RESUMEN
Neuraxial anaesthesia is widely utilised for elective caesarean section, but the prevalence of inadequate intra-operative anaesthesia is unclear. We aimed to determine the prevalence of inadequate neuraxial anaesthesia for elective caesarean section; prevalence of conversion from neuraxial anaesthesia to general anaesthesia following inadequate neuraxial anaesthesia; and the effect of mode of anaesthesia. We searched studies reporting inadequate neuraxial anaesthesia that used ≥ ED95 doses (effective dose in 95% of the population) of neuraxial local anaesthetic agents. Our primary outcome was the prevalence of inadequate neuraxial anaesthesia, defined as the need to convert to general anaesthesia; the need to repeat or abandon a planned primary neuraxial technique following incision; unplanned administration of intra-operative analgesia (excluding sedatives); or unplanned epidural drug supplementation. Fifty-four randomised controlled trials were included (3497 patients). The overall prevalence of requirement for supplemental analgesia or anaesthesia was 14.6% (95%CI 13.3-15.9%); 510 out of 3497 patients. The prevalence of general anaesthesia conversion was 2 out of 3497 patients (0.06% (95%CI 0.0-0.2%)). Spinal/combined spinal-epidural anaesthesia was associated with a lower overall prevalence of inadequate neuraxial anaesthesia than epidural anaesthesia (10.2% (95%CI 9.0-11.4%), 278 out of 2732 patients vs. 30.3% (95%CI 26.5-34.5%), 232 out of 765 patients). Further studies are needed to identify risk factors, optimise detection and management strategies and to determine long-term effects of inadequate neuraxial anaesthesia.
Asunto(s)
Anestesia Epidural , Anestesia Obstétrica , Anestesia Raquidea , Anestesia Epidural/efectos adversos , Anestesia General/efectos adversos , Anestesia Obstétrica/métodos , Anestesia Raquidea/efectos adversos , Cesárea , Femenino , Humanos , EmbarazoRESUMEN
BACKGROUND: This study aims to investigate the relationship between the birth experience and the risk of developing postpartum depression or post-traumatic stress disorder. METHODS: In this prospective, longitudinal, observational study, women were assessed at different time points for depression and post-traumatic stress disorder. The risk of depression or post-traumatic stress disorder based on patient characteristics and specific birth events was assessed within three months postpartum. RESULTS: We enrolled 600 women; 426 were eligible for postpartum assessment. At six weeks and three months postpartum, 15.9% and 12.7% screened positive for depression respectively. Positive post-traumatic stress disorder screenings at six weeks and three months postpartum were 6.2% and 5.1% respectively. Twenty-seven women (8.3%) with a negative screening at six weeks converted to a positive depression or post-traumatic stress disorder screening at three months. A pre-existing history of anxiety or depression was associated with an increased risk of developing depression (aOR 2.12, 95% CI 1.30 to 3.47) and post-traumatic stress (aOR 3.15, 95% CI 1.42 to 7.02) within three months postpartum. The risk of developing post-traumatic stress disorder within three months postpartum was also increased among patients experiencing their first delivery (aOR 2.55, 95% CI 1.10 to 5.88) or operative management of postpartum hemorrhage (aOR 4.44, 95% CI 1.16 to 17.02). CONCLUSION: Depression and post-traumatic stress symptoms either persisted or had new onset at three months postpartum. Mental health screening and postpartum follow-up after six weeks should be considered in high-risk patients who have a history of psychopathology, nulliparity, or undergo operative management of postpartum hemorrhage.
