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

The interaction between HER2 and aryl hydrocarbon receptor (AhR) signaling cascades during mammosphere formation of MCF-7 cells was studied. A newly established clonal MCF-7 cell line (HER2-5),stably overexpressing HER2,showed significantly enhanced levels of AhR mRNA and protein compared with MCF-7 cells. AhR was required for the HER2-mediated induction of interleukin-6 mRNA and for mammosphere formation in HER2-5 and MCF-7 cells. Mammosphere forming efficiency was suppressed by an AhR antagonist in a dose-dependent manner,as well as by knockdown of AhR. Taken together,these results indicate that AhR enhances mammosphere formation by human HER2-overexpressing breast cancer cells. View Publication

microRNAs (miRNAs) are small,non-coding RNAs that regulate expression of multiple genes. MiR-193a-3p functions as a tumor suppressor in many cancer types,but its effect on inducing specific anti-tumor immune responses is unclear. Therefore,we examined the effect of our lipid nanoparticle (LNP) formulated,chemically modified,synthetic miR-193a-3p mimic (INT-1B3) on anti-tumor immunity. INT-1B3 inhibited distant tumor metastasis and significantly prolonged survival. INT-1B3-treated animals were fully protected against challenge with autologous tumor cells even in absence of treatment indicating long-term immunization. Protection against autologous tumor cell challenge was hampered upon T cell depletion and adoptive T cell transfer abrogated tumor growth. Transfection of tumor cells with our miR-193a-3p mimic (1B3) resulted in tumor cell death and apoptosis accompanied by increased expression of DAMPs. Co-culture of 1B3-transfected tumor cells and immature DC led to DC maturation and these mature DC were able to stimulate production of type 1 cytokines by CD4+ and CD8+ T cells. CD4-CD8- T cells also produced type 1 cytokines,even in response to 1B3-transfected tumor cells directly. Live cell imaging demonstrated PBMC-mediated cytotoxicity against 1B3-transfected tumor cells. These data demonstrate for the first time that miR-193a-3p induces long-term immunity against tumor development via modulation of the tumor microenvironment and induction of immunogenic cell death. View Publication

Dishevelled-associated activator of morphogenesis1 (DAAM1) is a member of the evolutionarily conserved Formin family and plays a significant role in the malignant progression of various human cancers. This study aims to explore the clinical and biological significance of DAAM1 in pancreatic cancer. Multiple public datasets and an in-house cohort were utilized to assess the clinical relevance of DAAM1 in pancreatic cancer. The LinkedOmics platform was employed to perform enrichment analysis of DAAM1-associated molecular pathways in pancreatic cancer. Subsequently,a series of in vitro and in vivo experiments were conducted to evaluate the biological roles of DAAM1 in pancreatic cancer cells and its effects on intratumoral T cells. DAAM1 was found to be upregulated in pancreatic cancer tissues,with higher expression levels observed in tumor cells. Additionally,high expression of DAAM1 was associated with poor prognosis. DAAM1 acted as an oncogene in pancreatic cancer,and its inhibition suppressed tumor cell proliferation,migration,and invasion,while promoted apoptosis. Furthermore,DAAM1 was involved in the JAK1/STAT1 signaling pathway and regulated PD-L1 expression in pancreatic cancer cells. The inhibition of DAAM1 also significantly reduced the exhaustion levels of CD8+ T cells. In conclusion,DAAM1 functions as an oncogene and is immunologically implicated in pancreatic cancer,these findings suggest that DAAM1 may serve as a promising therapeutic target for the clinical management of pancreatic cancer. The online version contains supplementary material available at 10.1186/s12935-024-03631-8. View Publication

Chronic myeloid leukemia is a clonal multilineage myeloproliferative disease of stem cell origin characterized by the presence of the Bcr/Abl oncoprotein,a constitutively active tyrosine kinase. In previous studies,we have provided evidence that Bcr/Abl overexpression in leukemic cells increased their susceptibility to NK-mediated lysis by different mechanisms. In the present study,using UT-7/9 cells,a high level Bcr/Abl transfectant of UT-7 cells,we show that the treatment of Bcr/Abl target by imatinib mesylate (IM),a specific Abl tyrosine kinase inhibitor,hampers the formation of the NK/target immunological synapse. The main effect of IM involves an induction of surface GM1 ganglioside on Bcr/Abl transfectants that prevents the redistribution of MHC-related Ag molecules in lipid rafts upon interaction with NK cells. IM also affects cell surface glycosylation of targets,as assessed by binding of specific lectins resulting in the subsequent modulation of their binding to lectin type NK receptor,particularly NKG2D. In addition,we demonstrate that the tyrosine kinase activity repression results in a decrease of MHC-related Ags-A/B and UL-16-binding protein expression on Bcr/Abl transfectants UT-7/9. We show that NKG2D controls the NK-mediated lysis of UT-7/9 cells,and IM treatment inhibits this activating pathway. Taken together,our results show that the high expression of Bcr/Abl in leukemic cells controls the expression of NKG2D receptor ligands and membrane GM1 via a tyrosine kinase-dependent mechanism and that the modulation of these molecules by IM interferes with NK cell recognition and cytolysis of the transfectants. View Publication

