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

Both anti-tumoral and pro-tumoral effects of mesenchymal stem cells (MSCs) in preclinical treatment of ovarian cancer have been controversially demonstrated. In this study,we profiled the phenotypes of mouse compact bone-derived MSCs (CB-MSCs) and bone marrow-derived MSCs (BM-MSCs) and found that CB-MSCs expressed lower CD90 compared to BM-MSCs. We examined gene expression of immune regulating cytokines of CB-MSCs in 2D and 3D culture and under stimulation with TLR4 agonist LPS or immune activator VIC-008. Our data showed that when CB-MSCs were cultured in simulated in vivo 3D condition,CD90 expression was further decreased. Moreover,gene expressions of immune activating cytokines IL-12,IL-21,IFNgamma and a pro-inflammatory cytokine CXCL10 in CB-MSCs were increased in 3D culture whereas gene expression of anti-inflammatory cytokines IL-10 and CCL5 were downregulated. Stimulation of CB-MSCs by LPS or VIC-008 presented similar profile of the cytokine gene expressions to that in 3D culture which might benefit the anti-tumor efficacy of CD90low MSCs. The anti-tumor effects of CD90low CB-MSCs alone or in combination with VIC-008 were evaluated in a syngeneic orthotopic mouse model of ovarian cancer. Treatment that combines CB-MSCs and VIC-008 significantly decreased tumor growth and prolonged mouse survival. This was associated with the increase of activated anti-tumoral CD4+ and CD8+ T cells and the decrease of Treg cells in the tumor microenvironment. Taken together,our study demonstrates the synergistic anti-tumoral efficacy by application of CB-MSCs combined with immune activator VIC-008 and provides new insight into CD90low MSCs as a new anti-tumor arsenal. View Publication

BackgroundThe epoxyeicosatrienoic acids (EETs) are derivatives of the arachidonic acid metabolism with anti-inflammatory activities. However,their efficacy is limited due to the rapid hydrolysis by soluble epoxide hydrolase (sEH). Inhibition of sEH has been shown to stabilize the EETs and reduce neuroinflammation in A? mouse models of Alzheimer’s disease (AD). However,the role of the sEH-EET signaling pathway in other CNS cell types and neurodegenerative conditions are less understood.MethodsHere we investigated the mechanisms and functional role of the sEH-EET axis in tauopathy by treating PS19 mice with a small molecule sEH inhibitor TPPU and by crossing the PS19 mice with Ephx2 (gene encoding sEH) knockout mice. This was followed by single-nucleus RNA-sequencing (snRNA-seq),biochemical and immunohistochemical analysis,and behavioral assessments. Additionally,we examined the effects of the sEH-EET pathway in primary microglia cultures and human induced pluripotent stem cell (iPSC)-derived neurons exhibiting seeding-induced Tau inclusions.ResultssEH inhibition improved cognitive function,rescued neuronal cell loss,and reduced Tau pathology and microglial reactivity. snRNA-seq revealed that TPPU treatment upregulated genes involved in actin cytoskeleton and excitatory synaptic pathways. Treatment of human iPSC-derived neurons with TPPU enhanced synaptic density without affecting Tau accumulation,suggesting a cell-autonomous neuroprotective effect of sEH blockade. Furthermore,sEH inhibition reversed disease-associated and interferon-responsive microglial states in PS19 mice,while EET supplementation promoted Tau phagocytosis and clearance in primary microglia cultures.ConclusionThese findings demonstrate that sEH blockade or EET augmentation confers therapeutic benefit in neurodegenerative tauopathies by simultaneously targeting neuronal and microglial pathways.Supplementary InformationThe online version contains supplementary material available at 10.1186/s13024-025-00844-x. View Publication

Transforming growth factor-beta1 (TGF-beta1) is a pleiotropic cytokine with major in vitro effects on hematopoietic stem cells (HSCs) and lymphocyte development. Little is known about hematopoiesis from mice with constitutive TGF-beta1 inactivation largely because of important embryonic lethality and development of a lethal inflammatory disorder in TGF-beta1(-/-) pups,making these studies difficult. Here,we show that no sign of the inflammatory disorder was detectable in 8- to 10-day-old TGF-beta1(-/-) neonates as judged by both the number of T-activated and T-regulator cells in secondary lymphoid organs and the level of inflammatory cytokines in sera. After T-cell depletion,the inflammatory disease was not transplantable in recipient mice. Bone marrow cells from 8- to 10-day-old TGF-beta1(-/-) neonates showed strikingly impaired short- and long-term reconstitutive activity associated with a parallel decreased in vivo homing capacity of lineage negative (Lin(-)) cells. In addition an in vitro-reduced survival of immature progenitors (Lin(-) Kit(+) Sca(+)) was observed. Similar defects were found in liver cells from TGF-beta1(-/-) embryos on day 14 after vaginal plug. These data indicate that TGF-beta1 is a critical regulator for in vivo homeostasis of the HSCs,especially for their homing potential. View Publication

