RESUMO
Lymphatic vessels form a critical component in the regulation of human health and disease. While their functional significance is increasingly being recognized, the comprehensive heterogeneity of lymphatics remains uncharacterized. Here, we report the profiling of 33,000 lymphatic endothelial cells (LECs) in human lymph nodes (LNs) by single-cell RNA sequencing. Unbiased clustering revealed six major types of human LECs. LECs lining the subcapsular sinus (SCS) of LNs abundantly expressed neutrophil chemoattractants, whereas LECs lining the medullary sinus (MS) expressed a C-type lectin CD209. Binding of a carbohydrate Lewis X (CD15) to CD209 mediated neutrophil binding to the MS. The neutrophil-selective homing by MS LECs may retain neutrophils in the LN medulla and allow lymph-borne pathogens to clear, preventing their spread through LNs in humans. Our study provides a comprehensive characterization of LEC heterogeneity and unveils a previously undefined role for medullary LECs in human immunity.
Assuntos
Células Endoteliais/imunologia , Neutrófilos/imunologia , Animais , Moléculas de Adesão Celular/imunologia , Células Cultivadas , Humanos , Lectinas Tipo C/imunologia , Antígenos CD15/imunologia , Linfonodos/imunologia , Vasos Linfáticos/imunologia , Camundongos Endogâmicos C57BL , Receptores de Superfície Celular/imunologia , Inquéritos e QuestionáriosRESUMO
Lymphocyte recruitment maintains intestinal immune homeostasis but also contributes to inflammation. The orphan chemoattractant receptor GPR15 mediates regulatory T cell homing and immunosuppression in the mouse colon. We show that GPR15 is also expressed by mouse TH17 and TH1 effector cells and is required for colitis in a model that depends on the trafficking of these cells to the colon. In humans GPR15 is expressed by effector cells, including pathogenic TH2 cells in ulcerative colitis, but is expressed poorly or not at all by colon regulatory T (Treg) cells. The TH2 transcriptional activator GATA-3 and the Treg-associated transcriptional repressor FOXP3 robustly bind human, but not mouse, GPR15 enhancer sequences, correlating with receptor expression. Our results highlight species differences in GPR15 regulation and suggest it as a potential therapeutic target for colitis.
Assuntos
Colite/fisiopatologia , Colo/fisiopatologia , Regulação da Expressão Gênica , Receptores Acoplados a Proteínas G/metabolismo , Receptores de Retorno de Linfócitos/metabolismo , Receptores de Peptídeos/metabolismo , Animais , Células Cultivadas , Colite/imunologia , Colo/imunologia , Modelos Animais de Doenças , Elementos Facilitadores Genéticos/genética , Fatores de Transcrição Forkhead/metabolismo , Técnicas de Inativação de Genes , Humanos , Camundongos , Ligação Proteica , Receptores Acoplados a Proteínas G/genética , Receptores de Peptídeos/genética , Especificidade da EspécieRESUMO
Lymphocytes are recruited from blood by high-endothelial venules (HEVs). We performed transcriptomic analyses and identified molecular signatures that distinguish HEVs from capillary endothelium and that define tissue-specific HEV specialization. Capillaries expressed gene programs for vascular development. HEV-expressed genes showed enrichment for genes encoding molecules involved in immunological defense and lymphocyte migration. We identify capillary and HEV markers and candidate mechanisms for regulated recruitment of lymphocytes, including a lymph node HEV-selective transmembrane mucin; transcriptional control of functionally specialized carbohydrate ligands for lymphocyte L-selectin; HEV expression of molecules for transendothelial migration; and metabolic programs for lipid mediators of lymphocyte motility and chemotaxis. We also elucidate a carbohydrate-recognition pathway that targets B cells to intestinal lymphoid tissues, defining CD22 as a lectin-homing receptor for mucosal HEVs.
