RESUMO
We have recently reported that transcription factor Runx3 is required for pulmonary generation of CD8+ cytotoxic T lymphocytes (CTLs) that play a crucial role in the clearance of influenza A virus (IAV). To understand the underlying mechanisms, we determined the effects of Runx3 knockout (KO) on CD8+ T cell local expansion and phenotypes using an inducible general Runx3 KO mouse model. We found that in contrast to the lungs, Runx3 general KO promoted enlargement of lung-draining mediastinal lymph node (mLN) and enhanced CD8+ and CD4+ T cell expansion during H1N1 IAV infection. We further found that Runx3 deficiency greatly inhibited core 2 O-glycosylation of selectin ligand CD43 on activated CD8+ T cells but minimally affected the cell surface expression of CD43, activation markers (CD44 and CD69) and cell adhesion molecules (CD11a and CD54). Runx3 KO had a minor effect on lung effector CD8+ T cell death by IAV infection. Our findings indicate that Runx3 differently regulates CD8+ T cell expansion in mLNs and lungs by H1N1 IAV infection. Runx3 is required for CD43 core 2 O-glycosylation on activated CD8+ T cells, and the involved Runx3 signal pathway may mediate CD8+ T cell phenotype for pulmonary generation of CTLs.
Assuntos
Linfócitos T CD8-Positivos , Subunidade alfa 3 de Fator de Ligação ao Core , Vírus da Influenza A Subtipo H1N1 , Infecções por Orthomyxoviridae , Animais , Camundongos , Linfócitos T CD8-Positivos/imunologia , Linfócitos T CD8-Positivos/metabolismo , Proliferação de Células , Subunidade alfa 3 de Fator de Ligação ao Core/metabolismo , Subunidade alfa 3 de Fator de Ligação ao Core/genética , Glicosilação , Vírus da Influenza A Subtipo H1N1/imunologia , Leucossialina/metabolismo , Pulmão/virologia , Pulmão/metabolismo , Pulmão/imunologia , Pulmão/patologia , Linfonodos/metabolismo , Linfonodos/imunologia , Ativação Linfocitária/imunologia , Camundongos Endogâmicos C57BL , Camundongos Knockout , Infecções por Orthomyxoviridae/imunologia , Infecções por Orthomyxoviridae/metabolismo , Infecções por Orthomyxoviridae/virologiaRESUMO
BACKGROUND: To present an unusual case of abnormal LCA expression and CD43 in SCLC and to review the reported literature to avoid potential diagnostic pitfalls. CASE PRESENTATION: A 73-year-old male patient suffered from persistent back pain for more than one month. MRI revealed a compression fracture of the L1-L5 vertebra. A CT scan revealed multiple nodules and masses at the left root of the neck, lung hilum and mediastinum, and multiple areas of bony destruction of the ribs. Histology of the tumor revealed that small and round cells were arranged in nests with areas of necrosis. The tumor cells were round to ovoid with scant cytoplasm and indistinct cell borders. The nuclear chromatin was finely granular, and the nucleoli were absent or inconspicuous. Immunohistochemically, the tumor cells were positive for cytokeratin, TTF-1, POU2F3, LCA, and CD43. CONCLUSION: This report highlights a potential diagnostic pitfall in the diagnosis of SCLC, urges pathologists to exercise caution in cases of LCA and CD43 positivity and illustrates the need for further immunohistochemical studies to avoid misdiagnosis.
Assuntos
Leucossialina , Neoplasias Pulmonares , Humanos , Masculino , Idoso , Leucossialina/metabolismo , Neoplasias Pulmonares/diagnóstico , Neoplasias Pulmonares/patologia , Carcinoma de Pequenas Células do Pulmão/diagnóstico , Carcinoma de Pequenas Células do Pulmão/patologia , Carcinoma de Pequenas Células do Pulmão/metabolismo , Tomografia Computadorizada por Raios X , Imuno-Histoquímica , Biomarcadores Tumorais/metabolismoRESUMO
Staphylococcus aureus is a noteworthy pathogen in allergic diseases, as four staphylococcal exotoxins activate mast cells, a significant contributor to inflammation, in an IgE-independent manner. Although the adhesion of mast cells is an essential process for their immune responses, only a small number of exotoxins have been reported to affect the process. Here, we demonstrated that staphylococcal superantigen-like (SSL) 3, previously identified as a toll-like receptor 2 agonist, induced the adhesion of murine bone marrow-derived mast cells to culture substratum. SSL3-induced adhesion was mediated by fibronectin in an Arg-Gly-Asp (RGD) sequence-dependent manner, suggesting the integrins were involved in the process. Additionally, SSL3 was found to bind to an anti-adhesive surface protein CD43. SSL3 induced the adhesion of HEK293 cells expressing exogenous CD43, suggesting that CD43 is the target molecule for adhesion induced by SSL3. Evaluation of SSL3-derived mutants showed that the C-terminal region (253-326), specifically T285 and H307, are necessary to induce adhesion. SSL3 augmented the IL-13 production of mast cells in response to immunocomplex and SSL12. These findings reveal a novel function of SSL3, triggering cell adhesion and enhancing mast cell activation. This study would clarify the correlation between S. aureus and allergic diseases such as atopic dermatitis.
