RESUMEN
Sensorineural hearing loss affects one to two children out of 1000 born apparently healthy and 9% approximately of those born with the risk of different pathologies. The origin of premature deafness is unknown in 25% of children whereas it is genetic in some cases. Prevention and early diagnosis, possibly within six to eight months, aim to avoid deafness becoming a cause of disability. The aim of the present study was to establish the optimal preoperative assessment with a diagnostic imaging protocol involving the integrated use of CT and MRI in the selection of the candidates for cochlear implantation. Twenty children were assessed, divided into three different groups: A) those who had CT only; B) those who had only MR; C) those who had both CT and MRI. The purpose was to estimate diagnostic accuracy in preoperative planning and the role of imaging in the diagnostic protocol for children's deafness. The petrous pyramid was studied with a CT Picker PQ 6000 system to high resolution in the axial and coronal planes, and with an MR Intera Philips 0.5 T device by means of acquisition of sequences B-TFE T2 3D and MIP reconstructions on radial coronal plans. This study was completed successfully for morphological brain MRI examination to complete the diagnosis. The following abnormalities were found in six patients (30%): one case of incomplete partition (Mondini malformation); two cases of vestibular aqueduct enlargement; two cases with anomalous jugular bulb positioning; one case with cochlear ossification. In the remaining 70%: eight patients had no anatomical anomalies; CT and MRI imaging were normal in six patients with minor abnormalities disclosed at surgery (one case of the stapedial artery emerging from the promontory); three anatomical variants of the round window, and two abnormal course of the facial nerve). We emphasize the importance of integrated CT and MRI imaging in the study of children with sensorineural hearing loss. The combination of CT and MRI has been shown to be superior to either modality used alone in view of "risk-free" cochear implantation. High resolution computed tomography and magnetic resonance images obtained by B-TFE T2-weighted 3D sequences help the surgeon in planning the operation and predict operative difficulty and potential complications in paediatric cochlear implant candidates.
RESUMEN
Systems for the staging of individuals with human immunodeficiency virus type 1 (HIV-1) infection were developed 15 years ago. Subsequently, assays for quantitating HIV-1 RNA and immunophenotyping of lymphocyte subsets have been developed and validated. The utility of these assays for improved staging in early disease was evaluated in 256 HIV-infected adults (52% minority) with CD4 counts > or = 400 cells/microL followed in U.S. military medical centers before the highly active anti-retroviral therapy era. HIV viral load (RNA) was quantitated; the frequencies of select CD4+ immunophenotypes were determined in 112 subjects. The results were analyzed in relation to three outcome measures: death, first acquired immunodeficiency syndrome-defining opportunistic infection, and CD4 count < or = 200 cells/microL. Serum RNA level and CD4 count were each found to be predictive of all three outcomes. In addition, increases in the T-cell subsets CD28-CD4+ and CD29+CD26-CD4+ were found to be independently predictive of more rapid progression. The classification of early-stage HIV patients is improved by the quantitation of both viral RNA and T-lymphocyte subsets.
Asunto(s)
Infecciones por VIH/inmunología , VIH-1 , ARN Viral/sangre , Subgrupos de Linfocitos T , Adulto , Distribución de Chi-Cuadrado , Progresión de la Enfermedad , Femenino , Humanos , Recuento de Linfocitos , Masculino , Evaluación de Resultado en la Atención de Salud , Pronóstico , Modelos de Riesgos Proporcionales , Estadísticas no Paramétricas , Análisis de SupervivenciaRESUMEN
To aid our investigation of tubulin as an antileishmanial drug target, the effects of the mammalian antimicrotubule agents ansamitocin P3, taxol, and hemiasterlin on Leishmania donovani promastigotes were described. These drugs affected the assembly of purified leishmanial tubulin and inhibited the growth of L. donovani promastigotes at micromolar concentrations. When promastigotes were treated with these agents, mitotic partitioning of nuclear DNA and cytokinesis were usually inhibited. The spatial orientation of kinetoplasts was often disturbed, suggesting a role for microtubules in the segregation of these organelles during mitosis. Aberrant cell types produced in drug-treated cultures included parasites with one nucleus and two geometrically distinct kinetoplasts, parasites with multiple kinetoplasts, and cytoplasts containing a kinetoplast but no nucleus. A subset of unique cell types, parasites containing two nuclei, a spindle fiber, and two geometrically distinct kinetoplasts, were observed in hemiasterlin-treated cultures. Flow cytometric analysis of L. donovani promastigotes treated with these three drugs indicated a dramatic shift toward the G2 + M phase of the cell cycle, with some cells containing four times the amount of DNA present in G1. These results were used to evaluate the cellular effects of WR85915, an aromatic thiocyanate with in vitro antileishmanial and anti-tubulin activity, on L. donovani. Treatment of parasites with WR85915 did not produce the unusual cell types described above and did not cause the accumulation of parasites in G2 + M, suggesting that WR85915 acts on target(s) in Leishmania in addition to tubulin. These studies validate tubulin as a suitable antileishmanial drug target and provide criteria to assess the cellular mechanism of action of new candidate antileishmanial agents.
