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1.
J Immunol ; 212(8): 1257-1267, 2024 Apr 15.
Artigo em Inglês | MEDLINE | ID: mdl-38560813

RESUMO

The Canadian Society for Immunology (CSI) established a formal Equity, Diversity, and Inclusion (EDI) Committee with the goal of providing EDI advocacy and leadership within the CSI, as well as in the broader scientific community. A first task of this committee was to review the publicly available historical data on gender representation within the CSI's membership, leadership, award recipients, and conference chairs/presenters as a step in establishing a baseline reference point and monitoring the trajectory of future success in achieving true inclusion. We found that, except for overall membership and a specific subset of awards, all categories showed a historical bias toward men, particularly prior to 2010. Bias persists in various categories, evident even in recent years. However, we note an encouraging trend toward greater gender parity, particularly in the roles of President, symposium presenters, and workshop chairs, especially from 2017 onward. We present these findings as well as our recommendations to enhance inclusivity. These include a more comprehensive collection and secure storage of self-identification data, emphasis on EDI as an essential component of all annual meeting activities, and innovative measures of outreach, collaboration, and leadership with the aim of making the CSI a model for improving EDI in other professional research societies.


Assuntos
Distinções e Prêmios , Liderança , Feminino , Humanos , Masculino , Canadá , Estudos Retrospectivos , Sociedades Médicas
2.
Curr Oncol ; 30(7): 6411-6431, 2023 07 05.
Artigo em Inglês | MEDLINE | ID: mdl-37504332

RESUMO

FISH cytogenetics, TP53 sequencing, and IGHV mutational status are increasingly used as prognostic and predictive markers in chronic lymphocytic leukemia (CLL), particularly as components of the CLL International Prognostic Index (CLL-IPI) and in directing therapy with novel agents. However, testing outside of clinical trials is not routinely available in Canada. As a centralized CLL clinic at CancerCare Manitoba, we are the first Canadian province to evaluate clinical outcomes and survivorship over a long period of time, incorporating the impact of molecular testing and the CLL-IPI score. We performed a retrospective analysis on 1315 patients diagnosed between 1960 and 2018, followed over a 12-year period, where 411 patients had molecular testing and 233 patients had a known CLL-IPI score at the time of treatment. Overall, 40.3% (n = 530) of patients received treatment, and 47.5% (n = 252) of patients received multiple lines of therapy. High-risk FISH and CLL-IPI (4-10) were associated with higher mortality (HR 2.03, p = 0.001; HR 2.64, p = 0.002), consistent with other studies. Over time, there was an increase in the use of targeted agents in treated patients. The use of Bruton's tyrosine kinase inhibitors improved survival in patients with unmutated IGHV and/or TP53 aberrations (HR 2.20, p = 0.001). The major cause of death in patients who received treatment was treatment/disease-related (32%, n = 42) and secondary malignancies (57%, n = 53) in those who were treatment-naïve. Our data demonstrate the importance of molecular testing in determining survivorship in CLL and underpinning the likely immune differences in outcomes for those treated for CLL.


Assuntos
Leucemia Linfocítica Crônica de Células B , Humanos , Leucemia Linfocítica Crônica de Células B/tratamento farmacológico , Leucemia Linfocítica Crônica de Células B/genética , Estudos Retrospectivos , Canadá , Prognóstico , Mutação
3.
Leuk Res ; 131: 107315, 2023 08.
Artigo em Inglês | MEDLINE | ID: mdl-37263073

RESUMO

Interleukin-16 (IL-16) is a novel biomarker that has been implicated in many cancers as well as inflammatory diseases. In this study, we examined plasma levels of 30 cytokines and chemokines in chronic lymphocytic leukemia (CLL) and monoclonal B cell lymphocytosis (MBL) patients, and examined their association with disease stage, CLL biomarkers and T cell subsets. Interleukin 16 (IL-16) was identified as a relatively uncharacterized cytokine significantly elevated in CLL patients compared to healthy controls and MBL patients. Plasma levels of IL-16 were significantly elevated by Rai stage 0, increased by Rai stage 3-4, correlated strongly with lymphocyte count and were decreased after Ibrutinib treatment. CLL cells expressed IL-16 mRNA and spontaneously secreted IL-16 in vitro. CLL cells express IL-16 mRNA at significantly higher levels in lymphoid tissues than blood, and we observed that IL-16 release was increased in co-cultures of CLL and autologous CD4 + T cells. Elevated plasma IL-16 levels were associated with abnormalities in the immune microenvironment including multiple inflammatory cytokines and chemokines and expansion of type 1 follicular helper T cells. Taken together, our results identify IL-16 as a novel biomarker in CLL with potential functional roles in cellular interactions between CLL cells and T cells.


