Cellular immune responses in mice infected with the intestinal nematode Trichuris muris.

CBA and B10.BR mice show variation in immune response to the intestinal nematode Trichuris muris. CBA mice develop strong resistance, eliminating worms from the intestine; B10BR mice are permissive and develop chronic infections. It is already known that resistance and permissiveness reflect differential T helper responses. The data reported here show that resistant CBA mice express good antigen-specific lymphocyte proliferative responses to infection, whereas cells from B10.BR mice are relatively anergic, although still responsive to Concanavalin A (ConA). The possibility that the altered proliferative responsiveness seen in infected B10.BR mice reflected quantitative or qualitative changes in T helper cells was examined by comparing cytokine production and expression of cell surface markers (CD4, CD8, and CD28) in mesenteric lymph node cells and spleen cells from both strains and comparing these with the characteristics of cells from resistant CBA mice and from CBA mice that had been rendered permissive to infection by a combination of irradiation and corticosteroid treatment. As expected, cells from B10.BR mice produced high levels of interferon-gamma (IFN-gamma), whereas those from CBA mice released high levels of IL-5, whether stimulated with adult worm somatic antigens, excretory/secretory antigens, or ConA. Immunosuppressed CBA mice produced high levels of both IFN-gamma and IL-5 throughout the experiment. FACS analysis revealed a decrease of CD4+ and an initial increase in CD8+ cells in all infected mice. No major changes occurred in the relative proportion of CD28(+) cells. Further evaluation of the CD28 costimulatory molecule, measured as mean fluorescence intensity, displayed down-regulation in permissive and immunosuppressed mice. The data obtained suggest that lymphocyte unresponsiveness and permissiveness to T. muris infection may be associated with a down-regulation or an initially reduced expression of costimulatory CD28 molecules.     

Striking phenotypic and functional differences in lamina propria lymphocytes from the large and small intestine of mice

Although intraepithelial T lymphocytes of the large intestine (LI) are known to differ from those of the small intestine (SI) in phenotype and function, differences in LI and SI lamina propria (LP) lymphocyte populations have not been clearly established. In this work we found striking phenotypic differences between SI and LI LP lymphocyte populations from Balb/c mice analyzed by flow cytometry. In the LI most lymphocytes were B cells and the predominant T cells were TCR-alpha beta+, CD8+. In contrast, in the SI most T lymphocytes were CD4+ expressing TCR-alpha beta+, although a higher proportion expressed TCR-gamma delta+ than in the LI. In T cells the expression of adhesion molecules and cytokines was also different between SI and LI. The proportion of LP T cells expressing alpha4beta7 and L-selectin was higher in the LI than in the SI; whereas a greater proportion of cells expressing alpha(E)beta7 were detected in the SI than in LI. Higher proportions of T cells expressing L-selectin and alpha4beta1 were detected in the intraepithelial compartment of the LI than that of the SI, whereas the number of T cells expressing alpha(E)beta7 was much higher in the SI than in the LI. The proportion of T cells spontaneously producing IL-2, IFN gamma, and IL-4 at the intraepithelial and lamina propria, in the small and large intestine, was different indicating that distinctive functional features exist in the lymphocyte populations residing at the different intestinal compartments.     

The absence of resistance in congenitally athymic nude mice toward infection with the intestinal nematode, Trichuris muris: resistance restored by lymphoid cell transfer

Congenitally athymic (nude) mice maintained infections of Trichuris muris for at least 40 days post-infection, whereas phenotypically normal mice expelled the worms by 18 days post-infection. Complete worm expulsion occurred in nude mice which had been given spleen cells. On the other hand, a partial resistance to the infection was observed in nude mice which received thymus or mesenteric lymph node cells. No significant worm reduction was seen by injection of immune serum.     