The human WDR33 gene encodes three major isoforms. The canonical isoform WDR33v1 (V1) is a well-characterized nuclear mRNA polyadenylation factor,while the other two,WDR33v2 (V2) and WDR33v3 (V3),have not been studied. Here,we report that V2 and V3 are generated by alternative polyadenylation,and neither protein contains all seven WD (tryptophan-aspartic acid) repeats that characterize V1. Surprisingly,V2 and V3 are not polyadenylation factors but localize to the endoplasmic reticulum and interact with stimulator of interferon genes (STING),the immune factor that induces the cellular response to cytosolic double-stranded DNA. V2 suppresses interferon-β induction by preventing STING disulfide oligomerization but promotes autophagy,likely by recruiting WIPI2 isoforms. V3,on the other hand,functions to ncrease STING protein levels. Our study has not only provided mechanistic insights into STING regulation but also revealed that protein isoforms can be functionally completely unrelated,indicating that alternative mRNA processing is a more powerful mechanism than previously appreciated. View Publication

A mysterious feature of Crohn's disease (CD) is the extra-intestinal manifestation of creeping fat" (CrF) defined as expansion of mesenteric adipose tissue around the inflamed and fibrotic intestine. In the current study we explore whether microbial translocation in CD serves as a central cue for CrF development. We discovered a subset of mucosal-associated gut bacteria that consistently translocated and remained viable in CrF in CD ileal surgical resections and identified Clostridium innocuum as a signature of this consortium with strain variation between mucosal and adipose isolates suggesting preference for lipid-rich environments. Single-cell RNA sequencing characterized CrF as both pro-fibrotic and pro-adipogenic with a rich milieu of activated immune cells responding to microbial stimuli which we confirm in gnotobiotic mice colonized with C. innocuum. Ex vivo validation of expression patterns suggests C. innocuum stimulates tissue remodeling via M2 macrophages leading to an adipose tissue barrier that serves to prevent systemic dissemination of bacteria." View Publication

The cell-surface straight and branched repeats of N-acetyllactosamine (LacNAc) units,called poly-LacNAc chains,characterize the histo-blood group i and I antigens,respectively. The transition of straight to branched poly-LacNAc chain (i to I) is determined by the I locus,which expresses 3 IGnT transcripts,IGnTA,IGnTB,and IGnTC. Our previous investigation demonstrated that the i-to-I transition in erythroid differentiation is regulated by the transcription factor CCAAT/enhancer binding protein alpha (C/EBPalpha). In the present investigation,the K-562 cell line was used as a model to show that the i-to-I transition is determined by the phosphorylation status of the C/EBPalpha Ser-21 residue,with dephosphorylated C/EBPalpha Ser-21 stimulating the transcription of the IGnTC gene,consequently resulting in I branching. Results from studies using adult erythropoietic and granulopoietic progenitor cells agreed with those derived using the K-562 cell model,with lentiviral expression of C/EBPalpha in CD34(+) hematopoietic cells demonstrating that the dephosphorylated form of C/EBPalpha Ser-21 induced the expression of I antigen,granulocytic CD15,and also erythroid CD71 antigens. Taken together,these results demonstrate that the regulation of poly-LacNAc branching (I antigen) formation in erythropoiesis and granulopoiesis share a common mechanism,with dephosphorylation of the Ser-21 residue on C/EBPalpha playing the critical role. View Publication

The Peutz-Jegher syndrome tumor-suppressor gene encodes a protein-threonine kinase,LKB1,which phosphorylates and activates AMPK [adenosine monophosphate (AMP)-activated protein kinase]. The deletion of LKB1 in the liver of adult mice resulted in a nearly complete loss of AMPK activity. Loss of LKB1 function resulted in hyperglycemia with increased gluconeogenic and lipogenic gene expression. In LKB1-deficient livers,TORC2,a transcriptional coactivator of CREB (cAMP response element-binding protein),was dephosphorylated and entered the nucleus,driving the expression of peroxisome proliferator-activated receptor-gamma coactivator 1alpha (PGC-1alpha),which in turn drives gluconeogenesis. Adenoviral small hairpin RNA (shRNA) for TORC2 reduced PGC-1alpha expression and normalized blood glucose levels in mice with deleted liver LKB1,indicating that TORC2 is a critical target of LKB1/AMPK signals in the regulation of gluconeogenesis. Finally,we show that metformin,one of the most widely prescribed type 2 diabetes therapeutics,requires LKB1 in the liver to lower blood glucose levels. View Publication