Substantial progress has been made in the development of radiation countermeasures,resulting in the recent approval of several mitigators; however,there has yet to be an approved prophylactic radioprotectant. Research on countermeasure performance in mixed neutron and gamma radiation fields has also been scarce. Fibroblast-stimulating lipopeptide (FSL-1) is a novel synthetic agonist for toll-like receptor 2/6. In previous studies,the administration of FSL-1 before and after gamma radiation significantly improved survival outcomes for mice through the activation of the NF-κB pathway. In the current study,we tested FSL-1’s radioprotective abilities in a mixed radiation field that models one produced by a nuclear detonation in 11–14-week-old C57BL/6 male and female mice. We demonstrate that a single dose of 1.5 mg/kg of FSL-1 administered 12 h prior to 65% neutron 35% gamma mixed-field (MF) irradiation enhances survival,accelerates recovery of hematopoietic cell and stem cell populations,reduces inflammation,and protects innate immune function in mice. FSL-1’s ability to recover blood and protect immune functions is important in countering the high rate of incidence of sepsis caused by MF radiation’s damaging effects. These results demonstrate that FSL-1 is a promising prophylactic countermeasure where exposure to MF radiation is anticipated. View Publication

NOD.B10-H2(b) and NOD/LtJ mice manifest,respectively,many features of primary and secondary Sjögren's syndrome (SjS),an autoimmune disease affecting primarily the salivary and lacrimal glands leading to xerostomia (dry mouth) and xerophthalmia (dry eyes). B lymphocytes play a central role in the onset of SjS with clinical manifestations dependent on the appearance of autoantibodies reactive to multiple components of acinar cells. Previous studies with NOD.IL4(-/-) and NOD.B10-H2(b).IL4(-/-) mice suggest that the Th2 cytokine,IL-4,plays a vital role in the development and onset of SjS-like disease in the NOD mouse model. To investigate the molecular mechanisms by which IL-4 controls SjS development,a Stat6 gene knockout mouse,NOD.B10-H2(b).C-Stat6(-/-),was constructed and its disease profile was defined and compared with that of NOD.B10-H2(b).C-Stat6(+/+) mice. As the NOD.B10-H2(b).C-Stat6(-/-) mice aged from 4 to 24 wk,they exhibited leukocyte infiltration of the exocrine glands,produced anti-nuclear autoantibodies,and showed loss and gain of saliva-associated proteolytic enzymes,similar to NOD.B10-H2(b).C-Stat6(+/+) mice. In contrast,NOD.B10-H2(b).C-Stat6(-/-) mice failed to develop glandular dysfunction,maintaining normal saliva flow rates. NOD.B10-H2(b).C-Stat6(-/-) mice were found to lack IgG1 isotype-specific anti-muscarinic acetylcholine type-3 receptor autoantibodies. Furthermore,the IgG fractions from NOD.B10-H2(b).C-Stat6(-/-) sera were unable to induce glandular dysfunction when injected into naive recipient C57BL/6 mice. NOD.B10-H2(b).C-Stat6(-/-) mice,like NOD.B10-H2(b).IL4(-/-) mice,are unable to synthesize IgG1 Abs,an observation that correlates with an inability to develop end-stage clinical SjS-like disease. These data imply a requirement for the IL-4/STAT6-pathway for onset of the clinical phase of SjS-like disease in the NOD mouse model. View Publication

Here it is shown that the phenotype of adult mice lacking the first enhancer (DNA hypersensitive site I) and the distal promoter of the GATA-1 gene (neo Delta HS or GATA-1(low) mutants) reveals defects in mast cell development. These include the presence of morphologically abnormal alcian blue(+) mast cells and apoptotic metachromatic(-) mast cell precursors in connective tissues and peritoneal lavage and numerous (60-70% of all the progenitors) unique" trilineage cells committed to erythroid View Publication

Hematopoiesis occurs in close association with a complex network of cells loosely termed the hematopoietic microenvironment. Analysis of the mechanisms of microenvironmental regulation of hematopoiesis has been hindered by the complexity of the microenvironment as well as the heterogeneity of hematopoietic stem cells and early progenitor cells. We have established immortalized primate bone marrow-derived stromal cell lines to facilitate analysis of the interactions of hematopoietic cells with the microenvironment in a large animal species. One such line,PU-34,was found to produce a variety of growth factors,including an activity that stimulates the proliferation of an interleukin 6-dependent murine plasmacytoma cell line. A cDNA encoding the plasmacytoma stimulatory activity was isolated through functional expression cloning in mammalian cells. The nucleotide sequence contained a single long reading frame of 597 nucleotides encoding a predicted 199-amino acid polypeptide. The amino acid sequence of this cytokine,designated interleukin 11 (IL-11),did not display significant similarity with any other sequence in the GenBank data base. Preliminary biological characterization indicates that in addition to stimulating plasmacytoma proliferation,IL-11 stimulates the T-cell-dependent development of immunoglobulin-producing B cells and synergizes with IL-3 in supporting murine megakaryocyte colony formation. These properties implicate IL-11 as an additional multifunctional regulator in the hematopoietic microenvironment. View Publication