Assuntos
Capilares/metabolismo , Endotélio/metabolismo , Perfilação da Expressão Gênica , Linfócitos/metabolismo , Tecido Linfoide/irrigação sanguínea , Vênulas/metabolismo , Animais , Movimento Celular/genética , Células Endoteliais/metabolismo , Endotélio/citologia , Feminino , Citometria de Fluxo , Ontologia Genética , Linfonodos/irrigação sanguínea , Masculino , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos Transgênicos , Microscopia Confocal , Análise de Sequência com Séries de OligonucleotídeosRESUMO
BACKGROUND: Lymphedema is a global health problem with no effective drug treatment. Enhanced T-cell immunity and abnormal lymphatic endothelial cell (LEC) signaling are promising therapeutic targets for this condition. Sphingosine-1-phosphate (S1P) mediates a key signaling pathway required for normal LEC function, and altered S1P signaling in LECs could lead to lymphatic disease and pathogenic T-cell activation. Characterizing this biology is relevant for developing much needed therapies. METHODS: Human and mouse lymphedema was studied. Lymphedema was induced in mice by surgically ligating the tail lymphatics. Lymphedematous dermal tissue was assessed for S1P signaling. To verify the role of altered S1P signaling effects in lymphatic cells, LEC-specific S1pr1-deficient (S1pr1LECKO) mice were generated. Disease progression was quantified by tail-volumetric and -histopathologic measurements over time. LECs from mice and humans, with S1P signaling inhibition, were then cocultured with CD4 T cells, followed by an analysis of CD4 T-cell activation and pathway signaling. Last, animals were treated with a monoclonal antibody specific to P-selectin to assess its efficacy in reducing lymphedema and T-cell activation. RESULTS: Human and experimental lymphedema tissues exhibited decreased LEC S1P signaling through S1P receptor 1 (S1PR1). LEC S1pr1 loss-of-function exacerbated lymphatic vascular insufficiency, tail swelling, and increased CD4 T-cell infiltration in mouse lymphedema. LECs, isolated from S1pr1LECKO mice and cocultured with CD4 T cells, resulted in augmented lymphocyte differentiation. Inhibiting S1PR1 signaling in human dermal LECs promoted T-helper type 1 and 2 (Th1 and Th2) cell differentiation through direct cell contact with lymphocytes. Human dermal LECs with dampened S1P signaling exhibited enhanced P-selectin, an important cell adhesion molecule expressed on activated vascular cells. In vitro, P-selectin blockade reduced the activation and differentiation of Th cells cocultured with shS1PR1-treated human dermal LECs. P-selectin-directed antibody treatment improved tail swelling and reduced Th1/Th2 immune responses in mouse lymphedema. CONCLUSIONS: This study suggests that reduction of the LEC S1P signaling aggravates lymphedema by enhancing LEC adhesion and amplifying pathogenic CD4 T-cell responses. P-selectin inhibitors are suggested as a possible treatment for this pervasive condition.
Assuntos
Linfedema , Selectina-P , Humanos , Camundongos , Animais , Transdução de Sinais , Inflamação/patologia , Linfedema/patologiaRESUMO
Specialized stromal cells occupy and help define B- and T-cell domains, which are crucial for proper functioning of our immune system. Signaling through lymphotoxin and TNF receptors is crucial for the development of different stromal subsets, which are thought to arise from a common precursor. However, mechanisms that control the selective generation of the different stromal phenotypes are not known. Using in vitro cultures of embryonic mouse stromal cells, we show that retinoic acid-mediated signaling is important for the differentiation of precursors towards the Cxcl13pos follicular dendritic cell (FDC) lineage, and also blocks lymphotoxin-mediated Ccl19pos fibroblastic reticular cell lineage differentiation. Accordingly, at the day of birth we observe the presence of Cxcl13posCcl19neg/low and Cxcl13neg/lowCcl19pos cells within neonatal lymph nodes. Furthermore, ablation of retinoic acid receptor signaling in stromal precursors early after birth reduces Cxcl13 expression, and complete blockade of retinoic acid signaling prevents the formation of FDC networks in lymph nodes.
