bone cancer model. RESULTS: In a rat model of prostate cancer-induced bone pain, we confirmed tactile hypersensitivity using the von Frey test. Subsequently, we recorded intracellularly from dorsal root ganglion neurons in vivo in anesthetized animals. Neurons remained connected to their peripheral receptive terminals and were classified on the basis of action potential properties, responses to dorsal root stimulation, and to mechanical stimulation of the respective peripheral receptive fields. Neurons included C-, Aδ-, and Aß-fibre nociceptors, identified by their expression of substance P. We suggest that bone tumour may induce phenotypic changes in peripheral nociceptors and that these could contribute to bone cancer pain. CONCLUSIONS: This work represents a significant technical and conceptual advance in the study of peripheral nociceptor functions in the development of cancer-induced bone pain. This is the first study to report that changes in sensitivity and excitability of dorsal root ganglion primary afferents directly correspond to mechanical allodynia and hyperalgesia behaviours following prostate cancer cell injection into the femur of rats. Furthermore, our unique combination of techniques has allowed us to follow, in a single neuron, mechanical pain-related behaviours, electrophysiological changes in action potential properties, and dorsal root substance P expression. These data provide a more complete understanding of this unique pain state at the cellular level that may allow for future development of mechanism-based treatments for cancer-induced bone pain.
Assuntos
Neoplasias Ósseas/patologia , Dor do Câncer/patologia , Dor do Câncer/fisiopatologia , Fenômenos Eletrofisiológicos , Nociceptores/patologia , Potenciais de Ação , Animais , Neoplasias Ósseas/complicações , Modelos Animais de Doenças , Gânglios Espinais/patologia , Masculino , Modelos Neurológicos , Fibras Nervosas/patologia , Condução Nervosa , Osteólise/complicações , Osteólise/patologia , Limiar da Dor , Ratos , Fatores de TempoAssuntos
Neoplasias Ósseas/metabolismo , Dor do Câncer/metabolismo , Ácido Glutâmico/metabolismo , Sistema Nervoso Periférico/metabolismo , Células Receptoras Sensoriais/metabolismo , Animais , Comportamento Animal , Neoplasias Ósseas/fisiopatologia , Linhagem Celular Tumoral , Modelos Animais de Doenças , Feminino , Transplante de Neoplasias , Neurônios/metabolismo , Nociceptividade , Manejo da Dor , Medição da Dor , RNA Mensageiro/metabolismo , Ratos , Ratos Sprague-Dawley , Transdução de Sinais , Sulfassalazina/farmacologiaRESUMO
System Xc- is a cystine/glutamate antiporter that contributes to the maintenance of cellular redox balance. The human xCT (SLC7A11) gene encodes the functional subunit of system Xc-. Transcription factors regulating antioxidant defense mechanisms including system Xc- are of therapeutic interest, especially given that aggressive breast cancer cells exhibit increased system Xc- function. This investigation provides evidence that xCT expression is regulated by STAT3 and/or STAT5A, functionally affecting the antiporter in human breast cancer cells. Computationally analyzing two kilobase pairs of the xCT promoter/5' flanking region identified a distal gamma-activated site (GAS) motif, with truncations significantly increasing luciferase reporter activity. Similar transcriptional increases were obtained after treating cells transiently transfected with the full-length xCT promoter construct with STAT3/5 pharmacological inhibitors. Knock-down of STAT3 or STAT5A with siRNAs produced similar results. However, GAS site mutation significantly reduced xCT transcriptional activity, suggesting that STATs may interact with other transcription factors at more proximal promoter sites. STAT3 and STAT5A were bound to the xCT promoter in MDA-MB-231 cells, and binding was disrupted by pre-treatment with STAT inhibitors. Pharmacologically suppressing STAT3/5 activation significantly increased xCT mRNA and protein levels, as well as cystine uptake, glutamate release, and total levels of intracellular glutathione. Our data suggest that STAT proteins negatively regulate basal xCT expression. Blocking STAT3/5-mediated signaling induces an adaptive, compensatory mechanism to protect breast cancer cells from stress, including reactive oxygen species, by up-regulating xCT expression and the function of system Xc-. We propose that targeting system Xc- together with STAT3/5 inhibitors may heighten therapeutic anti-cancer effects.
