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1.
Gut ; 54(5): 623-9, 2005 May.
Article in English | MEDLINE | ID: mdl-15831905

ABSTRACT

BACKGROUND AND AIMS: The mechanisms by which commensal bacteria provoke intestinal inflammation in animal models of inflammatory bowel disease (IBD) remain incompletely defined, leading to increasing interest in the innate immune response of the colonic mucosa to bacterial colonisation. METHODS: Using gene expression profiling of colonic RNA from C.B17.SCID germ free mice and those colonised with altered Schaedler's flora, we investigated the innate immune response to bacterial colonisation in vivo. The two most consistently induced gene groups were RegIIIbeta and gamma as well as interferon gamma (IFN-gamma) response genes. RESULTS: Using quantitative reverse transcription-polymerase chain reaction, we showed that RegIIIbeta, RegIIIgamma, and IFN-gamma were constitutively expressed in the colon of conventionally housed SCID mice compared with either germ free SCID or conventionally housed BALB/c mice. Induction of these genes was reproduced by chronic monoassociation of germ free SCID mice with either of two separate gut commensal bacterial species-segmented filamentous bacteria and Schaedler's Escherichia coli. The cellular source for IFN-gamma on monoassociation of SCID mice with Schaedler's E coli was localised to a subset of intraepithelial natural killer (IENK) cells that express asialo-GM1. In vivo IFN-gamma immunoneutralisation studies failed to demonstrate any alteration in RegIIIbeta or gamma expression. CONCLUSIONS: Thus bacterial colonisation of the colon independently activates two distinct innate immune cell types at the mucosal interface with the colonic lumen, intestinal epithelial cells, and IENK cells, a response that may be regulated by the adaptive immune system. These innate immune responses may play a role in the pathogenesis of colitis in SCID adoptive transfer models in mice and possibly in patients with IBD.


Subject(s)
Inflammatory Bowel Diseases/microbiology , Interferon-gamma/biosynthesis , Proteins/metabolism , Animals , Colon/immunology , Colon/microbiology , Disease Models, Animal , Escherichia coli/immunology , Gene Expression Profiling/methods , Gene Expression Regulation/immunology , Germ-Free Life , Immunity, Mucosal , Inflammatory Bowel Diseases/immunology , Inflammatory Bowel Diseases/metabolism , Interferon-gamma/genetics , Intestinal Mucosa/immunology , Intestinal Mucosa/metabolism , Intestinal Mucosa/microbiology , Killer Cells, Natural/immunology , Mice , Mice, Inbred BALB C , Mice, SCID , Oligonucleotide Array Sequence Analysis/methods , Pancreatitis-Associated Proteins , Proteins/genetics , Reverse Transcriptase Polymerase Chain Reaction/methods
2.
Vaccine ; 22(7): 805-11, 2004 Feb 17.
Article in English | MEDLINE | ID: mdl-15040931

ABSTRACT

Although mechanisms operative in the induction and maintenance of specific, adaptive immunity, including 'cognate' B/T interactions, have been extensively studied and defined, we still know little about the mechanisms operative in developing and maintaining B- and T-cell dependent 'natural' immunity. Particularly, we are still rather ignorant concerning gut microbial/gut or systemic APC, T cell and B cell interactions that lead to lymphoid cell mediated 'natural' immunity: specific or broadly reactive, activation via TCR and BCR and/or via other receptors such as the TLR series, and whether T/B interactions are operative at this level? Here we will address: (1) the general role of gut microbes in the development and maintenance of the intestinal, humoral immune system; (2) the general role of gut microbes in the development of B1 cell mediated, 'natural' gut IgA and the dependence of these B1 cells on bystander T cell help; (3) the relative contributions of B1 versus B2 cells to gut 'natural' and specific IgA responses; (4) the role for particular 'normal' gut microbes in the initiation of inflammatory bowel diseases (IBD) in mice with a dysregulated immune system; and (5) the possible roles of gut microbes in facilitating oral tolerance, a mechanism likely operative in forestalling or ameliorating IBD. A central theme of this paper is to attempt to define the specificities of activated, functional CD4+ T cells in the gut for Ags of particular, usually benign gut microbes. We will also consider the still-unresolved issue of whether the contributions of B1-derived IgA in the gut to the 'natural' Ab pool are Ag-selected and driven to proliferation/differentiation or whether the main stimuli are not via BCRs but rather other receptors (TLRs, etc.). The main experimental approach has been to use antigen-free, germ-free, or gnotobiotic (mono- or oligo-associated with precisely known bacterial species) mice.


