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Results for “stem cell model of development”

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PubMed2002

Adult hematopoietic stem cells provide functional hemangioblast activity during retinal neovascularization.

Adults maintain a reservoir of hematopoietic stem cells that can enter the circulation to reach organs in need of regeneration. We developed a novel model of retinal neovascularization in adult mice to examine the role of hematopoietic stem cells in revascularizing ischemic retinas. Adult mice were durably engrafted with hematopoietic stem cells isolated from transgenic mice expressing green fluorescent protein. We performed serial long-term transplants, to ensure activity arose from self-renewing stem cells, and single hematopoietic stem-cell transplants to show clonality. After durable hematopoietic engraftment was established, retinal ischemia was induced to promote neovascularization. Our results indicate that self-renewing adult hematopoietic stem cells have functional hemangioblast activity, that is, they can clonally differentiate into all hematopoietic cell lineages as well as endothelial cells that revascularize adult retina. We also show that recruitment of endothelial precursors to sites of ischemic injury has a significant role in neovascularization.

Animals↗
PubMed2005

Different subsets of primary chronic myeloid leukemia stem cells engraft immunodeficient mice and produce a model of the human disease.

Xenograft models of chronic phase human chronic myeloid leukemia (CML) have been difficult to develop because of the persistence of normal hematopoietic stem cells in most chronic phase CML patients and the lack of methods to selectively isolate the rarer CML stem cells. To circumvent this problem, we first identified nine patients' samples in which the long-term culture-initiating cells were predominantly leukemic and then transplanted cells from these samples into sublethally irradiated NOD/SCID and NOD/SCID-beta2microglobulin-/- mice. This resulted in the consistent and durable (>5 months) repopulation of both host genotypes with similar numbers of BCR-ABL+/Ph+ cells. The regenerated leukemic cells included an initial, transient population derived from CD34+CD38+ cells as well as more sustained populations derived from CD34+CD38- progenitors, indicative of a hierarchy of transplantable leukemic cells. Analysis of the phenotypes produced revealed a reduced output of B-lineage cells, enhanced myelopoiesis with excessive production of erythroid and megakaropoietic cells and the generation of primitive (CD34+) leukemic cells displaying an autocrine IL-3 and G-CSF phenotype, all characteristics of primary CML cells. These findings demonstrate the validity of this xenograft model of chronic phase human CML, which should enable future investigation of disease pathogenesis and new approaches to therapy.

Animals↗
PubMed2001

Collagen IV synthesis is restricted to the enteroendocrine pathway during multilineage differentiation of human colorectal epithelial stem cells.

The human large intestine is lined by a rapidly renewing epithelial monolayer where cell loss is precisely balanced with cell production. The continuous supply of new cells is produced by undifferentiated multipotent stem cells via a coordinated program of proliferation and differentiation yielding three epithelial lineages: absorptive, goblet and enteroendocrine. Cell-matrix interactions have been suggested to be regulators of the multilineage differentiation program of the colorectal crypt but the expression of matrix proteins or their receptors does not appear to have the subtlety expected for this task. We have developed an in vitro model system of intestinal epithelial stem cells to facilitate the direct analysis of stem cells undergoing lineage commitment and differentiation. Using this culture system, we can now directly investigate the role of cell-matrix signalling in stem-cell decisions. In this study, collagen-IV synthesis has been followed in monolayers of multipotent cells that have been induced to differentiate into absorptive, goblet and enteroendocrine cells. Our experiments demonstrate that commitment to the enteroendocrine lineage is specifically accompanied by the expression of type-IV collagen that remains enteroendocrine-cell associated. Undifferentiated cells, absorptive cells and goblet cells do not express collagen IV. To confirm that the differential lineage-specific expression of collagen IV observed in the model system was representative of the in vivo situation, collagen-IV synthesis was analysed in isolated human colorectal crypts and tissue sections using immunocytochemistry and in situ hybridisation. These studies confirmed the in vitro findings, in that implementation of the enteroendocrine differentiation program involves synthesis and accumulation of a collagen-IV matrix. Thus, human colorectal enteroendocrine cells are unique in the colorectal crypt in that they assemble a cell-associated collagen-IV-rich matrix not observed on other colorectal epithelial cells. This study provides the first evidence for differential matrix synthesis between colorectal epithelial lineages in human colorectal epithelium. The specialised pericellular environment of the enteroendocrine cells might explain some of the unique phenotypic characteristics of this cell lineage. Furthermore, these findings suggest a potential mechanism whereby individual epithelial cells could modulate their cell-matrix signalling even while rapidly migrating in heterogeneous sheets over a shared basement membrane.

Cell Differentiation↗
PubMed1987

Transformation of a rat liver cell line: neoplastic phenotype and regulation of gamma glutamyl transpeptidase in tumour tissue.