Assuntos
Neoplasias da Mama/genética , Neoplasias da Mama/metabolismo , Fator de Transcrição STAT3/genética , Fator de Transcrição STAT3/metabolismo , Fator de Transcrição STAT5/genética , Fator de Transcrição STAT5/metabolismo , Transdução de Sinais/genética , Proteínas Supressoras de Tumor/genética , Proteínas Supressoras de Tumor/metabolismo , Linhagem Celular Tumoral , Cistina/metabolismo , Feminino , Ácido Glutâmico/metabolismo , Glutationa/metabolismo , Humanos , Oxirredução , Regiões Promotoras Genéticas/genética , RNA Mensageiro/genética , RNA Interferente Pequeno/genética , Espécies Reativas de Oxigênio/metabolismo , Ativação Transcricional/genética , Regulação para Cima/genéticaRESUMO
BACKGROUND: St John's wort (SJW; Hypericum perforatum) induces CYP3A4 that is involved in the metabolism of the hepatitis C virus (HCV) protease inhibitor boceprevir. Reduced boceprevir exposure and efficacy would contribute to therapeutic failure and increase the risk for resistance development. Boceprevir is co-administered with interferon/ribavirin, and depression has been described frequently in patients undergoing HCV treatment. Patients may purchase over-the-counter herbals to manage depression, and knowing the interaction between SJW and boceprevir is desirable. METHODS: This Phase I, open-label, three-period, cross-over pharmacokinetic study enrolled healthy males and females who, following consent and screening procedures, were randomized to receive SJW on days 1-14, SJW plus boceprevir (SJW on days 22-35 and together on days 31-35) and boceprevir on days 52-56, separated by 7 day washout periods, or the same treatment in the opposite order. Pharmacokinetic sampling was performed at the end of each phase. RESULTS: Seventeen (11 female) subjects completed the study and no serious adverse events were reported. Geometric mean ratios (GMRs) and 90% CIs for boceprevir (with SJW versus alone) AUC(0-8), C(max) and C8 were 0.91 (0.87-0.96), 0.94 (0.82-1.07) and 1.00 (0.79-1.27), respectively. GMRs and 90% CIs for hypericin, the active component of SJW, (with boceprevir versus alone) AUC(0-8), C(max) and C(8) were 1.23 (1.10-1.38), 1.32 (1.16-1.52) and 1.37 (1.19-1.58), respectively. CONCLUSIONS: SJW did not have a clinically significant effect on boceprevir plasma concentrations (or those of its metabolite), suggesting that SJW and boceprevir can be safely co-administered.
Assuntos
Antivirais/farmacocinética , Ativação Enzimática , Hypericum , Extratos Vegetais/farmacocinética , Prolina/análogos & derivados , Adulto , Antivirais/administração & dosagem , Estudos Cross-Over , Interações Medicamentosas , Feminino , Voluntários Saudáveis , Humanos , Masculino , Pessoa de Meia-Idade , Extratos Vegetais/administração & dosagem , Plasma/química , Prolina/administração & dosagem , Prolina/farmacocinéticaRESUMO
BACKGROUND: Liver kinase 1 (LKB1) is an important multi-tasking protein linked with metabolic signaling, also controlling polarity and cytoskeletal rearrangements in diverse cell types including cancer cells. Prolactin (PRL) and Signal transducer and activator of transcription (STAT) proteins have been associated with breast cancer progression. The current investigation examines the effect of PRL and STAT-mediated signaling on the transcriptional regulation of LKB1 expression in human breast cancer cells. METHODS: MDA-MB-231, MCF-7, and T47D human breast cancer cells, and CHO-K1 cells transiently expressing the PRL receptor (long form), were treated with 100 ng/ml of PRL for 24 hours. A LKB1 promoter-luciferase construct and its truncations were used to assess transcriptional changes in response to specific siRNAs or inhibitors targeting Janus activated kinase 2 (JAK2), STAT3, and STAT5A. Real-time PCR and Western blotting were applied to quantify changes in mRNA and protein levels. Electrophoretic mobility shift (EMSA) and chromatin immunoprecipitation (ChIP) assays were used to examine STAT3 and STAT5A binding to the LKB1 promoter. RESULTS: Consistent with increases in mRNA, the LKB1 promoter was up-regulated by PRL in MDA-MB-231 cells, a response that was lost upon distal promoter truncation. A putative GAS element that could provide a STAT binding site mapped to this region, and its mutation decreased PRL-responsiveness. PRL-mediated increases in promoter activity required signaling through STAT3 and STAT5A, also involving JAK2. Both STATs imparted basally repressive effects in MDA-MB-231 cells. PRL increased in vivo binding of STAT3, and more definitively, STAT5A, to the LKB1 promoter region containing the GAS site. In T47D cells, PRL down-regulated LKB1 transcriptional activity, an effect that was reversed upon culture in phenol red-free media. Interleukin 6, a cytokine activating STAT signaling in diverse cell types, also increased LKB1 mRNA levels and promoter activity in MDA-MB-231 cells. CONCLUSIONS: LKB1 is differentially regulated by PRL at the level of transcription in representative human breast cancer cells. Its promoter is targeted by STAT proteins, and the cellular estrogen receptor status may affect PRL-responsiveness. The hormonal and possibly cytokine-mediated control of LKB1 expression is particularly relevant in aggressive breast cancer cells, potentially promoting survival under energetically unfavorable conditions.
