RESUMEN
Kawasaki disease (KD) is an acute vasculitis affecting mainly infants and children. Human B cells express Toll-like receptor (TLR)-9, whose natural ligands are unmethylated cytosine-guanine dinucleotide (CpG) motifs characteristic of bacterial DNA. The aim of this study was to clarify the pathogenesis of KD analysing the activation status of peripheral blood mononuclear cells (PBMC), focusing on B lymphocyte activation and functions. Ten patients and 10 age-matched healthy donors were recruited from the Bambino Gesù Hospital of Rome, Italy and enrolled into this study. We determined phenotype profile and immunoglobulin (Ig) production of PBMC from KD patients and age-matched controls. We found that the frequency of CD19(+) B lymphocytes and CD19(+) /CD86(+) activated B lymphocytes from KD patients during the acute phase before therapy was increased significantly. Moreover, B lymphocytes of acute-phase KD patients were more prone to CpG oligodeoxynucleotide (ODN) activation compared with the age-matched controls, as assessed by a significant increase of the number of IgA-secreting cells (SC). In the same patients we found a marked increase of IgM, IgG, interleukin (IL)-6 and tumour necrosis factor (TNF)-α production compared with the control group. In addition, in two convalescent KD patients, conventional treatment with intravenous immunoglobulin (IVIG) restored the normal frequency of CD19(+) B cells, the number of IgA-, IgM- and IgG-SC and the production of IL-6 and TNF-α. Our findings indicate that the percentages of peripheral B lymphocytes of acute-phase KD patients are increased and are prone to bacterial activation in terms of increased numbers of IgA-SC and increased production of IL-6 and TNF-α inflammatory cytokines. Thus, our data support the hypothesis of an infectious triggering in KD.
Asunto(s)
Células Productoras de Anticuerpos/metabolismo , Células Productoras de Anticuerpos/patología , Inmunoglobulina A/metabolismo , Síndrome Mucocutáneo Linfonodular/inmunología , Receptor Toll-Like 9/agonistas , Células Productoras de Anticuerpos/efectos de los fármacos , Antígenos CD19/metabolismo , Linfocitos B/efectos de los fármacos , Linfocitos B/metabolismo , Linfocitos B/patología , Preescolar , Femenino , Humanos , Inmunoglobulina G/metabolismo , Inmunoglobulina M/metabolismo , Inmunoglobulinas Intravenosas/uso terapéutico , Lactante , Interleucina-17/metabolismo , Interleucina-6/metabolismo , Células Asesinas Naturales/patología , Recuento de Linfocitos , Masculino , Síndrome Mucocutáneo Linfonodular/terapia , Oligodesoxirribonucleótidos/farmacología , Linfocitos T/metabolismo , Linfocitos T/patología , Factor de Necrosis Tumoral alfa/metabolismoRESUMEN
Surgically removed benign and malignant human skin lesions of nonmelanocyte origin have been tested with monoclonal antibodies to la antigens, to the HLA-A,B antigenic molecular complex, and to melanoma-associated antigen(s) (MAA). MAA include a high-molecular-weight (HMW) MAA, a 115,000-molecular-weight MAA, a 94,000-molecular-weight MAA, and a cytoplasmic MAA. Indirect immunofluorescence was used as the assay system because of the limited amount of tissue available. When the amount of tissue available was sufficient, double determinant immunoassays (DDIA) were used to quantitate the level of the HMW MAA and of the cytoplasmic MAA. The results of the DDIA were in agreement with those of indirect immunofluorescence in more than 75% of the cases. Malignant skin tumors of various histiotypes displayed three types of changes: 1) appearance of la antigens and cytoplasmic MAA, 2) increase in the level of the HMW MAA, of a 115,000- and a 100,000-molecular-weight MAA, and 3) reduction in the level of HLA-A,B antigens and beta 2-microglobulin. A significant heterogeneity was found in the antigenic profile among various lesions of a given histiotype as well as among tumor cells within a given lesion.