Asunto(s)
Depresión Posparto/epidemiología , Parto/psicología , Trastornos por Estrés Postraumático/epidemiología , Adolescente , Adulto , Depresión Posparto/psicología , Femenino , Humanos , Estudios Longitudinales , Estudios Prospectivos , Trastornos Puerperales/epidemiología , Trastornos Puerperales/psicología , Factores de Riesgo , Trastornos por Estrés Postraumático/psicología , Encuestas y Cuestionarios , Adulto JovenRESUMEN
BACKGROUND: This systematic review and meta-analysis evaluates evidence for seven risk factors associated with failed conversion of labor epidural analgesia to cesarean delivery anesthesia. METHODS: Online scientific literature databases were searched using a strategy which identified observational trials, published between January 1979 and May 2011, which evaluated risk factors for failed conversion of epidural analgesia to anesthesia or documented a failure rate resulting in general anesthesia. RESULTS: 1450 trials were screened, and 13 trials were included for review (n=8628). Three factors increase the risk for failed conversion: an increasing number of clinician-administered boluses during labor (OR=3.2, 95% CI 1.8-5.5), greater urgency for cesarean delivery (OR=40.4, 95% CI 8.8-186), and a non-obstetric anesthesiologist providing care (OR=4.6, 95% CI 1.8-11.5). Insufficient evidence is available to support combined spinal-epidural versus standard epidural techniques, duration of epidural analgesia, cervical dilation at the time of epidural placement, and body mass index or weight as risk factors for failed epidural conversion. CONCLUSION: The risk of failed conversion of labor epidural analgesia to anesthesia is increased with an increasing number of boluses administered during labor, an enhanced urgency for cesarean delivery, and care being provided by a non-obstetric anesthesiologist. Further high-quality studies are needed to evaluate the many potential risk factors associated with failed conversion of labor epidural analgesia to anesthesia for cesarean delivery.
Asunto(s)
Analgesia Epidural/métodos , Analgesia Obstétrica/métodos , Anestesia Epidural/métodos , Anestesia Obstétrica/métodos , Cesárea , Trabajo de Parto , Anestesia General , Femenino , Humanos , Embarazo , Factores de RiesgoRESUMEN
BACKGROUND: Chronic ultraviolet (UV) exposure is a major environmental factor involved in extrinsic skin ageing (photo-ageing). Skin nerve fibres are significantly reduced in number following UV irradiation and new skincare compounds with neuroprotective effects are thus highly warranted. OBJECTIVES: We developed a new skincare formulation from a plant extract and evaluated its neuroprotective effects of ex vivo UV irradiation. MATERIALS AND METHODS: The new skincare emulsion was formulated from Echinacea purpurea extract and was enriched with antioxidants (patent no. PROV020110087075). Skin samples were obtained from 20 healthy patients enrolled for plastic surgery and were immediately treated with placebo (SPF 15) or test emulsions. Skin samples were exposed to UVA and UVB for 60 min. Nerve fibres were identified by immunofluorescence using a monoclonal antibody, anti-human CD56. Cell damage was quantified by image analysis. RESULTS: UVA and UVB significantly reduced (40-60%) densities of nerve endings in control samples treated with placebo (P < 0.001). Samples treated with test emulsion completely blocked UV-related effects on skin nerve endings. These neuroprotective effects were similarly observed regardless of age or tissue analysed (breast versus abdomen). CONCLUSIONS: Our new skincare formulation obtained from E. purpurea provides important neuroprotective effects of UV irradiation and could be used together with SPFs to prevent chronic deleterious effects of solar exposure.
Asunto(s)
Fármacos Neuroprotectores/farmacología , Envejecimiento de la Piel/efectos de los fármacos , Protectores Solares/farmacología , Adulto , Química Farmacéutica , Echinacea , Femenino , Humanos , Técnicas In Vitro , Persona de Mediana Edad , Fibras Nerviosas/efectos de los fármacos , Fibras Nerviosas/patología , Fibras Nerviosas/efectos de la radiación , Extractos Vegetales/farmacología , Piel/efectos de los fármacos , Piel/inervación , Piel/patología , Piel/efectos de la radiación , Envejecimiento de la Piel/efectos de la radiación , Rayos Ultravioleta/efectos adversos , Adulto JovenRESUMEN
BACKGROUND: Asthma is characterized by chronic inflammation of the airways with significant changes in leucocyte trafficking, cellular activation and tissue remodelling. Brain-derived neurotrophic factor (BDNF) has been involved with asthma and allergic diseases but its role as a severity marker in paediatric asthma has not been clinically assessed. OBJECTIVES: To evaluate plasma BDNF and inflammatory markers in order to address their relationships with disease severity in children (6-15 years) with controlled persistent asthma. METHODS: Children with persistent asthma were selected and lung function and skin prick tests were performed in all patients. Plasma BDNF levels and various inflammatory markers (CCL3, CCL11, CCL22, CCL24, CXCL8, CXCL9, CXCL10, soluble TNF receptors) were assessed by ELISAs. RESULTS: Subjects with moderate and severe asthma had higher BDNF levels than mild asthma and controls (P<0.001). The chemokines studied and soluble TNF receptors did not differ between the studied groups. CONCLUSION AND CLINICAL RELEVANCE: Our results indicate BDNF as a potential biomarker for clinical severity in children with asthma.