Assuntos
Leucemia Linfocítica Crônica de Células B , Linfocitose , Humanos , Leucemia Linfocítica Crônica de Células B/terapia , Interleucina-16 , Contagem de Linfócitos , Efeitos Psicossociais da Doença , Microambiente Tumoral
4.
Front Immunol ; 14: 1115244, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-37234154

RESUMO

Phosphoinositide 3-kinase delta (PI3Kδ) plays key roles in normal B cell activation and is chronically activated in malignant B cells. Targeting of PI3Kδ using FDA-approved drugs Idelalisib or Umbralisib has shown efficacy in treatment of multiple B cell malignancies. Duvelisib, an inhibitor targeting both PI3Kδ and PI3Kγ (PI3Kδγi) has also been used for treatment of several leukemias and lymphomas and was suggested to offer potential additional benefits in supressing T cell and inflammatory responses. Transcriptomics analyses indicated that while most B cell subsets predominantly express PI3Kδ, plasma cells upregulate PI3Kγ. We thus assessed whether PI3Kδγi treatment can impact chronic B cell activation in the context of an autoantibody-mediated disease. Using the TAPP1R218LxTAPP2R211L (TAPP KI) mouse model of lupus-like disease driven by dysregulated PI3K pathway activity, we performed 4 week PI3Kδγi treatments and found significant reduction in CD86+ B cells, germinal center B cells, follicular helper T cells and plasma cells in multiple tissues. This treatment also significantly attenuated the abnormally elevated serum levels of IgG isotypes observed in this model. The profile of autoantibodies generated was markedly altered by PI3Kδγi treatment, with significant reductions in IgM and IgG targeting nuclear antigens, matrix proteins and other autoantigens. Kidney pathology was also impacted, with reduced IgG deposition and glomerulonephritis. These results indicate that dual inhibition of PI3Kδ and PI3Kγ can target autoreactive B cells and may have therapeutic benefits in autoantibody-mediated disease.


Assuntos
Autoanticorpos , Fosfatidilinositol 3-Quinases , Camundongos , Animais , Fosfatidilinositol 3-Quinases/metabolismo , Classe I de Fosfatidilinositol 3-Quinases/metabolismo , Modelos Animais de Doenças , Autoanticorpos/metabolismo , Fosfatidilinositóis , Imunoglobulina G
6.
Cell Tissue Res ; 390(3): 429-439, 2022 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-36129532

RESUMO

Barth syndrome (BTHS) is a rare X-linked genetic disease caused by mutations in TAFAZZIN. The tafazzin (Taz) protein is a cardiolipin remodeling enzyme required for maintaining mitochondrial function. Patients with BTHS exhibit impaired mitochondrial respiratory chain and metabolic function and are susceptible to serious infections. B lymphocytes (B cells) play a vital role in humoral immunity required to eradicate circulating antigens from pathogens. Intact mitochondrial respiration is required for proper B-cell function. We investigated whether Taz deficiency in mouse B cells altered their response to activation by anti-cluster of differentiation 40 (anti-CD40) + interleukin-4 (IL-4). B cells were isolated from 3-4-month-old wild type (WT) or tafazzin knockdown (TazKD) mice and were stimulated with anti-CD40 + IL-4 for 24 h and cellular bioenergetics, surface marker expression, proliferation, antibody production, and proteasome and immunoproteasome activities determined. TazKD B cells exhibited reduced mRNA expression of Taz, lowered levels of cardiolipin, and impairment in both oxidative phosphorylation and glycolysis compared to WT B cells. In addition, anti-CD40 + IL-4 stimulated TazKD B cells expressed lower levels of the immunogenic surface markers, cluster of differentiation 86 (CD86) and cluster of differentiation 69 (CD69), exhibited a lower proliferation rate, reduced production of immunoglobulin M and immunoglobulin G, and reduced proteasome and immunoproteasome proteolytic activities compared to WT B cells stimulated with anti-CD40 + IL-4. The results indicate that Taz is required to support T-cell-dependent signaling activation of mouse B cells.


