技术资料

Upon immunogenic challenge,lymph nodes become mechanically stiff as immune cells activate and proliferate within their encapsulated environments,and with resolution,they reestablish a soft baseline state. Here we show that sensing these mechanical changes in the microenvironment requires the mechanosensor YAP. YAP is induced upon activation and suppresses metabolic reprogramming of effector T cells. Unlike in other cell types in which YAP promotes proliferation,YAP in T cells suppresses proliferation in a stiffness-dependent manner by directly restricting the translocation of NFAT1 into the nucleus. YAP slows T cell responses in systemic viral infections and retards effector T cells in autoimmune diabetes. Our work reveals a paradigm whereby tissue mechanics fine-tune adaptive immune responses in health and disease. View Publication

The molecular basis of the earliest neuronal changes that lead to Alzheimer's disease (AD) is unclear. Here,we analyze neural cells derived from sporadic AD (SAD),APOE4 gene-edited and control induced pluripotent stem cells (iPSCs). We observe major differences in iPSC-derived neural progenitor (NP) cells and neurons in gene networks related to neuronal differentiation,neurogenesis,and synaptic transmission. The iPSC-derived neural cells from SAD patients exhibit accelerated neural differentiation and reduced progenitor cell renewal. Moreover,a similar phenotype appears in NP cells and cerebral organoids derived from APOE4 iPSCs. Impaired function of the transcriptional repressor REST is strongly implicated in the altered transcriptome and differentiation state. SAD and APOE4 expression result in reduced REST nuclear translocation and chromatin binding,and disruption of the nuclear lamina. Thus,dysregulation of neural gene networks may set in motion the pathologic cascade that leads to AD. View Publication

Adipose tissues dynamically remodel their cellular composition in response to external cues by stimulating beige adipocyte biogenesis; however,the developmental origin and pathways regulating this process remain insufficiently understood owing to adipose tissue heterogeneity. Here,we employed single-cell RNA-seq and identified a unique subset of adipocyte progenitor cells (APCs) that possessed the cell-intrinsic plasticity to give rise to beige fat. This beige APC population is proliferative and marked by cell-surface proteins,including PDGFR$\alpha$,Sca1,and CD81. Notably,CD81 is not only a beige APC marker but also required for de novo beige fat biogenesis following cold exposure. CD81 forms a complex with $\alpha$V/$\beta$1 and $\alpha$V/$\beta$5 integrins and mediates the activation of integrin-FAK signaling in response to irisin. Importantly,CD81 loss causes diet-induced obesity,insulin resistance,and adipose tissue inflammation. These results suggest that CD81 functions as a key sensor of external inputs and controls beige APC proliferation and whole-body energy homeostasis. View Publication

Oncolytic virus therapy leads to immunogenic death of virus-infected tumor cells and this has been shown in preclinical models to enhance the cytotoxic T-lymphocyte response against tumor-associated antigens (TAAs),leading to killing of uninfected tumor cells. To investigate whether oncolytic virotherapy can increase immune responses to tumor antigens in human subjects,we studied T-cell responses against a panel of known myeloma TAAs using PBMC samples obtained from ten myeloma patients before and after systemic administration of an oncolytic measles virus encoding sodium iodide symporter (MV-NIS). Despite their prior exposures to multiple immunosuppressive antimyeloma treatment regimens,T-cell responses to some of the TAAs were detectable even before measles virotherapy. Measurable baseline T-cell responses against MAGE-C1 and hTERT were present. Furthermore,MV-NIS treatment significantly (P {\textless} 0.05) increased T-cell responses against MAGE-C1 and MAGE-A3. Interestingly,one patient who achieved complete remission after MV-NIS therapy had strong baseline T-cell responses both to measles virus proteins and to eight of the ten tested TAAs. Our data demonstrate that oncolytic virotherapy can function as an antigen agnostic vaccine,increasing cytotoxic T-lymphocyte responses against TAAs in patients with multiple myeloma,providing a basis for continued exploration of this modality in combination with immune checkpoint blockade. View Publication