Assuntos
Células Dendríticas Foliculares/metabolismo , Células Dendríticas Foliculares/fisiologia , Linfonodos/metabolismo , Linfonodos/fisiologia , Transdução de Sinais/fisiologia , Tretinoína/metabolismo , Animais , Diferenciação Celular/fisiologia , Linhagem da Célula/fisiologia , Camundongos , Camundongos Endogâmicos C57BL , Células Estromais/metabolismo , Células Estromais/fisiologiaRESUMO
BACKGROUND & AIMS: Intestinal microfold (M) cells are a unique subset of intestinal epithelial cells in the Peyer's patches that regulate mucosal immunity, serving as portals for sampling and uptake of luminal antigens. The inability to efficiently develop human M cells in cell culture has impeded studies of the intestinal immune system. We aimed to identify signaling pathways required for differentiation of human M cells and establish a robust culture system using human ileum enteroids. METHODS: We analyzed transcriptome data from mouse Peyer's patches to identify cell populations in close proximity to M cells. We used the human enteroid system to determine which cytokines were required to induce M-cell differentiation. We performed transcriptome, immunofluorescence, scanning electron microscope, and transcytosis experiments to validate the development of phenotypic and functional human M cells. RESULTS: A combination of retinoic acid and lymphotoxin induced differentiation of glycoprotein 2-positive human M cells, which lack apical microvilli structure. Upregulated expression of innate immune-related genes within M cells correlated with a lack of viral antigens after rotavirus infection. Human M cells, developed in the enteroid system, internalized and transported enteric viruses, such as rotavirus and reovirus, across the intestinal epithelium barrier in the enteroids. CONCLUSIONS: We identified signaling pathways required for differentiation of intestinal M cells, and used this information to create a robust culture method to develop human M cells with capacity for internalization and transport of viruses. Studies of this model might increase our understanding of antigen presentation and the systemic entry of enteric pathogens in the human intestine.
Assuntos
Diferenciação Celular/imunologia , Linfotoxina-alfa/metabolismo , Nódulos Linfáticos Agregados/imunologia , Transdução de Sinais/imunologia , Tretinoína/metabolismo , Animais , Apresentação de Antígeno/imunologia , Técnicas de Cultura de Células/métodos , Células Epiteliais/imunologia , Células Epiteliais/metabolismo , Humanos , Íleo/citologia , Íleo/imunologia , Imunidade nas Mucosas , Mucosa Intestinal/citologia , Mucosa Intestinal/imunologia , Camundongos , NF-kappa B/metabolismo , Organoides , Nódulos Linfáticos Agregados/citologia , Nódulos Linfáticos Agregados/metabolismo , Cultura Primária de Células , Proteínas Recombinantes/metabolismoRESUMO
Dendritic cells (DCs) are 'professional' antigen-presenting cells that are key in the regulation of immune responses. Here we characterize a unique subset of tolerogenic DCs that expressed the chemokine receptor CCR9 and migrated to the CCR9 ligand CCL25, a chemokine linked to the homing of T cells and DCs to the gut. CCR9(+) DCs were of the plasmacytoid DC (pDC) lineage, had an immature phenotype and rapidly downregulated CCR9 in response to maturation-inducing pDC-restricted Toll-like receptor ligands. CCR9(+) pDCs were potent inducers of regulatory T cell function and suppressed antigen-specific immune responses both in vitro and in vivo, including inhibiting acute graft-versus-host disease induced by allogeneic CD4(+) donor T cells in irradiated recipients. Our results identify a highly immunosuppressive population of pDCs present in lymphoid tissues.
Assuntos
Células Dendríticas/imunologia , Doença Enxerto-Hospedeiro/imunologia , Receptores CCR10/imunologia , Tolerância a Antígenos Próprios , Tolerância ao Transplante , Animais , Biomarcadores/metabolismo , Diferenciação Celular , Linhagem Celular Tumoral , Movimento Celular , Quimiocinas CC/metabolismo , Células Dendríticas/metabolismo , Regulação para Baixo , Intestinos/imunologia , Antígenos Comuns de Leucócito/metabolismo , Ligantes , Camundongos , Camundongos Congênicos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Receptores CCR10/metabolismo , Linfócitos T Reguladores/imunologia , Receptores Toll-Like/metabolismo , Receptor D6 de QuimiocinaRESUMO
UNLABELLED: Circulating factor VIII (FVIII) is derived from liver and from extrahepatic sources probably of endothelial origin, but the vascular sites of FVIII production remain unclear. Among organs profiled, only liver and lymph nodes (LNs) show abundant expression of F8 messenger RNA (mRNA). Transcriptomic profiling of subsets of stromal cells, including endothelial cells (ECs) from mouse LNs and other tissues, showed that F8 mRNA is expressed by lymphatic ECs (LECs) but not by capillary ECs (capECs), fibroblastic reticular cells, or hematopoietic cells. Among blood ECs profiled, F8 expression was seen only in fenestrated ECs (liver sinusoidal and renal glomerular ECs) and some high endothelial venules. In contrast, von Willebrand factor mRNA was expressed in capECs but not in LECs; it was coexpressed with F8 mRNA in postcapillary high endothelial venules. Purified LECs and liver sinusoidal ECs but not capECs from LNs secrete active FVIII in culture, and human and mouse lymph contained substantial FVIII: C activity. Our results revealed localized vascular expression of FVIII and von Willebrand factor and identified LECs as a major cellular source of FVIII in extrahepatic tissues.
