RESUMO
Heparin-induced thrombocytopenia (HIT) is a serious adverse drug reaction characterized by antibodies that recognize platelet factor 4/heparin complexes (PF4/H) and activate platelets to create a prothrombotic state. Although a high percentage of heparin-treated patients produce antibodies to PF4/H, only a subset also makes antibodies that are platelet activating (PA). A close correlation between PA antibodies and the likelihood of experiencing HIT has been demonstrated in clinical studies, but how PA (presumptively pathogenic) and nonactivating (NA) (presumptively benign) antibodies differ from each other at the molecular level is unknown. To address this issue, we cloned 7 PA and 47 NA PF4/H-binding antibodies from 6 patients with HIT and characterized their structural and functional properties. Findings showed that PA clones differed significantly from NA clones in possessing 1 of 2 heavy chain complementarity-determining region 3 (HCDR3) motifs, RX1-2R/KX1-2R/H (RKH) and YYYYY (Y5), in an unusually long complementarity-determining region 3 (≥20 residues). Mutagenic studies showed that modification of either motif in PA clones reduced or abolished their PA activity and that appropriate amino acid substitutions in HCDR3 of NA clones can cause them to become PA. Repertoire sequencing showed that the frequency of peripheral blood IgG+ B cells possessing RKH or Y5 was significantly higher in patients with HIT than in patients without HIT given heparin, indicating expansion of B cells possessing RKH or Y5 in HIT. These findings imply that antibodies possessing RKH or Y5 are relevant to HIT pathogenesis and suggest new approaches to diagnosis and treatment of this condition.
Assuntos
Regiões Determinantes de Complementaridade , Trombocitopenia , Humanos , Regiões Determinantes de Complementaridade/genética , Trombocitopenia/induzido quimicamente , Trombocitopenia/genética , Heparina , Anticorpos/efeitos adversos , Plaquetas/metabolismo , Fator Plaquetário 4RESUMO
The caspase recruitment domain family member (CARD)11-Bcl10-Malt1 signalosome controls TGF-ß-activated kinase 1 (TAK1) activation and regulates BCR-induced NF-κB activation. In this study, we discovered that CARD19 interacted with TAK1 and inhibited TAB2-mediated TAK1 ubiquitination and activation. Although CARD19 deficiency in mice did not affect B cell development, it enhanced clonal deletion, receptor editing, and anergy of self-reactive B cells, and it reduced autoantibody production. Mechanistically, CARD19 deficiency increased BCR/TAK1-mediated NF-κB activation, leading to increased expression of transcription factors Egr2/3, as well as the E3 ubiquitin ligases c-Cbl/Cbl-b, which are known inducers of B cell tolerance in self-reactive B cells. RNA sequencing analysis revealed that although CARD19 deficiency did not affect the overall Ag-induced gene expression in naive B cells, it suppressed BCR signaling and increased hyporesponsiveness of self-reactive B cells. As a result, CARD19 deficiency prevented Bm12-induced experimental systemic lupus erythematosus. In summary, CARD19 negatively regulates BCR/TAK1-induced NF-κB activation and its deficiency increases Egr2/3 and c-Cbl/Cbl-b expression in self-reactive B cells, thereby enhancing B cell tolerance.
Assuntos
NF-kappa B , Transdução de Sinais , Animais , Camundongos , NF-kappa B/metabolismo , Transdução de Sinais/fisiologia , MAP Quinase Quinase Quinases/metabolismo , UbiquitinaçãoRESUMO
The paraspeckle protein NONO is a multifunctional nuclear protein participating in the regulation of transcriptional regulation, mRNA splicing and DNA repair. However, whether NONO plays a role in lymphopoiesis is not known. In this study, we generated mice with global deletion of NONO and bone marrow (BM) chimeric mice in which NONO is deleted in all of mature B cells. We found that the global deletion of NONO in mice did not affect T-cell development but impaired early B-cell development in BM at pro- to pre-B-cell transition stage and B-cell maturation in the spleen. Studies of BM chimeric mice demonstrated that the impaired B-cell development in NONO-deficient mice is B-cell-intrinsic. NONO-deficient B cells displayed normal BCR-induced cell proliferation but increased BCR-induced cell apoptosis. Moreover, we found that NONO deficiency impaired BCR-induced activation of ERK, AKT, and NF-κB pathways in B cells, and altered BCR-induced gene expression profile. Thus, NONO plays a critical role in B-cell development and BCR-induced B-cell activation.
