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Since the start of the COVID-19 pandemic, in a short span of 3 years, vaccination against SARS-CoV-2 has resulted in the end of the pandemic. Patients with inborn errors of immunity (IEI) are at an increased risk for SARS-CoV-2 infection; however, serious illnesses and mortality, especially in primary antibody deficiencies (PADs), have been lower than expected and lower than other high-risk groups. This suggests that PAD patients may mount a reasonable effective response to the SARS-CoV-2 vaccine. Several studies have been published regarding antibody responses, with contradictory reports. The current study is, perhaps, the most comprehensive study of phenotypically defined various lymphocyte populations in PAD patients following the SARS-CoV-2 vaccine. In this study, we examined, following two vaccinations and, in a few cases, prior to and following the 1st and 2nd vaccinations, subsets of CD4 and CD8 T cells (Naïve, TCM, TEM, TEMRA), T follicular helper cells (TFH1, TFH2, TFH17, TFH1/17), B cells (naïve, transitional, marginal zone, germinal center, IgM memory, switched memory, plasmablasts, CD21low), regulatory lymphocytes (CD4Treg, CD8Treg, TFR, Breg), and SARS-CoV-2-specific activation of CD4 T cells and CD8 T cells (CD69, CD137), SARS-CoV-2 tetramer-positive CD8 T cells, and CD8 CTL. Our data show significant alterations in various B cell subsets including Breg, whereas only a few subsets of various T cells revealed alterations. These data suggest that large proportions of PAD patients may mount significant responses to the vaccine.
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INTRODUCTION: Regulatory lymphocytes (CD4+ T regulatory cells [Treg], CD8+ Treg, and B regulatory cells [Breg]) play a critical role in immune homeostasis and tolerance. Common variable immunodeficiency (CVID) is associated with increased susceptibility to infections and increased frequency of inflammatory and autoimmune diseases. CD4+ Treg cell abnormalities have been reported in CVID; however, CD8+ Treg cells have not been reported in CVID. The objective of this study was to evaluate CD4+ Treg and CD8+ Treg cells in CVID patients. METHODS: In 25 patients with CVID and age-matched healthy controls, Treg cells, evaluated in freshly isolated peripheral blood mononuclear cells (natural; nCD4+ Treg and nCD8+ Treg) and following in vitro activation with anti-CD3/CD28 monoclonal antibodies (induced; iCD4+ Treg and iCD8+ Treg) as well as Breg cells were analyzed with specific monoclonal antibodies and isotype controls using flow cytometry. RESULTS: The proportions of nCD4+ Treg (CD4+ CD127low CD25high FoxP3+), iCD4+ Treg (CD4+ CD127low CD25high FoxP3+), iCD8+ Treg (CD8+ CD25high CD183+ FoxP3+), and Breg (CD19+ CD24high CD38high) lymphocytes were significantly lower in patients with CVID than in controls. CONCLUSIONS: Altered regulatory lymphocytes may play a role in the pathogenesis and autoimmunity and inflammation associated with CVID.
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Linfocitos B Reguladores/inmunología , Linfocitos T CD8-positivos/inmunología , Inmunodeficiencia Variable Común/inmunología , Linfocitos T Reguladores/inmunología , Adolescente , Adulto , Anciano , Envejecimiento/inmunología , Autoinmunidad/inmunología , Femenino , Humanos , Inflamación/inmunología , Activación de Linfocitos/inmunología , Recuento de Linfocitos , Masculino , Persona de Mediana Edad , Adulto JovenRESUMEN
BACKGROUND: Progressive T cell decline in aged humans is associated with a deficiency of naïve (TN) and central memory (TCM) T cells. We have previously reported increased Tumor necrosis factor-α (TNF-α)-induced apoptosis in TN and TCM T cells in aged humans; however, the molecular basis of increased apoptosis remains to be defined. Since expression of TNF receptors (TNFRs) was reported to be comparable in young and aged, we investigated signaling events downstream of TNFRs to understand the molecular basis of increased TNF-α-induced apoptosis in aged TN and TCM CD8+ cells. RESULTS: The expression of TRAF-2 and RIP, phosphorylation of JNK, IKKα/ß, and IκBα, and activation of NF-κB activation were significantly decreased in TN and TCM CD8+ cells from aged subjects as compared to young controls. Furthermore, expression of A20, Bcl-xL, cIAP1, and FLIP-L and FLIP-S was significantly decreased in TN and TCM CD8+ cells from aged subjects. CONCLUSIONS: These data demonstrate that an impaired expression/function of molecules downstream TNFR signaling pathway that confer survival signals contribute to increased apoptosis of TN and TCM CD8+ cells in aged humans.
