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PubMedDate not supplied

Current methods for biopsy and treatment of potentially curable breast cancer.

Concepts regarding the best methods for doing biopsies and the most appropriate methods for treatment of patients with breast cancers have changed dramatically since the early nineteen seventies. Currently formal biopsies (incisional or excisional) are being done as two-step procedures with reliance on permanent sections for diagnosis. The advantages of this type of biopsy are discussed along with the increasing frequency of doing them under local anesthesia on an out-patient basis. Informal biopsies (fine needle aspiration or needle core) are becoming increasingly popular in the office and clinic and stereotactic biopsies are receiving increasing attention. The use of flow cell cytometry to complement cytologic diagnosis and the use of needle localization, with or without dye injection, for the biopsy of occult lesions are described. The need for determining by prognostic parameters and by scanning the remaining breast for residual cancers after radiolabeled monoclonal antibody injection, which in situ cancers can be managed by lesser procedures than total mastectomy and which early invasive cancers can be managed by just wide excision is discussed. The two main current local-regional therapy options for women with invasive breast cancers (some type of modified mastectomy with breast reconstruction and breast conservation with radiotherapy) are presented in detail. The present need of doing an axillary dissection in both for staging is discussed along with the future possibility of eliminating it by scanning with a gamma-camera the axillary and internal mammary nodal areas after radiolabeled monoclonal antibody injection.(ABSTRACT TRUNCATED AT 250 WORDS)

Biopsy↗
PubMed2000

Triggering the ExoS regulon of Pseudomonas aeruginosa: A GFP-reporter analysis of exoenzyme (Exo) S, ExoT and ExoU synthesis.

The ExoS regulon of Pseudomonas aeruginosa encodes diverse type III secreted effector proteins which have been shown to exert cytotoxic effects in cell culture experiments. However, little information exists about the environmental conditions and stimuli for upregulation of the ExoS regulon. Translational reporter fusion proteins of exoenzyme (Exo) S, ExoT and ExoU, as well as the type II secreted exotoxin A (ETA) to the green fluorescent protein (GFP), were constructed in order to compare exoprotein production under diverse growth conditions. Reporter protein activity was recorded by FACS-analysis and by conventional and confocal laser scanning microscopy. Low ion concentration induced co-ordinated upregulation of ExoS, ExoT and ExoU with a maximum effect at 37 degrees C. A dose-dependent upregulation was seen with human serum or increasing NaCl concentrations. A type III secretion-negative pcrD mutant of P. aeruginosa showed a weak ExoS response to environmental stimuli, compared with the parental strain, suggesting a negative regulatory mechanism. Co-culture with the mammalian cell lines J774A.1 or HeLa led to rapid upregulation of ExoS, ExoT and ExoU synthesis. These data suggest that the ExoS regulon of P. aeruginosa can be triggered by a variety of environmental signals as well as by cell contact with eukaryotic cells.

ADP Ribose Transferases↗
PubMed1997

Internalization of non-permeant materials into cultured mammalian cells by vortex-stirring in the presence of a high molecular weight polyacrylic acid: direct evidence of internalization and its dependence on the cell lines.

Laser scanning confocal fluorescence microscopic analysis provided a direct evidence of the internalization of non-permeant Lucifer Yellow and a dextran of MW 4400 into leukemia L1210 cells when the cells were vortex-stirred in the presence of a high molecular weight polyacrylic acid, A-119 (MW ca. 9 x 10(6)). The morphology of A-119-treated cells was probed by scanning electron microscopic analysis using 2% glutaraldehyde for fixation. The cells immediately after vortex-stirring with A-119 showed many blebs on the cell surface, indicative of local weakening of the plasma membrane. The blebby surface returned to normal within 5 min at 37 degrees C, but not completely at 0 degree C even after 20 min. Several cultured cell lines, murine splenocytes, and Ehrlich ascites carcinoma cells were particularly susceptible to permeabilization by this method, although the efficiency varied from one type of cell to another.

Acrylic Resins↗
PubMed1999

Probing the mechanism of transport and compartmentalisation of polyamines in mammalian cells.