Role of murine intestinal intraepithelial lymphocytes and lamina propria lymphocytes against primary and challenge infections with Cryptosporidium parvum

To investigate the role of intestinal lamina propria lymphocytes (LPL) and intraepithelial lymphocytes (IEL) in controlling Cryptosporidium parvum infection, changes in their phenotypes and functional properties were studied after induction of primary and challenge infections in immunocompetent mice. As shown by oocyst-shedding patterns, the challenge-infected group recovered more rapidly from infection than did the primary-infected group. In LPL, proportions of activated CD4+, CD25+, IgG1+, IgA+, and CD4+/IFN-gamma+ cells increased significantly in the primary-infected group compared with controls. In the challenge-infected group, proportions of these cells decreased. The antigen-specific IgA level was elevated significantly among LPL of both primary- and challenge-infected groups. Among IEL, proportions of activated CD8+, T cell receptor (TCR) gammadelta+, and CD8+/TCR gammadelta+ cells increased significantly in the challenge-infected group compared with controls and the primary-infected group; their cytotoxicity also was enhanced. However, the proportion of IEL expressing Th1 cytokines was lower than that among LPL in both infected groups. The results suggest that LPL play a more important role in protection against a primary infection with C. parvum, through the production of IFN-gamma and IgA, whereas IEL are more involved in protection against a challenge infection, through enhanced cytotoxicity.     

Impairment of primary expulsion of Trichuris muris in mice concurrently infected with Nematospiroides dubius

Primary immune expulsion of Trichuris muris was markedly delayed by concurrent infection with Nematospiroides dubius. Maximum delay of expulsion was dependent on size and timing of N. dubius infection relative to T. muris infection. In NIH mice infection with 400 N. dubius larvae immediately before or after T. muris infection was found to be most effective in suppressing expulsion. Infection on day 8 of T. muris infection, when mice are sensitized to T. muris, also impaired expulsion. From this evidence it is suggested that the larvae of N. dubius are immunosuppressive and that the efferent role of the immune response to T. muris is inhibited. The results are discussed in terms of non-specific immunosuppression and their relevance to the tropical disease situation is emphasized.     

Changes in the bacterial cecal flora of mice infected with Trichuris muris (Schrank, 1788).

Cecal microorganisms of mice were categorized and enumerated weekly during the developmental cycle of infection with the whipworm, Trichuris muris. The cecal bacterial population consisted of Escherichia coli, Proteus spp, Acinetobacter lwoffi (Mima polymorpha), aerobic lactobacilli, staphylococci, enterococci, and anaerobes (bacteroides, streptococci, and lactobacilli) in control and T. muris-infected mice. The aerobic lactobacilli and the anaerobes constituted the greatest number of organisms in both groups. In week three there was a decrease in the number of these organisms, and in week four fewer of these and of all other organisms in the worm-infected mice when compared to controls. The most significantly reduced bacterial counts were observed during the period of T. muris self-cure.     

The effect of Plasmodium berghei and Trypanosoma brucei infections on the immune expulsion of the nematode Trichuris muris from mice

The effects of concurrent P. berghei or T. brucei infections on the immune expulsion of primary and challenge infections of T. muris from CFLP strain mice have been examined. CFLP mice usually expel the nematode 18–21 days after a primary infection and within 4–6 days after a challenge infection. Both acute malaria and trypanosome infections initiated at the same time as the T. muris infection suppressed worm expulsion; when the protozoal infections were started 7 days after the T. muris infection worm expulsion was suppressed in a proportion of the mice. Acute trypanosome and malaria infections delayed the expulsion of a challenge infection from immune mice, but in the case of P. berghei the delay was short-lived.     

Mast cells from the human intestinal lamina propria. Isolation, histochemical subtypes, and functional characterization

With the use of a collagenase dispersion technique, cells were isolated from the lamina propria of the human small and large intestine. The cell suspensions contained 8% mast cells, which on average contained 1 to 2 pg of histamine/cell. With the use of histochemical procedures based upon fixative sensitivity and dye binding, which identify functionally distinct mast cell subtypes in the rat, dispersed human intestinal mast cells contained approximately equal proportions of two histochemical subtypes analogous to those in the rat. Whether these are functionally distinct as in the rat remains to be determined. The histochemically mixed mast cell populations from the human intestinal mucosa secreted histamine in a dose- and energy-dependent manner in response to anti-IgE and A23187, but not 48/80. Theophylline, doxantrazole, quercetin, and salbutamol all significantly inhibited anti-IgE-induced histamine secretion by human intestinal mast cells, but cromolyn sodium and the experimental antisecretory drugs, nedocromil sodium and FPL 52694, did not inhibit histamine secretion by the mast cell mixture to a statistically significant extent. Cromolyn sodium inhibited histamine secretion by 15 to 30%, and whether this reflected inhibition of one of the two histochemical mast cell subtypes to a greater extent than the other or all the cells to a minimal degree remains to be established. Control investigations of the intestinal cell isolation procedure indicated that these qualities did not reflect effects of the cell dispersal procedure. Further characterization and analysis of intestinal mast cells is essential to determine if functionally distinct mast cell subtypes exist in human tissues.     