Assuntos
Adesão Celular , Leucossialina , Mastócitos , Staphylococcus aureus , Superantígenos , Animais , Mastócitos/metabolismo , Mastócitos/imunologia , Camundongos , Humanos , Superantígenos/metabolismo , Staphylococcus aureus/metabolismo , Staphylococcus aureus/imunologia , Células HEK293 , Leucossialina/metabolismo , Proteínas de Bactérias/metabolismo , Interleucina-13/metabolismo , Camundongos Endogâmicos C57BLRESUMO
Newborns are unable to reach the adult-level humoral immune response partly due to the potent immunoregulatory role of IL-10. Increased IL-10 production by neonatal B cells has been attributed to the larger population of IL-10-producting CD43+ B-1 cells in neonates. Here, we show that neonatal mouse CD43- non-B-1 cells also produce substantial amounts of IL-10 following B cell antigen receptor (BCR) activation. In neonatal mouse CD43- non-B-1 cells, BCR engagement activated STAT5 under the control of phosphorylated forms of signaling molecules Syk, Btk, PKC, FAK, and Rac1. Neonatal STAT5 activation led to IL-6 production, which in turn was responsible for IL-10 production in an autocrine/paracrine fashion through the activation of STAT3. In addition to the increased IL-6 production in response to BCR stimulation, elevated expression of IL-6Rα expression in neonatal B cells rendered them highly susceptible to IL-6-mediated STAT3 phosphorylation and IL-10 production. Finally, IL-10 secreted from neonatal mouse CD43- non-B-1 cells was sufficient to inhibit TNF-α secretion by macrophages. Our results unveil a distinct mechanism of IL-6-dependent IL-10 production in BCR-stimulated neonatal CD19+CD43- B cells.
Assuntos
Interleucina-10 , Interleucina-6 , Animais , Camundongos , Animais Recém-Nascidos , Antígenos CD19/metabolismo , Interleucina-10/metabolismo , Interleucina-6/metabolismo , Fosforilação , Receptores de Antígenos de Linfócitos B/metabolismo , Fator de Transcrição STAT5/metabolismo , Leucossialina/imunologiaRESUMO
Nascent HIV-1 particles incorporate the viral envelope glycoprotein and multiple host transmembrane proteins during assembly at the plasma membrane. At least some of these host transmembrane proteins on the surface of virions are reported as pro-viral factors that enhance virus attachment to target cells or facilitate trans-infection of CD4+ T cells via interactions with non-T cells. In addition to the pro-viral factors, anti-viral transmembrane proteins are incorporated into progeny virions. These virion-incorporated transmembrane proteins inhibit HIV-1 entry at the point of attachment and fusion. In infected polarized CD4+ T cells, HIV-1 Gag localizes to a rear-end protrusion known as the uropod. Regardless of cell polarization, Gag colocalizes with and promotes the virion incorporation of a subset of uropod-directed host transmembrane proteins, including CD162, CD43, and CD44. Until recently, the functions of these virion-incorporated proteins had not been clear. Here, we review the recent findings about the roles played by virion-incorporated CD162, CD43, and CD44 in HIV-1 spread to CD4+ T cells.
Assuntos
Infecções por HIV/metabolismo , Receptores de Hialuronatos/metabolismo , Leucossialina/metabolismo , Glicoproteínas de Membrana/metabolismo , Membrana Celular/metabolismo , Infecções por HIV/genética , HIV-1/genética , HIV-1/metabolismo , HIV-1/patogenicidade , Interações Hospedeiro-Patógeno , Humanos , Receptores de Hialuronatos/genética , Leucossialina/genética , Glicoproteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Linfócitos T/metabolismo , Linfócitos T/virologia , Vírion/metabolismo , Montagem de Vírus , Ligação Viral , Produtos do Gene gag do Vírus da Imunodeficiência Humana/metabolismoRESUMO
Moreau score has been used to differentiate chronic lymphocytic leukemia (CLL) from other mature B-cell neoplasms. However, it showed limitations in Asian patients. Therefore, we conducted a new score system replacing CD5 and CD23 with CD43 and CD180 to evaluate its diagnostic value of CLL. 237 untreated samples diagnosed with mature B-cell neoplasms were collected and were randomly divided into an exploratory and a validation cohort by a 2:1 ratio. The expression of CD5, CD19, CD20, CD23, CD43, CD79b, CD180, CD200, FMC7, and surface immunoglobulin (SmIg) were analyzed among all the samples. A proposed score was developed based on the logistic regression model. The sensitivity and specificity of the proposed score were calculated by ROC curves. CD43/CD180, CD200, FMC7, and CD79b were included in our new CLL score, which showed a sensitivity of 91.8% and a specificity of 83.1%. These results were confirmed in a validation cohort with a sensitivity of 90.5% (p = 0.808) and a specificity of 79.5% (p = 0.639). In CD5 negative or CD23 negative CLL group, the new CLL score displayed improved sensitivity of 79.4% compared to Moreau score and CLLflow score (41.2% and 47.1%, respectively). In atypical CLL group, the new CLL score showed improved sensitivity of 84.2% compared to Moreau score and CLLflow score (61.4% and 64.9%, respectively). This proposed atypical CLL score helped to offer an accurate differentiation of CLL from non-CLL together with morphological and molecular methods, particularly in Chinese patients with atypical immunophenotype.