Assuntos
Aciltransferases , Linfócitos B , Síndrome de Barth , Cardiolipinas , Animais , Camundongos , Aciltransferases/deficiência , Aciltransferases/genética , Linfócitos B/metabolismo , Síndrome de Barth/genética , Síndrome de Barth/metabolismo , Cardiolipinas/metabolismo , Interleucina-4/farmacologia , Complexo de Endopeptidases do Proteassoma/metabolismo , Fatores de Transcrição/metabolismo , Antígenos CD40/metabolismo
7.
Front Immunol ; 13: 932627, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-35967356

RESUMO

Background: Despite immune cell dysregulation being an important event preceding the onset of rheumatoid arthritis (RA), the phenotype of T and B cells in preclinical RA is less understood. The aim of this study was to characterize T and B cell populations in RA patients and their autoantibody (aAb) negative and positive first-degree relatives (FDR). Methods: Cryopreserved peripheral blood mononuclear cells (PBMCs) collected at scheduled visits from aAb-(n=25), and aAb+ FDR (n=10) and RA patients (n=13) were thawed and stained using optimized antibody cocktails as per a specific 13-color T or B cell panel. Immunophenotyping was performed using a Cytoflex LX (Beckman-Coulter) flow cytometer and FlowJo software was used for analyzing the frequency of immune cell populations. Results: Multicolor flow cytometry experiments identified an increased TIGIT expression in circulating lymphocytes of aAb+ FDR and RA patients, relative to aAb- FDR (P<0.01). These TIGIT+ T cells exhibited a memory phenotype and expressed high levels of PD-1, ICOS, HLA-DR, CXCR3 and CXCR5. Moreover, increased TIGIT+ CD4 T cell frequency correlated with the frequency of PD-1+ CD4 T cells (r = 0.4705: P = 0.0043) and circulating levels of ACPA and RF. We also identified a decreased frequency of CD27+IgD- switched memory B cells in RA patients (P < 0.01), while increased frequency of TIGIT+ CD4 T cells in FDR correlated with the frequency of PD1+PTEN+ B cells (r = 0.6838, P = 0.0004) and autoantibody positivity (P = 0.01). Conclusion: We demonstrate TIGIT as a distinct CD4 T cell marker for differentiating aAb- FDR from aAb+FDR and might play a critical role in regulating T and B cell crosstalk in preclinical RA.


Assuntos
Artrite Reumatoide , Linfócitos T CD4-Positivos , Receptores Imunológicos , Artrite Reumatoide/genética , Artrite Reumatoide/imunologia , Autoanticorpos/genética , Autoanticorpos/imunologia , Linfócitos T CD4-Positivos/imunologia , Humanos , Leucócitos Mononucleares/imunologia , Receptor de Morte Celular Programada 1/imunologia , Receptores Imunológicos/genética , Receptores Imunológicos/imunologia , Subpopulações de Linfócitos T/imunologia
8.
FASEB J ; 36(8): e22443, 2022 08.
Artigo em Inglês | MEDLINE | ID: mdl-35816277