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IgMFcR (FcµR) are expressed on B cell and B cell subsets. Mice deficient in secreted IgM and FcµR share properties of impaired specific antibody response and autoimmunity with patient with selective IgM deficiency (SIGMD). Intravenous immunoglobulin (IGIV) regulates immune response, including modulation of IgGFc receptors. However, there are no data on the expression of FcµR in patients with SIGMD, and the effects of IGIV on FcµR. In this study, we investigated FcµR expression in naïve marginal zone (MZ), IgM memory, and class-switched memory B cells in patients with selective IgM deficiency and healthy controls. Furthermore, we examined the direct effect of IGIV on FcµR expression and on the upregulation of FcµR by TLR2 agonist (Pam3). Finally, we examined the effect of IVIG on spontaneously produced IgM and natural IgM anti-phosphorylcholine (PC) antibodies by B cells and B1 cells. FcµR expression is significantly decreased in MZ B cells in patients with SIGMD as compared to control. IGIV, at immunomodulatory concentrations, inhibited FcµR upregulation by Pam3 in MZ B cells, and IgM-depleted IGIV inhibited spontaneous secretion of natural IgM anti-PC antibodies and not total IgM by B1 cells. These data suggest that decreased FcµR expression on MZ B cells may play a role in the pathogenesis of SIGMD, and an inhibition of TLR-2-induced upregulation of FcµR by IGIV may be one of the mechanisms of its anti-inflammatory action. IGIV-induced inhibition of natural IgM antibodies may be one of the mechanisms of IGIV-induced immunoregulation.
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Subgrupos de Linfocitos B/inmunología , Inmunoglobulina M/metabolismo , Síndromes de Inmunodeficiencia/inmunología , Inmunoterapia/métodos , Receptores Fc/metabolismo , Animales , Autoinmunidad , Subgrupos de Linfocitos B/efectos de los fármacos , Células Cultivadas , Humanos , Inmunidad Humoral , Cambio de Clase de Inmunoglobulina , Inmunoglobulina M/genética , Inmunoglobulinas Intravenosas/uso terapéutico , Síndromes de Inmunodeficiencia/terapia , Memoria Inmunológica , Activación de Linfocitos , Ratones , Receptores Fc/genéticaRESUMEN
Coenzyme Q10, (CoQ10) an electron transporter and an antioxidant, protects a variety of cell types against oxidative stress and apoptosis. However, protective effect of CoQ10 on oxidative stress-induced apoptosis in lymphocytes has not been studied in detail. In this study, we investigated the effect of CoQ10 on oxidative stress-induced apoptosis in lymphocytes. An exposure of peripheral blood lymphocytes to oxidative stressors, rotenone or hydrogen peroxide, lead to apoptosis. Pre-treatment of lymphocytes with CoQ10 resulted in a significantly reduced level of oxidative stress-induced apoptosis, which was associated with decreased reactive oxygen species production, an inhibition of mitochondrial membrane depolarization, and inhibition of activation of caspase-9 and caspase-3. Furthermore, CoQ10 inhibited oxidative stress induced apoptosis in both CD4+ T, and CD8+ T, and CD19+ B cells. Our findings suggest that CoQ10 may provide new therapeutic strategies for preventing oxidative stress-induced cell death and dysfunction in lymphocytes and lymphocyte subsets.
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Chronic, low grade inflammation is a characteristic of old age. Innate immune system cells such as dendritic cells (DCs) from the elderly display a pro-inflammatory phenotype associated with increased reactivity to self. Lithium is a well-established anti-inflammatory agent used in the treatment of bipolar disorders. It has also been reported to reduce inflammation in DCs. Here, we investigated whether Lithium is effective in reducing the inflammatory responses in DCs from the elderly. The effect of Lithium Chloride (LiCl) was compared on the response of TLR4 agonist, LPS and TLR2 agonist, PAM3CSK4 stimulated aged and young DCs. LiCl enhanced the production of IL-10 in LPS stimulated young DCs. However, it did not affect TNF-α and IL-6 production. In contrast, in aged DCs, LiCl reduced the secretion of TNF-α and IL-6 in LPS stimulated DCs but did not increase IL-10. LiCl had no significant effect on PAM3CSK4 responses in aged and young DCs. LiCl treated DCs also displayed differences at the level of CD4 T cell priming and polarization. LPS-stimulated young DCs reduced IFN-γ secretion and biased the Th cell response towards Th2/Treg while LiCl treated aged DCs only reduced IFN-γ secretion but did not bias the response towards Th2/Treg. In summary, our data suggests that LiCl reduces inflammation in aged and young DCs via different mechanisms. Furthermore, the effect of LiCl is different on LPS and PAM3CSK4 responses.