Assuntos
NF-kappa B , Transdução de Sinais , Camundongos , Animais , Camundongos Knockout , NF-kappa B/metabolismo , Linfócitos B/metabolismo , Receptores de Antígenos de Linfócitos B , Proteínas de Ligação a DNA/metabolismo , Proteínas de Ligação a RNA/metabolismoRESUMO
Immune thrombocytopenia (ITP) is a platelet disorder. Pediatric and adult ITP have been associated with sialic acid alterations, but the pathophysiology of ITP remains elusive, and ITP is often a diagnosis of exclusion. Our analysis of pediatric ITP plasma samples showed increased anti-Thomsen-Friedenreich antigen (TF antigen) antibody representation, suggesting increased exposure of the typically sialylated and cryptic TF antigen in these patients. The O-glycan sialyltransferase St3gal1 adds sialic acid specifically on the TF antigen. To understand if TF antigen exposure associates with thrombocytopenia, we generated a mouse model with targeted deletion of St3gal1 in megakaryocytes (MK) (St3gal1MK-/-). TF antigen exposure was restricted to MKs and resulted in thrombocytopenia. Deletion of Jak3 in St3gal1MK-/- mice normalized platelet counts implicating involvement of immune cells. Interferon-producing Siglec H-positive bone marrow (BM) immune cells engaged with O-glycan sialic acid moieties to regulate type I interferon secretion and platelet release (thrombopoiesis), as evidenced by partially normalized platelet count following inhibition of interferon and Siglec H receptors. Single-cell RNA-sequencing determined that TF antigen exposure by MKs primed St3gal1MK-/- BM immune cells to release type I interferon. Single-cell RNA-sequencing further revealed a new population of immune cells with a plasmacytoid dendritic cell-like signature and concomitant upregulation of the immunoglobulin rearrangement gene transcripts Igkc and Ighm, suggesting additional immune regulatory mechanisms. Thus, aberrant TF antigen moieties, often found in pathological conditions, regulate immune cells and thrombopoiesis in the BM, leading to reduced platelet count.
Assuntos
Megacariócitos/patologia , Contagem de Plaquetas , Polissacarídeos/análise , Púrpura Trombocitopênica Idiopática/patologia , Adolescente , Animais , Antígenos Glicosídicos Associados a Tumores/análise , Criança , Pré-Escolar , Humanos , Lactente , Camundongos Endogâmicos C57BL , Sialiltransferases/análise , beta-Galactosídeo alfa-2,3-SialiltransferaseRESUMO
Acute graft-versus-host disease (aGVHD) is one major serious complication that is induced by alloreactive donor T cells recognizing host Ags and limits the success of allogeneic hematopoietic stem cell transplantation. In the current studies, we identified a critical role of Kras in regulating alloreactive T cell function during aGVHD. Kras deletion in donor T cells dramatically reduced aGVHD mortality and severity in an MHC-mismatched allogeneic hematopoietic stem cell transplantation mouse model but largely maintained the antitumor capacity. Kras-deficient CD4 and CD8 T cells exhibited impaired TCR-induced activation of the ERK pathway. Kras deficiency altered TCR-induced gene expression profiles, including the reduced expression of various inflammatory cytokines and chemokines. Moreover, Kras deficiency inhibited IL-6-mediated Th17 cell differentiation and impaired IL-6-induced ERK activation and gene expression in CD4 T cells. These findings support Kras as a novel and effective therapeutic target for aGVHD.
