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Cell Sorting (cell + sorting)
Kinds of Cell Sorting Terms modified by Cell Sorting Selected AbstractsRegulatory T cells in Graves' diseaseCLINICAL ENDOCRINOLOGY, Issue 4 2009Deshun Pan Summary Context, Graves' disease (GD) involves auto-immunity against thyroid cell antigens, but the reasons for induction of auto-immunity are uncertain. We wished to determine whether there was a deficiency of regulatory T cells in patients with active GD. Design, Venous blood samples were obtained from patients with GD before and after treatment, and controls, and peripheral blood mononuclear cells were prepared. Patients and measurements, Regulatory T cells were enumerated by Fluorescent Activated Cell sorting (FACS) in nineteen patients with untreated GD, 9 patients 6,8 weeks post RAI therapy, and 30 control subjects. Twenty-one patients with active GD prior to control of hyperthyroidism, 23 euthyroid controls without known autoimmune thyroid disease, and 10 patients who were euthyroid 6,12 months after RAI treatment were studied for expression of genes found in regulatory T cells by real-time Polymerase Chain reaction (PCR). Results, Percent distribution of CD4+, CD4+CD25+ and CD4+ CD25+int-hi CD127+lo regulatory T cells was similar in active GD patients and control subjects. The number of CD25+ and CD4+ CD25+int-hi CD127+lo cells was similar in GD patients and control subjects, but was lower in recently treated patients. Messenger RNA was prepared from PBMC, and reverse transcribed. Copy DNA abundance was evaluated by Real Time PCR using appropriate primers, for GAPDH (glyceraldehyde phosphate dehydrogenase) as a control housekeeping gene, and 5 genes related to function of regulatory T cells. Message RNA for Gadd45 alpha, Gadd45beta (growth arrest and damage inducible proteins), GITR (glucocorticoid inducible TNF receptor) and CD25 (IL-2R subunit) was more abundant in patients with active GD than in normal controls, and FoxP3 mRNA level was equal to that in controls. Message RNA levels in patients treated and euthyroid for 6 months were also greater than or equal to values in controls. Conclusion, This study provides evidence that there is no deficit in T regulatory cells during active GD, or during the months post therapy. [source] An examination of different fetal specific antibodies and magnetic activated cell sorting for the enrichment of fetal erythroblasts from maternal bloodCONGENITAL ANOMALIES, Issue 3 2002Xiao Xi Zhao ABSTRACT, The aim of the present study was to compare the rates of fetal cells obtained after separation from maternal blood by magnetic activated cell sorting (MACS) using different fetal specific antibodies, and to evaluate the potential role of this method in the prenatal diagnosis of fetal trisomies. Peripheral blood samples were obtained from 42 women carrying chromosomally normal fetuses and from 4 women with aneuploid fetuses (2 cases of 47,XX,+18 and 2 of 47,XY,+21) at 9,20 weeks of gestation. After fetal cells were enriched by MACS with three different monoclonal antibodies (GPA, CD71, CD14), fluorescence in situ hybridization (FISH) with chromosome X, and Y-specific probes was performed to detect the rates of fetal cells in the samples sorted. FISH with chromosome 13-, 18-, and 21-specific probes was carried out to compare proportions of cells with three-signal nuclei in chromosomally normal and abnormal groups. In male infants, X-and Y-positive cells were detected in 80%, 73.3%, and 66.6% of samples after the separation by antibodies CD14, GPA, and CD71, respectively. The percentage of nuclei with three signals was increased in pregnancies with trisomy, ranging between 2% and 5.18%. Pregnancies with normal fetuses showed 0 to 3.7% of nuclei with three signals. The data demonstrate that fetal cell detection varies depending on the antibodies used for cell sorting. This study provides further evidence on the feasibility of screening for fetal chromosomal abnormalities by enriching maternal blood for fetal cells and using FISH. [source] Optimization of a flow cytometry-based protocol for detection and phenotypic characterization of multipotent mesenchymal stromal cells from human bone marrowCYTOMETRY, Issue 6 2006Elena A. Jones Abstract Background: To study the biology of rare bone marrow (BM) multipotent mesenchymal stromal cells (MSCs), recognized protocols are needed. Colony-forming unit-fibroblast (CFU-F) assays have historically been used for the enumeration of MSCs. However, the need to isolate and further analyze MSCs requires new strategies based on cell surface markers. The purpose of this work was to verify the phenotype of BM MSCs in vivo and to develop flow cytometry-based methods for their evaluation. Methods: Pre-enrichment with D7-FIB-conjugated microbeads, cell sorting for CD45lowD7-FIB+LNGFR+ cells, and CFU-F assay were used to confirm the phenotype of BM MSCs in vivo. Further phenotypic characterization of MSCs was performed using three-color flow cytometry following pre-enrichment or by direct four-color flow cytometry. The sensitivity of direct flow cytometry/rare event analysis for the accurate enumeration of MSCs was validated using 85 samples from patients with neoplastic BM diseases. Results: In normal BM, a significant correlation was found between the frequencies of CFU-Fs and CD45lowD7-FIB+LNGFR+ cells (n = 19, R = 0.719, P = 0.001). Following cell sorting, ,15% of these cells were clonogenic. The same cells were enriched using LNGFR-based positive selection, CD45/Glycophorin A-based depletion, or plastic adherence. CD45lowD7-FIB+LNGFR+ cells expressed classic makers of cultured MSCs CD73/SH3 and CD105/SH2 and markers of stromal reticular cells CD106/VCAM and alkaline phosphatase. Novel markers were identified including leukemia inhibitory factor receptor and gp130. CD45lowD7-FIB+LNGFR+ cells were increased fourfold in the floating fat fraction of normal BM aspirates. Their frequency was decreased in chronic lymphocytic leukemia (threefold, n = 13, P = 0.049) and chronic myelogenous leukemia (ninefold, n = 11, P = 0.001) compared with that in age-matched controls (n = 26 and n = 31, respectively). Conclusions: This study demonstrates the usefulness of flow cytometry-based methods for the detection, enumeration and further phenotypic analysis of BM MSCs. These findings have broad applications for the future evaluation of BM MSCs in health and disease. © 2006 International Society for Analytical Cytology [source] Near-infrared dyes for six-color immunophenotyping by laser scanning cytometryCYTOMETRY, Issue 3 2002Andreas O.H. Gerstner Abstract Background To adequately analyze the complexity of the immune system and reduce the required sample volume for immunophenotyping in general, more measurable colors for the discrimination of leukocyte subsets are necessary. Immunophenotyping by the laser scanning cytometer (LSC), a slide-based