Subject(s)
B-Lymphocytes/immunology , Digestive System/microbiology , T-Lymphocytes/immunology , Animals , Antibody Formation/physiology , Immune Tolerance , Immunoglobulin A/biosynthesis , Immunoglobulin A/immunology , Mice
3.
J Immunol ; 167(7): 3677-81, 2001 Oct 01.
Article in English | MEDLINE | ID: mdl-11564781

ABSTRACT

In this study, we investigated the effect of morphine on the mucosal immune system using fragment cultures of ileal segments, Peyer's patches (PPs), and mesenteric lymph nodes. Mice were implanted s.c. with a morphine slow release pellet. Control groups received a naltrexone slow release pellet, a placebo pellet, or both a morphine and a naltrexone pellet. After 48 h, mice were orally immunized with cholera toxin (CT) and were boosted orally 1 wk later. Animals were sacrificed 1 wk after the booster immunization, and PPs, mesenteric lymph nodes, and ileal segments were cultured in 24-well plates for 12 days. Morphine resulted in a highly significant inhibition of CT-specific IgA and IgG production in fragment culture supernatants of all three tissues compared with placebo. Naltrexone blocked the reduction in Ab levels induced by morphine, indicating that the effect is opioid receptor mediated. Morphine did not significantly alter total IgA levels in any of the tissue culture supernatants. Morphine also inhibited CT-specific IgA and IgG levels in serum. By flow cytometry, morphine did not alter the lymphoid cell composition in PPs compared with placebo. The effect of morphine on TGF-beta, IL-5, and IL-6 mRNA expression in PPs and ileal segments was determined following oral immunization with CT. Morphine significantly decreased TGF-beta mRNA compared with that in the placebo group, and naltrexone blocked this effect. These results indicate that morphine inhibits Ag-specific IgA responses in gut-associated lymphoid tissue at least partially through the inhibition of TGF-beta, a putative IgA switch factor, in the gastrointestinal tract.


Subject(s)
Cholera Toxin/pharmacology , Immunity, Mucosal , Immunoglobulin A/biosynthesis , Intestinal Mucosa/immunology , Morphine/pharmacology , Transforming Growth Factor beta/biosynthesis , Administration, Oral , Animals , Cholera Toxin/administration & dosage , Drug Antagonism , Ileum/drug effects , Ileum/immunology , Immunoglobulin G/biosynthesis , Lymph Nodes/drug effects , Lymph Nodes/immunology , Mesentery , Mice , Mice, Inbred C3H , Organ Culture Techniques , Peyer's Patches/drug effects , Peyer's Patches/immunology , RNA, Messenger/biosynthesis , Transforming Growth Factor beta/genetics
4.
J Virol ; 75(12): 5482-90, 2001 Jun.
Article in English | MEDLINE | ID: mdl-11356955