RESUMO

Barth Syndrome (BTHS) is a rare X-linked genetic disorder caused by mutation in the TAFAZZIN gene. Tafazzin (Taz) deficiency in BTHS patients results in an increased risk of infections. Mesenchymal stem cells (MSCs) are well known for their immune-inhibitory function. We examined how Taz-deficiency in murine MSCs impact their ability to modulate the function of lipopolysaccharide (LPS)-activated wild type (WT) B lymphocytes. MSCs from tafazzin knockdown (TazKD) mice exhibited a reduction in mitochondrial cardiolipin compared to wild type (WT) MSCs. However, mitochondrial bioenergetics and membrane potential were unaltered. In contrast, TazKD MSCs exhibited increased reactive oxygen species generation and increased glycolysis. The increased glycolysis was associated with an elevated proliferation, phosphatidylinositol-3-kinase expression and expression of the immunosuppressive markers indoleamine-2,3-dioxygenase, cytotoxic T-lymphocyte-associated protein 4, interleukin-10, and cluster of differentiation 59 compared to controls. Inhibition of glycolysis with 2-deoxyglucose attenuated the TazKD-mediated increased expression of cytotoxic T-lymphocyte-associated protein 4 and interleukin-10. When co-cultured with LPS-activated WT B cells, TazKD MSCs inhibited B cell proliferation and growth rate and reduced B cell secretion of immunoglobulin M compared to controls. In addition, co-culture of LPS-activated WT B cells with TazKD MSCs promoted B cell differentiation toward interleukin-10 secreting plasma cells and B regulatory cells compared to controls. The results indicate that Taz deficiency in MSCs promote reprogramming of activated B lymphocytes toward immunosuppressive phenotypes.


Assuntos
Síndrome de Barth , Células-Tronco Mesenquimais , Aciltransferases/genética , Animais , Linfócitos B/metabolismo , Síndrome de Barth/genética , Síndrome de Barth/metabolismo , Interleucina-10/genética , Lipopolissacarídeos/toxicidade , Células-Tronco Mesenquimais/metabolismo , Camundongos , Fenótipo , Fatores de Transcrição/metabolismo
10.
FASEB J ; 35(12): e22023, 2021 12.
Artigo em Inglês | MEDLINE | ID: mdl-34767647

RESUMO

B lymphocytes are responsible for humoral immunity and play a key role in the immune response. Optimal mitochondrial function is required to support B cell activity during activation. We examined how deficiency of tafazzin, a cardiolipin remodeling enzyme required for mitochondrial function, alters the metabolic activity of B cells and their response to activation by lipopolysaccharide in mice. B cells were isolated from 3-month-old wild type or tafazzin knockdown mice and incubated for up to 72 h with lipopolysaccharide and cell proliferation, expression of cell surface markers, secretion of antibodies and chemokines, proteasome and immunoproteasome activities, and metabolic function determined. In addition, proteomic analysis was performed to identify altered levels of proteins involved in survival, immunogenic, proteasomal and mitochondrial processes. Compared to wild type lipopolysaccharide activated B cells, lipopolysaccharide activated tafazzin knockdown B cells exhibited significantly reduced proliferation, lowered expression of cluster of differentiation 86 and cluster of differentiation 69 surface markers, reduced secretion of immunoglobulin M antibody, reduced secretion of keratinocytes-derived chemokine and macrophage-inflammatory protein-2, reduced proteasome and immunoproteasome activities, and reduced mitochondrial respiration and glycolysis. Proteomic analysis revealed significant alterations in key protein targets that regulate cell survival, immunogenicity, proteasomal processing and mitochondrial function consistent with the findings of the above functional studies. The results indicate that the cardiolipin transacylase enzyme tafazzin plays a key role in regulating mouse B cell function and metabolic activity during activation through modulation of mitochondrial function.


Assuntos
Aciltransferases/fisiologia , Linfócitos B/patologia , Glicólise , Lipopolissacarídeos/toxicidade , Mitocôndrias/patologia , Proteoma/metabolismo , Animais , Linfócitos B/efeitos dos fármacos , Linfócitos B/imunologia , Linfócitos B/metabolismo , Feminino , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/imunologia , Mitocôndrias/metabolismo , Proteoma/análise , Proteoma/efeitos dos fármacos
11.
bioRxiv ; 2021 Sep 08.
Artigo em Inglês | MEDLINE | ID: mdl-34729562