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Envejecimiento/inmunología , Células Dendríticas/efectos de los fármacos , Células Dendríticas/inmunología , Interleucina-10/biosíntesis , Cloruro de Litio/farmacología , Adulto , Anciano , Anciano de 80 o más Años , Antígeno B7-1/metabolismo , Linfocitos T CD4-Positivos/citología , Linfocitos T CD4-Positivos/efectos de los fármacos , Linfocitos T CD4-Positivos/inmunología , Linfocitos T CD8-positivos/citología , Linfocitos T CD8-positivos/efectos de los fármacos , Linfocitos T CD8-positivos/inmunología , Proliferación Celular/efectos de los fármacos , Humanos , Inmunidad Innata/efectos de los fármacos , Inflamación/inmunología , Inflamación/prevención & control , Interleucina-6/biosíntesis , Lipopolisacáridos/farmacología , Lipoproteínas/farmacología , Receptor Toll-Like 2/agonistas , Receptor Toll-Like 4/agonistas , Factor de Necrosis Tumoral alfa/biosíntesis , Adulto JovenRESUMEN
Aging is associated chronic inflammation and autoimmunity, and increased levels of leptin. Increased levels of leptin are associated with inflammation and autoimmunity. We have recently reported that leptin activates B cells to induce secretion of proinflammatory and anti-inflammatory cytokines. Role of B cells and leptin in inflammation associated with aging has not been explored. In this study we demonstrate that leptin activates and induces significantly greater amount of IL-6, TNF-α, and IL-10 by B cells from aged humans as compared to young controls. This is associated with increased leptin-induced phosphorylation of STAT3 (signal transducer and activator of transcription-3) in B cells from aged humans as compared to young subjects. These data suggest that leptin-induced B cell-derived proinflammatory cytokines may play a role in chronic inflammation associated with human aging.
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BACKGROUND: With the recent implementation of bundling reimbursement policy, the use of intravenous (IV) iron preparations for the management of anemia in the end-stage renal disease (ESRD) population has dramatically increased. Iron overload increases the risk of infections in individuals with or without kidney disease. IV iron administration in ESRD patients impairs bacteriocidal capacity of polymorphonuclear leukocytes (PMNs) against Escherichia coli. These preparations consist of an elemental iron core and a carbohydrate shell. In addition to the iron core, the carbohydrate shell may affect PMNs. We therefore examined the effect of iron sucrose, a commonly used preparation, on phagocytic capacity of PMNs from a group of normal individuals against Gram-positive (Staphylococcus aureus) and Gram-negative (E. coli) bacteria. METHODS: Iron sucrose was added to heparinized blood samples at pharmacologically-relevant concentrations and incubated for 4 and 24 h at 37°C to simulate in vivo condition. Blood samples mixed with equal volume of saline solution served as controls. To isolate the effects of the carbohydrate shell, blood samples were co-treated with the iron chelator, desferrioxamine. RESULTS: Iron sucrose caused significant PMN apoptosis and dose-dependent suppression of phagocytic function against both Gram-positive and Gram-negative bacteria. These abnormalities were prevented by desferrioxamine which precluded contribution of the carbohydrate shell to the PMN dysfunction. CONCLUSIONS: At pharmacologically-relevant concentrations, iron sucrose promotes apoptosis and inhibits phagocytic activities of PMNs. The deleterious effect of iron sucrose is mediated by its elemental iron core, not its carbohydrate shell, and as such may be shared by other IV iron preparations.
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Anemia/terapia , Apoptosis , Compuestos Férricos/farmacología , Ácido Glucárico/farmacología , Hematínicos/farmacología , Neutrófilos/patología , Adulto , Carbohidratos/química , Escherichia coli/metabolismo , Femenino , Sacarato de Óxido Férrico , Citometría de Flujo/métodos , Humanos , Fallo Renal Crónico/complicaciones , Fallo Renal Crónico/terapia , Masculino , Neutrófilos/citología , Fagocitosis , Staphylococcus aureus/metabolismo , Factores de TiempoRESUMEN
Leptin, one of the adipokines, functions as a hormone and a cytokine. In this investigation, we show for the first time that leptin, in a concentration-dependent manner, activates human peripheral blood B cells to induce secretion of IL-6, IL-10, and TNF-α. Leptin increased B cells expressing CD25 and HLA-DR. Leptin induces phosphorylation of Janus activation kinase 2 (JAK2), signal transducer and activator of transcription 3 (STAT3), p38 mitogen-activated protein kinase (p38MAPK), and extracellular signal-regulated kinase (ERK1/2). Furthermore, leptin-induced cytokine secretion by B cells was blocked by inhibitors of JAK2, STAT3, p38MAPK, and ERK1/2. These data demonstrate that leptin activates human B cells to secrete cytokines via activation of JAK2/STAT3 and p38MAPK/ERK1/2 signaling pathways, which may contribute to its inflammatory and immunoregulatory properties.