ABSTRACT

Studies utilizing various immunodeficient mouse models of rotavirus (RV) infection demonstrated significant roles of RV-specific secretory immunoglobulin A (IgA), CD4+ T cells, and CD8+ T cells in the clearance of RV and protection from secondary infection. Secretion of small but detectable amounts of IgA in RV-infected alphabeta T-cell receptor knockout mice (11) and distinctive anatomical localization and physiology of B1 cells suggested that B1 cells might be capable of producing RV-specific intestinal IgA in a T-cell-independent fashion and, therefore, be responsible for ablation of RV shedding. We investigated the role of B1 cells in the resolution of primary RV infection using a SCID mouse model. We found that the adoptive transfer of unseparated peritoneal exudate cells ablates RV shedding and leads to the production of high levels of RV-specific intestinal IgA. In contrast, purified B1 cells do not ablate RV shedding and do not induce a T-cell-independent or T-cell-dependent, RV-specific IgA response but do secrete large amounts of polyclonal (total) intestinal IgA. Cotransfer of mixtures of purified B1 cells and B1-cell-depleted peritoneal exudate cells differing in IgA allotypic markers also demonstrated that B2 cells (B1-cell-depleted peritoneal exudate cells) and not B1 cells produced RV-specific IgA. To our knowledge, this is the first observation that B1 cells are unable to cooperate with CD4+ T cells and produce virus-specific intestinal IgA antibody. We also observed that transferred CD4+ T cells alone are capable of resolving RV shedding, although no IgA is secreted. These data suggest that RV-specific IgA may not be obligatory for RV clearance but may protect from reinfection and that effector CD4+ T cells alone can mediate the resolution of primary RV infection. Reconstitution of RV-infected SCID mice with B1 cells results in the outgrowth of contaminating, donor CD4+ T cells that are unable to clear RV, possibly because their oligoclonal specificities may be ineffective against RV antigens.


Subject(s)
B-Lymphocyte Subsets/immunology , CD4-Positive T-Lymphocytes/immunology , Rotavirus Infections/immunology , Rotavirus/immunology , Adoptive Transfer , Animals , Antibodies, Viral/blood , Disease Models, Animal , Flow Cytometry , Humans , Immunoglobulin A, Secretory/analysis , Intestines/immunology , Lymphoid Tissue/immunology , Mice , Mice, Inbred BALB C , Mice, SCID , Peritoneum/cytology , Peritoneum/immunology , Rotavirus/isolation & purification , Rotavirus Infections/virology
5.
Infect Immun ; 69(6): 3542-9, 2001 Jun.
Article in English | MEDLINE | ID: mdl-11349011

ABSTRACT

We used Listeria monocytogenes, a gram-positive, facultative intracellular bacterium, to study the gut mucosal immune responses following oral infection. We employed a germfree (GF) mouse model to try to accentuate the development of a humoral mucosal immune response in the gut, and we used oral colonization with one of the mutants, actA-negative (DeltaactA) L. monocytogenes, to restrict infection largely to the gut. The DeltaactA mutant was able to colonize the intestinal mucosa of formerly GF mice for long periods of time without causing disease while eliciting secretory immunoglobulin A (IgA) responses, as evidenced by gut tissue fragment culture assays. Flow cytometric analyses and immunohistochemical methods showed the development of only minimal germinal center reactions (GCR) in Peyer's patches and more robust GCR in mesenteric lymph nodes. Pronounced increases in total (natural) IgA production occurred in gut tissues by day 7 and were maintained for up to 90 days. Levels of specific IgA were modest in gut tissues on day 14, increased until day 76, and stabilized at day 90. We also observed a significant rise in serum IgA and IgG1 levels following oral infection by listeriae. Upon colonization, the organisms mainly infected the intestines and intestinal lumen, and we only sporadically observed few colony-forming bacteria in the liver and spleen. We observed a marked rise in IgA-secreting cells, including listeria-specific IgA antibody-secreting cells, in the lamina propria of the small intestine by enzyme-linked immunospot assays. To ascertain whether some of the IgA was specific for listeriae, we performed Western blot analysis to test the reactivity of IgA from fragment cultures to antigens in sonicates of L. monocytogenes. We detected IgA binding to antigenic proteins with molecular masses of 96, 60, 40, and 14 kDa in the Listeria sonicates.