BACKGROUND: Many mammalian cells possess an active polyamine uptake system but little is known about the molecular mechanism of this transporter. The fate of polyamines taken up from the medium and the relationship to polyamine homeostasis remains to be fully established. The aim of this study was to develop a range of modified polyamines, particularly ligands incorporating a fluorophore, to explore the structural tolerances of the polyamine transport system and to probe the intracellular location of polyamines acquired from the medium. RESULTS: We synthesised a wide range of polyamine analogues incorporating cytotoxic agents, fluorescent chromophores and bulky substituents. All of these analogues have been shown to be good competitive inhibitors of spermidine uptake in a range of mammalian cells. Direct evidence for uptake of the fluorescent polyamine analogues and their subcellular distribution was obtained from confocal laser scanning fluorescence microscopy, which showed that they accumulated in granular structures within the cytoplasm and not in the nucleus. We demonstrated that their uptake is through the polyamine transport system by showing that pretreatment with DFMO, a potent inhibitor of polyamine biosynthesis, led to enhanced uptake, and cells deficient in the polyamine transport system did not accumulate these polyamine analogues. CONCLUSIONS: The polyamine transport system has a surprisingly broad structural tolerance. Fluorophore-containing polyamine analogues derived from the extracellular pool are located in granular structures within the cytoplasm and not to any great extent in the nuclei of mammalian cells. These observations might be consistent with a mechanism involving receptor-mediated endocytosis, and the granular 'structures' seen might reflect polyamine compartmentalisation within vesicles.

Animals↗
PubMed2000

The DNA of annexin V-binding apoptotic cells is highly fragmented.

Jurkat leukemia cells induced to undergo apoptosis by treatment with an antibody against the Fas receptor have two annexin V (AV)-binding subpopulations: (a) single-positive cells that bind AV but not propidium iodide (PI); and (b) double-positive cells that bind AV and PI. The single-positive population is thought to represent an early stage of apoptosis. We have examined the relationship between AV binding and a classical characteristic of apoptosis, DNA fragmentation. Time course studies with Jurkat cells treated for 1, 2, or 4 h with anti-Fas indicated that the proportion of AV-binding cells was increased after 2 h. A significant increase in DNA fragmentation was observed only at 4 h as measured by the mean tail moment determined with the alkaline single cell gel electrophoresis (comet) assay. This correlation suggests a temporal relationship between the two parameters, but does not provide direct evidence of what happens in individual cells. We developed a method to measure fluorescent markers of cellular structure or function with a laser scanning cytometer and then perform the comet assay on the same cells. Cells in each AV-binding subpopulation were re-examined before and after electrophoresis. Most AV-/PI- cells had no DNA damage, although a few cells showed a pattern of damage characteristic for apoptosis. Double-positive cells all had damaged DNA; approximately half had the apoptotic pattern, and the rest had a pattern typical for necrosis. Nearly all of the single-positive cells had damaged DNA with the apoptotic pattern. Both AV-positive populations contained cells with little or no detectable DNA after electrophoresis, indicating that the DNA was highly fragmented. These results indicate that AV binding is an excellent marker for apoptotic cells, but that these cells already have fragmented DNA.

Annexin A5↗
PubMed1997

Laser scanning cytometric analysis of cyclin B1 in primary human malignancies.

Cyclins are key components of the cell cycle progression machinery. They activate their partner-dependent kinases (CDKs) and target them to respective substrate proteins within the cell. CDK-mediated phosphorylation of specific sets of proteins drives the cell through particular phases or checkpoints of the cell cycle. During unperturbed growth of normal cells, the timing of expression of several cyclins is discontinuous, occurring at discrete and well-defined periods of the cell cycle. Immunocytochemical detection of cyclins in relation to cell cycle position (DNA content) by multiparameter flow cytometric techniques has provided a new approach to cell cycle studies. This approach, like no other method, can be used to detect the "unscheduled" expression of cyclins, namely, the presentation of G1 cyclins by cells in G2/M and of G2/M cyclins by G1 cells, without the need for cell synchronization. By use of multiparameter flow cytometric and laser scanning cytometric analysis, we correlated the expression of cyclin B1 with cell cycle position in normal lymphocytes stimulated to proliferate by the mitogen phytohemagglutinin and in 28 primary human tumors of different organ and type. Eighteen of the 28 tumors expressed the cyclin B1 in more than 5% of cells (B1 positive), and the rest showed cyclin expression from 2.1 to 5% (B1 negative). In normal lymphocytes, the expression of cyclin B1 was restricted to very late S and G2 + M phases of the cell cycle. In 15 of 18 primary tumors studied, the expression of cyclin B1 was "unscheduled" (unrestricted to particular phases of the cycle). The data suggest that the "unscheduled" expression of cyclin B1 might be a common defect in neoplasia.

