搜索结果: 'methocult media formulations for mouse hematopoietic cells serum containing'

SummaryA tissue resident-like phenotype in tumor infiltrating T cells can limit systemic anti-tumor immunity. Enhanced systemic anti-tumor immunity is observed in head and neck cancer patients after neoadjuvant PD-L1 immune checkpoint blockade (ICB) and transforming growth factor β (TGF-β) neutralization. Using T cell receptor (TCR) sequencing and functional immunity assays in a syngeneic model of oral cancer,we dissect the relative contribution of these treatments to enhanced systemic immunity. The addition of TGF-β neutralization to ICB resulted in the egress of expanded and exhausted CD8+ tumor infiltrating lymphocytes (TILs) into circulation and greater systemic anti-tumor immunity. This enhanced egress associated with reduced expression of Itgae (CD103) and its upstream regulator Znf683. Circulating CD8+ T cells expressed higher Cxcr3 after treatment,an observation also made in samples from patients treated with dual TGF-β neutralization and ICB. These findings provide the scientific rationale for the use of PD-L1 ICB and TGF-β neutralization in newly diagnosed patients with carcinomas prior to definitive treatment of locoregional disease. Graphical abstract Highlights•TGF-β blockade reduces Znf683 and CD103 in αPDL1-activated TILs•Reduced TIL CD103 expression associates with egress into circulation•The addition of TGF-β blockade to αPDL1 enhances systemic anti-tumor immunity•Circulating CD8+ T cells express greater CXCR3 after dual TGF-β and PDL1 blockade Natural sciences; Biological sciences; Immunology ; Immune response; Systems biology; Cancer systems biology; Cancer View Publication

CD8+ T cells are central to targeting and eliminating cancer cells. Their function is critically supported by type 1 conventional dendritic cells (cDC1s),which both prime antigen‐specific CD8+ T cells in tumour‐draining lymph nodes (tdLNs) and sustain primed CD8+ T cells within tumours. Despite their importance,the spatiotemporal organisation of cDC1s within tumours and their diverse functional roles remain poorly understood. Here,we use scRNAseq and unbiased spatial analysis to construct a detailed map of cDC1 states and distribution within immunogenic mouse tumours during CD8+ T‐cell‐mediated rejection. We reveal two distinct cDC1 activation states characterised by differential expression of genes linked to anti‐tumour immunity,including Cxcl9 and Il12b. Strikingly,Il12b‐expressing cDC1s are CCR7+ and enriched at tumour borders,where they closely associate with stem‐like TCF1+ CD8+ T cells. In contrast,CCR7–Cxcl9‐expressing cDC1s are preferentially found within the tumour parenchyma alongside effector CD8+ T cells. Analysis of a published dataset of human tumours similarly reveals a spatial association between CCR7+ cDC1 and stem‐like TCF1+ CD8+ T cells. These findings uncover a highly spatially coordinated interaction between cDC1s and CD8+ T cells within tumours,shedding light on the intricate cellular dynamics that underpin effective anti‐tumour immunity. Using scRNAseq and spatial analysis,we analyse cDC1 states and spatial distribution in tumours during immune‐mediated rejection. We identify two cDC1 activation states,each occupying different regions and associated with distinct CD8+ T cell populations. This reveals the spatial organisation of cDC1 states that may be key to anti‐tumour immunity. View Publication

