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Biomedical subjects

G R Johnson

Publications and source records attributed to G R Johnson.

At least 217 records · Page 12Linked to original sources
PubMed1980

Detection of a new type of mouse eosinophil colony by Luxol-fast-blue staining.

Staining with Luxol-fast-blue was shown to be a satisfactory method for identifying mouse eosinophils. IN agar cultures of mouse marrow stimulated by pokeweed mitogen-stimulated spleen conditioned medium (SCM), 2-10 compact Luxol-fast-blue positive eosinophil colonies varied according to the batch of human plasma used in the culture medium. Similar colony-forming cells were detected in the spleen and peripheral blood but not in the thymus or lymph nodes. Some Luxol-fast-blue positive eosinophil colonies were stimulated to develop by crude mouse lung conditioned medium and by high concentrations of GM-CSF purified from this source. The cells forming Luxol-fast-blue positive eosinophil colonies sedimented more rapidly (5.5-6.5 mm/h) than the cells forming dispersed-eosinophil colonies 94.5 mm/h). Transfer studies using intact colonies or redispersed colony cells failed to demonstrate an interrelationship between the two types of eosinophil colonies and the cells forming Luxol-fast-blue positive eosinophil colonies appear to be a distinct subset of eosinophil percursors.

Animals↗
PubMed1979

Colony formation in agar by multipotential hemopoietic cells.

Agar cultures of CBA fetal liver, peripheral blood, yolk sac and adult marrow cells were stimulated by pokeweed mitogen-stimulated spleen conditioned medium. Two to ten percent of the colonies developing were mixed colonies, documented by light or electron microscopy to contain erythroid, neutrophil, macrophage, eosinophil and megakaryocytic cells. No lymphoid cells were detected. Mean size for 7-day mixed colonies was 1,800-7,300 cells. When 7-day mixed colonies were recloned in agar, low levels of colony-forming cells were detected in 10% of the colonies but most daughter colonies formed were small neutrophil and/or macrophage colonies. Injection of pooled 7-day mixed colony cells to irradiated CBA mice produced low numbers of spleen colonies, mainly erythroid in composition. Karyotypic analysis using the T6T6 marker chromosome showed that some of these colonies were of donor origin. With an assumed f factor of 0.2, the mean content of spleen colony-forming cells per 7-day mixed colony was calculated to vary from 0.09 to 0.76 according to the type of mixed colony assayed. The fetal and adult multipotential hemopoietic cells forming mixed colonies in agar may be hemopoietic stem cells perhaps of a special or fetal type.

Animals↗
PubMed1979

Interactions between purified GM-CSF, purified erythropoietin and spleen conditioned medium on hemopoietic colony formation in vitro.

Preincubation of C57BL adult marrow cells or CBA fetal liver cells with a 250-fold excess concentration of purified GM-CSF failed to reduce the frequency of cells forming eosinophil, megakaryocyte or erythroid colonies in subsequent agar cultures. When excess concentrations of purified GM-CSF were added to agar cultures stimulated by pokeweed mitogen-stimulated spleen conditioned medium (SCM), no reduction was observed in the frequency of eosinophil, megakaryocyte or erythroid colonies. Addition of 4 units of purified erythropoietin (EPO) to cultures of fetal liver or adult marrow cells stimulated by SCM increased the number of erythroid colonies but did not reduce the number of non-erythroid colonies or the non-erythroid content of mixed erythroid colonies. Although neither GM-CSF nor EPO alone was able to stimulate erythroid colony formation in agar cultures of fetal liver cells, small numbers of large erythroid colonies were stimulated to develop in cultures containing both purified regulators. Purified GM-CSF was also able to support the survival in vitro of a small proportion of erythroid colony-forming cells in fetal liver populations cultured initially in the absence of SCM and the survival of some eosinophil and megakaryocyte colony-forming cells in similar cultures of adult marrow cells. The results do not support the hypothesis that GM-CSF and EPO compete for a common pool of uncommitted progenitor cells. On the contrary, the data indicate that GM-CSF und EPO are able to collaborate in stimulating the proliferation of some erythropoietic cells. Furthermore, purified GM-CSF appears to be able to support temporarily the survival and/or initial proliferation of at least some cells forming erythroid, eosinophil and megakaryocyte colonies, even though GM-CSF is unable to stimulate the formation of colonies of these types.

Animals↗
PubMed1979

Peritoneal cell population of mice infected with Mesocestoides corti as a source of eosinophils.

A prominent feature of the inflammatory cellular response in the peritoneal cavity of Mesocestoides corti-infected mice is a marked and sustained increase in the number of eosinophils. In intact mice, the total number of nucleated cells in the peritoneal cavity rises from less than 5 X 10(6) to more than 50 X 10(6) and, at certain time points, in excess of 50% of these cells are eosinophils. Peritoneal eosinophils are absent in infected hypothymic nude (nu/nu) mice of three genotypes, and eosinophils counts can be elevated in infected nude mice by injection of peripheral lymphoid cells or thymocytes. The peritoneal cells of M. corti-infected mice are a convenient starting cell population for eosinophil purification.

