RESUMO
BACKGROUND: Despite the rising incidence of melanoma, medical students have progressively fewer opportunities to encounter patients with this important condition. Curricula tend to attach the greatest value to intellectual forms of learning. However, compared with intellectual learning, experiential learning affords students deep insights about a condition. Doctors who experience ill health are more empathic towards patients. However, opportunities to learn about cancer experientially are limited. Temporary transfer tattoos can simulate the ill health associated with melanoma. We reasoned that if doctors who have been sick are more empathic temporarily 'having' melanoma might have a similar effect. OBJECTIVES: To explore the impact of wearing a melanoma tattoo on medical students' understanding of patienthood and attitudes towards patients with melanoma. METHODS: Ten fourth-year medical students were recruited to a simulation. They wore a melanoma tattoo for 24 h and listened to a patient's account of receiving their diagnosis. Data were captured using audio diaries and face-to-face interviews, transcribed and analysed phenomenologically using the template analysis method. RESULTS: There were four themes: (i) melanoma simulation: opening up new experiences; (ii) drawing upon past experiences; (iii) a transformative introduction to patienthood; (iv) doctors in the making: seeing cancer patients in a new light. CONCLUSIONS: By means of a novel simulation, medical students were introduced to lived experiences of having a melanoma. Such an inexpensive simulation can prompt students to reflect critically on the empathetic care of such patients in the future.
Assuntos
Educação de Graduação em Medicina/métodos , Melanoma/psicologia , Neoplasias Cutâneas/psicologia , Estudantes de Medicina/psicologia , Adulto , Atitude do Pessoal de Saúde , Atitude Frente a Saúde , Feminino , Humanos , Masculino , Relações Médico-Paciente , Treinamento por Simulação , Tatuagem , Adulto JovemRESUMO
OBJECTIVE: To identify genetic associations with severity of radiographic damage in ankylosing spondylitis (AS). METHOD: We studied 1537 AS cases of European descent; all fulfilled the modified New York Criteria. Radiographic severity was assessed from digitised lateral radiographs of the cervical and lumbar spine using the modified Stoke Ankylosing Spondylitis Spinal Score (mSASSS). A two-phase genotyping design was used. In phase 1, 498 single nucleotide polymorphisms (SNPs) were genotyped in 688 cases; these were selected to capture >90% of the common haplotypic variation in the exons, exon-intron boundaries, and 5â kb flanking DNA in the 5' and 3' UTR of 74 genes involved in anabolic or catabolic bone pathways. In phase 2, 15 SNPs exhibiting p<0.05 were genotyped in a further cohort of 830 AS cases; results were analysed both separately and in combination with the discovery phase data. Association was tested by contingency tables after separating the samples into 'mild' and 'severe' groups, defined as the bottom and top 40% by mSASSS, adjusted for gender and disease duration. RESULTS: Experiment-wise association was observed with the SNP rs8092336 (combined OR 0.32, p=1.2×10(-5)), which lies within RANK (receptor activator of NFκB), a gene involved in osteoclastogenesis, and in the interaction between T cells and dendritic cells. Association was also found with the SNP rs1236913 in PTGS1 (prostaglandin-endoperoxide synthase 1, cyclooxygenase 1), giving an OR of 0.53 (p=2.6×10(-3)). There was no observed association between radiographic severity and HLA-B*27. CONCLUSIONS: These findings support roles for bone resorption and prostaglandins pathways in the osteoproliferative changes in AS.