Subject(s)
Bacterial Proteins/genetics , Immunity, Mucosal , Intestinal Mucosa/microbiology , Listeria monocytogenes/immunology , Listeriosis/immunology , Membrane Proteins/genetics , Mutation , Animals , Antibodies, Bacterial/blood , Blotting, Western , Disease Models, Animal , Flow Cytometry , Germ-Free Life , Immunization , Immunoglobulin A, Secretory/analysis , Immunohistochemistry , Intestinal Mucosa/immunology , Listeria monocytogenes/genetics , Listeria monocytogenes/growth & development , Listeriosis/microbiology , Mice , Mice, Inbred BALB C , Mice, SCID
6.
Infect Immun ; 69(6): 3611-7, 2001 Jun.
Article in English | MEDLINE | ID: mdl-11349021

ABSTRACT

As a member of the indigenous gut mucosal microbiota, segmented filamentous bacteria (SFB) colonize the guts of a variety of vertebrates and invertebrates. They are potent microbial stimuli of the gut mucosal immune system. In the small intestines of mice and rats, it has been observed that SFB are absent during the suckling period and appear in high numbers shortly after weaning, then quickly retreat to the cecum and large intestine. In this study, we explored whether this microecological phenomenon resulted from the interaction between SFB and the passively acquired maternal mucosal immunity and/or the actively acquired mucosal immunity. We set up a mouse model by reciprocal crossings and backcrossings of SFB-monoassociated, formerly germ-free, immunocompetent (+/+) BALB/c mice and immunodeficient (scid/scid) mice to produce pups which are either immunocompetent (scid/+) or immunodeficient (scid/scid) and are born either to immunocompetent (scid/+) mothers or to immunodeficient (scid/scid) mothers. We monitored the number of SFB on the mucosa of the small intestine in the four different groups of mice after birth, as well as the level of passively acquired antibodies, the active gut mucosal immune responses, and immunoglobulin A (IgA) coating of SFB in the gut. The results showed that, irrespective of whether the pups were scid/scid or scid/+, SFB could be found earlier on the mucosa of the small intestine in pups born to scid/scid mothers, appearing from day 13 and rapidly reaching a climax around weaning time on day 28, compared to the significantly delayed colonization in the pups of scid/+ mothers, starting from day 16 and peaking around days 28 to 32. After the climax, SFB quickly declined to very low levels in the small intestines of scid/+ pups of either scid/scid mothers or scid/+ mothers, whereas they remained at high levels in scid/scid pups at least until day 70, the last observation time in this study. The dynamic changes in SFB colonization of the small intestines of the different groups of pups may be related to the dynamic changes in the levels of SFB coated with secretory IgA (sIgA), which resulted from the significantly different levels of sIgA obtained from the mothers' milk during the suckling period and, later, of self-produced sIgA in the small intestine. Nevertheless, it is evident that the timing, localization, and persistence of colonization of the neonatal gut by SFB depends on the immune status of both mothers and pups.


Subject(s)
Bacteria, Anaerobic/growth & development , Gram-Positive Endospore-Forming Bacteria/growth & development , Immunoglobulin A, Secretory/immunology , Stomach/microbiology , Animals , Antibody Specificity , Bacteria, Anaerobic/immunology , Gram-Positive Endospore-Forming Bacteria/immunology , Immunity, Maternally-Acquired , Immunity, Mucosal , Immunoglobulin A, Secretory/metabolism , Intestinal Mucosa/microbiology , Mice , Mice, SCID , Stomach/immunology
9.
J Virol ; 73(9): 7633-40, 1999 Sep.
Article in English | MEDLINE | ID: mdl-10438854

ABSTRACT

Respiratory virus infections are a serious health challenge. A number of models that examine the nature of the respiratory immune response to particular pathogens exist. However, many pathogens that stimulate specific immunity in the lung are frequently not effective immunogens at other mucosal sites. A pathogen that is an effective respiratory as well as gastrointestinal immunogen would allow studies of the interaction between the mucosal sites. Reovirus (respiratory enteric orphan virus) serotype 1 is known to be an effective gut mucosal immunogen and provides a potential model for the relationship between the respiratory and the gut mucosal immune systems. In this study, we demonstrate that intratracheal immunization with reovirus 1/Lang (1/L) in C3H mice resulted in high titers of virus in the respiratory tract-associated lymphoid tissue (RALT). High levels of reovirus-specific immunoglobulin A were determined in the RALT fragment cultures. The major responding components of the bronchus-associated lymphoid tissue were the CD8(+) T lymphocytes. Cells from draining lymph nodes also exhibited lysis of reovirus-infected target cells after an in vitro culture. The present study also describes the distribution of transiently present CD4(+)/CD8(+) double-positive (DP) T cells in the mediastinal and tracheobronchial lymph nodes of RALT. CD4(+)/CD8(+) DP lymphocytes were able to proliferate in response to stimulation with viral antigen in culture. Furthermore, these cells exhibited lysis of reovirus-infected target cells after in vitro culture. These results establish reovirus 1/L as a viable model for future investigation of the mucosal immune response in the RALT and its relationship to the common mucosal immune system.