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Asma/sangre , Asma/fisiopatología , Factor Neurotrófico Derivado del Encéfalo/sangre , Índice de Severidad de la Enfermedad , Adolescente , Biomarcadores/sangre , Quimiocinas CC/sangre , Niño , Humanos , Receptores del Factor de Necrosis Tumoral/sangreRESUMEN
UNLABELLED: Due to an increasing number of skin diseases as a result of exposure to ultraviolet (UV) radiation, it is necessary to evaluate the effectiveness of new skin care formulations with broad-spectrum sunscreens. OBJECTIVES: This study aims to assess the status of nerve fibres in healthy human skin, to quantify effects of UV radiation on nerve endings, and to evaluate neuroprotective effects of new skin care formulations against UV exposure damage. METHODS: Samples were obtained from 34 female patients enrolled for plastic surgery and were immediately treated (10 min) with three emulsions: Cream 1, Cream 2 (placebo) and a sunscreen with sun protection factor 15 (SPF15). Control samples and those treated with the cream emulsions were exposed to UVA and UVB for 60 min. Nerve fibres were identified by immunofluorescence using a monoclonal antibody (anti-human CD56/NCAM). Cell damage was assessed by image analysis. RESULTS: Several cellular nervous structures were identified in the skin samples, including free nerve endings. UVA and UVB significantly decreased (40-60%) density of nerve endings in the control samples and those treated with placebo (Cream 2) or SPF15 (all P < 0.001). Cream 1 completely blocked effects of UV radiation on nerve endings (P > 0.05 vs. control). CONCLUSIONS: Quantification of cell damage induced by UV radiation provides useful information for identification of new skin care compounds with neuroprotective properties.
Asunto(s)
Fibras Nerviosas/efectos de los fármacos , Fibras Nerviosas/efectos de la radiación , Piel/efectos de los fármacos , Piel/efectos de la radiación , Rayos Ultravioleta/efectos adversos , Adulto , Femenino , Técnica del Anticuerpo Fluorescente , Humanos , Persona de Mediana Edad , Fibras Nerviosas/patología , Piel/patología , Protectores Solares/uso terapéutico , Adulto JovenRESUMEN
BACKGROUND: Human T-cell lymphotropic viruses (HTLV)-I/II have a special tropism for infecting T cells and inducing spontaneous lymphocyte proliferation. Leukaemia and neurological manifestations are associated with HTLV-I/II infections, and treatment is usually based on anti-inflammatory drugs including glucocorticoids. Although steroid resistance has been reported, it is unknown whether this condition is related to the infection itself or to the treatment. OBJECTIVE: We investigated whether spontaneous cell proliferation is associated with T-cell sensitivity to glucocorticoids. MATERIALS AND METHODS: Twenty-eight HTLV-I/II patients and 11 healthy age-matched controls took part in this study. Lymphocytes were isolated and cultured in vitro to measure spontaneous and mitogen-induced proliferation as well as cellular sensitivity to dexamethasone. RESULTS: Patients with HTLV-I/II infection showed similar stimulated and unstimulated T-cell proliferation as well as comparable sensitivity to dexamethasone in vitro. There were no group differences in the frequency of glucocorticoid responders versus non-responders. However, T cells of patients with spontaneous proliferation were unresponsive to mitogenic stimulation and were remarkably more resistant to dexamethasone than cells of patients with normal proliferation. CONCLUSION: These data suggest that the poor clinical response to steroids may be associated with spontaneous cell proliferation during HTLV infection.