Asunto(s)
Leishmania donovani/efectos de los fármacos , Maitansina/análogos & derivados , Maitansina/farmacología , Oligopéptidos/farmacología , Oxadiazoles/farmacología , Paclitaxel/farmacología , Tubulina (Proteína)/metabolismo , Animales , Antiprotozoarios/farmacología , Ciclo Celular/efectos de los fármacos , ADN Protozoario/análisis , Citometría de Flujo , Leishmania donovani/crecimiento & desarrollo , Leishmania donovani/ultraestructura , Microscopía Electrónica , Microscopía FluorescenteRESUMEN
Signals generated through CD28-B7 and CD40 ligand (CD40L)-CD40 interactions have been shown to be crucial for the induction of long-term allograft survivability. We have recently demonstrated that humanized anti-CD40L (hu5C8) prevents rejection of mismatched renal allografts in primates. To investigate potential mechanisms of CD40L-induced allograft acceptance, we coimmobilized hu5C8 with suboptimal amounts of anti-CD3 to stimulate CD4(+) T cells. We now report that anti-CD3/CD40L costimulation results in CD28-independent activation and subsequent deletion of resting T cells. Coligation of CD3 and CD40L increased expression of CD69, CD25, and CD54 on CD4(+) T cells. We also found that costimulation with anti-CD3/CD40L resulted in enhanced production of interleukin (IL)-10, interferon gamma, and tumor necrosis factor alpha but not IL-2 or IL-6. Interestingly, after several days, anti-CD3/CD40L-mediated activation was followed by apoptosis in a significant population of cells. Consistent with that observation, anti-CD3/CD40L did not enhance the antiapoptotic proteins Bcl-2 and Bcl-xL. Further, the addition of CD28 at 24 h failed to rescue those cells induced to die after costimulation with anti-CD3/CD40L. Together, these data suggest that the graft-sparing effect of hu5C8 in vivo may result in part from early and direct effects on CD4(+) T cells, including a vigorous induction of immunomodulatory cytokines and/or apoptosis of allograft-specific T cells.
Asunto(s)
Apoptosis , Linfocitos T CD4-Positivos/inmunología , Antígenos CD40/fisiología , Citocinas/biosíntesis , Glicoproteínas de Membrana/fisiología , Antígenos CD/análisis , Antígenos CD/fisiología , Linfocitos T CD4-Positivos/citología , Linfocitos T CD4-Positivos/efectos de los fármacos , Antígenos CD40/genética , Ligando de CD40 , Células Cultivadas , Citocinas/genética , Regulación de la Expresión Génica , Humanos , Inmunoglobulina G/farmacología , Interferón gamma/biosíntesis , Interferón gamma/genética , Interleucinas/biosíntesis , Interleucinas/genética , Activación de Linfocitos , Complejo Mayor de Histocompatibilidad , Glicoproteínas de Membrana/inmunología , Reacción en Cadena de la Polimerasa , Proteínas Recombinantes/metabolismo , Transducción de Señal , Transfección , Células Tumorales CultivadasRESUMEN
Infection by the human immunodeficiency virus type 1 (HIV-1) is known to cause a number of changes in the immunophenotypic profile of patients even in the early asymptomatic stages of disease. Such "surrogate markers" are known to correlate with the stage of HIV disease and often are predictive of outcomes. The best known of these is the absolute count of T helper lymphocytes, or CD4 cells, which undergoes a gradual decline as the virus infects greater and greater numbers of these cells (1). A number of other markers have been found, some of which also are predictive of outcome in many cases. These include prevalence and intensity of the CD38 marker on CD8 T cells, percentage of CD4 cells exhibiting loss of the CD26 and CD28 markers, and percentage of CD4 cells with the CD95 marker (2). The CD38 intensity of CD3/CD8 cells has, in fact, been more closely correlated with future disease progression in patients than the CD4 count (3). The following method is a comprehensive immunophenotyping panel that incorporates all of these markers and provides several parameters by which to monitor disease progression and advise clinicians on treatment options.
