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Clonal proliferation of multipotent stem/progenitor cells in the neonatal and adult salivary glands 总被引:7,自引:0,他引:7
Kishi T Takao T Fujita K Taniguchi H 《Biochemical and biophysical research communications》2006,340(2):544-552
Salivary gland stem/progenitor cells are thought to be present in intercalated ductal cells, but the fact is unclear. In this study, we sought to clarify if stem/progenitor cells are present in submandibular glands using colony assay, which is one of the stem cell assay methods. Using a low-density culture of submandibular gland cells of neonatal rats, we developed a novel culture system that promotes single cell colony formation. Average doubling time for the colony-forming cells was 24.7 (SD=+/-7.02)h, indicating high proliferative potency. When epidermal growth factor (EGF) and hepatocyte growth factor (HGF) were added to the medium, the number of clonal colonies increased greater than those cultured without growth factors (13.2+/-4.18 vs. 4.5+/-1.73). The RT-PCR and immunostaining demonstrated expressing acinar, ductal, and myoepithelial cell lineage markers. This study demonstrated the presence of the salivary gland stem/progenitor cells that are highly proliferative and multipotent in salivary glands. 相似文献
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Neural stem/progenitor cell (NSPC) transplantation is a promising therapy for spinal cord injury (SCI). However, little is known about NSPC from the adult human spinal cord as a donor source. We demonstrate for the first time that multipotent and self-renewing NSPC can be cultured, passaged and transplanted from the adult human spinal cord of organ transplant donors. Adult human spinal cord NSPC require an adherent substrate for selection and expansion in EGF (epidermal growth factor) and FGF2 (fibroblast growth factor) enriched medium. NSPC as an adherent monolayer can be passaged for at least 9 months and form neurospheres when plated in suspension culture. In EGF/FGF2 culture, NSPC proliferate and primarily express nestin and Sox2, and low levels of markers for differentiating cells. Leukemia inhibitory factor (LIF) promotes NSPC proliferation and significantly enhances GFAP expression in hypoxia. In differentiating conditions in the presence of serum, these NSPC show multipotentiality, expressing markers of neurons, astrocytes, and oligodendrocytes. Dibutyryl cyclic AMP (dbcAMP) significantly enhances neuronal differentiation. We transplanted the multipotent NSPC into SCI rats and show that the xenografts survive, are post-mitotic, and retain the capacity to differentiate into neurons and glia.Together, these findings reveal that multipotent self-renewing NSPC cultured and passaged from adult human spinal cords of organ transplant donors, respond to exogenous factors that promote selective differentiation, and survive and differentiate after transplantation into the injured spinal cord. 相似文献
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de la Fuente R Abad JL García-Castro J Fernández-Miguel G Petriz J Rubio D Vicario-Abejón C Guillén P González MA Bernad A 《Experimental cell research》2004,297(2):313-328
OBJECTIVE: To test the hypothesis that dedifferentiated adult human cartilage chondrocytes (HAC) are a true multipotent primitive population. METHODS: Studies to characterize dedifferentiated HAC included cell cycle and quiescence analysis, cell fusion, flow-FISH telomere length assays, and ABC transporter analysis. Dedifferentiated HAC were characterized by flow cytometry, in parallel with bone marrow mesenchymal stem cells (MSC) and processed lipoaspirate (PLA) cells. The in vitro differentiation potential of dedifferentiated HAC was studied by cell culture under several inducing conditions, in multiclonal and clonal cell populations. RESULTS: Long-term HAC cultures were chromosomically stable and maintained cell cycle dynamics while showing telomere shortening. The phenotype of dedifferentiated HAC was quite similar to that of human bone marrow MSC. In addition, this population expressed human embryonic stem cell markers. Multiclonal populations of dedifferentiated HAC differentiated to chondrogenic, osteogenic, adipogenic, myogenic, and neurogenic lineages. Following VEGF induction, dedifferentiated HAC expressed characteristics of endothelial cells, including AcLDL uptake. A total of 53 clonal populations of dedifferentiated HAC were efficiently expanded; 17 were able to differentiate to chondrogenic, osteogenic, and adipogenic lineages. No correlation was observed between telomere length or quiescent population and differentiation potential in the clones assayed. CONCLUSION: Dedifferentiated HAC should be considered a human multipotent primitive population. 相似文献
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Elisabeth Lehmberg Joseph A. Traina John A. Chakel Ray-Jen Chang Maria Parkman Michael T. McCaman Peter K. Murakami Vafa Lahidji Jeffrey W. Nelson William S. Hancock Eirik Nestaas Erno PungorJr. 《Journal of chromatography. B, Analytical technologies in the biomedical and life sciences》1999,732(2):7
