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E Sahar

Publications and source records attributed to E Sahar.

36 records · Page 2Linked to original sources
PubMed1988

T6 positive cells in the peripheral blood of burn patients: are they Langerhans cells precursors?

Peripheral blood mononuclear cells of 14 patients suffering thermal injury were separated by affinity chromatography on peanut agglutinin (PNA) coupled to Sepharose macrobeads. The resulting PNA positive subset was 14% of the total mononuclear population. About 30% of these cells were found to coexpress T6(CD1), Ia-like and the myeloid differentiation antigens My4(CDw14) and Mo1(CD11). In comparison, the PNA+ subset from normal blood donors (about 5% of total mononuclear cells) contained mature monocytes that were found to be T6 negative. Electron microscopic studies using immunogold labeling showed that the T6 positive cells were slightly smaller than monocytes but larger than the classical lymphocytes and had common morphologic features with the Langerhans cells of the skin. Considering that patients suffering extensive damage of the epidermis require fast renewal of all skin elements, it is possible that the cells we identified in their peripheral blood are the precursors of the Langerhans cells of the skin en route from bone marrow to the epidermis.

Adolescent↗
PubMed1987

Production of human pluripotent progenitor cell colony stimulating activity (CFU-GEMMCSA) in patients with myelodysplastic syndromes.

The present study was aimed at assessing the possible relationship between the T-lymphocyte abnormalities and the stem cell dysfunction in myelodysplastic syndromes (MDS), by investigating the production of specific stimulators of stem cell differentiation in such patients. Conditioned media from peripheral blood mononuclear cells (PBMNC) of MDS patients and healthy controls, prepared with or without phytohaemagglutinin (PHA), were assayed for their capacity to stimulate the in-vitro formation of multilineage colonies (CFU-GEMM) in target marrow cell cultures of healthy donors. Both PHA-induced DNA synthesis and T-cell subpopulation ratios (T4/T8) in patient cells were significantly lower than in controls. However, no impaired production of pluripotent progenitor cell colony stimulating activity (CFU-GEMMCSA) by PHA-stimulated and unstimulated PBMNC, could be found. Normal levels of activity were also produced by isolated T lymphocytes of MDS patients. Autologous serum neither enhanced nor suppressed the production of CFU-GEMMCSA. Our observations demonstrate that PHA-induced production of CFU-GEMMCSA is not directly correlated with DNA synthesis. Furthermore, we have found that both major T-cell subsets, defined by OKT4 and OKT8 monoclonal antibodies, are equally capable of producing CFU-GEMMCSA. The normal production of CFU-GEMMCSA by T cells of MDS patients suggests that this T-cell function is not an etiological factor in the stem cell disorder of myelodysplastic syndromes.

Anemia, Sideroblastic↗
PubMed1986

Marker and function analysis of natural killer and alloreactive T cells during early stages of dimethylbenzanthracene carcinogenesis. The immunity system during the precancer period.

The effect of the chemical carcinogen dimethylbenzanthracene (DMBA) on cellular immunity was studied at a 6-mg dose which induces adenocarcinomas and adenoacanthomas in more than 70% of BalB/c mice within 1 year after administration. DMBA caused a significant reduction of splenic natural killer (NK) activity and responsiveness to alloantigens in mixed lymphocyte reactions (MLR). These activities decreased soon after the carcinogen treatment and remained suppressed during the entire tumor induction period. There was a linear correlation between the reduction in NK activity and a selective decrease in the number of asialo GM1 positive cells in the spleen. However, cell sorting experiments using the flow cytometer have shown that the lytic activity per cell of asialo GM-1 positive cells in untreated mice and in DMBA-treated ones was similar. There was no correlation between the suppressed response of the T cells in MLR and the percentage of T cell subpopulations residing in the spleen of the DMBA-treated mice. The decrease in the number of NK cells and the reduced MLR activity in the spleen occurred simultaneously with a decrease in the potential of bone marrow precursor cells to reconstitute NK and MLR activity in the spleen of lethally irradiated mice. These results indicate that the carcinogen DMBA effects the immune system at various levels and either eliminates or inactivates precursor cells as well as mature lymphoid cells.

9,10-Dimethyl-1,2-benzanthracene↗
PubMed1986

Quantitation by flow cytometry of anthracycline drug uptake by peripheral blood and bone marrow cells in human leukemias.

