22 research outputs found
Improving characterisation of human Multipotent Stromal Cells cultured in 2D and 3D: Design and evaluation of primer sets for accurate gene expression normalisation.
Human Multipotent Stromal Cells (MSCs) are a valuable resource for regenerative medicine and are widely studied. They can be isolated from a variety of tissues and differentiate into multiple cell types (multi-potent). Many reports have been published using human MSCs and to be able to compare outcome, or be able to identify differences between MSCs, several cell surface markers have been proposed. Nevertheless, still many differences remain. Gene expression is known to be different between cell stage and origin. Furthermore, cells cultured on a culture dish (2D) show different gene expression profiles as compared to cells grown on scaffolds (3D). Even the RNA extraction method and the selection of genes used for normalisation have a role in gene expression profiling. To be able to compare gene expression data from samples cultured in different dimensions and RNA extracted using a variety of protocols we set out to define a set of reference genes suitable to normalise qPCR data from a very heterogeneous sample set. Hereto, Trizol was used to extract RNA from human MSCs cultured in 3D and 2D to validate newly designed and previously published primer sets. Subsequently, RNA from fresh human MSC samples and samples stored in RLT-buffer, Trizol or RNAlater was extracted using RNeasy and Trizol methods. All samples have been used to rank the candidate reference genes according to their stability after qPCR enabling identification of the most suitable reference gene(s) for normalisation of a heterogeneous sample set. The most stably expressed reference genes indicated superior normalisation of MSC marker gene expression over the least stable reference genes
Transforming Growth Factor Beta Promotes Inflammation and Tumorigenesis in Smad4âDeficient Intestinal Epithelium in a YAPâDependent Manner
Abstract Transforming growth factor beta (TGFâβ), a multifunctional cytokine, plays critical roles in immune responses. However, the precise role of TGFâβ in colitis and colitisâassociated cancer remains poorly defined. Here, it is demonstrated that TGFâβ promotes the colonic inflammation and related tumorigenesis in the absence of Smad family member 4 (Smad4). Smad4 loss in intestinal epithelium aggravates colitis and colitisâassociated neoplasia induced by dextran sulfate sodium (DSS) and azoxymethane/dextran sulfate sodium (AOM/DSS), leading to overâactivated immune responses and increased TGFâβ1 levels. In Smad4âdeficient organoids, TGFâβ1 stimulates spheroid formation and impairs intestinal stem cell proliferation and lineage specification. YAP, whose expression is directly upregulated by TGFâβ1 after Smad4 deletion, mediates the effect of TGFâβ1 by interacting with Smad2/3. Attenuation of YAP/TAZ prevents TGFâβ1âinduced spheroid formation in Smad4â/â organoids and alleviates colitis and colitisâassociated cancer in Smad4âdeficient mice. Collectively, these results highlight an integral role of the TGFâβ/Smad4 axis in restraining intestinal inflammation and tumorigenesis and suggest TGFâβ or YAP signaling as therapeutic targets for these gastrointestinal diseases intervention
EXPERIENCES IN DEVELOPING THE HIGH CURRENT DENSITY WAX-FILLED SUPERCONDUCTING SADDLE MAGNETS
La conception, la fabrication et les rĂŠsultats expĂŠrimentaux de trois aimants ("saddle magnets") sont dĂŠcrit(e)s. Les expĂŠriences pendant le dĂŠveloppement sont rĂŠsumĂŠes.The design, construction technology and test results of three model saddle magnets are described. The main research and development experiences are summarized
Environmental Pollutant Benzo[a]pyrene Induces Recurrent Pregnancy Loss through Promoting Apoptosis and Suppressing Migration of Extravillous Trophoblast