Asunto(s)
Dexametasona/farmacología , Glucocorticoides/farmacología , Infecciones por HTLV-I/sangre , Infecciones por HTLV-II/sangre , Linfocitos T/efectos de los fármacos , Adolescente , Adulto , Anciano , Estudios de Casos y Controles , Proliferación Celular , Células Cultivadas , Resistencia a Medicamentos , Femenino , Humanos , Activación de Linfocitos , Masculino , Persona de Mediana Edad , Transducción de Señal , Linfocitos T/fisiología , Linfocitos T/virologíaRESUMEN
OBJECTIVE AND DESIGN: Chronic glucocorticoid treatment is associated with pharmacological resistance. We investigated the auxiliary effects of fructose-1,6-bisphosphate (FBP) on dexamethasone (DEX)-related modulation of inflammation and T-cell proliferation. METHODS: Acute inflammation (pleurisy) was induced by injection of carrageenan into the pleural cavity of rats that were treated in vivo with DEX s. c. and FBP i. p. Peripheral blood mononuclear cells were isolated and T-cell sensitivity to FBP and DEX was evaluated in vitro. RESULTS: FBP and DEX reduced the exudate volume, protein concentration and neutrophils in the pleural cavity. However no synergistic effects were observed when these compounds were tested simultaneously. In contrast, both compounds dose-dependently and synergistically suppressed T-cell proliferation. CONCLUSION: These data suggest that FBP may be beneficial as auxiliary drug for the treatment of patients with acquired glucocorticoid resistance.
Asunto(s)
Dexametasona/farmacología , Fructosadifosfatos/farmacología , Inflamación/patología , Linfocitos T/efectos de los fármacos , Animales , Antiinflamatorios/farmacología , Carragenina/farmacología , Proliferación Celular , Relación Dosis-Respuesta a Droga , Femenino , Humanos , Leucocitos Mononucleares/metabolismo , Pleuresia/patología , Ratas , Ratas Wistar , Linfocitos T/metabolismoRESUMEN
Epstein-Barr virus (EBV) infection in vitro causes transformation of B cells and generates B lymphoblastoid cell lines (LCLs). These LCLs have been widely used for the diagnostic of several genetic metabolic disorders. However, up to now, efficiency of LCL generation has been based on misleading subjective analysis. In this study, quantitative analyses have been performed to indicate efficiency of B-cell transformation to measuring human lysosomal acid hydrolases associated with: GM1-gangliosidosis type I, Gaucher disease and mucopolysaccharidosis type I. Peripheral blood mononuclear cells were isolated from 13 subjects, and LCLs were produced by culturing them with EBV for 12 days. Activities of the enzymes beta-galactosidase, beta-glucosidase and alpha-iduronidase were measured before and after cryopreservation in liquid nitrogen for 30 days. Efficiency of the B-cell transformation was screened every 4 days by the enumeration of cell proliferation, cell counts and changes in granularity estimated by flow cytometry. We observed the generation of 13 LCLs. Cell transformation was confirmed by the gradual increase of cellular clusters, cell size and granularity. In addition, we determined that the activity of the enzymes mentioned above did not change following cryopreservation. These data suggest that our enumerative approach for screening of EBV-LCLs is efficient for the enzymatic determination of human lysosomal acid hydrolases and may thus replace misleading subjective analyses.
Asunto(s)
Transformación Celular Viral , Criopreservación , Herpesvirus Humano 4 , Iduronidasa/metabolismo , beta-Galactosidasa/metabolismo , beta-Glucosidasa/metabolismo , Adulto , Linfocitos B/enzimología , Linfocitos B/patología , Linfocitos B/virología , Línea Celular Tumoral , Proliferación Celular , Gangliosidosis GM1/diagnóstico , Gangliosidosis GM1/enzimología , Enfermedad de Gaucher/diagnóstico , Enfermedad de Gaucher/enzimología , Humanos , Recuento de Linfocitos , Lisosomas/enzimología , Mucopolisacaridosis I/diagnóstico , Mucopolisacaridosis I/enzimologíaRESUMEN
Cytokines are key modulators of the immune responses that take place in the inflamed synovium of arthritis patients. Consequently, substances that can reverse the inflammatory profile of the inflamed joint are potential tools for clinical management of the disease. Mycobacterial heat shock protein 70 (MTBHSP70) has been found to protect rats from experimentally induced arthritis through the induction of interleukin (IL)-10-producing T cells. In this study, we have demonstrated that MTBHSP70 induces IL-10 production in synoviocytes from arthritis patients and peripheral blood monoculear cells (PBMCs) from both patients and healthy controls. IL-10 production was accompanied by a decrease in tumour necrosis factor (TNF)-alpha production by synovial cells. Separation studies showed that the target cells were mainly monocytes. Accordingly, we observed that MTBHSP70 delayed maturation of murine bone marrow-derived dendritic cells. Our results suggest that MTBHSP may act on antigen-presenting cells (APCs) to modulate the cytokine response in arthritis and support an anti-inflammatory role for this protein, suggesting that it may be of therapeutic use in the modulation of arthritis.
