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

A causative relationship between chronic inflammation and cancer has been postulated for many years,and clinical observations and laboratory experiments support the hypothesis that inflammation contributes to tumor onset and progression. However,the precise mechanisms underlying the relationship are not known. We recently reported that the proinflammatory cytokine,interleukin-1beta,induces the accumulation and retention of myeloid-derived suppressor cells (MDSC),which are commonly found in many patients and experimental animals with cancer and are potent suppressors of adaptive and innate immunity. This finding led us to hypothesize that inflammation leads to cancer through the induction of MDSC,which inhibit immunosurveillance and thereby allow the unchecked persistence and proliferation of premalignant and malignant cells. We now report that host MDSC have receptors for prostaglandin E2 (PGE2) and that E-prostanoid receptor agonists,including PGE2,induce the differentiation of Gr1(+)CD11b(+) MDSC from bone marrow stem cells,whereas receptor antagonists block differentiation. BALB/c EP2 knockout mice inoculated with the spontaneously metastatic BALB/c-derived 4T1 mammary carcinoma have delayed tumor growth and reduced numbers of MDSC relative to wild-type mice,suggesting that PGE2 partially mediates MDSC induction through the EP2 receptor. Treatment of 4T1-tumor-bearing wild-type mice with the cyclooxygenase 2 inhibitor,SC58236,delays primary tumor growth and reduces MDSC accumulation,further showing that PGE2 induces MDSC and providing a therapeutic approach for reducing this tumor-promoting cell population. View Publication

Recent success in pancreatic islet transplantation has energized the field to discover an alternative source of stem cells with differentiation potential to beta cells. Generation of glucose-responsive,insulin-producing beta cells from self-renewing,pluripotent human ESCs (hESCs) has immense potential for diabetes treatment. We report here the development of a novel serum-free protocol to generate insulin-producing islet-like clusters (ILCs) from hESCs grown under feeder-free conditions. In this 36-day protocol,hESCs were treated with sodium butyrate and activin A to generate definitive endoderm coexpressing CXCR4 and Sox17,and CXCR4 and Foxa2. The endoderm population was then converted into cellular aggregates and further differentiated to Pdx1-expressing pancreatic endoderm in the presence of epidermal growth factor,basic fibroblast growth factor,and noggin. Soon thereafter,expression of Ptf1a and Ngn3 was detected,indicative of further pancreatic differentiation. The aggregates were finally matured in the presence of insulin-like growth factor II and nicotinamide. The temporal pattern of pancreas-specific gene expression in the hESC-derived ILCs showed considerable similarity to in vivo pancreas development,and the final population contained representatives of the ductal,exocrine,and endocrine pancreas. The hESC-derived ILCs contained 2%-8% human C-peptide-positive cells,as well as glucagon- and somatostatin-positive cells. Insulin content as high as 70 ng of insulin/mug of DNA was measured in the ILCs,representing levels higher than that of human fetal islets. In addition,the hESC-derived ILCs contained numerous secretory granules,as determined by electron microscopy,and secreted human C-peptide in a glucose-dependent manner. Disclosure of potential conflicts of interest is found at the end of this article. View Publication

Human stem cells are powerful tools by which to investigate molecular mechanisms of cell growth and differentiation under normal and pathological conditions. Hedgehog signaling,the dysregulation of which causes several pathologies,such as congenital defects and cancer,is involved in several cell differentiation processes and interferes with adipocyte differentiation of rodent cells. The present study was aimed at investigating the effect of Hedgehog pathway modulation on adipocyte phenotype using different sources of human mesenchymal cells,such as bone marrow stromal cells and human multipotent adipose-derived stem cells. We bring evidence that Hedgehog signaling decreases during human adipocyte differentiation. Inhibition of this pathway is not sufficient to trigger adipogenesis,but activation of Hedgehog pathway alters adipocyte morphology as well as insulin sensitivity. Analysis of glycerol-3-phosphate dehydrogenase activity and expression of adipocyte marker genes indicate that activation of Hedgehog signaling by purmorphamine impairs adipogenesis. In sharp contrast to reports in rodent cells,the maturation process,but not the early steps of human mesenchymal stem cell differentiation,is affected by Hedgehog activation. Hedgehog interferes with adipocyte differentiation by targeting CCAAT enhancer-binding protein alpha and peroxisome proliferator-activated receptor (PPAR) gamma2 expression,whereas PPARgamma1 level remains unaffected. Although Hedgehog pathway stimulation does not modify the total number of adipocytes,adipogenesis appears dramatically impaired,with reduced lipid accumulation,a decrease in adipocyte-specific markers,and acquisition of an insulin-resistant phenotype. This study indicates that a decrease in Hedgehog signaling is necessary but not sufficient to trigger adipocyte differentiation and unveils a striking difference in the adipocyte differentiation process between rodent and human mesenchymal stem cells. View Publication

