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1.
BMC Cancer ; 19(1): 700, 2019 Jul 16.
Artigo em Inglês | MEDLINE | ID: mdl-31311517

RESUMO

BACKGROUND: Multiple myeloma (MM), characterized by cancerous proliferation of plasmablasts (PB) and plasma cells (PC), remains incurable in many patients. Differentially expressed molecules between MM PCs and healthy PCs have been explored in order to identify novel targets for treating MM. In the present study, we searched for novel MM therapeutic targets by comparing mRNA expression patterns between the Mus musculus myeloma plasmablast-like SP 2/0 cell line and LPS-induced PB/PC. METHODS: Gene expression profiles of LPS-induced PB/PC and SP 2/0 cells were determined using RNA-sequencing. A predicted gene (Gm40600) was found to be expressed at a low level in SP 2/0 cells. To study the role of Gm40600 in malignant PC, Gm40600 cDNA was cloned into a lentiviral vector (LV201) containing a puromycin selectable marker that was then transfected into SP 2/0 cells. Stable Gm40600-expressing SP 2/0 cells were selected using puromycin. The effect of Gm40600 on SP 2/0 cell proliferation, cell cycle/apoptosis, and tumor progression was assessed by cell counting kit-8 (CCK8), flow cytometry (FACS), and the SP 2/0 isograft mouse model, respectively. The effect of Gm40600 on mRNA and protein expression was evaluated by RNA-sequencing and western blotting, respectively. RESULTS: We found that SP 2/0 cells expressed lower level of Gm40600 mRNA as compared to LPS-induced PB/PC. Overexpression of Gm40600 significantly suppressed SP 2/0 cell proliferation and isograft tumor progression in an isograft mouse model by promoting apoptosis. In addition, Gm40600 overexpression suppressed transcription of the gene encoding Bcl2. Gm40600 overexpression also reduced the expression of PC-associated transcription factors Blimp1 and Xbp1, which promote transcription of the gene that encodes Bcl2. CONCLUSIONS: Gm40600 reduced SP 2/0 cell proliferation and isograft tumor growth and progression by suppressing Blimp1 and Xbp1-mediated Bcl2 transcription to induce apoptosis. Thus, regulation of a human homolog of Gm40600, or associated factors, may be a potential therapeutic approach for treating MM.


Assuntos
Biomarcadores Tumorais/genética , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Fator 1 de Ligação ao Domínio I Regulador Positivo/genética , Proteína 1 de Ligação a X-Box/genética , Animais , Apoptose/genética , Linhagem Celular Tumoral , Progressão da Doença , Isoenxertos , Camundongos , Camundongos Transgênicos , Fator 1 de Ligação ao Domínio I Regulador Positivo/metabolismo , Regiões Promotoras Genéticas , Proteínas Proto-Oncogênicas c-bcl-2/genética , RNA Mensageiro/genética , Proteína 1 de Ligação a X-Box/metabolismo
2.
Oncol Rep ; 41(6): 3219-3232, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31002364

RESUMO

The inactivation of tumor suppressor gene positive regulatory domain containing I (PRDM1) and activation of signal transducer and activator of transcription 3 (STAT3) have been detected in the majority of extranodal NK/T­cell lymphoma, nasal type (EN­NK/T­NT) cases. In the present study, their association with and effects on the clinicopathologic features of EN­NK/T­NT are described. PRDM1 was revealed to be expressed in 19 out of 58 patients (32.8%) with EN­NK/T­NT, and phosphorylated STAT3 was overexpressed in 42 out of 58 (72.4%). Oncogenic pathways were investigated by NanoString encounter technology in 5 PRDM1(+) and 5 PRDM1(­) EN­NK/T­NT specimens. Multiple oncogenic pathways involved in cell apoptosis, cellcycle (CC) and angiogenesis were discriminately activated in EN­NK/T­NT cases, and in PRDM1(+) cases in particular. The sustained activation of the Janus kinase 3 (JAK)/STAT3 pathway was more pronounced. In addition, missense mutations in the SRC homology 2 domain of STAT3 were detected in 7 out of 37 EN­NK/T­NT cases (18.92%), and the acquired mutation was related to the activation of the JAK3/STAT3 pathway. The downregulation of PRDM1 and upregulation of phospho­STAT3 (Tyr705) were associated with angiocentric infiltration of EN­NK/T­NT (P=0.039). Notably, the prognosis of patients in the PRDM1(+)/STAT3 [mutated (mut­)] group was considerably improved than that of patients in the STAT3(mut+)/PRDM(­) group (P=0.037). In addition, the inhibition of NK/T cell lymphoma cell lines by Stattic and tofacitinib could suppress cell proliferation by inducing cell apoptosis or arresting the CC. The present results revealed that the JAK3/STAT3 oncogenic pathway and PRDM1 expression could stratify clinicopathologic features of EN­NK/T­NT. The inhibition of the JAK3/STAT3 pathway may serve as a treatment option for EN­NK/T­NT.


