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BACKGROUNDSepsis is a severe illness characterized by systemic and multiorgan reactive responses and damage. However,the impact of sepsis on the bone marrow,particularly on bone marrow mesenchymal stem cells (BMSCs),is less reported. BMSCs are critical stromal cells in the bone marrow microenvironment that maintain bone stability and hematopoietic homeostasis; however,the impairment caused by sepsis remains unknown.AIMTo investigate the effects of sepsis on BMSCs and the underlying mechanisms.METHODSBMSCs were obtained from healthy donors and patients with sepsis. We compared the self-renewal capacity,differentiation potential,and hematopoietic supportive ability in vitro. Senescence of septic BMSCs was assessed using β-galactosidase staining,senescence-associated secretory phenotype,intracellular reactive oxygen species levels,and the expression of P16 and P21. Finally,the changes in septic BMSCs after nicotinamide adenine dinucleotide (NAD) treatment were evaluated.RESULTSSeptic BMSCs showed decreased proliferation and self-renewal,bias towards adipogenic differentiation,and weakened osteogenic differentiation. Additionally,hematopoietic supportive capacity declines in sepsis. The levels of aging markers were significantly higher in the septic BMSCs. After NAD treatment,the proliferation capacity of septic BMSCs showed a recovery trend,with increased osteogenic and hematopoietic supportive capacities. Sepsis resulted in decreased expression of sirtuin 3 (SIRT3) in BMSCs,whereas NAD treatment restored SIRT3 expression,enhanced superoxide dismutase enzyme activity,reduced intracellular reactive oxygen species levels,maintained mitochondrial stability and function,and ultimately rejuvenated septic BMSCs.CONCLUSIONSepsis accelerates the aging of BMSCs,as evidenced by a decline in self-renewal and osteogenic capabilities,as well as weakened hematopoietic support functions. These deficiencies can be effectively reversed via the NAD/SIRT3/superoxide dismutase pathway. View Publication

Immune dysregulation-induced inflammation serves as a driving force in the progression of metabolic dysfunction-associated steatohepatitis (MASH),while the underlying cellular and molecular mechanisms remain largely uncharted. A Western diet (WD) is employed to construct mouse models of metabolic dysfunction associated steatotic liver disease (MASLD) or MASH. Mass cytometry identifies a c-kit+ cDC1 subset whose frequency is reduced in the livers of mice and patients with MASH compared with healthy controls. Adoptive cell transfer of c-kit+ cDC1 protects the progression of MASH. Moreover,analysis of gut microbe sequence shows that WD-fed mice and MASLD/MASH patients exhibit gut microbiota dysbiosis,with an elevated abundance of H2S-producing Desulfovibrio_sp. Transplanting of MASH-derived fecal flora,Desulfovibrio_sp.,or injecting H2S intraperitoneally into MASLD mice decreases the c-kit+cDC1 population and exacerbates liver inflammation. Mechanistically,H2S induces autophagic cell death of cDC1 in a c-kit-dependent manner in cDC-specific c-kit-/- and Atg5-/- mice. We thus uncover that microbiota-derived H2S triggers the autophagic cell death of c-kit+ cDC1 and ignites the liver inflammatory cascade in MASH. The immune regulatory mechanism for metabolic dysfunction-associated steatohepatitis (MASH) remains elusive. Here,the authors identify a c-kit+ cDC1 subset,which can be depleted by Desulfovibrio_sp.-induced H2S via autophagic cell death and contributing to uncontrolled inflammation for MASH progression. View Publication

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

Regulatory T cells (Treg) play an important role in regulating immune homeostasis in health and disease. Traditionally their suppressive function has been assayed by mixing purified cell populations,which does not provide an accurate picture of a physiologically relevant response. To overcome this limitation,we here develop ‘single cell suppression profiling of human Tregs’ (scSPOT). scSPOT uses a 52-marker CyTOF panel,a cell division detection algorithm,and a whole PBMC system to assess the effect of Tregs on all other cell types simultaneously. In this head-to-head comparison,we find Tregs having the clearest suppressive effects on effector memory CD8 T cells through partial division arrest,cell cycle inhibition,and effector molecule downregulation. Additionally,scSPOT identifies a Treg phenotypic split previously observed in viral infection and propose modes of action by the FDA-approved drugs Ipilimumab and Tazemetostat. scSPOT is thus scalable,robust,widely applicable,and may be used to better understand Treg immunobiology and screen for therapeutic compounds. Traditional regulatory T cell (Tregs) assays utilize mixture of purified cell population. Here the authors develop a ‘single cell suppression profiling of human Tregs’ (scSPOT) with 52-marker CyTOF panel,a cell division detection algorithm,and a whole PBMC system to assess Treg suppressive function on all cell types simultaneously. 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

Ginseng may improve the myelosuppression and intestinal microbiota disorder induced by cyclophosphamide (CY); however,the effect of ginseng components on hematopoietic stem cell (HSC) damage remains largely unexplored. The present study aimed to assess the protective effect of ginseng extract (GE),total ginsenosides (TG) and total polysaccharides (TP) from ginseng on the intestinal microflora and HSCs of model mice. In the present study,a mouse model of HSC damage induced by CY was constructed,intestinal microflora of fecal samples were sequenced using the 16S ribosomal RNA (rRNA) sequencing techniques,the differentially expressed genes (DEGs) of HSCs were analyzed using high-throughput RNA-sequencing,cell apoptosis and erythroid differentiation were detected using flow cytometry and the blood cell parameters were analyzed using a hematology analyzer. Analysis of the 16S rRNA in fecal samples showed that GE,TG and TP improved an imbalanced intestinal microflora,where the relative abundance of Lactobacillus intestinalis had a positive correlation with ginsenosides content. Specifically,TP significantly increased the expression of low-abundance microflora. Transcriptomic analysis results revealed 2,250,3,432 and 261 DEGs in the GE,TG and TP groups compared with those in the Model group,respectively. In the expression analysis of DEGs,both TG and GE were found to markedly increase the expression levels of Klf4,Hhex,Pbx1,Kmt2a,Mecom,Zc3h12a,Zbtb16,Lilr4b,Flt3 and Klf13. Furthermore,TG inhibited the apoptosis of HSCs by increasing the expression levels of Bcl2 and Mcl1,whilst decreasing the expression of Bax. By contrast,GE inhibited the apoptosis of HSCs by reducing the expression of Bax and Bad. Regarding erythroid differentiation and blood cell parameters,GE was found to significantly increase the expression of TER-119. In addition,GE and TG improved all blood cell parameters,including the count of white blood cells,neutrophils (NEUT),lymphocytes (LYMPH),red blood cells (RBC),hemoglobin (HGB) and reticulocyte and platelets (PLT),whereas TP could only improve the counts of LYMPH,RBC,HGB and PLT. The improvement effect of GE and TG on WBC,NEUT and Ret was superior to TP. In conclusion,TG may protect the hematopoiesis function of HSCs in a CY-induced mouse model of HSC damage,followed by GE. However,TP did not appear to improve HSC damage. Ginsenosides may therefore be considered essential ingredients in GE when protecting HSCs against damage. GE and TG exerted their protective effects on HSCs by inhibiting the apoptosis of HSCs whilst improving the imbalance of intestinal microflora. View Publication