Asunto(s)
Anticuerpos Monoclonales , Antígenos de Neoplasias/análisis , Antígenos HLA/análisis , Proteínas de Neoplasias/análisis , Neoplasias Cutáneas/inmunología , Anciano , Femenino , Feto , Humanos , Antígenos Específicos del Melanoma , Persona de Mediana Edad , Peso Molecular , Embarazo , Valores de Referencia , Piel/inmunología , Enfermedades de la Piel/inmunologíaRESUMEN
Indirect immunofluorescence and immunoperoxidase staining of surgically removed tissues of nonlymphoid origin with monoclonal antibodies to the heavy and light chain of HLA-A,B,C antigens have shown that they have a more restricted tissue distribution than previously assumed. HLA-A,B,C antigens were not detected in brain cortex, cerebellum, sympathetic ganglia, hypophysis, parathyroid gland, thyroid, exocrine pancreas, hepatocytes, sperm, seminiferous tubules, or skeletal or smooth muscle. Malignant transformation of cells may be associated with appearance, changes in cellular distribution of HLA-A,B,C antigens, and/or dissociation in the expression of the two subunits. Analysis of primary tumors and of autologous metastases showed heterogeneity in the expression of HLA-A,B,C antigens among lesions removed from different sites. The degree of heterogeneity did not correlate with the site of origin of metastases.
Asunto(s)
Antígenos HLA/análisis , Neoplasias/inmunología , Femenino , Antígenos HLA-A , Antígenos HLA-B , Antígenos HLA-C , Humanos , Linfocitos/inmunología , Macrófagos/inmunología , Masculino , Metástasis de la Neoplasia , Neoplasias/patología , Especificidad de Órganos , Valores de Referencia , Distribución TisularRESUMEN
A cytoplasmic glycoprotein, originally identified by the monoclonal antibody 465.12S in melanoma tumors, is significantly increased in epithelial cells of different histotype following transformation. In the present study we show that the cytoplasmic melanoma associated antigen (cyt-MAA) is drastically enhanced in lymphoid cells by polyclonal and allogeneic stimulation, as well as by transformation. Normal T-cells with helper and suppressor phenotype are far more susceptible than B-cells to this enhancement. However, among transformed lymphoid cells, the expression of the cyt-MAA does not correlate with lineage, but rather with stage of differentiation. Acute lymphoblastic leukemias represent the only exception, since in these lymphoid malignancies cyt-MAA levels are highly heterogeneous even within groups of phenotypically similar lesions. Thus, the expression of the cyt-MAA is shared by cells of distant embryological origin in early stages of their differentiation and/or during proliferation. Quantitation of the cyt-MAA may provide useful information for the classification of some lymphoid malignancies.
Asunto(s)
Antígenos de Neoplasias/análisis , Citoplasma/inmunología , Activación de Linfocitos , Melanoma/inmunología , Proteínas de Neoplasias/análisis , Línea Celular , Enfermedad de Hodgkin/inmunología , Humanos , Tejido Linfoide/inmunología , Trastornos Linfoproliferativos/inmunología , Antígenos Específicos del Melanoma , Mitógenos/farmacología , Peso Molecular , Linfocitos T/clasificación , Linfocitos T/inmunologíaRESUMEN
In this in vitro study, an evaluation of the importance of intracellular oxidative balance on cell-mediated cytotoxicity was performed by analyzing the effects of the antioxidant N-acetylcysteine (NAC), a specific thiol supplier, on natural killer (NK) cell-mediated cytotoxicity. The results obtained indicate that an enhancement of target cell (TC) killing can be detected when a pre-exposure of effector cells (EC) to NAC was performed. However, this effect seems to depend upon the TC type used. In fact, the increase of EC activity was detected against the differentiated U937 TC while no changes were detected by the same effectors against K562 cells. The mechanism of this enhancement seems to be ascribable to an increased ability of NAC-exposed NK cells to form conjugates (binding) which, in turn, appears to be due to a specific effect of NAC on actin microfilaments. A role for NAC as a cytoskeleton thiol-modifier contributing to the activation of effector cells can thus be hypothesized.