Oncogenic Kit mutations are found in somatic gastrointestinal (GI) stromal tumors (GISTs) and mastocytosis. A mouse model for the study of constitutive activation of Kit in oncogenesis has been produced by a knock-in strategy introducing a Kit exon 11-activating mutation into the mouse genome based on a mutation found in a case of human familial GIST syndrome. Heterozygous mutant KitV558Delta/+ mice develop symptoms of disease and eventually die from pathology in the GI tract. Patchy hyperplasia of Kit-positive cells is evident within the myenteric plexus of the entire GI tract. Neoplastic lesions indistinguishable from human GISTs were observed in the cecum of the mutant mice with high penetrance. In addition,mast cell numbers in the dorsal skin were increased. Therefore KitV558Delta/+ mice reproduce human familial GISTs,and they may be used as a model for the study of the role and mechanisms of Kit in neoplasia. Importantly,these results demonstrate that constitutive Kit signaling is critical and sufficient for induction of GIST and hyperplasia of interstitial cells of Cajal. View Publication

Spinal muscular atrophy (SMA),a motor neuron disease (MND) and one of the most common genetic causes of infant mortality,currently has no cure. Patients with SMA exhibit muscle weakness and hypotonia. Stem cell transplantation is a potential therapeutic strategy for SMA and other MNDs. In this study,we isolated spinal cord neural stem cells (NSCs) from mice expressing green fluorescent protein only in motor neurons and assessed their therapeutic effects on the phenotype of SMA mice. Intrathecally grafted NSCs migrated into the parenchyma and generated a small proportion of motor neurons. Treated SMA mice exhibited improved neuromuscular function,increased life span,and improved motor unit pathology. Global gene expression analysis of laser-capture-microdissected motor neurons from treated mice showed that the major effect of NSC transplantation was modification of the SMA phenotype toward the wild-type pattern,including changes in RNA metabolism proteins,cell cycle proteins,and actin-binding proteins. NSC transplantation positively affected the SMA disease phenotype,indicating that transplantation of NSCs may be a possible treatment for SMA. View Publication

Adeno-associated viral (AAV) vector-based gene therapy is gaining foothold as treatment for genetic neurological diseases with encouraging clinical results. Nonetheless,dose-dependent adverse events have emerged in recent clinical trials through mechanisms that remain unclear. We have modelled here the impact of AAV transduction in cell models of the human central nervous system (CNS),taking advantage of induced pluripotent stem cells. Our work uncovers vector-induced innate immune mechanisms that contribute to cell death. While empty AAV capsids were well tolerated,the AAV genome triggered p53-dependent DNA damage responses across CNS cell types followed by the induction of inflammatory responses. In addition,transgene expression led to MAVS-dependent activation of type I interferon responses. Formation of DNA damage foci in neurons and gliosis were confirmed in murine striatum upon intraparenchymal AAV injection. Transduction-induced cell death and gliosis could be prevented by inhibiting p53 or by acting downstream on STING- or IL-1R-mediated responses. Together,our work identifies innate immune mechanisms of vector sensing in the CNS that can potentially contribute to AAV-associated neurotoxicity. Subject terms: Neuroimmunology,Innate immunity,Neural stem cells View Publication

BACKGROUND/AIMS [corrected] Embryonic stem cells (ES cells) have the capacity to propagate indefinitely,maintain pluripotency,and differentiate into any cell type under defined conditions. As a result,they are considered to be the best model system for research into early embryonic development. AICA ribonucleotide (AICAR) is an activator of AMP-activated protein kinase (AMPK) that is thought to affect ES cell function,but its role in ES cell fate decision is unclear. METHODS In this study,we performed microarray analysis to investigate AICAR downstream targets and further understand its effect on ES cells. RESULTS Our microarray data demonstrated that AICAR can significantly up-regulate pluripotency-associated genes and down-regulate differentiation-associated transcription factors. Although AICAR cannot maintain ES cell identity without LIF,it can antagonize the action of RA-induced differentiation. Using those differentially expressed genes identified,we performed gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis with the Database for Annotation,Visualization and Integrated Discovery (DAVID) online system. AICAR was not only shown to influence the AMPK pathway,but also act on other signaling pathways such as BMP,MAPK and TGF-β,to maintain the stemness of J1 ES cells. Furthermore,AICAR modulated ES cell epigenetic modification by altering the expression of epigenetic-associated proteins,including Dnmt3a,Dnmt3b,Smarca2,Mbd3,and Arid1a,or through regulating the transcription of long intervening non-coding RNA (lincRNA). CONCLUSION Taken together,our work suggests that AICAR is capable of maintaining ES cell self-renewal and pluripotency,which could be useful in future medical treatment. View Publication