Animals↗
PubMed1979

Separation of functionally distinct human granulocyte-macrophage colony-stimulating factors.

Human placental conditioned medium (HPCM) contans colony-stimulating factors (CSFs) required for the growth in vitro of neutrophilic granulocyte-macrophage (GM) and eosinophilic (EO) progenitor cells from human bone marrow. Fractionation of CSFs in HPCM was achieved by manipulation of the elution conditions on a column of phenyl-Sepharose. After equilibration of the phenyl-Sepharose column at high ionic strength (1 M ammonium sulfate), all of the CSF bound; one species of GM-CSF (alpha) and all of the elutable EO-CSF were eluted from the column simply by reducing the salt concentration, whereas the second species of GM-CSF (beta) was free of EO-CSF and was eluted only by increasing the concentration of tehylene glycol in the elution buffer. The two GM-CSFs were functionally distinct. GM-CSF alpha preferentially stimulated colony formation by day 14 of culture, and there was a decreased proportion of neutrophil colonies and increased proportion of macrophage colonies as the strength of the stimulus was decreased; GM-CSF beta, on the other hand, preferentially stimulated colony formation by day 7 of culture, and the proportion of neutrophil colonies was high (average 80%) and independent of the concentration of GM-CSF beta. GM-CSF alpha and GM-CSF beta were indistinguishable on the basis of apparent molecular size on tel filtration columns (molecular weight 30,000), charge properties on isoelectric focusing beds (isoelectric point, 4.9), and were not related to each other as a sialoglycoprotein is related to its asialo form. Adherent cell removal of the target bone marrow cells (to remove colony-stimulating cells) suggested that both GM-CSFs acted directly rather than by stimulating the production of GM-CSF. Mixing and titration experiments indicated that the differences in functional specificities of the two GM-CSFs (and the lack of EO-CSF associated with GM-CSF beta) were not due to the presence of specific inhibitory molecules or lower absolute levels of CSF in one fraction relative to the other. These two species of GM-CSF should be useful in separately enumerating subpopulations of different GM-progenitor cells inhuman hemopoietic disorders.

Bone Marrow Cells↗
PubMed1978

Monoclonal origin of B lymphocyte colony-forming cells in spleen colonies formed by multipotential hemopoietic stem cells.

Spleen colonies produced by transplanting lethally irradiated mice with either 12 day fetal liver or adult bone marrow cells were found to contain B- lymphocyte colony-forming cells (BL-CFC) . The proportion of BL-CFC positive spleen colonies did not increase substantially between 8 and 14 days after transplantation, the range being 18-45 percent. However, the absolute number of BL-CFC per spleen colony varied considerably (between 1 and 10,318), although the majority of colonies contained less than 200 BL-CFC. Irrespective of the time after transplantation, smaller spleen colonies were found to have a higher frequency of BL-CFC than larger spleen colonies. To determine the possible clonal origin of BL-CFC from spleen colony- forming unit (CFU-S), CBA mice were injected with equal numbers of CBA and CBA T(6)/T(6) fetal liver or adult bone marrow cells. Analysis of 7-15-day spleen colonies demonstrated that 90 percent were either exclusively T(6) positive or T(6) negative and approximately equal numbers ofboth colony types were observed. B-lymphocyte colonies were grown and successfully karyotyped from 19 spleen colonies. When compared with the original spleen colony karyotype the B-lymphocyte colony cells karyotype was identical in all 19 cases. In 3 of the 19 colonies analyzed a mixture of T(6) positive and T(6) negative karyotypes was present and identical proportions of the karyotypes were present in the pooled B-lymphocyte colony cells and spleen colony cells. The data indicate that the B-lymphocyte colony-forming cells detected in spleen colonies are genuine members of the hemopoietic clone derived from the initiating hemopoietic stem cell (CFU-S).

Animals↗
PubMed1978

Surface immunoglobulin phenotype of murine spleen cells which form B cell colonies in agar.

Separation of IgD-bearing B lymphocytes from the spleens of 3-week-old or adult mice has been achieved using cellular affinity columns and an anti-delta allotype serum. The results demonstrate that the majority of cells capable of forming B lymphocyte colonies in agar are delta-positive although they lose detectable IgD upon stimulation. It is proposed that these cells are the descendants of IgD-negative cells found in the fetal liver and neonatal spleen.

Animals↗
PubMed1978

The anti-delta suppressed mouse.