Adrenal Gland Neoplasms↗
PubMed2006

Multispectral imaging in biology and medicine: slices of life.

Multispectral imaging (MSI) is currently in a period of transition from its role as an exotic technique to its being offered in one form or another by all the major microscopy manufacturers. This is because it provides solutions to some of the major challenges in fluorescence-based imaging, namely ameliorating the consequences of the presence of autofluorescence and the need to easily accommodate relatively high levels of signal multiplexing. MSI, which spectrally characterizes and computationally eliminates autofluorescence, enhances the signal-to-background dramatically, revealing otherwise obscured targets. While this article concentrates on examples derived from liquid-crystal tunable filter-based technology, the intent is to showcase the advantages of multispectral imaging in general. Some technologies used to generate multispectral images are compatible with only particular optical configurations, such as point-scanning laser confocal microscopy. Band-sequential approaches, such as those afforded by liquid-crystal tunable filters (LCTFs), can be conveniently coupled with a variety of imaging modalities, which, in addition to fluorescence microscopy, include brightfield (nonfluorescent) microscopy as well as small-animal, noninvasive in-vivo imaging. Brightfield microscopy is the chosen format for histopathology, which relies on immunohistochemistry to provide molecularly resolved clinical information. However, in contrast to fluorescent labels, multiple chromogens, if they spatially overlap, are much harder to separate and quantitate, unless MSI approaches are used. In-vivo imaging is a rapidly growing field with applications in basic biology, drug discovery, and clinical medicine. The sensitivity of fluorescence-based in-vivo imaging, as with fluorescence microscopy, can be limited by the presence of significant autofluorescence, a limitation which can be overcome through the utilization of MSI.

Animals↗
PubMed2000

The organization of replicon clusters is not affected by polyamine depletion.

Earlier investigations have shown that polyamine depletion affects DNA replication negatively. DNA is synthesized in replicons which are gathered in replicon clusters. DNA replication is initiated simultaneously in every replicon of a replicon cluster. By pulse labeling cells with the thymidine analog bromodeoxyuridine and then detecting bromodeoxyuridine in situ with immunofluorescence, replicon clusters can be studied. We have used this method to investigate the effects of 2-difluoromethylornithine (DFMO)- and 4-amidinoindan-1-one 2'-amidinohydrazone (CGP 48664)-mediated polyamine depletion on the organization of replicon clusters. The cells were studied by fluorescence microscopy and confocal laser scanning microscopy. Our studies give at hand that neither the number nor the distribution of replicon clusters were affected even after 4 days of treatment with 5 mM DFMO or 20 microM CGP 48664, indicating that polyamine depletion did not affect the organization of replicon clusters. However, the fluorescence intensity of the replicon clusters was much lower in inhibitor-treated cells. The results indicate that the impaired DNA replication observed in polyamine-depleted cells is not due to an effect on the initiation step of DNA replication, but rather on the elongation process. To confirm that it is possible to observe changes in the organization of replicon clusters using bromodeoxyuridine, we treated the cells with various drugs that affect DNA replication. Aphidicolin, which inhibits DNA elongation, gave results similar to those of DFMO and CGP 48664.

Amanitins↗
PubMed1991

Fluorescent probes as tools to assess the receptor for the urokinase-type plasminogen activator on tumor cells.

Flow cytofluorometric protocols (FACScan) are described for the rapid and quantitative real-time analysis of binding of FITC-pro-u-PA to cell surface receptors (u-PAR) on living, resting, and also on PMA-stimulated human monocytic U937 cells. Binding of pro-u-PA was visualized by CLSM. This fairly new technique is superior over conventional fluorescence microscopy and is an alternative to electron microscopic approaches. Both flow cytofluorometry and confocal laser scanning microscopy allow the analysis quantitatively and with high-sensitivity binding of FITC-pro-u-PA to single suspended or adherent cells. By CLSM u-PA/u-PAR were found to be located in heterogeneously distributed discrete patches at the cell surface on U937 and not inside the cell. This is in agreement with previous studies by Hansen et al, who applied radioiodinated u-PA and electron microscopy to locate u-PAR on microvilli of fixed U937 cells. By flow cytofluorometry, it was possible to quantify the time-dependent and temperature-dependent binding of FITC-pro-u-PA to living single U937. Apparent saturation of u-PAR was achieved at 5 nM FITC-pro-u-PA for both nonstimulated and PMA-stimulated U937 cells. Half saturation of u-PAR was also determined. Nonstimulated U937 was 0.7 nM, and PMA-stimulated U937 was 1.1 nM of FITC-pro-u-PA. This increase in half-saturation concentration in PMA-stimulated cells is paralleled by a steep increase in binding sites (3.6-fold). The use of fluoresceinated reference beads is recommended to verify changes in affinity and binding sites. Using CLSM or flow cytofluorometry, it is also possible to study the structure relationship of u-PA/u-PAR in the presence of competitive binding analogues or inhibitors. Fluorescence techniques will also permit the identification of u-PAR-positive cells in blood, ascitic fluid, or biopsies obtained from cancer patients.