Hepatocellular carcinoma (HCC) is among the most common causes of cancer death in men. Whether or not a longitudinal follow-up of circulating tumor cells (CTCs) before and at different time points during systemic/targeted therapy is useful for monitoring the treatment response of patients with locally advanced or metastatic HCC has been evaluated in this study. Blood samples (n = 104) were obtained from patients with locally advanced or metastatic HCC (n = 30) for the enrichment of CTCs by a negative selection method. Analysis of the blood samples from patients with defined disease status (n = 81) revealed that those with progressive disease (PD,n = 37) had significantly higher CTC counts compared to those with a partial response (PR) or stable disease (SD; n = 44 for PR + SD,p = 0.0002). The median CTC count for patients with PD and for patients with PR and SD was 50 (interquartile range 21-139) and 15 (interquartile range 4-41) cells/mL of blood,respectively. A longitudinal analysis of patients (n = 17) after a series of blood collections demonstrated that a change in the CTC count correlated with the patient treatment response in most of the cases and was particularly useful for monitoring patients without elevated serum alpha-fetoprotein (AFP) levels. Sequential CTC enumeration during treatment can supplement standard medical tests and benefit the management of patients with locally advanced or metastatic HCC,in particular for the AFP-low cases. View Publication

The enteric pathogen Lawsonia intracellularis is one of the main causes of diarrhea and compromised weight gain in pigs worldwide. Traditional cell-line cultures have been used to study L. intracellularis pathogenesis. However,these systems fail to reproduce the epithelial changes observed in the intestines of L. intracellularis-infected pigs,specifically,the changes in intestinal cell constitution and gene expression. A more physiologically accurate and state-of-the-art model is provided by swine enteroids derived from stem cell-containing crypts from healthy pigs. The objective of this study was to verify the feasibility of two-dimensional swine enteroids as in vitro models for L. intracellularis infection. We established both three- and two-dimensional swine enteroid cultures derived from intestinal crypts. The two-dimensional swine enteroids were infected by L. intracellularis in four independent experiments. Enteroid-infected samples were collected 3 and 7 d postinfection for analysis using real-time quantitative PCR and L. intracellularis immunohistochemistry. In this study,we show that L. intracellularis is capable of infecting and replicating intracellularly in two-dimensional swine enteroids derived from ileum. View Publication

Short-chain fatty acids (SCFAs) butyrate and propionate are metabolites from dietary fiber's fermentation by gut microbiota that can affect differentiation or functions of T cells,macrophages and dendritic cells. We show here that at low doses these SCFAs directly impact B cell intrinsic functions to moderately enhance class-switch DNA recombination (CSR),while decreasing at higher doses over a broad physiological range,AID and Blimp1 expression,CSR,somatic hypermutation and plasma cell differentiation. In human and mouse B cells,butyrate and propionate decrease B cell Aicda and Prdm1 by upregulating select miRNAs that target Aicda and Prdm1 mRNA-3'UTRs through inhibition of histone deacetylation (HDAC) of those miRNA host genes. By acting as HDAC inhibitors,not as energy substrates or through GPR-engagement signaling in these B cell-intrinsic processes,these SCFAs impair intestinal and systemic T-dependent and T-independent antibody responses. Their epigenetic impact on B cells extends to inhibition of autoantibody production and autoimmunity in mouse lupus models. View Publication

Rabies is a zoonotic neurological infection caused by lyssavirus that continues to result in devastating loss of human life. Many aspects of rabies pathogenesis in human neurons are not well understood. Lack of appropriate ex-vivo models for studying rabies infection in human neurons has contributed to this knowledge gap. In this study,we utilize advances in stem cell technology to characterize rabies infection in human stem cell-derived neurons. We show key cellular features of rabies infection in our human neural cultures,including upregulation of inflammatory chemokines,lack of neuronal apoptosis,and axonal transmission of viruses in neuronal networks. In addition,we highlight specific differences in cellular pathogenesis between laboratory-adapted and field strain lyssavirus. This study therefore defines the first stem cell-derived ex-vivo model system to study rabies pathogenesis in human neurons. This new model system demonstrates the potential for enabling an increased understanding of molecular mechanisms in human rabies,which could lead to improved control methods. View Publication