Assuntos
Reabsorção Óssea/genética , Vértebras Cervicais/diagnóstico por imagem , Estudos de Associação Genética , Vértebras Lombares/diagnóstico por imagem , Osteogênese/genética , Espondilite Anquilosante/diagnóstico por imagem , Espondilite Anquilosante/genética , Adulto , Ciclo-Oxigenase 1/genética , Éxons/genética , Feminino , Genótipo , Haplótipos/genética , Humanos , Masculino , Pessoa de Meia-Idade , Polimorfismo de Nucleotídeo Único/genética , Radiografia , Receptor Ativador de Fator Nuclear kappa-B/genética , Índice de Gravidade de DoençaRESUMO
BACKGROUND: Low concentrations of leptin secondary to low body fat or other modulators are thought to be a key signal whereby an energy deficit suppresses the reproductive axis in exercising women resulting in functional hypothalamic amenorrhea (FHA). The purpose of this study was to first examine leptin concentrations in exercising women with and without FHA to address whether there is a threshold concentration of leptin below which reproductive function is suppressed. Secondly, we examined the role of adiposity and other possible modulators of leptin to ascertain whether leptin regulation differs depending on reproductive status. METHODS: This study assessed 50 exercising, premenopausal women (aged 18-30 years) over the course of one menstrual cycle (eumenorrheic women) or one 28-day monitoring period (amenorrheic women). Quantification of daily urinary ovarian steroids and menstrual history were used to determine menstrual status. Body composition was assessed using dual energy X-ray absorptiometry, and leptin was determined by enzyme-linked immunoassay. Key modulators of leptin such as serum insulin concentration, carbohydrate intake, glucose availability, indirect indices of sympathetic nervous activity and other factors were assessed using linear regression. RESULTS: Percentage body fat (%BF) (21.0 ± 1.0 versus 26.8 ± 0.7%; P < 0.001) and leptin concentration (4.8 ± 0.8 versus 9.6 ± 0.9 ng/ml; P < 0.001) were lower in the exercising women with amenorrhea (ExAmen; n = 24) compared with the exercising ovulatory women (ExOvul; n = 26). However, the ranges in leptin were similar for each group (ExAmen: 0.30-16.98 ng/ml; ExOvul: 2.57-18.28 ng/ml), and after adjusting for adiposity the difference in leptin concentration was no longer significant. Significant predictors of log leptin in ExAmen included %BF (ß = 0.826, P < 0.001), log insulin (ß = 0.308, P = 0.012) and log glycerol (ß = 0.258, P = 0.030), but in ExOvul only %BF predicted leptin. CONCLUSIONS These data suggest that leptin concentrations per se are not associated with FHA in exercising women, but the modulation of leptin concentrations may differ depending on reproductive status.
Assuntos
Amenorreia/sangue , Exercício Físico , Leptina/sangue , Ciclo Menstrual/sangue , Absorciometria de Fóton , Adiposidade , Adolescente , Adulto , Amenorreia/epidemiologia , Amenorreia/urina , Glicemia/análise , Estudos Transversais , Carboidratos da Dieta/administração & dosagem , Estrona/análogos & derivados , Estrona/urina , Exercício Físico/fisiologia , Feminino , Glucuronídeos/urina , Glicerol/sangue , Humanos , Insulina/sangue , Hormônio Luteinizante/sangue , Hormônio Luteinizante/urina , Ciclo Menstrual/urina , Pregnanodiol/análogos & derivados , Pregnanodiol/urina , Adulto JovemRESUMO
We have characterized the amino acid sequences of over 20 endogenous peptides bound by a soluble analog of H-2Dd, H-2Dds. Synthetic analogs corresponding to self, viral, tumor, or motif peptides were then tested for their ability to bind to H-2Dd by serologic epitope induction assays using both purified soluble protein and cell surface H-2Dd. The dominant primary sequence motif included glycine at position 2, proline at position 3, and a hydrophobic COOH terminus: leucine, isoleucine, or phenylalanine at position 9 or 10. Ancillary support for high affinity binding was contributed by a positively charged residue at position 5. Three-dimensional computer models of H-2Dds/peptide complexes, based on the crystallographic structure of the human HLA-B27/peptide complex, showed that the basic residue at position 5 was in position to form a salt bridge with aspartic acid at position 156, a polymorphic residue of the H-2Dd heavy (H) chain. Analysis of 28 such models, including 17 based on nonamer self-peptides, revealed considerable variation in the structure of the major histocompatibility complex (MHC) surrounding peptide residue 1, depending on the size and charge of the side chain. Interactions between the side chains of peptide residues 5 and 7, and 6 and 8 commonly occurred. Those peptide positions with limited sequence variability and least solvent accessibility may satisfy structural requirements for high affinity binding of the peptide to the MHC class I H chain, whereas the highly variable positions of the peptide (such as positions 4, 6, and 8) may contribute more to the T cell epitopes.