In this study we dissected retinal organoid morphogenesis in human embryonic stem cell (hESC)-derived cultures and established a convenient method for isolating large quantities of retinal organoids for modeling human retinal development and disease. Epithelialized cysts were generated via floating culture of clumps of Matrigel/hESCs. Upon spontaneous attachment and spreading of the cysts,patterned retinal monolayers with tight junctions formed. Dispase-mediated detachment of the monolayers and subsequent floating culture led to self-formation of retinal organoids comprising patterned neuroretina,ciliary margin,and retinal pigment epithelium. Intercellular adhesion-dependent cell survival and ROCK-regulated actomyosin-driven forces are required for the self-organization. Our data supports a hypothesis that newly specified neuroretina progenitors form characteristic structures in equilibrium through minimization of cell surface tension. In long-term culture,the retinal organoids autonomously generated stratified retinal tissues,including photoreceptors with ultrastructure of outer segments. Our system requires minimal manual manipulation,has been validated in two lines of human pluripotent stem cells,and provides insight into optic cup invagination in vivo. View Publication

Genome-wide association studies (GWASs) have reported many single nucleotide polymorphisms (SNPs) associated with psychiatric disorders,but knowledge is lacking regarding molecular mechanisms. Here we show that risk alleles spanning multiple genes across the 10q24.32 schizophrenia-related locus are associated in the human brain selectively with an increase in the expression of both BLOC-1 related complex subunit 7 (BORCS7) and a previously uncharacterized,human-specific arsenite methyltransferase (AS3MT) isoform (AS3MT(d2d3)),which lacks arsenite methyltransferase activity and is more abundant in individuals with schizophrenia than in controls. Conditional-expression analysis suggests that BORCS7 and AS3MT(d2d3) signals are largely independent. GWAS risk SNPs across this region are linked with a variable number tandem repeat (VNTR) polymorphism in the first exon of AS3MT that is associated with the expression of AS3MT(d2d3) in samples from both Caucasians and African Americans. The VNTR genotype predicts promoter activity in luciferase assays,as well as DNA methylation within the AS3MT gene. Both AS3MT(d2d3) and BORCS7 are expressed in adult human neurons and astrocytes,and they are upregulated during human stem cell differentiation toward neuronal fates. Our results provide a molecular explanation for the prominent 10q24.32 locus association,including a novel and evolutionarily recent protein that is involved in early brain development and confers risk for psychiatric illness. View Publication

Reaction with peroxynitrite at pH 7.4 and 37 degrees C was found to increase the 8-oxodeoxyguanosine levels in calf thymus DNA 35- 38-fold. This oxidation of deoxyguanosine,as well as the peroxynitrite-mediated nitration of tyrosine to 3-nitrotyrosine,was significantly inhibited by ascorbic acid,glutathione and (-)-epigallocatechin gallate,a polyphenolic antioxidant present in tea. For 50% inhibition of the oxidation of deoxyguanosine to 8-oxodeoxyguanosine,1.1,7.6 or 0.25 mM ascorbate,glutathione or (-)-epigallocatechin gallate,respectively,was required. For 50% inhibition of tyrosine nitration,the respective concentrations were 1.4,4.6 or 0.11 mM. Thus,(-)-epigallocatechin gallate is a significantly better inhibitor of both reactions than either ascorbate or glutathione. Reaction of (-)-epigallocatechin gallate with peroxynitrite alone resulted in the formation of a number of products. Ultraviolet spectra of two of these suggest that the tea polyphenol and/or its oxidation products are nitrated by peroxynitrite. View Publication

Protein modification by small ubiquitin-related modifier proteins (SUMOs) controls diverse cellular functions. Dysregulation of SUMOylation or deSUMOylation processes has been implicated in the development of cancer and neurodegenerative diseases. However,no small-molecule inhibiting protein SUMOylation has been reported so far. Here,we report inhibition of SUMOylation by ginkgolic acid and its analog,anacardic acid. Ginkgolic acid and anacardic acid inhibit protein SUMOylation both in vitro and in vivo without affecting in vivo ubiquitination. Binding assays with a fluorescently labeled probe showed that ginkgolic acid directly binds E1 and inhibits the formation of the E1-SUMO intermediate. These studies will provide not only a useful tool for investigating the roles of SUMO conjugations in a variety of pathways in cells,but also a basis for the development of drugs targeted against diseases involving aberrant SUMOylation. View Publication