The ontogeny of IgD was examined in neonatal to adult mice by immunofluorescence. The first delta-positive cells appeared in the spleen at about 3 days of age, while lymph nodes from 4 to 5-day-old mice already contained 10--20% delta-positive B cells. In order to study the function of IgD, we developed a system in which the appearance of delta-positive cells was suppressed by treatment of mice from birth with an anti-delta allotype serum. Immunofluorescence studies showed that very few delta-positive cells could be found in the spleen and lymph nodes of such suppressed mice, and also that the percentage of mu-positive cells was reduced by about 50%. The recovery from suppression was rapid, indicating that potential delta-positive cells were modulated rather than killed. Studies of the immune response of suppressed mice in vitro and in vivo showed that suppression had little effect on the IgM response but caused a definite reduction in the IgG response.

Aging↗
PubMed1978

Nature of cells forming erythroid colonies in agar after stimulation by spleen conditioned medium.

Erythroid colony formation in agar cultures of CBA cells was stimulated by the addition of pokeweed mitogen-stimulated C57BL spleen conditioned medium. Both 48-hour colonies ("48-hour benzidine-positive aggregates") and day 7 large burst or unicentric erythroid colonies ("erythroid colonies") developed, together with many neutrophil and/or macrophage colonies. In CBA mice, the cells forming erythroid colonies occurred with maximum frequency (650/10(5) cells) in 10- to 11-day-old yolk sac and fetal liver but were present also in fetal blood, spleen and bone marrow. The frequency of these cells fell sharply with increasing age and only occasional cells (2/10(5) cells) were present in adult marrow. A marked strain variation was noted, CBA mice having the highest levels of erythroid colony-forming cells. The erythroid colony-forming cells in 12-day CBA fetal liver were radiosensitive (DO 110-125 rads), mainly in cycle and were non-adherent, light density, cells sedimenting with a peak velocity of 6-9 mm/hr. These properties are similar to those of other hemopoietic progenitor cells in fetal tissues. The relationship of these apparently erythropoietin-independent erythroid colony-forming cells to those forming similar colonies after stimulation by erythropoietin remains to be determined.

Animals↗
PubMed1978

Haemopoietic stem cells in the foetal mouse thymus.

The presence of haemopoietic stem cells (HSC) in the foetal mouse thymus was assessed to determine whether all cells which enter the developing organ are precommitted to thymocyte differentiation, or if stem cell multipotentiality still exists. The Till and McCulloch spleen colony assay was used to delineate foetal-thymus derived HSC in lethally irradiated recipients. Of the range examined, between 13 days of gestation to birth, a peak of stem cell activity occurs in 15-day foetal thymus. The surface colonies produced by the thymus-derived HSC are small compared to colonies produced by the liver derived HSC, although well within the range of the latter. Histologically, five types of colonies were identifiable which were produced by the thymus-derived HSC, indicating that these cells retain the potential to form a wide range of differentiated colonies.

Animals↗
PubMed1978

Molecular and biological properties of a macrophage colony-stimulating factor from mouse yolk sacs.

A colony-stimulating factor (M-CSF) has been partially purified and concentrated from mouse yolk sac-conditioned medium (YSCM). M-CSF appeared to preferentially stimulate CBA bone marrow granulocyte-macrophage progenitor cells (GM-CFC) to differentiate to form macrophage colonies in semisolid agar cultures. By comparison, colony-stimulating factor (GM-CSF) from mouse lung-conditioned medium (MLCM) stimulated the formation of granulocytic, mixed granulocytic-macrophage, and pure macrophage colonies. Mixing experiments indicated that both M-CSF and GM-CSF stimulated all of the GM-CFC but that the smaller CFC were more sensitive to GM-CSF and that the larger CFC were more sensitive to M-CSF. Almost all developing "clones" stimulated initially with M-CSF continued to develop when transferred to cultures containing GM-CSF. In the converse situation, only 50% of GM-CSF prestimulated "clones" survived when transferred to cultures containing M-CSF. All clones initially stimulated by M-CSF or transferred to cultures stimulated by M-CSF contained macrophages after 7 days of culture. These results suggest that there is a population of cells (GM-CFC) that are capable of differentiating to form both granulocytes and macrophages, but, once these cells are activated by a specific CSF (e.g. M-CSF), they are committed to a particular differentiation pathway. The pattern of CFC differentiation was not directly related to the rate of proliferation: cultures maximally stimulated by M-CSF produced mostly macrophage colonies, but the presence of small amounts of GM-CSF produced granulocytic cells in 30% of the colonies. Gel filtration, polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate, and affinity chromatography with concanavalin A-Sepharose indicated that M-CSF from yolk sacs was a glycoprotein with an apparent molecular weight of 60,000. There was some heterogeneity of the carbohydrate portion of the molecule as evidenced by chromatography on concanavalin A-Sepharose.

Animals↗