Subject(s)
Antibodies, Viral/immunology , CD4-Positive T-Lymphocytes/immunology , CD8-Positive T-Lymphocytes/immunology , Orthoreovirus/immunology , Animals , Cell Division , Cell Line , Cells, Cultured , Cytotoxicity, Immunologic , Digestive System/virology , Drug Administration Routes , Humans , Immunoglobulin A/immunology , Lymphoid Tissue/virology , Male , Mice , Mice, Inbred C3H , Mucous Membrane/immunology , Orthoreovirus/growth & development , Phenotype , Respiratory System/virology , Vaccination
10.
Am J Clin Nutr ; 69(5): 1046S-1051S, 1999 05.
Article in English | MEDLINE | ID: mdl-10232647

ABSTRACT

The normal colonization of the mammalian intestine with commensal microbes is hypothesized to drive the development of the humoral and cellular mucosal immune systems during neonatal life and to maintain the physiologically normal steady state of inflammation in the gut throughout life. Neonatal conventionally reared mice and germ-free, deliberately colonized adult mice (gnotobiotic mice) were used to examine the efficacy of certain intestinal microbes.


Subject(s)
Intestines/microbiology , Animals , Animals, Newborn , Humans , Immunoglobulin Isotypes/analysis , Intestinal Mucosa/immunology , Intestines/growth & development , Intestines/immunology
11.
Infect Immun ; 67(4): 1992-2000, 1999 Apr.
Article in English | MEDLINE | ID: mdl-10085047

ABSTRACT

Segmented filamentous bacteria (SFB) are autochthonous bacteria inhabiting the intestinal tracts of many species, including humans. We studied the effect of SFB on the mucosal immune system by monoassociating formerly germfree C3H/HeN mice with SFB. At various time points during 190 days of colonization, fragment cultures of small intestine and Peyer's patches (PP) were analyzed for total immunoglobulin A (IgA) and SFB-specific IgA production. Also, phenotypic changes indicating germinal center reactions (GCRs) and the activation of CD4(+) T cells in PP were determined by using fluorescence-activated cell sorter analyses. A second group of SFB-monoassociated mice was colonized with a gram-negative commensal, Morganella morganii, to determine if the mucosal immune system was again stimulated and to evaluate the effect of prior colonization with SFB on the ability of M. morganii to translocate to the spleen and mesenteric lymph nodes. We found that SFB stimulated GCRs in PP from day 6 after monoassociation, that GCRs only gradually waned over the entire length of colonization, that natural IgA production was increased to levels 24 to 63% of that of conventionally reared mice, and that SFB-specific IgA was produced but accounted for less than 1.4% of total IgA. Also, the proportion of CD4(+), CD45RBlow T cells, indicative of activated cells, gradually increased in the PP to the level found in conventionally reared mice. Secondary colonization with M. morganii was able to stimulate GCRs anew, leading to a specific IgA antibody response. Previous stimulation of mucosal immunity by SFB did not prevent the translocation of M. morganii in the double-colonized mice. Our findings generally indicate that SFB are one of the single most potent microbial stimuli of the gut mucosal immune system.