Asunto(s)
Acetilcisteína/farmacología , Citotoxicidad Inmunológica , Células Asesinas Naturales/metabolismo , Proteínas de Microfilamentos , Actinas/metabolismo , Membrana Celular/metabolismo , Humanos , Células Asesinas Naturales/efectos de los fármacos , Células Asesinas Naturales/ultraestructura , Glicoproteínas de Membrana/metabolismo , Microscopía Fluorescente , Monocitos/metabolismo , Compuestos de Sulfhidrilo , Células Tumorales Cultivadas/metabolismoRESUMEN
Synthetic peptides containing amino acid sequence 218-238 of the core protein p24 of human immunodeficiency virus type 1 (HIV-1) and progressively shorter sequences at its C-terminus, were tested for their effect on antigen dependent in vitro responses of peripheral blood lymphocytes (PBL) from normal human donors. A peptide as short as 7 amino acids, corresponding to a highly conserved sequence, was able to inhibit in a dose-dependent manner the induction of a specific primary antibody response to the sheep red cell (SRC) antigen, as well as the proliferative response to recall microbial antigens. The results of this study constitute additional evidence of the immunoinhibitory effects of HIV components and may help to unravel some of the pathogenic mechanisms of AIDS. Moreover, they are of potential relevance for the development of immunoprophylactic and therapeutic strategies.
Asunto(s)
Proteína p24 del Núcleo del VIH/química , Linfocitos/inmunología , Secuencia de Aminoácidos , Regulación hacia Abajo , Epítopos/química , Anticuerpos Anti-VIH/biosíntesis , Humanos , Inmunosupresores/química , Inmunosupresores/farmacología , Técnicas In Vitro , Indometacina/farmacología , Activación de Linfocitos , Linfocitos/efectos de los fármacos , Datos de Secuencia Molecular , Oligopéptidos/química , Oligopéptidos/inmunología , Oligopéptidos/farmacología , Péptidos/química , Péptidos/inmunología , Péptidos/farmacologíaRESUMEN
We investigated the role of some gp120 peptides on the apoptosis induction in malignant T cell lines. We took advantage of recent findings reporting that three major regions of gp120 are important for CD4 binding. They consist of residues 256-262 in the C2 domain, residues 368-389 in the C3 domain, and residues 421-457 in C4 domain. We used a peptide from C2 domain (aa 250-263) the homologous major histocompatibility complex (MHC) class II peptide (aa 135-155) and three peptides from domain C4 (aa 414-434; 419-430; 428-445). We selected for this study the following human cell lines: CEM and Jurkat, two lymphoblastoid CD4-positive T cell line and U937, a myelomonocytic CD4 positive cell line. We demonstrated that the CD4-positive T cell lines, in the presence of gp120 250-263 peptide and DR 135-155 peptide, can be induced to accelerate apoptosis, while no effect in apoptosis induction was observed in the presence of 414-424 gp120 peptide. Interestingly, we have shown by fluorescence study, that the small sequence 414-419 must be responsible for the inhibition of binding of gp120 to the CD4 molecule. Indeed while 414-424 gp120 peptide is very efficient in CD4-gp120 binding inhibition, no effect is observed in the presence of either 419-430 or 428-445 peptide.
Asunto(s)
Apoptosis/efectos de los fármacos , Apoptosis/inmunología , Proteína gp120 de Envoltorio del VIH/farmacología , VIH-1/inmunología , Péptidos/farmacología , Secuencia de Aminoácidos , Unión Competitiva/inmunología , Linfocitos T CD4-Positivos/inmunología , Citometría de Flujo , Proteína gp120 de Envoltorio del VIH/inmunología , Humanos , Células Jurkat , Leucemia Promielocítica Aguda , Linfoma de Células B Grandes Difuso , Datos de Secuencia Molecular , Péptidos/inmunología , Unión Proteica/inmunologíaRESUMEN
In the complex mechanism of adhesion, internalization, and infection of cells by human immunodeficiency virus (HIV) viral particles, a determinant role is played by the viral envelope glycoprotein gp120, which binds to CD4 receptors of T cells and monocytes. We tested the ability of a panel of 7- to 12-residue synthetic peptides, selected from the region 414-434 of the HIV-1 gp120, to inhibit the binding of the viral protein to CD4 receptors of cultured human lymphoid cells. The assay was based on the observation that the binding of gp120 to the receptors interferes with the binding of a specific anti-CD4 monoclonal antibody, as a result of the masking of the antibody epitope; thus, we tested whether preincubation of cells with the peptides before gp120 addition might restore the recognition of the CD4 molecule by the antibody. High expression of CD4 receptors was thus assumed as indication that the binding of the viral protein had been inhibited. Maximum activity was displayed by a 9-residue peptide located near the amino terminal end of the 414-434 fragment. In addition, several fragments deduced from other viral proteins, possessing partial amino acid sequence homology with the HIV gp120 fragment, exhibited a similar type of interaction with the CD4 receptor. All active peptides contain the Cys residue (position 423 of gp120). This residue is essential, although not sufficient, for inhibiting gp120 binding, as few other amino acid residues within the fragment play a complementary role in increasing or decreasing the inhibitory ability.