Asunto(s)
Artritis/inmunología , Citocinas/análisis , Células Dendríticas/inmunología , Proteínas HSP70 de Choque Térmico/inmunología , Interleucina-10/inmunología , Líquido Sinovial/inmunología , Adulto , Animales , Artritis Reactiva/inmunología , Artritis Reumatoide/inmunología , Proteínas Bacterianas , Células Cultivadas , Ensayo de Inmunoadsorción Enzimática/métodos , Femenino , Humanos , Interferón gamma/biosíntesis , Lipopolisacáridos/inmunología , Masculino , Ratones , Ratones Endogámicos C57BL , Persona de Mediana Edad , Monocitos/inmunología , Fitohemaglutininas/inmunología , Linfocitos T/inmunología , Factor de Necrosis Tumoral alfa/biosíntesisRESUMEN
Previous animal research suggests that progesterone may have powerful neuroprotective effects in traumatic brain injury (TBI). This experiment tested the hypothesis that progesterone levels correlate with decreased cerebral edema in male rats with bilateral medial frontal cortex injuries. Three groups of male Sprague-Dawley rats were used: injured given progesterone (4 mg/kg), injured given vehicle (oil), and uninjured controls given vehicle. Progesterone or vehicle was administered intraperitoneally at 1, 6, and 24 h postinjury. At 48 h postinjury, the rats were killed, brains extracted, and assayed for edema. Percent difference in water content of the area surrounding the lesion was compared to posterior cortex. A strong inverse relationship was found between serum progesterone levels and percent cerebral edema; the higher the progesterone levels, the lower the percent edema. Both progesterone and oil-treated animals had some edema compared to sham-operated controls. The brains of the injured animals given control solution were higher in water content than either the uninjured group or injured progesterone-treated rats 48 h postinjury. These findings confirm that progesterone significantly decreases cerebral edema after TBI in adult male subjects.
Asunto(s)
Edema Encefálico/sangre , Edema Encefálico/tratamiento farmacológico , Lesiones Encefálicas/sangre , Lesiones Encefálicas/tratamiento farmacológico , Progesterona/sangre , Progesterona/farmacología , Animales , Masculino , Fármacos Neuroprotectores/sangre , Fármacos Neuroprotectores/farmacología , Ratas , Ratas Sprague-Dawley , Análisis de RegresiónRESUMEN
Acute psychological stress is associated with important changes in circulating cell populations and reductions in cell-mediated immune responses. However, the mechanisms underlying these phenomena are poorly understood. In this study, we investigated (i) acute and chronic restraint stress effects in Sprague-Dawley rats on peripheral lymphocyte subsets and (ii) adhesion molecule (beta2 integrins) expression and (iii) also determined whether glucocorticoids could underlie stress-related changes in cellular redistribution. We observed time-dependent changes in lymphocyte distribution including decreased (-21%) percentages of peripheral T helper cells and increased (88%) NK cell numbers following acute brief restraint. Acute stress was also found to overall upregulate beta2-integrin (CD11a and CD11b) expression on T cells and to raise (1049%) plasma corticosterone levels. However, this stress response was found habituated (-75% vs. acute) in the animals previously exposed to chronic restraint stress. Stress effects on circulating lymphocytes were not observed in animals previously exposed to chronic intermittent restraint stress or chronically stressed animals re-exposed to the same stressor. Our results indicate that 1) stress alters lymphocyte distribution, 2) that adhesion molecules may be involved in stress-induced alterations of T-cell distribution and 3) that these changes may be related to circulating glucocorticoids and subjected to adaptation with repeated stress exposure.