cytometric technology, combines cell detection based on multiple colors with their subsequent visualization without the need for physical cell sorting. In the present study, the filter setting of the LSC was adapted for the measurement of the far-red emitting dye cyanine 7 (Cy7), thereby increasing the number of measurable commercially available fluorochromes. Methods The optical filters of the LSC were replaced,photomultiplier (PMT) 3/allophycocyanin (APC): 740-nm dichroic long pass, and 670-/55-nm bandpass; PMT 4/Cy7: 810-/90-nm bandpass. Peripheral blood leukocytes were stained directly by fluorochrome-labeled antibodies or by indirect staining. The tandem dyes of Cy7 (phycoerythrin [PE]-Cy7, APC-Cy7) and the fluorochromes fluorescein isothiocyanate (FITC), PE, PE-Cy5, and APC were tested alone and in different combinations. Results With the new filter combination and tandem fluorochromes, Cy7 was measurable at 488-nm (argon laser) or 633-nm (helium-neon laser) excitation. Resolution was in the range of FITC for PE-Cy7 but approximately 30% lower for APC-Cy7; spillover into the respective donor fluorochrome channel for both tandem dyes was prominent. A six-color panel for leukocyte subtyping was designed. Conclusions With this adaptation, it is possible to measure the tandem conjugates PE-Cy7 and APC-Cy7. This new setup opens the way for six-color immunophenotyping by LSC. Cytometry 48:115,123, 2002. © 2002 Wiley-Liss, Inc. [source] Cellular and molecular dissection of pluripotent adult somatic stem cells in planariansDEVELOPMENT GROWTH & DIFFERENTIATION, Issue 1 2010Norito Shibata Freshwater planarians, Plathelminthes, have been an intriguing model animal of regeneration studies for more than 100 years. Their robust regenerative ability is one of asexual reproductive capacity, in which complete animals develop from tiny body fragments within a week. Pluripotent adult somatic stem cells, called neoblasts, assure this regenerative ability. Neoblasts give rise to not only all types of somatic cells, but also germline cells. During the last decade, several experimental techniques for the analysis of planarian neoblasts at the molecular level, such as in situ hybridization, RNAi and fluorescence activated cell sorting, have been established. Moreover, information about genes involved in maintenance and differentiation of neoblasts has been accumulated. One of the molecular features of neoblasts is the expression of many RNA regulators, which are involved in germline development in other animals, such as vasa and piwi family genes. In this review, we introduce physiological and molecular features of the neoblast, and discuss how germline genes regulate planarian neoblasts and what differences exist between neoblasts and germline cells. [source] Single-cell gene profiling of planarian stem cells using fluorescent activated cell sorting and its "index sorting" function for stem cell researchDEVELOPMENT GROWTH & DIFFERENTIATION, Issue 1 2010Tetsutaro Hayashi To achieve an integrated understanding of the stem cell system of planarians at both the cellular and molecular levels, we developed a new method by combining "fluorescent activated cell sorting (FACS) index sorting" analysis and single-cell reverse transcription,polymerase chain reaction (RT,PCR) to detect the gene expression and cell cycle state of stem cells simultaneously. Single cells were collected using FACS, and cDNAs of each cell were used for semi-quantitative RT,PCR. The results were plotted on the FACS sorting profile using the "index sorting" function, which enabled us to analyze the gene expression in combination with cell biological data (such as cell cycle phase) for each cell. Here we investigated the adult stem cells of planarians using this method and obtained findings suggesting that the stem cells might undergo commitment during S to G2/M phase. This method could be a powerful and straightforward tool for examining the stem cell biology of not only planarians but also other organisms, including vertebrates. [source] Tissue surface tensions guide in vitro self-assembly of rodent pancreatic islet cellsDEVELOPMENTAL DYNAMICS, Issue 8 2007Dongxuan Jia Abstract The organization of endocrine cells in pancreatic islets is established through a series of morphogenetic events involving cell sorting, migration, and re-aggregation processes for which intercellular adhesion is thought to play a central role. In animals, these morphogenetic events result in an islet topology in which insulin-secreting cells form the core, while glucagon, somatostatin, and pancreatic polypeptide-secreting cells segregate to the periphery. Isolated pancreatic islet cells self-assemble in vitro into pseudoislets with the same cell type organization as native islets. It is widely held that differential adhesion between cells of the pancreatic islets generates this specific topology. However, this differential adhesion has never been rigorously quantified. In this manuscript, we use tissue surface tensiometry to measure the cohesivity of spherical aggregates from three immortalized mouse pancreatic islet cell lines. We show that, as predicted by the differential adhesion hypothesis, aggregates of the internally segregating INS-1 and MIN6 beta-cell lines are substantially more cohesive than those of the externally segregating ,-TC line. Furthermore, we show that forced overexpression of P-cadherin by ,-TC cells significantly perturbs the sorting process. Collectively, the data indicate that differential adhesion can drive the in vitro organization of immortalized rodent pancreatic islet cells. Developmental Dynamics 236:2039,2049, 2007. © 2007 Wiley-Liss, Inc. [source] Molecular characterization of conditionally immortalized cell lines derived from mouse early embryonic inner earDEVELOPMENTAL DYNAMICS, Issue 4 2004John A. Germiller Abstract Inner ear sensory hair cells (HCs), supporting cells (SCs), and sensory neurons (SNs) are hypothesized to develop from common progenitors in the early embryonic otocyst. Because little is known about the molecular signals that control this lineage specification, we derived a model system of early otic development: conditionally immortalized otocyst (IMO) cell lines from the embryonic day 9.5 Immortomouse. This age is the earliest stage at which the otocyst can easily be separated from surrounding mesenchymal, nervous system, and epithelial cells. At 9.5 days post coitum, there are still pluripotent cells in the otocyst, allowing for the eventual identification of both SN and HC precursors,and possibly an elusive inner ear stem cell. Cell lines derived from primitive precursor cells can also be used as blank canvases for transfections of genes that can affect lineage decisions as the cells differentiate. It is important, therefore, to characterize the "baseline state" of these cell lines in as much detail as possible. We characterized seven representative "precursor-like" IMO cell populations and the uncloned IMO cells, before cell sorting, at the molecular level by polymerase chain reaction (PCR) and immunocytochemistry (IHC), and one line (IMO-2B1) in detail by real-time quantitative PCR and IHC. Many of the phenotypic markers characteristic of differentiated HCs or SCs were detected in IMO-2B1 proliferating cells, as well as during differentiation for up to 30 days in culture. These IMO cell lines represent a unique model system for studying early stages of inner ear development and determining the consequences of affecting key molecular events in their differentiation. Developmental Dynamics 231:815,827, 2004. © 2004 Wiley-Liss, Inc. [source] FACS-array gene expression analysis during early development of mouse telencephalic interneuronsDEVELOPMENTAL NEUROBIOLOGY, Issue 4 2008Eric D. Marsh Abstract Cortical interneuron dysfunction has been implicated in multiple human disorders including forms of epilepsy, mental retardation, and autism. Although significant advances have been made, understanding the biologic basis of these disorders will require a level of anatomic, molecular, and genetic detail of interneuron development that currently does not exist. To further delineate the pathways modulating interneuron development we performed fluorescent activated cell sorting (FACs) on genetically engineered mouse embryos that selectively express green fluorescent protein (GFP) in developing interneurons followed by whole genome microarray expression profiling on the isolated cells. Bioinformatics analysis revealed expression of both predicted and unexpected genes in developing cortical interneurons. Two unanticipated pathways discovered to be up regulated prior to interneurons differentiating in the cortex were ion channels/neurotransmitters and synaptic/vesicular related genes. A significant association of neurological disease related genes to the population of developing interneurons was found. These results have defined new and potentially important data on gene expression changes during the development of cortical interneurons. In addition, these data can be mined to uncover numerous novel genes involved in the generation of interneurons and may suggest genes/pathways potentially involved in a number of human neurological disorders. © 2008 Wiley Periodicals, Inc. Develop Neurobiol, 2008. [source] Cover Picture: Electrophoresis 6/2008ELECTROPHORESIS, Issue 6 2008Article first published online: 18 MAR 200 Regular issues provide a wide range of research and review articles covering all aspects of electrophoresis. Here you will find cutting-edge articles on methods and theory, instrumentation, nucleic acids, CE and CEC, miniaturization and microfluidics, proteomics and two-dimensional electrophoresis. In addition, issue no. 6 features a series of 7 important papers on "Microfluidics and Miniaturization" dealing with LCD-based optoelectronic tweezers, cell sorting, single cell clone analysis and cultivation, integrated ITP stacking and gel electrophoresis, floating injection, electrokinetic flows on thermosensitive surfaces and flow velocity measurement in CE chip instruments. [source] A micropillar-integrated smart microfluidic device for specific capture and sorting of cellsELECTROPHORESIS, Issue 24 2007Yan-Jun Liu Abstract An integrated smart microfluidic device consisting of nickel micropillars, microvalves, and microchannels was developed for specific capture and sorting of cells. A regular hexagonal array of nickel micropillars was integrated on the bottom of a microchannel by standard photolithography, which can generate strong induced magnetic field gradients under an external magnetic field to efficiently trap superparamagnetic beads (SPMBs) in a flowing stream, forming a bed with sufficient magnetic beads as a capture zone. Fluids could be manipulated by programmed controlling the integrated air-pressure-actuated microvalves, based on which in situ bio-functionalization of SPMBs trapped in the capture zone was realized by covalent attachment of specific proteins directly to their surface on the integrated microfluidic device. In this case, only small volumes of protein solutions (62.5,nL in the capture zone; 375,nL in total volume needed to fill the device from inlet A to the intersection of outlet channels F and G) can meet the need for protein! The newly designed microfluidic device reduced greatly chemical and biological reagent consumption and simplified drastically tedious manual handling. Based on the specific interaction between wheat germ agglutinin (WGA) and N -acetylglucosamine on the cell membrane, A549 cancer cells were effectively captured and sorted on the microfluidic device. Capture efficiency ranged from 62 to 74%. The integrated microfluidic device provides a reliable technique for cell sorting. [source] Analysis of electrokinetic transport of a spherical particle in a microchannelELECTROPHORESIS, Issue 4 2007Harikrishnan N. Unni Abstract Electrokinetically driven microfluidic devices that are used for biological cell/particle manipulation (e.g., cell sorting, separation) involve electrokinetic transport of these particles in microchannels whose dimension is comparable with particles' size. This paper presents an analytical study on electrokinetic transport of a charged spherical particle in a charged parallel-plate microchannel. Under the thin electric double-layer assumption, solutions in closed-form solutions for the particle velocity and disturbed electrical and fluid velocity fields are obtained for plane-symmetric (along the channel centerline) and asymmetric (off the channel centerline) motions of a sphere in a parallel-plate microchannel. The effects of relative particle size and eccentricity (i.e., off the centerline distance) on a particle's translational and rotational velocities are analyzed. [source] Bacterioplankton assemblages transforming dissolved organic compounds in coastal seawaterENVIRONMENTAL MICROBIOLOGY, Issue 8 2007Xiaozhen Mou Summary To characterize bacterioplankton functional assemblages that transform specific components of the coastal seawater dissolved organic carbon (DOC) pool, bromodeoxyuridine (BrdU) was used to label the bacterioplankton cells that were active following addition of single-DOC model compounds: two organic osmolytes [dimethylsulfoniopropionate (DMSP) and glycine betaine (GlyB)] and two aromatic monomers [para -hydroxybenzoic acid (pHBA) and vanillic acid (VanA)]. Bacterial populations were analysed based on in situ fluorescent immunodetection of BrdU incorporation followed by fluorescence-activated cell sorting (FACS). Sorted cells were then characterized by 16S rDNA-based analysis. Populations with high BrdU incorporation level (HI) developed within 8 h of introduction of 100 nM model compound. Terminal restriction fragment length polymorphisms (T-RFLP) analysis indicated that the HI populations in all four amendments