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M F Heyworth

Publications and source records attributed to M F Heyworth.

At least 37 records · Page 2Linked to original sources
PubMed1989

Generation and characterization of monoclonal antibodies against Giardia muris trophozoites.

Mouse monoclonal antibodies (mAb) were produced against Giardia muris trophozoite surface antigens. To generate B-cell hybridomas, P3/NS1/1-Ag4-1 myeloma cells were fused with splenic lymphocytes from BALB/c mice that had been immunized parenterally with G. muris trophozoites. Hybridoma culture supernatants were screened for mAb by flow cytometry of G. muris trophozoites incubated with culture supernatant followed by fluorescein-conjugated anti-mouse IgG and IgM. Flow cytometry showed three types of trophozoite staining by mAb: (i) bright staining of greater than 90% of trophozoites, with aggregation of the organisms; (ii) bright staining of approximately 90% of trophozoites, with little or no aggregation; (iii) dull staining of approximately 20% of trophozoites, without aggregation. Western blotting of mAb on G. muris trophozoite antigens separated by polyacrylamide gel electrophoresis showed that a mAb exhibiting the third of these flow cytometry staining patterns recognized trophozoite antigens of MW approximately 31,000 and 35,000. Immunoprecipitation studies indicated that the same mAb specifically precipitated two 125I-labelled trophozoite surface antigens of MW approximately 30,000. Monoclonal antibodies generated in this study may facilitate the purification and biochemical characterization of trophozoite antigens that are targets for protective intestinal antibody in G. muris-infected mice.

Animals↗
PubMed1988

Generation of L3T4-/Ly-2- T cells in Peyer's patches of mice treated with monoclonal antibody against helper T cells.

The effect of in vivo administration of anti-L3T4 monoclonal antibody on the generation of an L3T4-/Ly-2- (CD4-/CD8-) population of Thy-1.2+ cells was examined in Peyer's patches of mice by two-color flow cytometry. Female BALB/c mice aged 8 wk were given 4-6 weekly injections of either anti-L3T4 (1 mg/wk) or phosphate-buffered saline (control group), and dispersed Peyer's patch cells analyzed for the presence and absence of L3T4 and Ly-2 on Thy-1.2+ cells. In anti-L3T4 treated mice, L3T4-/Ly-2- T cells accounted for 25-30% of the Thy-1.2+ cells, whereas in control mice these cells represented only 3-4% of the T cells. The remaining 70-75% of the Thy-1.2+ cells after anti-L3T4 treatment were Ly-2+ and not L3T4+. The L3T4-/Ly-2- population of Thy-1.2+ cells is a novel subset which has not been previously found in Peyer's patches and is amplified when helper T cells are depleted.

Animals↗
PubMed1988

Enzyme-linked immunosorbent assay for Giardia-specific IgA in mouse intestinal secretions.

We describe an ELISA for trophozoite-specific IgA in the intestinal secretions of mice infected with Giardia muris. Using this method, trophozoite-specific IgA was demonstrated in intestinal secretions of Giardia-infected immunocompetent BALB/c mice. Such antibody was undetectable in the intestinal secretions of Giardia-infected athymic (nude) mice. Immunocompetent BALB/c mice are able to clear G. muris infection whereas nude mice are not. The study provides evidence that the chronicity of G. muris infection in nude mice results from lack of intestinal trophozoite-specific IgA in these animals. By means of the ELISA, trophozoite-specific IgA was demonstrated in intestinal secretions from immunocompetent mice in the absence of protease inhibitors.

Animals↗
PubMed1988

Modulation of lymphocyte subsets in Peyer's patches of mice treated with monoclonal antibody against helper T-cells.