An RP-HPLC assay was developed for a recombinant adenovirus type 5. During chromatography, intact adenovirus dissociated into its structural components (DNA and proteins) and the viral proteome was separated yielding a characteristic fingerprint. The individual components were identified by matrix-assisted laser desorption ionization time-of-flight mass spectroscopy, N-terminal sequencing and amino acid composition. The assay was utilized to measure adenovirus particle concentration through quantification of structural proteins. Each structural protein provided independent measurement of virus concentration allowing verification of accuracy. The assay sensitivity is at or below 2·108 particles. Contrary to the benchmark spectrophotometric assay, the RP-HPLC assay was shown to be insensitive to contaminants common for partially purified adenovirus preparations. 相似文献
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Generation and expansion of multipotent mesenchymal progenitor cells from cultured human pancreatic islets 总被引:1,自引:0,他引:1
Gallo R Gambelli F Gava B Sasdelli F Tellone V Masini M Marchetti P Dotta F Sorrentino V 《Cell death and differentiation》2007,14(11):1860-1871
Cellular models and culture conditions for in vitro expansion of insulin-producing cells represent a key element to develop cell therapy for diabetes. Initial evidence that human beta-cells could be expanded after undergoing a reversible epithelial-mesenchymal transition has been recently negated by genetic lineage tracing studies in mice. Here, we report that culturing human pancreatic islets in the presence of serum resulted in the emergence of a population of nestin-positive cells. These proliferating cells were mainly C-peptide negative, although in the first week in culture, proliferating cells, insulin promoter factor-1 (Ipf-1) positive, were observed. Later passages of islet-derived cells were Ipf-1 negative and displayed a mesenchymal phenotype. These human pancreatic islet-derived mesenchymal (hPIDM) cells were expanded up to 10(14) cells and were able to differentiate toward adipocytes, osteocytes and chondrocytes, similarly to mesenchymal stem/precursor cells. Interestingly, however, under serum-free conditions, hPIDM cells lost the mesenchymal phenotype, formed islet-like clusters (ILCs) and were able to produce and secrete insulin. These data suggest that, although these cells are likely to result from preexisting mesenchymal cells rather than beta-cells, hPIDM cells represent a valuable model for further developments toward future replacement therapy in diabetes. 相似文献
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Pax6 is required for the multipotent state of retinal progenitor cells 总被引:29,自引:0,他引:29
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Lee MW Moon YJ Yang MS Kim SK Jang IK Eom YW Park JS Kim HC Song KY Park SC Lim HS Kim YJ 《Biochemical and biophysical research communications》2007,358(2):637-643
Umbilical cord blood (UCB) is a rich source of hematopoietic stem cells, with practical and ethical advantages. To date, the presence of other stem cells in UCB remains to be established. We investigated whether other stem cells are present in cryopreserved UCB. Seeded mononuclear cells formed adherent colonized cells in optimized culture conditions. Over a 4- to 6-week culture period, colonized cells gradually developed into adherent mono-layer cells, which exhibited homogeneous fibroblast-like morphology and immunophenotypes, and were highly proliferative. Isolated cells were designated 'multipotent progenitor cells (MPCs)'. Under appropriate conditions for 2 weeks, MPCs differentiated into neural tissue-specific cell types, including neuron, astrocyte, and oligodendrocyte. Differentiated cells presented their respective markers, specifically, NF-L and NSE for neurons, GFAP for astrocytes, and myelin/oligodendrocyte for oligodendrocytes. In this study, we successfully isolated MPCs from cryopreserved UCB, which differentiated into the neural tissue-specific cell types. These findings suggest that cryopreserved human UCB is a useful alternative source of neural progenitor cells, such as MPCs, for experimental and therapeutic applications. 相似文献
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Pei-Min Chen Men-Luh Yen Ko-Jiunn Liu Huey-Kang Sytwu B-Linju Yen 《Journal of biomedical science》2011,18(1):49
In recent years, a large number of studies have contributed to our understanding of the immunomodulatory mechanisms used by
multipotent mesenchymal stem cells (MSCs). Initially isolated from the bone marrow (BM), MSCs have been found in many tissues
but the strong immunomodulatory properties are best studied in BM MSCs. The immunomodulatory effects of BM MSCs are wide,
extending to T lymphocytes and dendritic cells, and are therapeutically useful for treatment of immune-related diseases including
graft-versus-host disease as well as possibly autoimmune diseases. However, BM MSCs are very rare cells and require an invasive
procedure for procurement. Recently, MSCs have also been found in fetal-stage embryo-proper and extra-embryonic tissues, and
these human fetal MSCs (F-MSCs) have a higher proliferative profile, and are capable of multilineage differentiation as well
as exert strong immunomodulatory effects. As such, these F-MSCs can be viewed as alternative sources of MSCs. We review here
the current understanding of the mechanisms behind the immunomodulatory properties of BM MSCs and F-MSCs. An increase in our
understanding of MSC suppressor mechanisms will offer insights for prevalent clinical use of these versatile adult stem cells
in the near future. 相似文献
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Azan G Low WC Wendelschafer-Crabb G Ikramuddin S Kennedy WR 《Cell and tissue research》2011,344(2):217-225
Putative neural stem cells have been identified within the enteric nervous system (ENS) of adult rodents and cultured from