Assuntos
Antígenos CD/análise , Biomarcadores Tumorais/análise , Leucemia Linfocítica Crônica de Células B/diagnóstico , Leucossialina/análise , Antígenos CD19/análise , Antígenos CD20/análise , Antígenos CD5/análise , Antígenos CD79/análise , Diagnóstico Diferencial , Citometria de Fluxo/métodos , Glicoproteínas/análise , Humanos , Imunofenotipagem , Leucemia Linfocítica Crônica de Células B/metabolismo , Modelos Logísticos , Linfoma de Células B/diagnóstico , Linfoma de Células B/metabolismo , Curva ROC , Receptores de Antígenos de Linfócitos B/análise , Receptores de IgE/análise , Sensibilidade e EspecificidadeRESUMO
Humoral immune responses depend on the generation of high-affinity antigen-specific antibodies. Germinal center (GC) B cells are the cornerstone of this response in peripheral lymphoid organs. High purities of GC B cells, and also naive B cells, are required for accurate analysis in downstream assays to yield essential knowledge on immunity. This protocol lays out quick and easy steps to purify GC B cells from spleens of immunized mice or B cells from naive animals by negative selection using MACS. For complete details on the use and execution of this protocol, please refer to Ramezani-Rad et al. (2020).
Assuntos
Separação Celular/métodos , Separação Imunomagnética/métodos , Animais , Linfócitos B/citologia , Centro Germinativo/citologia , Imunidade Humoral , Imunização , Leucossialina/imunologia , Camundongos , Baço/imunologiaRESUMO
Cells release diverse types of extracellular vesicles (EVs), which transfer complex signals to surrounding cells. Specific markers to distinguish different EVs (e.g. exosomes, ectosomes, enveloped viruses like HIV) are still lacking. We have developed a proteomic profiling approach for characterizing EV subtype composition and applied it to human Jurkat T cells. We generated an interactive database to define groups of proteins with similar profiles, suggesting release in similar EVs. Biochemical validation confirmed the presence of preferred partners of commonly used exosome markers in EVs: CD81/ADAM10/ITGB1, and CD63/syntenin. We then compared EVs from control and HIV-1-infected cells. HIV infection altered EV profiles of several cellular proteins, including MOV10 and SPN, which became incorporated into HIV virions, and SERINC3, which was re-routed to non-viral EVs in a Nef-dependent manner. Furthermore, we found that SERINC3 controls the surface composition of EVs. Our workflow provides an unbiased approach for identifying candidate markers and potential regulators of EV subtypes. It can be widely applied to in vitro experimental systems for investigating physiological or pathological modifications of EV release.
Assuntos
Vesículas Extracelulares/metabolismo , Infecções por HIV/metabolismo , Proteoma/metabolismo , Células Cultivadas , Células HEK293 , HIV-1 , Humanos , Células Jurkat , Leucossialina/metabolismo , Glicoproteínas de Membrana/metabolismo , RNA Helicases/metabolismoRESUMO
Cell-cell contact participates in the process of mesenchymal stromal cell (MSC)-mediated T cell modulation and thus contributes to MSC-based therapies for various inflammatory diseases, especially T cell-mediated diseases. However, the mechanisms underlying the adhesion interactions between MSCs and T cells are still poorly understood. In this study, we explored the interaction between MSCs and T cells and found that activated T cells could rapidly adhere to MSCs, leading to significant reduction of TNF-α and IFN-γ mRNA expression. Furthermore, TCR-proximal signaling in activated T cells was also dramatically suppressed in the MSC co-culture, resulting in weakened Ca2+ signaling. MSCs rapidly suppressed TCR signaling and its downstream signaling in a cell-cell contact-dependent manner, partially through the ICAM-1/CD43 adhesion interaction. Blockade of either ICAM-1 on MSCs or CD43 on T cells significantly reversed this rapid suppression of proinflammatory cytokine expression in T cells. Mechanistically, MSC-derived ICAM-1 likely disrupts CD43-mediated TCR microcluster formation to limit T cell activation. Taken together, our results reveal a fast mechanism of activated T cell inhibition by MSCs, which provides new clues to unravel the MSC-mediated immunoregulatory mechanism for aGVHD and other severe acute T cell-related diseases.