Assuntos
Neoplasias da Mama/metabolismo , Proteínas Serina-Treonina Quinases/genética , Fator de Transcrição STAT3/genética , Fator de Transcrição STAT5/genética , Proteínas Supressoras de Tumor/genética , Quinases Proteína-Quinases Ativadas por AMP , Neoplasias da Mama/genética , Neoplasias da Mama/patologia , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Células MCF-7 , Prolactina/metabolismo , Proteínas Serina-Treonina Quinases/biossíntese , Fator de Transcrição STAT3/metabolismo , Fator de Transcrição STAT5/metabolismo , Transdução de Sinais , Proteínas Supressoras de Tumor/metabolismoRESUMO
Cancer pain is a well-documented and prevalent healthcare problem, with current treatment strategies often failing to achieve acceptable efficacy. One of the major difficulties in treating cancer pain owes to the complex interplay between the cancer microenvironment, cancer therapy, and the body's own responses to these biochemical changes. A better understanding of the molecular pathways of nociception that are activated during cancer progression and treatment is necessary for better pain management and increased quality of life. This article reviews the current research that implicates oxidative stress as an important target for attenuating cancer pain. Sources of oxidative stress are first established, followed by a discussion of the various pathways that are affected by oxidative stress and that ultimately cause cancer pain.
Assuntos
Antineoplásicos/efeitos adversos , Antioxidantes/uso terapêutico , Ácido Glutâmico/metabolismo , Neoplasias/complicações , Dor Nociceptiva/etiologia , Estresse Oxidativo , Antineoplásicos/administração & dosagem , Progressão da Doença , Feminino , Humanos , Masculino , Neoplasias/tratamento farmacológico , Neoplasias/metabolismo , Dor Nociceptiva/tratamento farmacológico , Dor Nociceptiva/metabolismo , Manejo da Dor , Qualidade de VidaRESUMO
This article is one of a series of 4 that reports on a task of the NanoRelease Food Additive project of the International Life Science Institute Center for Risk Science Innovation and Application to identify, evaluate, and develop methods that are needed to confidently detect, characterize, and quantify intentionally produced engineered nanomaterials (ENMs) released from food along the alimentary tract. This particular article focuses on the problem of detecting ENMs in food, paying special attention to matrix interferences and how to deal with them. In this review, an in-depth analysis of the literature related to detection of ENMs in complex matrices is presented. The literature review includes discussions of sampling methods, such as centrifugation and ENM extraction. Available analytical methods, as well as emerging methods, are also presented. The article concludes with a summary of findings and an overview of potential knowledge gaps and targets for method development in this area.
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Objective: Nav1.8 expression is restricted to sensory neurons; it was hypothesized that aberrant expression and function of this channel at the site of injury contributed to pathological pain. However, the specific contributions of Nav1.8 to neuropathic pain are not as clear as its role in inflammatory pain. The aim of this study is to understand how Nav1.8 present in peripheral sensory neurons regulate neuronal excitability and induce various electrophysiological features on neuropathic pain. Methods: To study the effect of changes in sodium channel Nav1.8 kinetics, Hodgkin-Huxley type conductance-based models of spiking neurons were constructed using the NEURON v8.2 simulation software. We constructed a single-compartment model of neuronal soma that contained Nav1.8 channels with the ionic mechanisms adapted from some existing small DRG neuron models. We then validated and compared the model with our experimental data from in vivo recordings on soma of small dorsal root ganglion (DRG) sensory neurons in animal models of neuropathic pain (NEP). Results: We show that Nav1.8 is an important parameter for the generation and maintenance of abnormal neuronal electrogenesis and hyperexcitability. The typical increased excitability seen is dominated by a left shift in the steady state of activation of this channel and is further modulated by this channel's maximum conductance and steady state of inactivation. Therefore, modified action potential shape, decreased threshold, and increased repetitive firing of sensory neurons in our neuropathic animal models may be orchestrated by these modulations on Nav1.8. Conclusion: Computational modeling is a novel strategy to understand the generation of chronic pain. In this study, we highlight that changes to the channel functions of Nav1.8 within the small DRG neuron may contribute to neuropathic pain.