Assuntos
Antígenos H-2/metabolismo , Peptídeos/imunologia , Sequência de Aminoácidos , Animais , Autoantígenos/química , Simulação por Computador , Cristalografia por Raios X , Antígenos HIV/química , Técnicas In Vitro , Ligantes , Camundongos , Dados de Sequência Molecular , Peptídeos/química , Peptídeos/metabolismo , Ligação Proteica , Conformação Proteica , Estrutura Secundária de Proteína , Estrutura Terciária de Proteína , Relação Estrutura-AtividadeRESUMO
The injection of naked plasmid DNA directly into the muscle cells of mice has been shown to induce potent humoral and cellular immune responses. The generation of a cytotoxic T lymphocyte (CTL) response after plasmid DNA injection may involve the presentation of the expressed antigen in the context of the injected myocytes' endogenous major histocompatibility (MHC)-encoded class I molecules or may use the MHC molecules of bone marrow-derived antigen presenting cells (APC) which are capable of providing co-stimulation as well. To resolve which cell type provides the specific restricting element for this method of vaccination we generated parent-->F1 bone marrow chimeras in which H-2bxd recipient mice received bone marrow that expressed only H-2b or H-2d MHC molecules. These mice were injected intramuscularly with naked plasmid DNA that encoded the nucleoprotein from the A/PR/8/34 influenza strain, which as a single antigen has epitopes for both H-2Db and H-2Kd. The resulting CTL responses were restricted to the MHC haplotype of the bone marrow alone and not to the second haplotype expressed by the recipient's myocytes. The role of somatic tissues that express protein from injected plasmids may be to serve as a reservoir for that antigen which is then transferred to the APC. Consequently, our data show that the mechanism of priming in this novel method for vaccination uses the MHC from bone marrow-derived APC, which are efficient at providing all of the necessary signals for priming the T cell.
Assuntos
Nucleoproteínas/imunologia , Proteínas de Ligação a RNA , Linfócitos T Citotóxicos/imunologia , Vacinação/métodos , Vacinas de DNA/imunologia , Proteínas do Core Viral/imunologia , Vacinas Virais/imunologia , Animais , Células Apresentadoras de Antígenos/imunologia , Medula Óssea/imunologia , Feminino , Antígenos de Histocompatibilidade/imunologia , Injeções Intramusculares , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Proteínas do Nucleocapsídeo , Nucleoproteínas/genética , Plasmídeos/imunologia , Quimera por Radiação , Proteínas do Core Viral/genéticaRESUMO
The majority of T cell responses are restricted to peptide antigens bound by polymorphic major histocompatibility complex (MHC) molecules. However, peptide antigens can be presented to T cells by murine non-MHC-encoded CD1d (mCD1) molecules, and human T cell lines specific for nonpeptide antigens presented on CD1 isoforms have been identified. It is shown here that antigen-specific, mCD1-restricted lymphocytes can be generated in vivo by immunizing mice with a combination of plasmids encoding chicken ovalbumin, murine CD1d, and costimulatory molecules. Splenocytes from immunized mice have CD1d-restricted, MHC- unrestricted, ovalbumin-specific cytolytic activity that can be inhibited by anti-CD1 antibodies as well as a competing CD1-binding peptide. These results suggest a physiologic role for murine CD1d to present exogenous protein antigens.
Assuntos
Antígenos CD1/imunologia , Linfócitos T Citotóxicos/metabolismo , Sequência de Aminoácidos , Animais , Anticorpos/imunologia , Anticorpos/farmacologia , Apresentação de Antígeno/fisiologia , Antígenos CD8/imunologia , Imunização , Complexo Principal de Histocompatibilidade/imunologia , Camundongos , Dados de Sequência Molecular , Ovalbumina/genética , Ovalbumina/imunologia , Peptídeos/imunologia , Peptídeos/farmacologia , Plasmídeos/genética , Plasmídeos/imunologia , Ligação Proteica , Baço/imunologia , Linfócitos T Citotóxicos/imunologia , Transfecção/genéticaRESUMO
To gain insight into the rules that govern the binding of endogenous and viral peptides to a given major histocompatibility complex (MHC) class I molecule, we characterized the amino acid sequences of a set of self peptides bound by a soluble analogue of murine H-2Ld, H-2Lds. We tested corresponding synthetic peptides quantitatively for binding in several different assays, and built three-dimensional computer models of eight peptide/H-2Lds complexes, based on the crystallographic structure of the human HLA-B27/peptide complex. Comparison of primary and tertiary structures of bound self and antigenic peptides revealed that residues 2 and 9 were not only restricted in sequence and tolerant of conservative substitutions, but were spatially constrained in the three-dimensional models. The degree of sequence variability of specific residues in MHC-restricted peptides reflected the lack of structural constraint on those amino acids. Thus, amino acid residues that define a peptide motif represent side chains required or preferred for a close fit with the MHC class I heavy chain.
