SEARCH · PubMed Health

Results for “Cell Tracking”

Explore indexed PubMed citations for clinical trials, systematic reviews and public health research. Read source abstracts and follow each citation to its original PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 217 records · Page 12Linked to original sources

PubMed1987

Methodology for detection of heterogeneity of cell locomotory phenotypes in three-dimensional gels.

A computer-driven, three-dimensional, optical, cell-tracking system was used to quantify the locomotory phenotypes of Mos-11 cells in a collagen gel. Factor analysis was used as a statistical means to make the complex variables generated by the system more tractable to analysis. Heterogeneity in locomotory behavior (two independent locomotory phenotypes) was detected among a sample of 54 cells.

Cell Line↗

PubMed2004

Directed differentiation of rhesus monkey ES cells into pancreatic cell phenotypes.

Embryonic stem cells (ES) can self-replicate and differentiate into all cell types including insulin-producing, beta-like cells and could, therefore, be used to treat diabetes mellitus. To date, results of stem cell differentiation into beta cells have been debated, largely due to difficulties in defining the identity of a beta cell. We have recently differentiated non-human primate (rhesus) embryonic stem (rES) cell lines into insulin producing, beta-like cells with the beta cell growth factor, Exendin-4 and using C-peptide as a phenotype marker. Cell development was characterized at each stage by gene and protein expression. Insulin, NKX6.1 and glucagon mRNA were expressed in stage 4 cells but not in early undifferentiated cells. We concluded that rES cells could be differentiated ex vivo to insulin producing cells. These differentiated rES cells could be used to develop a non-human primate model for evaluating cell therapy to treat diabetes. To facilitate the identification of beta-like cells and to track the cells post-transplantation, we have developed a marker gene construct: fusing the human insulin promoter (HIP) to the green fluorescent protein (GFP) gene. This construct was transfected into stage 3 rES derived cells and subsequent GFP expression was identified in C-peptide positive cells, thereby substantiating endogenous insulin production by rES derived cells. Using this GFP detection system, we will enrich our population of insulin producing rES derived cells and track these cells post-transplantation in the non-human primate model.

Animals↗

PubMed1999

Fluorescent labeling of blood cells for evaluation of retinal and choroidal circulation.

BACKGROUND AND OBJECTIVE: To develop a method of staining and tracking cells in vivo with a scanning laser ophthalmoscope (SLO) to evaluate the retinal and choroidal circulations. MATERIALS AND METHODS: Twelve pigmented male rats were used. Staining of leukocytes. Two commercially available nucleic acid stains SYTO 16 and SYTO 59 (Molecular Probes, Eugene, OR) were used to stain leukocytes. Blood (0.5 mL) was withdrawn from the subject animal and placed in a heparinized tube to which phosphate buffered solution was added (1.0 mL buffer/0.5 mL blood). Ten microliters of solution of SYTO 16 or SYTO 59 in alcohol (5 mM) was added. Staining of Erythrocytes. The lipophilic carbocyanine dye indocarbocyanine (D-307, Molecular Probes) was used to stain erythrocytes. Blood (0.1 mL) was withdrawn from the subject and handled as above; plasma and leukocytes were removed with a pipette. HEPES buffer was added to the remaining erythrocytes (1.4 mL buffer added to 0.1 mL blood). Ten microliters of D-307 in alcohol (5 mM) was added to the 1.5 mL buffered blood. The final concentration of dyes in the buffered blood mixtures was 0.03 mM. The stained cells were injected intravenously in the subject animal. The eyes were observed with SLO; the resident helium-neon laser was used to excite the SYTO 59 and the resident argon laser for SYTO 16. The He-Ne laser was used to excite the D-307-stained erythrocytes. RESULTS: The movement of the leukocytes and erythrocytes in the retinal and choroidal vessels was clearly distinguishable from the surrounding tissue. CONCLUSION: By selecting different dyes for staining leukocytes or erythrocytes and choosing appropriate laser wavelengths, the hemodynamics of blood cells can be observed and evaluated in the retina and choroid.