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Linfocitos B , Leptina , Receptores de Leptina/metabolismo , Transducción de Señal/inmunología , Autoinmunidad , Linfocitos B/efectos de los fármacos , Linfocitos B/inmunología , Linfocitos B/metabolismo , Células Cultivadas , Relación Dosis-Respuesta a Droga , Ensayo de Inmunoadsorción Enzimática , Humanos , Inflamación/inmunología , Inflamación/metabolismo , Interleucina-10/análisis , Interleucina-10/biosíntesis , Interleucina-6/análisis , Interleucina-6/biosíntesis , Janus Quinasa 2/análisis , Janus Quinasa 2/biosíntesis , Leptina/farmacología , Activación de Linfocitos , Proteína Quinasa 3 Activada por Mitógenos/análisis , Proteína Quinasa 3 Activada por Mitógenos/biosíntesis , Proteína Quinasa 6 Activada por Mitógenos/análisis , Proteína Quinasa 6 Activada por Mitógenos/biosíntesis , Proteínas Quinasas Activadas por Mitógenos/análisis , Proteínas Quinasas Activadas por Mitógenos/biosíntesis , Fosforilación/efectos de los fármacos , Receptores de Leptina/inmunología , Factor de Transcripción STAT3/análisis , Factor de Transcripción STAT3/biosíntesis , Factor de Necrosis Tumoral alfa/análisis , Factor de Necrosis Tumoral alfa/biosíntesis , Proteínas Quinasas p38 Activadas por Mitógenos/análisis , Proteínas Quinasas p38 Activadas por Mitógenos/biosíntesisRESUMEN
Aging is associated with increased susceptibility to microbial infections, and monocytes play an important role in microbial defense. In this study, we have identified and compared four subpopulations of monocytes (CD14(++(high))CD16(-), CD14(+(low))CD16(-), CD14(++(high))CD16(+), and CD14(+(low))CD16(+)) in the peripheral blood of young and aged subjects with regard to their numbers, cytokine production, TLR expression, and phosphorylation of ERK1/2 in response to pam3Cys a TLR-1/2 ligand. Proportions and numbers of CD14(++(high))CD16(+) and CD14(+(low))CD16(+) monocytes were significantly increased, whereas proportions of CD14(+(low))CD16(-) monocytes were decreased in aged subjects as compared to young subjects. In aged subjects, IL-6 production by all four subsets of monocytes was significantly decreased, whereas TNF-α production was decreased in monocyte subsets, except the CD14(+(low))CD16(-) subset. A significantly reduced expression of TLR1 was observed in CD14(++(high))CD16(+) and CD14(+(low))CD16(+) monocyte subsets in aged subjects. Furthermore, following pam3Cys stimulation, ERK1/2 phosphorylation was significantly lower in CD14(+(low))CD16(+), CD14(++(high))CD16(+), and CD14(+(low))CD16(-) subsets of monocytes from aged subjects. This is the first study of four subpopulations of monocytes in aging, which demonstrates that their functions are differentially impaired with regard to the production of cytokines, expression of TLR, and signaling via the ERK-MAPK pathway. Finally, changes in the number of monocyte subsets, and impairment of TLR1 expression, TNF-α production, and EK1/2 phosphorylation was more consistent in CD16(+) monocyte subsets regardless of expression of CD14(high) or CD14(+low), therefore highlighting the significance of further subdivision of monocytes into four subpopulations.
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Envejecimiento/inmunología , Infecciones Bacterianas/inmunología , Macrófagos/inmunología , Proteína Quinasa 3 Activada por Mitógenos/metabolismo , Receptores Toll-Like/metabolismo , Adulto , Anciano , Anciano de 80 o más Años , Envejecimiento/patología , Antígenos CD/genética , Antígenos CD/metabolismo , Infecciones Bacterianas/patología , Diferenciación Celular/inmunología , Células Cultivadas , Regulación de la Expresión Génica/inmunología , Humanos , Interleucina-6/inmunología , Proteína Quinasa 3 Activada por Mitógenos/inmunología , Transducción de Señal/inmunología , Receptores Toll-Like/genética , Receptores Toll-Like/inmunologíaRESUMEN
INTRODUCTION: Streptococcus pneumoniae is one of the most common infectious pathogens in common variable immunodeficiency (CVID). Both innate and adaptive immune response appears to play a role in defense against S. pneumoniae. In mice, it has been established that TLR2 and macrophages-derived cytokines (IL-6, TNF-alpha) play a crucial role in defense against S. pneumoniae. In humans, monocyte/macrophage-derived cytokines in response to S. pneumoniae have not been studied. Patients with CVID respond poorly to Pneumovax-23 (containing all capsular polysaccharides) vaccination. FINDINGS: In this study, we show that Pneumovax-23, in a concentration and time-dependent manner, induced secretion of IL-6, IL-10, and TNF-alpha by monocytes and not by B cells or T cells from healthy controls. Furthermore, Pneumovax-23-induced secretion of IL-6 and TNF-alpha was significantly less in patients with CVID as compared with controls. In addition, Pneumovax-23-induced upregulation of TLR2 in all four subsets of monocytes; however, differences between control and CVID were not significant. CONCLUSION: Pneumovax-23-induced monocytes-derived cytokine production is impaired in CVID, which may play an important role in increased susceptibility of CVID patients to S. pneumoniae infection.