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PURPOSE: To evaluate the sagittal alignment of the lumbar spine in patients with degenerative spondylolisthesis at the L4-5 level. METHODS: Patients with untreated degenerative spondylolisthesis at L4-5 were retrospectively identified from the clinical practice of spine surgeons at an academic medical center. All patients had standing X-rays that were reviewed by the senior surgeon to confirm the presence of degenerative spondylolisthesis at L4-5. Radiographs were analyzed for the following: lumbar lordosis (LL), lower lumbar lordosis (L4-S1; LLL), L5-S1 lordosis, pelvic incidence (PI), and pelvic tilt (PT). From these measurements, lumbar distribution index (LLL/LL × 100; LDI), ideal lumbar lordosis (PI × 0.62 + 29; ILL), PI-LL mismatch, and relative lumbar lordosis (LL-ILL; RLL) were calculated. These parameters were used to evaluate the sagittal alignment of the lumbar spine. Normal alignment was defined based on previous studies and clinical experience. RESULTS: 117 participants met inclusion criteria, with an average age of 67.2 years. The majority of the cohort demonstrated hypolordotic sagittal alignment of the L5-S1 segment when assessed in relation to ILL, PI, and LL (73.5%, 61.5%, and 50.4% respectively). Evaluation of the lower lumbar spine (L4-S1) demonstrated normal sagittal alignment when evaluated via LDI and LLL (65%, 52.1%, respectively), suggesting the presence of compensatory hyperextension at L4-5 in response to the L5-S1 hypolordosis. Consequently, normal sagittal alignment of the regional lumbar spine was maintained when evaluated using LL, PI-LL mismatch, and RLL (51.3%, 47%, and 62.4% respectively). CONCLUSIONS: This study demonstrates that there is a high incidence of relative hypolordosis at the L5-S1 level among patients who present with degenerative spondylolisthesis at L4-5. The L5-S1 hypolordosis is associated with L4-5 hyperlordosis, such that the lower lumbar lordosis (L4-S1; LLL) and regional lumbar lordosis (LL) are still within normal range. It is probable that L5-S1 hypolordosis was the initial pathologic event that incited compensatory L4-5 hyperlordosis, which in turn may have led to facet degeneration and laxity, and eventually to development of spondylolisthesis.
Assuntos
Lordose , Espondilolistese , Humanos , Idoso , Espondilolistese/diagnóstico por imagem , Espondilolistese/cirurgia , Lordose/diagnóstico por imagem , Lordose/complicações , Estudos Retrospectivos , Vértebras Lombares/diagnóstico por imagem , Vértebras Lombares/cirurgia , RadiografiaRESUMO
BACKGROUND: Ovarian cancer is characterized by high rates of metastasis and therapeutic resistance. Many chemotherapeutic agents rely on the induction of oxidative stress to cause cancer cell death, thus targeting redox regulation is a promising strategy to overcome drug resistance. METHODS: We have used a tetracycline-inducible Ets-1 overexpression model derived from 2008 ovarian cancer cells in the present study. To examine the role of Ets-1 in glutathione regulation we have measured intracellular reactive oxygen species and glutathione levels, as well as glutathione peroxidase enzyme activity. Glutathione synthesis was limited using transsulfuration or Sx(c)- pathway blocking agents, and glutamate release was measured to confirm Sx(c)- blockade. Cell viability following drug treatment was assessed via crystal violet assay. Oxidative stress was induced through glucose oxidase treatment, which produces hydrogen peroxide by glucose oxidation. The protein expressions of redox-related factors were measured through western blotting. RESULTS: Overexpression of Ets-1 was associated with decreased intracellular ROS, concomitantly with increased intracellular GSH, GPX antioxidant activity, and Sx(c)- transporter activity. Under basal conditions, inhibition of the transsulfuration pathway resulted in decreased GSH levels and GPX activity in all cell lines, whereas inhibition of Sx(c)- by sulfasalazine decreased GPX activity in Ets-1-expressing cells only. However, under oxidative stress the intracellular GSH levels decreased significantly in correlation with increased Ets-1 expression following sulfasalazine treatment. CONCLUSIONS: In this study we have identified a role for proto-oncogene Ets-1 in the regulation of intracellular glutathione levels, and examined the effects of the anti-inflammatory drug sulfasalazine on glutathione depletion using an ovarian cancer cell model. The findings from this study show that Ets-1 mediates enhanced Sx(c)- activity to increase glutathione levels under oxidative stress, suggesting that Ets-1 could be a promising putative target to enhance conventional therapeutic strategies.