Assuntos
Janus Quinase 3/genética , Linfoma de Células T/tratamento farmacológico , Fator 1 de Ligação ao Domínio I Regulador Positivo/genética , Fator de Transcrição STAT3/genética , Apoptose/efeitos dos fármacos , Carcinogênese/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Feminino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Linfoma de Células T/genética , Linfoma de Células T/patologia , Masculino , Pessoa de Meia-Idade , Mutação/genética , Fosforilação/efeitos dos fármacos , Piperidinas/farmacologia , Pirimidinas/farmacologia , Pirróis/farmacologia
3.
Med Sci Monit ; 25: 3032-3040, 2019 Apr 25.
Artigo em Inglês | MEDLINE | ID: mdl-31019190

RESUMO

BACKGROUND T follicular helper (Tfh) cells are a subgroup of activated CD4+ T cells in the germinal centers of secondary lymphoid organs, they play critical roles in the development of many chronic autoimmune inflammatory diseases. The aim of this study was to investigate whether circulating Tfh cells contribute to the development of rheumatoid arthritis (RA). MATERIAL AND METHODS Thirty patients fulfilled the diagnosis criteria that was established by the American College of Rheumatology and 30 healthy controls were recruited. The frequency of Tfh cells in patients and collagen-induced arthritis (CIA) in DBA/1J mice were analyzed by flow cytometry. The serum IL-21 level was examined by enzyme-linked immunosorbent assay (ELISA). The mRNA expression of Blimp-1 and Bcl-6 were detected by qRT-PCR. RESULTS RA patients had more CD4⁺PD-1⁺CXCR5⁺ Tfh cells in peripheral blood compared with healthy controls, and CIA in DBA/1J mice showed similar results. Higher mRNA expression of Bcl-6 and lower Blimp-1 mRNA expression were observed in patients with RA compared to healthy controls, and the expression level of IL-21 was higher in RA patients, which was also seen in CIA mice. Furthermore, the spleen CD4⁺ICOS⁺CXCR5⁺ Tfh cells in CIA mice show significantly higher frequency than that in the control mice. The percentage of CD4⁺PD-1⁺CXCR5⁺ Tfh cells was correlated positively with the values of erythrocyte sedimentation rate (ESR) (r=0.968, P<0.001), rheumatoid factor (RF) (r=0.962, P<0.001), C-reactive protein (CRP) (r=0.953, P<0.001), and anti-cyclic citrullinated peptide antibodies (ACPA) (r=0.966, P<0.001), and the level of serum interleukin (IL)-21 in RA patients showed positive correlation with ESR (r=0.982, P<0.001), RF (r=0.959, P<0.001), CRP (r=0.951, P<0.001), and ACPA (r=0.971, P<0.001) as well. CONCLUSIONS The activated Tfh cells in the peripheral blood may be responsible for the development of RA.


Assuntos
Artrite Reumatoide/imunologia , Receptor de Morte Celular Programada 1/imunologia , Receptores CXCR5/imunologia , Linfócitos T Auxiliares-Indutores/imunologia , Adulto , Idoso , Animais , Artrite Reumatoide/sangue , Autoanticorpos/imunologia , Proteína C-Reativa/imunologia , Proteína C-Reativa/metabolismo , Linfócitos T CD4-Positivos/imunologia , Feminino , Citometria de Fluxo , Humanos , Proteína Coestimuladora de Linfócitos T Induzíveis/metabolismo , Interleucinas/sangue , Interleucinas/imunologia , Masculino , Camundongos , Camundongos Endogâmicos DBA , Pessoa de Meia-Idade , Fator 1 de Ligação ao Domínio I Regulador Positivo/genética , Fator 1 de Ligação ao Domínio I Regulador Positivo/metabolismo , Receptor de Morte Celular Programada 1/metabolismo , Proteínas Proto-Oncogênicas c-bcl-6/genética , Proteínas Proto-Oncogênicas c-bcl-6/metabolismo , Receptores CXCR5/sangue , Fator Reumatoide/metabolismo , Subpopulações de Linfócitos T/imunologia , Subpopulações de Linfócitos T/metabolismo , Linfócitos T Auxiliares-Indutores/metabolismo
4.
BMC Cancer ; 19(1): 81, 2019 Jan 17.
Artigo em Inglês | MEDLINE | ID: mdl-30654767

RESUMO

BACKGROUND: Age-related genetic changes in lymphocyte subsets are not currently well documented. BACH2 is a transcription factor that plays an important role in immune-mediated homeostasis by tightly regulating PRDM1 expression in both B-cells and T-cells. BACH2 gene expression is highly sensitive to DNA damage in aged mice. This concept led us to investigate the variation in BACH2 and also PRDM1 expression in major lymphocyte subsets with age. METHODS: Lymphocyte subsets from 60 healthy donors, aged from 20 to 90 years, and 41 untreated chronic lymphocytic leukemia patients were studied. BACH2 and PRDM1 gene expression was analyzed by real-time quantitative PCR. BACH2 gene expression was correlated with its protein expression. Lymphocyte apoptosis was evaluated after intracellular oxidative stress-inducing etoposide treatment of T and B cells. RESULTS: Our analysis shows BACH2 mRNA downregulation with age in healthy donor CD4+, CD8+ T-cells and CD19+ B-cells. Decreased BACH2 expression was also correlated with an age-related reduction in CD8 + CD28+ T-cells. We found a strong correlation between age-related BACH2 downregulation and decreased CD4+ T-cell and CD19+ B-cell apoptosis. PRDM1, as expected, was significantly upregulated in CD4+ T-cells, CD8+ T-cells and CD19+ B-cells, and inversely correlated with BACH2. A comparison of untreated chronic lymphocytic leukemia patients with age-matched healthy donors reveals that BACH2 mRNA expression was further reduced in CD4+ T-cells, CD8+ T-cells and leukemic-B cells. PRDM1 gene expression was consequently significantly upregulated in CD4+ and CD8+ T-cells in chronic lymphocytic leukemia patients but not in their leukemic B-cells. CONCLUSION: Overall, our data suggest that BACH2 and PRDM1 genes are significantly correlated with age in human immune cells and may be involved in immunosenescence.