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Antígenos CD18/metabolismo , Subgrupos Linfocitarios/metabolismo , Estrés Psicológico/metabolismo , Enfermedad Aguda , Glándulas Suprarrenales/citología , Glándulas Suprarrenales/fisiología , Animales , Recuento de Células , Enfermedad Crónica , Corticosterona/sangre , Inmunofenotipificación , Células Asesinas Naturales/citología , Antígeno-1 Asociado a Función de Linfocito/biosíntesis , Subgrupos Linfocitarios/citología , Antígeno de Macrófago-1/biosíntesis , Masculino , Tamaño de los Órganos , Ratas , Ratas Sprague-Dawley , Restricción Física/psicología , Linfocitos T Colaboradores-Inductores/citología , Linfocitos T Colaboradores-Inductores/metabolismo , Timo/citología , Timo/fisiología , Factores de Tiempo , Regulación hacia ArribaRESUMEN
Psychological stress has been associated with activation of the hypothalamic-pituitary-adrenal (HPA) axis and impaired cell-mediated immune (CMI) responses. There is also evidence suggesting that intermittent chronic stress differentially alters CMI across different immune compartments, but the mechanisms underlying this phenomenon have not been explored in detail. In the present study, we investigated (i) acute and chronic restraint stress effects in Sprague-Dawley rats on both peripheral blood lymphocyte (PBL) and splenocyte mitogen-induced proliferation and (ii) also determined whether differential stress effects within these immune compartments might reflect alterations in lymphocyte sensitivity to glucocorticoids. It was found that while acute stress exposure significantly raised plasma corticosterone levels (1048% vs. controls, P<.001), this response was attenuated in the animals previously exposed to chronic intermittent stress (-79.66% vs. acute; P<.001). Acute stress increased phytohemagglutinin (PHA)-induced lymphocyte proliferation in the spleen (69.04%, P=.01) and suppressed PBL proliferation (-45.52%, P<.001). Neither of these changes were observed following chronic stress. We also demonstrated that reexposure to the stressor rapidly increased splenocyte sensitivity to in vitro dexamethasone (P<.05) and corticosterone (P<.05) in chronically stressed rats. Our data (1) confirm that acute stress is associated with compartment-specific changes in CMI function, (2) indicate that chronic stress is associated with habituated endocrine and immune responses and (3) that stressor exposure rapidly alters splenocyte sensitivity to glucocorticoids and we suggest that the latter may contribute to differential stress effects across immune compartments.
Asunto(s)
Glucocorticoides/fisiología , Inmunidad/fisiología , Estrés Psicológico/inmunología , Enfermedad Aguda , Glándulas Suprarrenales/patología , Animales , División Celular/fisiología , Enfermedad Crónica , Corticosterona/sangre , Glucocorticoides/farmacología , Linfocitos/efectos de los fármacos , Linfocitos/fisiología , Masculino , Tamaño de los Órganos , Ratas , Ratas Sprague-Dawley , Restricción Física , Estrés Psicológico/patología , Timo/patologíaRESUMEN
In a previous study, Haemophilus ducreyi was found in the pustule and dermis of samples obtained at the clinical end point in the human model of infection. To understand the kinetics of localization, we examined infected sites at 0, 24, and 48 h after inoculation and at the clinical end point. Immediately after inoculation, bacteria were found predominantly in the dermis but also in the epidermis. Few bacteria were detectable at 24 h; however, by 48 h, bacteria were readily seen in the pustule and dermis. H. ducreyi was associated with polymorphonuclear leukocytes and macrophages in the pustule and at its base, but was not associated with T cells, Langerhans' cells, or fibroblasts. H. ducreyi colocalized with collagen and fibrin but not laminin or fibronectin. Association with phagocytes, collagen, and fibrin was seen as early as 48 h and persisted at the pustular stage of disease. Optical sectioning by confocal microscopy and transmission electron microscopy both failed to demonstrate intracellular H. ducreyi. These data identify collagen and fibrin as potentially important targets of adherence in vivo and strongly suggest that H. ducreyi remains extracellular throughout infection and survives by resisting phagocytic killing in vivo.