Impaired accumulation of antigen-specific CD8 lymphocytes in chemokine CCL25-deficient intestinal epithelium and lamina propria

CCL25 and CCR9 constitute a chemokine/receptor pair involved in T cell development and in gut-associated immune responses. In this study, we generated CCL25(-/-) mice to answer questions that could not be addressed with existing CCR9(-/-) mice. Similar phenotypes were observed for both CCL25(-/-) and CCR9(-/-) mice, consistent with the notion that CCL25 and CCR9 interact with each other exclusively. We assessed the requirement for CCL25 in generating CCR9(high) CD8 intestinal memory-phenotype T cells and the subsequent accumulation of these cells within effector sites. TCR-transgenic naive CD8 T cells were transferred into wild-type or CCL25-deficient hosts. Oral sensitization with Ag allowed these naive donor cells to efficiently differentiate into CCR9(high) memory-phenotype cells within the mesenteric lymph nodes of wild-type hosts. This differentiation event occurred with equal efficiency in the MLN of CCL25-deficient hosts, demonstrating that CCL25 is not required to induce the CCR9(high) memory phenotype in vivo. However, we found that CCL25 deficiency severely impaired the Ag-dependent accumulation of donor-derived CD8 T cells within both lamina propria and epithelium of the small intestine. Thus, although CCL25 is not necessary for generating memory-phenotype CD8 T cells with "gut-homing" properties, this chemokine is indispensable for their trafficking to the small intestine.     

Cells containing IgE in the intestinal mucosa of mice infected with the nematode parasite Trichinella spiralis are predominantly of a mast cell lineage

To determine whether IgE+ cells in the intestinal mucosa of nematode-infected mice were of a mast cell or a lymphocyte lineage, the intestinal mucosae of mast cell-deficient w/wv mice were examined for IgE+ cells after inoculation with Trichinella spiralis muscle-stage larvae. Immunofluorescence staining techniques were used to detect IgE associated with cells in the intestinal mucosa. Comparisons were made among four strains of mice, w/wv (mast cell-deficient), +/+ (normal congenic littermates of w/wv), BALB/c, and SJL, that were either uninfected controls or inoculated with T. spiralis. Tissue sections from the small intestine of T. spiralis-infected BALB/c, SJL, and +/+ mice were fixed in ethanol and were stained with an affinity-purified F(ab')2 rabbit anti-mouse IgE followed by FITC goat anti-rabbit IgG. Large numbers of cells in the intestinal mucosa exhibited bright fluorescence. When other sections of intestines from these mice were processed in Carnoy's fixative and were stained with alcian blue at low pH (a metachromatic stain for mast cells) or alcian blue followed by immunofluorescence staining for IgE, large numbers of mast cells were observed in the intestinal mucosa, and 70 to 90% stained positively for IgE. There was a considerable number of cells in the intestinal mucosa which were IgE+ but which did not stain with alcian blue. Few alcian blue-positive cells and no IgE+ staining cells were present in the intestinal mucosa of control, uninfected +/+, BALB/c, and SJL mice. To determine whether these IgE+ alcian blue-negative cells were of a lymphocyte or a mast cell lineage, the mast cell-deficient w/wv mouse strain was examined after infection with T. spiralis. In contrast to BALB/c, SJL, or +/+ mice, few cells in the intestinal mucosa of T. spiralis-infected w/wv mice stained with alcian blue or were positive for IgE. However, when the IgE response in the MLN of the w/wv mice was compared to the IgE response of BALB/c, SJL, and +/+ mice, numerous IgE+ cells, but no alcian blue-positive cells, were observed in the parenchyma of the MLN from all four strains of T. spiralis-infected mice. In addition, flow microfluorometric analysis of MLN cells stained for surface IgE in suspension showed a comparable proportion of IgE-bearing cells, which were mostly B lymphocytes, among all four strains of T. spiralis-infected mice.(ABSTRACT TRUNCATED AT 400 WORDS)     

Proliferative kinetics of large and small intraepithelial lymphocytes in the small intestine of the mouse.