Acute myeloid leukaemia (AML) is a haematological malignancy characterised by the accumulation of transformed myeloid progenitors in the bone marrow. Piplartine (PL),also known as piperlongumine,is a pro-oxidant small molecule extracted from peppers that has demonstrated antineoplastic potential in solid tumours and other haematological malignancies. In this work,we explored the potential of PL to treat AML through the use of a combination of cellular and molecular analyses of primary and cultured leukaemia cells in vitro and in vivo. We showed that PL exhibits in vitro cytotoxicity against AML cells,including CD34 + leukaemia-propagating cells,but not healthy haematopoietic progenitors,suggesting anti-leukaemia selectivity. Mechanistically,PL treatment increased reactive oxygen species (ROS) levels and induced ROS-mediated apoptosis in AML cells,which could be prevented by treatment with the antioxidant scavenger N -acetyl-cysteine and the pancaspase inhibitor Z-VAD(OMe)-FMK. PL treatment reduced NFKB1 gene transcription and the level of NF-κB p65 (pS536),which was depleted from the nucleus of AML cells,indicating suppression of NF-κB p65 signalling. Significantly,PL suppressed AML development in a mouse xenograft model,and its combination with current AML treatments (cytarabine,daunorubicin and azacytidine) had synergistic effects,indicating translational therapeutic potential. Taken together,these data position PL as a novel anti-AML candidate drug that can target leukaemia stem/progenitors and is amenable to combinatorial therapeutic strategies. Subject terms: Acute myeloid leukaemia,Cancer stem cells,Pharmacology View Publication

Induced pluripotent stem cell (iPSC)-derived mesenchymal stromal cells (iMSCs) offer a promising alternative to primary mesenchymal stromal cells (MSCs) and their derivatives,particularly extracellular vesicles (EVs),for use in advanced therapy medicinal products. In this study we evaluated the immunomodulatory and regenerative potential of iMSCs as well as iMSC-EVs,alongside primary human umbilical cord-derived mesenchymal stromal cells (hUCMSCs). Our findings demonstrate that iMSCs exhibit comparable abilities to hUCMSCs in regulating lymphocyte proliferation and inducing an anti-inflammatory phenotype in monocytes. We also observed decreased TNFα levels and increased IL-10 induction,indicating a potential mechanism for their immunomodulatory effects. Furthermore,iMSC-EVs also showed effective immunomodulation by inhibiting T cell proliferation and inducing macrophage polarization similar to their parental cells. Additionally,iMSC-EVs exhibited pro-regenerative potential akin to hUCMSC-EVs in in vitro scratch assays. Notably,priming iMSCs with pro-inflammatory cytokines significantly enhanced the immunomodulatory potential of iMSC-EVs. These results underscore the considerable promise of iMSCs and iMSCs-EVs as an alternate source for MSC-derived therapeutics,given their potent immunomodulatory and regenerative properties. The online version contains supplementary material available at 10.1038/s41598-024-75956-3. View Publication

Natural killer (NK) cells are an important innate defense against malignancies,and exogenous sources of NK cells have been developed as anti-cancer agents. Nevertheless,the apparent limitations of NK cells in clearing cancers have suggested that their efficacy might be augmented by combination with other treatments. We have developed cell-penetrating peptides that target the transcription factors ATF5,CEBPB,and CEBPD and that promote apoptotic cancer cell death both in vitro and in vivo without apparent toxicity to non-transformed cells. We report here that one such peptide,Dpep,significantly sensitizes a variety of tumor cell types to the cytotoxic activity of the NK cell line,NK-92MI. Such sensitization requires pre-exposure of tumor cells to Dpep and does not appear due to effects of Dpep on NK cells themselves. Our findings suggest that Dpep acts in this context to lower the apoptotic threshold of tumor cells to NK cell toxicity. Additionally,while Dpep pre-treatment does not prevent tumor cells from causing NK cell “inactivation”,it sensitizes cancer cells to repeated rounds of exposure to fresh NK cells. These findings thus indicate that Dpep pre-treatment is an effective strategy to sensitize cancer cells to the cytotoxic actions of NK cells. View Publication

Acute myeloid leukemia (AML) is an aggressive clonal disorder of hematopoietic stem cells (HSCs) and primitive progenitors that blocks their myeloid differentiation,generating self-renewing leukemic stem cells (LSCs). Here,we show that the mRNA m6A reader YTHDF2 is overexpressed in a broad spectrum of human AML and is required for disease initiation as well as propagation in mouse and human AML. YTHDF2 decreases the half-life of diverse m6A transcripts that contribute to the overall integrity of LSC function,including the tumor necrosis factor receptor Tnfrsf2,whose upregulation in Ythdf2-deficient LSCs primes cells for apoptosis. Intriguingly,YTHDF2 is not essential for normal HSC function,with YTHDF2 deficiency actually enhancing HSC activity. Thus,we identify YTHDF2 as a unique therapeutic target whose inhibition selectively targets LSCs while promoting HSC expansion. View Publication