Assuntos
Antígenos H-2/química , Antígenos H-2/metabolismo , Sequência de Aminoácidos , Animais , Ligação Competitiva , Cromatografia Líquida de Alta Pressão , Epitopos/química , Antígenos H-2/genética , Antígeno de Histocompatibilidade H-2D , Cinética , Complexo Principal de Histocompatibilidade , Camundongos , Modelos Moleculares , Dados de Sequência Molecular , Peptídeos/síntese química , Peptídeos/química , Conformação Proteica , Homologia de Sequência de AminoácidosRESUMO
T cell stimulation by the human immunodeficiency virus 1 gp160-derived peptide p18 presented by H-2Dd class I major histocompatibility complex molecules in a cell-free system was found to require proteolytic cleavage. This extracellular processing was mediated by peptidases present in fetal calf serum. In vitro processing of p18 resulted in a distinct reverse phase high performance liquid chromatography profile, from which a biologically active product was isolated and sequenced. This peptide processing can be specifically blocked by the angiotensin-1 converting enzyme (ACE) inhibitor captopril, and can occur by exposing p18 to purified ACE. The ability of naturally occurring extracellular proteases to convert inactive peptides to T cell antigens has important implications for understanding cytotoxic T lymphocyte responses in vivo, and for rational peptide vaccine design.
Assuntos
Produtos do Gene env/metabolismo , Antígenos H-2/imunologia , HIV-1/imunologia , Antígenos de Histocompatibilidade Classe I/imunologia , Peptidil Dipeptidase A/sangue , Precursores de Proteínas/metabolismo , Sequência de Aminoácidos , Animais , Captopril/farmacologia , Carboxipeptidases/antagonistas & inibidores , Carboxipeptidases/metabolismo , Sistema Livre de Células , Produtos do Gene env/imunologia , Antígenos H-2/genética , Antígenos H-2/isolamento & purificação , Proteína gp160 do Envelope de HIV , Antígeno de Histocompatibilidade H-2D , Antígenos de Histocompatibilidade Classe I/genética , Antígenos de Histocompatibilidade Classe I/isolamento & purificação , Humanos , Cinética , Células L , Camundongos , Dados de Sequência Molecular , Fragmentos de Peptídeos/síntese química , Fragmentos de Peptídeos/metabolismo , Precursores de Proteínas/imunologia , Linfócitos T/imunologia , Transfecção , Microglobulina beta-2/metabolismoRESUMO
OBJECTIVES: The I kappaB kinase (IKK)-related kinase IKKepsilon regulates type I interferon expression and responses as well as proinflammatory mediator production. We examined the role of IKKepsilon in arthritis and its ability to enhance the therapeutic response to systemic interferon (IFN) beta therapy in passive murine K/BxN arthritis. METHODS: IKKepsilon(-/-), IFN alpha(approximately)beta R(-/-) and wild type mice were given K/BxN serum and treated with polyinosinic polycytidylic acid (poly(I:C)), IFN beta, or normal saline. Clinical response and histological scores were assessed. Gene expression in the paws was measured by quantitative PCR. Serum interleukin 1a receptor agonist (IL1Ra) and IL10 were measured by ELISA and multiplex bead array. RESULTS: Arthritis was almost completely blocked in wild type mice if arthritogenic K/BxN serum and the Toll-like receptor (TLR)3 ligand, poly(I:C), were coadministered at the onset of the model, but not in established disease. Mice deficient in IFN alpha(approximately)beta R had an accelerated course of arthritis, and did not respond to poly(I:C). IKKepsilon null mice had a modest decrease in clinical arthritis compared with heterozygous mice. Low doses of IFN beta that were ineffective in wild type mice significantly decreased clinical arthritis in IKKepsilon null mice. Articular chemokine gene expression was reduced in the IKKepsilon(-/-) mice with arthritis and secreted IL1Ra (sIL1Ra) mRNA was significantly increased. Serum levels of IL1Ra were increased in low dose IFN beta-treated IKKepsilon(-/-) mice. CONCLUSIONS: Subtherapeutic doses of IFN beta enhance the anti-inflammatory effects of IKKepsilon deficiency, possibly by increasing production of IL1Ra and unmasking the antichemokine effects. Combination therapy with low dose IFN beta and an IKKepsilon inhibitor might improve efficacy of either agent alone and offers a novel approach to RA.