The proliferative kinetics of the intraepithelial lymphocytes (IL) of the mouse intestine have been evaluated. By inducing mitotic arrest it was found that large IL - constituting about 50% of the IL - showed a mitotic rate of 2.3. Autoradiographic results obtained after two different schedules of 3H-thymidine injections showed that 30% of the large IL were in DNA synthesis, and that the large IL were renewed at a rate comparable to that of blast cells from Peyer's patches, mesenteric lymph nodes and thoracic duct lymph. The small IL were renewed very rapidly compared to small lymphocytes of peripheral lymphoid tissues, although small lymphocytes with lifespans of several weeks were also present in the epithelial sheet. By the use of intestinal perfusion, in vivo, it was estimated that the loss of lymphocytes from intestinal villi into the lumen of the gut was negligible, and it is concluded that the most probable kinetic model for the majority of IL is: B and T lymphoblasts invade the epithelium and undergo mitosis. B lymphoblasts give rise predominantly to plasma cells, and T lymphoblasts give rise to small lymphocytes - probably long-lived - which reenter the circulation.     

Adaptive immune responses are dispensable for isolated lymphoid follicle formation: antigen-naive, lymphotoxin-sufficient B lymphocytes drive the formation of mature isolated lymphoid follicles

Isolated lymphoid follicles (ILFs) are recently appreciated members of the mucosal immune system. The architecture, composition, and inducible nature of these structures indicates that these structures are tertiary lymphoid structures. The process leading to the formation of tertiary lymphoid structures, lymphoid neogenesis, has been observed in a number of inflammatory and autoimmune conditions. Given this association, there is considerable interest in identifying the factors promoting lymphoid neogenesis, and understanding the steps in this process. Using murine ILF formation as a model, we have examined the roles of different cellular sources of lymphotoxin (LT) and the adaptive immune response in lymphoid neogenesis. In this study, we report that, although other cellular sources of LT may supplant B lymphocytes in the formation of immature ILFs (loosely organized clusters of B lymphocytes), LT-sufficient B lymphocytes are required for the progression of immature ILFs to mature ILFs (organized lymphoid aggregates with a follicle-associated epithelium). ILF formation occurs in the absence of T lymphocytes and Ag-specific B lymphocyte responses, and ILF B lymphocytes express elevated levels of LT in the absence of antigenic stimulation. Consistent with a role for chemokines inducing LT expression in Ag-naive B lymphocytes, and a chemokine-driven positive-feedback loop driving mature ILF formation, mature ILFs express elevated levels of B lymphocyte chemoattractant in the absence of Ag-specific B lymphocyte stimulation. These observations indicate that ILFs contain Ag-naive lymphocytes, and suggest that events occurring within ILFs shape subsequent immune responses mediated by these lymphocytes.     

Role of intraepithelial lymphocytes in mucosal immune responses of mice experimentally infected with Cryptosporidium parvum

In order to investigate the role of intestinal intraepithelial lymphocytes (IELs) in host defense against Cryptosporidium parvum infection, conventionally bred immunocompetent (ImCT) ICR mice and immunosuppressed (ImSP) littermates were infected orally with 10(6) C. parvum oocysts. Then fecal oocyst excretion, the number and location of IELs, and their T lymphocyte subsets were observed on days 4, 7, 10, 13, 16, and 20 postinfection (PI). Uninfected ImCT and ImSP mice were used as controls. The starting point of oocyst excretion was day 4 PI in both ImCT- and ImSP-infected mice. The highest oocyst excretion occurred on day 7 PI in both groups, though the number of oocysts excreted was 3 times greater in ImSP than in ImCT mice. In ImCT mice, IELs greatly increased in number on days 16 and 20 PI (P < 0.05), but the increase was minimal in ImSP mice. IELs changed their location from the basal area to intermediate and apical areas of villous epithelial cells during the early stage of infection. In ImCT-infected mice, IEL phenotypes also changed; whereas CD4+ cells increased temporarily on day 7 PI (P < 0.05), CD8+ cells increased significantly on days 16 and 20 PI (P < 0.05). The results strongly suggest that IELs play a significant role in host defense against C. parvum infection, with helper T cells initiating control of the infection and cytotoxic T cells eliminating the parasites.     