RESUMO

Barth Syndrome (BTHS) is a rare X-linked genetic disorder caused by mutation in the TAFAZZIN gene which encodes the cardiolipin (CL) transacylase tafazzin (Taz). Taz deficiency in BTHS patients results in reduced CL in their tissues and a neutropenia which contributes to the risk of infections. However, the impact of Taz deficiency in other cells of the immune system is poorly understood. Mesenchymal stem cells (MSCs) are well known for their immune inhibitory function. We examined whether Taz-deficiency in murine MSCs impacted their ability to modulate lipopolysaccharide (LPS)-activated wild type (WT) murine B lymphocytes. MSCs from tafazzin knockdown (TazKD) mice exhibited a 50% reduction in CL compared to wild type (WT) MSCs. However, mitochondrial oxygen consumption rate and membrane potential were unaltered. In contrast, TazKD MSCs exhibited increased glycolysis compared to WT MSCs and this was associated with elevated proliferation, phosphatidylinositol-3-kinase expression and expression of the immunosuppressive markers indoleamine-2,3-dioxygenase, cytotoxic T-lymphocyte-associated protein 4, interleukin-10, and cluster of differentiation 59. When co-cultured with LPS-activated WT B cells, TazKD MSCs inhibited B cell proliferation and growth rate and reduced B cell secretion of IgM to a greater extent than B cells co-cultured with WT MSCs. In addition, co-culture of LPS-activated WT B cells with TazKD MSCs induced B cell differentiation toward potent immunosuppressive phenotypes including interleukin-10 secreting plasma cells and B regulatory cells compared to activated B cells co-cultured with WT MSCs. These results indicate that Taz deficiency in MSCs enhances MSCs-mediated immunosuppression of activated B lymphocytes.

12.
J Immunol ; 207(5): 1401-1410, 2021 09 01.
Artigo em Inglês | MEDLINE | ID: mdl-34380646

RESUMO

PI3Kδ is critical in generating humoral and regulatory immune responses. In this study, we determined the impact of PI3Kδ in immunity to Trypanosoma congolense, an African trypanosome that can manipulate and evade Ab responses critical for protection. Upon infection with T. congolense, PI3KδD910A mice lacking PI3Kδ activity paradoxically show a transient enhancement in early control of parasitemia, associated with impaired production of regulatory IL-10 by B cells in the peritoneum. C57BL/6 wild-type (WT) mice treated with the PI3Kδ inhibitor (PI3Kδi) Idelalisib showed a similar transient decrease in parasitemia associated with reduced IL-10. Strikingly, however, we find that PI3KδD910A mice were ultimately unable to control this infection, resulting in uncontrolled parasitemia and death within 2 wk. Assessment of humoral responses revealed delayed B cell activation, impaired germinal center responses, and compromised Ab responses to differing degrees in PI3KδD910A and PI3Kδi-treated mice. To test the role of Abs, we administered serum from WT mice to PI3KδD910A mice and found that lethality was prevented by postinfection serum. Interestingly, serum from naive WT mice provided partial protection to PI3KδD910A mutants, indicating an additional role for natural Abs. Together our findings suggest that although PI3Kδ drives immune regulatory responses that antagonize early control of parasite growth in the peritoneum, it is also required for generation of Abs that are critical for protection from systemic trypanosome infection. The essential role of PI3Kδ for host survival of African trypanosome infection contrasts with findings for other pathogens such as Leishmania, underlining the critical importance of PI3Kδ-dependent humoral immunity in this disease.


Assuntos
Linfócitos B/imunologia , Classe I de Fosfatidilinositol 3-Quinases/metabolismo , Trypanosoma congolense/fisiologia , Tripanossomíase Africana/imunologia , Animais , Classe I de Fosfatidilinositol 3-Quinases/genética , Imunidade Humoral , Imunomodulação , Interleucina-10/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Parasitemia
13.
Front Oncol ; 11: 674492, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33996605

RESUMO

Follicular helper T cells (TFH) have specialized properties in promoting normal B cell activation but their role in chronic lymphocytic leukemia (CLL) is unknown. We find that TFH cells are elevated in CLL patients and are phenotypically abnormal, expressing higher levels of PD-1, TIGIT, CD40L, IFNγ and IL-21, and exhibiting abnormal composition of TFH1, TFH2 and TFH17 subsets. Frequencies of CD4-positive T cells expressing TFH1 markers and IL-21 were positively correlated with patient lymphocyte counts and RAI stage, suggesting that accumulation of abnormal TFH cells is concomitant with expansion of the leukemic B cell clone. Treatment with ibrutinib led to normalization of TFH frequencies and phenotype. TFH cells identified in CLL bone marrow display elevated expression of several functional markers compared to blood TFH cells. CLL T cell-B cell co-culture experiments revealed a correlation of patient TFH frequencies with functional ability of their CD4-positive T cells to promote CLL proliferation. Conversely, CLL cells can preferentially activate the TFH cell subset in co-culture. Together our results indicate that CLL development is associated with expansion of abnormal TFH populations that produce elevated levels of cytokines and costimulatory molecules which may help support CLL proliferation.