Asunto(s)
Adhesión Bacteriana , Colágeno/fisiología , Fibrina/fisiología , Haemophilus ducreyi/fisiología , Fagocitos/microbiología , Adulto , Femenino , Humanos , Macrófagos/microbiología , Masculino , Persona de Mediana Edad , Neutrófilos/microbiología , Piel/microbiología , Linfocitos T/microbiologíaRESUMEN
Haemophilus ducreyi expresses a peptidoglycan-associated lipoprotein (PAL) that exhibits extensive homology to Haemophilus influenzae protein 6. We constructed an isogenic PAL mutant (35000HP-SMS4) by the use of a suicide vector that contains lacZ as a counterselectable marker. H. ducreyi 35000HP-SMS4 and its parent, 35000HP, had similar growth rates in broth and similar lipooligosaccharide profiles. 35000HP-SMS4 formed smaller, more transparent colonies than 35000HP and, unlike its parent, was hypersensitive to antibiotics. Complementation of the mutant in trans restored the parental phenotypes. To test whether expression of PAL is required for virulence, nine human volunteers were experimentally infected. Each subject was inoculated with two doses (41 to 89 CFU) of live 35000HP and one dose of heat-killed bacteria on one arm and with three doses (ranging from 28 to 800 CFU) of live 35000HP-SMS4 on the other arm. Papules developed at similar rates at sites inoculated with the mutant or parent but were significantly smaller at mutant-inoculated sites than at parent-inoculated sites. The pustule formation rate was 72% (95% confidence interval [CI], 46.5 to 90.3%) at 18 parent sites and 11% (95% CI, 2.4 to 29.2%) at 27 mutant sites (P < 0.0001). The rates of recovery of H. ducreyi from surface cultures were 8% (n = 130; 95% CI, 4.3 to 14.6%) for parent-inoculated sites and 0% (n = 120; 95% CI, 0.0 to 2.5%) for mutant-inoculated sites (P < 0.001). H. ducreyi was recovered from six of seven biopsied parent-inoculated sites and from one of three biopsied mutant-inoculated sites. Confocal microscopy confirmed that the bacteria present in a mutant inoculation site pustule lacked a PAL-specific epitope. Although biosafety regulations precluded our testing the complemented mutant in humans, these results suggest that expression of PAL facilitates the ability of H. ducreyi to progress to the pustular stage of disease.
Asunto(s)
Proteínas de la Membrana Bacteriana Externa , Infecciones por Haemophilus/etiología , Haemophilus ducreyi/patogenicidad , Lipoproteínas/metabolismo , Peptidoglicano/metabolismo , Proteoglicanos , Adulto , Proteínas de Escherichia coli , Femenino , Haemophilus ducreyi/efectos de los fármacos , Haemophilus ducreyi/genética , Humanos , Antígenos Comunes de Leucocito/análisis , Lipoproteínas/genética , Masculino , Pruebas de Sensibilidad Microbiana , Persona de Mediana Edad , Sistemas de Lectura Abierta , Peptidoglicano/genética , Fenotipo , VirulenciaRESUMEN
To localize Haemophilus ducreyi in vivo, human subjects were experimentally infected with H. ducreyi until they developed a painful pustule or for 14 days. Lesions were biopsied, and biopsy samples were fixed in 4% paraformaldehyde, and cryosectioned. Sections were stained with polyclonal anti-H. ducreyi antiserum or H. ducreyi-specific monoclonal antibodies (MAbs) and fluorescently tagged secondary antibodies and examined by confocal microscopy. We identified H. ducreyi in 16 of 18 pustules but did not detect bacteria in the one papule examined. H. ducreyi was observed as individual cells and in clumps or chains. Staining with MAbs 2D8, 5C9, 3B9, 2C7, and 9D12 demonstrated that H. ducreyi expresses the major pilus subunit, FtpA, the 28-kDa outer membrane protein Hlp, the 18-kDa outer membrane protein PAL, and the major outer membrane protein (MOMP) or OmpA2 in vivo. By dual staining with polyclonal anti-H. ducreyi antiserum and MAbs that recognize human skin components, we observed bacteria within the neutrophilic infiltrates of all positively staining pustules and in the dermis of 10 of 16 pustules. We were unable to detect bacteria associated with keratinocytes in the samples examined. The data suggest that H. ducreyi is found primarily in association with neutrophils and in the dermis at the pustular stage of disease in the human model of infection.
Asunto(s)
Haemophilus ducreyi/aislamiento & purificación , Adulto , Anticuerpos Monoclonales/inmunología , Chancroide/microbiología , Dermis/microbiología , Fluoresceína-5-Isotiocianato/metabolismo , Colorantes Fluorescentes/metabolismo , Humanos , Microscopía Confocal , Microscopía Fluorescente/métodos , Piel/microbiologíaRESUMEN
Haemophilus ducreyi expresses fine tangled pili, which are composed predominantly of a major subunit (FtpA). Confocal microscopy showed that an FtpA-specific monoclonal antibody bound to bacteria in biopsy samples obtained from infected human volunteers. To test the role of pili in pathogenesis, an isogenic mutant (35000HP-SMS1) was constructed by insertionally inactivating ftpA. 35000HP-SMS1 did not express FtpA and was nonpiliated but was otherwise identical to its parent, 35000HP. Seven healthy adults were challenged on the upper arm with the isogenic isolates in a double-blinded, escalating dose-response study. Sites inoculated with the mutant produced papules and pustules at rates similar to the rates observed at sites inoculated with the parent. The recovery rate of H. ducreyi from cultures and the histopathology of biopsy samples obtained from pustules inoculated with 35000HP or 35000HP-SMS1 were similar. Although pili are expressed in vivo, FtpA is not required for pustule formation in the human challenge model.