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Vacunas Bacterianas/farmacología , Inmunodeficiencia Variable Común/inmunología , Citocinas/biosíntesis , Macrófagos/metabolismo , Infecciones Neumocócicas/inmunología , Streptococcus pneumoniae/inmunología , Animales , Antígenos Bacterianos/inmunología , Linfocitos B/inmunología , Cápsulas Bacterianas/inmunología , Células Cultivadas , Inmunodeficiencia Variable Común/complicaciones , Inmunodeficiencia Variable Común/tratamiento farmacológico , Citocinas/genética , Citocinas/metabolismo , Humanos , Macrófagos/inmunología , Macrófagos/patología , Ratones , Infecciones Neumocócicas/etiología , Infecciones Neumocócicas/prevención & control , Linfocitos T/inmunología , Receptor Toll-Like 2/genética , Receptor Toll-Like 2/inmunología , Receptor Toll-Like 2/metabolismoRESUMEN
Aging is associated with progressive T-cell deficiency and increased incidence of infections, cancer and autoimmunity. In this comprehensive study, we have compared the gene expression profiles in CD8+ T cells from aged and young healthy subjects using Affymetrix microarray Human Genome U133A-2 GeneChips. A total of 5.2% (754) of the genes analyzed had known functions and displayed statistically significant age-associated expression changes. These genes were involved in a broad array of complex biological processes, mainly in nucleic acid and protein metabolism. Functional groups, in which downregulated genes were overrepresented, were the following: RNA transcription regulation, RNA and DNA metabolism, intracellular (Golgi, endoplasmic reticulum and nuclear) transportation, signaling transduction pathways (T-cell receptor, Ras/MAPK, JNK/Stat, PI3/AKT, Wnt, TGFbeta, insulin-like growth factor and insulin), and the ubiquitin cycle. In contrast, the following functional groups contained more up-regulated genes than expected: response to oxidative stress and cytokines, apoptosis, and the MAPKK signaling cascade. These age-associated gene expression changes may be responsible for impaired DNA replication, RNA transcription, and signal transduction, possibly resulting in instability of cellular and genomic integrity, and alterations of growth, differentiation, apoptosis and anergy in human aged CD8+ T cells.
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Envejecimiento , Linfocitos T CD8-positivos/citología , Linfocitos T CD8-positivos/inmunología , Expresión Génica , Adulto , Anciano , Anciano de 80 o más Años , Linfocitos T CD8-positivos/metabolismo , Femenino , Humanos , Masculino , Análisis de Secuencia por Matrices de Oligonucleótidos , Transducción de Señal , Adulto JovenRESUMEN
BACKGROUND: End-stage renal disease (ESRD) results in increased susceptibility to infections, impaired response to vaccination and diffuse B-cell lymphopenia. However, the precise nature and mechanism of ESRD-induced B-cell lymphopenia remains unclear. Therefore, we studied the distribution of major B-cell subsets, B-cell growth, differentiation and survival factors, IL-7 and BAFF, and their receptors in 21 haemodialysis patients and 21 controls. METHODS: Innate B1 cells (CD19+, CD5+), conventional B2 cells (CD19+, CD5-), newly formed transitional B cells (CD19+, CD10+, CD27-), naïve B cells (CD19+, CD27-) and memory B cells (CD19+, CD27+) and BAFF receptor were quantified by flow cytometry. Plasma IL-7, BAFF, IL-6, TNF-alpha and IL-10 were measured by ELISA. RESULTS: The ESRD group exhibited significant reductions of all B-cell subpopulations except for transitional B cells that were less severely affected. No significant difference was found in B-cell apoptosis between the ESRD and control groups. Moreover, plasma IL-7 and BAFF levels were elevated in ESRD patients, therefore excluding their deficiencies as a possible culprit. However, BAFF receptor expression was significantly reduced in transitional but not mature B cells in the ESRD group. Interestingly, B-cell activation with the TLR9 agonist resulted in significantly greater production of IL-6 and TNF alpha but not IL-10 in the ESRD group. CONCLUSIONS: Thus, despite elevation of B-cell growth, differentiation and survival factors, ESRD patients exhibited diffuse reduction of B-cell subpopulations. This was associated with the down-regulation of BAFF receptor in transitional B cells. The latter can, in part, contribute to B-cell lymphopenia by promoting resistance to the biological actions of BAFF that is a potent B-cell differentiation and survival factor.