Assuntos
Células Endoteliais/metabolismo , Endotélio Linfático/metabolismo , Endotélio Vascular/metabolismo , Fator VIII/biossíntese , Regulação da Expressão Gênica/fisiologia , Fator de von Willebrand/biossíntese , Animais , Capilares/citologia , Capilares/metabolismo , Células Endoteliais/citologia , Endotélio Linfático/citologia , Endotélio Vascular/citologia , Feminino , Humanos , Glomérulos Renais/irrigação sanguínea , Glomérulos Renais/citologia , Glomérulos Renais/metabolismo , Fígado/irrigação sanguínea , Fígado/citologia , Fígado/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Especificidade de Órgãos , Vênulas/citologia , Vênulas/metabolismoRESUMO
Dipeptidyl peptidase-4 (DPP-4) inhibitors are a well-known and novel class of oral antihyperglycaemic drugs. DPP-4 inhibition facilitates ulcer healing in patients with diabetes. However, the actual mechanisms, which are independent of lower blood glucose levels, are still unknown. Therefore, the aim of this study was to analyse the effect of the DPP-4 inhibitor sitagliptin on wound healing through a glucose-independent pathway. In this study, DPP-4 inhibitors facilitate keratinocyte differentiation and the proliferation, increase blood flow in the cutaneous of wounds in healthy C57BL/6 mice. Additionally, the administration of the DPP-4 inhibitor ameliorates wound healing and enhances adiponectin expression in healthy C57BL/6 mice. Taken together, our results reveal a protective role for the DPP-4 inhibitor sitagliptin in wound healing by regulating adiponectin and phospho-eNOS levels in keratinocytes. Based on these results, the DPP-4 inhibitor may have therapeutic potential for healing wounds through a diabetes-independent mechanism.
Assuntos
Adiponectina/metabolismo , Inibidores da Dipeptidil Peptidase IV/farmacologia , Reepitelização/efeitos dos fármacos , Fluxo Sanguíneo Regional/efeitos dos fármacos , Fosfato de Sitagliptina/farmacologia , Animais , Barreira Hematoencefálica/metabolismo , Diferenciação Celular/efeitos dos fármacos , Linhagem Celular , Proliferação de Células/efeitos dos fármacos , Dipeptidil Peptidase 4/sangue , Peptídeo 1 Semelhante ao Glucagon/sangue , Queratinócitos/fisiologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Óxido Nítrico Sintase Tipo III/metabolismo , Fosforilação , Pele/irrigação sanguínea , Pele/lesões , Ferimentos e Lesões/metabolismo , Ferimentos e Lesões/patologiaRESUMO
Homing of murine dendritic epidermal T cells (DETCs) from the thymus to the skin is regulated by specific trafficking receptors during late embryogenesis. Once in the epidermis, Vγ3δ1 TCR DETCs are maintained through self-renewal and participate in wound healing. GPR15 is an orphan G protein-linked chemoattractant receptor involved in the recruitment of regulatory T cells to the colon. Here we show that GPR15 is highly expressed on fetal thymic DETC precursors and on recently recruited DETCs, and mediates the earliest seeding of the epidermis, which occurs at the time of establishment of skin barrier function. DETCs in GPR15(-/-) mice remain low at birth, but later participation of CCR10 and CCR4 in DETC homing allows DETCs to reach near normal levels in adult skin. Our findings establish a role for GPR15 in skin lymphocyte homing and suggest that it may contribute to lymphocyte subset targeting to diverse epithelial sites.
