Bombesin affects mucosal immunity and gut-associated lymphoid tissue in intravenously fed mice.

BACKGROUND: Our prior studies show that intravenous (IV) total parenteral nutrition (TPN) produces atrophy of the small intestine-related gut-associated lymphoid tissue and significant decreases in intestinal IgA levels, the major system of mucosal immunity. Others have noted increased small intestinal permeability, bacterial adherence and translocation, and decreased IgA levels in TPN-fed animals. Bombesin, a neuropeptide, may play a regulatory role in mucosal immunity. It is not clear whether bombesin attenuates the TPN-associated gut-associated lymphoid tissue atrophy. OBJECTIVE: To examine the effect of bombesin on gut-associated lymphoid tissue integrity and function during IV TPN feeding. DESIGN: Randomized animal study. SETTING: A university laboratory. MATERIALS AND METHODS: Male ICR mice weighing 25 to 30 g were randomized to chow plus IV saline solution (n = 12), IV TPN (n = 12), or IV TPN plus bombesin (15 micrograms/kg, administered intramuscularly three times a day) (n = 12). Animals were killed after 5 days of receiving the experimental diet. Total small intestinal IgA level was quantified by enzyme-linked immunosorbent assay. Lymphocytes were isolated from Peyer's patches, intraepithelial spaces, and lamina propria and were stained with specific antibodies for B and T cells and for T-cell expression of CD4 and CD8 by flow cytometric analysis. Data were analyzed by analysis of variance. RESULTS: Bombesin prevented the IV TPN decreases in (1) total cell yield and B-cell yield from the Peyer's patches, intraepithelial spaces, and lamina propria; (2) T-cell yield in the intraepithelial spaces and lamina propria; and (3) small intestinal IgA levels. Bombesin also reversed IV TPN decreases in CD4+ and CD8+ T cells in the intraepithelial spaces and Peyer's patches and prevented the decrease in the CD4/CD8 ratio in the lamina propria. CONCLUSION: Bombesin prevents the TPN-associated atrophy and dysfunction of gut-associated lymphoid tissue, supporting the concept of close neuroimmunologic interaction.

Effects of parenteral and enteral nutrition on gut-associated lymphoid tissue.

Changes in mucosal defense have been implicated as important factors affecting infections complications in critically ill patients. To study the effects of nutrient administration on gut-associated lymphatic tissue (GALT), ICR mice were randomized to receive chow plus intravenous saline, intravenous feeding of a total parenteral nutrition (TPN) solution, or enteral feeding of the same TPN solution. In a second series of experiments, a more complex enteral diet (Nutren) was compared with chow feeding and enteral TPN. After 5 days of feeding with experimental diets, lymphocytes were harvested from the mesenteric lymph nodes (MLNs), Peyer's patches (PPs), lamina propria (LP) cells, and intraepithelial (IE) spaces of the small intestine to determine cell yields and phenotypes. Small intestinal washings, gallbladder contents, and sera were collected and analyzed for immunoglobulin A (IgA) levels. In both series of experiments, there were no significant changes within the MLNs. There were significant decreases in total cell yields from the PPs, IE spaces, and LP in animals fed with TPN solution, either enterally or parenterally, as compared with chow-fed mice. Total T cells were decreased in both TPN-fed groups in the PPs and LP, whereas total B cells were decreased in the PP, IE, and LP populations. Total cell numbers remained normal in the Nutrenfed group, except for a decrease in LP T cells. CD4+ LP cells decreased significantly with a reduction in the CD4/CD8 ratio in mice fed TPN solution either intravenously or enterally, whereas IgA recovery from small intestinal washings was significantly decreased in the same groups.(ABSTRACT TRUNCATED AT 250 WORDS)

cis-dominant regulation of CD4 and CD8 gene expression in rat/mouse T cell heterohybridomas.

