技术资料

Characterizing the HIV-1 reservoir in blood and tissues is crucial for the development of curative strategies. Using an HIV Tat mRNA-containing lipid nanoparticle (Tat-LNP) in combination with panobinostat,we show that p24+ cells from blood and lymph nodes exhibit distinct phenotypes. Blood p24+ cells are found in both central/transitional (TCM/TTM) and effector memory subsets,mostly lack CXCR5 expression and are enriched in GZMA+ cells. In contrast,most lymph node p24+ cells display a TCM/TTM phenotype,with approximately 50% expressing CXCR5 and nearly all lacking GZMA expression. Furthermore,germinal center T follicular helper cells do not appear to harbor the translation-competent reservoir in long-term suppressed individuals. Near full-length HIV-1 sequencing in longitudinal samples from matched blood,lymph nodes,and gut indicates that clones of infected cells,including those carrying an inducible provirus,persist and spread across various anatomical compartments. Finally,uniform genetic diversity across sites suggests the absence of ongoing replication in tissues under treatment. Here,Pardons and Lambrechts et al show that HIV-1 reservoirs in blood and lymph nodes differ phenotypically. Furthermore, germinal center T follicular helper cells do not harbor the inducible reservoir in long-term suppressed individuals. Infected clones can spread across tissues and persist without active replication. View Publication

Liver metastasis (LM) poses a significant challenge in cancer treatment,with limited available therapeutic options and poor prognosis. Understanding the dynamics of tumor microenvironment (TME) and immune interactions is crucial for developing effective treatments. We find that WNT11 promoted CD8+ T-cell exclusion and suppression,which was correlated with poor prognosis in LM. Mechanistically,WNT11-overexpressing tumor cells directly reduce CD8+ T-cell recruitment and activity by decreasing CXCL10 and CCL4 expression through CAMKII-mediated β-catenin/AFF3 downregulation. WNT11-overexpressing tumor cells promote immunosuppressive macrophage polarization by inducing IL17D expression via the CAMKII/NF-κB pathway,which result in CD8+ T-cell suppression. Moreover,CAMKII inhibition increases the efficacy of anti-PD-1 therapy in mouse model of LM. Serum expression of WNT11 is identified as a potential minimally invasive biomarker in the management of colorectal cancer-LM with immunotherapy. Our findings highlight WNT11/CAMKII axis as a critical regulator of the TME and a promising target for immunotherapy in patients with LM. Activation of the WNT/β-catenin signaling pathway has been associated with immune evasion in several cancer types. Here the authors show that expression of WNT11,a member of the non-canonical WNT signaling pathway,is associated with CD8 + T cell exclusion and resistance to immune checkpoint inhibitors in liver metastasis. View Publication

Type I Interferons (IFN-I) are central to host protection against viral infections,with plasmacytoid dendritic cells (pDC) being the most significant source,yet pDCs lose their IFN-I production capacity following an initial burst of IFN-I,resulting in susceptibility to secondary infections. The underlying mechanisms of these dynamics are not well understood. Here we find that viral infection reduces the capacity of pDCs to engage both oxidative and glycolytic metabolism. Mechanistically,we identify lactate dehydrogenase B (LDHB) as a positive regulator of pDC IFN-I production in mice and humans; meanwhile,LDHB deficiency is associated with suppressed IFN-I production,pDC metabolic capacity,and viral control following infection. In addition,preservation of LDHB expression is sufficient to partially retain the function of otherwise exhausted pDCs,both in vitro and in vivo. Furthermore,restoring LDHB in vivo in pDCs from infected mice increases IFNAR-dependent,infection-associated pathology. Our work thus identifies a mechanism for balancing immunity and pathology during viral infections,while also providing insight into the highly preserved infection-driven pDC inhibition. Plasmacytoid dendritic cells (pDC) are the major IFN-I-producing cells,but this production returns to baseline soon after viral infection. Here the authors show that this decrease in IFN-I production and related pDC functions may be attributed to suppressed oxidative and glycolytic metabolism of pDCs,with lactate dehydrogenase B identified as a regulator. View Publication