Assuntos
Linfócitos T CD8-Positivos/imunologia , Doença Enxerto-Hospedeiro/imunologia , Efeito Enxerto vs Leucemia/imunologia , Transplante de Células-Tronco Hematopoéticas , Proteínas Proto-Oncogênicas p21(ras)/deficiência , Células Th17/imunologia , Aloenxertos , Animais , Linhagem Celular Tumoral , Doença Enxerto-Hospedeiro/genética , Efeito Enxerto vs Leucemia/genética , Interleucina-6/genética , Interleucina-6/imunologia , Sistema de Sinalização das MAP Quinases/genética , Sistema de Sinalização das MAP Quinases/imunologia , Camundongos , Camundongos Transgênicos , Proteínas Proto-Oncogênicas p21(ras)/imunologiaRESUMO
Heparin-induced thrombocytopenia is a relatively common drug-induced immune disorder that can have life-threatening consequences for affected patients. Immune complexes consisting of heparin, platelet factor 4 (PF4), and PF4/heparin-reactive Abs are central to the pathogenesis of heparin-induced thrombocytopenia. Regulatory T (Treg) cells are a subpopulation of CD4 T cells that play a key role in regulating immune responses, but their role in controlling PF4/heparin-specific Ab production is unknown. In the studies described in this article, we found that Foxp3-deficient mice lacking functional Treg cells spontaneously produced PF4/heparin-specific Abs. Following transplantation with bone marrow cells from Foxp3-deficient but not wild-type mice, Rag1-deficient recipients also produced PF4/heparin-specific Abs spontaneously. Adoptively transferred Treg cells prevented spontaneous production of PF4/heparin-specific Abs in Foxp3-deficient mice and inhibited PF4/heparin complex-induced production of PF4/heparin-specific IgGs in wild-type mice. Treg cells suppress immune responses mainly through releasing anti-inflammatory cytokines, such as IL-10. IL-10-deficient mice spontaneously produced PF4/heparin-specific Abs. Moreover, bone marrow chimeric mice with CD4 T cell-specific deletion of IL-10 increased PF4/heparin-specific IgG production upon PF4/heparin complex challenge. Short-term IL-10 administration suppresses PF4/heparin-specific IgG production in wild-type mice. Taken together, these findings demonstrate that Treg cells play an important role in suppressing PF4/heparin-specific Ab production.
Assuntos
Formação de Anticorpos , Heparina/imunologia , Imunoglobulina G/imunologia , Fator Plaquetário 4/imunologia , Linfócitos T Reguladores/imunologia , Animais , Fatores de Transcrição Forkhead/deficiência , Fatores de Transcrição Forkhead/imunologia , Heparina/genética , Imunoglobulina G/genética , Interleucina-10/deficiência , Interleucina-10/imunologia , Camundongos , Camundongos Knockout , Fator Plaquetário 4/genética , Linfócitos T Reguladores/citologiaRESUMO
Developing solid polymer electrolytes (SPEs) is a promising approach to realize practical dendrite-free lithium metal batteries (LMBs). Tuning the nanoscale polymer network chemsitry is of critical importance for SPE design. In this work, we took lessons from the rubber chemistry and developed a series of comb-chain crosslinker-based SPEs (ConSPEs) using a preformed polymer as the multifunctional crosslinker. The high-functionality crosslinker increased the connectivity of nanosized cross-linked domains, which led to a robust network with dramatically improved toughness and superior lithium dendrite resistance even at a current density of 2 mA cm-2. The uniform and flexile network also dramatically improved the anodic stability to over 5.3 V versus Li/Li+. Additive-free, all-solid-state LMBs with the ConSPE showed high discharge capacity and stable cycling up to 10 C rate, and could be stably cycled at 25 °C. Our results demonstrated that ConSPEs are promising for high-performance and dendrite-free LMBs.
RESUMO
AIM: The fragile X mental retardation protein (FMRP), the product of the FMR1 gene, is responsible for the fragile X syndrome (FXS). FMRP regulates miRNA expression and is involved in miRNA-mediated gene silencing. However, the question of whether FMRP is, in turn, regulated by miRNAs remains unanswered. MAIN METHODS: We detected the FMRP expression pattern by in situ hybridization. MiR-315 overexpression and knockout models were generated by germ-line transformation and ends-out homologous recombination, respectively. Western blotting and immunohistochemistry were used to detect Drosophila FMRP (dFMRP) and a Luciferase reporter assay was used to confirm the regulation of dfmr1 mRNA by mir-315. Synaptic structural quantification and electrophysiological methods were used to compare synaptic functions among groups. KEY FINDINGS: Here, we determined that the transcription product of dFMR1, the Drosophila homologue of FMR1, is a direct target of miR-315. MiR-315 is mainly expressed in the nervous system of Drosophila. Flies overexpressing miR-315 showed pupation defects and reduced hatching rates. A homozygous miR-315 knockout status is embryonic lethal in flies. These observations indicate that miR-315 is a key regulator of the Drosophila nervous system. Furthermore, computational prediction and cell-based luciferase and in vivo assays demonstrated that dfmr1 is directly targeted by miR-315. Lastly, using the neuromuscular junction as a model, we found that miR-315 regulates synaptic structure and transmission by targeting dfmr1. SIGNIFICANCE: These findings provide compelling evidence that miR-315 targets dfmr1 in the Drosophila nervous system, acting as a regulatory factor for the fine-tuned modulation of FMRP expression.