Assuntos
Antirreumáticos/uso terapêutico , Artrite Experimental/tratamento farmacológico , Quinase I-kappa B/fisiologia , Interferon beta/uso terapêutico , Animais , Artrite Experimental/enzimologia , Artrite Experimental/patologia , Modelos Animais de Doenças , Progressão da Doença , Avaliação Pré-Clínica de Medicamentos/métodos , Quinase I-kappa B/deficiência , Indutores de Interferon/uso terapêutico , Proteína Antagonista do Receptor de Interleucina 1/biossíntese , Proteína Antagonista do Receptor de Interleucina 1/genética , Proteína Antagonista do Receptor de Interleucina 1/fisiologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Poli I-C/uso terapêutico , Proteínas Recombinantes/uso terapêutico , Índice de Gravidade de Doença , Resultado do TratamentoRESUMO
The critical discriminatory event in the activation of T lymphocytes bearing alpha beta T cell receptors (TCRs) is their interaction with a molecular complex consisting of a peptide bound to a major histocompatibility complex (MHC)-encoded class I or class II molecule on the surface of an antigen-presenting cell. The kinetics of binding were measured of a purified TCR to molecular complexes of a purified soluble analog of the murine MHC class I molecule H-2Ld (sH-2Ld) and a synthetic octamer peptide p2CL in a direct, real-time assay based on surface plasmon resonance. The kinetic dissociation rate of the MHC-peptide complex from the TCR was rapid (2.6 x 10(-2) second-1, corresponding to a half-time for dissociation of approximately 27 seconds), and the kinetic association rate was 2.1 x 10(5) M-1 second-1. The equilibrium constant for dissociation was approximately 10(-7) M. These values indicate that TCRs must interact with a multivalent array of MHC-peptide complexes to trigger T cell signaling.
Assuntos
Antígenos H-2/metabolismo , Complexo Principal de Histocompatibilidade , Oligopeptídeos/metabolismo , Receptores de Antígenos de Linfócitos T alfa-beta/metabolismo , Sequência de Aminoácidos , Animais , Técnicas Biossensoriais , Antígeno de Histocompatibilidade H-2D , Cinética , Camundongos , Dados de Sequência Molecular , SolubilidadeRESUMO
Vaccination with naked DNA elicits cellular and humoral immune responses that have a T helper cell type 1 bias. However, plasmid vectors expressing large amounts of gene product do not necessarily induce immune responses to the encoded antigens. Instead, the immunogenicity of plasmid DNA (pDNA) requires short immunostimulatory DNA sequences (ISS) that contain a CpG dinucleotide in a particular base context. Human monocytes transfected with pDNA or double-stranded oligonucleotides containing the ISS, but not those transfected with ISS-deficient pDNA or oligonucleotides, transcribed large amounts of interferon-alpha, interferon-beta, and interleukin-12. Although ISS are necessary for gene vaccination, they down-regulate gene expression and thus may interfere with gene replacement therapy by inducing proinflammatory cytokines.
Assuntos
Resistência a Ampicilina/genética , Formação de Anticorpos , Citocinas/biossíntese , DNA/imunologia , Plasmídeos/imunologia , Vacinação , beta-Galactosidase/imunologia , Sequência de Aminoácidos , Animais , Sequência de Bases , Ilhas de CpG , DNA/química , DNA/genética , Feminino , Regulação da Expressão Gênica , Vetores Genéticos , Humanos , Injeções Intradérmicas , Interferons/biossíntese , Interleucina-12/biossíntese , Camundongos , Camundongos Endogâmicos BALB C , Dados de Sequência Molecular , Monócitos/imunologia , Plasmídeos/genética , Células Th1/imunologia , TransfecçãoRESUMO
OBJECTIVE: Epistaxis is the most common ENT emergency. This study aimed to assess one-year mortality rates in patients admitted to a large teaching hospital. METHOD: This study was a retrospective case note analysis of all patients admitted to the Queen Elizabeth University Hospital in Glasgow with epistaxis over a 12-month period. RESULTS: The one-year overall mortality for a patient admitted with epistaxis was 9.8 per cent. The patients who died were older (mean age 77.2 vs 68.8 years; p = 0.002), had a higher Cumulative Illness Rating Scale-Geriatric score (9.9 vs 6.7; p < 0.001) and had a higher performance status score (2 or higher vs less than 2; p < 0.001). Other risk factors were a low admission haemoglobin level (less than 128 g/dl vs 128 g/dl or higher; p = 0.025), abnormal coagulation (p = 0.004), low albumin (less than 36 g/l vs more than 36 g/l; p < 0.001) and longer length of stay (p = 0.046). CONCLUSION: There are a number of risk factors associated with increased mortality after admission with epistaxis. This information could help with risk stratification of patients at admission and enable the appropriate patient support to be arranged.