Assuntos
Citocinas/metabolismo , Molécula 1 de Adesão Intercelular/metabolismo , Leucossialina/metabolismo , Ativação Linfocitária/imunologia , Células-Tronco Mesenquimais/metabolismo , Receptores de Antígenos de Linfócitos T/metabolismo , Linfócitos T/metabolismo , Biomarcadores , Cálcio , Comunicação Celular/imunologia , Humanos , Ativação Linfocitária/genética , Células-Tronco Mesenquimais/imunologia , Ligação Proteica , Transdução de Sinais , Linfócitos T/imunologiaRESUMO
Sialic acid (Sia)-binding immunoglobulin-like lectin 7 (Siglec-7) is an inhibitory receptor primarily expressed on natural killer (NK) cells and monocytes. Siglec-7 is known to negatively regulate the innate immune system through Sia binding to distinguish self and nonself; however, a counter-receptor bearing its natural ligand remains largely unclear. Here, we identified a counter-receptor of Siglec-7 using K562 hematopoietic carcinoma cells presenting cell surface ligands for Siglec-7. We affinity-purified the ligands using Fc-ligated recombinant Siglec-7 and diSia-dextran polymer, a strong inhibitor for Siglec-7. We then confirmed the counter-receptor for Siglec-7 as leukosialin (CD43) through mass spectrometry, immunoprecipitation, and proximity labeling. Additionally, we demonstrated that the cytotoxicity of NK cells toward K562 cells was suppressed by overexpression of leukosialin in a Siglec-7-dependent manner. Taken together, our data suggest that leukosialin on K562 is a counter-receptor for Siglec-7 on NK cells and that a cluster of the Sia-containing glycan epitope on leukosialin is key as trans-ligand for unmasking the cis-ligand.
Assuntos
Antígenos de Diferenciação Mielomonocítica/metabolismo , Células K562/metabolismo , Lectinas/metabolismo , Leucossialina/metabolismo , Antígenos de Diferenciação Mielomonocítica/genética , Linhagem Celular Tumoral , Cromatografia de Afinidade/métodos , Humanos , Células Matadoras Naturais/metabolismo , Lectinas/genética , Leucossialina/imunologia , Ligantes , Proteínas de Membrana/metabolismo , Monócitos/metabolismo , Polissacarídeos/metabolismo , Lectinas Semelhantes a Imunoglobulina de Ligação ao Ácido Siálico/genética , Lectinas Semelhantes a Imunoglobulina de Ligação ao Ácido Siálico/metabolismoRESUMO
Aberrant expression of CD43 in malignant tumors of nonhematopoietic origin such as those from lung, cervix, colon, and breast has been shown to correlate with poor prognosis, providing tumor cells with enhanced motility, anchorage-independent growth, and in vivo tumor size, while protecting the cells of NK lysis and apoptosis. To further characterize the role of CD43 in cell transformation, we tested whether interfering its expression modified the capacity of the A549 non-small cell lung cancer cells to secrete molecules contributing to malignancy. The proteomic analysis of the secretome of serum-starved A549 cells revealed that cells expressing normal levels of CD43 released significantly high levels of molecules involved in extracellular matrix organization, angiogenesis, platelet degranulation, collagen degradation, and inflammation, as compared to CD43 RNAi cells. This data reveals a novel and unexpected role for CD43 in lung cancer development, mainly in remodeling the tumor microenvironment.
Assuntos
Matriz Extracelular/metabolismo , Leucossialina/metabolismo , Neoplasias Pulmonares/irrigação sanguínea , Neoplasias Pulmonares/metabolismo , Neovascularização Patológica/metabolismo , Células A549 , Inativação Gênica , Humanos , Metaloproteinase 2 da Matriz/metabolismo , Metaloproteinase 9 da Matriz/metabolismo , NF-kappa B/metabolismo , Fator de Transcrição STAT3/metabolismo , Microambiente TumoralRESUMO
BACKGROUND: T-cell acute lymphoblastic leukemia (T-ALL) is an aggressive disease with a poor cure rate for relapsed/resistant patients. Due to the lack of T-cell restricted targetable antigens, effective immune-therapeutics are not presently available and the treatment of chemo-refractory T-ALL is still an unmet clinical need. To develop novel immune-therapy for T-ALL, we generated an afucosylated monoclonal antibody (mAb) (ahuUMG1) and two different bispecific T-cell engagers (BTCEs) against UMG1, a unique CD43-epitope highly and selectively expressed by T-ALL cells from pediatric and adult patients. METHODS: UMG1 expression was assessed by immunohistochemistry (IHC) on a wide panel of normal tissue microarrays (TMAs), and by flow cytometry on healthy peripheral blood/bone marrow-derived cells, on 10 different T-ALL cell lines, and on 110 T-ALL primary patient-derived cells. CD43-UMG1 binding site was defined through a peptide microarray scanning. ahuUMG1 was generated by Genetic Glyco-Engineering technology from a novel humanized mAb directed against UMG1 (huUMG1). BTCEs were generated as IgG1-(scFv)2 constructs with bivalent (2+2) or monovalent (2+1) CD3ε arms. Antibody dependent cellular cytotoxicity (ADCC), antibody dependent cellular phagocytosis (ADCP) and redirected T-cell cytotoxicity assays were analysed by flow cytometry. In vivo antitumor activity of ahUMG1 and UMG1-BTCEs was investigated in NSG mice against subcutaneous and orthotopic xenografts of human T-ALL. RESULTS: Among 110 T-ALL patient-derived samples, 53 (48.1%) stained positive (24% of TI/TII, 82% of TIII and 42.8% of TIV). Importantly, no expression of UMG1-epitope was found in normal tissues/cells, excluding cortical thymocytes and a minority (<5%) of peripheral blood T lymphocytes. ahUMG1 induced strong ADCC and ADCP on T-ALL cells in vitro, which translated in antitumor activity in vivo and significantly extended survival of treated mice. Both UMG1-BTCEs demonstrated highly effective killing activity against T-ALL cells in vitro. We demonstrated that this effect was specifically exerted by engaged activated T cells. Moreover, UMG1-BTCEs effectively antagonized tumor growth at concentrations >2 log lower as compared with ahuUMG1, with significant mice survival advantage in different T-ALL models in vivo. CONCLUSION: Altogether our findings, including the safe UMG1-epitope expression profile, provide a framework for the clinical development of these innovative immune-therapeutics for this still orphan disease.