BACKGROUND Cell-surface mucins are expressed in apical epithelial cells of the respiratory tract,and contribute a crucial part of the innate immune system. Despite anti-inflammatory or antiviral functions being revealed for certain cell-surface mucins such as MUC1,the roles of other mucins are still poorly understood,especially in viral infections. METHODS To further identify mucins significant in influenza infection,we screened the expression of mucins in human nasal epithelial cells infected by H3N2 influenza A virus. RESULTS We found that the expression of MUC15 was significantly upregulated upon infection,and specific only to active infection. While MUC15 did not interact with virus particles or reduce viral replication directly,positive correlations were observed between MUC15 and inflammatory factors in response to viral infection. Given that the upregulation of MUC15 was only triggered late into infection when immune factors (including cytokines,chemokines,EGFR and phosphorylated ERK) started to peak and plateau,MUC15 may potentially serve an immunomodulatory function later during influenza viral infection. CONCLUSIONS Our study revealed that MUC15 was one of the few cell-surface mucins induced during influenza infection. While MUC15 did not interact directly with influenza virus,we showed that its increase coincides with the peak of immune activation and thus MUC15 may serve an immunomodulatory role during influenza infection. View Publication

BACKGROUND Asthma is a heterogenous disease characterized by chronic inflammation and airway remodeling. An increase in the severity of airway remodeling is associated with a more severe form of asthma. There is increasing interest in the epithelial to mesenchymal transition process and mechanisms involved in the differentiation and repair of the airway epithelium,especially as they apply to severe asthma. Growing evidence suggests that Epithelial-Mesenchymal transition (EMT) could contribute to airway remodeling and fibrosis in asthma. Severe asthmatic patients with remodeled airways have a neutrophil driven inflammation. Neutrophils are an important source of TGF-$\beta$1,which plays a role in recruitment and activation of inflammatory cells,extracellular matrix (ECM) production and fibrosis development,and is a potent inducer of EMT. OBJECTIVE As there is little data examining the contribution of neutrophils and/or their mediators to the induction of EMT in airway epithelial cells,the objective of this study was to better understand the potential role of neutrophils in severe asthma in regards to EMT. METHODS We used an in vitro system to investigate the neutrophil-epithelial cell interaction. We obtained peripheral blood neutrophils from severe asthmatic patients and control subjects and examined for their ability to induce EMT in primary airway epithelial cells. RESULTS Our data indicate that neutrophils from severe asthmatic patients induce changes in morphology and EMT marker expression in bronchial epithelial cells consistent with the EMT process when co-cultured. TGF-$\beta$1 levels in the culture medium of severe asthmatic patients were increased compared to that from co-cultures of non-asthmatic neutrophils and epithelial cells. CONCLUSIONS AND CLINICAL RELEVANCE As an inducer of EMT and an important source of TGF-$\beta$1,neutrophils may play a significant role in the development of airway remodeling and fibrosis in severe asthmatic airways. View Publication