A continuously dividing but non-transformed rat cell line was transformed by ras oncogenes and by aflatoxin. Transformed lines grew as solid tumours in nude mice with a short latency period. Different phenotypes were obtained from the transformed clones, and covered a wide spectrum of tumour types. While the commonest tumour type obtained was undifferentiated with unorganized spindle shaped and rounded cells, tumours with well formed trabeculae and adenomatous differentiation were also obtained. Gamma glutamyl transpeptidase (GGT), which showed an increase in transformed cells in tissue culture showed a variable expression in nude mouse tumours. While GGT positive cells were seen in all types of tumours they followed no consistent pattern and GGT negative areas were seen with a high frequency. However, in tumours showing adenomatous differentiation it was confined to the membrane on the luminal surface of the glandular structures. The different types of tumours resulting from the transformation of a single cell line provide a model of in vivo tumour development, supporting the possibility of a single stem cell origin of different types of liver neoplasms. This system also demonstrates the complexity of the regulation of GGT in tumour tissue. The influence of GGT on tumorigenesis has also been studied.

Adenoma↗
PubMedDate not supplied

Effects of oxygen transport on 3-d human mesenchymal stem cell metabolic activity in perfusion and static cultures: experiments and mathematical model.

Human mesenchymal stem cells (hMSCs) have unique potential to develop into functional tissue constructs to replace a wide range of tissues damaged by disease or injury. While recent studies have highlighted the necessity for 3-D culture systems to facilitate the proper biological, physiological, and developmental processes of the cells, the effects of the physiological environment on the intrinsic tissue development characteristics in the 3-D scaffolds have not been fully investigated. In this study, experimental results from a 3-D perfusion bioreactor system and the static culture are combined with a mathematical model to assess the effects of oxygen transport on hMSC metabolism and proliferation in 3-D constructs grown in static and perfusion conditions. Cells grown in the perfusion culture had order of magnitude higher metabolic rates, and the perfusion culture supports higher cell density at the end of cultivation. The specific oxygen consumption rate for the constructs in the perfusion bioreactor was found to decrease from 0.012 to 0.0017 micromol/10(6) cells/h as cell density increases, suggesting intrinsic physiological change at high cell density. BrdU staining revealed the noneven spatial distribution of the proliferating cells in the constructs grown under static culture conditions compared to the cells that were grown in the perfusion system. The hypothesis that the constructs in static culture grow under oxygen limitation is supported by higher Y(L/G) in static culture. Modeling results show that the oxygen tension in the static culture is lower than that of the perfusion unit, where the cell density was 4 times higher. The experimental and modeling results show the dependence of cell metabolism and spatial growth patterns on the culture environment and highlight the need to optimize the culture parameters in hMSC tissue engineering.

Biological Transport↗
PubMed2005

Reconstitution of hematopoiesis following intrauterine transplantation of stem cells.

In utero hematopoietic stem cell transplantation is an entirely nonmyeloablative approach to achieve mixed hematopoietic chimerism and associated donor-specific tolerance. This chapter provides the rationale and methodologic detail for the administration of stem cells to the "preimmune" mouse fetus by a variety of routes. The development of murine model systems for in utero transplantation has accelerated progress in the field of in utero hematopoietic stem cell transplantation. Creative use of these models should also have experimental application to the fields of fetal gene therapy, stem cell biology, and developmental biology.

Animals↗
PubMed2006

Muscling in on stem cells.

Skeletal muscle is one of the few adult tissues that possesses the capacity for regeneration (restoration of lost functional tissue) as opposed to repair. This capacity is due to the presence of 'muscle stem cells' known as satellite cells. Detailed investigation of these cells over the past 50 years has revealed that both these and other cells within the skeletal muscle complex are capable of regenerating both muscle and other cell types as well. Here, we review this information, and suggest that skeletal muscle is an exciting reservoir of cells for regenerating skeletal muscle itself, as well as other cell types.

Adipose Tissue↗
PubMed2006

Routine culture and differentiation of human embryonic stem cells.

Human embryonic stem cells provide both an in vitro model of human development and a potential source of cells for treatment of degenerative, metabolic, or traumatic disorders. This chapter describes techniques for routine maintenance and differentiation of human embryonic stem cells in culture.

Animals↗
PubMed1997

Myelination following transplantation of EGF-responsive neural stem cells into a myelin-deficient environment.