Assuntos
Proteínas de Drosophila/genética , Drosophila melanogaster/embriologia , Drosophila melanogaster/genética , Proteína do X Frágil da Deficiência Intelectual/genética , Regulação da Expressão Gênica no Desenvolvimento , Animais , Sistema Nervoso/embriologia , Sistema Nervoso/metabolismo , Neurogênese , Junção Neuromuscular/genética , Sinapses/genéticaRESUMO
Pre-T cell receptor (TCR) signaling is required for pre-T cell survival, proliferation, and differentiation from the CD4 and CD8 double negative (DN) to the double positive (DP) stage. However, the pre-TCR signal transduction pathway is not fully understood and the signaling molecules involved have not been completely identified. Phospholipase Cγ (PLCγ) 1 is an important signaling molecule that generates two second messengers, diacylglycerol and inositol 1,4,5-trisphosphate, that are important to mediate PKC activation and intracellular Ca2+ flux in many signaling pathways. Previously, we have shown that PLCγ1 is important for TCR-mediated signaling, development and T-cell activation, but the role of PLCγ1 in pre-TCR signal transduction and pre-T cell development is not known. In this study, we demonstrated that PLCγ1 expression level in pre-T cells was comparable to that in mature T cells. Deletion of PLCγ1 prior to the pre-TCR signaling stage partially blocked the DN3 to DN4 transition and reduced thymic cellularity. We also demonstrated that deletion of PLCγ1 impaired pre-T cell proliferation without affecting cell survival. Further study showed that deficiency of PLCγ1 impaired pre-TCR mediated Ca2+ flux and Erk activation. Thus our studies demonstrate that PLCγ1 is important for pre-TCR mediated signal transduction and pre-T cell development.
Assuntos
Diferenciação Celular , Fosfolipase C gama/metabolismo , Células Precursoras de Linfócitos T/citologia , Células Precursoras de Linfócitos T/metabolismo , Receptores de Antígenos de Linfócitos T/metabolismo , Transdução de Sinais , Animais , Biomarcadores , Cálcio/metabolismo , Diferenciação Celular/genética , Diferenciação Celular/imunologia , Proliferação de Células , Sobrevivência Celular/genética , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Expressão Gênica , Genótipo , Ativação Linfocitária/genética , Ativação Linfocitária/imunologia , Camundongos , Camundongos Transgênicos , Fosfolipase C gama/deficiência , Fosfolipase C gama/genética , Fosforilação , Células Precursoras de Linfócitos T/imunologia , Receptores de Antígenos de Linfócitos T/genética , Receptores de Antígenos de Linfócitos T alfa-beta/genética , Receptores de Antígenos de Linfócitos T alfa-beta/metabolismo , Timócitos/citologia , Timócitos/imunologia , Timócitos/metabolismoRESUMO
The three major Ras members, Kras, Hras, and Nras, are highly homologous and individual Ras genes can have distinct biological functions. Embryonic lethality of Kras-deficient mice precludes study of the biological functions of this Ras family member. In this study, we generated and examined mice with hematopoietic-specific deletion of Kras and bone marrow (BM) chimeric mice with B cell-specific targeted deletion of Kras. Hematopoietic-specific deletion of Kras impaired early B cell development at the pre-B cell stage and late B cell maturation, resulting in the reduction of BM pre-, immature, and mature B cells and peripheral follicular, marginal zone, and B1 mature B cells. In contrast, Kras deficiency did not affect T cell development. Studies of BM chimeric mice with B cell-specific deletion of Kras demonstrated that Kras deficiency intrinsically impaired B cell development. Kras deficiency reduced BCR-induced B cell proliferation and survival. Furthermore, Kras deficiency specifically impaired pre-BCR- and BCR-induced activation of the Raf-1/MEK/ERK pathway in pre-B and mature B cells, respectively. Thus, Kras is the unique Ras family member that plays a critical role in early B cell development and late B cell maturation through controlling the Raf-1/MEK/ERK pathway.