Assuntos
Movimento Celular/imunologia , Células Dendríticas/imunologia , Receptores de Antígenos de Linfócitos T gama-delta/imunologia , Receptores Acoplados a Proteínas G/imunologia , Pele/imunologia , Linfócitos T Reguladores/imunologia , Animais , Movimento Celular/genética , Células Dendríticas/citologia , Camundongos , Camundongos Knockout , Receptores de Antígenos de Linfócitos T gama-delta/genética , Receptores CCR4/genética , Receptores CCR4/imunologia , Receptores Acoplados a Proteínas G/genética , Pele/citologia , Linfócitos T Reguladores/citologia , Timo/citologia , Timo/imunologiaRESUMO
Stromal cells within the tumor tissue promote immune evasion as a critical strategy for cancer development and progression, but the underlying mechanisms remain poorly understood. In this study, we explore the role of endothelial cells (ECs) in the regulation of the immunosuppressive tumor microenvironment. Using mouse pancreatic ductal adenocarcinoma (PDAC) models, we found that canonical Notch signaling in endothelial cells suppresses the recruitment of antitumor T cells and promotes tumor progression by inhibiting the pro-inflammatory functions of cancer-associated fibroblasts (CAFs). Abrogation of endothelial Notch signaling modulates EC-derived angiocrine factors to enhance the pro-inflammatory activities of CAFs, which drive CXCL9/10-CXCR3-mediated T cell recruitment to inhibit tumor growth. Additionally, abrogation of endothelial Notch unleashed interferon gamma responses in the tumor microenvironment, upregulated PDL1 expression on tumor cells, and sensitized PDAC to PD1-based immunotherapy. Collectively, these data uncover a pivotal role of endothelial cells in shaping the immunosuppressive microenvironment, and suggest the potential of targeting EC-CAF interaction as a novel therapeutic modality to boost antitumor immunity.
RESUMO
The estuarine plastisphere, a novel ecological habitat in the Anthropocene, has garnered global concerns. Recent geochemical evidence has pointed out its potential role in influencing nitrogen biogeochemistry. However, the biogeochemical significance of the plastisphere and its mechanisms regulating nitrogen cycling remain elusive. Using 15N- and 13C-labelling coupled with metagenomics and metatranscriptomics, here we unveil that the plastisphere likely acts as an underappreciated nitrifying niche in estuarine ecosystems, exhibiting a 0.9 ~ 12-fold higher activity of bacteria-mediated nitrification compared to surrounding seawater and other biofilms (stone, wood and glass biofilms). The shift of active nitrifiers from O2-sensitive nitrifiers in the seawater to nitrifiers with versatile metabolisms in the plastisphere, combined with the potential interspecific cooperation of nitrifying substrate exchange observed among the plastisphere nitrifiers, collectively results in the unique nitrifying niche. Our findings highlight the plastisphere as an emerging nitrifying niche in estuarine environment, and deepen the mechanistic understanding of its contribution to marine biogeochemistry.
Assuntos
Bactérias , Biofilmes , Estuários , Nitrificação , Água do Mar , Água do Mar/microbiologia , Bactérias/metabolismo , Bactérias/genética , Biofilmes/crescimento & desenvolvimento , Ecossistema , Microbiota/fisiologia , Metagenômica , Filogenia , Ciclo do Nitrogênio , Nitrogênio/metabolismo , Isótopos de Nitrogênio/metabolismoRESUMO
BACKGROUND: Lymphedema is a global health problem with no effective drug treatment. Enhanced T cell immunity and abnormal lymphatic endothelial cell (LEC) signaling are promising therapeutic targets for this condition. Sphingosine-1-phosphate (S1P) mediates a key signaling pathway required for normal LEC function, and altered S1P signaling in LECs could lead to lymphatic disease and pathogenic T cell activation. Characterizing this biology is relevant for developing much-needed therapies. METHODS: Human and mouse lymphedema was studied. Lymphedema was induced in mice by surgically ligating the tail lymphatics. Lymphedematous dermal tissue was assessed for S1P signaling. To verify the role of altered S1P signaling effects in lymphatic cells, LEC-specific S1pr1 -deficient ( S1pr1 LECKO ) mice were generated. Disease progression was quantified by tail-volumetric and -histopathological measurements over time. LECs from mice and humans, with S1P signaling inhibition, were then co-cultured with CD4 T cells, followed by an analysis of CD4 T cell activation and pathway signaling. Finally, animals were treated with a monoclonal antibody specific to P-selectin to assess its efficacy in reducing