Amino Acid Sequence↗
PubMed1997

Influence of N-methylformamide on the intracellular transport of doxorubicin.

The polar solvent N-methylformamide proved to be capable of enhancing the cytotoxic potential of various antitumoral compounds, both in vitro and in vivo. In many cases, this ability depended on the sequence of treatment, and the enhancement of the cytotoxic effect occurred only when N-methylformamide administration succeeded anticancer drug treatment. The results obtained in the present study indicate that N-methylformamide interferes with the mechanisms of intracellular transport and efflux of the antitumoral drug doxorubicin. In particular, laser scanning confocal microscopy observations performed on melanoma cells (M14) after N-methylformamide administration revealed evident alterations of the microtubular network, including numerous interruptions of the microtubules. Moreover, when doxorubicin-treated cells were recovered in the presence of the polar solvent, the normal efflux of the anthracyclinic antibiotic appeared to be hampered, and the drug was localized mainly in well delimited perinuclear regions. Double staining experiments demonstrated the colocalization of the doxorubicin molecules and the WGA-stained regions as well as a close structural relationship between them and the microtubule system. These results indicate that N-methylformamide interferes with the doxorubicin transport inducing a damage in the microtubular network and the consequent persistence and entrapment of the drug in the regions likely occupied by the Golgi apparatus of tumor cells. This finding could account for the chemosensitizing properties exerted by N-methylformamide.

Antineoplastic Agents↗
PubMed2004

Analysis of the fluorescence of monodansylcadaverine-positive cytoplasmic structures during 7-ketocholesterol-induced cell death.

OBJECTIVE: To analyze multilamellar cytoplasmic structures by confocal laser scanning microscopy (CLSM) combined with factor analysis of biomedical image sequences (FAMIS). STUDY DESIGN: After treatment of U937 cells with 7-ketocholesterol (7-keto), cytoplasmic alterations were assessed with monodansylcadaverine (MDC). By ultraviolet excitation of a confocal laser scanning microscope (UV-CLSM), spectral sequences were performed to characterize 7-keto and MDC distribution inside cells. FAMIS was used to transform the image sequences in factor curves and images. RESULTS: By UV-CLSM, 7-keto fluorescence was detected together with MDC, which revealed morphologic cytoplasmic changes in cells. The factor images obtained from confocal image sequences emphasized the view of these results. These data are in agreement with biochemical characterizations of MDC-positive structures. CONCLUSION: The combined use of confocal microscopy and FAMIS allowed us to detect MDC-positive cytoplasmic structures in 7-keto-treated cells and to colocalize MDC and 7-keto distribution. This new method confirms the usefulness of MDC as a marker of oxysterol-induced cell death.

Benzimidazoles↗
PubMed2005

Application of a static fluorescence-based cytometer (the CellScan) in basic cytometric studies, clinical pharmacology, oncology and clinical immunology.

The CellScan apparatus is a laser scanning cytometer enabling repetitive fluorescence intensity (FI) and polarization (FP) measurements in living cells, as a means of monitoring lymphocyte activation. The CellScan may serve as a tool for diagnosis of rheumatoid arthritis (RA) and systemic lupus erythematosus (SLE) as well as other autoimmune diseases by monitoring FP changes in peripheral blood lymphocytes (PBLs) following exposure to autoantigenic stimuli. Changes in FI and FP in atherosclerotic patients' PBLs following exposure to various stimuli have established the role of the immune system in atherosclerotic disease. The CellScan has been evaluated as a diagnostic tool for drug-allergy, based on FP reduction in PBLs following incubation with allergenic drugs. FI and FP changes in cancer cells have been found to be well correlated with the cytotoxic effect of anti-neoplastic drugs. In conclusion, the CellScan has a variety of applications in cell biology, immunology, cancer research and clinical pharmacology.