Assuntos
Envelhecimento/imunologia , Fatores de Transcrição de Zíper de Leucina Básica/metabolismo , Leucemia Linfocítica Crônica de Células B/imunologia , Subpopulações de Linfócitos/metabolismo , Fator 1 de Ligação ao Domínio I Regulador Positivo/metabolismo , Adulto , Idoso , Idoso de 80 Anos ou mais , Fatores de Transcrição de Zíper de Leucina Básica/genética , Fatores de Transcrição de Zíper de Leucina Básica/imunologia , Senescência Celular/imunologia , Regulação para Baixo/imunologia , Feminino , Voluntários Saudáveis , Humanos , Leucemia Linfocítica Crônica de Células B/sangue , Leucemia Linfocítica Crônica de Células B/patologia , Subpopulações de Linfócitos/imunologia , Masculino , Pessoa de Meia-Idade , Fator 1 de Ligação ao Domínio I Regulador Positivo/genética , Fator 1 de Ligação ao Domínio I Regulador Positivo/imunologia , RNA Mensageiro/metabolismo , Regulação para Cima/imunologia , Adulto Jovem
5.
Nat Immunol ; 20(3): 288-300, 2019 03.
Artigo em Inglês | MEDLINE | ID: mdl-30692620

RESUMO

Although tissue-resident memory T cells (TRM cells) have been shown to regulate host protection in infectious disorders, their function in inflammatory bowel disease (IBD) remains to be investigated. Here we characterized TRM cells in human IBD and in experimental models of intestinal inflammation. Pro-inflammatory TRM cells accumulated in the mucosa of patients with IBD, and the presence of CD4+CD69+CD103+ TRM cells was predictive of the development of flares. In vivo, functional impairment of TRM cells in mice with double knockout of the TRM-cell-associated transcription factors Hobit and Blimp-1 attenuated disease in several models of colitis, due to impaired cross-talk between the adaptive and innate immune system. Finally, depletion of TRM cells led to a suppression of colitis activity. Together, our data demonstrate a central role for TRM cells in the pathogenesis of chronic intestinal inflammation and suggest that these cells could be targets for future therapeutic approaches in IBD.


Assuntos
Linfócitos T CD8-Positivos/imunologia , Colite/imunologia , Memória Imunológica/imunologia , Fator 1 de Ligação ao Domínio I Regulador Positivo/imunologia , Fatores de Transcrição/imunologia , Animais , Linfócitos T CD8-Positivos/metabolismo , Células Cultivadas , Doença Crônica , Colite/genética , Colite/metabolismo , Citocinas/genética , Citocinas/imunologia , Citocinas/metabolismo , Modelos Animais de Doenças , Perfilação da Expressão Gênica , Humanos , Memória Imunológica/genética , Doenças Inflamatórias Intestinais/genética , Doenças Inflamatórias Intestinais/imunologia , Doenças Inflamatórias Intestinais/metabolismo , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos Transgênicos , Fator 1 de Ligação ao Domínio I Regulador Positivo/deficiência , Fator 1 de Ligação ao Domínio I Regulador Positivo/genética , Fatores de Transcrição/deficiência , Fatores de Transcrição/genética
6.
EMBO J ; 38(2)2019 01 15.
Artigo em Inglês | MEDLINE | ID: mdl-30498131

RESUMO

The transcription factor Blimp1 is not only an essential regulator of plasma cells, but also a risk factor for the development of autoimmune disease in humans. Here, we demonstrate in the mouse that the Prdm1 (Blimp1) gene was partially activated at the chromatin and transcription level in early B cell development, although mature Prdm1 mRNA did not accumulate due to posttranscriptional regulation. By analyzing a mouse model that facilitated ectopic Blimp1 protein expression throughout B lymphopoiesis, we could demonstrate that Blimp1 impaired B cell development by interfering with the B cell gene expression program, while leading to an increased abundance of plasma cells by promoting premature plasmablast differentiation of immature and mature B cells. With progressing age, these mice developed an autoimmune disease characterized by the presence of autoantibodies and glomerulonephritis. Hence, these data identified ectopic Blimp1 expression as a novel mechanism, through which Blimp1 can act as a risk factor in the development of autoimmune disease.


Assuntos
Linfócitos B/metabolismo , Glomerulonefrite/metabolismo , Fator 1 de Ligação ao Domínio I Regulador Positivo/genética , Fator 1 de Ligação ao Domínio I Regulador Positivo/metabolismo , Animais , Autoanticorpos/metabolismo , Linfócitos B/citologia , Diferenciação Celular , Modelos Animais de Doenças , Feminino , Redes Reguladoras de Genes , Glomerulonefrite/genética , Humanos , Masculino , Camundongos , Ativação Transcricional
7.
J Immunol ; 202(1): 79-92, 2019 01 01.
Artigo em Inglês | MEDLINE | ID: mdl-30478092

RESUMO

The role of retinoid-related orphan receptor γ t (RORγt) in Th17 cell differentiation has been well established; however, how it regulates other T cell lineages is still not clearly understood. In this study, we report that in mice, while promoting Th17 cell differentiation, RORγt inhibited IL-10 production by T cells, thereby preserving the pathogenicity of Th17 cells. Treatment with RORγt-specific inhibitor suppressed Th17 cell signature cytokines, but promoted IL-10 production. RORγt inhibitor-treated Th17 cells induce less severe colitis compared with control Th17 cells. Mechanistically, the RORγt inhibitor induced T cell expression of Blimp-1 (encoded by Prdm1). Prdm1-/- T cells produced significantly fewer IL-10 when treated with RORγt inhibitor compared with wild-type T cells. Furthermore, RORγt inhibitor-treated Prdm1-/- Th17 cells induce more severe colitis compared with RORγt inhibitor-treated wild-type Th17 cells. Collectively, our studies reveal a novel mechanism by which RORγt drives and maintains pathogenic Th17 cell development by inhibiting IL-10 production.