Assuntos
Linfócitos B/citologia , Ativação Linfocitária/imunologia , Proteínas Proto-Oncogênicas p21(ras)/imunologia , Transdução de Sinais/imunologia , Animais , Linfócitos B/imunologia , Western Blotting , Diferenciação Celular/imunologia , Proliferação de Células/fisiologia , Ensaio de Desvio de Mobilidade Eletroforética , Citometria de Fluxo , Linfopoese/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Células Precursoras de Linfócitos B/citologia , Células Precursoras de Linfócitos B/imunologiaRESUMO
Antibodies specific for platelet factor 4 (PF4)/heparin complexes are central to the pathogenesis of heparin-induced thrombocytopenia. Marginal zone B cells appear to be the source of such antibodies, but whether T-cell help is required is unclear. Here, we showed that induction of PF4/heparin-specific antibodies by PF4/heparin complexes was markedly impaired in mice depleted of CD4 T cells by anti-CD4 antibodies. Furthermore, Rag1-deficient recipient mice produced PF4/heparin-specific antibodies upon PF4/heparin challenge when reconstituted with a mixture of wild-type splenic B cells and splenocytes from B-cell-deficient (µMT) mice but not splenocytes from T- and B-cell-deficient (Rag1 knockout) mice. Lastly, mice with B cells lacking CD40, a B-cell costimulatory molecule that helps T-cell-dependent B-cell responses, displayed a marked reduction of PF4/heparin-specific antibody production following PF4/heparin challenge. Together, these findings show that helper T cells play a critical role in production of PF4/heparin-specific antibodies.
Assuntos
Formação de Anticorpos , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD4-Positivos/metabolismo , Heparina/imunologia , Fator Plaquetário 4/imunologia , Transferência Adotiva , Animais , Especificidade de Anticorpos , Linfócitos B/imunologia , Modelos Animais de Doenças , Heparina/efeitos adversos , Heparina/química , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/imunologia , Humanos , Imunização , Depleção Linfocítica , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Fator Plaquetário 4/química , Trombocitopenia/sangue , Trombocitopenia/etiologia , Trombocitopenia/imunologia , Quimeras de TransplanteRESUMO
Immune complexes consisting of heparin, platelet factor 4 (PF4), and PF4/heparin-reactive antibodies are central to the pathogenesis of heparin-induced thrombocytopenia (HIT). It is as yet unclear what triggers the initial induction of pathogenic antibodies. We identified B cells in peripheral blood of healthy adults that produce PF4/heparin-specific antibodies following in vitro stimulation with proinflammatory molecules containing deoxycytosine-deoxyguanosine (CpG). Similarly, B cells from unmanipulated wild-type mice produced PF4/heparin-specific antibodies following in vitro or in vivo CpG stimulation. Thus, both healthy humans and mice possess preexisting inactive/tolerant PF4/heparin-specific B cells. The findings suggest that breakdown of tolerance leads to PF4/heparin-specific B-cell activation and antibody production in patients developing HIT. Consistent with this concept, mice lacking protein kinase Cδ (PKCδ) that are prone to breakdown of B-cell tolerance produced anti-PF4/heparin antibodies spontaneously. Therefore, breakdown of tolerance can lead to PF4/heparin-specific antibody production, and B-cell tolerance may play an important role in HIT pathogenesis.
