搜索结果: 'gmp t cell activator'

Increased levels of intestinal bile acids (BAs) are a risk factor for colorectal cancer (CRC). Here,we show that the convergence of dietary factors (high-fat diet) and dysregulated WNT signaling (APC mutation) alters BA profiles to drive malignant transformations in Lgr5-expressing (Lgr5+) cancer stem cells and promote an adenoma-to-adenocarcinoma progression. Mechanistically,we show that BAs that antagonize intestinal farnesoid X receptor (FXR) function,including tauro-$\beta$-muricholic acid (T-$\beta$MCA) and deoxycholic acid (DCA),induce proliferation and DNA damage in Lgr5+ cells. Conversely,selective activation of intestinal FXR can restrict abnormal Lgr5+ cell growth and curtail CRC progression. This unexpected role for FXR in coordinating intestinal self-renewal with BA levels implicates FXR as a potential therapeutic target for CRC. View Publication

The cancer stem cell (CSC) hypothesis implicates the development of new therapeutic approaches to target the CSC population. Characterization of the pathways that regulate CSCs activity will facilitate the development of targeted therapies. We recently reported that the enzymatic activity of ALDH1,as measured by the ALDELFUOR assay,can be utilized to isolate normal and malignant breast stem cells in both primary tumors and cell lines. In this study,utilizing a tumorsphere assay,we have demonstrated the role of retinoid signaling in the regulation of breast CSCs self-renewal and differentiation. Utilizing the gene set enrichment analysis (GSEA) algorithm we identified gene sets and pathways associated with retinoid signaling. These pathways regulate breast CSCs biology and their inhibition may provide novel therapeutic approaches to target breast CSCs. View Publication

In an attempt to better understand and control the processes that regulate stem cell fate,we have set out to identify small molecules that induce neuronal differentiation in embryonic stem cells (ESCs). A high-throughput phenotypic cell-based screen of kinase-directed combinatorial libraries led to the discovery of TWS119,a 4,6-disubstituted pyrrolopyrimidine that can induce neurogenesis in murine ESCs. The target of TWS119 was shown to be glycogen synthase kinase-3beta (GSK-3beta) by both affinity-based and biochemical methods. This study provides evidence that GSK-3beta is involved in the induction of mammalian neurogenesis in ESCs. This and such other molecules are likely to provide insights into the molecular mechanisms that control stem cell fate,and may ultimately be useful to in vivo stem cell biology and therapy. View Publication

Most studies of cancer stem cells (CSC) involve the inoculation of cells from human tumors into immunosuppressed mice,preventing an assessment on the immunologic interactions and effects of CSCs. In this study,we examined the vaccination effects produced by CSC-enriched populations from histologically distinct murine tumors after their inoculation into different syngeneic immunocompetent hosts. Enriched CSCs were immunogenic and more effective as an antigen source than unselected tumor cells in inducing protective antitumor immunity. Immune sera from CSC-vaccinated hosts contained high levels of IgG which bound to CSCs,resulting in CSC lysis in the presence of complement. CTLs generated from peripheral blood mononuclear cells or splenocytes harvested from CSC-vaccinated hosts were capable of killing CSCs in vitro. Mechanistic investigations established that CSC-primed antibodies and T cells were capable of selective targeting CSCs and conferring antitumor immunity. Together,these proof-of-concept results provide a rationale for a new type of cancer immunotherapy based on the development of CSC vaccines that can specifically target CSCs. View Publication

Understanding how hematopoietic stem cells (HSCs) are generated and the signals that control this process is a crucial issue for regenerative medicine applications that require in vitro production of HSC. HSCs emerge during embryonic life from an endothelial-like cell population that resides in the aorta-gonad-mesonephros (AGM) region. We show here that β-catenin is nuclear and active in few endothelial nonhematopoietic cells closely associated with the emerging hematopoietic clusters of the embryonic aorta during mouse development. Importantly,Wnt/β-catenin activity is transiently required in the AGM to generate long-term HSCs and to produce hematopoietic cells in vitro from AGM endothelial precursors. Genetic deletion of β-catenin from the embryonic endothelium stage (using VE-cadherin-Cre recombinase),but not from embryonic hematopoietic cells (using Vav1-Cre),precludes progression of mutant cells toward the hematopoietic lineage; however,these mutant cells still contribute to the adult endothelium. Together,those findings indicate that Wnt/β-catenin activity is needed for the emergence but not the maintenance of HSCs in mouse embryos. View Publication

Since the 1960s,the stem cells have been extensively studied including embryonic stem cells,neural stem cells,bone marrow hematopoietic stem cells,and mesenchymal stem cells. In the recent years,several stem cells have been initially used in the treatment of diseases,such as in bone marrow transplant. At the same time,isolation and culture experimental technologies for stem cell research have been widely developed in recent years. In addition,molecular imaging technologies including optical molecular imaging,positron emission tomography,single-photon emission computed tomography,and computed tomography have been developed rapidly in recent the 10 years and have also been used in the research on disease mechanism and evaluation of treatment of disease related with stem cells. This paper will focus on recent typical isolation,culture,and observation techniques of stem cells followed by a concise introduction. Finally,the current challenges and the future applications of the new technologies in stem cells are given according to the understanding of the authors,and the paper is then concluded. View Publication