Assuntos
Colite/imunologia , Interleucina-10/metabolismo , Intestinos/imunologia , Membro 3 do Grupo F da Subfamília 1 de Receptores Nucleares/metabolismo , Células Th17/imunologia , Animais , Diferenciação Celular , Linhagem da Célula , Células Cultivadas , Repressão Epigenética , Ativação Linfocitária , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Membro 3 do Grupo F da Subfamília 1 de Receptores Nucleares/genética , Fator 1 de Ligação ao Domínio I Regulador Positivo/genética
8.
Int J Mol Med ; 43(2): 1094-1104, 2019 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-30483767

RESUMO

Modulation of differentiation of dendritic cells (DCs), which are derived from bone marrow cells, may influence their maturation and consequently regulate their ability to present antigens to alloreactive T lymphocytes. B lymphocyte­induced maturation protein­1 (Blimp1) is a master regulator of immunocyte differentiation, which has been investigated for its effect on DCs. In the present study, a lentivirus was used as a vector to transduce Blimp1­short hairpin (sh)RNA into primary bone marrow cells during their differentiation to DCs. Lentiviral­mediated Blimp1­shRNA (lenti­shRNA­Blimp1) had a transduction efficiency of >60% in DC precursors. Lenti­shRNA­Blimp1 significantly downregulated the expression levels of Blimp1 and modulated the expression of its target proteins, including class II major histocompatibility complex (MHC) transactivator, c­myc and interleukin­6. Although lenti­shRNA­Blimp1 did not interfere with the differentiation of bone marrow cells to DCs, it inhibited DC maturation by decreasing the expression of surface MHC­II molecules, but not the expression of MHC­I molecules and co­stimulatory molecules [cluster of differentiation (CD)80/CD86]. Subsequently, alloreactive T cell proliferation was alleviated and regulatory T cells were expanded in response to lenti­shRNA­Blimp1. A toxicity assay indicated that the morphology and proliferation of cultured DCs were mildly influenced by the lentiviral vector, indicating that the use of alternative vectors with minimal or no toxicity could be investigated in future studies. In conclusion, transduction with lenti­shRNA­Blimp1 modulated the maturation of DCs via MHC­II molecule suppression and inhibited alloreactive T cell activation. The present findings supported the application of Blimp1­based intervention as a novel approach to induce immature DCs for further immunological research.


Assuntos
Diferenciação Celular , Células Dendríticas/citologia , Células Dendríticas/metabolismo , Regulação da Expressão Gênica no Desenvolvimento , Fator 1 de Ligação ao Domínio I Regulador Positivo/genética , Animais , Células da Medula Óssea/citologia , Células da Medula Óssea/metabolismo , Ativação Linfocitária/imunologia , Masculino , Camundongos , RNA Mensageiro/genética , RNA Interferente Pequeno/genética , Subpopulações de Linfócitos T/imunologia , Subpopulações de Linfócitos T/metabolismo , Transdução Genética
9.
Front Immunol ; 9: 2470, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30410493

RESUMO

Follicular helper T cells (Tfh) are specialized helper T cells that are predominantly located in germinal centers and provide help to B cells. The development and differentiation of Tfh cells has been shown to be regulated by transcription factors, such as B-cell lymphoma 6 protein (Bcl-6), signal transducer and activator of transcription 3 (STAT3) and B lymphocyte-induced maturation protein-1 (Blimp-1). In addition, cytokines, including IL-21, have been found to be important for Tfh cell development. Moreover, several epigenetic modifications have also been reported to be involved in the determination of Tfh cell fate. The regulatory network is complicated, and the number of novel molecules demonstrated to control the fate of Tfh cells is increasing. Therefore, this review aims to summarize the current knowledge regarding the molecular regulation of Tfh cell development and differentiation at the protein level and at the epigenetic level to elucidate Tfh cell biology and provide potential targets for clinical interventions in the future.


Assuntos
Diferenciação Celular/genética , Centro Germinativo/imunologia , Linfócitos T Auxiliares-Indutores/imunologia , Animais , Epigênese Genética , Regulação da Expressão Gênica , Humanos , Interleucinas/metabolismo , Fator 1 de Ligação ao Domínio I Regulador Positivo/genética , Fator 1 de Ligação ao Domínio I Regulador Positivo/metabolismo , Proteínas Proto-Oncogênicas c-bcl-6/genética , Proteínas Proto-Oncogênicas c-bcl-6/metabolismo , Fator de Transcrição STAT3/genética , Fator de Transcrição STAT3/metabolismo
10.
Medicine (Baltimore) ; 97(44): e13103, 2018 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-30383696