Asunto(s)
Antígenos CD4/metabolismo , Proteína gp120 de Envoltorio del VIH/metabolismo , Fragmentos de Péptidos/metabolismo , Secuencia de Aminoácidos , Sustitución de Aminoácidos , Unión Competitiva , Línea Celular , Proteína gp120 de Envoltorio del VIH/química , Humanos , Datos de Secuencia Molecular , Fragmentos de Péptidos/química , Fragmentos de Péptidos/farmacología , Proteínas Virales/químicaRESUMEN
The aim of this study was to assess the antibody reactivity in HIV-infected subjects against an HIV-1 p24 sequence, p226 (aa226-237), including a seven amino acid epitope showing immunosuppressive activity in vitro and to evaluate the relationship between anti-peptide antibody levels and disease progression. Sera of HIV-infected subjects, at different stages of disease, were compared to control sera in a retrospective evaluation. Recombinant HIV-1 p24 and p24- and control-peptides were used in an enzyme immunoassay as targets for antibodies present in the sera. Antibodies directed against the whole p24 protein and its peptides were found in all the sera studied but at different levels. The anti-p226 reactivity was not significantly different at different clinical stages. Nevertheless, it was inversely correlated to the reactivity directed against the whole protein, that was lower in subjects characterized by low CD4 cell numbers.
Asunto(s)
Anticuerpos Anti-VIH/sangre , Proteína p24 del Núcleo del VIH/inmunología , Infecciones por VIH/sangre , Inmunosupresores/farmacología , Secuencia de Aminoácidos , Recuento de Linfocito CD4 , Anticuerpos Anti-VIH/biosíntesis , Proteína p24 del Núcleo del VIH/farmacología , Infecciones por VIH/inmunología , VIH-1/inmunología , Humanos , Activación de Linfocitos , Datos de Secuencia MolecularRESUMEN
Oxidized LDL (ox-LDL) have been involved in the pathogenesis of several human diseases including dermatological pathologies. Oxidative modification of low-density lipoproteins (LDL) is accompanied by both extensive degradation of its polyunsaturated fatty acids and production of lipoperoxides. These highly reactive products induce an intracellular oxidative stress with a variety of cytotoxic effects. In order to evaluate cellular damage induced by oxidative stress in epidermal cells, a human epidermoid carcinoma cell line in culture (A 431) was used as experimental model. Cell treatment with UV-oxidized LDL resulted in cytostatic and cytotoxic effects characterized by morphological and functional alterations: inhibition of cell proliferation, modifications of cytoskeleton network, microtubular derangement, loss of cell-cell and cell-substrate contacts, cell detachment and cell death by apoptosis. The ox-LDL-induced alterations were almost completely prevented by pre-incubating cells with alpha-tocopherol. The results presented here could be of relevance for a better comprehension of the pathogenic mechanisms of several human diseases, including dermatological pathologies, and could indicate that antioxidants such as alpha-tocopherol could represent an important therapeutic challenge in the maintenance of cell and tissue homeostasis in the long run.
Asunto(s)
Lipoproteínas LDL/fisiología , Vitamina E/farmacología , Células Epiteliales/efectos de los fármacos , Células Epiteliales/metabolismo , Células Epiteliales/ultraestructura , Humanos , Lipoproteínas LDL/efectos de la radiación , Microscopía Electrónica de Rastreo , Estrés Oxidativo , Fracciones Subcelulares/efectos de los fármacos , Fracciones Subcelulares/metabolismo , Células Tumorales Cultivadas , Rayos UltravioletaRESUMEN
Peripheral blood mononuclear cells from HIV-infected subjects have been demonstrated by different methods to die by apoptosis after short time in culture. In the present study the percentages of apoptotic cells have been measured by propidium iodide staining and flow cytometry in PBMC from healthy controls (15) and HIV-infected subjects with asymptomatic (10) or advanced (15) disease, with or without anti-viral treatment. The percentage of apoptosis significantly correlated with clinical stage (CDCII: 15.85% +/- 9.17, CDCIV: 22.6% +/- 5.97, P < 0.001) and the CD4/CD8 CD3 cell ratio. R = -0.57, P = 0.012), while no differences were found in relation to AZT therapy. By adding IL-2 to the cultures the percentages of apoptosis of PBMC from HIV-infected patients were significantly reduced in all experiments.
