技术资料

SummaryThe existing suite of therapies for bone diseases largely act to prevent further bone loss but fail to stimulate healthy bone formation and repair. We describe an endogenous osteopeptide (PEPITEM) with anabolic osteogenic activity,regulating bone remodeling in health and disease. PEPITEM acts directly on osteoblasts through NCAM-1 signaling to promote their maturation and formation of new bone,leading to enhanced trabecular bone growth and strength. Simultaneously,PEPITEM stimulates an inhibitory paracrine loop: promoting osteoblast release of the decoy receptor osteoprotegerin,which sequesters RANKL,thereby limiting osteoclast activity and bone resorption. In disease models,PEPITEM therapy halts osteoporosis-induced bone loss and arthritis-induced bone damage in mice and stimulates new bone formation in osteoblasts derived from patient samples. Thus,PEPITEM offers an alternative therapeutic option in the management of diseases with excessive bone loss,promoting an endogenous anabolic pathway to induce bone remodeling and redress the imbalance in bone turnover. Graphical abstract Highlights•PEPITEM exerts anabolic osteogenic activity to regulate osteoblast-osteoclast coupling•PEPITEM acts directly on osteoblasts to promote formation of new and stronger bone•PEPITEM stimulates an inhibitory paracrine loop via OPG to limit bone resorption•PEPITEM therapy halts disease-induced bone loss in vivo Lewis and Frost et al. identify the anabolic activity of an endogenous osteopeptide (PEPITEM),revealing the cellular and molecular mechanisms by which PEPITEM regulates bone remodeling in vitro and in preclinical disease models,to promote new bone formation. They suggest that PEPITEM offers an alternative therapeutic option for bone loss diseases. View Publication

Bacteria such as the oral microbiome member Peptostreptococcus anaerobius can exacerbate colorectal cancer (CRC) development. Little is known regarding whether these immunomodulatory bacteria also affect antitumour immune checkpoint blockade therapy. Here we show that administration of P. anaerobius abolished the efficacy of anti-PD1 therapy in mouse models of CRC. P. anaerobius both induced intratumoral myeloid-derived suppressor cells (MDSCs) and stimulated their immunosuppressive activities to impair effective T cell responses. Mechanistically,P. anaerobius administration activated integrin α2β1–NF-κB signalling in CRC cells to induce secretion of CXCL1 and recruit CXCR2+ MDSCs into tumours. The bacterium also directly activated immunosuppressive activity of intratumoral MDSCs by secreting lytC_22,a protein that bound to the Slamf4 receptor on MDSCs and promoted ARG1 and iNOS expression. Finally,therapeutic targeting of either integrin α2β1 or the Slamf4 receptor were revealed as promising strategies to overcome P. anaerobius-mediated resistance to anti-PD1 therapy in CRC. Interactions between Peptostreptococcus anaerobius and host cells promote recruitment and activation of myeloid-derived suppressor cells,leading to anti-PD1 immune checkpoint inhibitor resistance and exacerbated colorectal cancer in mice. View Publication

Haematopoietic stem cell (HSC) transplantation (HSCT) is the only curative treatment for a broad range of haematological malignancies,but the standard of care relies on untargeted chemotherapies and limited possibilities to treat malignant cells after HSCT without affecting the transplanted healthy cells1. Antigen-specific cell-depleting therapies hold the promise of much more targeted elimination of diseased cells,as witnessed in the past decade by the revolution of clinical practice for B cell malignancies2. However,target selection is complex and limited to antigens expressed on subsets of haematopoietic cells,resulting in a fragmented therapy landscape with high development costs2–5. Here we demonstrate that an antibody–drug conjugate (ADC) targeting the pan-haematopoietic marker CD45 enables the antigen-specific depletion of the entire haematopoietic system,including HSCs. Pairing this ADC with the transplantation of human HSCs engineered to be shielded from the CD45-targeting ADC enables the selective eradication of leukaemic cells with preserved haematopoiesis. The combination of CD45-targeting ADCs and engineered HSCs creates an almost universal strategy to replace a diseased haematopoietic system,irrespective of disease aetiology or originating cell type. We propose that this approach could have broad implications beyond haematological malignancies. An antibody–drug conjugate that targets the pan-haematopoietic marker CD45 combined with transplanted stem cells engineered to be shielded from it can eradicate leukaemic cells while preserving haematopoiesis. View Publication