RESUMO

RATIONALE: The abnormal cell types in chronic myeloid leukemia (CML) and monoclonal gammopathy of uncertain (MGUS) are quite different, being myeloid and plasma cells, respectively. The coexistence of CML and MGUS is an uncommon event, which is seldom reported in literature. PATIENT CONCERNS: A 52-year-old female was diagnosed with CML in April 2001. From November 2006, the patient started on imatinib mesylate and kept a complete hematologic and cytogenetic response for nearly 11 years. During her follow-up on July 7, 2017, thrombocytopenia (35*109/L) was found. Bone marrow aspiration revealed 6% plasma cell infiltration. Serum immunoelectrophoresis revealed 1.24 g/dL of serum monoclonal (M) protein of IgG-κ type. DIAGNOSIS: MGUS was diagnosed because of absence of anemia, hypercalcemia, lytic bone lesions, or renal failure. Immune thrombocytopenia (ITP) was also diagnosed in this patient following the detection of antiplatelet autoantibodies. Complex karyotype and missense mutation in PRDM1 were identified. INTERVENTIONS: Because of her obvious decrease of platelets, she started treatment with thalidomide and prednisone. OUTCOMES: Three months later, bone marrow aspirate showed disappearance of plasma cells. There developed an abrupt decrease in IgG and the absence of M-spike in serum immunoelectrophoresis. The platelet count kept normal during 1 year follow-up. LESSONS: Karyotypic event and gene mutation found in this case may be the initiation of disease transformation. Administration of thalidomide and prednisone proved effective in this patient.


Assuntos
Leucemia Mielogênica Crônica BCR-ABL Positiva/complicações , Gamopatia Monoclonal de Significância Indeterminada/diagnóstico , Prednisolona/uso terapêutico , Talidomida/uso terapêutico , Feminino , Humanos , Mesilato de Imatinib/uso terapêutico , Leucemia Mielogênica Crônica BCR-ABL Positiva/tratamento farmacológico , Pessoa de Meia-Idade , Gamopatia Monoclonal de Significância Indeterminada/tratamento farmacológico , Gamopatia Monoclonal de Significância Indeterminada/genética , Mutação , Fator 1 de Ligação ao Domínio I Regulador Positivo/genética , Prevalência , Resultado do Tratamento
11.
J Immunol ; 201(11): 3343-3351, 2018 12 01.
Artigo em Inglês | MEDLINE | ID: mdl-30348736

RESUMO

Vaccination has been the most effective way to prevent or reduce infectious diseases; examples include the eradication of smallpox and attenuation of tetanus and measles. However, there is a large segment of the population that responds poorly to vaccines, in part because they are immunocompromised because of disease, age, or pharmacologic therapy and are unable to generate long-term protection. Specialized proresolving mediators are endogenously produced lipids that have potent proresolving and anti-inflammatory activities. Lipoxin B4 (LXB4) is a member of the lipoxin family, with its proresolving effects shown in allergic airway inflammation. However, its effects on the adaptive immune system, especially on human B cells, are not known. In this study, we investigated the effects of LXB4 on human B cells using cells from healthy donors and donors vaccinated against influenza virus in vitro. LXB4 promoted IgG Ab production in memory B cells and also increased the number of IgG-secreting B cells. LXB4 enhanced expression of two key transcription factors involved in plasma cell differentiation, BLIMP1 and XBP1. Interestingly, LXB4 increased expression of cyclooxygenase-2 (COX2), an enzyme that is required for efficient B cell Ab production. The effects of LXB4 are at least partially COX2-dependent as COX2 inhibitors attenuated LXB4-stimulated BLIMP1 and Xpb-1 expression as well as IgG production. Thus, our study reveals for the first time, to our knowledge, that LXB4 boosts memory B cell activation through COX2 and suggests that LXB4 can serve as a new vaccine adjuvant.


Assuntos
Adjuvantes Imunológicos/metabolismo , Anticorpos Antivirais/metabolismo , Linfócitos B/imunologia , Ciclo-Oxigenase 2/metabolismo , Vacinas contra Influenza/imunologia , Influenza Humana/imunologia , Lipoxinas/metabolismo , Imunidade Adaptativa , Formação de Anticorpos , Diferenciação Celular , Células Cultivadas , Ciclo-Oxigenase 2/genética , Humanos , Memória Imunológica , Fator 1 de Ligação ao Domínio I Regulador Positivo/genética , Fator 1 de Ligação ao Domínio I Regulador Positivo/metabolismo , Regulação para Cima , Vacinação , Proteína 1 de Ligação a X-Box/genética , Proteína 1 de Ligação a X-Box/metabolismo
12.
Cancer Discov ; 8(12): 1632-1653, 2018 12.
Artigo em Inglês | MEDLINE | ID: mdl-30274972

RESUMO

TET2 somatic mutations occur in ∼10% of diffuse large B-cell lymphomas (DLBCL) but are of unknown significance. Herein, we show that TET2 is required for the humoral immune response and is a DLBCL tumor suppressor. TET2 loss of function disrupts transit of B cells through germinal centers (GC), causing GC hyperplasia, impaired class switch recombination, blockade of plasma cell differentiation, and a preneoplastic phenotype. TET2 loss was linked to focal loss of enhancer hydroxymethylation and transcriptional repression of genes that mediate GC exit, such as PRDM1. Notably, these enhancers and genes are also repressed in CREBBP-mutant DLBCLs. Accordingly, TET2 mutation in patients yields a CREBBP-mutant gene-expression signature, CREBBP and TET2 mutations are generally mutually exclusive, and hydroxymethylation loss caused by TET2 deficiency impairs enhancer H3K27 acetylation. Hence, TET2 plays a critical role in the GC reaction, and its loss of function results in lymphomagenesis through failure to activate genes linked to GC exit signals. SIGNIFICANCE: We show that TET2 is required for exit of the GC, B-cell differentiation, and is a tumor suppressor for mature B cells. Loss of TET2 phenocopies CREBBP somatic mutation. These results advocate for sequencing TET2 in patients with lymphoma and for the testing of epigenetic therapies to treat these tumors.See related commentary by Shingleton and Dave, p. 1515.This article is highlighted in the In This Issue feature, p. 1494.