Treatment of mice with anti-L3T4, a monoclonal antibody directed against helper T-cells, impairs clearance of intestinal Giardia muris infection. The present study examined the effect of anti-L3T4 treatment on mouse Peyer's patch cytoarchitecture and on the distribution of T-cell subsets within microenvironments of the follicle. Female BALB/c mice, aged 8 weeks, were given 4-7 weekly injections of either anti-L3T4 (1 mg/wk) or PBS (control group), and Peyer's patches were examined by immunohistochemistry or flow cytometry. In anti-L3T4-treated mice, Peyer's patch follicles (B-cell areas) were about two thirds the size of follicles in controls, and virtually all the size difference occurred in germinal centers. Peyer's patches were depleted of L3T4+ cells, yet the proportion of Thy-1.2+ (all T) cells was not decreased correspondingly, and the distribution of Thy-1.2+ cells in the patches was similar to that in control mice. In anti-L3T4-treated mice, Thy-1.2+ cells consisted of (a) Ly-2+ (cytotoxic/suppressor T) cells, and (b) a population of Thy-1.2+ cells that were neither L3T4+ nor Ly-2+. After treatment, Ly-2+ cells accounted for most of the T-cells in interfollicular areas and were also scattered in follicles, in germinal centers, and below the dome epithelium--in areas where L3T4+ cells predominated in control mice. Thy-1.2+ cells that were L3T4- and Ly-2- were mainly localized below the dome epithelium. These shifts indicate complex interrelationships among different lymphocyte subsets in Peyer's patches.

Animals↗
PubMed1988

Time-course of Giardia muris infection in male and female immunocompetent mice.

The time-course of acute Giardia muris infection was compared in male and female immunocompetent BALB/c mice that had not previously been exposed to the parasite. No sex-related difference was observed in the time-course of the infection in these mice. Sexually mature mice of both sexes excreted substantial numbers of G. muris cysts (greater than 10(4)/2 hr) over a longer period than did sexually immature mice.

Aging↗
PubMed1987

Clearance of Giardia muris infection requires helper/inducer T lymphocytes.

The aim of this work was to identify the T cell subset (helper/inducer or cytotoxic/suppressor) that plays a major part in the clearance of Giardia muris infection. BALB/c mice were selectively depleted of helper/inducer (Th/i) or cytotoxic/suppressor (Tc/s) T cells, by treatment with rat IgG monoclonal antibodies directed against the Th/i cell surface antigen L3T4, or against the Tc/s cell surface antigen Ly-2, and were infected with G. muris cysts. While mice depleted of Tc/s cells cleared G. muris infection at a normal rate, mice depleted of Th/i cells did not clear the infection. Depletion of the relevant T cell subset was confirmed by flow cytometry and by immunohistochemistry. The data indicate that helper/inducer T cells, but not cytotoxic T cells, are of major importance for elimination of G. muris from the mouse intestine.

Animals↗
PubMed1987

T-lymphocyte subsets in nude mice with Giardia muris infection.

The aim of this study was to compare the relative percentages of T-lymphocyte subsets in athymic (nude) mice and immunocompetent BALB/c mice, with and without Giardia muris infection. Suspension of mononuclear leukocytes from blood, spleen, and Peyer's patches of uninfected mice were incubated with fluorescent monoclonal antibodies (MAbs) directed against mouse T-lymphocytes (anti-Thy-1.2 MAb), T helper/inducer lymphocytes (anti-L3T4 MAb), or T suppressor/cytotoxic lymphocytes (anti-Ly-2 MAb), and examined by fluorescence microscopy to quantify each of these cell populations. While the percentages of Thy-1.2+, L3T4+, and Ly-2+ lymphocytes are all reduced in nude mice, L3T4+ lymphocytes were found to be especially depleted, resulting in a reversed L3T4+/Ly-2+ ratio (less than 1) in all of the nude mouse tissues examined. BALB/c and nude mouse Peyer's patch T-cell subsets were quantified at various times during enteric Giardia muris infection. Unlike immunocompetent BALB/c mice, nude mice have an impaired ability to clear this infection. It was found that clearance of G. muris infection is associated with a Peyer's patch L3T4+/Ly-2+ ratio of greater than or equal to 2, and that this infection does not alter the percentages of Peyer's patch Thy-1.2+, L3T4+, or Ly-2+ lymphocytes in BALB/c mice or nude mice. The data obtained in the present work and in other studies suggest that the impaired capacity of nude mice to clear G. muris infection results from deficiency of L3T4+ lymphocytes.

Animals↗
PubMed1986

Response of Peyer's patch lymphocyte subsets to Giardia muris infection in BALB/c mice. I. T-cell subsets.