14.
Int J Mol Sci ; 22(4)2021 Feb 12.
Artigo em Inglês | MEDLINE | ID: mdl-33673180

RESUMO

Bam32 (B cell adaptor molecule of 32 kDa) functions in the immune responses of various leukocytes. However, the role of neutrophil Bam32 in inflammation is entirely unknown. Here, we determined the role of Bam32 in chemokine CXCL2-induced neutrophil chemotaxis in three mouse models of neutrophil recruitment. By using intravital microscopy in the mouse cremaster muscle, we found that transmigrated neutrophil number, neutrophil chemotaxis velocity, and total neutrophil chemotaxis distance were increased in Bam32-/- mice when compared with wild-type (WT) mice. In CXCL2-induced mouse peritonitis, the total emigrated neutrophils were increased in Bam32-/- mice at 2 but not 4 h. The CXCL2-induced chemotaxis distance and migration velocity of isolated Bam32-/- neutrophils in vitro were increased. We examined the activation of small GTPases Rac1, Rac2, and Rap1; the levels of phospho-Akt2 and total Akt2; and their crosstalk with Bam32 in neutrophils. The deficiency of Bam32 suppressed Rap1 activation without changing the activation of Rac1 and Rac2. The pharmacological inhibition of Rap1 by geranylgeranyltransferase I inhibitor (GGTI298) increased WT neutrophil chemotaxis. In addition, the deficiency of Bam32, as well as the inhibition of Rap1 activation, increased the levels of CXCL2-induced Akt1/2 phosphorylation at Thr308/309 in neutrophils. The inhibition of Akt by SH-5 attenuated CXCL2-induced adhesion and emigration in Bam32-/- mice. Together, our results reveal that Bam32 has a suppressive role in chemokine-induced neutrophil chemotaxis by regulating Rap1 activation and that this role of Bam32 in chemokine-induced neutrophil recruitment relies on the activation of PI3K effector Akt.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Quimiocinas/metabolismo , Lipoproteínas/metabolismo , Infiltração de Neutrófilos , Neutrófilos/metabolismo , Transdução de Sinais , Proteínas Adaptadoras de Transdução de Sinal/genética , Animais , Quimiocinas/genética , Lipoproteínas/genética , Camundongos , Camundongos Knockout , Fosfatidilinositol 3-Quinases/genética , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/genética , Proteínas Proto-Oncogênicas c-akt/metabolismo , Proteínas rho de Ligação ao GTP/genética , Proteínas rho de Ligação ao GTP/metabolismo
15.
J Immunol ; 206(5): 1013-1026, 2021 03 01.
Artigo em Inglês | MEDLINE | ID: mdl-33462138

RESUMO

There is currently no effective vaccine against leishmaniasis because of the lack of sufficient knowledge about the Ags that stimulate host-protective and long-lasting T cell-mediated immunity. We previously identified Leishmania phosphoenolpyruvate carboxykinase (PEPCK, a gluconeogenic enzyme) as an immunodominant Ag that is expressed by both the insect (promastigote) and mammalian (amastigote) stages of the parasite. In this study, we investigated the role of PEPCK in metabolism, virulence, and immunopathogenicity of Leishmania major We show that targeted loss of PEPCK results in impaired proliferation of L. major in axenic culture and bone marrow-derived macrophages. Furthermore, the deficiency of PEPCK results in highly attenuated pathology in vivo. BALB/c mice infected with PEPCK-deficient parasites failed to develop any cutaneous lesions despite harboring parasites at the cutaneous site of infection. This was associated with a dramatic reduction in the frequency of cytokine (IFN-γ, IL-4, and IL-10)-producing CD4+ T cells in spleens and lymph nodes draining the infection site. Cells from mice infected with PEPCK-deficient parasites also produced significantly low levels of these cytokines into the culture supernatant following in vitro restimulation with soluble Leishmania Ag. PEPCK-deficient parasites exhibited significantly greater extracellular acidification rate, increased proton leak, and decreased ATP-coupling efficiency and oxygen consumption rates in comparison with their wild-type and addback counterparts. Taken together, these results show that PEPCK is a critical metabolic enzyme for Leishmania, and its deletion results in altered metabolic activity and attenuation of virulence.