Assuntos
Diferenciação Celular/genética , Proteínas de Ligação a DNA/genética , Centro Germinativo/metabolismo , Linfoma Difuso de Grandes Células B/genética , Plasmócitos/metabolismo , Proteínas Proto-Oncogênicas/genética , Animais , Proteína de Ligação a CREB/genética , Proteína de Ligação a CREB/metabolismo , Proteínas de Ligação a DNA/metabolismo , Epigênese Genética/genética , Perfilação da Expressão Gênica/métodos , Centro Germinativo/patologia , Células-Tronco Hematopoéticas/metabolismo , Humanos , Hiperplasia , Linfoma Difuso de Grandes Células B/metabolismo , Linfoma Difuso de Grandes Células B/patologia , Camundongos Knockout , Camundongos Transgênicos , Mutação , Plasmócitos/patologia , Fator 1 de Ligação ao Domínio I Regulador Positivo/genética , Fator 1 de Ligação ao Domínio I Regulador Positivo/metabolismo , Proteínas Proto-Oncogênicas/metabolismo
13.
Nat Commun ; 9(1): 4292, 2018 10 16.
Artigo em Inglês | MEDLINE | ID: mdl-30327475

RESUMO

Early mammalian development entails transit through naive pluripotency towards post-implantation epiblast, which subsequently gives rise to primordial germ cells (PGC), the founding germline population. To investigate these cell fate transitions, we developed a compound-reporter to track cellular identity in a model of PGC specification (PGC-like cells; PGCLC), and coupled it with genome-wide CRISPR screening. We identify key genes both for exit from pluripotency and for acquisition of PGC fate, and characterise a central role for the transcription regulators Nr5a2 and Zfp296 in germline ontogeny. Abrogation of these genes results in widespread activation (Nr5a2-/-) or inhibition (Zfp296-/-) of WNT pathway factors in PGCLC. This leads to aberrant upregulation of the somatic programme or failure to activate germline genes, respectively, and consequently loss of germ cell identity. Our study places Zfp296 and Nr5a2 as key components of an expanded PGC gene regulatory network, and outlines a transferable strategy for identifying critical regulators of complex cell fate decisions.


Assuntos
Repetições Palindrômicas Curtas Agrupadas e Regularmente Espaçadas , Regulação da Expressão Gênica no Desenvolvimento , Células-Tronco Pluripotentes/citologia , Células-Tronco Pluripotentes/fisiologia , Animais , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Proteínas de Ligação a DNA/genética , Desenvolvimento Embrionário/genética , Genes Reporter , Proteínas de Fluorescência Verde/genética , Camundongos , Camundongos Transgênicos , Fator 1 de Ligação ao Domínio I Regulador Positivo/genética , Receptores Citoplasmáticos e Nucleares/genética , Proteínas Repressoras/genética , Proteínas Wnt/genética
14.
Int J Mol Sci ; 19(10)2018 Oct 19.
Artigo em Inglês | MEDLINE | ID: mdl-30347759

RESUMO

The PR/SET domain gene family (PRDM) encodes 19 different transcription factors that share a subtype of the SET domain [Su(var)3-9, enhancer-of-zeste and trithorax] known as the PRDF1-RIZ (PR) homology domain. This domain, with its potential methyltransferase activity, is followed by a variable number of zinc-finger motifs, which likely mediate protein⁻protein, protein⁻RNA, or protein⁻DNA interactions. Intriguingly, almost all PRDM family members express different isoforms, which likely play opposite roles in oncogenesis. Remarkably, several studies have described alterations in most of the family members in malignancies. Here, to obtain a pan-cancer overview of the genomic and transcriptomic alterations of PRDM genes, we reanalyzed the Exome- and RNA-Seq public datasets available at The Cancer Genome Atlas portal. Overall, PRDM2, PRDM3/MECOM, PRDM9, PRDM16 and ZFPM2/FOG2 were the most mutated genes with pan-cancer frequencies of protein-affecting mutations higher than 1%. Moreover, we observed heterogeneity in the mutation frequencies of these genes across tumors, with cancer types also reaching a value of about 20% of mutated samples for a specific PRDM gene. Of note, ZFPM1/FOG1 mutations occurred in 50% of adrenocortical carcinoma patients and were localized in a hotspot region. These findings, together with OncodriveCLUST results, suggest it could be putatively considered a cancer driver gene in this malignancy. Finally, transcriptome analysis from RNA-Seq data of paired samples revealed that transcription of PRDMs was significantly altered in several tumors. Specifically, PRDM12 and PRDM13 were largely overexpressed in many cancers whereas PRDM16 and ZFPM2/FOG2 were often downregulated. Some of these findings were also confirmed by real-time-PCR on primary tumors.


Assuntos
Regulação Neoplásica da Expressão Gênica , Neoplasias/genética , Domínios PR-SET , Fator 1 de Ligação ao Domínio I Regulador Positivo/genética , Transcriptoma , Bases de Dados Genéticas , Humanos , Taxa de Mutação , Fator 1 de Ligação ao Domínio I Regulador Positivo/química , Fator 1 de Ligação ao Domínio I Regulador Positivo/metabolismo
15.
J Dermatol Sci ; 92(2): 151-161, 2018 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-30197274