Diurnal changes in intraepithelial lymphocytes (IELs) in the small intestine of mice.

Diurnal changes in small intestinal intraepithelial lymphocytes (IELs) were examined in 8- to 10-week-old BALB/cA male mice. The ratio of T cell subsets expressing CD8 alpha alpha homodimer/CD8 alpha beta heterodimer was found to be higher in the dark period than that in the light period. Increased expression of Thy-1.2 on gamma delta T cells was also observed in the light period. No significant changes were found in other subsets. This is the first report to document diurnal changes in the small intestinal IELs in mice.     

Identification of intestinal M cells in isolated lymphoid follicles and Peyer’s patches of the Angora rabbit

The presence, distribution, and localization of M cells in isolated lymphoid follicles (ILF) and in follicle-associated epithelia (FAE) covering Peyer’s patches (PP) in Angora rabbits were investigated by immunohistochemistry and electron microscopy. Although PP could macroscopically be identified along the length of the mucosal and serosal surfaces of jejunum and ileum, the presence of ILF could only be located microscopically. Typical M cells in FAE were detected within the periphery of the dome regions of the PP, and immature columnar M cells in the FAE resided in the vicinity of the crypts. M cells in the FAE of both ILF and PP showed vimentin-positive reaction. M cells in the FAE of ILF were morphologically similar to the immature M cells found in the FAE of PP. Typical mature M cells were also observed in the FAE of a few ILF. In contrast to FAE of PP, numerous goblet cells were observed in the FAE of many ILF. Moreover, among intestinal villi, we noticed villi-like solitary lymphoid structures that showed abundant lymphocytes in their lamina propria and that were surrounded with vimentin-positive cells and goblet cells. Thus, the occurrence of copious immature M cells and goblet cells, in addition to the detection of villi-like solitary lymphoid structures full of lymphocytes in the FAE of many ILF, indicate that ILF do not complete their immunological maturation in contrast to PP. Various antigenic stimulations conceivably induce the formation and maturation of ILF along the length of the small intestine. The morphological resemblance between ILF M cells and PP M cells suggests that these two types of cells perform similar or the same immunological functions.     

Prenatal blockage of lymphotoxin beta receptor and TNF receptor p55 signaling cascade resulted in the acceleration of tissue genesis for isolated lymphoid follicles in the large intestine

Signaling by lymphotoxin (LT) and TNF is essential for the organogenesis of secondary lymphoid tissues in systemic and mucosal compartments. In this study, we demonstrated that the progeny of mice treated with fusion protein of LTbetaR and IgGFc (LTbetaR-Ig) or LTbetaR-Ig plus TNFR55-Ig (double Ig) showed significantly increased numbers of isolated lymphoid follicles (ILF) in the large intestine. Interestingly, double Ig treatment accelerated the maturation of large intestinal ILF. Three-week-old progeny of double Ig-treated mice showed increased numbers of ILF in the large intestine, but not in the small intestine. Furthermore, alteration of intestinal microflora by feeding of antibiotic water did not affect the increased numbers of ILF in the large intestine of double Ig-treated mice. Most interestingly, mice that developed numerous ILF also had increased levels of activation-induced cytidine deaminase expression and numbers of IgA-expressing cells in the lamina propria of the large intestine. Taken together, these results suggest that ILF formation in the large intestine is accelerated by blockage of LTbetaR and TNFR55 signals in utero, and ILF, like colonic patches, might play a role in the induction of IgA response in the large intestine.     

Infection of immunosuppressed mice with the abomasal nematode parasite of ruminants, Haemonchus contortus

Infective larvae of Haemonchus contortus established in mice either immunosuppressed with the corticosteroid, dexamethasone, or the cytotoxic drug, cyclophosphamide, or treated with the histamine H2 inhibitor, cimetidine. Infections persisted for as long as the immunosuppressive treatment (7 days) and growth of larvae was similar to that seen in sheep. Virtually no larvae survived in untreated mice. Accordingly, it would appear that adaptive immunity is an important barrier against primary infection by H. contortus in mice and is a determinant of host-range for this parasite. Antibody raised in either sheep or mice against soluble extracts of adult H. contortus precipitated with different but overlapping sets of worm antigens. This suggests that the unique antigens recognized by the mouse compared with the sheep are crucial for the rapid protective responses which prevent primary infection.