lymphedema and T cell activation. RESULTS: Human and experimental lymphedema tissues exhibited decreased LEC S1P signaling through S1PR1. LEC S1pr1 loss-of-function exacerbated lymphatic vascular insufficiency, tail swelling, and increased CD4 T cell infiltration in mouse lymphedema. LECs, isolated from S1pr1 LECKO mice and co-cultured with CD4 T cells, resulted in augmented lymphocyte differentiation. Inhibiting S1PR1 signaling in human dermal LECs (HDLECs) promoted T helper type 1 and 2 (Th1 and Th2) cell differentiation through direct cell contact with lymphocytes. HDLECs with dampened S1P signaling exhibited enhanced P-selectin, an important cell adhesion molecule expressed on activated vascular cells. In vitro , P-selectin blockade reduced the activation and differentiation of Th cells co-cultured with sh S1PR1 -treated HDLECs. P-selectin-directed antibody treatment improved tail swelling and reduced Th1/Th2 immune responses in mouse lymphedema. CONCLUSION: This study suggests that reduction of the LEC S1P signaling aggravates lymphedema by enhancing LEC adhesion and amplifying pathogenic CD4 T cell responses. P-selectin inhibitors are suggested as a possible treatment for this pervasive condition. Clinical Perspective: What is New?: Lymphatic-specific S1pr1 deletion exacerbates lymphatic vessel malfunction and Th1/Th2 immune responses during lymphedema pathogenesis. S1pr1 -deficient LECs directly induce Th1/Th2 cell differentiation and decrease anti-inflammatory Treg populations. Peripheral dermal LECs affect CD4 T cell immune responses through direct cell contact.LEC P-selectin, regulated by S1PR1 signaling, affects CD4 T cell activation and differentiation.P-selectin blockade improves lymphedema tail swelling and decreases Th1/Th2 population in the diseased skin.What Are the Clinical Implications?: S1P/S1PR1 signaling in LECs regulates inflammation in lymphedema tissue.S1PR1 expression levels on LECs may be a useful biomarker for assessing predisposition to lymphatic disease, such as at-risk women undergoing mastectomyP-selectin Inhibitors may be effective for certain forms of lymphedema.
RESUMO
The hematopoietic niche is a supportive microenvironment composed of distinct cell types, including specialized vascular endothelial cells that directly interact with hematopoietic stem and progenitor cells (HSPCs). The molecular factors that specify niche endothelial cells and orchestrate HSPC homeostasis remain largely unknown. Using multi-dimensional gene expression and chromatin accessibility analyses in zebrafish, we define a conserved gene expression signature and cis-regulatory landscape that are unique to sinusoidal endothelial cells in the HSPC niche. Using enhancer mutagenesis and transcription factor overexpression, we elucidate a transcriptional code that involves members of the Ets, Sox, and nuclear hormone receptor families and is sufficient to induce ectopic niche endothelial cells that associate with mesenchymal stromal cells and support the recruitment, maintenance, and division of HSPCs in vivo. These studies set forth an approach for generating synthetic HSPC niches, in vitro or in vivo, and for effective therapies to modulate the endogenous niche.
Assuntos
Nicho de Células-Tronco , Fatores de Transcrição , Animais , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Células Endoteliais/metabolismo , Peixe-Zebra/genética , Peixe-Zebra/metabolismo , Regulação da Expressão GênicaRESUMO
"Plastisphere", microbial communities colonizing plastic debris, has sparked global concern for marine ecosystems. Microbiome inhabiting this novel human-made niche has been increasingly characterized; however, whether the plastisphere holds crucial roles in biogeochemical cycling remains largely unknown. Here we evaluate the potential of plastisphere in biotic and abiotic denitrification and nitrous oxide (N2O) production in estuaries. Biofilm formation provides anoxic conditions favoring denitrifiers. Comparing with surrounding bulk water, plastisphere exhibits a higher denitrifying activity and N2O production, suggesting an overlooked N2O source. Regardless of plastisphere and bulk water, bacterial and fungal denitrifications are the main regulators for N2O production instead of chemodenitrification. However, the contributions of bacteria and fungi in the plastisphere are different from those in bulk water, indicating a distinct N2O production pattern in the plastisphere. These findings pinpoint plastisphere as a N2O source, and provide insights into roles of the new biotope in biogeochemical cycling in the Anthropocene.