Assuntos
Causas de Morte , Epistaxe/diagnóstico , Epistaxe/mortalidade , Mortalidade Hospitalar/tendências , Hospitalização/estatística & dados numéricos , Distribuição por Idade , Idoso , Idoso de 80 Anos ou mais , Estudos de Coortes , Epistaxe/terapia , Feminino , Hospitais Universitários , Humanos , Incidência , Masculino , Pessoa de Meia-Idade , Estudos Retrospectivos , Escócia , Índice de Gravidade de Doença , Distribuição por Sexo , Estatísticas não Paramétricas , Análise de SobrevidaRESUMO
Immunostimulatory DNA sequences (ISS) contain unmethylated CpG dinucleotides within a defined motif. Immunization with ISS-based vaccines has been shown to induce high antigen-specific cytotoxic lymphocyte (CTL) activity and a Th1-biased immune response. We have developed a novel ISS-based vaccine composed of ovalbumin (OVA) chemically conjugated to ISS-oligodeoxynucleotide (ODN). Protein-ISS conjugate (PIC) is more potent in priming CTL activity and Th1-biased immunity than other ISS-based vaccines. Cytotoxic lymphocyte activation by ISS-ODN-based vaccines is preserved in both CD4-/- and MHC class II-/- gene-deficient animals. Furthermore, PIC provides protection against a lethal burden of OVA-expressing tumor cells in a CD8+ cell-dependent manner. These results demonstrate that PIC acts through two unique mechanisms: T-helper-independent activation of CTL and facilitation of exogenous antigen presentation on MHC class I. This technology may have clinical applications in cancer therapy and in stimulating host defense in AIDS and chronic immunosuppression.
Assuntos
Antígenos de Neoplasias/uso terapêutico , Vacinas Anticâncer/uso terapêutico , Neoplasias Experimentais/terapia , Vacinas de DNA/uso terapêutico , Vacinas Sintéticas/uso terapêutico , Animais , Apresentação de Antígeno , Fosfatos de Dinucleosídeos/imunologia , Antígenos de Histocompatibilidade Classe I , Camundongos , Camundongos Mutantes , Linfócitos T Citotóxicos/imunologia , Linfócitos T Auxiliares-Indutores/imunologia , VacinaçãoRESUMO
The blending together of synthetic chemistry with natural product biosynthesis represents a potentially powerful approach to synthesis; to enable this, further synthetic tools and methodologies are needed. To this end, we have explored the first Sonogashira cross-coupling to halotryptophans in water. Broad reaction scope is demonstrated and we have explored the limits of the scope of the reaction. We have demonstrated this methodology to work excellently in the modification of model tripeptides. Furthermore, through precursor directed biosynthesis, we have generated for the first time a new to nature brominated natural product bromo-cystargamide, and demonstrated the applicability of our reaction conditions to modify this novel metabolite.
RESUMO
BACKGROUND: Glucose-6-phosphate isomerase and collagen type II antibody-induced arthritis models (K/BxN and CAIA, respectively) have an inflammatory and a post-inflammatory phase. Both phases display robust tactile allodynia. In previous work, inflammatory phase allodynia was reversed by gabapentin and ketorolac, whereas in late phase only gabapentin was effective. Here, we sought to determine if the effects of these two drugs during the early and late phases of the two arthritis models were observed in the conditioned place preference (CPP) paradigm, indicating a differential drug effect on the aversive state. METHODS: Male C57BL/6 mice received K/BxN serum intraperitoneally, while male BALB/c mice received collagen type II antibody cocktail intravenously. After onset of inflammation and allodynia, we assessed effects of i.p. gabapentin (100 mg/kg) or ketorolac (15 mg/kg) using a CPP paradigm: 2 days adaptation, 2 days conditioning (vehicle in morning and drug in afternoon), preference testing on day 5. RESULTS: Consistent with the effects upon allodynia, both gabapentin and ketorolac produced a preference for the drug-paired compartment in the early phase of the K/BxN model, while gabapentin, but not ketorolac, resulted in a place preference during late phase. In the CAIA model, consistent with differential effects upon allodynia, gabapentin produced a preference in the early phase and a trend in the late phase, whereas ketorolac was ineffective at either time. CONCLUSIONS: CPP validated the aversive state in the inflammatory and post-inflammatory phases of the K/BxN and CAIA arthritis models and correspondence between the anti-hyperpathic pharmacology as defined by thresholds and CPP.