were composed of bacteria from the same major taxa (phylum and subphylum levels), but the relative abundance of each differed. High-resolution clone libraries (each containing ,200 clones) showed that the HI populations in the GlyB and VanA amendments consisted of both metabolic generalists and specialists within the , -Proteobacteria (mainly members of the Roseobacter clade), , -Proteobacteria and , -Proteobacteria (mainly members of Altermonadaceae, Chromatiaceae, Oceanospirillaceae and Pseudomonadaceae). The presence of members of OM60/241, OM185, SAR11, SAR86 and SAR116 in the HI populations indicated that members of these groups can assimilate the model DOC compounds, providing some of the first glimpses into heterotrophy by members of these poorly understood environmental clusters. [source] Procoagulant and inflammatory response of virus-infected monocytesEUROPEAN JOURNAL OF CLINICAL INVESTIGATION, Issue 10 2002J. J. M. Bouwman Abstract Background Monocytes play a prominent role in inflammation, coagulation and atherosclerosis by their ability to produce tissue factor (TF) and cytokines. The aim of the present study was to establish whether virus-infected monocytes initiate coagulation. In addition, the production of cytokines by monocytes may accelerate the chronic process of atherosclerosis and may contribute to coronary syndromes by eliciting plaque instability. Materials and methods Monocytes were isolated by Vacutainer®, BD Biosciences, Alphen aan den Rijn, Netherlands and subsequent magnetic cell sorting (MACS®, Milteny Biotec, Bergish Gladbach, Germany). Coagulation times in normal pooled plasma and Factor VII-deficient plasma were measured after infection with cytomegalovirus (CMV), Chlamydia pneumoniae (Cp) and influenza A\H1N1. Anti-TF antibodies were added to neutralize TF expressed on monocytes. Interleukins (IL) 6, 8 and 10 were measured in the supernatants. Results Chlamydia pneumoniae- and CMV-infected monocytes decreased the clotting time by 60%, and influenza-infected monocytes by 19%, as compared to uninfected monocytes. Procoagulant activity was absent when Factor VII-deficient plasma or anti-TF antibodies were used. Monocytes produced both IL-6 and IL-8 after infection with CMV (317 pg mL,1 and 250 pg mL,1) or Cp (733 pg mL,1 and 268 pg mL,1). Similar results were obtained for influenza virus-infected monocytes, but the levels of both cytokines were 3,5-fold higher (1797 pg mL,1 and 725 pg mL,1). Interleukin-10 was not produced by infected monocytes. Conclusion The procoagulant activity of virus-infected monocytes is TF-dependent. Although influenza infection did not generate a significant reduction in clotting time, the pronounced expression of IL-6 and IL-8 may induce local and/or systemic inflammatory reactions, which may be associated with plaque rupture and atherosclerosis. The lack of production of the anti-inflammatory cytokine IL-10 may even accelerate these processes. [source] Activated CD1d-restricted natural killer T cells secrete IL-2: innate help for CD4+CD25+ regulatory T cells?EUROPEAN JOURNAL OF IMMUNOLOGY, Issue 4 2005Shuiping Jiang Abstract CD4+CD25+ and CD1d-restricted natural killer T (NKT) cells are thymus-derived self-reactive regulatory T,cells that play a key role in the control of pathological immune responses. Little is known about functional cooperation between innate regulatory NKT,cells and adaptive CD4+CD25+ regulatory cells. Here we show that human CD4+V,24+V,11+ (CD4+ NKT) cells isolated from peripheral blood by flow cytometric cell sorting secrete substantial amounts of IL-2 after stimulation with dendritic cells (DC) and ,-Galactosylceramide. When cocultured with CD4+CD25+ cells, CD4+ NKT,cells promoted moderate proliferation of CD4+CD25+ cells. The proliferation of CD4+CD25+ T,cells was due to soluble IL-2 produced by activated CD4+ NKT,cells. The expanded CD4+CD25+ cells remained anergic and retained their potent suppressive properties. These findings indicate that unlike conventional CD4+ and CD8+ T,cells, which are susceptible to CD4+CD25+ regulatory cell suppression, NKT,cells promote CD4+CD25+ regulatory cell proliferation. These data raise the possibility that NKT,cells can function as helper cells to CD4+CD25+ regulatory T,cells, thereby providing a link between the two naturally occurring populations of regulatory T,cells. [source] Isolation and characterization of neural precursor cells from the Sox1,GFP reporter mouseEUROPEAN JOURNAL OF NEUROSCIENCE, Issue 7 2005Perrine Barraud Abstract We have made use of a reporter mouse line in which enhanced green fluorescence protein (GFP) is inserted into the Sox1 locus. We show that the GFP reporter is coexpressed with the Sox1 protein as well as with other known markers for neural stem and progenitor cells, and can be used to identify and isolate these cells by fluorescence-activated cell sorting (FACS) from the developing or adult brain and from neurosphere cultures. All neurosphere-forming cells with the capacity for multipotency and self-renewal reside in the Sox1,GFP-expressing population. Thus, the Sox1,GFP reporter system is highly useful for identification, isolation and characterization of neural stem and progenitor cells, as well as for the validation of alternative means for isolating neural stem and progenitor cells. Further, transplantation experiments show that Sox1,GFP cells isolated from the foetal brain give rise to neurons and glia in vivo, and that many of the neurons display phenotypic characteristics appropriate for the developing brain region from which the Sox1,GFP precursors were derived. On the other hand, Sox1,GFP cells isolated from the adult subventricular zone or expanded neurosphere cultures gave rise almost exclusively to glial cells following transplantation. Thus, not all Sox1,GFP cells possess the same capacity for neuronal differentiation in vivo. [source] Expression of psoriasis-associated fatty acid-binding protein in senescent human dermal microvascular endothelial cellsEXPERIMENTAL DERMATOLOGY, Issue 9 2004Moon Kyung Ha Abstract:, Aging is associated with the progressive pathophysiologic modification of endothelial cells. In vitro endothelial cell senescence is accompanied by proliferative activity failure and by perturbations in gene and protein expressions. Moreover, this cellular senescence in culture has been proposed to reflect processes that occur in aging organisms. In order to observe the changing patterns of protein expression in senescent human dermal microvascular endothelial cells (HDMECs), proteins obtained from both early- and late-passaged HDMECs were separated by two-dimensional electrophoresis, visualized by silver staining, and quantified by image processing. Proteins of interest were extracted by in-gel digestion with trypsin and quantified by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS), by searching the National Center for Biotechnology Information