Objects. To investigate the effects of environmental pollutant benzo(a)pyrene (BaP) and its metabolite benzo[a]pyrene-7,8-diol-9,10-epoxide (BPDE) on human trophoblasts and on murine miscarriages. Methods. The implantation sites, fetus resorption, and abnormal fetuses were studied in pregnant mice treated with different doses of BaP by oral gavage from day 1 to day 10 of gestation. Additionally, apoptosis and related signaling pathway, and the migration and invasion of trophoblasts, were assessed before and after exposure of BPDE in Swan 71 trophoblast cell. Besides, the migration and invasion, and its related signaling pathway, were assessed in villi obtained from women. Results. We observed a concentration-dependent incidence of abnormal murine fetuses, beginning with 0.1âmg/kg BaP; with a BaP concentration of 2âmg/kg, no fetuses developed. Correspondingly, a BPDE concentration-dependent apoptosis of human trophoblasts. Beginning with 0.5âÎźM BPDE exposure, Bax/Caspase-3 were increased and Bcl-2 decreased. Furthermore, BPDE also inhibited, in a dose-dependent manner, the migration of villous explants from elective abortion women, consistent with the reduced migration of villous explants from women with recurrent pregnancy loss (RPL), and reduced the cell immigration in Swan 71 trophoblasts, in a dose-dependent manner measured by transwell assays. Conclusions. Our study results provide mechanistic insight to the effect of BPDE on trophoblast dysfunction through enhanced cell apoptosis and inhibited migration, providing further experimental evidence to the causative links between BaP exposure and PRL
Effect of Ion Source Current on the Microstructure and Properties of Cr-DLC Coatings Prepared by Ion Beam-Assisted Arc Ion Plating
Correction: The cyclooxygenase-expressing mesenchyme resists intestinal epithelial injury by paracrine signaling
The cyclooxygenase-expressing mesenchyme resists intestinal epithelial injury by paracrine signaling
Abstract Paracrine signals play pivotal roles in organ homeostasis. Mesenchymal stromal cells (MSCs) play a key role in regulating epithelium homeostasis in the intestine while their paracrine effects are poorly characterized. Here, we identified prostaglandin E2 (PGE2) secreted by cyclooxygenase (COX)-expressing MSCs as a vital factor to maintain the intestinal mucosal barrier. We found that MSCs-induced organoid swelling through paracrine effect in vitro, a process due to enhanced water adsorption and is mediated by the COX-PGE2-EP4 axis. To further explore the regulatory effect of this axis on the intestinal epithelial barrier in vivo, we established the conditional knockout mouse model to specifically delete COX in MSCs and found that PGE2 reduction downregulated the gene Muc2 and induced a gastric metaplasia-like phenotype. Moreover, PGE2 defects increased the susceptibility of intestinal epithelium to colitis. Our study uncovers the paracrine signaling of COX-expressing MSCs in intestinal mucosal barrier maintenance, providing a basis for understanding the role of mesenchymal cells in the pathophysiological function of the intestine
Shanghai cognitive intervention of mild cognitive impairment for delaying progress with longitudinal evaluation-a prospective, randomized controlled study (SIMPLE): rationale, design, and methodology
Abstract Background Mild cognitive impairment is an early stage of Alzheimerâs disease. Increasing evidence has indicated that cognitive training could improve cognitive abilities of MCI patients in multiple cognitive domains, making it a promising therapeutic approach for MCI. However, the effect of long-time training has not been widely explored. It is also necessary to evaluate the extent how it could reduce the convertion rate from MCI to AD. Methods/design The SIMPLE study is a multicenter, randomized, single-blind prospective clinical trial assessing the effects of computerized cognitive training on different cognitive domains in MCI patients. It is carried out in 7 centers in China. The study population includes patients aged 50â85, and they are randomly allocated to the training or control group. The primary outcome is to compare the conversion rate of MCI within 36-month follow-up. Structural and functional MRI will be used to interpret the effect of cognitive training. The cognitive training comprises a variety of games related with cognitive domains such as attention, memory, visualspatial ability and executive function. We cautiously set 50% reduction in the rate of conversion as estimated effect. With 80â90% statistical power and 12% as the overall probability of conversion within the study period, 600â800 patients are finally required in the study. The first patent has been recruited in April 2017. Discussion Previous studies suggested the benefit of cognitive training for MCI, but neither long-time nor Chinese culture were investigated. The SIMPLE designs and utilizes an improved computerized cognitive training approach and assesses its effects on MCI progress. In addition, neural activities explaining the effects on cognition function changes will be revealed, which could in turn to imply more useful therapeutic approaches. Trial registration ClinicalTrials.gov Identifier: NCT03119051