Assuntos
Aminas/uso terapêutico , Analgésicos/uso terapêutico , Artrite/tratamento farmacológico , Ácidos Cicloexanocarboxílicos/uso terapêutico , Hiperalgesia/tratamento farmacológico , Cetorolaco/uso terapêutico , Ácido gama-Aminobutírico/uso terapêutico , Animais , Artrite/etiologia , Modelos Animais de Doenças , Gabapentina , Glucose-6-Fosfato Isomerase , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BLRESUMO
BACKGROUND: Chemotherapeutic agents, such as cisplatin, are known to induce a persistent polyneuropathy. The mechanisms underlying the development of this pain are complex, and have only been investigated rodent models using male animals, despite an equivalent presentation of neuropathy between the sexes, clinically. METHODS: Male and female C57Bl/6, Tlr3(-/-) Tlr4(-/-) , Myd88(-/-) , Trif(lps2) and Myd88(-/-) /Trif(lps2) mice received 6 i.p. injections of cisplatin (2.3 mg/kg/day) every other day over the course of 2 weeks. Changes in tactile threshold were monitored during this time, continuing through day 23, using von Frey filaments. RESULTS: Male WT mice develop a persistent tactile allodynia resulting from cisplatin administration. Female mice develop an initial allodynia, but thresholds return to baseline by day 23. Deletion of TLR3, TLR4, MyD88 and Trif/MyD88 protects animals from the development of cisplatin-induced polyneuropathy, and there are no sex differences. Trif(lps2) male mice show a persistent tactile allodynia following cisplatin administration, while female mice show a reduced allodynia, and remain higher in threshold than their male counterparts. On day 18, animals were given the analgesic gabapentin, and thresholds were tested 45 min after. Gabapentin was effective in transiently reversing mechanical allodynia in those mice with lowered thresholds. CONCLUSIONS: It is important to continue examining both sexes in various pain models, as a mononeuropathy and polyneuropathy show sex differences in pain development and the role of TLR signalling.
Assuntos
Antineoplásicos/toxicidade , Cisplatino/toxicidade , Hiperalgesia/induzido quimicamente , Mononeuropatias/induzido quimicamente , Neuralgia/induzido quimicamente , Limiar da Dor/efeitos dos fármacos , Polineuropatias/induzido quimicamente , Transdução de Sinais/efeitos dos fármacos , Receptores Toll-Like/metabolismo , Aminas/administração & dosagem , Aminas/farmacologia , Analgésicos/administração & dosagem , Analgésicos/farmacologia , Animais , Antineoplásicos/administração & dosagem , Cisplatino/administração & dosagem , Ácidos Cicloexanocarboxílicos/administração & dosagem , Ácidos Cicloexanocarboxílicos/farmacologia , Modelos Animais de Doenças , Feminino , Gabapentina , Hiperalgesia/tratamento farmacológico , Hiperalgesia/etiologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Mononeuropatias/complicações , Neuralgia/tratamento farmacológico , Neuralgia/etiologia , Polineuropatias/complicações , Fatores Sexuais , Ácido gama-Aminobutírico/administração & dosagem , Ácido gama-Aminobutírico/farmacologiaRESUMO
TRPM8 is the molecular sensor for cold; however, the physiological role of TRPM8+ neurons at mucosal surfaces is unclear. Here we evaluated the distribution and peptidergic properties of TRPM8+ fibers in naive and inflamed colons, as well as their role in mucosal inflammation. We found that Trpm8(-/-) mice were hypersusceptible to dextran sodium sulfate (DSS)-induced colitis, and that Trpm8(-/-) CD11c+ DCs (dendritic cells) showed hyperinflammatory responses to toll-like receptor (TLR) stimulation. This was phenocopied in calcitonin gene-related peptide (CGRP) receptor-deficient mice, but not in substance P receptor-deficient mice, suggesting a functional link between TRPM8 and CGRP. The DSS phenotype of CGRP receptor-deficient mice could be adoptively transferred to wild-type (WT) mice, suggesting that CGRP suppresses the colitogenic activity of bone marrow-derived cells. TRPM8+ mucosal fibers expressed CGRP in human and mouse colon. Furthermore, neuronal CGRP contents were increased in colons from naive and DSS-treated Trpm8(-/-) mice, suggesting deficient CGRP release in the absence of TRPM8 triggering. Finally, treatment of Trpm8(-/-) mice with CGRP reversed their hyperinflammatory phenotype. These results suggest that TRPM8 signaling in mucosal sensory neurons is indispensable for the regulation of innate inflammatory responses via the neuropeptide CGRP.