The fallopian tube undergoes extensive molecular changes during the menstrual cycle and menopause. We use single-cell RNA and ATAC sequencing to construct a comprehensive cell atlas of healthy human fallopian tubes during the menstrual cycle and menopause. Our scRNA-seq comparison of 85,107 pre- and 46,111 post-menopausal fallopian tube cells reveals substantial shifts in cell type frequencies,gene expression,transcription factor activity,and cell-to-cell communications during menopause and menstrual cycle. Menstrual cycle dependent hormonal changes regulate distinct molecular states in fallopian tube secretory epithelial cells. Postmenopausal fallopian tubes show high chromatin accessibility in transcription factors associated with aging such as Jun,Fos,and BACH1/2,while hormone receptors were generally downregulated,a small proportion of secretory epithelial cells had high expression of ESR2,IGF1R,and LEPR. While a pre-menopausal secretory epithelial gene cluster is enriched in the immunoreactive molecular subtype,a subset of genes expressed in post-menopausal secretory epithelial cells show enrichment in the mesenchymal molecular type of high-grade serous ovarian cancer. The fallopian tube undergoes extensive cellular and molecular changes during the menstrual cycle and aging. Here,Weigert et al. present a single-cell atlas of the normal human fallopian tube revealing the transition of secretory epithelial cells throughout the menstrual cycle and menopause. View Publication

Despite significant advancements in targeted- and immunotherapies,millions of patients with cancer still succumb to the disease each year. In renal cell carcinoma,up to 25% of metastatic patients do not respond to first-line therapies. This reality underscores the urgent need for innovative or repurposed therapies to effectively treat these patients. Patient-derived organoids represent a promising model for evaluating treatment efficacy and toxicity,offering a potential breakthrough in personalized medicine. However,utilizing organoid models for drug screening presents several challenges. Our protocol aims to address these obstacles by outlining a practical approach to successfully isolate and cultivate patient-derived renal cell carcinoma and kidney organoids for treatment screening purposes. Graphical abstract Patient-derived organoids represent a promising model for evaluating treatment efficacy and toxicity,offering a potential breakthrough in personalized medicine. Nowak-Sliwinska and colleagues present a detailed protocol for obtaining kidney and kidney cancer organoids for drug development and precision oncology. View Publication

In dynamic systems like the circulatory system,establishing localized cytokine gradients is challenging. Upon lipopolysaccharide (LPS) stimulation,we observed that monocytes release numerous migrasomes enriched with inflammatory cytokines,such as TNF-α and IL-6. These cytokines are transported into migrasomes via secretory carriers,leading to their immediate exocytosis or eventual release from detached migrasomes. We successfully isolated TNF-α and IL-6-enriched,monocyte-derived migrasomes from the blood of LPS-treated mice. Total secretion analysis revealed a substantial amount of TNF-α and IL-6 released in a migrasome-packaged form. Thus,detached,monocyte-derived migrasomes represent a type of extracellular vesicle highly enriched with cytokines. Physiologically,these cytokine-laden migrasomes rapidly accumulate at local sites of inflammation,effectively creating a concentrated source of cytokines. Our research uncovers novel mechanisms for cytokine release and delivery,providing new insights into immune response modulation. View Publication

BackgroundAcute respiratory distress syndrome (ARDS) is a life-threatening condition associated with the inflammatory activation of alveolar macrophages. Here,we examined the role of circVAPA in regulating inflammasome activation and macrophage inflammatory polarization in an ARDS model.MethodscircVAPA expression levels were analyzed in macrophages isolated from healthy controls and patients with ARDS. In vitro cell models of mouse alveolar macrophages and an in vivo mouse ARDS model were established through Lipopolysaccharide (LPS) stimulation. The effects of circVAPA knockdown on macrophage inflammatory polarization,inflammasome activation,and pulmonary tissue damage were investigated in both cell and animal models. The interaction between circVAPA and downstream factors was verified through a luciferase reporter assay and by silencing circVAPA.ResultscircVAPA upregulation in alveolar macrophages was associated with the inflammation in ARDS patients. circVAPA was also upregulated in LPS-stimulated mouse alveolar macrophages (MH-S cells). Additionally,circVAPA knockdown attenuated the inflammatory activation of MH-S cells and reduced the expression of pyroptosis-related proteins. circVAPA silencing also mitigated the inflammatory effects of LPS-stimulated MH-S cells on lung epithelial cells (MLE-12),and alleviated the inflammatory damage in the pulmonary tissue of ARDS mouse model. We further showed that miR-212-3p/Sirt1 axis mediated the functional role of circVAPA in the inflammatory polarization of MH-S cells.ConclusionOur data suggest that circVAPA promotes inflammasome activity and macrophage inflammation by modulating miR-212-3p/Sirt1 axis in ARDS. Targeting circVAPA may be employed to suppress the inflammatory activation of alveolar macrophages in ARDS.Supplementary InformationThe online version contains supplementary material available at 10.1007/s10735-024-10312-3. View Publication