Asunto(s)
Apoptosis/inmunología , Infecciones por VIH/sangre , Infecciones por VIH/inmunología , VIH-1 , Interleucina-2/farmacología , Leucocitos Mononucleares/inmunología , Leucocitos Mononucleares/patología , ADN/sangre , Citometría de Flujo , Humanos , Técnicas In Vitro , Leucocitos Mononucleares/metabolismoRESUMEN
TLRs are a family of molecules that function as sensors for the detection of pathogens. TLR-9, expressed on B cells and pDCs, recognizes CpG motifs of unmethylated bacterial DNA and plays a role in the development of autoimmunity. The present study was designed to investigate the effects of IFN-alpha in combination with CpG ODN on the activation of CD27(-) naïve B cells and on Ig production. We provide evidence that CpG ODN not only induces a total and T-dependent, specific IgM response by naïve B cells but also their phenotypic differentiation in plasma cells, as demonstrated by the up-regulation of CD38 expression. We found that TLR-9 stimulation with CpG ODN induces IL-1beta, TNF-alpha, IL-10, and IL-6 production. Interestingly, we also found that CpG ODN induces naïve B cell maturation into memory cells, as demonstrated by the induction of CD27, AID mRNA expression, and IgG production. More importantly, our results demonstrate that IFN-alpha amplifies the inductive effect of CpG ODN on naïve B activation and on Ig production through a mechanism involving TLR-9/MyD88-dependent signaling. Moreover, we found that IFN-alpha enhances the frequency of CpG ODN-induced memory B cells. Our results may contribute to clarify the events promoting IFN-alpha-induced amplification of naïve B cell activation via TLR-9 for a better understanding of the pathogenesis of autoimmune disorders and may guide treatments targeting this pathway within B cells.
Asunto(s)
Formación de Anticuerpos/inmunología , Linfocitos B/inmunología , Linfocitos B/metabolismo , Interferón-alfa/metabolismo , Activación de Linfocitos/inmunología , Receptor Toll-Like 9/inmunología , ADP-Ribosil Ciclasa 1/metabolismo , Formación de Anticuerpos/efectos de los fármacos , Linfocitos B/efectos de los fármacos , Diferenciación Celular/efectos de los fármacos , Diferenciación Celular/inmunología , Células Cultivadas , Citidina Desaminasa/genética , Citidina Desaminasa/metabolismo , Citocinas/metabolismo , Humanos , Inmunoglobulina G/metabolismo , Inmunoglobulinas/metabolismo , Memoria Inmunológica/efectos de los fármacos , Memoria Inmunológica/inmunología , Interferón-alfa/farmacología , Activación de Linfocitos/efectos de los fármacos , Glicoproteínas de Membrana/metabolismo , Oligodesoxirribonucleótidos/farmacología , Fenotipo , Células Plasmáticas/efectos de los fármacos , Células Plasmáticas/inmunología , Células Plasmáticas/metabolismo , Receptor Toll-Like 9/agonistas , Miembro 7 de la Superfamilia de Receptores de Factores de Necrosis Tumoral/genética , Miembro 7 de la Superfamilia de Receptores de Factores de Necrosis Tumoral/metabolismoRESUMEN
The in vitro effect of single or combined doses of zidovudine (AZT) and dideoxycytidine (ddC) on the production and utilization of interleukin-2 (IL-2) by normal human peripheral blood mononuclear cells (PBMC) was evaluated by measuring IL-2 concentrations in supernatants from PHA-stimulated PBMC cultures. Drugs were added at the beginning of the culture period and left throughout. Whereas AZT alone (1 and 10 microM) caused only a slight increase, ddC alone (1 and 10 microM) and combined AZT/ddC (1 + 1 and 10 + 10 microM) caused a considerable increase. IL-2 gene expression in the drug-treated PBMC did not increase. This finding suggested that the increased supernatant IL-2 accumulations might be caused by a drug-induced down-regulation of the IL-2 receptor alpha (IL-2R alpha, CD25). AZT decreased IL-2R alpha expression, but only slightly. In contrast, ddC alone and combined AZT/ddC decreased the CD25 molecules in a marked and dose-dependent manner. They also markedly reduced IL-2R alpha gene expression. These findings show that the dideoxynucleoside drugs tested left PHA-induced IL-2 gene activation unchanged but decreased IL-2R alpha gene activation, thus down-regulating IL-2R alpha cell-surface protein expression.