RESUMEN
Cells within an organism undergo two common forms of cell death. Sudden injury resulting from physical or chemical insult leads to a form of cell death called necrosis. A more subtle programmed form of cell death is termed apoptosis. Apoptosis describes a genetically encoded pathway that plays an important role in regulating the immune response (1,2). Apoptotic cell death is characterized by distinct biochemical and morphologic changes and the fragmentation of DNA into nucleosomal-sized multimers (3). Apoptosis plays a crucial role in viral infections and in the host response to viral insult (4).
RESUMEN
Previous studies have revealed that the expression of CD38 on CD8+ T cells is a strong predictor of disease progression in human immunodeficiency virus (HIV)-infected individuals. Those studies were performed using fresh patient samples over an extended trial period. After demonstrating the validity of assay results on cryopreserved cells, we performed a retrospective study using frozen cell samples to determine the predictive value of CD38 expression in patients with CD4 counts above 400 cells/microl. The CD38 expression as measured by antibody binding capacity and the CD38 median channel were shown to be associated with time to new opportunistic infection or death (both P < 0.001). These results suggest that CD38 expression on CD8+ T cells, whether fresh or frozen, provides a useful predictor of HIV disease progression.
Asunto(s)
Antígenos CD , Antígenos de Diferenciación/análisis , Linfocitos T CD8-positivos/química , Infecciones por VIH/sangre , NAD+ Nucleosidasa/análisis , ADP-Ribosil Ciclasa , ADP-Ribosil Ciclasa 1 , Adulto , Biomarcadores , Conservación de la Sangre , Criopreservación , Progresión de la Enfermedad , Femenino , Infecciones por VIH/patología , Humanos , Masculino , Glicoproteínas de Membrana , Persona de Mediana EdadRESUMEN
We have assessed the functional effects of a panel of CTLA-4 mAbs on resting human CD4+ T cells. Our results demonstrate that some CTLA-4 mAbs can inhibit proliferative responses of resting CD4+ cells and cell cycle transition from G0 to G1. The inhibitory effects of CTLA-4 were evident within 4 h, at a time when cell surface CTLA-4 expression remained undetectable. Other CTLA-4 mAbs had no detectable inhibitory effects, indicating that binding of Ab to CTLA-4 alone is not sufficient to mediate down-regulation of T cell responses. Interestingly, while IL-2 production was shut off, inhibitory anti-CTLA-4 mAbs permitted induction and expression of the cell survival gene bcl-X(L). Consistent with this observation, cells remained viable and apoptosis was not detected after CTLA-4 ligation.
Asunto(s)
Antígenos de Diferenciación/fisiología , Linfocitos T CD4-Positivos/fisiología , Inmunoconjugados , Interleucina-2/metabolismo , Proteínas Proto-Oncogénicas c-bcl-2/fisiología , Abatacept , Anticuerpos Monoclonales , Antígenos CD , Antígeno CTLA-4 , Ciclo Celular , Células Cultivadas , Expresión Génica , Humanos , Activación de Linfocitos , ARN Mensajero/genética , Transducción de Señal , Proteína bcl-XRESUMEN
The International Society of Analytical Cytology (ISAC) Biohazard Working Group presents guidelines for sorting of unfixed cells, including known biohazardous samples, using jet-in-air, deflected-droplet cell sorters. There is a risk that personnel operating these instruments could become exposed to droplets and aerosols containing biological agents present in the samples. The following guidelines can aid in the prevention of exposures of laboratory personnel to pathogens contained in the sort samples. The document provides biosafety recommendations for sample handling, operator training and protection, laboratory facility design, and instrument setup and maintenance. In addition, it describes in detail methods for assessment of instrument aerosol containment. Recommendations provided here may also help laboratories to obtain institutional and/or regulatory agency approval for sorting of unfixed and known biohazardous samples.