Affinity Labels↗

PubMed2004

Motion gradient vector flow: an external force for tracking rolling leukocytes with shape and size constrained active contours.

Recording rolling leukocyte velocities from intravital microscopic video imagery is a critical task in inflammation research and drug validation. Since manual tracking is excessively time consuming, an automated method is desired. This paper illustrates an active contour based automated tracking method, where we propose a novel external force to guide the active contour that takes the hemodynamic flow direction into account. The construction of the proposed force field, referred to as motion gradient vector flow (MGVF), is accomplished by minimizing an energy functional involving the motion direction, and the image gradient magnitude. The tracking experiments demonstrate that MGVF can be used to track both slow- and fast-rolling leukocytes, thus extending the capture range of previously designed cell tracking techniques.

Algorithms↗

PubMed1998

Characterization of Involution during Sea Urchin Gastrulation Using Two-Photon Excited Photorelease and Confocal Microscopy.

: Sea urchin embryos have served as a model system for the investigation of many concepts in developmental biology. Their gastrulation consists of two stages; primary invagination, where part of the epithelium invaginates into the blastocoel, and secondary invagination, where the archenteron elongates to completely traverse the blastocoel. Primary invagination involves proliferation of cells within the vegetal plate during primary invagination, but until recently, it was assumed that the larval gastrointestinal (GI) tract developed without further involution of epithelial cells. To investigate rigorously the contribution of epithelial cell involution during archenteron and GI tract development in the sea urchin, Lytechinus variegatus, we developed a new method for cell tracking based on two-photon excited photorelease of caged fluorophores. Single-cell embryos were injected with caged dye and two-photon excitation uncaging was employed to mark small groups of cells throughout gastrulation. Two-photon excitation allowed for noninvasive, three-dimensionally resolved uncaging inside living cells with minimal biological damage. Cellular involution into the archenteron was observed throughout primary and secondary invagination, and the larval intestine was formed by further involution of cells following secondary invagination, which is inconsistent with the traditional model of sea urchin gastrulation. Further, as two-photon excitation microscopy becomes accessible to many researchers, the novel techniques described here will be broadly applicable to development of other invertebrate and vertebrate embryos.

Journal Article↗

PubMed1997

AtLTP1 luciferase expression during carrot somatic embryogenesis.

The carrot (Daucus carota L.) EP2 gene encodes a Lipid Transfer Protein (LTP) which is expressed during protoderm formation in developing embryos. To develop a vital reporter system for gene expression during somatic embryo development a 1.1 kB fragment of the Arabidopsis thaliana LTP1 promoter was fused to the firefly luciferase (LUC) coding sequence. The AtLTP1 luciferase expression pattern in transformed carrot suspension cultures was identical to the expression pattern of the endogenous carrot EP2 gene. Cell tracking experiments revealed that all somatic embryos were derived from AtLTP1 luciferase expressing cell clusters. However, not all cell clusters that expressed the AtLTP1 luciferase reporter gene developed into a somatic embryo, suggesting that initiation of an embryogenic pathway in tissue culture does not always lead to development of a somatic embryo.

Antigens, Plant↗

PubMed2005

Promises and pitfalls of stem cell therapy for promotion of bone healing.