Subject(s)
Clostridium/immunology , Intestinal Mucosa/immunology , Animals , Enterobacteriaceae/immunology , Female , Germinal Center , Lymph Nodes/immunology , Mesentery , Mice , Mice, Inbred BALB C , Mice, Inbred C3H , Spleen/immunology
12.
Dev Immunol ; 6(1-2): 13-8, 1998.
Article in English | MEDLINE | ID: mdl-9716901

ABSTRACT

GALT can be subdivided into several compartments: (a) Peyer's patches (PP); (b) lamina propria (LP); and (c) intraepithelial leukocyte (IEL) spaces. The B-cell follicles of PP are quiescent in neonatal and germ-free (GF) adult mice. Germinal centers (GC), including sIgA+ blasts, appear in the B follicles of formerly GF adult mice about 10-14 days after monoassociation with various gut commensal bacteria. The GC wax and wane over about a 3-week period, although the bacterial colonizers remain in the gut at high density. Neonatal mice, born of conventionally reared (CV), immunocompetent mothers, display GC reactions in PP postweaning, although pups of SCID mothers display precocious GC reactions at about 14 days of life. Normally, gut colonization of neonates with segmented filamentous bacteria (SFB) leads to explosive development of IgA plasmablasts in LP shortly after weaning. Commensal gut bacteria and the immunocompetency of mothers also appears to control the rate of accumulation of primary B cells from "virgin" B cells in neonates. Enteric reovirus infection by the oral route can cause the activation of CD8+ T cells in the interfollicular regions of PP and the appearance of virus-specific precursor cytotoxic T lymphocytes (pCTL) in the IEL spaces. Such oral stimulation can also lead to "activation" of both CTL and natural killer (NK) cells in the IEL spaces. More normally, colonization of the gut with SFB also leads to similar activations of NK cells and "constitutively" cytotoxic T cells.


Subject(s)
Bacteria/immunology , Intestines/immunology , Intestines/microbiology , Lymphoid Tissue/physiology , Viruses/immunology , Animals , Cell Movement , Humans , Immunoglobulin A/physiology , Intestines/virology , Lymphocytes/physiology , Mice
13.
Can J Microbiol ; 44(12): 1177-82, 1998 Dec.
Article in English | MEDLINE | ID: mdl-10347864

ABSTRACT

Unlike most other indigenous bacteria, segmented filamentous bacteria (SFB) are potent activators of the mucosal immune system. SFB are strongly anchored to the epithelial cells of the small intestine where they have a preference for mucosal lymphoid epithelium. Since SFB are only present in high numbers shortly after weaning, it was investigated whether an SFB-induced immune reaction results in the removal of these bacteria from the small intestine. A correlation was found between age and colonization levels in the small intestines of SFB monoassociated Swiss mice. Five-week-old athymic BALB/c (nu/nu) mice showed lower colonization levels than their heterozygous littermates, but the opposite was found at the age of 12 weeks. However, SFB inoculation of germfree Swiss mice resulted in higher colonization levels in 5-week-old mice when compared with 4-month-old mice. We conclude that SFB colonization levels in the small intestine are likely influenced by the activity of the mucosal immune system. However, an additional age-dependent factor that modulates SFB colonization levels cannot be excluded.


Subject(s)
Bacteria/immunology , Intestinal Mucosa/immunology , Intestine, Small/immunology , Peyer's Patches/microbiology , Age Factors , Animals , Bacteria/ultrastructure , Germ-Free Life , Immunocompromised Host , Immunoglobulin A/analysis , Intestinal Mucosa/microbiology , Intestinal Mucosa/ultrastructure , Intestine, Small/microbiology , Intestine, Small/ultrastructure , Mice , Mice, Inbred BALB C , Mice, Nude , Microscopy, Electron, Scanning
14.
J Virol ; 71(5): 3431-6, 1997 May.
Article in English | MEDLINE | ID: mdl-9094613

ABSTRACT

Previous studies have found that intraepithelial lymphocytes (IEL) contain virus-specific cytotoxic T lymphocytes (CTL) that increase dramatically during the course of virus infection. In the present study, the T-cell receptor (TCR) V beta pattern used by IEL against reovirus enteric infection was investigated both in conventional and in germfree mice. IEL were isolated by a modified rapid method, and their expression of 13 TCR V betas was examined by flow cytometric analysis. The virus-specific CTL activity of each TCR V beta subset was assessed by subtraction with coated Dyna beads by a nonradioactive assay. There was a preferential perturbation of TCR V betas following virus challenge, including increases in cells expressing V beta7, -12, -14, and -17 in conventional mice and V beta2, -12, and -17 in germfree mice. In conventionally reared mice, IEL maintained and restimulated in culture had a preferential use of TCR V beta9, -12, and -17. TCR V beta2 and -17 subfamilies were found amplified in all conditions. Furthermore, TCR V beta12 and -17 accounted for 37 and 77% of the virus-specific CTL activity, respectively, after in vitro restimulation. This study provides evidence that virus-specific CTL activity may be due to the oligoclonal expansion of TCR V beta subfamilies in IEL. Our findings suggest that in vivo infection selectively presents few T-cell epitopes and that the correct identification of these T-cell epitopes would increase the likelihood of success when designing subunit vaccines.