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Factor Activador de Células B/metabolismo , Receptor del Factor Activador de Células B/metabolismo , Subgrupos de Linfocitos B/patología , Interleucina-7/metabolismo , Fallo Renal Crónico/metabolismo , Fallo Renal Crónico/patología , Adulto , Anciano , Apoptosis/fisiología , Linfocitos B/patología , Estudios de Casos y Controles , Diferenciación Celular/fisiología , Proliferación Celular , Femenino , Humanos , Interleucina-6/metabolismo , Fallo Renal Crónico/terapia , Masculino , Persona de Mediana Edad , Diálisis Renal , Receptor Toll-Like 9/metabolismo , Factor de Necrosis Tumoral alfa/metabolismoRESUMEN
Prostate cancer is one of the most frequent cancers among men in the United States. In the current study, we examined the susceptibility of the human LNCaP prostate cancer cells to the apoptotic effect of marina crystal minerals (MCM), a crystallized mixture of minerals and trace elements from sea water, in vitro. Cancer cells were cultured with MCM at different concentrations (0-1000 microg/ml). At day 3, MCM exhibited a marked apoptotic effect on LNCaP cells as measured by propidium iodide (PI) and Giemsa-stained cytospin preparations. The apoptotic effect was dose-dependent: a 2-fold increase in the level of apoptosis over the control cells was observed at low concentrations of 50 microg/ml, maximial (3-fold) at a concentration of 500-1000 microg/ml. The mechanism by which MCM induces apoptosis was examined. Flow cytometery analysis demonstrated that mitochondrial membrane potential was significantly decreased post-treatment with MCM at low concentrations of 100 microg/ml. In addition, Western blot analysis showed that the level of anti-apoptotic molecule Bcl-2 in LNCaP cells decreased upon treatment with MCM. We conclude that MCM has an apoptotic effect toward human prostate cancer cells, which may suggest its broader potential as an innovative, non-toxic anti-cancer agent.
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Antineoplásicos/farmacología , Apoptosis/efectos de los fármacos , Minerales/farmacología , Neoplasias de la Próstata/patología , Western Blotting , Línea Celular Tumoral , Forma de la Célula/efectos de los fármacos , Relación Dosis-Respuesta a Droga , Citometría de Flujo , Humanos , Masculino , Potencial de la Membrana Mitocondrial/efectos de los fármacos , Minerales/aislamiento & purificación , Neoplasias de la Próstata/metabolismo , Proteínas Proto-Oncogénicas c-bcl-2/metabolismo , Agua de Mar/químicaRESUMEN
BACKGROUND: Several studies have examined the correlation between iron oxidation and H(2)O(2) degradation. The present study was carried out to examine the protective effects of MRN-100 against stress-induced apoptosis in murine splenic cells in vitro. MRN-100, or HydroFerrate fluid, is an iron-based beverage composed of bivalent and trivalent ferrates. METHODS: Splenic lymphocytes from mice were cultured in the presence or absence of MRN-100 for 2 hrs and were subsequently exposed to hydrogen peroxide (H(2)O(2) ) at a concentration of 25 microM for 14 hrs. Percent cell death was examined by flow cytometry and trypan blue exclusion. The effect of MRN-100 on Bcl-2 and Bax protein levels was determined by Western blot. RESULTS: Results show, as expected, that culture of splenic cells with H(2)O(2) alone results in a significant increase in cell death (apoptosis) as compared to control (CM) cells. In contrast, pre-treatment of cells with MRN-100 followed by H(2)O(2) treatment results in significantly reduced levels of apoptosis.In addition, MRN-100 partially prevents H(2)O(2) -induced down-regulation of the anti-apoptotic molecule Bcl-2 and upregulation of the pro-apoptotic molecule Bax. CONCLUSION: Our findings suggest that MRN-100 may offer a protective effect against oxidative stress-induced apoptosis in lymphocytes.