Asunto(s)
Didesoxinucleósidos/farmacología , Regulación hacia Abajo/genética , Proteínas/efectos de los fármacos , Receptores de Interleucina-2/genética , Receptores de Interleucina-2/fisiología , Antígenos de Superficie , Secuencia de Bases , División Celular , Expresión Génica , Regulación de la Expresión Génica , Humanos , Interleucina-2/biosíntesis , Interleucina-2/genética , Leucocitos Mononucleares/citología , Activación de Linfocitos/efectos de los fármacos , Datos de Secuencia Molecular , Fitohemaglutininas/farmacología , Biosíntesis de Proteínas , Receptores de Interleucina-2/biosíntesis , Receptores de Interleucina-2/efectos de los fármacos , Activación Transcripcional , Zalcitabina/farmacología , Zidovudina/farmacologíaRESUMEN
We have previously demonstrated that exogenous Nef protein induced activation of normal human T cells up-regulating IL-15 production by monocytes. Since HIV-1 infection results in the early impairment of immune functions we decided to evaluate if Nef is able to modulate the induction of a specific antibody response. Human peripheral blood mononuclear cells from healthy donors were induced in vitro to mount a specific antibody response to the Candida albicans antigen. We show that Nef inhibited, in a dose-dependent manner, the induction of the anti-C. albicans antibody response. The ability of an anti-Nef antibody to prevent such inhibition indicates that the effect was indeed Nef-specific. In the Nef-treated cultures an early increase of IL-15 production was observed and the addition of anti-IL-15 antibody abrogated the Nef-induced inhibitory effect. Moreover the addition of IL-15 to the cultures inhibited, as well as Nef, the induction of the specific antibody response. Thus, our results suggest that Nef may inhibit the induction of a specific antibody response by an early up-regulation of IL-15 production. A better comprehension of this phenomenon may be important for unravelling some aspects of the B cell defects in HIV infection.
Asunto(s)
Anticuerpos Antifúngicos/inmunología , Candida albicans/inmunología , Productos del Gen nef/inmunología , VIH-1/inmunología , Interleucina-15/biosíntesis , Células Cultivadas , Productos del Gen nef/fisiología , Humanos , Terapia de Inmunosupresión , Interleucina-15/inmunología , Interleucina-15/farmacología , Leucocitos Mononucleares/citología , Leucocitos Mononucleares/efectos de los fármacos , Leucocitos Mononucleares/inmunología , Leucocitos Mononucleares/microbiología , Regulación hacia Arriba , Productos del Gen nef del Virus de la Inmunodeficiencia HumanaRESUMEN
Several recent reports have provided evidence that Nef enhances human immunodeficiency virus HIV infectivity, and in vitro experiments with the nef gene have demonstrated the possible role of Nef in modulating immune responses. Exogenous Nef has been demonstrated to induce proliferation of normal human peripheral blood mononuclear cells (PBMC) and to enhance HIV-1 replication. The aim of this study was to evaluate the biological mechanisms by which Nef, used as exogenous protein, modulates cellular activation. We showed that exogenous Nef protein induces the proliferation of unstimulated and suboptimally stimulated normal human PBMC, while it has no effect on the proliferation of optimally stimulated PBMC. Moreover, the activating effect of exogenous Nef on PBMC proliferation was associated with an increase of IFN-gamma, TNF-alpha, and IL-6 production, while, surprisingly, IL-2 production was not affected by Nef. More importantly we showed, for the first time, that Nef exerts its activating effects on PBMC proliferation through IL-15 synthesis induction by monocyte/macrophage population. In conclusion, we found that exogenous Nef protein (i) induces activation of normal PBMC, increasing their proliferative response; (ii) modulates cytokine production; (iii) exerts its activating effects through IL-15 synthesis induction; and (iv) exerts these effects entering monocyte/macrophages. Our results might suggest that Nef enhances the rate of viral replication by a novel mechanism involving the production of IL-15.