Assuntos
Formação de Anticorpos/imunologia , Anticoagulantes/efeitos adversos , Linfócitos B/imunologia , Heparina/efeitos adversos , Fator Plaquetário 4/metabolismo , Proteína Quinase C-delta/fisiologia , Trombocitopenia/imunologia , Adulto , Animais , Anticoagulantes/metabolismo , Linfócitos B/metabolismo , Linfócitos B/patologia , Células Cultivadas , Heparina/metabolismo , Humanos , Tolerância Imunológica , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Fator Plaquetário 4/imunologia , Prognóstico , Trombocitopenia/induzido quimicamente , Trombocitopenia/metabolismoRESUMO
Heparin-induced thrombocytopenia (HIT) is an immune-mediated disorder that can cause fatal arterial or venous thrombosis/thromboembolism. Immune complexes consisting of platelet factor 4 (PF4), heparin, and PF4/heparin-reactive antibodies are central to the pathogenesis of HIT. However, the B-cell origin of HIT antibody production is not known. Here, we show that anti-PF4/heparin antibodies are readily generated in wild-type mice on challenge with PF4/heparin complexes, and that antibody production is severely impaired in B-cell-specific Notch2-deficient mice that lack marginal zone (MZ) B cells. As expected, Notch2-deficient mice responded normally to challenge with T-cell-dependent antigen nitrophenyl-chicken γ globulin but not to the T-cell-independent antigen trinitrophenyl-Ficoll. In addition, wild-type, but not Notch2-deficient, B cells plus B-cell-depleted wild-type splenocytes adoptively transferred into B-cell-deficient µMT mice responded to PF4/heparin complex challenge. PF4/heparin-specific antibodies produced by wild-type mice were IgG2b and IgG3 isotypes. An in vitro class-switching assay showed that MZ B cells were capable of producing antibodies of IgG2b and IgG3 isotypes. Lastly, MZ, but not follicular, B cells adoptively transferred into B-cell-deficient µMT mice responded to PF4/heparin complex challenge by producing PF4/heparin-specific antibodies of IgG2b and IgG3 isotypes. Taken together, these data demonstrate that MZ B cells are critical for PF4/heparin-specific antibody production.
Assuntos
Formação de Anticorpos , Autoanticorpos/imunologia , Linfócitos B/imunologia , Heparina/imunologia , Fator Plaquetário 4/imunologia , Trombocitopenia/imunologia , Transferência Adotiva , Animais , Anticoagulantes/efeitos adversos , Anticoagulantes/imunologia , Células Apresentadoras de Antígenos/imunologia , Autoanticorpos/sangue , Linfócitos B/química , Coagulantes/efeitos adversos , Coagulantes/imunologia , Citometria de Fluxo , Heparina/efeitos adversos , Imunização , Switching de Imunoglobulina/imunologia , Imunoglobulina G/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Fator Plaquetário 4/efeitos adversos , Receptor Notch2/fisiologia , Trombocitopenia/induzido quimicamente , Trombocitopenia/diagnósticoRESUMO
The infiltration and cytotoxicity of chimeric antigen receptor (CAR)-T cells are crucial for effective elimination of solid tumors. While metallo-immunotherapy is a promising strategy that can activate the antitumor immunity, its role in promoting CAR-T cell therapy remains elusive. The first single-element nanomaterial based on chromium nanoparticles (Cr NPs) for cancer photo-metallo-immunotherapy has been reported previously. Herein, an extended study using biodegradable polydopamine as a versatile carrier for these nanoparticles, enabling synergistic CAR-T cell therapy, is reported. The results show that these nanocomposites with or without further encapsulation of the anticancer drug alpelisib can promote the CAR-T cell migration and antitumor effect. Upon irradiation with near-infrared light, they caused mild hyperthermia that can "warm" the "cold" tumor microenvironment (TME). The administration of B7-H3 CAR-T cells to NOD severe combined immunodeficiency gamma mice bearing a human hepatoma or PIK3CA-mutated breast tumor can significantly inhibit the tumor growth after the induction of tumor hyperthermia by the nanocomposites and promote the secretion of serum cytokines, including IL-2, IFN-γ, and TNF-α. The trivalent Cr3+ ions, which are the major degradation product of these nanocomposites, can increase the CXCL13 and CCL3 chemokine expressions to generate tertiary lymphoid structures (TLSs) in the tumor tissues, facilitating the CAR-T cell infiltration.