UNLABELLED: There is promise in combining stem cells with allogeneic bone matrix to promote bone healing. Murine bone marrow, peripheral blood, and compact bone cells were transplanted ectopically under the kidney capsule in mice, alone or in combination with allogeneic matrix products: powder and putty to determine their bone forming potential in comparison to transplanted femoral bone fragments and long-term cultured bone marrow cells. The end point was the amount of bone formed as determined by quantitative histology. Mononuclear cells from marrow, peripheral blood, or bone alone transplanted under the kidney capsule did not form bone. Mononuclear cell populations did not combine readily with matrix products and there was in vivo migration of the transplanted combinations. Kidney subcapsular transplanted cultured bone marrow cells formed bone in proportion to the culture period, but after 9 weeks, the extent was only 20% by area of that of similarly transplanted femoral bone fragments. An inductive stimulus for bone formation seemed necessary. Osteoprogenitor cells were not detected in significant numbers in blood unless high doses of cytokines were administered. A better definition of the optimal cell populations and manipulations required for promotion of bone healing is needed along with new (transplant) models that allow for cell tracking. Much work remains to overcome current pitfalls in the use of stem cells to promote allograft integration and bone healing. LEVEL OF EVIDENCE: Therapeutic study, Level V (expert opinion). See the Guidelines for Authors for a complete description of levels of evidence.

Animals↗

PubMed1993

Repeated administration of Sigma ligands alters the population activity of rat midbrain dopaminergic neurons.

Acute and repeated administration of antipsychotic drugs produce distinctive profiles of electrophysiological effects on the population activity of midbrain dopaminergic (DA) neurons which correlate with their clinical effects. Sigma receptors have been hypothesized to be involved in psychosis and in the efficacy of antipsychotic drugs, but little is known about the effects of repeated treatment with sigma ligands on the activity of midbrain DA neuronal populations. In the present study, the cells-per-track cell-sampling method was used to evaluate the effects of 3 sigma ligands on the numbers of spontaneously active A9 and A10 DA neurons in chloral hydrate-anesthetized rats. One-hour pretreatment with either (+)-pentazocine (10 mg/kg, i.p.), DTG (2 mg/kg, i.p.), or JO 1784 (1 or 10 mg/kg, s.c.) did not alter the number of spontaneously active DA neurons encountered per electrode track. Repeated treatment (21 daily injections) with (+)-pentazocine (1 or 10 mg/kg) or DTG (0.2 or 2 mg/kg) increased the number of A10 DA cells per track; JO 1784 (10 mg/kg but not 1 mg/kg) moderately decreased the number of active A9 DA cells and increased the firing rate of A10 DA neurons. The effect of JO 1784 on A9 DA neurons was not due to depolarization inactivation. The effects of all 3 sigma ligands differ from those of antipsychotic drugs, all of which inactivate A10 DA neurons after repeated treatment. Clinical studies are necessary to determine if selective sigma ligands will provide a novel alternative to DA antagonists in the treatment of psychosis.

Animals↗

PubMed1984

HZE effects on mammalian cells.

In track segment experiments cell survival and chromosome aberrations of mammalian cells have been measured for various heavy ion beams between helium and uranium in the energy range between 0.5 and 960 MeV/u, corresponding to a velocity range of 0.03 to 0.87 C, and an LET spectrum from 10 to 15 000 keV/micrometers. At low LET, the cross section (sigma) for cell killing increases with increasing LET and shows a common curve for all ions regardless of the atomic number. This indicates that in this region the track structure of the different ions is of only a minor influence, and it is rather the total energy transfer, which is important for cell killing. At higher LET values, deviations from a common sigma-LET curve can be observed which indicate a saturation effect. The saturation of the lighter ions occurs at lower LET values than for the heavier ions. These findings are also confirmed by the chromosome data, where the efficiency for the induction of chromosomal aberrations for high LET particles depends on the track structure and is nearly independent of LET. In the heavier beams (Z > or = 10) individual particles cause multiple chromosome breaks in mitotic cells.

Animals↗

PubMed2003

New TCR transgenic model for tracking allospecific CD4 T-cell activation and tolerance in vivo.

