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Biomedical subjects

H R MacDonald

Publications and source records attributed to H R MacDonald.

At least 343 records · Page 19Linked to original sources
PubMed1975

Serum potentiation of granulocyte and macrophage colony formation in vitro.

Addition of serum or plasma from many species increased the number and size of neutrophilic granulocyte and macrophage colonies formed in vitro by mouse bone marrow cells after stimulation by a variety of preparations containing colony stimulating factor (CSF). The active factor in serum was heat-labile, non-dialyzable and eluted from G-100 columns in the serum albumin region. Serum non-selectively affected all cells potentially capable of forming granulocytic and macrophage colonies but did not potentiate the formation of eosinophilic or megakaryocytic colonies. The potentiating activity of mouse serum was increased by prior injection of the mice with endotoxin but was not affected by prior whole-body irradiation. Colony potentiating activity in plasma from patients with myeloid of lymphoid leukemia did not differ from that of normal human plasma.

Animals↗
PubMed1975

Generation of cytotoxic T lymphocytes in vitro. VI. Effect of cell density on response in mixed leukocyte cultures.

Reexposure of day 14 murine mixed leukocyte culture (MLC) populations to the original irradiated allogeneic stimulating spleen cells has previously been found to result in the ratpid generation of cytolytic T lymphocytes (CTL) associated with a net increase in cultured cell number. Under the experimental conditions used, day 5 MLC cells appeared unable to respond to the allogeneic stimulus. In order to characterize further the development of the potential for anamnestic reactivity during the course of MLC, C57BL/6 spleen cells were incubated with irradiated (1000 rads) DBA/2 spleen cells (primary MLC) for up to 3 weeks. At various time intervals after the onset of the primary MLC, the surviving cells were collected and reexposed, at varying cell concentrations, to irradiated DBA/2 spleen cells (secondary MLC). At daily intervals thereafter, CTL activity was assessed using a quantitative 51Cr-release assay system. A paradoxic effect of responding cell concentration on generation of CTL activity was observed; relatively greater increase in CTL activity was observed as the concentration of responding cells was decreased over a 100-fold range. This effect was more pronounced with responding cells reexposed to antigen after primary MLC for 20 days, but was observed even with normal cells. The apparent unresponsiveness of day 5 MLC cells to alloantigen restimulation could be overcome by simple dilution of responding cells. Cytotoxic activity at the time of restimulation with antigen seems to be a major factor determining the magnitude of the secondary response. Since intact cells bearing alloantigens are required for the generation of CTL in MLC, residual cytotoxic cells reduce the effective antigenic stimulus by destroying stimulating cells. This effect of concentration of responding cells on generation of CTL in MLC complicates interpretation of experiments investigating the role of "inhibitor" and "helper" cell in cell-mediated immune responses occurring in vitro. Under optimal conditions, the highest CTL activity and the largest increase in total cell number was observed 4 days after restimulation of day 10 MLC cells. On a per cell basis, the lytic activity was up to 4 times greater than that observed at the peak of a primary response, and the number of viable cells recovered was nearly 20 times higher than that at the onset. Such secondary MLC are thus a convenient source of lymphoid cells selected primarily on the basis of proliferation induced by alloantigens.

Animals↗
PubMed1974

Generation of cytotoxic T lymphocytes in vitro. III. Velocity sedimentation studies of the differentiation and fate of effector cells in long-term mixed leukocyte cultures.

Separation of cells by velocity sedimentation at unit gravity was utilized to investigate the physical properties of cytotoxic thymus-derived lymphocytes (CTL) generated in long-term mixed leukocyte cultures (MLC). In kinetic studies, CTL were found almost exclusively in the large cell fractions at the peak of the response on day 4, whereas the majority of CTL in day 14 MLC had the sedimentation properties of small lymphocytes. Reculture until day 14 of cells fractionated on the basis of size on day 4 indicated that the small CTL were derived exclusively from cells which had been large on day 4. Re-exposure of day 14 MLC cells to the original stimulating alloantigens resulted in significant cell proliferation and rapid regeneration of CTL activity. Cell fractionation experiments demonstrated that the cells in the day 14 MLC population which responded to the secondary allogeneic stimulus were small T lymphocytes, and that these cells rapidly developed into large, highly cytotoxic CTL following stimulation. Moreover, by restimulating on day 14 fractions which were selected on the basis of size on day 4, it was found that the responding small lymphocytes were themselves the progeny of cells which were large at the peak of the response. Since CTL and CTL progenitors showed concomitant changes in physical properties with time, the possibility exists that they belong to the same cell lineage, and hence that CTL can differentiate into cells which are no longer cytotoxic, but capable of mounting an anamnestic response.

Animals↗
PubMedDate not supplied

Differentiation in vitro of Lyt 2+ thymocytes from embryonic Lyt 2- precursors.

In mice, the thymus is regarded as being the primary anatomical site for the generation of immunologically competent T lymphocytes. Such cells comprise approximately 20% of the cells in the thymus and share with T lymphocytes from peripheral lymphoid tissues certain phenotypic properties defined by anti-Lyt antibodies. Thus, most immunocompetent T cells are either Lyt 1+2+ or Lyt 1+2- with the former cells being restricted to recognizing antigen in association with class I (H-2K, D) major histocompatability complex (MHC) products and the latter to class II (H-21) MHC products. Although evidence suggests that Lyt 1+2+ cells are generated from Lyt 1+2- precursor the independent development of two separate Lyt-defined lineages of thymocytes could not be ruled out. Here, the acquisition of Lyt 2 antigen by Lyt 2- cells from late embryonic and early postnatal thymuses is directly demonstrated. Furthermore, by combining cell cycle and Lyt phenotype analysis on a flow microfluorometer, the role of cell division in this differentiation process has been investigated.