SummaryAerobic exercise training (AET) has emerged as a strategy to reduce cancer mortality,however,the mechanisms explaining AET on tumor development remain unclear. Tumors escape immune detection by generating immunosuppressive microenvironments and impaired T cell function,which is associated with T cell mitochondrial loss. AET improves mitochondrial content and function,thus we tested whether AET would modulate mitochondrial metabolism in tumor-infiltrating lymphocytes (TIL). Balb/c mice were subjected to a treadmill AET protocol prior to CT26 colon carcinoma cells injection and until tumor harvest. Tissue hypoxia,TIL infiltration and effector function,and mitochondrial content,morphology and function were evaluated. AET reduced tumor growth,improved survival,and decreased tumor hypoxia. An increased CD8+ TIL infiltration,IFN-γ and ATP production promoted by AET was correlated with reduced mitochondrial loss in these cells. Collectively,AET decreases tumor growth partially by increasing CD8+ TIL effector function through an improvement in their mitochondrial content and function. Graphical abstract Highlights•Exercise training reduces tumor growth and improves survival in colorectal cancer•Trained mice present tumors with less hypoxia and higher CD8+ T cells infiltration•The production of IFNγ by CD8+ TIL is increased in exercise-trained mice•CD8+ TIL from trained mice show higher mitochondrial density and function Natural sciences; Biological sciences; Biochemistry; Physiology; Immunology; Systems biology; Cancer systems biology View Publication

SummarySepsis survivors are at high risk for infection-related rehospitalization and mortality for years following the resolution of the acute septic event. These infection-causing microorganisms generally do not cause disease in immunocompetent hosts,suggesting that the post-septic immune response is compromised. Given the importance of CD4 T cells in the development of long-lasting protective immunity,we analyzed their post-septic function. Here we showed that sepsis induced chronic increased and non-specific production of IL-17 by CD4 T cells,resulting in the inability to mount an effective immune response to a secondary pneumonia challenge. Altered cell function was associated with metabolic reprogramming,characterized by mitochondrial dysfunction and increased glycolysis. This metabolic reprogramming began during the acute septic event and persisted long after sepsis had resolved. Our findings reveal cell metabolism as a potential therapeutic target. Given the critical role of cell metabolism in the physiological and pathophysiological processes of immune cells,these findings reveal a potential new therapeutic target to help mitigate sepsis survivors’ susceptibility to secondary infections. Graphical abstract Highlights•Sepsis survivors demonstrate dysfunctional CD4 T cell immunity•Sepsis induces persistent mitochondrial dysfunction in CD4 T cells•Post-septic CD4 T cells are highly glycolytic and exhibit a Th17 phenotype•Sepsis impairs the CD4 T cell recall response Physiology; Molecular biology; Immunology; Components of the immune system View Publication

Recently developed small-molecule inhibitors of the lysosomal protease dipeptidyl peptidase 1 (DPP1),also known as cathepsin C (CatC),can suppress suppurative inflammation in vivo by blocking the processing of zymogenic (pro-) forms of neutrophil serine proteases (NSPs),including neutrophil elastase,proteinase 3,and cathepsin G. DPP1 also plays an important role in activating granzyme serine proteases that are expressed by cytotoxic T lymphocytes (CTL) and natural killer (NK) cells. Therefore,it is critical to determine whether DPP1 inhibition can also cause off-target suppression of CTL/NK-cell-mediated killing of virus-infected or malignant cells. Herein,we demonstrate that the processing of human granzymes A and B,transitioning from zymogen to active proteases,is not solely dependent on DPP1. Thus,the killing of target cells by primary human CD8+ T cells,NK cells,and gene-engineered anti-CD19 CAR T cells was not blocked in vitro even after prior exposure to high concentrations of the reversible DPP1 inhibitor brensocatib. Consistent with this observation,the turnover of model granzyme A/B peptide substrates in the human CTL/NK cell lysates was not significantly reduced by brensocatib. In contrast,preincubation with brensocatib almost entirely abolished (>90%) both the cytotoxic activity of mouse CD8+ T cells and granzyme substrate turnover. Overall,our finding that the effects of DPP1 inhibition on human cytotoxic lymphocytes are attenuated in comparison to those of mice indicates that granzyme processing/activation pathways differ between mice and humans. Moreover,the in vitro data suggest that human subjects treated with reversible DPP1 inhibitors,such as brensocatib,are unlikely to experience any appreciable deficits in CTL/NK-cell-mediated immunities. View Publication