The clinical success of stem cell therapy for myocardial repair hinges on a better understanding of cardiac fate mechanisms. We have identified small molecules involved in cardiac fate by screening a chemical library for activators of the signature gene Nkx2.5,using a luciferase knockin bacterial artificial chromosome (BAC) in mouse P19CL6 pluripotent stem cells. We describe a family of sulfonyl-hydrazone (Shz) small molecules that can trigger cardiac mRNA and protein expression in a variety of embryonic and adult stem/progenitor cells,including human mobilized peripheral blood mononuclear cells (M-PBMCs). Small-molecule-enhanced M-PBMCs engrafted into the rat heart in proximity to an experimental injury improved cardiac function better than control cells. Recovery of cardiac function correlated with persistence of viable human cells,expressing human-specific cardiac mRNAs and proteins. Shz small molecules are promising starting points for drugs to promote myocardial repair/regeneration by activating cardiac differentiation in M-PBMCs. View Publication

Hematopoiesis is a carefully controlled process that is regulated by complex networks of transcription factors that are,in part,controlled by signals resulting from ligand binding to cell-surface receptors. To further understand hematopoiesis,we have compared gene expression profiles of human erythroblasts,megakaryocytes,B cells,cytotoxic and helper T cells,natural killer cells,granulocytes,and monocytes using whole genome microarrays. A bioinformatics analysis of these data was performed focusing on transcription factors,immunoglobulin superfamily members,and lineage-specific transcripts. We observed that the numbers of lineage-specific genes varies by 2 orders of magnitude,ranging from 5 for cytotoxic T cells to 878 for granulocytes. In addition,we have identified novel coexpression patterns for key transcription factors involved in hematopoiesis (eg,GATA3-GFI1 and GATA2-KLF1). This study represents the most comprehensive analysis of gene expression in hematopoietic cells to date and has identified genes that play key roles in lineage commitment and cell function. The data,which are freely accessible,will be invaluable for future studies on hematopoiesis and the role of specific genes and will also aid the understanding of the recent genome-wide association studies. View Publication

T-cell genome engineering holds great promise for cell-based therapies for cancer,HIV,primary immune deficiencies,and autoimmune diseases,but genetic manipulation of human T cells has been challenging. Improved tools are needed to efficiently knock out" genes and "knock in" targeted genome modifications to modulate T-cell function and correct disease-associated mutations. CRISPR/Cas9 technology is facilitating genome engineering in many cell types� View Publication

MicroRNA expression profiling in human liver progenitor cells following hepatocytic differentiation identified miR-122 and miR-194 as the microRNAs most strongly upregulated during hepatocytic differentiation of progenitor cells. MiR-194 was also highly upregulated following hepatocytic differentiation of human embryonic stem cells (hESCs). Overexpression of miR-194 in progenitor cells accelerated their differentiation into hepatocytes,as measured by morphological features such as canaliculi and expression of hepatocytic markers. Overexpression of miR-194 in hESCs induced their spontaneous differentiation,a phenotype accompanied with accelerated loss of the pluripotent factors OCT4 and NANOG and decrease in mesoderm marker HAND1 expression. We then identified YAP1 as a direct target of miR-194. Inhibition of YAP1 strongly induced hepatocytic differentiation of progenitor cells and YAP1 overexpression reversed the miR-194-induced hepatocytic differentiation of progenitor cells. In conclusion,we identified miR-194 as a potent inducer of hepatocytic differentiation of progenitor cells and further identified YAP1 as a mediator of miR-194's effects on hepatocytic differentiation and liver progenitor cell fate. Stem Cells 2016;34:1284-1296. View Publication

T cells can be re-directed to kill cancer cells using chimeric antigen receptors (CARs) or T cell receptors (TCRs). This approach,however,is constrained by the rarity of tumor-specific single antigens. Targeting antigens also found on bystander tissues can cause life-threatening adverse effects. A powerful way to enhance ON-target activity of therapeutic T cells is to engineer them to require combinatorial antigens. Here,we engineer a combinatorially activated T cell circuit in which a synthetic Notch receptor for one antigen induces the expression of a CAR for a second antigen. These dual-receptor AND-gate T cells are only armed and activated in the presence of dual antigen tumor cells. These T cells show precise therapeutic discrimination in vivo-sparing single antigen bystander" tumors while efficiently clearing combinatorial antigen "disease" tumors. This type of precision dual-receptor circuit opens the door to immune recognition of a wider range of tumors. VIDEO ABSTRACT." View Publication

Current long term cryopreservation of cell stocks routinely requires the use of liquid nitrogen (LN2),because commonly used cryopreservation media containing cell membrane permeating cryoprotectants are thermally unstable when frozen at higher storage temperatures,e.g. -80 °C. This instability leads to ice recrystallization,causing progressive loss of cell viability over time under the storage conditions provided by most laboratory deep freezers. The dependency on LN2 for cell storage significantly increases operational expense and raises issues related to impaired working efficiency and safety. Here we demonstrate that addition of Ficoll 70 to cryoprotectant solutions significantly improves system thermal stability at the working temperature (˜-80 °C) of laboratory deep freezers. Moreover,a medium comprised of Ficoll 70 and dimethyl sulfoxide (DMSO) in presence or absence of fetal bovine serum (FBS) can provide reliable cryopreservation of various kinds of human and porcine pluripotent stem cells at -80 °C for periods that extend up to at least one year,with the post-thaw viability,plating efficiency,and full retention of pluripotent phenotype comparable to that achieved with LN2 storage. These results illustrate the practicability of a promising long-term cryopreservation method that completely eliminates the need for LN2. View Publication