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Apoptosis/efectos de los fármacos , Compuestos de Hierro/farmacología , Linfocitos/efectos de los fármacos , Estrés Oxidativo/efectos de los fármacos , Bazo/citología , Animales , Bebidas , Western Blotting , Calcio/metabolismo , Células Cultivadas , Femenino , Citometría de Flujo , Peróxido de Hidrógeno/farmacología , Compuestos de Hierro/administración & dosificación , Linfocitos/metabolismo , Ratones , Ratones Endogámicos C57BL , Óxido Nítrico/metabolismo , Oxidación-Reducción , Proteínas Proto-Oncogénicas c-bcl-2/metabolismo , Azul de Tripano/metabolismo , Proteína X Asociada a bcl-2/metabolismoRESUMEN
The immune responses of naive and different memory subsets of CD4(+) and CD8(+) T cells to human herpesvirus 6 (HHV-6) have not been previously investigated. We show that HHV-6A induces cell division, as measured by 5,6-carboxyfluorescein succinimidyl ester dye and flow cytometry, predominantly in two populations of effector memory CD4(+) and CD8(+) T cells (T(EM) and T(EMRA)); naïve (T(N)) and central memory (T(CM)) CD4(+) and CD8(+) T cells showed almost no cell division. In contrast, HHV-6A induced apoptosis primarily in T(N) and T(CM) CD4(+) and CD8(+) T cells, whereas T(EM) and T(EMRA) CD4(+) and CD8(+) T cells were resistant to HHV-6A-induced apoptosis. HHV-6A-induced apoptosis was associated with activation of caspase-8, caspase-9, and caspase-3, suggesting the involvement of death receptor and mitochondrial signaling pathways. In addition, HHV-6A induced secretion of interleukin-6 (IL-6), tumor necrosis factor alpha (TNF-alpha), IL-8, and gamma interferon by peripheral blood mononuclear cells; TNF-alpha secretion was observed exclusively from CCR7(+) (T(N) plus T(CM)) CD4(+) T cells. These data show that HHV-6 differentially influences the functions of naïve T cells and different subsets of memory CD4(+) and CD8(+) T cells, which in part may be due to differential susceptibility to HHV-6A-induced apoptosis.
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Apoptosis/inmunología , Linfocitos T CD4-Positivos/inmunología , Linfocitos T CD8-positivos/inmunología , Herpesvirus Humano 6/inmunología , Inmunidad Innata/inmunología , Memoria Inmunológica/inmunología , Subgrupos de Linfocitos T/inmunología , Linfocitos T CD4-Positivos/citología , Linfocitos T CD4-Positivos/metabolismo , Linfocitos T CD8-positivos/citología , Linfocitos T CD8-positivos/metabolismo , División Celular/inmunología , Células Cultivadas , Citocinas/biosíntesis , Citocinas/inmunología , Humanos , Proteína Cofactora de Membrana/inmunología , Proteínas Proto-Oncogénicas c-bcl-2/metabolismo , Subgrupos de Linfocitos T/citología , Regulación hacia Arriba/inmunologíaRESUMEN
We have recently demonstrated that non-metastatic human breast cancer cell lines undergo apoptosis following phagocytosis of S. cerevisiae. In this study, we investigated the apoptotic effect of heat-killed yeast against human metastatic breast cancer (MBC) cells, MDA-MB-231 in vitro, and the underlying mechanistic bases of this effect. Results show that monolayer MDA-MB-231 cells phagocytized yeast (50% at 16 h) and underwent apoptosis (32% compared with 7.6% of untreated cells, representing a 4.2-fold increase). The increase in apoptosis was associated with an elevation of [Ca2+]I. Addition of 2-aminoethoxydiphenyl borate (2APB), a pharmacological inhibitor of Ca2+ release from the endoplasmic reticulum, effectively diminished yeast-induced apoptosis. Furthermore, yeast caused a substantial decrease in expression of Bcl-2 and an increase in Bax resulting in alteration in the Bax:Bcl-2 ratio. However, yeast had no effect on NO levels. In conclusion, yeast induces apoptosis of human MBC cells in vitro by a mechanism involving intracellular Ca2+ and Bax:Bcl-2.
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Apoptosis/fisiología , Neoplasias de la Mama/microbiología , Calcio/metabolismo , Proteínas Proto-Oncogénicas c-bcl-2/metabolismo , Saccharomyces cerevisiae/fisiología , Proteína X Asociada a bcl-2/metabolismo , Western Blotting , Neoplasias de la Mama/metabolismo , Neoplasias de la Mama/patología , Línea Celular Tumoral , Citoplasma/metabolismo , Femenino , Citometría de Flujo , Humanos , Fagocitosis/fisiologíaRESUMEN
Previous studies have shown that thymic extracts possess antitumor and antimetastatic properties, but the mechanisms are not completely understood. Therefore, in this study the ability of the gross thymic extract Thymax to induce apoptosis in human breast cancer cell line (MCF-7) cells in vitro was evaluated. Tumor cells were cultured with different concentrations of Thymax for 24 h and the apoptotic response was assessed by propidium iodide and TUNEL assays. Activation of caspases and changes in mitochondrial membrane potential (MMP) were monitored by flow cytometry and the expression of Bcl-2 and Bax was determined by Western blot analysis. Thymax induced apoptosis in monolayer MCF-7 cells in a dose-dependent manner; at concentrations of 2.5, 5 and 10% (v/v) it caused 9%, 10% and 25% apoptosis, respectively, as compared to 6% for control cancer cells without treatment. The induction of apoptosis by Thymax was associated with activation of caspases 8 and 9, and the addition of a pan caspase inhibitor partially inhibited Thymax-induced apoptosis by 20%. In addition, the MMP was decreased significantly at Thymax concentrations of 5%-20%, which was associated with a decrease in the protein expression of Bcl-2 and an increase in Bax. These results suggest that Thymax exerts its effects via the mitochondrial pathway of apoptosis and may represent a new class of adjuvants for the treatment of breast cancer.