protein-sequence database. More than 2000 spots were detected by 2D electrophoresis within a linear pH range of 3,10. Twenty-two major differentially expressed spots were observed in serially passaged HDMECs and identified with high confidence by MALDI-TOF-MS. One of these spots was found to be a 14,15 kDa psoriasis-associated fatty acid-binding protein (PA-FABP) with high affinity for long-chain fatty acids. The expression of PA-FABP was confirmed to be elevated in senescent HDMECs (passage 20) by fluorescence-activated cell sorting (FACS), confocal laser microscopy, and by immunohistochemistry in aged human skin tissue. Our results suggest that the overexpression of FABP in cultured senescent HDMECs is closely related to skin aging. [source] Secreted CREG inhibits cell proliferation mediated by mannose 6-phosphate/insulin-like growth factor II receptor in NIH3T3 fibroblastsGENES TO CELLS, Issue 9 2008Ya-Ling Han Cellular repressor of E1A-stimulated genes (CREG) is a recently described glycoprotein that plays a critical role in keeping cells or tissues in mature, homeostatic states. To understand the relationship between CREG and its membrane receptor, mannose 6-phosphate/insulin-like growth factor II receptor (M6P/IGF2R), we first generated stable NIH3T3 fibroblasts by transfection of pDS_shCREGs vectors, which produced an approximately 80% decrease in CREG levels both in the lysate and in the media. We used fluorescence activated cell sorting and a bromide deoxyuridine incorporation assay to identify whether CREG knockdown promoted the cell proliferation associated with the increase of IGF-II in NIH3T3 fibroblasts. Proliferation was markedly inhibited in a concentration-dependent manner by re-addition of recombinant CREG protein into the media, and this was mediated by the membrane receptor M6P/IGF2R. We subsequently confirmed the direct interaction of CREG and M6P/IGF2R by both immunoprecipitation-Western blotting and immunofluorescence staining. We found that expression of CREG correlated with localization of the receptor in NIH3T3 fibroblasts but did not affect its expression. Our findings indicated that CREG might act as a functional regulator of M6P/IGF2R to facilitate binding and trafficking of IGF-II endocytosis, leading to growth inhibition. [source] In vivo investigation of CD133 as a putative marker of cancer stem cells in Hep-2 cell lineHEAD & NECK: JOURNAL FOR THE SCIENCES & SPECIALTIES OF THE HEAD AND NECK, Issue 1 2009Xu Dong Wei PhD Abstract Background Mounting evidence suggests that most tumors consist of a heterogeneous population of cells with a subset population that has the exclusive tumorigenic ability. They are called cancer stem cells (CSCs). CSCs can self-renew to generate additional CSCs and also differentiate to generate phenotypically diverse cancer cells with limited proliferative potential. They have been identified in a variety of tumors. In this study, we identify the marker of CSCs in the established human laryngeal tumor Hep-2 cell line in vivo. Our in vitro experiment shown as CD133, a 5-transmembrane glycoprotein expressed in Hep-2 cell line. CD133 was supposed as a candidate of CSC in laryngeal carcinoma. In this study, the expression of CD133 was detected in a Hep-2 cell line. Applying the magnetic cell sorting (MACS) technology, we reported the results of purifying CD133 positive cells from a Hep-2 cell line. Three-type cells' tumor-forming ability was examined in vivo to identify the marker of CSCs in Hep-2 cell line. Methods CD133 was selected as a putative marker of CSC in laryngeal carcinoma, Hep-2 cell lines. Flow cytometry was used to detect the expression of CD133 in the Hep-2 cell line. Immunomagnetic beads were applied to purify CD133-positive cells. CD133(+), CD133(,) tumor cells, and unsorted Hep-2 cells were injected into severe combined immune deficiency (SCID) mice individually to observe tumor-forming ability. Results Only a small proportion (3.15% ± 0.83%) of cells in the Hep-2 cell line express the CD133 marker. In comparison with CD133(,) tumor cells and unsorted cells, CD133(+) cells possess a marked capacity for tumor formation in vivo (p <.05). Conclusion CD133 is 1 of the markers for CSCs in human laryngeal tumors of the Hep-2 cell line. Work on the characterization of these cells provides a powerful tool to investigate the tumorigenic process in the larynx and to develop therapies targeting the CSC. © 2008 Wiley Periodicals, Inc. Head Neck, 2009 [source] CXC chemokine ligand 4 (Cxcl4) is a platelet-derived mediator of experimental liver fibrosis,HEPATOLOGY, Issue 4 2010Mirko Moreno Zaldivar Liver fibrosis is a major cause of morbidity and mortality worldwide. Platelets are involved in liver damage, but the underlying molecular mechanisms remain elusive. Here, we investigate the platelet-derived chemokine (C-X-C motif) ligand 4 (CXCL4) as a molecular mediator of fibrotic liver damage. Serum concentrations and intrahepatic messenger RNA of CXCL4 were measured in patients with chronic liver diseases and mice after toxic liver injury. Platelet aggregation in early fibrosis was determined by electron microscopy in patients and by immunohistochemistry in mice. Cxcl4,/, and wild-type mice were subjected to two models of chronic liver injury (CCl4 and thioacetamide). The fibrotic phenotype was analyzed by histological, biochemical, and molecular analyses. Intrahepatic infiltration of immune cells was investigated by fluorescence-activated cell sorting, and stellate cells were stimulated with recombinant Cxcl4 in vitro. The results showed that patients with advanced hepatitis C virus,induced fibrosis or nonalcoholic steatohepatitis had increased serum levels and intrahepatic CXCL4 messenger RNA concentrations. Platelets were found directly adjacent to collagen fibrils. The CCl4 and thioacetamide treatment led to an increase of hepatic Cxcl4 levels, platelet activation, and aggregation in early fibrosis in mice. Accordingly, genetic deletion of Cxcl4 in mice significantly reduced histological and biochemical liver damage in vivo, which was accompanied by changes in the expression of fibrosis-related genes (Timp-1 [tissue inhibitor of matrix metalloproteinase 1], Mmp9 [matrix metalloproteinase 9], Tgf -, [transforming growth factor beta], IL10 [interleukin 10]). Functionally, Cxcl4,/, mice showed a strongly decreased infiltration of neutrophils (Ly6G) and CD8+ T cells into the liver. In vitro, recombinant murine Cxcl4 stimulated the proliferation, chemotaxis, and chemokine expression of hepatic stellate cells. Conclusion: The results underscore an important role of platelets in chronic liver damage and imply a new target for antifibrotic therapies. (HEPATOLOGY 2010.) [source] Ammonia impairs neutrophil phagocytic function in liver disease,HEPATOLOGY, Issue 4 2008Debbie L. Shawcross Hyperammonemia is a feature of liver failure, which is associated with increased risk of infection. The