Assuntos
Leishmania major/metabolismo , Leishmania major/patogenicidade , Leishmaniose Cutânea/parasitologia , Fosfoenolpiruvato/metabolismo , Fatores de Virulência/metabolismo , Animais , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD4-Positivos/parasitologia , Citocinas/imunologia , Feminino , Imunidade Celular/imunologia , Leishmania major/imunologia , Leishmaniose Cutânea/imunologia , Macrófagos/parasitologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos SCID , Camundongos Transgênicos , Fosfoenolpiruvato/imunologia , Proteínas de Protozoários/imunologia , Proteínas de Protozoários/metabolismo , Fatores de Virulência/imunologia
16.
J Immunol ; 205(12): 3237-3245, 2020 12 15.
Artigo em Inglês | MEDLINE | ID: mdl-33288538

RESUMO

PI3Ks activate critical signaling cascades and have multifaceted regulatory functions in the immune system. Loss-of-function and gain-of-function mutations in the PI3Kδ isoform have revealed that this enzyme can substantially impact immune responses to infectious agents and their products. Moreover, reports garnered from decades of infectious disease studies indicate that pharmacologic inhibition of the PI3K pathway could potentially be effective in limiting the growth of certain microbes via modulation of the immune system. In this review, we briefly highlight the development and applications of PI3K inhibitors and summarize data supporting the concept that PI3Kδ inhibitors initially developed for oncology have immune regulatory potential that could be exploited to improve the control of some infectious diseases. This repurposing of existing kinase inhibitors could lay the foundation for alternative infectious disease therapy using available therapeutic agents.


Assuntos
Doenças Transmissíveis/tratamento farmacológico , Reposicionamento de Medicamentos , Terapia de Alvo Molecular , Fosfatidilinositol 3-Quinases/imunologia , Inibidores de Fosfoinositídeo-3 Quinase/uso terapêutico , Animais , Doenças Transmissíveis/imunologia , Humanos
18.
Front Immunol ; 11: 1028, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32536926

RESUMO

B cell adaptor molecule of 32 kDa (Bam32), known as dual adapter for phosphotyrosine and 3-phosphoinositides 1 (DAPP1), has been implicated in regulating lymphocyte proliferation and recruitment during inflammation. However, its role in neutrophils during inflammation remains unknown. Using intravital microscopy, we examined the role of Bam32 in formyl peptide receptor agonist WKYMVm-induced permeability changes in post-capillary venules and assessed simultaneously neutrophil adhesion and emigration in cremaster muscles of Bam32-deficient (Bam32-/-) and wild-type (WT) control mice. We observed significantly reduced WKYMVm-induced microvascular hyperpermeability accompanied by markedly decreased neutrophil emigration in Bam32-/- mice. The Bam32-specific decrease in WKYMVm-induced hyperpermeability was neutrophil-dependent as this was verified in bone marrow transplanted chimeric mice. We discovered that Bam32 was critically required for WKYMVm-induced intracellular and extracellular production of reactive oxygen species (ROS) in neutrophils. Pharmacological scavenging of ROS eliminated the differences in WKYMVm-induced hyperpermeability between Bam32-/- and WT mice. Deficiency of Bam32 decreased WKYMVm-induced ERK1/2 but not p38 or JNK phosphorylation in neutrophils. Inhibition of ERK1/2 signaling cascade suppressed WKYMVm-induced ROS generation in WT neutrophils and microvascular hyperpermeability in WT mice. In conclusion, our study reveals that Bam32-dependent, ERK1/2-involving ROS generation in neutrophils is critical in WKYMVm-induced microvascular hyperpermeability during neutrophil recruitment.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Permeabilidade Capilar/efeitos dos fármacos , Permeabilidade Capilar/fisiologia , Lipoproteínas/metabolismo , Neutrófilos/metabolismo , Oligopeptídeos/farmacologia , Proteínas Adaptadoras de Transdução de Sinal/deficiência , Proteínas Adaptadoras de Transdução de Sinal/genética , Animais , Transplante de Medula Óssea , Permeabilidade Capilar/imunologia , Adesão Celular/efeitos dos fármacos , Adesão Celular/imunologia , Adesão Celular/fisiologia , Lipoproteínas/deficiência , Lipoproteínas/genética , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Sistema de Sinalização das MAP Quinases/imunologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Infiltração de Neutrófilos/efeitos dos fármacos , Infiltração de Neutrófilos/imunologia , Infiltração de Neutrófilos/fisiologia , Neutrófilos/efeitos dos fármacos , Neutrófilos/imunologia , Espécies Reativas de Oxigênio/metabolismo , Receptores de Formil Peptídeo/agonistas , Quimeras de Transplante/imunologia , Quimeras de Transplante/fisiologia , Vênulas/efeitos dos fármacos , Vênulas/imunologia , Vênulas/fisiologia
19.
Biochem Biophys Res Commun ; 527(1): 207-212, 2020 06 18.
Artigo em Inglês | MEDLINE | ID: mdl-32446368