Age Factors↗
PubMedDate not supplied

Expression of interleukin-2 receptors as a differentiation marker on intrathymic stem cells.

The thymus is regarded as the primary site for T-cell lymphopoiesis, but very little is known about the lineage inter-relationships of cells within that organ. At least four subpopulations of mouse thymocytes can be defined on the basis of staining with monoclonal antibodies directed against the T-cell differentiation antigens Lyt-2 and L3T4 (ref. 2). Thus immunocompetent (medullary) thymocytes, like peripheral T cells, express either Lyt-2 (cytotoxic phenotype) or L3T4 (helper phenotype) but not both, whereas non-functional (cortical) thymocytes express both markers. In addition, a small subpopulation comprising 2-3% of cells in the thymus and expressing neither Lyt-2 nor L3T4 has recently been described. The latter cells have the properties of intrathymic 'stem cells' in that they are the first to appear in the embryonic thymus and at least some can be shown to give rise, both in vivo (ref. 4. and our unpublished data) and in vitro, to other thymocyte subpopulations. We show here that 50% of Lyt-2-/L3T4- cells in the adult thymus express receptors for the polypeptide growth hormone interleukin-2 (IL-2) whereas other cells in the thymus do not. Furthermore, immunohistochemical localization studies on frozen sections indicate a disperse distribution of IL-2 receptor-positive cells in both the cortex and medulla. These novel findings have potential implications in the context of current models of differentiation pathways within the thymus.

Age Factors↗
PubMedDate not supplied

Similarities between interleukin-2 receptor number and affinity on activated B and T lymphocytes.

Interleukin-2 (IL-2) is a T-cell-derived polypeptide hormone of 133 amino acids which exerts its growth-promoting activity via a surface receptor. Originally, IL-2 was believed to be a unique growth factor for activated T cells; more recent studies, however, have demonstrated that certain B-cell tumours as well as normal activated B lymphocytes express a surface molecule which is recognized by monoclonal antibodies directed against the IL-2 receptor. Furthermore, we and others have shown recently that activated B cells proliferate in response to either immunoaffinity-purified or recombinant IL-2. These controversial findings prompted us to undertake a detailed quantitative comparison of IL-2 receptor expression on activated B and T cells. We show here, using biosynthetically labelled IL-2(3H-IL-2) and anti-IL-2 receptor antibody (3H-PC61) that activated B and T cells express both high-affinity (apparent dissociation constant, Kd approximately 20 pM) and low-affinity (Kd approximately 1,000 pM) IL-2 receptors. Binding of IL-2 to both classes of receptor is inhibited by the monoclonal anti-IL-2 receptor antibody PC61. B blasts express half as many total IL-2 binding sites or PC61 binding sites as T blasts, and the ratio of the number of low- to high-affinity receptors for each cell type is approximately 10:1. Immunoprecipitation analysis of surface-labelled blasts indicates that B and T cells have IL-2 receptors of similar relative molecular mass. Taken together, these data suggest strongly that IL-2 can act as a growth hormone for both B and T lymphocytes.

Animals↗
PubMedDate not supplied

Recombinant-derived interleukin-1 alpha stabilized against specific deamidation.

Recombinant-derived human interleukin-1 alpha (IL-1 alpha), purified from Escherichia coli, was resolved by isoelectric focusing on polyacrylamide gels into two species of isoelectric points (pI) 5.45 and 5.20, which constituted approximately 75% and approximately 25% of the total IL-1 alpha protein respectively. The pI 5.45 and pI 5.20 species were separated by chromatofocusing and subjected to N-terminal sequence analysis. The pI 5.45 species contained the expected Asn residue at position 36 of the mature protein sequence whereas the pI 5.20 species contained an Asp residue at the same position. A mutant protein in which Asn-36 was substituted for a Ser residue was isolated from E. coli and shown to be homogeneous on isoelectric focusing analysis with a pI = 5.45. 1H-n.m.r. and circular dichroism analyses of wild-type and the mutant IL-1 alpha indicated a similar conformation which was also indicated by the identical receptor binding affinities of IL-1 alpha with Asn, Asp or Ser in position 36. The mutant protein was stabilized against specific base-catalysed and temperature-induced deamidation, and may be more suitable than the wild-type position for physical and structural studies.

Amino Acids↗
PubMedDate not supplied

Endometrial cancer patients have a significant risk of harboring isolated tumor cells in histologically negative lymph nodes.

In this study, we examine the prevalence of finding isolated tumor cells (ITCs) in negative lymph nodes of endometrial cancer patients using immunohistochemistry. Seventy-six endometrial cancer patients with lymph nodes histologically negative for metastatic disease were examined. Nodal tissue sections were stained with anticytokeratin antibodies AE-1 and CAM 5.2. Nodes with single or groups of cells (two to four cells) < or =0.2 mm and showing cytokeratin reactivity were positive for ITCs. Findings were compared to features of the primary tumor and patient outcome. ITCs were present in 31 of 1712 lymph nodes. Fifteen (19.7%) patients had ITC-positive nodes. ITCs involved only pelvic nodes in nine cases, only para-aortic nodes in five cases, and pelvic and para-aortic in one case. Tumor in adnexa was the only pathologic feature associated with nodal ITCs (P= 0.0485). All 15 patients with nodal ITCs were alive at follow-up. One (6.7%) patient suffered recurrent disease but was alive at last encounter. Disease recurred in 5 (8.8%) of 57 patients without nodal ITCs. Two are alive without disease, two alive with disease, and one died from her cancer. In summary, a significant proportion of endometrial cancer patients have ITCs detected by immunohistochemistry in histologically negative regional lymph nodes.

Adult↗