Assuntos
Peptídeo Relacionado com Gene de Calcitonina/imunologia , Colite/imunologia , Imunidade Inata , Mucosa Intestinal/imunologia , Células Receptoras Sensoriais/imunologia , Canais de Cátion TRPM/imunologia , Animais , Peptídeo Relacionado com Gene de Calcitonina/deficiência , Peptídeo Relacionado com Gene de Calcitonina/genética , Colite/induzido quimicamente , Colite/genética , Colite/patologia , Colo/imunologia , Colo/patologia , Células Dendríticas/imunologia , Células Dendríticas/patologia , Sulfato de Dextrana , Células Epiteliais/imunologia , Células Epiteliais/patologia , Feminino , Regulação da Expressão Gênica , Humanos , Imunidade nas Mucosas , Mucosa Intestinal/patologia , Masculino , Camundongos Endogâmicos C57BL , Camundongos Knockout , Fenótipo , Receptores da Neurocinina-1/deficiência , Receptores da Neurocinina-1/genética , Receptores da Neurocinina-1/imunologia , Células Receptoras Sensoriais/patologia , Transdução de Sinais , Canais de Cátion TRPM/deficiência , Canais de Cátion TRPM/genéticaRESUMO
A variety of approaches to antitumor therapy are currently being explored that use both antigen-encoding DNA and noncoding nucleotides as a component of gene vaccination. Among the specific strategies reviewed are a construct that fuses a single-chain variable fragment (scFv) that incorporates both the variable-region genes necessary to encode the idiotypic determinants with fragment C (FrC) of tetanus toxin; a novel vector system using herpes simplex virus 1 (HSV-1) for in vivo gene delivery; the possibility of eliciting hyperacute xenograft response to treat human cancer; and the use of gene gun-mediated granulocyte-macrophage colony-stimulating factor (GM-CSF) cDNA-based tumor cell vaccines. The protection provided by DNA vaccination against viral diseases such as influenza suggested a role for such vaccines against cancer. However, unlike vaccines against infectious diseases, cancer vaccines are therapeutic, rather than prophylactic. With multiple myeloma, for example, it is possible that the optimal timing of administration of such a vaccine is during a remission that has been induced by traditional therapies, to eliminate residual disease. DNA cancer vaccines are designed to activate immune responses to tumor antigens to which the immune system has already been exposed. To do so, the vaccines must first overcome immune tolerance that may have already developed to the tumor. There is increasing evidence that tumor antigens, unlike viral or bacterial antigens, do not consistently activate an immune response. One major factor in determining whether a reaction occurs appears to be whether antigen presentation is accompanied by danger signals. With viral or bacterial infection, the accompanying tissue destruction and inflammation produce costimulatory signals that promote T-cell activation. However, inflammatory and tissue-destructive processes are absent during initial tumor transformation. The typical outcome may be immunologic tolerance.
Assuntos
Mieloma Múltiplo/imunologia , Mieloma Múltiplo/terapia , Vacinas de DNA/imunologia , Terapia Genética , Humanos , Mieloma Múltiplo/genéticaRESUMO
To examine the molecular interactions between major histocompatibility complex (MHC)-encoded molecules and peptides, monoclonal antibodies (mAbs), or T cell receptors, we have developed model systems employing genetically engineered soluble MHC class I molecules (MHC-I), synthetic peptides, purified mAbs, and engineered solubilizable T cell receptors. Direct binding assays based on immobilization of one of the interacting components to the dextran modified gold biosensor surface of a surface plasmon resonance (SPR) detector have been developed for each of these systems. The peptide binding site of the MHC-I molecule can be sterically mapped by evaluation of a set of peptides immobilized through the thiol group of cysteine substitutions at each peptide position. Kinetic binding studies indicate that the MHC-I/peptide interaction is characterized by a low to moderate apparent kass (approximately 5000-60000 M-1 s-1) and very small kdis (approximately 10(-4)-10(-6) s-1) consistent with the biological requirement for a long cell surface residence time to permit engagement with T cell receptors. Several mAb directed against different MHC-I epitopes were examined, and kinetic parameters of their interaction with MHC molecules were determined. These showed characteristic moderate association rate constants and moderate dissociation rate constants (kass approximately 10(4)-10(6) M-1 s-1 and kdis approximately 10(-2)-10(-4) s-1), characteristic of many antibody/protein antigen interactions. The interaction of an anti-idiotypic anti-TCR mAb with its purified cognate TCR was of moderate affinity and revealed kinetic binding similar to that of the anti-MHC mAbs. The previously determined interaction of a purified T cell receptor with its MHC-I/peptide ligand is characterized by kinetic constants more similar to those of the antibody/antigen interaction than of the MHC-I/peptide interaction, but is remarkable for rapid dissociation rates (apparent kdis approximately 10(-2) s-1). Such binding studies of reactions involving the MHC-I molecules offer insight into the mechanisms responsible for the initial specific events required for the stimulation of T cells.