To determine whether expression of CD4 and CD8 molecules on T cells is determined-solely by transacting regulators, we examined heterohybridomas derived from the fusion of a rat CD4+ T cell line and the CD4- CD8- mouse thymoma BW5147. The majority of hybrid offspring expressed rat CD4. However, a fraction of the cell lines obtained expressed not only rat CD4 but also various amounts of mouse CD4 and CD8 molecules from both species. Cloning of two of these heterogeneous lines revealed that expression of all four Ag varied not only between different clones but also within clonal populations. The expression of Ag not present on the parental cells suggested an alteration in the normally stable regulatory mechanisms present in those cells. Moreover, a lack of concordant expression between the rat and mouse loci was observed, indicating that active and silent homologous loci can exist together in single nuclei. Expression of CD4 and CD8 in these cells, therefore, cannot be solely mediated by trans-acting diffusible regulators but must also depend on cis-dominant effects on the loci themselves. The phenotypic heterogeneity of clonal populations was found to result from unpredictable shifts, both positive and negative, in the expression of CD4 and CD8 over time, indicating that the cis-dominant effects were only quasistable. Preliminary examinations of the density of 5-methylcytosine within the CD4 and CD8 loci in various phenotypic populations separated by FACS from within heterogeneous clones revealed a correlation between surface expression of the mouse CD8 protein and a lack of methylation around the mouse CD8 gene. In contrast, the CD4 gene remained extensively methylated regardless of its surface expression.

Roles of CD4+ and CD8+ T cells in murine contact sensitivity revealed by in vivo monoclonal antibody depletion.

mAb specific for murine CD4+ and CD8+ T cell subsets were utilized to determine the populations participating in delayed-in-time, cutaneous hypersensitivity responses in BALB/c mice. In vivo depletions of these T cell phenotypes revealed that delayed-type hypersensitivity to cellular and protein Ag were mediated by CD4+ effector cells, whereas CD8+ cells down-regulated such responses. Similar depletions in mice prior to sensitization with the hapten 1-fluoro-2,4-dinitrobenzene demonstrated a more complex pattern of cell participation in contact sensitivity (CS) responses. Depletion of CD4+ cells resulted in strikingly enhanced ear swelling, indicating not only an important effector role for CD8+ cells but also a down-regulatory role for some CD4+ cells; depletion of CD8+ cells revealed that some CD4+ cells also act as CS effectors. In vitro depletion of immune lymph node cells with the same mAb before adoptive transfer confirmed CS effector roles for both subsets, and also suggested that at least some CD4+ suppressors act on the efferent limb of the CS response, perhaps by down-regulating the activity of CD8+ effector cells. Partial in vivo depletion with small amounts anti-CD4 mAb and subsequent flow cytometric analysis of residual CD4+ cells was consistent with the hypothesis that CD4+ CS effector cells express a higher density of the CD4 antigen than do CD4+ suppressor cells, raising the possibility that these two functionally distinct CD4+ populations might be separable on the basis of their surface expression of CD4.

Phagocytic cell responses to in vivo and in vitro exposure to the Lyme disease spirochete.

An experimental skin lesion induced in rabbits by the bite of infected adult Ixodes dammini showed dense dermal interstitial inflammatory cell infiltrates composed of mononuclear cells (histiocytes and lymphocytes) and granulocytes. The prevalence of phagocytic cells in this experimental lesion motivated a study on the interactions of macrophages and neutrophils with Lyme disease spirochetes. Interactions as measured by uptake of radiolabeled spirochetes and by indirect immunofluorescence were enhanced by opsonization of spirochetes with immune serum and not significantly decreased by heat inactivation of the same. Phagocytosis was inhibited by treatment of cells with Cytochalasin B. Adherence of opsonized spirochetes to neutrophils was decreased by blocking Fc receptors with heat-aggregated IgG, suggesting an important role for this receptor.

The effect of age on the infection and immunoresponsiveness of mice to Babesia microti.

5he effect of age on the immunological responses to Babesia microti infection in the mouse was investigated. Aged mice experienced reduced and delayed peak parasitemias compared to younger animals; however, the old mice failed to clear the parasites from the blood and experienced fluctuating parasitemias until death. Babesiosis produced suppression of responses to nonspecific B and T cell mitogens concomitant with rising autoantibody plaque forming cells reactive with untreated and bromelain modified mouse erythrocytes. Similar observations of increased susceptibility to babesiosis with age and immunosuppression have been made in human babesiosis. Thus, the murine model for this hemoprotozoan infection is faithful to the human immunological responses.