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Separación Celular , Contención de Riesgos Biológicos , Guías como Asunto , Administración de la Seguridad , Citometría de Flujo , Humanos , Fijación del TejidoRESUMEN
Progression to AIDS in asymptomatic HIV-infected individuals is characterized by a gradual but progressive loss of CD4+ T cells. While the mechanisms underlying this decline are currently unknown, recent evidence suggests that these cells are abnormally sensitive to apoptosis in response to activation signals. Recent work has implicated downregulation of Bcl-2 with the increased spontaneous apoptosis in lymphocytes from HIV-infected patients. We have evaluated the roles of the apoptosis-protective proteins Bcl-2 and Bcl-x in stimulated PBMC from asymptomatic HIV-infected and HIV-uninfected individuals. We found that Bcl-2 was constitutively expressed in PBMC from both HIV-infected and uninfected samples. However, Bcl-x induction was delayed and responses were decreased in stimulated HIV-infected samples. Additionally, single-cell intracellular staining of Bcl-x revealed a significant inverse correlation between PWM-induced Bcl-x expression and apoptosis (r = -0.695, P = 0.05). This was confirmed at the single-cell level in direct experiments when stimulated cells were sorted based on Bcl-x induction and then measured for apoptosis. Furthermore, low Bcl-x expression was not due to reduced lymphocyte activation following PWM stimulation. Our data indicate that the induction of Bcl-x is markedly impaired in asymptomatic HIV-infected patients and that stimuli which induce inadequate expression of Bcl-x are associated with increased levels of apoptosis in these cells.
Asunto(s)
Apoptosis/inmunología , Infecciones por VIH/inmunología , Leucocitos Mononucleares/metabolismo , Proteínas Proto-Oncogénicas c-bcl-2/biosíntesis , Antígenos CD/biosíntesis , Antígenos de Diferenciación de Linfocitos T/biosíntesis , Apoptosis/efectos de los fármacos , Femenino , Infecciones por VIH/metabolismo , Humanos , Células Jurkat , Lectinas Tipo C , Leucocitos Mononucleares/inmunología , Activación de Linfocitos/efectos de los fármacos , Masculino , Mitógenos de Phytolacca americana/farmacología , Proteínas Proto-Oncogénicas c-bcl-2/efectos de los fármacos , Proteína bcl-XRESUMEN
Peripheral blood mononuclear cells from many asymptomatic HIV-infected patients exhibit defects in cytokine production and impaired proliferative responses in vitro but the mechanisms underlying this subclinical immune deficiency are controversial. To determine whether abnormalities in the earliest events following receptor engagement may help to explain the in vitro immune dysfunction, we measured the inducibility of the early activation marker CD69 in T cells from asymptomatic HIV-infected individuals in response to stimulation with anti-CD2 or anti-CD3 mAb. In a whole blood assay, we found that induction of CD69 was markedly impaired in CD4+ T cells from later-stage HIV-infected patients (CD4 counts 200-400/mm3) compared to uninfected controls. Among early stage patients (CD4 > 400/mm3), a subset (29%) had impaired CD69 induction. CD69 responses were equally depressed after stimulation through the CD3 or CD2 receptor pathways. Survey of a panel of immunophenotypic markers and propensity for apoptosis demonstrated a significant association between depressed induction of CD69 and decreased percentages of CD4+CD26+ and CD4+CD95+ cells but no association with the level of apoptosis. These data indicate that defects in T lymphocyte activation through CD3 and CD2 can be measured within hours of receptor stimulation in asymptomatic HIV-infected individuals and might be useful to monitor as an indicator of immune function in these patients.
Asunto(s)
Antígenos CD/biosíntesis , Antígenos de Diferenciación de Linfocitos T/biosíntesis , Linfocitos T CD4-Positivos/inmunología , Infecciones por VIH/inmunología , Biomarcadores , Infecciones por VIH/sangre , Humanos , Lectinas Tipo C , FenotipoRESUMEN
The susceptibility of highly purified human CD34+ cells to monocytotropic (Ba-L) and lymphotropic (A018-post) strains of human immunodeficiency virus-1 (HIV-1) was examined. Liquid cultures initiated with fresh immunomagnetically purified CD34+ cells using the K6.1 CD34 monoclonal antibody (MoAb) (K6.1/CD34+) were positive for HIV expression 2 weeks after exposure to HIV-1 Ba-L. These cells were initially greater than 90% CD34+ and had undetectable monocyte contamination by flow-cytometric staining and side-scatter analyses, respectively, and undetectable T-cell contamination by CD3 polymerase chain reaction (PCR) analysis. However, secondary CD34+ liquid cultures reselected from the primary liquid cultures 24 hours after HIV exposure by panning with the ICH3 CD34 MoAb (ICH3/CD34+) and maintained for an additional 14 days were negative for HIV expression. The ICH3-unbound cells were positive for both spliced and unspliced HIV RNA when exposed to HIV-1 Ba-L, and were DNA PCR positive when exposed to either monocytotropic or lymphotropic HIV-1. To further test that CD34+ cells were not infectible by HIV-1, we exposed K6.1/CD34+ cells continuously to HIV-1 in a culture system capable of maintaining and expanding primitive CD34+ cells. HIV-exposed K6.1/CD34+ cells proliferated and expanded as efficiently as uninfected cultures. However, when reselected magnetically using the K6.1 CD34 MoAb after expansion for 7 days, bound K6.1/CD34+ cells were again negative for HIV-1 expression, whereas unbound cells were positive for HIV-1 expression. These findings suggest that a sequential CD34+ cell-selection process, in which the two selections are separated by a brief culture period, can yield a population of CD34+ cells that are not infected with HIV-1. This process may be useful in the design of stem or progenitor cell-based transplantation therapies for HIV infection.