RESUMO

BACKGROUND: B lymphocyte-induced maturation protein-1 (BLIMP1) is a transcriptional repressor, and plays a crucial role in the regulation of development and functions of various immune cells. Currently, there is limited understanding about the regulation of BLIMP1 expression in keratinocytes and crosstalk between EGFR and BLIMP1 in skin homeostasis. OBJECTIVE: The aim of the study was to investigate the regulation and functional link between EGFR and BLIMP1 in human epidermal keratinocytes. METHODS: Immunoblotting and Q-PCR were used to determine the molecular mechanism of BLIMP1 expression induced by EGFR in primary human epidermal keratinocytes (NHEK) and HaCaT cells. In functional assay, effects of BLIMP1 knockdown on EGFR-mediated cytokine production, differentiation, and migration in NHEK were evaluated by Q-PCR, ELISA, immunoblotting, and/or wound-healing assay. RESULTS: EGFR activation by EGFR ligands could upregulate the protein and mRNA levels of BLIMP1 in NHEK and HaCaT cells. This effect was dependent on PKC, p38, and ERK activation. Additionally, the stability of BLIMP1 protein was under the control of the proteasome and lysosome degradation systems. EGF could also upregulate BLIMP1 expression in skin squamous cell carcinomas. In addition, BLIMP1 knockdown enhanced the EGFR-mediated IL8, CXCL5 and IL6 gene expression and keratinocyte migration, but reduced the EGFR-mediated suppression of differentiation marker K10. CONCLUSIONS: Our findings shed new insights into the regulation of BLIMP1 expression by EGFR-mediated gene transcription and proteasome/lysosome-mediated degradation in keratinocytes. Functionally, BLIMP1 is a negative regulator of EGF-induced inflammation and migration in keratinocytes, and exerts a gene-specific regulation on keratinocyte differentiation.


Assuntos
Queratinócitos/fisiologia , Lisossomos/metabolismo , Fator 1 de Ligação ao Domínio I Regulador Positivo/genética , Complexo de Endopeptidases do Proteassoma/metabolismo , Diferenciação Celular/fisiologia , Linhagem Celular Tumoral , Movimento Celular/fisiologia , Células Cultivadas , Citocinas/imunologia , Família de Proteínas EGF/metabolismo , Receptores ErbB/metabolismo , Técnicas de Silenciamento de Genes , Humanos , Fator 1 de Ligação ao Domínio I Regulador Positivo/metabolismo , Proteólise , RNA Mensageiro/metabolismo , Regulação para Cima
16.
Cell Rep ; 24(10): 2682-2693.e6, 2018 09 04.
Artigo em Inglês | MEDLINE | ID: mdl-30184502

RESUMO

Primordial germ cells (PGCs) are fate determined from pluripotent epiblasts. Signaling pathways and transcriptional regulators involved in PGC formation have been identified, but detailed molecular mechanisms of PGC fate determination remains poorly understood. Using RNAi screening, we identified histone deacetylase 3 (HDAC3) as a regulator of PGC formation. Hdac3 deficiency resulted in decreased nascent PGCs in vitro and in vivo, and somatic developmental genes were de-repressed by Hdac3 knockdown during PGC induction. We also demonstrated BLIMP1-dependent enrichment of HDAC3 and deacetylation of H3 and H4 histones in the somatic developmental genes in epiblast-like cells. In addition, the HDAC3/BLIMP1-targeted somatic gene products were enriched in PGC determinant genes; overexpression of these gene products in PGC-like cells in culture resulted in repression of PGC determinant genes. We propose that selective recruitment of HDAC3 to somatic genes by BLIMP1 and subsequent repression of these somatic genes are crucial for PGC fate determination.


Assuntos
Células Germinativas/metabolismo , Histona Desacetilases/metabolismo , Fator 1 de Ligação ao Domínio I Regulador Positivo/metabolismo , Acetilação , Animais , Regulação da Expressão Gênica no Desenvolvimento , Histona Desacetilases/genética , Histonas/metabolismo , Camundongos , Fator 1 de Ligação ao Domínio I Regulador Positivo/genética , Transdução de Sinais/genética , Transdução de Sinais/fisiologia , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo
17.
Nat Commun ; 9(1): 3555, 2018 09 03.
Artigo em Inglês | MEDLINE | ID: mdl-30177845

RESUMO

T-cells are crucial in maintanence of intestinal homeostasis, however, it is still unclear how microbiota metabolites regulate T-effector cells. Here we show gut microbiota-derived short-chain fatty acids (SCFAs) promote microbiota antigen-specific Th1 cell IL-10 production, mediated by G-protein coupled receptors 43 (GPR43). Microbiota antigen-specific Gpr43-/- CBir1 transgenic (Tg) Th1 cells, specific for microbiota antigen CBir1 flagellin, induce more severe colitis compared with wide type (WT) CBir1 Tg Th1 cells in Rag-/- recipient mice. Treatment with SCFAs limits colitis induction by promoting IL-10 production, and administration of anti-IL-10R antibody promotes colitis development. Mechanistically, SCFAs activate Th1 cell STAT3 and mTOR, and consequently upregulate transcription factor B lymphocyte-induced maturation protein 1 (Blimp-1), which mediates SCFA-induction of IL-10. SCFA-treated Blimp1-/- Th1 cells produce less IL-10 and induce more severe colitis compared to SCFA-treated WT Th1 cells. Our studies, thus, provide insight into how microbiota metabolites regulate Th1 cell functions to maintain intestinal homeostasis.