Assuntos
Anticorpos Biespecíficos/farmacologia , Anticorpos Monoclonais Humanizados/farmacologia , Antineoplásicos Imunológicos/farmacologia , Leucossialina/agonistas , Leucemia-Linfoma Linfoblástico de Células T Precursoras/tratamento farmacológico , Linfócitos T/efeitos dos fármacos , Animais , Especificidade de Anticorpos , Proliferação de Células/efeitos dos fármacos , Citotoxicidade Imunológica/efeitos dos fármacos , Epitopos , Feminino , Humanos , Células Jurkat , Leucossialina/metabolismo , Ativação Linfocitária/efeitos dos fármacos , Camundongos Endogâmicos NOD , Camundongos SCID , Fagocitose/efeitos dos fármacos , Leucemia-Linfoma Linfoblástico de Células T Precursoras/genética , Leucemia-Linfoma Linfoblástico de Células T Precursoras/imunologia , Leucemia-Linfoma Linfoblástico de Células T Precursoras/metabolismo , Linfócitos T/imunologia , Linfócitos T/metabolismo , Microambiente Tumoral , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Glyco-immune checkpoint receptors, molecules that inhibit immune cell activity following binding to glycosylated cell-surface antigens, are emerging as attractive targets for cancer immunotherapy. Defining biologically relevant ligands that bind and activate such receptors, however, has historically been a significant challenge. Here, we present a CRISPRi genomic screening strategy that allowed unbiased identification of the key genes required for cell-surface presentation of glycan ligands on leukemia cells that bind the glyco-immune checkpoint receptors Siglec-7 and Siglec-9. This approach revealed a selective interaction between Siglec-7 and the mucin-type glycoprotein CD43. Further work identified a specific N-terminal glycopeptide region of CD43 containing clusters of disialylated O-glycan tetrasaccharides that form specific Siglec-7 binding motifs. Knockout or blockade of CD43 in leukemia cells relieves Siglec-7-mediated inhibition of immune killing activity. This work identifies a potential target for immune checkpoint blockade therapy and represents a generalizable approach to dissection of glycan-receptor interactions in living cells.
Assuntos
Antígenos de Diferenciação Mielomonocítica/metabolismo , Repetições Palindrômicas Curtas Agrupadas e Regularmente Espaçadas/genética , Genoma Humano , Lectinas/metabolismo , Polissacarídeos/metabolismo , Motivos de Aminoácidos , Antígenos de Diferenciação Mielomonocítica/química , Linhagem Celular Tumoral , Membrana Celular/metabolismo , Glicopeptídeos/metabolismo , Humanos , Sinapses Imunológicas/metabolismo , Células Matadoras Naturais/metabolismo , Lectinas/química , Leucossialina/química , Leucossialina/metabolismo , Ligantes , Ligação ProteicaRESUMO
Objectives: CD43 can be useful in routine flow cytometry. We conducted a systematic review aiming to describe when CD43 is used by flow cytometry in malignant hematology and to determine its value in these settings. Methods: Systematic review of MEDLINE (search 'CD43' AND 'flow cytometry,' starting in 2010). Results: Twenty-one of 103 entries retrieved were included in this systematic review. CD43 is used in three settings: 1) in the classification of mature B cell lymphoproliferative disorders, 2) as part of a strategy to quantify residual disease in chronic lymphocytic leukemia (CLL) and 3) to help classify CD10-positive B cell populations. In this section, the published data is summarized, the clinical usefulness in each of these settings is evaluated and illustrative cases are shown. Conclusion: CD43 has a growing role in the diagnosis and management of B cell malignancies; it has become essential for the classification of B cell lymphoproliferative disorders and may be of help in the differential diagnosis of CD10-positive lymphomas by FC. It is also required for optimal quantification of CLL residual disease, which will soon be used to guide therapeutic decisions.