Allergy and Immunology↗
PubMed1999

Cytofluorometry and fluorescence confocal laser scanning microscopy (CLSM) in the study of neutral lipid dynamics in Paramecium primaurelia mating types during cell line development.

BACKGROUND: In Paramecium primaurelia, an exconjugant cell can produce two lines with different mating capacities. Mating type II cells can form a higher food vacuole number and digest the nutrient taken up in a shorter time; thus, mating type II cells grow at a faster rate than do mating type I cells. The present study was done to determine whether cells that ingest more nutrients also have a larger amount of storage lipids. METHODS: Quantitative and qualitative determinations of neutral lipids were obtained by means of cytofluorometry and fluorescence confocal laser scanning microscopy (CLSM), respectively, by using nile red on cells in different physiologic states. RESULTS: Lipid droplet number and neutral lipid content were higher in mating type II cells than in mating type I cells in the early logarithmic growth phase (i.e., immature well-fed cells). These values were reversed during the middle and the late logarithmic phases and became equal in the stationary phase (i.e., mature starved cells). In well-fed cells maintained with food excess, differences in neutral lipid content between the two mating types also were present in mature cells. CONCLUSIONS: Although differences between mating type I and mating type II lines were not correlated to cell size, a relation was found between lipid content and food ingestion capacity. A depletion of bacteria in the culture medium could be responsible for the lack of differences in mature starved cells. CLSM allowed us to gather volume information about the lipid droplet distribution within the cell.

Animals↗
PubMed1995

Immunofluorescence and confocal laser scanning microscopy studies of osteoblast growth and phenotypic expression in three-dimensional degradable synthetic matrices.

In the development of three-dimensional cell-polymer synthetic matrices for tissue regeneration, visualization of cells growing in these porous structures can be difficult. The focus of this study was the development and use of a novel method that would allow for visualization of osteoblasts inside opaque matrices. The morphologic responses and phenotypic characterization of osteoblasts as they attach, spread, and migrate through a porous three-dimensional biodegradable polymer-ceramic matrix in vitro were studied using immunofluorescence and confocal laser scanning microscopy (CLSM). CLSM offers several advantages over the most commonly used imaging methods [traditional light microscopy and scanning electron microscopy (SEM)]. CLSM filters out-of-focus background and provides more structural details of cells. In addition, CLSM does not require extensive sample preparation as does SEM. When used in conjunction with fluorescence-labeled antibodies to identify cells and their products, it can characterize morphology of growing cells and successfully determine phenotypic function. Using monoclonal antibody to osteocalcin, a bone cell-specific protein, cells throughout the matrix were found to have preserved osteoblast-like phenotype with growth. The morphology of cells throughout the matrix was found to be similar to osteoblast cells grown on tissue culture polystyrene and consisted of spread polygonal forms. Using the technique of CLSM with immunofluorescent antibodies, we have demonstrated for the first time that these three-dimensional degradable polymer matrices can support osteoblast growth and phenotypic expression throughout its structure.

Alkaline Phosphatase↗
PubMed2005

Visualizing regulatory T cell control of autoimmune responses in nonobese diabetic mice.

The in vivo mechanism of regulatory T cell (T(reg) cell) function in controlling autoimmunity remains controversial. Here we have used two-photon laser-scanning microscopy to analyze lymph node priming of diabetogenic T cells and to delineate the mechanisms of T(reg) cell control of autoimmunity in vivo. Islet antigen-specific CD4(+)CD25(-) T helper cells (T(H) cells) and T(reg) cells swarmed and arrested in the presence of autoantigens. These T(H) cell activities were progressively inhibited in the presence of increasing numbers of T(reg) cells. There were no detectable stable associations between T(reg) and T(H) cells during active suppression. In contrast, T(reg) cells directly interacted with dendritic cells bearing islet antigen. Such persistent T(reg) cell-dendritic cell contacts preceded the inhibition of T(H) cell activation by dendritic cells, supporting the idea that dendritic cells are central to T(reg) cell function in vivo.

Adoptive Transfer↗
PubMed2001

Detection and quantification of small numbers of circulating tumour cells in peripheral blood using laser scanning cytometer (LSC).