aims of the present study were to determine in vitro, in rats fed an ammoniagenic diet and in patients with cirrhosis, whether induction of hyperammonemia results in neutrophil dysfunction. As hyperammonemia produces cell swelling, we explored the role of the osmoregulating, p38 mitogen-activated protein kinase (p38MAPK) pathway in mediating this neutrophil dysfunction. Neutrophils were isolated from blood of healthy volunteers and incubated with either 75 ,M ammonia or phosphate-buffered saline. Both groups were studied under hyponatremic conditions and/or with the addition of p38MAPK modulators. Neutrophil phagocytosis was measured in naive rats and rats fed an ammoniagenic diet and in patients with stable cirrhosis given placebo (n = 8) or an amino acid solution inducing hyperammonemia (n = 8). Cell volume and phagocytosis was analyzed by fluorescent-activated cell sorting using fluorescein isothiocyanate,labeled E. coli. p38MAPK phosphorylation was measured by western blotting. In healthy neutrophils incubated with ammonia and in rats fed an ammoniagenic diet, neutrophils showed evidence of swelling, impaired phagocytosis, and increased spontaneous oxidative burst compared to controls. Phagocytosis was significantly impaired in patients with induced hyperammonemia compared to placebo. The effects of hyperammonemia and hyponatremia were synergistic. The p38MAPK intracellular signaling pathways were activated in healthy neutrophils exposed to ammonia in association with increased burst activity. Neutrophil phagocytic dysfunction was abrogated by the addition of a p38MAPK agonist. Conclusion: Ammonia produces neutrophil swelling and impairs neutrophil phagocytosis. The p38MAPK intracellular signaling pathway has been shown to be important in mediating the ammonia-induced neutrophil dysfunction. (HEPATOLOGY 2008.) [source] Activation of hepatic stellate cells after phagocytosis of lymphocytes: A novel pathway of fibrogenesis,HEPATOLOGY, Issue 3 2008Nidal Muhanna Increased CD8-T lymphocytes and reduced natural killer (NK) cells contribute to hepatic fibrosis. We have characterized pathways regulating the interactions of human hepatic stellate cells (HSCs) with specific lymphocyte subsets in vivo and in vitro. Fluorescence-activated cell sorting (FACS) was used to characterize human peripheral blood lymphocytes (PBLs) and intrahepatic lymphocytes (IHLs) obtained from healthy controls and from patients with either hepatitis B virus (HBV) or hepatitis C virus (HCV) with advanced fibrosis. Liver sections were analyzed by immunohistochemistry and confocal microscopy. To investigate in vitro interactions, PBLs from healthy controls or patients with HCV cirrhosis were co-cultured with an immortalized human HSC line (LX2 cells) or with primary HSCs. Significant alterations in lymphocyte distribution were identified in IHLs but not PBLs. The hepatic CD4/CD8 ratio and NK cells were significantly reduced in HBV/HCV patients. Expression of alpha-smooth muscle actin and infiltration of CD4, CD8, and NK cells were readily apparent in liver sections from patients with cirrhosis but not in healthy controls. Lymphocytes from each subset were in proximity to HSCs primarily within the periportal regions, and some were directly attached or engulfed. In culture, HSC activation was stimulated by HCV-derived CD8-subsets but attenuated by NK cells. Confocal microscopy identified lymphocyte phagocytosis within HSCs that was completely prevented by blocking intracellular adhesion molecule 1 (ICAM-1) and integrin molecules, or by irradiation of HSCs. LX2 knockdown of either Cdc42 or Rac1 [members of the Rho-guanosine triphosphatase (GTPase) family] prevented both phagocytosis and the activation of HSC by HCV-derived lymphocytes. Conclusion: The CD4/CD8 ratio and NK cells are significantly decreased in livers with advanced human fibrosis. Moreover, disease-associated but not healthy lymphocytes are engulfed by cultured HSCs, which is mediated by the Rac1 and Cdc42 pathways. Ingestion of lymphocytes by HSCs in hepatic fibrosis is a novel and potentially important pathway regulating the impact of lymphocytes on the course of hepatic fibrosis. (HEPATOLOGY 2008.) [source] Naturally occurring polyphenolic antioxidants modulate IgE-mediated mast cell activationIMMUNOLOGY, Issue 4 2000S.-S. Chen Summary Reactive oxygen species (ROS) are known to modulate activities of a host of kinases, phosphatases and transcription factors. Rutin and chlorogenic acid (CGA) are the major polyphenolic antioxidants present in the small molecular fraction of smokeless tobacco leaf extracts, as ascertained by reverse-phase high-pressure liquid chromatography (HPLC) and mass spectrometry. Levels of intracellular ROS in resting versus antigen,immunoglobulin E (IgE)-challenged murine mast cells were measured at 510 nm by fluorescence-activated cell sorting (FACS) using carboxy-dichlorofluorescein (DCFH-DA). Enhanced ROS production was observed in IgE-sensitized mast cells following antigenic challenge. Rutin and CGA reduced ROS levels in antigen,IgE-activated mast cells. Concomitantly, they also profoundly inhibited histamine release by these activated mast cells. In contrast, rutin and CGA augmented the inducible cytokine messages, i.e. interleukin (IL)-10, IL-13, interferon-, (IFN-,), IL-6 and tumour necrosis factor-, (TNF-,) in IgE-sensitized mast cells following antigen challenge. This study indicates that tobacco polyphenolic antioxidants that quench intracellular ROS, differentially affect two effector functions of antigen,IgE-activated mast cells. This model system may be employed to determine the molecular target of polyphenols. The potential role of these polyphenolic antioxidants on IgE-mediated allergy in vivo depends on a balance of their differential effects on mast cell activation. [source] Dipeptidyl peptidase expression during experimental colitis in miceINFLAMMATORY BOWEL DISEASES, Issue 8 2010Roger Yazbeck PhD Abstract Background: We have previously demonstrated that inhibition of dipeptidyl peptidase (DP) activity partially attenuates dextran sulfate sodium (DSS) colitis in mice. The aim of this study was to further investigate the mechanisms of this protection. Materials and Methods: Wildtype (WT) and DPIV,/, mice consumed 2% DSS in drinking water for 6 days to induce colitis. Mice were treated with saline or the DP inhibitors Ile-Pyrr-(2-CN)*TFA or Ile-Thia. DP mRNA and enzyme levels were measured in the colon. Glucagon-like peptide (GLP)-2 and GLP-1 concentrations were determined by radioimmunoassay, regulatory T-cells (Tregs) by fluorescence activated cell sorting (FACS) on FOXp3+T cells in blood, and neutrophil infiltration assessed by myeloperoxidase (MPO) assay. Results: DP8 and DP2 mRNA levels were increased (P < 0.05) in WT+saline mice compared to untreated WT mice with colitis. Cytoplasmic DP enzyme activity was increased (P < 0.05) in DPIV,/, mice at day 