Humanized mice are limited in terms of modeling human immunity,particularly with regards to antibody responses. Here we constructed a humanized (THX) mouse by grafting non-γ-irradiated,genetically myeloablated KitW-41J mutant immunodeficient pups with human cord blood CD34+ cells,followed by 17β-estradiol conditioning to promote immune cell differentiation. THX mice reconstitute a human lymphoid and myeloid immune system,including marginal zone B cells,germinal center B cells,follicular helper T cells and neutrophils,and develop well-formed lymph nodes and intestinal lymphoid tissue,including Peyer’s patches,and human thymic epithelial cells. These mice have diverse human B cell and T cell antigen receptor repertoires and can mount mature T cell-dependent and T cell-independent antibody responses,entailing somatic hypermutation,class-switch recombination,and plasma cell and memory B cell differentiation. Upon flagellin or a Pfizer-BioNTech coronavirus disease 2019 (COVID-19) mRNA vaccination,THX mice mount neutralizing antibody responses to Salmonella or severe acute respiratory syndrome coronavirus 2 Spike S1 receptor-binding domain,with blood incretion of human cytokines,including APRIL,BAFF,TGF-β,IL-4 and IFN-γ,all at physiological levels. These mice can also develop lupus autoimmunity after pristane injection. By leveraging estrogen activity to support human immune cell differentiation and maturation of antibody responses,THX mice provide a platform to study the human immune system and to develop human vaccines and therapeutics. Humanized mice have been a valuable tool for modeling human immunology but are limited in their ability to model human antibody responses. Here the authors present their THX humanized mouse that does model human antibody responses and test its suitability for vaccination and autoimmunity studies. View Publication

Acute systemic inflammation critically alters the function of the immune system,often promoting myelopoiesis at the expense of lymphopoiesis. In the thymus,systemic inflammation results in acute thymic atrophy and,consequently,impaired T-lymphopoiesis. The mechanism by which systemic inflammation impacts the thymus beyond suppressing T-cell development is still unclear. Here,we describe how the synergism between TL1A and IL-18 suppresses T-lymphopoiesis to promote thymic myelopoiesis. The protein levels of these two cytokines were elevated in the thymus during viral-induced thymus atrophy infection with murine cytomegalovirus (MCMV) or pneumonia virus of mice (PVM). In vivo administration of TL1A and IL-18 induced acute thymic atrophy,while thymic neutrophils expanded. Fate mapping with Ms4a3-Cre mice demonstrated that thymic neutrophils emerge from thymic granulocyte-monocyte progenitors (GMPs),while Rag1-Cre fate mapping revealed a common developmental path with lymphocytes. These effects could be modeled ex vivo using neonatal thymic organ cultures (NTOCs),where TL1A and IL-18 synergistically enhanced neutrophil production and egress. NOTCH blockade by the LY411575 inhibitor increased the number of neutrophils in the culture,indicating that NOTCH restricted steady-state thymic granulopoiesis. To promote myelopoiesis,TL1A,and IL-18 synergistically increased GM-CSF levels in the NTOC,which was mainly produced by thymic ILC1s. In support,TL1A- and IL-18-induced granulopoiesis was completely prevented in NTOCs derived from Csf2rb-/- mice and by GM-CSFR antibody blockade,revealing that GM-CSF is the essential factor driving thymic granulopoiesis. Taken together,our findings reveal that TL1A and IL-18 synergism induce acute thymus atrophy while promoting extramedullary thymic granulopoiesis in a NOTCH and GM-CSF-controlled manner. View Publication