The detection of circulating tumour cells disseminated from solid tumours requires extremely sensitive methods. Molecular genetic methods, which are most sensitive, are not applicable to solid tumours because no tumour-specific genetic markers are available. Detection of disseminated tumour cells by immunocytochemistry is time-consuming, whereas fluorimetry is fast and quantitative. The laser scanning cytometer (LSC) provides an automated microscopic procedure for screening up to 5x10(4) cells in suitable time. Using this system together with an enrichment procedure which allows up to ten thousand-fold enrichment, we have quantified minimal numbers of tumour cells. In a model system, breast cancer cell line cells diluted into peripheral blood mimicked seeding of tumour cells into the periphery. After staining with fluorochrome-conjugated anti-epithelial antibody, slides were screened for positive events directly or after enrichment with antibody-coated magnetic beads. One positive cell was unequivocally detectable in 10(4) cells and 50 out of 60 tumour cells were reliably recovered from a 20 ml blood volume, equal to 1-2 cells per 10(7), after magnetic bead enrichment. This method allows quantitation of tumour cells in peripheral blood and bone marrow in reasonable time and will, for the first time, enable extensive investigation of the seeding behaviour of tumours.

Breast Neoplasms↗
PubMed1996

Detection of cellulose with improved specificity using laser-based instruments.

Specific detection of cellulose has not been possible using laser based instruments such as laser scanning confocal microscopes (LSCM) and fluorescently activated cell sorters (FACS). Common cellulose dyes are nonspecific and/or nonexcitable with common lasers. Furthermore, many lasers emit wavelengths that overlap with autofluorescence from chlorophyll and other plant molecules. We demonstrate that a cellulase and an isolated bacterial cellulose binding domain (CBD) conjugated to fluorescent dyes can be used for laser detection of cellulose with improved specificity. Cell walls of differentiating tracheary elements and spores of Dictyostelium discoideum were tested in this study. For double labeling, autofluorescence interfering with the rhodamine signal was eliminated by collecting each excitation channel separately followed by computer recombination or by using a narrow band pass barrier filter allowing simultaneous channel collection. Using these methods, cellulose and microtubules tagged with a monoclonal antibody to alpha-tubulin were effectively colocalized in chlorophyll-containing tracheary elements using a LSCM. Also, Dictyostelium discoideum spores labeled or unlabeled with CBD-FITC were separated into two populations by FACS indicating that this tag should be useful in future mutagenesis experiments. Therefore, the presence or absence of cellulose can now be analyzed using common lasers.

Animals↗
PubMed1997

In vitro differentiation of neural progenitor cells from prenatal rat brain: common cell surface glycoprotein on three glial cell subsets.

Glial progenitor cell differentiation and cell lineage relationships during brain development are complex hierarchical processes depending on genetic programming, cell-cell interactions, and microenvironmental factors. The identification of precursor cell-specific antigens provides a tool for the study of both normal development and deviations from lineage-specific differentiation associated with malignant transformation. Monoclonal antibody (mAb) RB13-6 recognizes a 130-kDa cell surface glycoprotein (gp130RB13-6) expressed by a subset of 9OAcGD3-positive glial precursor cells scattered in the rat neuroepithelium on prenatal day (PRD) 13. During prenatal development the fraction of gp130RB13-6-positive fetal brain cells (FBC) decreased from about 18% (PRD 14) to about 1.5% (PRD 22), coinciding with increasing fractions of more mature cell types, as indicated by the elevated expression of p24RB21-15, another cell surface determinant specified by mAb RB21-15 (Kindler-Röhrborn et al.; Differentiation 30:53-60, 1985) and other neural cell type-specific markers. Accordingly, gp130RB13-6 positive precursor cells were localized in the ventricular zones throughout brain development. Concomitant with their formation and in the adult rat brain, ependymal layers lining the ventricular surface, choroid plexus, and the leptomeninges were intensely labeled by anti-gp130RB13-6 mAb. As visualized by confocal laser scanning microscopy of FBC cultures from PRD 13, gp130RB13-6 was coexpressed with the RC1 antigen by progenitor cells morphologically resembling radial glia cells. In addition, a very small subpopulation of astrocytes coexpressing gp130RB13-6 and glial fibrillary acidic protein (GFAP; < 5%) occurred 3 days after seeding. Primary FBC cultures from PRD 18 contained an increased subset of astrocytes coexpressing gp130RB13-6 and GFAP (approximately 25% of all gp130RB13-6 expressing cells), apparently generated from gp130RB13-6-positive precursors. Corresponding to in vivo conditions, ciliated ependymal cells but also microglial cells/macrophages and leptomeningeal cells showed strong expression of gp130RB13-6 in culture. We thus present a new glycoprotein on the cell surfaces of a glial progenitor cell subset for further studies of cell lineage relationships between radial glia cells, astrocytes, and ependymal cells.

Animals↗