6 of DSS, while DP2 activity was increased (P < 0.05) in WT mice with colitis. GLP-1 (63%) and GLP-2 (50%) concentrations increased in WT+Ile-Pyrr-(2-CN)*TFA mice compared to day-0 controls. MPO activity was lower in WT+Ile-Thia and WT+Ile-Pyrr-(2-CN)*TFA treated mice compared to WT+saline (P < 0.001) at day 6 colitis. Conclusions: DP expression and activity are differentially regulated during DSS colitis, suggesting a pathophysiological role for these enzymes in human inflammatory bowel disease (IBD). DP inhibitors impaired neutrophil recruitment and maintenance of the Treg population during DSS-colitis, providing further preclinical evidence for the potential therapeutic use of these inhibitors in IBD. Finally, DPIV appears to play a critical role in mediating the protective effect of DP inhibitors. Inflamm Bowel Dis 2010 [source] Establishment, characterization and drug sensitivity testing in primary cultures of human thymoma and thymic carcinomaINTERNATIONAL JOURNAL OF CANCER, Issue 12 2008Volker Ehemann Abstract Thymomas and thymic carcinomas are peculiar epithelial tumors of the anterior mediastinum. They may show aggressive clinical behavior and are a paradigm for the interaction between the tumor and the immune system. So far, adequate functional studies enabling a better understanding of this malignancy have not been performed, since human thymoma/thymic carcinoma cell lines have not been available. Here, the authors describe the establishment, characterization and functional analyses of epithelial cell lines from a Type B1-thymoma and a poorly differentiated thymic carcinoma. By Fluorescence-activated cell sorting (FACS) analyses, both cell lines were aneuploid. The aneuploid cell fraction of the thymic carcinoma cell line was characterized by a high proliferation index of 55.9%, in contrast to a lower proliferation rate of the aneuploid cell fraction of the thymoma (19.7%). Array-based comparative genomic hybridization (aCGH) and conventional cytogenetic analysis of the thymoma revealed only minor imbalances whereas the thymic carcinoma was characterized by a complex karyotype in the hyperdiploid range that was readily defined with multicolor FISH (mFISH). Application of a selective COX-2 inhibitor reduced cell viability in both cell lines in a dose-dependent manner. In conclusion, these first cell lines of a thymoma and a CD5-positive thymic carcinoma are useful tools for further in vitro studies of cellular, molecular and genetic aspects of the disease and for functional tests to evaluate new therapeutic targets. © 2008 Wiley-Liss, Inc. [source] Development and clinical application of nucleated red blood cell counting and staging on the automated haematology analyser XE-2100TMINTERNATIONAL JOURNAL OF LABORATORY HEMATOLOGY, Issue 1 2003F.-S. Wang Summary We initially developed a new flow cytometric (FCM) reference method for the enumeration and staging of nucleated red blood cells (NRBC) in 1997 [Wang et al., 1998 (XIth International Symposium on Technological Innovations in Laboratory Haematology, Banff, Canada, 1998); Tsuji et al., 1999 (Cytometry, 1999)]. The method used CD45 antibody and propidium iodide staining to separate NRBCs from other cells. Accuracy and precision were enhanced because larger numbers of cells were counted than was possible with the manual method. We also developed a method for automated NRBC counting on a haematology analyser, the XE-2100 (Wang, 1988). NRBC were separated from other cells using a special lysing buffer and a fluorescent dye. The XE-2100 was found to detect peripheral and cord blood NRBC accurately and precisely when compared with cell morphology or FCM control methods. The FCM NRBC staging method was established through the identification of different NRBC populations following the novel staining and lysing method. To evaluate the method further, we sorted samples containing NRBCs using a FACSort and investigated NRBC staging on the Sysmex XE-2100TM based on the cell sorting results. Data were analysed using special software (ida). First, we used the data in various parameter combinations. We then established gates to classify the NRBC populations. Finally, we analysed blood specimens from patients with different types of diseases to explore possible clinical applications. [source] Profiling bacterial survival through a water treatment process and subsequent distribution systemJOURNAL OF APPLIED MICROBIOLOGY, Issue 1 2005D. Hoefel Abstract Aims:, To profile fractions of active bacteria and of bacteria culturable with routine heterotrophic plate count (HPC) methods through a typical water treatment process and subsequent distribution system. In doing so, investigate how water treatment affects both bacterial abundance and diversity, and reveal the identities of active bacteria not detected by traditional HPC culture. Methods and Results:, Profiling active fractions was performed by flow cytometric cell sorting of either membrane-intact (BacLightTM kit) or enzymatically active (carboxyfluorescein diacetate, CFDA) bacteria, followed by eubacterial 16S rDNA-directed PCR and denaturing gradient gel electrophoresis (DGGE). Water treatment significantly reduced active bacterial numbers detected by the BacLightTM kit and CFDA assay by 2·89 and 2·81 log respectively. Bacterial diversity was also reduced from >20 DGGE bands in the active fractions of reservoir water to only two bands in the active fractions of finished water. These two bands represented Stenotrophomonas maltophila, initially culturable by HPC, and a Burkholderia -related species. Both species maintained measurable traits of physiological activity in distribution system bulk water but were undetected by HPC. Conclusions:, Flow cytometric cell sorting with PCR-DGGE, to assess water treatment efficacy, identified active bacteria from a variety of major phylogenetic groups undetected by routine HPC. Following treatment S. maltophila and a Burkholderia -related species retained activity and entered distribution undetected by HPC. Significance and Impact of the Study:, Methods used here demonstrate how water treatment operators can better monitor water treatment plant efficacy and assess distribution system instability by the detection and identification of active bacteria recalcitrant to routine HPC culture. [source] A novel direct aerodynamically assisted threading methodology for generating biologically viable microthreads encapsulating living primary cellsJOURNAL OF APPLIED POLYMER SCIENCE, Issue 2 2008Sumathy Arumuganathar Abstract In a recent discovery, coaxial electrospinning was explored to encapsulate living organisms within a continuous bio-polymeric microthread from which active biological scaffolds were fabricated (Townsend-Nicholson and Jayasinghe, Biomacromolecules 2006, 7, 3364). The cells were demonstrated to have gone through all expected cellular activity without their viability being compromised. These