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The highly conserved MicroRNA-9 (miR-9) family consists of three members. We discovered that miR-9-1 deletion reduced mature miR-9 expression, causing 43% of the mice to display smaller size and postweaning lethality. MiR-9-1-deficient mice with growth defects experienced severe lymphopenia, but other blood cells were unaffected. The lymphopenia wasn't due to defects in hematopoietic progenitors, as mutant bone marrow (BM) cells underwent normal lymphopoiesis after transplantation into wild-type recipients. Additionally, miR-9-1-deficient mice exhibited impaired osteoblastic bone formation, as mutant mesenchymal stem cells (MSCs) failed to differentiate into osteoblastic cells (OBs). RNA sequencing revealed reduced expression of master transcription factors for osteoblastic differentiation, Runt-related transcription factor 2 (Runx2) and Osterix (Osx), and genes related to collagen formation, extracellular matrix organization, and cell adhesion, in miR-9-1-deficient MSCs. Follistatin (Fst), an antagonist of bone morphogenetic proteins (BMPs), was found to be a direct target of miR-9-1. Its deficiency led to the up-regulation of Fst, inhibiting BMP signaling in MSCs, and reducing IL-7 and IGF-1. Thus, miR-9-1 controls osteoblastic regulation of lymphopoiesis by targeting the Fst/BMP/Smad signaling axis.
Assuntos
Linfopenia , MicroRNAs , Animais , Camundongos , Linfopoese/genética , Proteínas Morfogenéticas Ósseas/metabolismo , Diferenciação Celular , MicroRNAs/genética , MicroRNAs/metabolismo , Osteogênese/genética , Osteoblastos/metabolismoRESUMO
The practical application of lithium-metal batteries (LMBs) is hindered by the lithium dendrite formation during cycling. In this work, we report a multilayered solid polymer electrolyte (SPE) formed by sandwiching a comb-chain cross-linker-based network SPE (ConSPE) film with a linear poly(ethylene oxide) (PEO) SPE coating. Benefiting from the drastically different lithium dendrite resisting properties of the ConSPE and linear PEO SPE, the lithium dendrite growth in the multilayered SPEs could be tuned, with the linear PEO SPE effectively serving as a sacrificial layer to accommodate the lithium dendrite growth. Symmetrical lithium cells with the multilayered SPE exhibited an extended short-circuit time â¼4.1 times that for the single-layer ConSPE at a high current density of 1.5 mA cm-2. Li/LiFePO4 batteries with multilayered SPEs delivered superior cycling performance at extremely high C-rates of 2C and 10C. Our multilayered SPE architecture, therefore, opens up a new gateway for advancing SPE design for future LMBs.
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C-type lectin-like molecule-1 (CLL1) is preferentially expressed on acute myeloid leukemia (AML) stem cells and AML blasts, which can be considered as AML-associated antigen. Anti-CLL1-based CAR-T cells exhibited effective tumor-killing capacity in vitro and in AML-bearing mouse model. In this report, eight children with relapsed or refractory AML (R/R-AML) were recruited for a phase 1/2 clinical trial of autologous anti-CLL1 CAR-T cell immunotherapy. The objectives of this clinical trial were to evaluate the safety and the preliminary efficacy of anti-CLL1 CAR-T cell treatment. Patients received one dose of autologous anti-CLL1 CAR-T cells after lymphodepletion conditioning. After CAR-T treatment, patients developed grade 1-2 cytokine release syndrome (CRS) but without any lethal events. 4 out of 8 patients achieved morphologic leukemia-free state (MLFS) and minimal residual disease (MRD) negativity, 1 patient with MLFS and MRD positivity, 1 patient achieved complete remission with incomplete hematologic recovery (CRi) but MRD positivity, 1 patient with partial remission (PR), and 1 patient remained at stable disease (SD) status but had CLL1-positive AML blast clearance. These results suggested that anti-CLL1-based CAR-T cell immunotherapy can be considered as a well-tolerated and effective option for treating children with R/R-AML.