Assuntos
Colite/imunologia , Ácidos Graxos Voláteis/imunologia , Microbioma Gastrointestinal/imunologia , Interleucina-10/imunologia , Receptores Acoplados a Proteínas-G/imunologia , Células Th1/imunologia , Animais , Ácidos Graxos Voláteis/metabolismo , Flagelina/imunologia , Expressão Gênica , Técnicas de Inativação de Genes , Homeostase , Camundongos , Fator 1 de Ligação ao Domínio I Regulador Positivo/genética , Receptores Acoplados a Proteínas-G/genética , Fator de Transcrição STAT3/imunologia , Serina-Treonina Quinases TOR/imunologia , Regulação para Cima
18.
Front Immunol ; 9: 2053, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30250473

RESUMO

In response to external stimuli, naïve B cells proliferate and take on a range of fates important for immunity. How their fate is determined is a topic of much recent research, with candidates including asymmetric cell division, lineage priming, stochastic assignment, and microenvironment instruction. Here we manipulate the generation of plasmablasts from B lymphocytes in vitro by varying CD40 stimulation strength to determine its influence on potential sources of fate control. Using long-term live cell imaging, we directly measure times to differentiate, divide, and die of hundreds of pairs of sibling cells. These data reveal that while the allocation of fates is significantly altered by signal strength, the proportion of siblings identified with asymmetric fates is unchanged. In contrast, we find that plasmablast generation is enhanced by slowing times to divide, which is consistent with a hypothesis of competing timed stochastic fate outcomes. We conclude that this mechanistically simple source of alternative fate regulation is important, and that useful quantitative models of signal integration can be developed based on its principles.


Assuntos
Linfócitos B/fisiologia , Plasmócitos/fisiologia , Células Precursoras de Linfócitos B/fisiologia , Animais , Relógios Biológicos , Antígenos CD40/metabolismo , Diferenciação Celular , Divisão Celular , Linhagem da Célula , Células Cultivadas , Feminino , Imunização , Ativação Linfocitária , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Fator 1 de Ligação ao Domínio I Regulador Positivo/genética , Processos Estocásticos
19.
Proc Natl Acad Sci U S A ; 115(41): E9630-E9639, 2018 10 09.
Artigo em Inglês | MEDLINE | ID: mdl-30257949

RESUMO

Plasma cell differentiation involves coordinated changes in gene expression and functional properties of B cells. Here, we study the role of Mzb1, a Grp94 cochaperone that is expressed in marginal zone (MZ) B cells and during the terminal differentiation of B cells to antibody-secreting cells. By analyzing Mzb1 -/- Prdm1 +/gfp mice, we find that Mzb1 is specifically required for the differentiation and function of antibody-secreting cells in a T cell-independent immune response. We find that Mzb1-deficiency mimics, in part, the phenotype of Blimp1 deficiency, including the impaired secretion of IgM and the deregulation of Blimp1 target genes. In addition, we find that Mzb1 -/- plasmablasts show a reduced activation of ß1-integrin, which contributes to the impaired plasmablast differentiation and migration of antibody-secreting cells to the bone marrow. Thus, Mzb1 function is required for multiple aspects of plasma cell differentiation.


Assuntos
Células da Medula Óssea/metabolismo , Diferenciação Celular/fisiologia , Integrina beta1/metabolismo , Chaperonas Moleculares/metabolismo , Plasmócitos/metabolismo , Fator 1 de Ligação ao Domínio I Regulador Positivo/metabolismo , Animais , Células da Medula Óssea/citologia , Imunoglobulina M/genética , Imunoglobulina M/metabolismo , Integrina beta1/genética , Camundongos , Camundongos Knockout , Chaperonas Moleculares/genética , Plasmócitos/citologia , Fator 1 de Ligação ao Domínio I Regulador Positivo/genética
20.
Front Immunol ; 9: 1828, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30131810

RESUMO

Plasmacytoid dendritic cells (pDCs) are a specialized subset of DCs capable of rapidly producing copious amounts of type I IFN (IFN-I) in response to viral infections. The mechanism regulating rapid production of IFN-I after pDCs are exposed to viral nucleic acids remains elusive. Here, we show that the transcription factor Blimp-1 is promptly induced in pDCs after exposure to TLR7 and TLR9 ligands via a unique Ras-related C3 botulinum toxin substrate (Rac)-mediated pathway. Deletion of the Prdm1 gene encoding Blimp-1 impaired production of IFN-I, but not other cytokines, upon viral infection or treatment with CpG DNA in pDCs. Accordingly, mice lacking Blimp-1 in DCs failed to produce IFN-I after CpG stimulation and did not mount proper antiviral responses following flavivirus infection. The development of pDCs in bone marrow as well as the induction of several activation markers, such as CD86, CD69, and MHCII, by CpG stimulation was generally not affected by the absence of Blimp-1. Mechanistically, we found that Blimp-1 controls the activation of IKKα and IRF7 by directly suppressing interleukin-1 receptor-associated kinase 3 (Irak3), a negative regulator of TLR signaling, in pDCs. Together, we identify a Blimp-1-dependent pathway that rapidly facilitates IFN-I production by relieving interleukin-1 receptor-associated kinase M, encoded by Irak3, in pDCs.


Assuntos
Células Dendríticas/imunologia , Células Dendríticas/metabolismo , Interferon Tipo I/biossíntese , Quinases Associadas a Receptores de Interleucina-1/metabolismo , Fator 1 de Ligação ao Domínio I Regulador Positivo/metabolismo , Transdução de Sinais , Animais , Regulação da Expressão Gênica , Interações Hospedeiro-Patógeno , Humanos , Imunofenotipagem , Fator Regulador 7 de Interferon/metabolismo , Quinases Associadas a Receptores de Interleucina-1/genética , Glicoproteínas de Membrana/metabolismo , Camundongos , Camundongos Knockout , Modelos Biológicos , Fator 1 de Ligação ao Domínio I Regulador Positivo/genética , Ligação Proteica , Receptor 7 Toll-Like/metabolismo , Receptor Toll-Like 9/metabolismo
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