Subject(s)
Intestinal Diseases/immunology , Intestines/immunology , Receptors, Antigen, T-Cell, alpha-beta/analysis , Reoviridae Infections/immunology , T-Lymphocytes/immunology , Animals , Epithelium/immunology , Mice , Mice, Inbred BALB C , Mice, Inbred C3H , T-Lymphocytes, Cytotoxic/immunology
15.
J Virol ; 71(4): 2844-50, 1997 Apr.
Article in English | MEDLINE | ID: mdl-9060640

ABSTRACT

Purified reovirus serotype 1, encapsulated in biodegradable aqueous microcapsules, was found to bypass maternal antibody passively transferred by suckling to neonates. Genetically identical, immunocompetent F1 scid/+ mice were generated by the reciprocal crosses of C.B17 scid/scid and normal congenic +/+ adult mice. The immunocompetent +/+ dams were either orally infected with reovirus prior to mating or not. Thus, these immunocompetent F1 pups developed either in the absence or in presence of passively transferred maternal immunity. The F1 mice were orally immunized on day 10 with either live virus, microencapsulated reovirus, or empty microcapsules plus live virus. The immune responses were assessed in the neonatal gut-associated lymphoid tissues (GALT). Examination of reovirus specific immunoglobulin A in the serum and GALT, taken on days 7, 14, and 21 postimmunization, clearly demonstrated that microencapsulated reovirus could bypass the normal effect of maternal antibodies, passively acquired by suckling, to inhibit active priming of neonates by oral route. These observations seem relevant to the development of efficacious oral vaccines that also allow passive, protective immunity via suckled maternal antibodies while permitting active oral immunization of neonates.


Subject(s)
Antibodies, Viral/immunology , Immunity, Maternally-Acquired , Orthoreovirus/immunology , Reoviridae Infections/prevention & control , Administration, Oral , Alginates/chemistry , Animals , Animals, Newborn , Capsules , Female , Glucuronic Acid , Hexuronic Acids , Immunization, Passive , Immunoglobulin A/blood , Immunoglobulin A/immunology , Male , Mice , Mice, SCID , Neutralization Tests , Spermine/chemistry , Vaccination
16.
Zentralbl Bakteriol ; 283(4): 431-65, 1996 Apr.
Article in English | MEDLINE | ID: mdl-8737943

ABSTRACT

An International Study Group on New Antimicrobial Strategies (ISGNAS) has been formed in response to the recognition that development of microbial resistance to antibiotics is becoming a serious, world-wide problem. The group met in 1993 for the first time to discuss the feasibility of developing rational alternatives to the use of antibiotics and prepared, as a result, a comprehensive overview of normal (physiological) mechanisms involved in the control of potentially pathogenic (oppotunistic) microorganisms. One objective of ISGNAS is to understand the conditions which allow opportunistic microbes present among the symbionts to cause an infection. There is a need for more coherent information concerning the habitat, growth requirements and host and pathogen properties which allow opportunistic pathogens to cause life-threatening infections. In particular, information is urgently being sought to understand the complexity of the interactions between the vast number of microbial species, and the interactions between the microbes and their host. Another goal is to inspire and enable basic and clinical research that will lead to the development of new therapies for regulating colonization, translocation and infection by opportunistic micro-organisms in patients during periods of decreased resistance. With a sufficient amount of knowledge of how healthy individuals keep opportunistic micro-organisms under control, it may become feasible for physicians to maintain host resistance and inter-microbial factors involved in the containment of opportunistic microbes. Therapies aimed at boostering natural resistance mechanisms will be of critical importance to individuals whose resistance has been compromised as a result of another clinical condition.