We have developed an adoptive transfer model system to visualize the dynamics of alloantigen-specific CD4+ T-cell activation in vivo. Using TCR-transgenic (tg) mice reactive to I-Ab(m12), we studied the clonal expansion and differentiation of alloreactive T cells by tracking the fate of adoptively transferred TCR-tg CD4+ T cells in syngeneic mice transplanted with skin grafts expressing I-Ab(m12). Following transplantation, alloantigen-specific TCR-tg CD4+ T-cell expansion was observed initially in the draining lymph nodes followed by the spleen. TCR-tg CD4+ T cells up-regulated CD69 and CD25 expression, developed an effector/memory surface phenotype and produced IFN-gamma in response to alloantigen ex vivo. Furthermore, we validate the model system as a means for studying the effects of tolerogenic regimens on alloreactive CD4+ T cells, demonstrating that CTLA4Ig inhibits alloantigen-dependent clonal expansion and effector function of TCR-tg CD4+ T cells in vivo. We describe the first model for tracking alloreactive CD4+ T-cell activation in vivo. It provides a powerful tool for studying CD4+ T-cell mediated alloimmune responses and mechanisms of tolerance induction in vivo.

Adoptive Transfer↗

PubMed2004

Cell population dynamics model for deconvolution of murine embryonic stem cell self-renewal and differentiation responses to cytokines and extracellular matrix.

Stem cell self-renewal versus differentiation fate decisions are difficult to characterize and analyze due to multiple competing rate processes occurring simultaneously among heterogeneous cell subpopulations. To address this challenge, we describe a mathematical model for cell population dynamics that allows flow cytometry measurement of population distributions of molecular markers to be deconvoluted in terms of subpopulation-specific rate parameters distinguishing commitment to differentiation, proliferation of differentiated cells, and proliferation of undifferentiated cells (i.e., self-renewal). We validate this model-based parameter determination by means of dedicated, independent cell-tracking studies. Our approach facilitates interpretation of relationships underlying effects of external cues on cell responses in differentiating cultures via intracellular signals.

Animals↗

PubMed2003

Introduction of cell markers into germ layer tissues of the mouse gastrula by whole embryo electroporation.

We have optimized the technique of electroporation for introducing genetic markers into cells of the gastrulating mouse embryo to follow cell fates, tissue movement, and lineage differentiation. Using a plate-needle electrode combination and specific route of plasmid delivery, labeling could be targeted to discrete regions of the epiblast or the endoderm of the late gastrula. Among the various types of fluorescent and chromogenic reporter constructs tested, those driven by CMV promoter are efficient and strong expression can be detected as soon as 2-3 h after electroporation. The efficacy of marking cell lineages by CRE-mediated activation of reporters proved to be inefficient for tracking cell lineages due to an obligatory 8-9-h lag from the electroporation of constructs to the expression of reporter. This significant time lag also raises concern of the temporal precision at which tissue- or stage-specific knock-out or activation of genetic activity may be achieved by the Cre-loxP mechanism.

Alkaline Phosphatase↗

PubMed2000

Migration of dendritic cells within 3-D collagen lattices is dependent on tissue origin, state of maturation, and matrix structure and is maintained by proinflammatory cytokines.

The function of dendritic cells (DC) depends on active migration through three-dimensional (3-D) extracellular matrices. We have analyzed the migration of murine DC from different tissue origins within 3-D collagen lattices through the use of time-lapse videomicroscopy and single-cell tracking. Directly after incorporation, 50-90% of DC from the spleen (spDC) and Langerhans cells freshly isolated from the epidermis (fLC) displayed active motility in these matrices. Whereas mature spDC showed multilateral pseudopod dynamics as well as fast and heterogeneous migration, immature fLC displayed a spherical shape with faint membrane processes and very homogenous, slow migration characteristics. In the absence of external stimuli, migration of both, spDC and fLC, vanished after >36 h due to cell death. Maintaining fLC viability by external granulocyte-macrophage colony-stimulating factor or tumor necrosis factor alpha prolonged migration up to 5 days. During this period fLC transformed into mature cells with large dendrites, thereby developing a heterogeneous migration pattern more similar to spDC. In randomly polymerized collagen matrices cell paths were without preferential orientation. In contrast, in artificially aligned lattices directional paths in accordance with the forced fiber orientation were observed. Thus, migration is an inherent property of DC, largely influenced by tissue origin, degree of maturity, and the 3-D structure of the environment.