Assuntos
Microbiota , Plásticos , Bactérias , Humanos , Óxido Nitroso , ÁguaRESUMO
Immunoglobulin family and carbohydrate vascular addressins encoded by Madcam1 and St6gal1 control lymphocyte homing into intestinal tissues, regulating immunity and inflammation. The addressins are developmentally programmed to decorate endothelial cells lining gut post-capillary and high endothelial venules (HEV), providing a prototypical example of organ- and segment-specific endothelial specialization. We identify conserved NKX-COUP-TFII composite elements (NCCE) in regulatory regions of Madcam1 and St6gal1 that bind intestinal homeodomain protein NKX2-3 cooperatively with venous nuclear receptor COUP-TFII to activate transcription. The Madcam1 element also integrates repressive signals from arterial/capillary Notch effectors. Pan-endothelial COUP-TFII overexpression induces ectopic addressin expression in NKX2-3+ capillaries, while NKX2-3 deficiency abrogates expression by HEV. Phylogenetically conserved NCCE are enriched in genes involved in neuron migration and morphogenesis of the heart, kidney, pancreas and other organs. Our results define an NKX-COUP-TFII morphogenetic code that targets expression of mucosal vascular addressins.
Assuntos
Células Endoteliais , Veias , Morfogênese/genética , Artérias , Movimento CelularRESUMO
Recombinant FVIII formulated in PEG-ylated liposomes (rFVIII-PEG-Lip) was reported to increase the bleed-free days from 7 to 13 days (at 35 IU/kg rFVIII) in severe hemophilia A patients. To understand the underlying mechanism, we sought to recapitulate its efficacy in hemophilia A mice. Animals treated with rFVIII-PEG-Lip achieved approximately 30% higher survival relative to rFVIII after tail vein transection inflicted 24 hours after dosing. The efficacy of rFVIII-PEG-Lip represents an approximately 2.5-fold higher "apparent" FVIII activity, which is not accounted for by its modestly increased (13%) half-life. The enhanced efficacy requires complex formation between rFVIII and PEG-Lip before the administration. Furthermore, PEG-Lip associates with the majority of platelets and monocytes in vivo, and results in increased P-selectin surface expression on platelets in response to collagen. Rotational thromboelastometry (ROTEM) analysis of whole blood from rFVIII-PEG-Lip-treated animals at 5 minutes up to 72 hours after dosing recapitulated the 2- to 3-fold higher apparent FVIII activity. The enhanced procoagulant activity is fully retained in plasma unless microparticles are removed by ultracentrifugation. Taken together, the efficacy of rFVIII-PEG-Lip is mediated mainly by its sensitization of platelets and the generation of procoagulant microparticles that may express sustained high-affinity receptors for FVIII.
Assuntos
Fator VIII/administração & dosagem , Hemofilia A/tratamento farmacológico , Polietilenoglicóis/administração & dosagem , Animais , Modelos Animais de Doenças , Avaliação Pré-Clínica de Medicamentos , Fator VIII/metabolismo , Meia-Vida , Hemofilia A/mortalidade , Hemofilia A/patologia , Lipossomos , Substâncias Macromoleculares/administração & dosagem , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Polietilenoglicóis/química , Polietilenoglicóis/metabolismo , Ligação Proteica , Proteínas Recombinantes/administração & dosagem , Proteínas Recombinantes/metabolismo , Especificidade por Substrato , Análise de Sobrevida , Resultado do TratamentoRESUMO
GPR15 is a chemoattractant receptor that facilitates colon homing of regulatory and effector CD4+ T cells in health and colitis. The molecular mechanisms that control GPR15 expression are not fully known. Here we report the presence of two highly conserved aryl hydrocarbon receptor (AHR) binding sequences in a 3' enhancer of GPR15, leading us to investigate AHR function in regulating GPR15 expression. Using luciferase reporter assays, we show that AHR activation increased GPR15 expression and requires both the AHR binding sites. Consistent with a transcriptional regulatory role, treatment with AHR agonists induce GPR15 expression on human CD4+ T cells. Using AHR-deficient mice, we demonstrate that the lack of AHR signaling drastically reduces GPR15 expression on effector/memory and Foxp3+ CD4+ T cells. In mixed bone marrow chimeras of AHR-deficient and wildtype cells, GPR15 expression was similarly diminished on AHR-deficient CD4+ effector/memory and regulatory T cells in the colon and small intestine. Furthermore, administration of AHR agonists upregulated GPR15 expression on CD4+ effector/memory T cells and increased their homing capability, especially to the colon. Collectively, our studies reveal a novel function of the AHR in regulation of GPR15 expression and increased colon trafficking of CD4+ T cells expressing GPR15.