Asunto(s)
Antígenos CD34/análisis , VIH-1/fisiología , Células Madre Hematopoyéticas/virología , Secuencia de Bases , Células de la Médula Ósea , Diferenciación Celular , División Celular/efectos de los fármacos , Células Cultivadas , Citometría de Flujo , Células Madre Hematopoyéticas/citología , Humanos , Separación Inmunomagnética , Datos de Secuencia Molecular , Reacción en Cadena de la Polimerasa , Replicación ViralRESUMEN
A variety of deficiencies in T cell activation have been described in HIV-1 infection. To determine whether one component of Ag receptor signal transduction might be impaired and contribute to the immunopathology of HIV infection, we tested CD4 cells from patients with early to mid-stage HIV infection for TCR-induced calcium mobilization. There was no detectable difference between patients and controls in the mean CD4 cell calcium response or in the fraction of responding CD4 cells after cross-linking the TCR with OKT3 Ab. In addition, in HIV-infected patients, there was no correlation between calcium mobilization and the CD4 cell count. These results indicate that there are no intrinsic impairments of Ag receptor calcium signaling in circulating CD4 cells from HIV-infected patients with more than 400 CD4 cells/mm3, although abnormalities in patients with later stage infections cannot be excluded.
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Calcio/fisiología , Infecciones por VIH/inmunología , VIH-1/inmunología , Receptores de Antígenos de Linfocitos T/inmunología , Transducción de Señal/inmunología , Adulto , Linfocitos T CD4-Positivos/metabolismo , Femenino , VIH-1/metabolismo , Humanos , Masculino , Persona de Mediana EdadRESUMEN
Neutrophil chemiluminescence (CL) as a measure of oxygen-dependent killing activity was evaluated in 3 groups of patients: (a) 63 patients with multiple myeloma (MM); (b) 31 subjects with monoclonal gammopathy of undetermined significance (MGUS); (c) 32 healthy controls. Neutrophil CL response was shown to be significant reduced both in patients with MM (p less than 0.001) and in subjects with MGUS (p less than 0.001). A significant difference was also observed between the results obtained in MM and those of MGUS (p less than 0.001). Treated MM patients showed a more severe impairment of neutrophil chemiluminescence response than that observed in untreated patients (p less than 0.001). It is suggested that the impairment of neutrophil CL response, possibly related to decreased killing activity, may play a role, along with other known causes, in the increased susceptibility to infection observed in MM patients.
Asunto(s)
Mieloma Múltiple/metabolismo , Neutrófilos/metabolismo , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Ciclofosfamida/uso terapéutico , Humanos , Mediciones Luminiscentes , Melfalán/uso terapéutico , Mieloma Múltiple/tratamiento farmacológico , Proteínas de Mieloma/farmacología , Neutrófilos/efectos de los fármacos , Consumo de Oxígeno/efectos de los fármacos , Paraproteinemias/metabolismo , Peroxidasa/sangre , Prednisona/uso terapéutico , Acetato de Tetradecanoilforbol/farmacología , Vincristina/uso terapéutico , Zimosan/farmacologíaRESUMEN
Idiopathic (autoimmune) thrombocytopenic purpura (ATP) is accepted to be a disorder resulting from accelerated platelet destruction attributed to an autoimmune process. The patient whose case is presented in this article was first seen by a dentist. The oral findings have been documented as the case was followed for 3 years through acute exacerbations, pregnancy, and delivery of an infant with thrombocytopenia. The patient was managed with intermittent steroid therapy and splenectomy.