Asunto(s)
Productos del Gen nef/fisiología , VIH-1/fisiología , Interleucina-15/biosíntesis , División Celular/fisiología , Células Cultivadas , Productos del Gen nef/inmunología , VIH-1/inmunología , Humanos , Leucocitos Mononucleares/citología , Leucocitos Mononucleares/inmunología , Proteínas Recombinantes/inmunología , Proteínas Recombinantes/farmacología , Células U937 , Productos del Gen nef del Virus de la Inmunodeficiencia HumanaRESUMEN
Fine-needle aspiration cytology (FNAC) is currently used in evaluating the nature of breast nodules. In the present study we have examined whether monoclonal antibodies (MAbs) to breast-tumor-associated antigens (BTAA) can be employed in FNAC for the diagnosis of breast cancer. For this purpose we have used 2 murine MAbs recognizing 2 distinct BTAA expressed by breast tumors, irrespective of their histotype, in an indirect avidin-biotin immunohistochemical technique on aspirate smears. The results of this study show that, while benign lesions are consistently negative, tumor cells containing aspirates are reactive with at least one MAb in 95% of the cases. Thus, selected MAbs to BTAA may be a powerful diagnostic tool in conjunction with conventional cytology, and because of the objective interpretation of the immunohistochemical findings, FNAC of the breast can eventually be extended outside specialized institutions.
Asunto(s)
Anticuerpos Monoclonales , Antígenos de Neoplasias/análisis , Neoplasias de la Mama/diagnóstico , Adulto , Anciano , Antígenos de Neoplasias/inmunología , Biopsia con Aguja , Neoplasias de la Mama/inmunología , Neoplasias de la Mama/patología , Femenino , Histocitoquímica , Humanos , Persona de Mediana EdadRESUMEN
This study investigates the in vitro effects of oxidized low-density lipoproteins (ox-LDL), 'physiological' pro-oxidants, N-acetylcysteine (NAC), a free radical scavenger and glutathione precursor, and their combination on human peripheral blood mononuclear cell functions. We found that treatment with ox-LDL induced a significant down-regulation of proliferative response to mitogens, antigens and interleukin-2. Lipid extracts from ox-LDL were able to reproduce the same effect as the lipoprotein. On the other hand, NAC exposure induced a significant up-regulation of proliferative responses to all the stimuli used. Moreover, we showed that natural killer (NK) cell-mediated cytotoxic activity was significantly down-regulated by ox-LDL while treatment with NAC induced a significant up-regulation of NK-cell activity. Finally, we found that ox-LDL and NAC exerted opposite effects on the cytokine network, interfering both at the protein secretion level and the messenger RNA synthesis level. More importantly, when NAC was used in combination with ox-LDL the proliferative responses, NK-cell-mediated cytotoxic activity and cytokine production were restored to values comparable to controls. These data indicate that ox-LDL and NAC modulate immune functions, exerting opposite effects reflecting their pro-oxidant and antioxidant behaviours. Our results add new insights to the key role played by redox imbalance as a modulator of immune system homeostasis and suggest that an antioxidant drug such as NAC could be useful against pathologies associated with an increase in lipid peroxidation.
Asunto(s)
Acetilcisteína/farmacología , Depuradores de Radicales Libres/farmacología , Leucocitos Mononucleares/efectos de los fármacos , Lipoproteínas LDL/farmacología , Estrés Oxidativo/inmunología , División Celular/efectos de los fármacos , Células Cultivadas , Citocinas/biosíntesis , Citotoxicidad Inmunológica/efectos de los fármacos , Regulación hacia Abajo/efectos de los fármacos , Humanos , Células Asesinas Naturales/efectos de los fármacos , Células Asesinas Naturales/inmunología , Leucocitos Mononucleares/inmunología , Activación de Linfocitos/efectos de los fármacos , Mitógenos/inmunología , Oxidación-Reducción , Regulación hacia Arriba/efectos de los fármacosRESUMEN
We investigated the effect of N-acetylcysteine (NAC) on normal human B cell functions. We found that NAC significantly inhibited both the induction of the specific antibody response to the T-dependent antigen Candida albicans and T-dependent pokeweed mitogen (PWM)-induced polyclonal Ig production. NAC did not induce either cell death due to a non-specific toxicity or apoptosis. The NAC-induced inhibitory effect might be a functional consequence of: (i) a down-regulation of the expression on the B cell surface of CD40 and CD27 co-stimulatory molecules and (ii) a down-regulation of interleukin (IL-4) production. In contrast, NAC up-regulated interferon-gamma (IFN-gamma) production. NAC did not induce any effect on the T cell-independent B cell polyclonal activation system. These results indicate that NAC down-regulates T dependent B cell activation and leads to T helper cell type 1 (Th1) polarization.