Epidermal growth factor (EGF)-responsive stem cells have been identified in the murine central nervous system. These cells can be isolated from the brain and maintained in an undifferentiated state in vitro in the presence of EGF. After removing EGF, the cells cease mitosis and can be induced to differentiate into neurons, astrocytes, and oligodendrocytes. We demonstrate that when the undifferentiated stem cells (nestin-positive) are injected into the myelin-deficient rat spinal cord, they respond to cues within the mutant CNS and differentiate into myelinating oligodendrocytes, in contrast to their behavior in vitro, where they mainly form astrocytes. The cells provide a valuable model system for the study of the development of early oligodendrocytes from multipotent neural stem cells. Because these cells are influenced to divide using growth factors, rather than oncogenes, and because they appear to make appropriate lineage decisions when transplanted into a mutant environment, they may provide an excellent source of cells for a variety of future therapies using cellular transplantation.

Animals↗
PubMed1994

Bone marrow culture and transduction of stem cells in a miniature swine model.

Recombinant retroviral vectors, engineered to express the beta-chain gene of swine major histocompatibility complex class II DR, were developed for the genetic modification of swine hematopoietic stem cells (HSC). The expression of these vectors in swine bone marrow has been studied both in culture and after bone marrow transplantation. In addition, myeloid progenitor colony assays were performed on swine umbilical cord blood as part of a study to identify alternative sources of HSC for somatic gene transfer, revealing the presence of both granulocyte macrophage colony forming-units (CFU-GM) and CFU-Mix at frequencies comparable to those found in juvenile swine bone marrow.

Age Factors↗
PubMed1999

Role of c-kit receptor tyrosine kinase in development of oval cells in the rat 2-acetylaminofluorene/partial hepatectomy model.

Oval cells that develop in the rat 2-acetylaminofluorene/partial hepatectomy (AAF/PH) model express the c-kit receptor tyrosine kinase (KIT) and its ligand, stem cell factor (SCF). We investigated the role of the SCF/KIT system in the development of oval cells using Ws/Ws rats, whose c-kit kinase activity was severely impaired owing to a small deletion in the kinase domain. On days 7, 9, and 13 after PH in the AAF/PH model, the development of oval cells was remarkably suppressed in Ws/Ws rats when compared with that of the control normal (+/+) rats. However, oval cells that developed in Ws/Ws rats expressed marker proteins of oval cells, such as alpha-fetoprotein (AFP), cytokeratin-19 (CK-19), and flt-3 receptor tyrosine kinase, similar to those of +/+ rats. Furthermore, labeling with [3H]-thymidine and immunostaining of Ki-67 showed that the proliferative activity of oval cells that developed in Ws/Ws rats was comparable with that of +/+ rats. The present results indicate that the signal transduction of the SCF/KIT system plays a crucial role in the development of oval cells, at least, in the rat AAF/PH model, and suggest that KIT-mediated signal transduction plays only a small role in determining the phenotype and in the proliferative activity of oval cells.

2-Acetylaminofluorene↗
PubMed2006

Embryonic stem cells: a novel tool for the study of antiangiogenesis and tumor-induced angiogenesis.

Major research initiatives in antiangiogenesis research have been undertaken to control angiogenic diseases such as polyarthritis, psoriasis, endometriosis, and diabetic retinopathy, and inhibition of tumor-induced angiogenesis has emerged as one of the most promising anti-cancer therapies currently available. Although several quantitative in vivo (i.e., animal models) as well as in vitro (i.e., pure endothelial cell cultures) angiogenesis assays have been described, the development of novel angiogenesis assays with organotypic culture systems that take into account oxygen and nutrient gradients, depth-dependent changes in intracellular pH and a redox state similar to that found in a natural tissue microenvironment are necessary to investigate blood vessel growth. Embryonic stem cells of mouse and human origin have the capacity to develop into three-dimensional tissues with functional capillaries, and this model system represents an excellent in vitro model for antiangiogenesis research. Upon confrontation of stem cells by co-culture with multicellular tumor spheroids, tumor-induced angiogenesis, i.e., the invasion of endothelial host-derived cells into a tumor tissue, can also be monitored. The current review provides an overview of embryonic stem cells as novel tools for antiangiogenesis research and outlines the use of confrontation cultures for the study of tumor-induced angiogenesis.

Angiogenesis Inhibitors↗
PubMed2003

Label-retaining cells in the rat pancreas: location and differentiation potential in vitro.

Islets of Langerhans are micro-organs scattered throughout the pancreas that contain insulin-producing cells, called beta-cells. Although new light has been recently shed on beta-cell development, information on the phenotype and location of beta-stem cells remains scarce. Here, we provide evidence that beta-stem cells are slow-cycling cells located within and around the islets of Langerhans. First, using a bromodeoxyuridine (BrdU) pulse/chase approach, we detected BrdU-retaining cells in vivo in the islet area of rat pancreata. These cells were negative for endocrine markers but expressed Pdx1, a marker for pancreatic stem cells. Next, using an in vitro model that mimicked endocrine cell development, we found that BrdU-retaining cells were capable of differentiating into beta-cells. Taken together, these observations demonstrate that BrdU retention is a property of beta-stem cells.