Germinal center (GC) formation,which is an integrant part of humoral immunity,involves energy-consuming metabolic reprogramming. Rag-GTPases are known to signal amino acid availability to cellular pathways that regulate nutrient distribution such as the mechanistic target of rapamycin complex 1 (mTORC1) pathway and the transcription factors TFEB and TFE3. However,the contribution of these factors to humoral immunity remains undefined. Here,we show that B cell-intrinsic Rag-GTPases are critical for the development and activation of B cells. RagA/RagB deficient B cells fail to form GCs,produce antibodies,and to generate plasmablasts during both T-dependent (TD) and T-independent (TI) humoral immune responses. Deletion of RagA/RagB in GC B cells leads to abnormal dark zone (DZ) to light zone (LZ) ratio and reduced affinity maturation. Mechanistically,the Rag-GTPase complex constrains TFEB/TFE3 activity to prevent mitophagy dysregulation and maintain mitochondrial fitness in B cells,which are independent of canonical mTORC1 activation. TFEB/TFE3 deletion restores B cell development,GC formation in Peyer’s patches and TI humoral immunity,but not TD humoral immunity in the absence of Rag-GTPases. Collectively,our data establish the Rag GTPase-TFEB/TFE3 pathway as a likely mTORC1 independent mechanism to coordinating nutrient sensing and mitochondrial metabolism in B cells. Rag-GTPases play roles in sensing nutrient availability,and it is not fully known how they contribute to energy-consuming immunological processes such as the B cell response. Here authors show that genomic deletion fo RagA/RagB distrupts both T-dependent and T-independent humoral immune responses,independent of mechanistic target of rapamycin complex 1 but involving the transcription factors TFEB and TFE3. View Publication

Primary antiphospholipid syndrome (PAPS) is a life-threatening clotting disorder mediated by pathogenic autoantibodies. Here we dissect the origin of self-reactive B cells in human PAPS using peripheral blood and bone marrow of patients with triple-positive PAPS via combined single-cell RNA sequencing,B cell receptors (BCR) repertoire profiling,CITEseq analysis and single cell immortalization. We find that antiphospholipid (aPL)-specific B cells are present in the naive compartment,polyreactive,and derived from the natural repertoire. Furthermore,B cells with aPL specificities are not eliminated in patients with PAPS,persist until the memory and long-lived plasma cell stages,likely after defective germinal center selection,while becoming less polyreactive. Lastly,compared with the non-PAPS cells,PAPS B cells exhibit distinct IFN and APRIL signature as well as dysregulated mTORC1 and MYC pathways. Our findings may thus elucidate the survival mechanisms of these autoreactive B cells and suggest potential therapeutic targets for the treatment of PAPS. Primary antiphospholipid syndrome (PAPS) is a clotting disorder attributed to autoreactive antibodies produced by B cells. Here the authors show,using single cell omics and B cell repertoire data,that autoreactive B cells originate from the natural B cell repertoire and escape germinal center selection to persist in PAPS patient via potential dysregulation of mTORC1 and MYC pathways. View Publication

Crystalline silica (CS) particle exposure leads to silicosis which is characterized as progressive fibrosis. Fibroblasts are vital effector cells in fibrogenesis. Emerging studies have identified immune sentinel roles for fibroblasts in chronic disease,while their immune-modulatory roles in silicosis remain unclear. Herein,we show that group 2 innate lymphoid cell (ILC2) conversion to ILC1s is closely involved in silicosis progression,which is mediated by activated fibroblasts via interleukin (IL)−18. Mechanistically,Notch3 signaling in mechanics-activated fibroblasts modulates IL-18 production via caspase 1 activity. The mouse-specific Notch3 knockout in fibroblasts retards pulmonary fibrosis progression that is linked to attenuated ILC conversion. Our results indicate that activated fibroblasts in silicotic lungs are regulators of ILC2–ILC1 conversion,associated with silicosis progression via the Notch3–IL-18 signaling axis. This finding broadens our understanding of immune-modulatory mechanisms in silicosis,and indicates potential therapeutic targets for lung fibrotic diseases. Crystalline silica particle exposure in the airways can lead to lung silicosis and progressive fibrosis. Here the authors use mouse silicosis models to show mechanics activated fibroblasts promote conversion of ILC2 to ILC1-like cells pulmonary fibrosis and that this is associated with a Notch3-IL-18 signalling pathway. View Publication