Asunto(s)
Acetilcisteína/farmacología , Formación de Anticuerpos/efectos de los fármacos , Antígenos CD40/metabolismo , Miembro 7 de la Superfamilia de Receptores de Factores de Necrosis Tumoral/metabolismo , Células Productoras de Anticuerpos/efectos de los fármacos , Células Productoras de Anticuerpos/inmunología , Antígenos , Antioxidantes/farmacología , Linfocitos B/citología , Linfocitos B/efectos de los fármacos , Linfocitos B/inmunología , Supervivencia Celular/efectos de los fármacos , Citocinas/biosíntesis , Regulación hacia Abajo/efectos de los fármacos , Humanos , Técnicas In Vitro , Interferón gamma/biosíntesis , Interleucina-4/biosíntesis , Activación de Linfocitos/efectos de los fármacos , Células TH1/efectos de los fármacos , Células TH1/inmunología , Regulación hacia Arriba/efectos de los fármacosRESUMEN
In this paper, we show that the antioxidant N-acetyl-cysteine (NAC) is capable of enhancing the adhesion properties of the epithelial cell line A431 and of the lymphocytic cells with cytotoxic activity from human peripheral blood: the natural killer (NK) cells. This effect leads to an increased efficiency of A431 cells to form a monolayer and of NK cells to kill their targets. In both cases a specific effect of NAC was found in the distribution of those molecules of the cytoskeleton which are generally involved in cell substrate and cell-to-cell contact region formation, e.g., the actin microfilaments. NAC could thus behave as a drug influencing certain cytoskeleton-dependent cell processes in a non-histotype dependent manner.
Asunto(s)
Acetilcisteína/farmacología , Antioxidantes/farmacología , Adhesión Celular/efectos de los fármacos , Depuradores de Radicales Libres/farmacología , Células Asesinas Naturales/inmunología , Linfocitos/fisiología , Carcinoma de Células Escamosas , Línea Celular , Células Cultivadas , Citoesqueleto/efectos de los fármacos , Citoesqueleto/fisiología , Citoesqueleto/ultraestructura , Citotoxicidad Inmunológica/efectos de los fármacos , Epitelio/efectos de los fármacos , Epitelio/fisiología , Glutatión/metabolismo , Humanos , Células Asesinas Naturales/efectos de los fármacos , Linfocitos/efectos de los fármacos , Células Tumorales CultivadasRESUMEN
The in vitro effect of single or combined doses of zidovudine (AZT) and dideoxycytidine (ddC) on PHA-activated human peripheral blood mononuclear cells (PBMC) proliferative response and lymphoblastoid T cell line CEM cell growth was evaluated. Clinically relevant amounts (0.1, 1 and 10 microM) of AZT, ddC and AZT/ddC combination (10 + 10 microM) inhibited 3H TdR uptake in both cell models in a dose-dependent manner. The inhibitory effect on cell growth was confirmed by counting the amount of viable CEM cells recovered after 24, 48 and 72 h exposure to the drugs. On equimolar basis, ddC was considerably more efficient than AZT although the latter potentiates the activity of the former Flow cytometric analysis of PBMC and CEM cells exposed to the dideoxynucleosides revealed a decrease in the rate of DNA synthesis (rate of passage through the S phase of the cell cycle) and a reduced number of cell generations, the latter assessed by measuring the halving of the fluorescent probe 5-6-carboxyfluorescein diacetate succinimidyl ester by flow cytometry. The analysis of CEM cells recovered after exposure to ddC or AZT/ddC combination (10 + 10 microM), showed that in addition to perturbing cell cycle progression, ddC, and most efficiently the AZT/ddC combination, induced cell death by apoptosis. The latter was manifested as enhanced side scatter and decreased, sub-G1, DNA content by flow cytometry, and as DNA breakdown in nucleosomal fragments by gel electrophoresis. Present findings indicate that clinically relevant concentrations of dideoxynucleosides reduce cell growth by hampering DNA replication and inducing apoptosis.