Assuntos
Neoplasias Hematológicas/diagnóstico , Leucemia Linfocítica Crônica de Células B/diagnóstico , Leucossialina/análise , Transtornos Linfoproliferativos/diagnóstico , Animais , Linfócitos B/patologia , Citometria de Fluxo/métodos , Humanos , Neoplasia Residual/diagnósticoRESUMO
Targeted cancer therapy with monoclonal antibodies has proven successful for different cancer types but is limited by the availability of suitable antibody targets. CD43s, a unique sialylated form of CD43 expressed by hematologic malignancies, is a recently identified target and antibodies interacting with CD43s may have therapeutic potential against acute myeloid leukemia (AML) and myelodysplastic syndrome. CD43s is recognized by the human antibody AT1413, that was derived from a high-risk AML patient who successfully cleared leukemia after allogeneic stem cell transplantation. Here we observed that AT1413 binds also to certain non-hematopoietic tumor cells, particularly melanoma and breast cancer. AT1413 immune precipitated CD43s from melanoma cells confirming that it recognizes the same target on melanoma as on AML. AT1413 induced antibody-dependent cellular cytotoxicity against short-term cultured patient-derived melanoma samples. However, AT1413 was unable to affect the growth of melanoma cells in vivo. To increase the efficacy of AT1413 as a therapeutic antibody, we generated two different formats of bispecific T-cell engaging antibodies (TCEs): one binding bivalently (bTCE) and the other monovalently (knob-in-hole; KiH) to both CD43s and CD3ε. In vitro, these TCEs redirected T-cell cytotoxicity against melanoma cells with differences in potencies. To investigate their effects in vivo, we grafted mice that harbor a human immune system with the melanoma cell line A375. Treatment with both AT1413 bTCE and AT1413 KiH significantly reduced tumor outgrowth in these mice. These data indicate a broad therapeutic potential of AT1413 that includes AML and CD43s-expressing solid tumors that originate from CD43-negative tissues.
Assuntos
Anticorpos Biespecíficos/farmacologia , Anticorpos Monoclonais/farmacologia , Antineoplásicos Imunológicos/farmacologia , Complexo CD3/imunologia , Leucossialina/imunologia , Melanoma/terapia , Ácido N-Acetilneuramínico/química , Linfócitos T/imunologia , Animais , Apoptose , Proliferação de Células , Citotoxicidade Imunológica , Feminino , Humanos , Técnicas In Vitro , Melanoma/imunologia , Melanoma/patologia , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID , Células Tumorais Cultivadas , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
We examined whether active fruit naringin can reduce the risk of BPA-mediated neurotoxicity in L-NAME induced hypertensive rats and whether the modulation could be linked to improvement of brain NO signaling. Male albino rats were randomly distributed into eight (n = 7) groups. Group I was control animals, Group II was orally-treated with L-NAME, Group III was orally treated with 100 mg/kg BPA, Group IV was orally-treated with L-NAME +100 mg/kg BPA. Group V was orally-administered with L-NAME +80 mg/kg NAR. Group VI was orally-administered with 100 mg/kg BPA +80 mg/kg NAR. Group VII was orally-administered with L-NAME+100 mg/kg BPA +80 mg/kg NAR. Lastly, group VIII was orally-treated with 80 mg/kg NAR. The treatment lasted for 14 days. Sub-acute exposure to L-NAME and BPA induced hypertension and mediated-neuroinflammation at CA-2 and CA-4 of hippocampus cells. It was evident by increase in PDE-51 and enzymes of ATP hydrolysis (ATPase, ADPase and AMPase) with corresponding upsurge in cholinergic (AChE and BuChE), dopaminergic (MAO-A) and adenosinergic (ADA) enzymes as well as movement disorder. The hypertensive-mediated neurotoxicity was related to alteration of NO signaling and higher release of pro-inflammatory cytokines (TNF-α and IL-1ß), apoptotic proteins (P53 and caspace-9) and facilitated entry of T-lymphocytes (CD43+) into CNS through blood brain barrier potentiated by antigen presenting cells. Hence, these features of BPA-mediated neurotoxicity in L-NAME induced hypertensive rats were prohibited by co-administration of NAR through production of neuro-inflammatory mediators, stabilizing neurotransmitter enzymes, normalizing NO signaling and improving brain histology.
Assuntos
Flavanonas/farmacologia , Hipocampo/efeitos dos fármacos , Hipertensão/complicações , Mediadores da Inflamação/metabolismo , Neurônios/efeitos dos fármacos , Fármacos Neuroprotetores/farmacologia , Síndromes Neurotóxicas/prevenção & controle , Nucleotídeos/deficiência , Estresse Oxidativo/efeitos dos fármacos , Animais , Anti-Inflamatórios/farmacologia , Células Apresentadoras de Antígenos/efeitos dos fármacos , Células Apresentadoras de Antígenos/imunologia , Células Apresentadoras de Antígenos/metabolismo , Antioxidantes/farmacologia , Apoptose/efeitos dos fármacos , Compostos Benzidrílicos , Modelos Animais de Doenças , Hipocampo/metabolismo , Hipocampo/patologia , Hipertensão/metabolismo , Hipertensão/fisiopatologia , Leucossialina/metabolismo , Locomoção/efeitos dos fármacos , Masculino , Neurônios/imunologia , Neurônios/metabolismo , Neurônios/patologia , Síndromes Neurotóxicas/imunologia , Síndromes Neurotóxicas/metabolismo , Síndromes Neurotóxicas/patologia , Óxido Nítrico/metabolismo , Fenóis , Ratos Wistar , Linfócitos T/efeitos dos fármacos , Linfócitos T/imunologia , Linfócitos T/metabolismoRESUMO