Asunto(s)
Chancroide/etiología , Fimbrias Bacterianas/fisiología , Haemophilus ducreyi/patogenicidad , Adulto , Método Doble Ciego , Femenino , Haemophilus ducreyi/aislamiento & purificación , Humanos , Mutación , Fenotipo , VirulenciaRESUMEN
Caring for the chronically ill is associated with chronic distress. In view of the adverse effects of distress on cellular immune function, such distress may have implications for health. Indeed, it has been proposed that the hypothalamic-pituitary-adrenal (HPA) axis is a potential psychobiological mediator of these effects. In this study, we observed that elderly caregivers experienced greater distress and increased salivary cortisol than non-caregivers. In addition, caregivers had blunted mitogen-induced lymphocyte proliferation, lower mitogen-induced IL-2 production, and reduced lymphocyte sensitivity to glucocorticoids. These results indicate that chronic distress is associated with impaired cell-mediated immunity which is, in turn, associated with elevated basal steroid levels and altered steroid immunoregulation at the level of the lymphocyte.
Asunto(s)
Cuidadores , Glucocorticoides/farmacología , Activación de Linfocitos/efectos de los fármacos , Estrés Fisiológico/inmunología , Anciano , Enfermedad Crónica , Demencia , Femenino , Humanos , Sistema Hipotálamo-Hipofisario/fisiología , Interleucina-2/biosíntesis , Masculino , Fenómenos Fisiológicos de la Nutrición , Sistema Hipófiso-Suprarrenal/fisiologíaRESUMEN
We developed an enzyme-linked immunosorbent assay-based assay to assess Haemophilus ducreyi binding to extracellular matrix (ECM) proteins. H. ducreyi 35000HP bound to fibronectin, laminin, and type I and III collagen but not to type IV, V, or VI collagen or elastin. Isogenic strains with mutations in ftpA or losB bound as well as the parent, suggesting that neither pili nor full-length lipooligosaccharide is required for H. ducreyi to bind to ECM proteins.
Asunto(s)
Adhesión Bacteriana/fisiología , Proteínas de la Matriz Extracelular/metabolismo , Haemophilus ducreyi/fisiología , Adulto , Adhesión Bacteriana/genética , Colágeno/metabolismo , Ensayo de Inmunoadsorción Enzimática/métodos , Fibronectinas/metabolismo , Fimbrias Bacterianas/genética , Fimbrias Bacterianas/fisiología , Haemophilus ducreyi/genética , Haemophilus ducreyi/patogenicidad , Humanos , Técnicas In Vitro , Laminina/metabolismo , Lipopolisacáridos/metabolismo , Mutación , Unión Proteica , Piel/metabolismo , Piel/microbiologíaRESUMEN
A bactericidal assay was developed in order to test the effect of hyperimmune rabbit sera on the viability of serum-resistant Haemophilus ducreyi 35000HP. Testing of several lots of rabbit complement and time course experiments showed that the serum-sensitive H. ducreyi CIPA77 was killed efficiently by 25% complement at 35 degrees C in 3 h. We hypothesized that incubation of 35000HP under these conditions with the appropriate bactericidal antibody would kill this strain. A panel of high titre rabbit antisera was developed and tested against 35000HP. The panel included antisera raised to whole cells, total membranes, Sarkosyl-insoluble outer membrane proteins, the H. ducreyi lipoprotein, and the peptidoglycan-associated lipoprotein. None of the antisera convincingly showed bactericidal activity. The bactericidal assay was also used to determine the effect of normal human serum (NHS) on isogenic mutants of 35000HP. 35000HP-RSM2, an Omegakan insertion mutant that expresses a truncated lipooligosaccharide, was as resistant to NHS as its parent. A mutant deficient in expression of the major outer membrane protein (35000. 60) was sensitive to NHS. We conclude that 35000HP is relatively resistant to normal and hyperimmune sera, and that the major outer membrane protein contributes to this resistance.