Initial cellular events in the intestinal immune response occur within Peyer's patches and are subject to complex regulation by T cells. The aim of this study was to analyze the response of murine Peyer's patch T, T-helper (Th), and T-suppressor (Ts) cells to Giardia muris infection. Immunocompetent BALB/c mice were infected with G. muris cysts and, at serial times during the infection, Peyer's patch leukocyte suspensions were incubated with fluorescent monoclonal antibodies that identified murine leukocytes, T, Th, or Ts lymphocytes. These suspensions were examined by flow cytometry to quantify each T-cell subset as a percentage of total leukocytes. Total Peyer's patch leukocytes more than doubled in number during the course of G. muris infection and returned to control levels as the infection was cleared. The percentages of Peyer's patch Th and Ts lymphocytes were 34.1 +/- 0.8% (mean +/- SEM) and 6.2 +/- 0.3%, respectively, in the absence of infection, and did not change significantly during Giardia infection. The Th/Ts ratio in Peyer's patches was 5.6 +/- 0.2 in uninfected BALB/c mice and also did not change significantly during clearance of G. muris. We conclude that Peyer's patch leukocytes double in number in response to G. muris infection in immunocompetent mice, G. muris infection does not lead to altered percentages of Peyer's patch T, Th, or Ts lymphocytes, and clearance of G. muris infection is associated with a Peyer's patch Th/Ts ratio of greater than 5.

Animals↗
PubMed1986

Response of Peyer's patch lymphocyte subsets to Giardia muris infection in BALB/c mice. II. B-cell subsets: enteric antigen exposure is associated with immunoglobulin isotype switching by Peyer's patch B cells.

The aim of this study was to quantify the response of Peyer's patch B cells, surface IgA-bearing (sIgA) B cells, and surface IgM-bearing (sIgM) B cells to Giardia muris infection. Following infection of a cohort of immunocompetent BALB/c mice with G. muris cysts, Peyer's patch cell suspensions were prepared at serial time points during the infection, incubated with fluorescein-conjugated monoclonal antibodies directed against murine leukocytes, B cells, sIgA B cells, sIgM B cells, or T cells, and analyzed by flow cytometry. Of total Peyer's patch leukocytes, the percentages of B cells, sIgA B cells, and sIgM B cells in uninfected BALB/c mice were 64.7 +/- 2.0% (mean +/- SEM), 30.3 +/- 1.5%, and 52.5 +/- 2.4%, respectively. The total number of Peyer's patch leukocytes increased significantly (1.8 X) during G. muris infection, and returned to control levels as the infection was cleared. The percentages of Peyer's patch T and total B cells did not change significantly during Giardia infection. However, sequential changes were observed in the percentages and numbers of sIgM and sIgA B cells during the infection. Peyer's patch sIgM B cells rapidly increased in percentage and number, reaching maximum levels 1 week after cyst inoculation. After remaining constant the first week, the number of Peyer's patch sIgA B cells increased during the second week of G. muris infection, reaching a maximum level 11-14 days after cyst inoculation. The data support the hypothesis that immunoglobulin isotype switching in Peyer's patches is induced by antigen exposure.

Animals↗
PubMed1986

Antibody response to Giardia muris trophozoites in mouse intestine.

The protozoan parasite Giardia muris colonizes the mouse small intestinal lumen. This parasite is cleared immunologically from the intestine of normal mice. In contrast, T-lymphocyte-deficient (nude) mice have an impaired immunological response to G. muris and become chronically infected. In the present study, trophozoites were harvested from the intestinal lumen of immunocompetent BALB/c mice and nude mice and examined for surface-bound mouse immunoglobulins by immunofluorescence microscopy. Immunoglobulin A (IgA) and IgG, but not IgM, were detected on trophozoites obtained from BALB/c mice, from day 10 of the infection onwards. Trophozoites from nude mice showed very little evidence of surface-bound mouse immunoglobulin at any time during the 5-week period immediately following infection of these animals with G. muris cysts. Intestinal G. muris infection was cleared by the BALB/c mice but not by the nude animals. The data suggest that parasite-specific IgA and IgG bind to G. muris trophozoites in the intestinal lumen of immunocompetent BALB/c mice. Intestinal antibodies that bind to trophozoite surfaces are likely to play an important part in the clearance of G. muris infection by immunocompetent mice. The inability of nude mice to clear this infection at a normal rate is likely to be due to impairment of Giardia-specific intestinal antibody production.

Animals↗
PubMed1986

Clearance of Giardia muris infection in mice deficient in natural killer cells.