BACKGROUND: The Matutes score (MS) was proposed to differentiate chronic lymphocytic leukemia (CLL) from other B-cell non-Hodgkin lymphomas (B-NHLs). However, ambiguous immunophenotypes are common and remain a diagnostic challenge. Therefore, we evaluated the diagnostic benefit of measuring CD200 and CD43 expression together with the standard MS antigens. METHODS: 138 lymphoma patient samples and a validation cohort of 138 additive samples were classified according to the standard MS and further assigned with one or two additional points, for high CD200 and/or CD43 expression levels. The "classical" MS and the "Matutes score-extended" (MS-e) were categorized as high (4-5/6-7), intermediate (2-3/4-5), and low (0-1/0-3). Samples were reclassified into the MS-e with focus on ambiguous cases with an intermediate "classical" MS. RESULTS: A total of 35 of 138 (25.4%) patient samples were assigned to the intermediate MS group and confirmed by histopathological reports as CLL (14/40.0%) and B-NHLs other than CLL (21/60%). MS-e analysis identified 13 of 14 (92.9%) of CLL cases (MS-e 4-5) and 18/21 (85.7%) non-CLL cases (MS-e ≤ 3) correctly. Overall, the sensitivity of the CLL diagnosis was significantly increased by application of MS-e compared to the "classical" MS (98.8% vs. 82.7%; p = 0.0009), while specificity of both methods was almost equal (94.7% vs. 98.3%; p = 0.4795). Of note, sole measurement of CD43 and CD200 on B-cells sufficiently differentiated CLL from non-CLL with a test accuracy superior to the "classical" MS (F1 score 96.2 vs. 93.6). CONCLUSION: CD200 and CD43 have a high informative value in diagnostic immunophenotyping and facilitate the separation of CLL from other B-NHLs particularly in ambiguous cases.
Assuntos
Antígenos CD/imunologia , Leucemia Linfocítica Crônica de Células B/diagnóstico , Leucossialina/imunologia , Linfoma de Células B/diagnóstico , Antígenos CD/isolamento & purificação , Linfócitos B/imunologia , Linfócitos B/patologia , Biomarcadores Tumorais/imunologia , Diferenciação Celular/genética , Diferenciação Celular/imunologia , Diagnóstico Diferencial , Feminino , Regulação da Expressão Gênica , Humanos , Imunofenotipagem/métodos , Leucemia Linfocítica Crônica de Células B/imunologia , Leucemia Linfocítica Crônica de Células B/patologia , Leucossialina/isolamento & purificação , Linfoma de Células B/imunologia , Linfoma de Células B/patologia , MasculinoRESUMO
BACKGROUND: Sphingosine-1-phosphate (S1P) is a bioactive metabolite of sphingolipids and produced by sphingosine kinases (SphK1 and SphK2). SphK1/S1P pathway is implicated in the progression of chronic kidney disease. However, the role of SphK1/S1P pathway in renal injury in hypertension has not been reported. This study tested the hypothesis that SphK1/S1P pathway mediates the kidney damage in DOCA-salt hypertensive mice. METHODS: Male wild type (WT) C57BL6 and SphK1 knockout (KO) mice were subjected to unilateral nephrectomy, subcutaneous implant containing 50 mg of deoxycorticosterone acetate (DOCA) and 1% NaCl drinking water for 7 weeks. At the end of experiments, blood pressure data, 24 h urine and kidney samples were collected. Renal mRNA levels of SphK1 were measured by real-time RT-PCR. Markers for fibrogenesis and immune cell infiltration in kidneys were detected using Western blot and immunohistochemistray analysis, respectively. The glomerular morphological changes were examined in kidney tissue slides stained with Periodic-Acid Schiff. Four groups were studied: wild type control (WT-C), WT-DOCA, KO-C and KO-DOCA. RESULTS: The renal SphK1 mRNA expression was significantly upregulated in WT-DOCA mice, whereas this upregulation of renal SphK1 mRNA was blocked in KO-DOCA mice. There was no difference in DOCA-salt-induced hypertension between WT and KO mice. The urinary albumin was increased in both DOCA-salt groups. However, the albuminuria was significantly lower in KO-DOCA than in WT-DOCA group. There were increases in glomerulosclerosis indices in both DOCA-salt groups, whereas the increases were also significantly lower in KO-DOCA than in WT-DOCA mice. Renal protein levels of α-smooth muscle actin were upregulated in both DOCA-salt groups, but the increase was significant lower in KO-DOCA than in WT-DOCA group. The increased staining areas of collagen detected by Sirius Red-staining in kidney tissue sections were also attenuated in KO-DOCA compared with WT-DOCA mice. In contrast, the increased infiltration of CD43+ (a T cell marker) or CD68+ (a macrophage marker) cells in DOCA-salt kidneys showed no significant difference between WT-DOCA and KO-DOCA mice. CONCLUSIONS: SphK1/S1P signaling pathway mediates kidney damage in DOCA-salt hypertensive mice independent of blood pressure and immune modulation.