Assuntos
Fatores de Transcrição Hélice-Alça-Hélice Básicos , Regulação da Expressão Gênica , Receptores de Hidrocarboneto Arílico , Receptores Acoplados a Proteínas G , Receptores de Peptídeos , Humanos , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Sítios de Ligação , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD4-Positivos/metabolismo , Fatores de Transcrição Forkhead , Fator de Transcrição GATA3/metabolismo , Regiões Promotoras Genéticas , Ligação Proteica , Receptores de Hidrocarboneto Arílico/metabolismo , Receptores Acoplados a Proteínas G/genética , Receptores Acoplados a Proteínas G/metabolismo , Receptores de Peptídeos/genética , Receptores de Peptídeos/metabolismo , Subpopulações de Linfócitos T/imunologia , Subpopulações de Linfócitos T/metabolismo , Animais , CamundongosRESUMO
Immunoglobulin A (IgA) provides protection against pathogens at mucosal surfaces. Chemotactic responses have been hypothesized to target IgA plasma cells involved in mucosal immune responses. We show here that thymus-expressed chemokine (TECK, CCL25) is a potent and selective chemoattractant for IgA antibody-secreting cells (ASC), efficiently recruiting IgA-producing cells from spleen, Peyer's patches, and mesenteric lymph node. Cells secreting IgA antibody in response to rotavirus, an intestinal pathogen, also respond well. In contrast, IgG- and IgM-ASC respond poorly. Epithelial cells in the small intestines, a principal site of IgA-ASC localization and IgA production in the body, highly and selectively express TECK. The migration of IgA-ASC to the intestinal epithelial cell chemokine TECK may help target IgA-producing cells to the gut wall, thus helping define and segregate the intestinal immune response.
Assuntos
Linfócitos B/imunologia , Quimiocinas CC/imunologia , Quimiotaxia de Leucócito/imunologia , Imunoglobulina A/imunologia , Animais , Linfócitos B/citologia , Feminino , Imunoglobulina A/biossíntese , Linfonodos/citologia , Linfonodos/imunologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Nódulos Linfáticos Agregados/citologia , Nódulos Linfáticos Agregados/imunologia , Baço/citologia , Baço/imunologiaRESUMO
Single-cell transcriptomics promise to revolutionize our understanding of the vasculature. Emerging computational methods applied to high-dimensional single-cell data allow integration of results between samples and species and illuminate the diversity and underlying developmental and architectural organization of cell populations. Here, we illustrate these methods in the analysis of mouse lymph node (LN) lymphatic endothelial cells (LEC) at single-cell resolution. Clustering identifies five well-delineated subsets, including two medullary sinus subsets not previously recognized as distinct. Nearest neighbor alignments in trajectory space position the major subsets in a sequence that recapitulates the known features and suggests novel features of LN lymphatic organization, providing a transcriptional map of the lymphatic endothelial niches and of the transitions between them. Differences in gene expression reveal specialized programs for (1) subcapsular ceiling endothelial interactions with the capsule connective tissue and cells; (2) subcapsular floor regulation of lymph borne cell entry into the LN parenchyma and antigen presentation; and (3) pathogen interactions and (4) LN remodeling in distinct medullary subsets. LEC of the subcapsular sinus floor and medulla, which represent major sites of cell entry and exit from the LN parenchyma respectively, respond robustly to oxazolone inflammation challenge with enriched signaling pathways that converge on both innate and adaptive immune responses. Integration of mouse and human single-cell profiles reveals a conserved cross-species pattern of lymphatic vascular niches and gene expression, as well as specialized human subsets and genes unique to each species. The examples provided demonstrate the power of single-cell analysis in elucidating endothelial cell heterogeneity, vascular organization, and endothelial cell responses. We discuss the findings from the perspective of LEC functions in relation to niche formations in the unique stromal and highly immunological environment of the LN.