The inherent fluorescence of the anthracycline drugs can be combined with flow cytometry to obtain a convenient and rapid method for the quantitation of anthracycline drug uptake by human leukemic cells. A good correlation exists between the average cellular fluorescence intensity and the amount of drug associated with the cell as measured by extraction. Cellular incorporation of adriamycin and daunomycin was determined in peripheral blood and bone marrow cells of human leukemic patients after standardized in vitro exposures. Differences in uptake were found between the different cell types, with cells of lymphatic origin incorporating less of the drugs than nonlymphatic cells. Preliminary observations made in two patients with nonlymphatic leukemia showed a correlation between the in vitro uptake of adriamycin and the clinical response.

Aged↗
PubMed1983

Maturation rates and transition probabilities of cycling cells.

The kinetics of cell cycling are analyzed in terms of equations incorporating a DNA synthesis rate continuously varying along S phase and a portion of G1 phase through which the cells advance in a probabilistic fashion. These equations are solved analytically for several steady state as well as nonsteady state populations of cells. Experiments are described illustrating how these results can be applied to obtain both the rate of DNA synthesis at each point along S-phase and the probability of transition through the nondeterministic portion of G1.

Animals↗
PubMed1983

Rapid identification of Streptococcus pyogenes by flow cytometry.

Flow cytometry combined with immunofluorescence of Streptococcus pyogenes was used to assay bacteria suspended in buffer solution and in saliva derived from throat swabs of healthy volunteers. The method allowed the enumeration of as few as 5 X 10(3) and 5 X 10(4) CFU per milliliter of buffer and saliva respectively. Controls including Streptococcus salivarius instead of Streptococcus pyogenes or buffer instead of specific antibodies confirmed the specificity of the detection of Streptococcus pyogenes in the samples. The results suggest that flow cytometry may serve as a basis for an automated reliable method for the diagnosis of streptococcal infections.

Culture Media↗
PubMed1983

A conjugate of adriamycin and monoclonal antibodies to Thy-1 antigen inhibits human neuroblastoma cells in vitro.

Antibodies to the differentation antigen Thy-1 were linked via a dextran-hydrazide bridge to adriamycin. These monoclonal antibodies, which were previously prepared against first-trimester human fetal cells, reacted with several human brain tissue tumors such as neuroblastoma, glioma, and leiomyosarcoma through a common differentiation antigen, Thy-1. The conjugates were tested in vitro for their cytotoxic effects on a neuroblastoma cell line LA-N-I. The conjugate maintained its full drug and antibody activities. Moreover, the specific conjugate exhibited a higher efficacy in inhibition of RNA synthesis and was more cytotoxic than the nonspecific Ig-drug conjugate. Fluorescent microscopy showed that the specific conjugate was able to penetrate the cell membrane. Both the specific antibody and the drug were also observed to accumulate in the cell nucleus. Flow microfluorometric analysis of cellular DNA traverse showed that the specific antibody-drug conjugate caused a higher accumulation of neuroblastoma cells in stage G2 of the cell cycle than did the free drug, perhaps indicating a more efficient prevention of the progression of cells through mitosis.

Antibodies, Monoclonal↗
PubMed1982

Isolation of mouse x-chromosome specific DNA from an x-enriched lambda phage library derived from flow sorted chromosomes.

A lambda phage library enriched in X(7) chromosomal material has been constructed from flow sorted chromosomes isolated from mice carrying the Cattanach translocation T(X;7)1Ct. The flow sorted fraction that was cloned contained 40% X(7) chromosomes, so that the resulting lambda phage library should be more than 10-fold enriched for X chromosomal DNA. Approximately 100,000 lambda phage clones were obtained; of these, at least 80% were recombinant. Three quarters of recombinants were positive for mouse repetitive DNA as detected either by phage plaque filter hybridization or by Southern blotting. Recombinant DNA inserts were prepared from some of the remaining nonrepetitive phage fraction. The X-chromosome specificity of cloned DNA inserts was tested by hybridization to DNA from mouse-hamster somatic cell hybrids that had retained all or most of the mouse X as the only mouse chromosome and by comparison of the extent of hybridization to DNA from male and female mice. Out of nine cloned unique sequence segments successfully examined thus far, two were presumably derived from the X. Possession of phage library highly enriched for mouse X DNA should facilitate molecular studies of the control of X chromosome gene expression.

Animals↗
PubMed1982

Flow cytometric characterization of the response of Fanconi's anemia cells to mitomycin C treatment.