Assuntos
Hipertensão/genética , Rim/metabolismo , Lisofosfolipídeos/metabolismo , Fosfotransferases (Aceptor do Grupo Álcool)/genética , RNA Mensageiro/metabolismo , Insuficiência Renal Crônica/genética , Esfingosina/análogos & derivados , Actinas/metabolismo , Albuminúria/metabolismo , Animais , Antígenos CD/metabolismo , Antígenos de Diferenciação Mielomonocítica/metabolismo , Western Blotting , Colágeno/metabolismo , Acetato de Desoxicorticosterona/toxicidade , Modelos Animais de Doenças , Fibrose , Hipertensão/induzido quimicamente , Hipertensão/metabolismo , Hipertensão/patologia , Imuno-Histoquímica , Rim/patologia , Leucossialina/metabolismo , Macrófagos/metabolismo , Masculino , Camundongos , Camundongos Knockout , Mineralocorticoides/toxicidade , Nefrectomia , Insuficiência Renal Crônica/metabolismo , Insuficiência Renal Crônica/patologia , Transdução de Sinais , Cloreto de Sódio na Dieta/toxicidade , Esfingosina/metabolismo , Linfócitos T/metabolismoRESUMO
HIV-1 particles incorporate various host transmembrane proteins in addition to viral Env glycoprotein during assembly at the plasma membrane. In polarized T cells, HIV-1 structural protein Gag localizes to the plasma membrane of uropod, a rear-end protrusion. Notably, uropod transmembrane proteins PSGL-1 and CD43 cocluster specifically with Gag assembling at the plasma membrane even in cells that do not form uropods. Recent reports have shown that expression of either PSGL-1 or CD43 in virus-producing cells reduces the infectivity of progeny virions and that HIV-1 infection reduces the cell surface expression of these proteins. However, the mechanisms for both processes remain to be determined. In this study, we found that virion incorporation of PSGL-1 and CD43 closely correlates with diminished virion infectivity. PSGL-1 and CD43 inhibited virus attachment to CD4+ cells irrespective of the presence of Env. These proteins also inhibited virion attachment to CD4- lymphoid organ fibroblastic reticular cells that mediate transinfection of CD4+ T cells. Consistent with the possibility that highly extended extracellular domains of these proteins physically block virus-cell attachment, the inhibitory effect of PSGL-1 required its full-length ectodomain. HIV-1 encoding Gag mutants that are defective in either coclustering with these host proteins or ESCRT-dependent particle release failed to reduce PSGL-1 on surface of infected cells. This study reveals an anti-HIV-1 mechanism that suppresses virus-cell attachment and a previously unappreciated process of HIV-1-mediated down-regulation of host antiviral proteins, both of which likely require virion incorporation of these proteins.
Assuntos
Infecções por HIV/imunologia , HIV-1/imunologia , Interações Hospedeiro-Patógeno/genética , Leucossialina/genética , Glicoproteínas de Membrana/genética , Vírion/genética , Produtos do Gene gag do Vírus da Imunodeficiência Humana/metabolismo , Buffy Coat/citologia , Regulação para Baixo , Técnicas de Inativação de Genes , Células HEK293 , Infecções por HIV/virologia , HIV-1/genética , HIV-1/metabolismo , Células HeLa , Voluntários Saudáveis , Interações Hospedeiro-Patógeno/imunologia , Humanos , Mutação , Domínios Proteicos/genética , Linfócitos T/imunologia , Montagem de Vírus/genética , Montagem de Vírus/imunologia , Ligação Viral , Replicação Viral/genética , Produtos do Gene gag do Vírus da Imunodeficiência Humana/genética , Produtos do Gene gag do Vírus da Imunodeficiência Humana/imunologiaRESUMO
Pediatric patients receiving solid organ transplants may develop lymphoproliferative diseases, including graft-versus-host disease (GvHD) and posttransplant lymphoproliferative diseases (PTLDs). We characterized lesions in 11 clinically ill NOD.Cg-Prkdcscid Il2rgtm1Wjl/SzJ (NSG) mice that received pediatric-patient-derived solid tumors (PDXs) and developed immunodeficiency-associated lymphoproliferations comparable to GvHD and PTLDs over a period of 46 to 283 days after implantation. Lymphoproliferations were diffusely positive for human-specific biomarkers, including NUMA1, CD45, and CD43, but lacked immunoreactivity for murine CD45. Human immune cells were CD3-positive, with subsets having immunoreactivity for CD4 and CD8 as well as PAX5, CD79a, and IRF4, resulting from populations of human T and B cells present within the xenotransplants. Tissues and organs infiltrated included mucocutaneous zones (oral cavity and perigenital and perianal regions), haired skin, tongue, esophagus, forestomach, thyroid, salivary glands, lungs, liver, kidneys, spleen, lymph nodes, bone marrow, and brain. In 4 of 5 mice with PTLD, Epstein-Barr virus (EBV)-encoded small RNAs (EBERs) were detected by in situ hybridization in PAX5+ human B cells associated with the PDX (n = 1/4) or with engrafted human immune cells at other anatomic locations (n = 4/11). One of the 4 mice had an EBV-associated human large B-cell lymphoma. NSG mice receiving xenotransplants can develop combinations of GvHD, EBV-driven PTLD, and B-cell lymphoma similar to those occurring in human pediatric patients. Therefore, pediatric xenotransplants should undergo histopathologic and immunohistochemical assessment upon collection to ensure that the specimen is not a lymphoma and does not contain lymphoma cells because these neoplasms can morphologically mimic small round blue cell pediatric solid tumors.