human myenteric plexus. We conducted studies to identify neural stem cells or progenitor cells within the submucosa of adult
human ENS. Jejunum tissue was removed from adult human subjects undergoing gastric bypass surgery. The tissue was immunostained,
and confocal images of ganglia in the submucosal plexus were collected to identify protein gene product 9.5 (PGP 9.5) - immunoractive
neurons and neuronal progenitor cells that coexpress PGP 9.5 and nestin. In addition to PGP-9.5-positive/nestin-negative neuronal
cells within ganglia, we observed two other types of cells: (1) cells in which PGP 9.5 and nestin were co-localized, (2) cells
negative for both PGP 9.5 and nestin. These observations suggest that the latter two types of cells are related to a progenitor
cell population and are consistent with the concept that the submucosa of human adult ENS contains stem cells capable of maintenance
and repair within the peripheral nervous system. 相似文献
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Di Palma S Stange D van de Wetering M Clevers H Heck AJ Mohammed S 《Journal of proteome research》2011,10(8):3814-3819
In proteomics, multidimensional liquid chromatography combined with mass spectrometry has become a standard technique to reduce sample complexity and tackle the vast dynamic range. Such fractionation is necessary to obtain a comprehensive analysis of biological samples such as tissues and cell lines. However, extensive fractionation comes at the expense of sample losses, hampering the analysis of limited material. We previously described a highly sensitive multidimensional chromatographic strategy based on a combination of hydrophilic interaction liquid chromatography and reversed phase chromatography, which allows proteomic analysis with minimal sample losses. Here we apply this strategy to the analysis of a limited number of FACS-sorted colon stem cells extracted from mouse intestine, obtaining a proteome coverage comparable to current methods that generally require 100-fold more starting material. We propose that this alternative multidimensional chromatographic technology will find ample application such as in the analysis of distinct cellular populations obtained by laser microdissection. 相似文献
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Stefan Radtke André Görgens Bing Liu Peter A. Horn 《Cell cycle (Georgetown, Tex.)》2016,15(4):540-545
A major goal in haematopoietic stem cell (HSC) research is to define conditions for the expansion of HSCs or multipotent progenitor cells (MPPs). Since human HSCs/MPPs cannot be isolated, NOD/SCID repopulating cell (SRC) assays emerged as the standard for the quantification of very primitive haematopoietic cell. However, in addition to HSCs/MPPs, lympho-myeloid primed progenitors (LMPPs) were recently found to contain SRC activities, challenging this assay as clear HSC/MPP readout. Because our revised model of human haematopoiesis predicts that HSCs/MPPs can be identified as CD133+CD34+ cells containing erythroid potentials, we investigated the potential of human mesenchymal and conventional murine stromal cells to support expansion of HSCs/MPPs. Even though all stromal cells supported expansion of CD133+CD34+ progenitors with long-term myeloid and long-term lymphoid potentials, erythroid potentials were exclusively found within erythro-myeloid CD133lowCD34+ cell fractions. Thus, our data demonstrate that against the prevailing assumption co-cultures on human mesenchymal and murine stromal cells neither promote expansion nor maintenance of HSCs and MPPs. 相似文献
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Jacobs JM Mottaz HM Yu LR Anderson DJ Moore RJ Chen WN Auberry KJ Strittmatter EF Monroe ME Thrall BD Camp DG Smith RD 《Journal of proteome research》2004,3(1):68-75
Recent multidimensional liquid chromatography MS/MS studies have contributed to the identification of large numbers of expressed proteins for numerous species. The present study couples size exclusion chromatography of intact proteins with the separation of tryptically digested peptides using a combination of strong cation exchange and high resolution, reversed phase capillary chromatography to identify proteins extracted from human mammary epithelial cells (HMECs). In addition to conventional conservative criteria for protein identifications, the confidence levels were additionally increased through the use of peptide normalized elution times (NET) for the liquid chromatographic separation step. The combined approach resulted in a total of 5838 unique peptides identified covering 1574 different proteins with an estimated 4% gene coverage of the human genome, as annotated by the National Center for Biotechnology Information (NCBI). This database provides a baseline for comparison against variations in other genetically and environmentally perturbed systems. Proteins identified were categorized based upon intracellular location and biological process with the identification of numerous receptors, regulatory proteins, and extracellular proteins, demonstrating the usefulness of this application in the global analysis of human cells for future comparative studies. 相似文献
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André G?rgens Stefan Radtke Peter A Horn Bernd Giebel 《Cell cycle (Georgetown, Tex.)》2013,12(22):3478-3482