DNA flow histogram analysis, using 33342 Hoechst as a stain, has been used to detect the effect of the potentially bifunctional alkylating agent, mitomycin C (MMC) on dermal fibroblasts from patients with Fanconi's anemia (FA), a hereditary human disease characterized by pancytopenia, hypersensitivity to DNA-crosslinking agents, congenital abnormalities and a predisposition for neoplasia. At 24 or 48 hr after a 2-hr exposure to 0.05 or 0.10 micrograms/ml MMC, (3)HdT incorporation was reduced to a greater extent in FA cells than in normal cells. Cells sorted from the last half of S phase showed a slightly greater inhibition of (3)HdT incorporation than did those sorted from the first half of S. Fanconi's anemia cells exhibited a marked accumulation in the G(2) + M peak of flow histograms following exposure to MMC. Twenty-four hr after treatment with .0.5 micrograms/ml MMC, the G(2) + M fraction of FA cells (eight lines) increased to more than 0.5 from a control value of approximately 0.02. Both normals (six lines) and heterozygotes (eight lines) showed, on the average, much less of a G(2) + M increment than did FA cells, even after exposure to 0.1 micrograms/ml MMC. Examination of cells sorted from the G(2) + M peak revealed that MMC-treated FA cells were blocked prior to mitosis. To determine whether the response of FA cells was specific for bifunctional alkylating agent, cells were also treated with ethylmethanesulfonate, a monofunctional agent. Twenty-four hours after exposure to 0.25 or 0.5 mg/ml ethylmethanesulfonate, FA and normal cells showed similar, small increases in the G(2) + M peak. The results suggest the utility of flow cytometry in the diagnostic evaluation of fibroblasts from patients suspected of having Fanconi's anemia.

Alkylating Agents↗
PubMed1982

Cytogenetic and flow cytometric studies of cells from patients with Fanconi's anemia.

Cells from patients wtih Fanconi's anemia are unusually sensitive to agents which are capable of crosslinking DNA. This increased sensitivity can be detected both by cytogenetic and flow cytometric methods. An elevated frequency of chromosome aberrations, which is further exaggerated by exposure of cells to DNA crosslinking agents, is a general feature of Fanconi's anemia. Information about the formation of sister chromatid exchanges in this disease is less consistent. Cytogenetic analysis of cells from patients with Fanconi's anemia can be compromised by a low mitotic index. This is reflected in an accumulation of cells In the G2 phase of the cycle, after exposure to the bifunctional alkylating agent, mitomycin C. New methods for differentiating individuals with Fanconi's anemia from unaffected individuals should be of empirical use and might also facilitate mechanistic studies of this disease.

Anemia, Aplastic↗
PubMed1981

Sister-chromatid exchange induction in Chinese hamster ovary cells by 8-methoxypsoralen and brief pulses of laser light: assessment of the relative importance of 8-methoxypsoralen--DNA monoadducts and crosslinks.

Brief (less than or equal to 20 nsec) 5-15 mJ pulses of near-ultraviolet laser light were employed to promote, in a controlled manner, the reaction between 8-methoxypsoralen and the DNA of Chinese hamster ovary (CHO) cells. According to Johnston et al. [21], a single brief laser pulse should induce only monoadducts between 8 methoxypsoralen and DNA, while multiple laser pulses are needed to induce 8-methoxypsoralen--DNA crosslinks. In the present study, a single laser pulse was found sufficient to cause a significant elevation in the sister-chromatid exchange (SCE) frequency in CHO cells exposed to 0.75-1.2 X 10(-5) M 8-methoxypsoralen. Additional laser pulses caused less than proportional further increments in SCEs, although the total yield of 8-methoxypsoralen adducts in similarly exposed solutions of isolated DNA increased linearly. Higher numbers of laser pulses induced detectable amounts of 8-methoxypsoralen--DNA crosslinks and distorted the DNA flow histogram of CHO cells in a manner consistent with retarded cell cycle traverse. It is concluded that 8-methoxypsoralen--DNA monoadducts are sufficient to induce SCEs in CHO cells, while 8-methoxypsoralen--DNA crosslinks might have relatively little overall effect on SCE frequencies. These results, together with previous data correlating SCE induction and mutagenesis [10] are discussed in the context of strategies for evaluating psoralen derivatives for their suitability in the phototherapy of skin disorders.

Animals↗
PubMed1980

Interactions between pairs of DNA-binding dyes: results and implications of chromosome analysis.