Assuntos
Transplante de Células-Tronco Hematopoéticas , Leucemia Mieloide Aguda , Receptores de Antígenos Quiméricos , Camundongos , Animais , Imunoterapia Adotiva/efeitos adversos , Imunoterapia Adotiva/métodos , Leucemia Mieloide Aguda/tratamento farmacológico , Lectinas Tipo C , Síndrome da Liberação de CitocinaRESUMO
The Switch/Sugar Non-Fermenting (SWI/SNF) nucleosome remodeling complexes play important roles in normal development and in the development of various cancers. Core subunits of the SWI/SNF complexes have been shown to have oncogenic roles in acute myeloid leukemia. However, the roles of the unique targeting subunits, including that of Arid2 and Arid1b, in AML leukemogenesis are not well understood. Here, we used conditional knockout mouse models to elucidate their role in MLL-AF9 leukemogenesis. We uncovered that Arid2 has dual roles; enhancing leukemogenesis when deleted during leukemia initiation and yet is required during leukemia maintenance. Whereas, deleting Arid1b in either phase promotes leukemogenesis. Our integrated analyses of transcriptomics and genomic binding data showed that, globally, Arid2 and Arid1b regulate largely distinct sets of genes at different disease stages, respectively, and in comparison, to each other. Amongst the most highly dysregulated transcription factors upon their loss, Arid2 and Arid1b converged on the regulation of Etv4/Etv5, albeit in an opposing manner while also regulating distinct TFs including Gata2,Tcf4, Six4, Irf4 and Hmgn3. Our data demonstrate the differential roles of SWI/SNF subunits in AML leukemogenesis and emphasize that cellular context and disease stage are key in determining their functions during this process.
Assuntos
Leucemia , Fatores de Transcrição , Animais , Carcinogênese/genética , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Humanos , Leucemia/genética , Camundongos , Camundongos Knockout , Proteína de Leucina Linfoide-Mieloide/genética , Proteínas de Fusão Oncogênica , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismoRESUMO
B7-H3 is over-expressed in multiple types of solid tumors, making it an ideal target for chimeric antigen receptor (CAR)-T therapy. Here, we first report a case of multiple basal cell carcinoma (BCC) patient treated with humanized monoclonal anti-B7-H3 CAR-T cells through direct intratumoral injection. After three dose-escalated injections, the lesion in the abdomen decreased by 40% in volume, shrank from bulging to flat, but was not eradicated completely. The large lesion in the forehead became dry from original ulcer and bleeding. The adverse events observed were itching, myalgia, and redness. Immunohistochemistry analysis demonstrated that B7-H3-positive tumor cells and B7-H3 expression intensity were reduced after injections of CAR-T cells. The number of infiltrating CD3 T cells increased significantly but mainly located outside the tumor region. Subsequently, high levels of TGF-ß in the tumor area were observed, suggesting that solid tumor microenvironment may hinder the infiltration and effect of CAR-T cells. In summary, in this particular case report, intratumoral injection of B7-H3 CAR-T cells partially controls tumor growth in the BCC patient with minor adverse events. The efficacy and safety of B7-H3 CAR-T therapy need to be further investigated with a larger cohort of patients. Although only one clinical case is reported here, the anti-B7-H3 CAR-T cell therapy should be considered as a treatment option for solid tumors in the future. This clinical trial was registered at the Chinese Clinical Trial Registry (www.chictr.org.cn) with registration number ChiCTR2100044386.
RESUMO
Acute myeloid leukemia (AML) is driven by mutations that occur in numerous combinations. A better understanding of how mutations interact with one another to cause disease is critical to developing targeted therapies. Approximately 50% of patients that harbor a common mutation in NPM1 (NPM1cA) also have a mutation in the cohesin complex. As cohesin and Npm1 are known to regulate gene expression, we sought to determine how cohesin mutation alters the transcriptome in the context of NPM1cA. We utilized inducible Npm1cAflox/+ and core cohesin subunit Smc3flox/+ mice to examine AML development. While Npm1cA/+;Smc3Δ/+ mice developed AML with a similar latency and penetrance as Npm1cA/+ mice, RNA-seq suggests that the Npm1cA/+; Smc3Δ/+ mutational combination uniquely alters the transcriptome. We found that the Rac1/2 nucleotide exchange factor Dock1 was specifically upregulated in Npm1cA/+;Smc3Δ/+ HSPCs. Knockdown of Dock1 resulted in decreased growth and adhesion and increased apoptosis only in Npm1cA/+;Smc3Δ/+ AML. Higher Rac activity was also observed in Npm1cA/+;Smc3Δ/+ vs. Npm1cA/+ AMLs. Importantly, the Dock1/Rac pathway is targetable in Npm1cA/+;Smc3Δ/+ AMLs. Our results suggest that Dock1/Rac represents a potential target for the treatment of patients harboring NPM1cA and cohesin mutations and supports the use of combinatorial genetics to identify novel precision oncology targets.