Immunocompetent C57BL/6J mice and beige mice (which are deficient in natural killer cells) were infected with Giardia muris. Both types of mice cleared G. muris infection at similar rates. This observation suggests that clearance of G. muris parasites from the mouse intestine is not mediated by natural killer cells.

Animals↗
PubMed1985

Harvesting of leukocytes from intestinal lumen in murine giardiasis and preliminary characterization of these cells.

The aims of this study were to develop a method for harvesting leukocytes from the mouse small intestinal lumen and to identify leukocytes which enter the intestinal lumen of mice infected with Giardia muris. Giardia-infected and uninfected BALB/c mice were anesthetized, and the small intestine was removed. The intestinal lumen was then flushed with nutrient medium, and the luminal washings were found to contain substantial numbers of mononuclear leukocytes. The number of these cells harvested from the intestinal lumen of 24 mice infected with Giardia muris was 22 +/- 3 X 10(4) (mean value +/- standard error). Most of the cells were lymphocytes, although small numbers of macrophages were also obtained. By staining with monoclonal antibodies, approximately 50% of the intraluminal leukocytes were shown to be T lymphocytes. Similar numbers of mononuclear leukocytes were obtained from the intestinal lumen of Giardia-infected and uninfected mice. However, T lymphocytes and other mononuclear cells from infected animals were frequently observed in contact with Giardia trophozoites. This observation is consistent with the hypothesis that intraluminal leukocytes play a part in gastrointestinal immune defense.

Animals↗
PubMed1985

Comparison of leukocytes obtained from the intestinal lumen of Giardia-infected immunocompetent mice and nude mice.

Previous studies have suggested that Giardia infections are cleared immunologically, but have not defined the mechanism of clearance. The aim of the present work was to compare subpopulations of leukocytes harvested from the intestinal lumen of immunocompetent BALB/c mice, which clear Giardia muris infection rapidly, with those of immunodeficient nude mice, which become chronically infected with Giardia muris. Leukocytes were obtained from the intestinal lumen of Giardia-infected mice, and subpopulations of these cells were quantified after immunofluorescent staining with monoclonal antibodies. Identical numbers of leukocytes were harvested from the intestinal lumen of Giardia-infected immunocompetent mice and nude mice, but the number of these leukocytes bearing the T-lymphocyte antigen Thy-1.2 was smaller in nude mice than in immunocompetent mice. In contrast, no striking differences were observed between the numbers of luminal cytotoxic/suppressor T lymphocytes or macrophages in Giardia-infected nude versus immunocompetent mice. The findings suggest that clearance of Giardia muris infection is not mediated by cytotoxic T lymphocytes or macrophages. Subsequently, T lymphocytes and T-lymphocyte subsets were quantified in cell suspensions prepared from Peyer's patches of immunocompetent BALB/c mice and nude mice. It was found that nude mice have a profound deficiency of Peyer's patch helper/inducer T lymphocytes. This deficiency may account for the inability of nude mice to clear Giardia muris infection at a normal rate.

Animals↗
PubMed1982

Clinical experience with antilymphocyte serum.

Many different antilymphocytic antisera have been used clinically, and the properties of any particular type of ALS are not necessarily identical to those of any other type. Nevertheless, it is possible to draw certain general conclusions about the effects of ALS in human subjects. ALS administration has often been shown to reduce the number of circulating E-rosette-positive lymphocytes, although the precise mechanisms by which this reduction occurs are not known. Using a combined technique of E-rosette formation and immunofluorescence, heterologous immunoglobulin has been demonstrated on T and non-T lymphocytes from patients receiving non-selective ALS. Fifteen years' experience has failed to provide convincing support for the view that ALS (including immunoglobulin prepared from the whole antiserum) prolongs human renal allograft survival. It is not yet known whether ALS is a useful immunosuppressive agent in cardiac transplantation. One observation of possible clinical interest is that bone marrow regeneration has occurred in a number of patients with aplastic anemia who have been treated with ALS. No satisfactory method has been developed for monitoring the dose of ALS in human subjects. Appropriate studies may determine whether monoclonal antilymphocytic antibodies are clinically useful, for example in prolonging the survival of transplanted organs, in preventing or treating graft-versus-host disease, or in treating lymphoma, leukemia, or aplastic anemia.

Anemia, Aplastic↗