A number of DNA-binding dyes, with spectral properties making them suitable as components of energy donor-acceptor pairs, are described. If such pairs are used to stain metaphase chromosomes, and if the energy acceptor (e.g., actinomycin D or methyl green) has a binding specificity opposite to the binding or fluorescence specificity of the donor (e.g., 33258 Hoechst, quinacrine or chromomycin A3), contrast in donor fluorescence can be enhanced, leading to patterns selectively highlighting standard or reverse chromosome bands or particular polymorphic regions. Such results presumably reflect chromosomal regions enriched in 10-20 base pair clusters to which the donor binds and fluoresces but to which the acceptor cannot bind. For other pairs, involving counterstains such as netropsin or echinomycin, which are not suitable as energy acceptors, specific changes observed in polymorphic region fluorescence are most likely due to binding competition between dyes. Dye pairs producing contrast by either method can be used to differentiate between homologous chromosomes or to facilitate detection of specific chromosomal rearrangements. Preliminary data indicate that contrast enhancement generated in fixed metaphase chromosomes spread on microscopic slides can also be observed in suspensions of unfixed metaphase chromosomes, reinforcing the expectation that the methodology described will be of use in flow cytometry.

Animals↗
PubMed1980

Energy transfer and binding competition between dyes used to enhance staining differentiation in metaphase chromosomes.

The ability of electronic energy transfer and direct binding competition between pairs of dyes to enhance contrast in human or bovine metaphase chromosome staining patterns is illustrated, and the relative effectiveness of these two mechanism compared. The existence of energy transfer between quinacrine or 33258 Hoechst and 7-amino-actinomycin D in doubly stained chromosomes is demonstrated directly by microfluorometry. The ability of the dyes 7-amino-actinomycin D, methyl green, or netropsin, acting as counterstains, to displace quinacrine, 33258 Hoechst, or chromomycin A3 from chromosomes, is estimated by quantitative analysis of energy transfer data, by photobleaching of the counterstains, or by selective removal of counterstains by appropriate synthetic polynucleotides. Effects on the fluorescence of soluble 33258 Hoechst-DNA complexes due to energy transfer or binding displacement, by actinomycin D or netropsin, respectively, are further differentiated by nanosecond fluorescence decay measurements. Examples are presented of dye combinations for which (a) energy transfer is the primary mechanism operative, (b) binding competition exists, with consequences reinforcing those due to energy transfer, or (c) binding competition is the most important interaction. These analyses of mechanisms responsible for contrast enhancement in doubly stained chromosomes are used to derive information about the relationship between chromosome composition and banding patterns.

Animals↗
PubMed1979

Pairs of fluorescent dyes as probes of DNA and chromosomes.

If two fluorescent dyes with different binding or fluorescence specificities are used simultaneously to stain DNA or chromosomes, the ratio of their fluorescent signals can provide information about base composition or base analogue substitution. Energy transfer between such dye pairs, possible if the fluorescence spectrum of one overlaps the absorption spectrum of the other, can modify observed fluorescence. Microfluorometric measurements were used to document the occurrence of energy transfer between quinacrine or 33258 Hoechst as energy donor and ethidium or 7-aminoactinomycin D as acceptor when used jointly to stain cytologic preparations of human metaphase chromosomes. Use of 7-aminoactinomycin D, a dye with G-C binding specificity, as energy acceptor permitted the identification of human chromosome regions presumptively enriched for clusters of A-T base pairs, based on the resistance of A-T specific fluorescence, from quinacrine or 33258 Hoechst, to energy transfer dependent quenching. The results provide information about basic structural features of metaphase chromosomes, and the associated methodology may prove useful in accentuating specific fluorescent polymorphic chromosome regions.

Cells, Cultured↗
PubMed1978

Enhancement of banding patterns in human metaphase chromosomes by energy transfer.

Intermolecular energy transfer between appropriately chosen pairs of dyes can be used to induce or enhance banding patterns in human metaphase chromosomes. Energy transfer, calibrated by fluorometric studies on soluble dye.DNA complexes, can also be detected by photometric measurements on cytological preparations of metaphase chromosomes stained with pairs of fluorochromes. If a fluorescent dye with one type of binding or quantum yield specificity (e.g., quinacrine, 33258 Hoechst, or chromomycin A(3)) is employed together with a counterstain (e.g., actinomycin D, 7-aminoactinomycin D, or methyl green) exhibiting a complementary base pair binding specificity and satisfying spectral overlap criteria for energy transfer, contrast in fluorescence from the first dye is enhanced in specific subsets of standard chromosome bands. Extensive energy transfer presumably suppress donor fluorescence except in chromosomal region containing clusters of at least 20 base pairs predominantly of one type, within which the donor but not the acceptor can bind and fluoresce. Quinacrine-bright polymorphic regions are especially resistant to fluorescence quenching by counterstains with G.C binding specificity, strengthening the evidence that these latter regions are highly enriched for A.T base pair clusters. The ability to highlight selectively many such polymorphic regions may prove of further, practical, utility in a number of cytogenetic problems.

Chromosomes, Human↗