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Apoptosis/efectos de los fármacos , Neoplasias de la Mama/tratamiento farmacológico , Neoplasias de la Mama/patología , Mitocondrias/efectos de los fármacos , Extractos del Timo/farmacología , Clorometilcetonas de Aminoácidos/farmacología , Neoplasias de la Mama/metabolismo , Caspasa 8/metabolismo , Caspasa 9/metabolismo , Inhibidores de Caspasas , Línea Celular Tumoral , Activación Enzimática/efectos de los fármacos , Citometría de Flujo , Humanos , Potencial de la Membrana Mitocondrial/efectos de los fármacos , Mitocondrias/fisiología , Proteínas Proto-Oncogénicas c-bcl-2/biosíntesis , Proteína X Asociada a bcl-2/biosíntesisRESUMEN
BACKGROUND: MGN-3/Biobran, a modified form of arabinoxylan from rice bran, is a potent biological response modifier (BRM). Our previous studies demonstrated that MGN-3 sensitizes human leukemia cells to death receptor [CD95]-induced apoptosis [Ghoneum M, Gollapudi S. MGN-3 sensitizes human T cell leukemia cells to death receptor (CD95)-induced apoptosis. Cancer Lett 2003;201:41-9]. In this study, we evaluated the chemo-sensitizing activity of MGN-3 against human breast cancer cells (BCCs) in vitro. METHODS: BCCs (MCF-7 and HCC70 cells) were cultured with different concentrations of daunorubicin (DNR) (from 1x10(-9) to 1x10(-6)M) in the presence or absence of selected concentrations of MGN-3 (100-1000mug/ml) for 3 days. Cancer cell survival was determined by MTT assay and drug accumulation was determined by flow cytometry. RESULTS: Treatment with MGN-3 increased susceptibility of BCCs to DNR (5.5-fold for MCF-7 and 2.5-fold for HCC70 cells) as compared to BCCs treated with DNR alone. The sensitizing effect of MGN-3 was associated with increased accumulation of DNR in cancer cells. CONCLUSIONS: Our data demonstrate that MGN-3 is an effective chemo-sensitizer and may represent a potential novel adjuvant for the treatment of breast cancer.
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Antineoplásicos/uso terapéutico , Neoplasias de la Mama/tratamiento farmacológico , Daunorrubicina/uso terapéutico , Xilanos/uso terapéutico , Línea Celular Tumoral , Supervivencia Celular , Quimioterapia Adyuvante , Daunorrubicina/farmacocinética , Relación Dosis-Respuesta a Droga , Sinergismo Farmacológico , HumanosRESUMEN
Aging is associated with a decrease in naïve (T(N)) and central memory (T(CM)), and an accumulation of effector memory (T(EM) and T(EMRA)) T cell subsets. Previously, we have demonstrated an increased sensitivity of T(N) and T(CM) CD4+ and CD8+ T cells in aging to TNF-alpha-induced apoptosis. In this investigation, we examined whether similar differential sensitivity is applicable to CD95-mediated apoptosis. We show that T(N) and T(CM) CD4+ and CD8+ T cells from aged subjects are significantly more sensitive to CD95-mediated apoptosis. Increased apoptosis is associated with increased activation of caspase-8 and caspase-3. Both caspase-8 and caspase-3 inhibitors blocked CD95-mediated apoptosis and activation of caspase-8 and caspase-3 in T(N) and T(CM) CD4+ and CD8+ T cells. No significant difference was observed in apoptosis or in activation of caspase-8 and caspase-3 in T(EM) and T(EMRA) CD4+ and CD8+ T cells between young and aged subjects; both populations were relatively and comparably resistant to CD95-mediated apoptosis and caspase activation. No correlation was observed between the sensitivity/resistance of any of the subsets of CD4+ or CD8+T cells to CD95-mediated apoptosis and the expression of CD95. Our data suggest that increased CD95-mediated apoptosis of T(N) and T(CM) CD8+ and CD4+ T cells may play a role in their decline in human aging.