Martin F. Heyworth

Impact of aging on gastrointestinal mucosal immunity

There is considerable evidence that the mucosal or secretory immune response in the gastrointestinal tract is compromised by aging. The generation of a mucosal immune response is an extremely complex process that involves antigenic stimulation of a specific subpopulation of immunologically competent cells in the Peyers patches, differentiation and migration of these cells to the small intestinal lamina propria, initiation and regulation of local antibody production in the intestinal wall, and mucosal epithelial cell receptor-mediated transport of antibodies to the intestinal lumen. Available data suggest that gastrointestinal mucosal immunosenescence reflects deficits in: (1) the differentiation and/or migration (homing) of immunoglobulin A immunoblasts to the intestinal lamina propria, and (2) the initiation and/or regulation of local antibody production. The significant age-related increases in the incidence and severity of gastrointestinal infectious diseases, coupled with the potential for immunopharmacologic manipulation of the mucosal immune compartment, substantiate the merit of studies designed to resolve the etiology of mucosal immunodeficiency in the elderly.

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 Peyers patches of immunocompetent BALB/c mice and nude mice. It was found that nude mice have a profound deficiency of Peyers patch helper/inducer T lymphocytes. This deficiency may account for the inability of nude mice to clear Giardia muris infection at a normal rate.

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 withGiardia muris. Giardia-infected and uninfectedBALB/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 withGiardia muris was 22±3×104 (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 ofGiardia-infected and uninfected mice. However, T lymphocytes and other mononuclear cells from infected animals were frequently observed in contact withGiardia trophozoites. This observation is consistent with the hypothesis that intraluminal leukocytes play a part in gastrointestinal immune defense.

Lymphocyte Migration from Peyer’s Patches by Diapedesis through M Cells into the Intestinal Lumen

Lymphocytes and other leukocytes are found in the intestinal lumen in a variety of pathological conditions, and also in the absence of disease. Although Mitchison (1) found few lymphocytes in luminal washings from rabbit intestinal loops, Kotani et al. (2) found large numbers of lymphocytes in the lumen of rat intestinal loops, and calculated that a number of lymphocytes equal to the blood lymphocyte count entered the intestinal lumen five times per day. In jejunal biopsies from patients with giardiasis, and in mice infected with Giardia muris, we have observed intraluminal lymphocytes in close association with Giardia trophozoites (3,4). These lymphocytes are numerous over Peyer’s patch lymphoid follicles. Leukocytes have also been found over Peyer’s patches and appendix lymphoid follicles in the rabbit intestinal lumen (5). In mice Abe and Ito found twice as many intraepithelial lymphocytes over Peyer’s patch follicle domes as over villi (6). Over half of these dome intraepithelial lymphocytes lay above the level of epithelial cell nuclei suggesting migration into the lumen, whereas only 7% of intraepithelial lymphocytes over villi lay above epithelial nuclei. After hydrocortisone administration intraepithelial lymphocytes dropped to one third but the proportion of lymphocytes above the level of epithelial nuclei remained unchanged, consistent with continued movement toward the lumen (6).

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 Peyers 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 Peyers patches were examined by immunohistochemistry or flow cytometry. In anti-L3T4-treated mice, Peyers 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. Peyers 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 Peyers patches.

Immunology of the Gastrointestinal Tract and Liver

This book contains 11 chapters. Some of the chapter titles are: T cells and Other Non-B Lymphocytes; Mucosal Mast Cells and IgE; Genetic Aspects of Gastrointestinal Immunology; Immunological Functions of the Liver; Lymphocyte Migration and Mucosal Immunity; and Immunoglobulin Circulation and Secretion.