Animals↗

PubMed2006

Neural stem cell transplant survival in brains of mice: assessing the effect of immunity and ischemia by using real-time bioluminescent imaging.

PURPOSE: To use bioluminescent imaging in a murine transplant model to monitor the in vivo responses of transplanted luciferase-gene-positive neural progenitor cells (NPCs) to host immunity and ischemia. MATERIALS AND METHODS: All animal studies were conducted according to institutional guidelines, with approval of the Subcommittee on Research Animal Care. Cranial windows were created in all animals, and all animals underwent NPC (C17.2-Luc-GFP-gal) transplantation into the right basal ganglia. An observational study was performed on C57 BL/6 (n = 5), nude (n = 4), and CD-1 (n = 4) mice, with bioluminescent imaging performed at days 7, 11, and 14 after transplantation. A study on the effects of ischemia was performed in a similar manner, but with the following differences: On day 9 after transplantation, the C57 BL/6 mice underwent 18 minutes of transient forebrain ischemia by means of temporary bilateral carotid occlusions (n = 6). A control group of C57 BL/6 mice underwent sham surgery (n = 6). Bioluminescent imaging was performed on the ischemic animals and control animals at days 7, 9, 11, and 14. Repeated-measures analysis of variance or Student t test was used to compare the means of the luciferase activities. RESULTS: In vivo cell tracking demonstrated that (a) C17.2-Luc-GFP-gal NPCs survived and proliferated better in the T-cell deficient nude mice than in the immunocompetent C57 BL/6 or CD-1 mice, in which progressive immune mediated cell loss was shown, and (b) transient forebrain ischemia appeared, unexpectedly, to act as a short-term stimulus to transplanted NPC growth and survival in immunocompetent mice. CONCLUSION: Immune status and host immunity can have an influence on NPC graft survival, and these changes can be noninvasively assessed with bioluminescent imaging in this experimental model.

Animals↗

PubMed2002

Evidence for incorporation of free-floating mesothelial cells as a mechanism of serosal healing.

Regeneration of the mesothelium is unlike that of other epithelial-like surfaces, as healing does not occur solely by centripetal migration of cells from the wound edge. The mechanism of repair of mesothelium is controversial, but it is widely accepted, without compelling evidence, that pluripotent cells beneath the mesothelium migrate to the surface and differentiate into mesothelial cells. In this study we examined an alternative hypothesis, using in vivo cell-tracking studies, that repair involves implantation, proliferation and incorporation of free-floating mesothelial cells into the regenerating mesothelium. Cultured mesothelial cells, fibroblasts and peritoneal lavage cells were DiI- or PKH26-PCL-labelled and injected into rats immediately following mesothelial injury. Implantation of labelled cells was assessed on mesothelial imprints using confocal microscopy, and cell proliferation was determined by proliferating cell nuclear antigen immunolabelling. Incorporation of labelled cells, assessed by the formation of apical junctional complexes, was shown by confocal imaging of zonula occludens-1 protein. Labelled cultured mesothelial and peritoneal lavage cells, but not cultured fibroblasts, implanted onto the wound surface 3, 5 and 8 days after injury. These cells proliferated and incorporated into the regenerated mesothelium, as demonstrated by nuclear proliferating cell nuclear antigen staining and membrane-localised zonula occludens-1 expression, respectively. Furthermore, immunolocalisation of the mesothelial cell marker HBME-1 demonstrated that the incorporated, labelled lavage-derived cells were mesothelial cells and not macrophages as it had previously been suggested. This study has clearly shown that serosal healing involves implantation, proliferation and incorporation of free-floating mesothelial cells into the regenerating mesothelium.

Animals↗

PubMed1989

Fluorescent in vivo tracking of hematopoietic cells. Part I. Technical considerations.