Patient-derived organoids (PDOs) have recently emerged as robust preclinical models; however,their potential to predict clinical outcomes in patients has remained unclear. We report on a living biobank of PDOs from metastatic,heavily pretreated colorectal and gastroesophageal cancer patients recruited in phase 1/2 clinical trials. Phenotypic and genotypic profiling of PDOs showed a high degree of similarity to the original patient tumors. Molecular profiling of tumor organoids was matched to drug-screening results,suggesting that PDOs could complement existing approaches in defining cancer vulnerabilities and improving treatment responses.We compared responses to anticancer agents ex vivo in organoids and PDO-based orthotopic mouse tumor xenograft models with the responses of the patients in clinical trials. Our data suggest that PDOs can recapitulate patient responses in the clinic and could be implemented in personalized medicine programs. View Publication

PURPOSE Imatinib,the breakpoint cluster region protein (BCR)/Abelson murine leukemia viral oncogene homolog (ABL) inhibitor,is widely used to treat chronic myeloid leukemia (CML). However,imatinib resistance develops in many patients. Therefore,new drugs with improved therapeutic effects are urgently needed. Berberine (BBR) is a potent BCR-ABL inhibitor for imatinib-sensitive and -resistant CML. EXPERIMENTAL DESIGN Protein structure analysis and virtual screening were used to identify BBR targets in CML. Molecular docking analysis,surface plasmon resonance imaging,nuclear magnetic resonance assays,and thermoshift assays were performed to confirm the BBR target. The change in BCR-ABL protein expression after BBR treatment was assessed by Western blotting. The effects of BBR were assessed in vitro in cell lines,in vivo in mice,and in human CML bone marrow cells as a potential strategy to overcome imatinib resistance. RESULTS We discovered that BBR bound to the protein tyrosine kinase domain of BCR-ABL. BBR inhibited the activity of BCR-ABL and BCR-ABL with the T315I mutation,and it also degraded these proteins via the autophagic lysosome pathway by recruiting E3 ubiquitin-protein ligase LRSAM1. BBR inhibited the cell viability and colony formation of CML cells and prolonged survival in CML mouse models with imatinib sensitivity and resistance. CONCLUSIONS The results show that BBR directly binds to and degrades BCR-ABL and BCR-ABL T315I via the autophagic lysosome pathway by recruiting LRSAM1. The use of BBR is a new strategy to improve the treatment of patients with CML with imatinib sensitivity or resistance. View Publication

We recently demonstrated that the sympathetic nervous system can be voluntarily activated following a training program consisting of cold exposure,breathing exercises,and meditation. This resulted in profound attenuation of the systemic inflammatory response elicited by lipopolysaccharide (LPS) administration. Herein,we assessed whether this training program affects the plasma metabolome and if these changes are linked to the immunomodulatory effects observed. A total of 224 metabolites were identified in plasma obtained from 24 healthy male volunteers at six timepoints,of which 98 were significantly altered following LPS administration. Effects of the training program were most prominent shortly after initiation of the acquired breathing exercises but prior to LPS administration,and point towards increased activation of the Cori cycle. Elevated concentrations of lactate and pyruvate in trained individuals correlated with enhanced levels of anti-inflammatory interleukin (IL)-10. In vitro validation experiments revealed that co-incubation with lactate and pyruvate enhances IL-10 production and attenuates the release of pro-inflammatory IL-1$\beta$ and IL-6 by LPS-stimulated leukocytes. Our results demonstrate that practicing the breathing exercises acquired during the training program results in increased activity of the Cori cycle. Furthermore,this work uncovers an important role of lactate and pyruvate in the anti-inflammatory phenotype observed in trained subjects. View Publication

As SARS-CoV-2 infections and death counts continue to rise,it remains unclear why some individuals recover from infection,whereas others rapidly progress and die. Although the immunological mechanisms that underlie different clinical trajectories remain poorly defined,pathogen-specific antibodies often point to immunological mechanisms of protection. Here,we profiled SARS-CoV-2-specific humoral responses in a cohort of 22 hospitalized individuals. Despite inter-individual heterogeneity,distinct antibody signatures resolved individuals with different outcomes. Although no differences in SARS-CoV-2-specific IgG levels were observed,spike-specific humoral responses were enriched among convalescent individuals,whereas functional antibody responses to the nucleocapsid were elevated in deceased individuals. Furthermore,this enriched immunodominant spike-specific antibody profile in convalescents was confirmed in a larger validation cohort. These results demonstrate that early antigen-specific and qualitative features of SARS-CoV-2-specific antibodies point to differences in disease trajectory,highlighting the potential importance of functional antigen-specific humoral immunity to guide patient care and vaccine development. View Publication