Despite recent advances in immunotherapies targeting single tumor-associated antigens,patients with multiple myeloma eventually relapse. ISB 2001 is a CD3+ T cell engager (TCE) co-targeting BCMA and CD38 designed to improve cytotoxicity against multiple myeloma. Targeting of two tumor-associated antigens by a single TCE resulted in superior cytotoxic potency across a variable range of BCMA and CD38 tumor expression profiles mimicking natural tumor heterogeneity,improved resistance to competing soluble factors and exhibited superior cytotoxic potency on patient-derived samples and in mouse models. Despite the broad expression of CD38 across human tissues,ISB 2001 demonstrated a reduced T cell activation profile in the absence of tumor cells when compared to TCEs targeting CD38 only. To determine an optimal first-in-human dose for the ongoing clinical trial (NCT05862012),we developed an innovative quantitative systems pharmacology model leveraging preclinical data,using a minimum pharmacologically active dose approach,therefore reducing patient exposure to subefficacious doses of therapies. Perro and colleagues develop a CD3+ T cell engager co-targeting BCMA and CD38 to improve immunotherapy for multiple myeloma,demonstrate cytotoxicity in patient-derived samples and murine models and develop a quantitative systems pharmacology model. View Publication

BackgroundMacrophage-based cell therapies have shown modest success in clinical trials,which can be attributed to their phenotypic plasticity,where transplanted macrophages get reprogrammed towards a pro-tumor phenotype. In most tumor types,including melanoma,the balance between antitumor M1-like and tumor-promoting M2-like macrophages is critical in defining the local immune response with a higher M1/M2 ratio favoring antitumor immunity. Therefore,designing novel strategies to increase the M1/M2 ratio in the TME has high clinical significance and benefits macrophage-based cell therapies.MethodsIn this study,we reprogrammed antitumor and proinflammatory macrophages ex-vivo with HDAC6 inhibitors (HDAC6i). We administered the reprogrammed macrophages intratumorally as an adoptive cell therapy (ACT) in the syngeneic SM1 murine melanoma model and patient-derived xenograft bearing NSG-SGM3 humanized mouse models. We phenotyped the tumor-infiltrated immune cells by flow cytometry and histological analysis of tumor sections for macrophage markers. We performed bulk RNA-seq profiling of murine bone marrow-derived macrophages treated with vehicle or HDAC6i and single-cell RNA-seq profiling of SM1 tumor-infiltrated immune cells to determine the effect of intratumor macrophage ACT on the tumor microenvironment (TME). We further analyzed the single-cell data to identify key cell-cell interactions and trajectory analysis to determine the fate of tumor-associated macrophages post-ACT.ResultsMacrophage ACT resulted in diminished tumor growth in both mouse models. We also demonstrated that HDAC6 inhibition in macrophages suppressed the polarization toward tumor-promoting phenotype by attenuating STAT3-mediated M2 reprogramming. Two weeks post-transplantation,ACT macrophages were viable,and inhibition of HDAC6 rendered intratumor transplanted M1 macrophages resistant to repolarization towards protumor M2 phenotype in-vivo. Further characterization of tumors by flow cytometry,single-cell transcriptomics,and single-cell secretome analyses revealed a significant enrichment of antitumor M1-like macrophages,resulting in increased M1/M2 ratio and infiltration of CD8 effector T-cells. Computational analysis of single-cell RNA-seq data for cell-cell interactions and trajectory analyses indicated activation of monocytes and T-cells in the TME.ConclusionsIn summary,for the first time,we demonstrated the potential of reprogramming macrophages ex-vivo with HDAC6 inhibitors as a viable macrophage cell therapy to treat solid tumors.Supplementary InformationThe online version contains supplementary material available at 10.1186/s13046-024-03182-w. View Publication