The adaptation of the clustered regularly interspaced short palindromic repeats (CRISPR) and CRISPR associated endonuclease 9 (CRISPR-Cas9) machinery from prokaryotic organisms has resulted in a gene editing system that is highly versatile,easily constructed,and can be leveraged to generate human cells knocked out (KO) for a specific gene. While standard transfection techniques can be used for the introduction of CRISPR-Cas9 expression cassettes to many cell types,delivery by this method is not efficient in many primary cell types,including primary human airway epithelial cells (AECs). More efficient delivery in AECs can be achieved through lentiviral-mediated transduction,allowing the CRISPR-Cas9 system to be integrated into the genome of the cell,resulting in stable expression of the nuclease machinery and increasing editing rates. In parallel,advancements have been made in the culture,expansion,selection,and differentiation of AECs,which allow the robust generation of a bulk edited AEC population from transduced cells. Applying these methods,we detail here our latest protocol to generate mucociliary epithelial cultures knocked out for a specific gene from donor-isolated primary human basal airway epithelial cells. This protocol includes methods to: (1) design and generate lentivirus which targets a specific gene for KO with CRISPR-Cas9 machinery,(2) efficiently transduce AECs,(3) culture and select for a bulk edited AEC population,(4) molecularly screen AECs for Cas9 cutting and specific sequence edits,and (5) further expand and differentiate edited cells to a mucociliary airway epithelial culture. The AEC knockouts generated using this protocol provide an excellent primary cell model system with which to characterize the function of genes involved in airway dysfunction and disease. View Publication

TOX is a DNA-binding factor required for development of CD4(+) T cells,natural killer T cells and regulatory T cells. Here we document that both natural killer (NK) cell development and lymphoid tissue organogenesis were also inhibited in the absence of TOX. We found that the development of lymphoid tissue-inducer cells,a rare subset of specialized cells that has an integral role in lymphoid tissue organogenesis,required TOX. Tox was upregulated considerably in immature NK cells in the bone marrow,consistent with the loss of mature NK cells in the absence of this nuclear protein. Thus,many cell lineages of the immune system share a TOX-dependent step for development. View Publication

Blood consists of different cell populations with distinct functions and correspondingly,distinct gene expression profiles. In this study,global miRNA expression profiling was performed across a panel of nine human immune cell subsets (neutrophils,eosinophils,monocytes,B cells,NK cells,CD4 T cells,CD8 T cells,mDCs and pDCs) to identify cell-type specific miRNAs. mRNA expression profiling was performed on the same samples to determine if miRNAs specific to certain cell types down-regulated expression levels of their target genes. Six cell-type specific miRNAs (miR-143; neutrophil specific,miR-125; T cells and neutrophil specific,miR-500; monocyte and pDC specific,miR-150; lymphoid cell specific,miR-652 and miR-223; both myeloid cell specific) were negatively correlated with expression of their predicted target genes. These results were further validated using an independent cohort where similar immune cell subsets were isolated and profiled for both miRNA and mRNA expression. miRNAs which negatively correlated with target gene expression in both cohorts were identified as candidates for miRNA/mRNA regulatory pairs and were used to construct a cell-type specific regulatory network. miRNA/mRNA pairs formed two distinct clusters in the network corresponding to myeloid (nine miRNAs) and lymphoid lineages (two miRNAs). Several myeloid specific miRNAs targeted common genes including ABL2,EIF4A2,EPC1 and INO80D; these common targets were enriched for genes involved in the regulation of gene expression (ptextless9.0E-7). Those miRNA might therefore have significant further effect on gene expression by repressing the expression of genes involved in transcriptional regulation. The miRNA and mRNA expression profiles reported in this study form a comprehensive transcriptome database of various human blood cells and serve as a valuable resource for elucidating the role of miRNA mediated regulation in the establishment of immune cell identity. View Publication

Human blastocyst-derived,pluripotent cell lines are described that have normal karyotypes,express high levels of telomerase activity,and express cell surface markers that characterize primate embryonic stem cells but do not characterize other early lineages. After undifferentiated proliferation in vitro for 4 to 5 months,these cells still maintained the developmental potential to form trophoblast and derivatives of all three embryonic germ layers,including gut epithelium (endoderm); cartilage,bone,smooth muscle,and striated muscle (mesoderm); and neural epithelium,embryonic ganglia,and stratified squamous epithelium (ectoderm). These cell lines should be useful in human developmental biology,drug discovery,and transplantation medicine. View Publication

Human embryonic stem cells (hESCs) have numerous potential biomedical applications owing to their unique abilities for self-renewal and pluripotency. Successful clinical application of hESCs and derivatives necessitates the culture of these cells in a fully defined environment. We have developed a novel peptide-based surface that uses a high-affinity cyclic RGD peptide for culture of hESCs under chemically defined conditions. View Publication

Hematopoietic stem cell (HSC) quiescence is a tightly regulated process crucial for hematopoietic regeneration,which requires a healthy and supportive microenvironmental niche within the bone marrow (BM). Here,we show that deletion of Ptpn21,a protein tyrosine phosphatase highly expressed in HSCs,induces stem cell egress from the niche due to impaired retention within the BM. Ptpn21-/- HSCs exhibit enhanced mobility,decreased quiescence,increased apoptosis,and defective reconstitution capacity. Ptpn21 deletion also decreased HSC stiffness and increased physical deformability,in part by dephosphorylating Spetin1 (Tyr246),a poorly described component of the cytoskeleton. Elevated phosphorylation of Spetin1 in Ptpn21-/- cells impaired cytoskeletal remodeling,contributed to cortical instability,and decreased cell rigidity. Collectively,these findings show that Ptpn21 maintains cellular mechanics,which is correlated with its important functions in HSC niche retention and preservation of hematopoietic regeneration capacity. View Publication