Visceral white adipose tissues (WAT) regulate systemic lipid metabolism and inflammation. Dysfunctional WAT drive chronic inflammation and facilitate atherosclerosis. Adipose tissue-associated macrophages (ATM) are the predominant immune cells in WAT,but their heterogeneity and phenotypes are poorly defined during atherogenesis. The scavenger receptor CD36 mediates ATM crosstalk with other adipose tissue cells,driving chronic inflammation. Here,we combined the single-cell RNA sequencing technique with cell metabolic and functional assays on major WAT ATM subpopulations using a diet-induced atherosclerosis mouse model (Apoe-null). We also examined the role of CD36 using Apoe/Cd36 double-null mice. Based on transcriptomics data and differential gene expression analysis,we identified a previously undefined group of ATM displaying low viability and high lipid metabolism and labeled them as “unhealthy macrophages”. Their phenotypes suggest a subpopulation of ATM under lipid stress. We also identified lipid-associated macrophages (LAM),which were previously described in obesity. Interestingly,LAM increased 8.4-fold in Apoe/Cd36 double-null mice on an atherogenic diet,but not in Apoe-null mice. The increase in LAM was accompanied by more ATM lipid uptake,reduced adipocyte hypertrophy,and less inflammation. In conclusion,CD36 mediates a delicate balance between lipid metabolism and inflammation in visceral adipose tissues. Under atherogenic conditions,CD36 deficiency reduces inflammation and increases lipid metabolism in WAT by promoting LAM accumulation. View Publication

The integrity of the blood–labyrinth barrier (BLB) is essential for inner ear homeostasis,regulating the ionic composition of endolymph and perilymph and preventing harmful substance entry. Endothelial hyperpermeability,central in inflammatory and immune responses,is managed through complex intercellular communication and molecular signaling pathways. Recent studies link BLB permeability dysregulation to auditory pathologies like acoustic trauma,autoimmune inner ear diseases,and presbycusis. Polymorphonuclear granulocytes (PMNs),or neutrophils,significantly modulate vascular permeability,impacting endothelial barrier properties. Neutrophil extracellular traps (NETs) are involved in diseases with autoimmune and autoinflammatory bases. The present study evaluated the impact of NETs on a BLB cellular model using a Transwell® setup. Our findings revealed a concentration-dependent impact of NETs on human inner ear-derived endothelial cells. In particular,endothelial permeability markers increased,as indicated by reduced transepithelial electrical resistance,enhanced dextran permeability,and downregulated junctional gene expression (ZO1,OCL,and CDH5). Changes in cytoskeletal architecture were also observed. These preliminary results pave the way for further research into the potential involvement of NETs in BLB impairment and implications for auditory disorders. View Publication

This study sheds light on the pivotal role of the oncoprotein DEK in B-cell lymphoma. We reveal DEK expression correlates with increased tumor proliferation and inferior overall survival in cases diagnosed with low-grade B-cell lymphoma (LGBCL). We also found significant correlation between DEK expression and copy number alterations in LGBCL tumors,highlighting a novel mechanism of LGBCL pathogenesis that warrants additional exploration. To interrogate the mechanistic role of DEK in B-cell lymphoma,we generated a DEK knockout cell line model,which demonstrated DEK depletion caused reduced proliferation and altered expression of key cell cycle and apoptosis-related proteins,including Bcl-2,Bcl-xL,and p53. Notably,DEK depleted cells showed increased sensitivity to apoptosis-inducing agents,including venetoclax and staurosporine,which underscores the therapeutic potential of targeting DEK in B-cell lymphomas. Overall,our study contributes to a better understanding of DEK’s role as an oncoprotein in B-cell lymphomas,highlighting its potential as both a promising therapeutic target and a novel biomarker for aggressive LGBCL. Further research elucidating the molecular mechanisms underlying DEK-mediated tumorigenesis could pave the way for improved treatment strategies and better clinical outcomes for patients with B-cell lymphoma. View Publication

Centrioles define centrosome structure and function. Deregulation of centriole numbers can cause developmental defects and cancer. The p53 tumor suppressor limits the growth of cells lacking or harboring additional centrosomes and can be engaged by the “mitotic surveillance” or the “PIDDosome pathway”,respectively. Here,we show that early B cell progenitors frequently present extra centrioles,ensuing their high proliferative activity and related DNA damage. Extra centrioles are efficiently cleared during B cell maturation. In contrast,centriole loss upon Polo-like kinase 4 (Plk4) deletion causes apoptosis and arrests B cell development. This defect can be rescued by co-deletion of Usp28,a critical component of the mitotic surveillance pathway,that restores cell survival and maturation. Centriole-deficient mature B cells are proliferation competent and mount a humoral immune response. Our findings imply that progenitor B cells are intolerant to centriole loss but permissive to centriole amplification,a feature potentially facilitating their malignant transformation. Centrioles organize chromosome segregation,migration,and the immune synapse. Here,Schapfl et al. show that B cell progenitors tolerate centriole overamplification,but not loss,while mature B cells can mount a humoral immune response in their absence. View Publication