RESUMO

SH2 domain-containing inositol 5'-phosphatase (SHIP) has critical functions in regulating signal transduction. In additional to its lipid phosphatase activity, SHIP engages in multiple protein-protein interactions, which can serve to localize either SHIP or its binding partners to a particular subcellular domain. Knock-out and knock-down studies have elucidated that SHIP negatively regulates the accumulation of F-actin in leukocytes, usually resulting in inhibition of actin dependent cellular activities such as spreading and migration. Here, we demonstrate that overexpression of SHIP inhibits B cell antigen receptor (BCR)-mediated cell spreading in murine and human B cell lines. B cell stimulation via the BCR or pervanadate induces an interaction between SHIP and Nck, an adaptor protein known to promote actin polymerization. Using a fluorescence recovery after photobleaching (FRAP) assay, we demonstrate that overexpression of SHIP slows F-actin dynamics in BCR-stimulated B cells and this can be overcome by co-overexpression of Nck. Our data supports a role for SHIP in limiting actin turnover and suggests it may do so in part by sequestering Nck.


Assuntos
Actinas/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Linfócitos B/metabolismo , Inositol Polifosfato 5-Fosfatases/metabolismo , Proteínas Oncogênicas/metabolismo , Animais , Humanos , Inositol Polifosfato 5-Fosfatases/genética , Camundongos , Receptores de Antígenos de Linfócitos B/metabolismo , Células Tumorais Cultivadas , Domínios de Homologia de src
20.
Cancers (Basel) ; 11(10)2019 Oct 09.
Artigo em Inglês | MEDLINE | ID: mdl-31601046

RESUMO

: The phosphatidyl-inositol 3 kinase (PI3K) δ inhibitor, idelalisib (IDE), is a potent inhibitor of the B-cell receptor pathway and a novel and highly effective agent for the treatment of chronic lymphocytic leukemia (CLL). We evaluated the activities of IDE in comparison to bendamusine (BEN), a commonly used alkylating agent, in primary CLL cells ex vivo. In contrast to BEN, IDE was cytotoxic to cells from extensively-treated patients, including those with a deletion (del)17p. Cross-resistance was not observed between BEN and IDE, confirming their different modes of cytotoxicity. Marked synergy was seen between BEN and IDE, even in cases that were resistant to BEN or IDE individually, and those with deletion (del) 17p. CD40L/interleukin 4 (IL4) co-treatment mimicking the CLL microenvironment increased resistance to IDE, but synergy was retained. PI3Kδ-deficient murine splenic B cells were more resistant to IDE and showed reduced synergy with BEN, thus confirming the importance of functional PI3Kδ protein. Although IDE was observed to induce γH2AX, IDE did not enhance activation of the DNA damage response nor DNA repair activity. Interestingly, IDE decreased global RNA synthesis and was antagonistic with 5,6-Dichlorobenzimidazole 1-b-D-ribofuranoside (DRB), an inhibitor of transcription. These findings add to the increasingly complex cellular effects of IDE, and B cell receptor (BCR) inhibitors in general, in CLL.

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