We report a new technology for in vivo tracking of hematopoietic cells, using fluorescent lipophilic probes. Because the probe is irreversibly bound in the lipids of the cell membrane; substantial numbers of dye molecules can be incorporated per cell and thus substantial signal to noise can be achieved. Although this technology can be used for all hematopoietic cells, these first findings are reported on red blood cells (RBCs) owing to the importance of the membrane to RBC function and integrity. We demonstrated that labeling 10% of the RBCs of a rabbit and reinjecting them into the animal makes possible the tracking of these cells at various times after injection. Furthermore, the labeling appears not to affect in vivo cell lifetime or cellular volume changes in response to hypotonic shock. The single cell fluorescence intensity of the labeled RBCs remains relatively constant for 60 days, and an immune response appears not to be generated against labeled cells. That labeled RBCs have lifetime kinetics in vivo, as shown in other studies, indicates that the membranes are functioning normally and are unaltered by the labeling technology. The technology we present is also applicable to white blood cells, bone marrow, and platelets.

Animals↗

PubMed2001

Deformation-enhanced fluctuations in the red cell skeleton with theoretical relations to elasticity, connectivity, and spectrin unfolding.

To assess local elasticity in the red cell's spectrin-actin network, nano-particles were tethered to actin nodes and their constrained thermal motions were tracked. Cells were both immobilized and controllably deformed by aspiration into a micropipette. Since the network is well-appreciated as soft, thermal fluctuations even in an unstressed portion of network were expected to be many tens of nanometers based on simple equipartition ideas. Real-time particle tracking indeed reveals such root-mean-squared motions for 40-nm fluorescent beads either tethered to actin directly within a cell ghost or connected to actin from outside a cell via glycophorin. Moreover, the elastically constrained displacements are significant on the scale of the network's internodal distance of approximately 60-80 nm. Surprisingly, along the aspirated projection-where the network is axially extended by as much as twofold or more-fluctuations in the axial direction are increased by almost twofold relative to motions in the unstressed network. The molecular basis for such strain softening is discussed broadly in terms of force-driven transitions. Specific considerations are given to 1) protein dissociations that reduce network connectivity, and 2) unfolding kinetics of a localized few of the red cell's approximately 10(7) spectrin repeats.

Actins↗

PubMed1998

Cell migration strategies in 3-D extracellular matrix: differences in morphology, cell matrix interactions, and integrin function.

Cell migration in extracellular matrix is a complex process of adhesion and deadhesion events combined with cellular strategies to overcome the biophysical resistance imposed by three-dimensionally interconnected matrix ligands. Using a 3-D collagen matrix migration model in combination with computer-assisted cell tracking for reconstruction of migration paths and confocal microscopy, we investigated molecular principles governing cell-matrix interactions and migration of different cell types. Highly invasive MV3 melanoma cells and fibroblasts are large and highly polarized cells migrating at low speed (0.1-0.5 microm/min) and at high directional persistence. MV3 melanoma cells utilize adhesive migration strategies as characterized by high beta1 integrin surface expression, beta1 integrin clustering at interactions with matrix fibers, and beta1 integrin-mediated adhesion for force generation and migration. In contrast, T lymphocytes and dendritic cells are highly mobile cells of lower beta1 integrin expression migrating at 10- to 40-fold higher velocities, and directionally unpredictable path profiles. This migration occurs in the absence of focal adhesions and largely independent of beta1 integrin-mediated adhesion. Whereas cell-matrix interactions of migrating tumor cells result in traction and reorientation of collagen fibers, partial matrix degradation, and pore formation, leukocytes form transient and short-lived interactions with the collagen lacking structural proteolysis and matrix remodeling. In conclusion, the 3-D extracellular matrix provides a spatially complex and biomechanically demanding substrate for cell migration, thereby differing from cell migration across planar ligands. Highly adhesive and integrin-dependent migration strategies detected in morphologically large and slowly migrating cells may result in reorganization of the extracellular matrix, whereas leukocytes favor largely integrin-independent, rapid, and flexible migration strategies lacking typical focal adhesions and structural matrix remodeling.

Cell Adhesion↗