The steps to leukemia following an in utero fusion of MLL (HRX,ALL-1) to a partner gene in humans are not known. Introduction of the Mll-AF9 fusion gene into embryonic stem cells results in leukemia in mice with cell-type specificity similar to humans. In this study we used myeloid colony assays,immunophenotyping,and transplantation to evaluate myelopoiesis in Mll-AF9 mice. Colony assays demonstrated that both prenatal and postnatal Mll-AF9 tissues have significantly increased numbers of CD11b(+)/CD117(+)/Gr-1(+/-) myeloid cells,often in compact clusters. The self-renewal capacity of prenatal myeloid progenitors was found to decrease following serial replating of colony-forming cells. In contrast,early postnatal myeloid progenitors increased following replating; however,the enhanced self-renewal of early postnatal myeloid progenitor cells was limited and did not result in long-term cell lines or leukemia in vivo. Unlimited replating,long-term CD11b/Gr-1(+) myeloid cell lines,and the ability to produce early leukemia in vivo in transplantation experiments,were found only in mice with overt leukemia. Prenatal Mll-AF9 tissues had reduced total (mature and progenitor) CD11b/Gr-1(+) cells compared with wild-type tissues. Colony replating,immunophenotyping,and cytochemistry suggest that any perturbation of cellular differentiation from the prenatal stage onward is partial and largely reversible. We describe a novel informative in vitro and in vivo model system that permits study of the stages in the pathogenesis of Mll fusion gene leukemia,beginning in prenatal myeloid cells,progressing to a second stage in the postnatal period and,finally,resulting in overt leukemia in adult animals. View Publication

Murine megakaryocytes (MKs) are defined by CD41/CD61 expression and acetylcholinesterase (AChE) activity; however,their stages of differentiation in bone marrow (BM) have not been fully elucidated. In murine lineage-negative (Lin(-))/CD45(+) BM cells,we found CD41(+) MKs without AChE activity (AChE(-)) except for CD41(++) MKs with AChE activity (AChE(+)),in which CD61 expression was similar to their CD41 level. Lin(-)/CD41(+)/CD45(+)/AChE(-) MKs could differentiate into AChE(+),with an accompanying increase in CD41/CD61 during in vitro culture. Both proplatelet formation (PPF) and platelet (PLT) production for Lin(-)/CD41(+)/CD45(+)/AChE(-) MKs were observed later than for Lin(-)/CD41(++)/CD45(+)/AChE(+) MKs,whereas MK progenitors were scarcely detected in both subpopulations. GeneChip and semiquantitative polymerase chain reaction analyses revealed that the Lin(-)/CD41(+)/CD45(+)/AChE(-) MKs are assigned at the stage between the progenitor and PPF preparation phases in respect to the many MK/PLT-specific gene expressions,including beta1-tubulin. In normal mice,the number of Lin(-)/CD41(+)/CD45(+)/AChE(-) MKs was 100 times higher than that of AChE(+) MKs in BM. When MK destruction and consequent thrombocytopenia were caused by an antitumor agent,mitomycin-C,Lin(-)/CD41(+)/CD45(+)/AChE(-) MKs led to an increase in AChE(+) MKs and subsequent PLT recovery with interleukin-11 administration. It was concluded that MKs in murine BM at least in part consist of immature Lin(-)/CD41(+)/CD45(+)/AChE(-) MKs and more differentiated Lin(-)/CD41(++)/CD45(+)/AChE(+) MKs. Immature Lin(-)/CD41(+)/CD45(+)/AChE(-) MKs are a major MK population compared with AChE(+) MKs in BM and play an important role in rapid PLT recovery in vivo. View Publication

BACKGROUND: Human embryonic stem cells (hESC) provide a unique model to study early events in human development. The hESC-derived cells can potentially be used to replace or restore different tissues including neuronal that have been damaged by disease or injury.backslashnbackslashnMETHODOLOGY AND PRINCIPAL FINDINGS: The cells of two different hESC lines were converted to neural rosettes using adherent and chemically defined conditions. The progenitor cells were exposed to retinoic acid (RA) or to human recombinant basic fibroblast growth factor (bFGF) in the late phase of the rosette formation. Exposing the progenitor cells to RA suppressed differentiation to rostral forebrain dopamine neural lineage and promoted that of spinal neural tissue including motor neurons. The functional characteristics of these differentiated neuronal precursors under both,rostral (bFGF) and caudalizing (RA) signals were confirmed by patch clamp analysis.backslashnbackslashnCONCLUSIONS/SIGNIFICANCE: These findings suggest that our differentiation protocol has the capacity to generate region-specific and electrophysiologically active neurons under in vitro conditions without embryoid body formation,co-culture with stromal cells and without presence of cells of mesodermal or endodermal lineages. View Publication