Three important goals of hematopoietic stem cell research are to understand of how hematopoietic stem cells (HSCs) self-renew, how lineage commitment takes place, and how HSCs can be expanded ex vivo. Research in this area requires a reliable model of hematopoiesis. Performing detailed functional analyses of human hematopoietic progenitor subsets, we recently gained evidence for new hematopoietic lineage relationships.1 According to our data, neutrophils belong to the same branch of the hematopoietic tree as lymphocytes. In contrast, eosinophils and basophils derive from another branch, the erythro-myeloid branch. Here, after introducing the newly proposed hematopoietic model, we discuss its consequences for the identification and expansion of human multipotent progenitors and suggest a fast and reliable method to screen for multipotent hematopoietic cells in vitro. 相似文献
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A method for the quantitative determination of ammonium in human urine by high-performance liquid chromatography (HPLC) is described. After making fluorescent substances with fluorescamine, they were separated and quantified by their fluorometric intensity. The intensity (as measured by peak height) was linear between 0.5 and 5.0 micrograms, and coefficients of variation for elution time and peak height on replicate analysis of the standard were 0.15 and 4.2%, respectively. Recoveries of added ammonium were 96.5 and 97.3%, respectively, on 2.0 and 3.0 micrograms by this method. Detection limit of this method was 0.2 microgram. There was good agreement between the proposed HPLC method (X) and ion chromatographic method (Y), giving the relationships as Y = 0.956X + 0.012, r = 0.991. 相似文献
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Shinoda K Sugimoto M Yachie N Sugiyama N Masuda T Robert M Soga T Tomita M 《Journal of proteome research》2006,5(12):3312-3317
We developed a computational method to predict the retention times of peptides in HPLC using artificial neural networks (ANN). We performed stepwise multiple linear regressions and selected for ANN input amino acids that significantly affected the LC retention time. Unlike conventional linear models, the trained ANN accurately predicted the retention time of peptides containing up to 50 amino acid residues. In 834 peptides, there was a strong correlation (R2 = 0.928) between measured and predicted retention times. We demonstrated the utility of our method by the prediction of the retention time of 121,273 peptides resulting from LysC-digestion of the Escherichia coli proteome. Our approach is useful for the proteome-wide characterization of peptides and the identification of unknown peptide peaks obtained in proteome analysis. 相似文献
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For more than a decade, we have known that the human brain harbors progenitor cells capable of becoming mature neurons in the adult human brain. Since the original landmark article by Eriksson et al. in 1998 (Nat Med 4:1313-1317), there have been many studies investigating the effect that depression, epilepsy, Alzheimer's disease, Huntington's disease, and Parkinson's disease have on the germinal zones in the adult human brain. Of particular interest is the demonstration that there are far fewer progenitor cells in the hippocampal subgranular zone (SGZ) compared with the subventricular zone (SVZ) in the human brain. Furthermore, the quantity of progenitor cell proliferation in human neurodegenerative diseases differs from that of animal models of neurodegenerative diseases; there is minimal progenitor proliferation in the SGZ and extensive proliferation in the SVZ in the human. In this review, we will present the data from a range of human and rodent studies from which we can compare the amount of proliferation of cells in the SVZ and SGZ in different neurodegenerative diseases. 相似文献
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BACKGROUND: Multipotent adult progenitor cells (MAPC) comprise interesting candidates for myocardial regeneration because of a broad differentiation ability and immune privilege. We aimed to compare the improvement of cardiac function by syngeneic and allogeneic MAPC produced on a large scale using a platform optimized from MAPC research protocols. METHODS: Myocardial infarction was induced in Lewis rats by direct left anterior descending ligation followed immediately by direct injection into the infarct border zone of either Sprague-Dawley or Lewis MAPC from large-scale expansions. Echocardiography was performed to evaluate improvement in cardiac function, and immunohistochemistry was performed to identify MAPC within the infarct zone. RESULTS: Significant increases were observed in functional performance in animals transplanted with expanded MAPC compared with saline controls, with no significant differences between the syngeneic and allogeneic groups. Immunostaining demonstrated significant engraftment of expanded MAPC at 1 day after acute myocardial infarction, with <10% of either syngeneic or allogeneic cells remaining at 6 weeks. At this point there was no evidence of myocardial regeneration. However, a significant increase in vascular density within the infarct zone in MAPC-transplanted animals was observed, and MAPC were found to produce high levels of VEGF in culture. DISCUSSION: These findings support a model in which delivery of expanded MAPC following acute myocardial infarction results in improvement in cardiac function because of paracrine effects resulting in vascular density increases, as well as potentially other trophic effects, supporting newly injured cardiac myocytes. Thus transplantation with MAPC may represent a promising therapeutic strategy with application in the stimulation of neovascularization in ischemic heart disease. 相似文献