biologically active threads and scaffolds have direct and tremendous applicability from regenerative to therapeutic medicine. Currently these post-processed cells as composite threads and scaffolds are being investigated in-depth at a cellular level to establish if the processing methodology has any affect on the cellular make-up. We now demonstrate a competing non-electric field driven approach for fabricating composite threads and scaffolds influenced only by a differential pressure. We refer to this novel composite thread to scaffold fabrication methodology as coaxial aerodynamically assisted bio-threading (CAABT). Our investigations firstly, demonstrate that this technique can process handle living organisms without biologically perturbing them in anyway. Secondly the process is elucidated as possessing the ability to form composite active threads from which biologically viable scaffolds are formed. Finally our study employs florescent activated cell sorting (FACScan), a method by which the cellular dynamics and viability are quantified on control and threaded cellular samples at two prescribed time points. In parallel with FACScan, optical comparison of cellular morphology at three time points within a period of three weeks is carried out to photographically observe any changes in the post-processed cellular phenotype. Our developmental investigations into this novel aerodynamically assisted threading methodology has unearthed a unique biomicrofabrication approach, which joins cell electrospinning in the cell threading to scaffold fabrication endeavor. © 2007 Wiley Periodicals, Inc. J Appl Polym Sci, 2008 [source] RANK Expression as a Cell Surface Marker of Human Osteoclast Precursors in Peripheral Blood, Bone Marrow, and Giant Cell Tumors of BoneJOURNAL OF BONE AND MINERAL RESEARCH, Issue 9 2006Gerald J Atkins Abstract RANK expression in vivo on hematopoietic subsets including pre-osteoclasts, identified by monoclonal antibodies, has not been described. We describe the lineages that express RANK in bone marrow, peripheral blood, and GCTs. We show that CD14+RANKhigh cells constitute a circulating pre-osteoclast pool. Introduction: The expression of RANK by subsets of hematopoietic cells has not been adequately studied in humans. While attributed to the monocytoid lineage, the phenotype of the pre-osteoclast (pre-OC) with respect to RANK expression in vivo remains unclear. We tested monoclonal antibodies (MAbs) raised against the extracellular domain of recombinant human RANK for reactivity with normal peripheral blood (PB) and bone marrow (BM) mononuclear cells (PBMNCs and BMMNCs, respectively). We also tested reactivity with giant cell tumor cells (GCT), a confirmed source of pre-OC and mature OCs. Materials and Methods: Human PBMNCs, BMMNCs, and GCT cells were analyzed for reactivity with anti-RANK MAbs by flow cytometry in combination with hematopoietic lineage restricted markers. GCTs were also analyzed by immunofluorescence. CD14+ monocytoid cells were sorted by fluorescence-activated cell sorting (FACS) based on their relative RANK expression and cultured under OC-forming conditions. Results: RANK+ cells were detected similarly by three independent anti-RANK MAbs. One MAb (80736) immunoprecipitated RANK,RANKL complexes from surface-biotinylated GCT lysates. Using dual-color flow cytometry, RANK was detected on CD14+ (monocytoid), CD19+ (B-lymphoid), CD56+ (NK cell), and glycophorin A+ erythroid progenitors. Minor populations of both CD3+ T lymphocytes and BM CD34+ hematopoietic progenitors also expressed cell surface RANK. In GCTs, RANK expression was identified on mononuclear CD45+CD14+,V,3+c-Fms+ cells, likely to be committed pre-OC, and on multinucleated CD45+,V,3+TRACP+ OCs. Importantly, sorted CD14+RANKhigh PBMNCs treated with recombinant RANKL and macrophage-colony stimulating factor (M-CSF) gave rise to approximately twice the number of osteoclasts than RANKmid or RANKlow cells. Conclusions: These results suggest that committed monocytoid RANK+ pre-OCs are represented in the marrow and circulate in the periphery, forming a pool of cells capable of responding rapidly to RANKL. The ability to reliably detect committed pre-OC in peripheral blood could have important clinical applications in the management of diseases characterized by abnormal osteoclastic activity. [source] Fibrodysplasia Ossificans Progressiva (FOP), a Disorder of Ectopic Osteogenesis, Misregulates Cell Surface Expression and Trafficking of BMPRIA,,JOURNAL OF BONE AND MINERAL RESEARCH, Issue 7 2005Lourdes Serrano de la Peña Abstract FOP is a disorder in which skeletal muscle is progressively replaced with bone. FOP lymphocytes, a model system for exploring the BMP pathway in these patients, exhibit a defect in BMPRIA internalization and increased activation of downstream signaling, suggesting that altered BMP receptor trafficking underlies ectopic bone formation in this disease. Introduction: Fibrodysplasia ossificans progressiva (FOP) is a severely disabling disorder characterized by progressive heterotopic ossification of connective tissues. Whereas the genetic defect and pathophysiology of this condition remain enigmatic, BMP4 mRNA and protein are overexpressed, and mRNAs for a subset of secreted BMP antagonists are not synthesized at appropriate levels in cultured lymphocytes from FOP patients. These data suggest involvement of altered BMP signaling in the disease. In this study, we investigate whether the abnormality is associated with defective BMP receptor function in lymphocytes. Materials and Methods: Cell surface proteins were quantified by fluorescence-activated cell sorting (FACS). Protein phosphorylation was assayed by immunoprecipitation and immunoblotting. Protein synthesis and degradation were examined by [35S]methionine labeling and pulse-chase assays. mRNA was detected by RT-PCR. Results: FOP lymphocytes expressed 6-fold higher levels of BMP receptor type IA (BMPRIA) on the cell surface compared with control cells and displayed a marked reduction in ligand-stimulated internalization and degradation of BMPRIA. Moreover, in control cells, BMP4 treatment increased BMPRIA phosphorylation, whereas BMPRIA showed ligand-insensitive constitutive phosphorylation in FOP cells. Our data additionally support that the p38 mitogen-activated protein kinase (MAPK) signaling pathway is a major BMP signaling pathway in these cell lines and that expression of inhibitor of DNA binding and differentiation 1 (ID-1), a transcriptional target of BMP signaling, is enhanced in FOP cells. Conclusions: These data extend our previous observations of misregulated BMP4 signaling in FOP lymphocytes and show that cell surface overabundance and constitutive phosphorylation of BMPRIA are associated with a defect in receptor internalization. Altered BMP receptor trafficking may play a significant role in FOP pathogenesis. [source] | ||||||||||||||||||||||||||||||||||