Animals↗
PubMed2006

New insights to vascular smooth muscle cell and pericyte differentiation of mouse embryonic stem cells in vitro.

OBJECTIVE: The molecular mechanisms that regulate pericyte differentiation are not well understood, partly because of the lack of well-characterized in vitro systems that model this process. In this article, we develop a mouse embryonic stem (ES) cell-based angiogenesis/vasculogenesis assay and characterize the system for vascular smooth muscle cell (VSMC) and pericyte differentiation. METHODS AND RESULTS: ES cells that were cultured for 5 days on OP9 stroma cells upregulated their transcription of VSMC and pericyte selective genes. Other SMC marker genes were induced at a later time point, which suggests that vascular SMC/pericyte genes are regulated by a separate mechanism. Moreover, sequence analysis failed to identify any conserved CArG elements in the vascular SMC and pericyte gene promoters, which indicates that serum response factor is not involved in their regulation. Gleevec, a tyrosine kinase inhibitor that blocks platelet-derived growth factor (PDGF) spell-receptor signaling, and a neutralizing antibody against transforming growth factor (TGF) beta1, beta2, and beta3 failed to inhibit the induction of vascular SMC/pericyte genes. Finally, ES-derived vascular sprouts recruited cocultured MEF cells to pericyte-typical locations. The recruited cells activated expression of a VSMC- and pericyte-specific reporter gene. CONCLUSIONS: We conclude that OP9 stroma cells induce pericyte differentiation of cocultured mouse ES cells. The induction of pericyte marker genes is temporally separated from the induction of SMC genes and does not require platelet-derived growth factor B or TGFbeta1 signaling.

Animals↗
PubMed1999

Immunodeficient mice as models of human hematopoietic stem cell engraftment.

The past year has brought forth some exciting developments in the use of murine xenotransplantation systems to study the biology and transduction of human hematopoietic stem cells. The effects of cytokines have been studied by injection into the mice or by treatment of the cell inoculum prior to injection. The importance of the cell cycle and integrin expression has been evaluated. New methods of gene therapy have been tested in xenograft models - including cell cycle manipulation and a promising new lentiviral vector system, based on HIV.

Animals↗
PubMed2004

Formation of human hepatocytes by human hematopoietic stem cells in sheep.

We took advantage of the proliferative and permissive environment of the developing preimmune fetus to develop a noninjury large animal model in sheep, in which the transplantation of defined populations of human hematopoietic stem cells resulted in the establishment of human hematopoiesis and led to the formation of significant numbers of long-lasting, functional human liver cells, with some animals exhibiting levels as high as 20% of donor (human) hepatocytes 11 months after transplantation. A direct correlation was found between hepatocyte activity and phenotype of transplanted cells, cell dose administered, source of cells used on a cell-per-cell basis (bone marrow, cord blood, mobilized peripheral blood), and time after transplantation. Human hepatocytes generated in this model retained functional properties of normal hepatocytes, constituted hepatic functional units with the presence of human endothelial and biliary duct cells, and secreted human albumin that was detected in circulation. Transplanting populations of hematopoietic stem cells can efficiently generate significant numbers of functional hepatic cells in this noninjury large animal model and thus could be a means of ameliorating or curing genetic diseases in which a deficiency of liver cells or their products threatens the life of the fetus or newborn.

ADP-ribosyl Cyclase↗
PubMed2002

Embryonic stem cells and mouse models of human syndromes: examples from the T-box gene family.

Synergism between the fields of reproduction and development has long been an important factor for advances in each field, and the development of gene modification technology in mammals has progressed hand in hand with advances in reproductive technology. The discovery and exploitation of embryonic stem cells for producing targeted gene mutations in the mouse, combined with advances in genome analysis, has provided a means of producing mouse models of human genetic diseases, including congenital defects. This review presents examples of how gene targeting and reproductive biology techniques are being applied to the production of mouse models for specific human developmental syndromes caused by mutations in genes of the T-box transcription factor gene family. The ulnar-mammary syndrome and the DiGeorge syndrome are two developmental syndromes that are currently being explored in this way.

Animals↗
PubMed2005

Reparative osteogenesis during transplantation of mesenchymal stem cells.

Reparative osteogenesis was studied after xenotransplantation of suspension cell graft from human mesenchymal stem cells. A model of experimental damage to rat femoral diaphysis was developed. The state of animals was satisfactory and non-depressed in the early and late postoperation period. We revealed no local pathological reactions and complications. Administration of mesenchymal stem cells into the area of bone defect accelerated and improved regeneration. Unilateral transplantation of the cell graft stimulated regeneration in the contralateral limb due to acceleration of bone tissue maturation. On day 90 after treatment the bone regenerate was completely developed in the area of defect in animals of various groups. The newly formed bone tissue was well integrated into the bone organ.

Animals↗