Subject(s)
Opportunistic Infections/prevention & control , Adjuvants, Immunologic/therapeutic use , Antibodies/immunology , Humans , Immunization, Passive , Intestines/immunology , Intestines/microbiology , Macrophages/immunology , Nutritional Physiological Phenomena
17.
Infect Immun ; 64(2): 616-23, 1996 Feb.
Article in English | MEDLINE | ID: mdl-8550216

ABSTRACT

We transferred peritoneal cells from BALB/c mice into C.B17 scid/scid mice. Six to eight months after injection, only cells with the B1 phenotype were retained in the spleens and peritoneal cavities of these mice. The lamina propria of the intestine contained many peritoneal, donor-derived, immunoglobulin A (IgA)-producing cells. The mesenteric lymph nodes of these mice were found to be a major site of proliferation and generation of IgA plasmablasts. We established eight IgA-producing hybridomas from the mesenteric lymph nodes of such mice, and all the hybridomas reacted with different but partially overlapping fecal bacterial populations. Cloning and sequencing of the VH genes of these hybridomas showed that two hybridomas utilized germ line-encoded VH genes while the VH genes of the six hybridomas showed somatic mutations, some of which are indicative of an antigen-driven selection process.


Subject(s)
Antibodies, Monoclonal/genetics , B-Lymphocytes/immunology , Bacteria/immunology , Feces/microbiology , Genes, Immunoglobulin , Immunoglobulin A/genetics , Immunoglobulin Heavy Chains/genetics , Immunoglobulin Variable Region/genetics , Animals , Base Sequence , Flow Cytometry , Hybridomas/immunology , Immunoglobulin A/immunology , Mice , Mice, Inbred BALB C , Mice, SCID , Molecular Sequence Data , Mutation , Peritoneal Cavity/cytology
18.
Infect Immun ; 63(10): 3904-13, 1995 Oct.
Article in English | MEDLINE | ID: mdl-7558298

ABSTRACT

We have employed a germfree mouse model to study the development and persistence of a humoral mucosal immune response to a gram-negative murine commensal organism, Morganella morganii. M. morganii bacteria rapidly colonize the gut, resulting in hypertrophy of Peyer's patches (PP), including germinal center reactions (GCR), and the development of specific immunoglobulin A (IgA) responses detected in vitro in PP fragment cultures and by ELISPOT assays of lamina propria cells. The GCR peaks 14 days after infection and begins to wane thereafter. Upon colonization, the organisms successfully translocate to the mesenteric lymph node and spleen, but the number of translocating bacteria begins to drop with the onset of a specific IgA response. A clonal B-cell microculture technique was used to determine the frequency of specific IgA plasmablasts and IgA memory cells. The frequencies of preplasmablasts were seen to be higher in the earlier stages of germinal center development, whereas the frequencies of antigen-specific memory cells appeared to remain at a relatively constant level even after 193 days postmonoassociation. We suggest that a successful secretory IgA response can attenuate chronic stimulation of GCR even though the bacteria persist in the gut. The observed developing hyporesponsiveness to a chronically present commensal organism may be relevant to the use of bacterial vectors for mucosal immunization.


Subject(s)
Antibodies, Bacterial/biosynthesis , Immunoglobulin A, Secretory/biosynthesis , Intestinal Mucosa/immunology , Intestines/microbiology , Animals , Immunologic Memory , Lymph Nodes/microbiology , Mice , Mice, Inbred C3H , Organ Culture Techniques , Peyer's Patches/immunology , Phosphorylcholine/immunology , Spleen/microbiology
19.
Int Immunol ; 7(6): 911-8, 1995 Jun.
Article in English | MEDLINE | ID: mdl-7577799

ABSTRACT

Reciprocal crossings of C.B17 scid/scid and congenic BALB/c (+/+) mice generate genetically identical, immunocompetent F1 scid/+ mice that develop in either the absence or influence of passively transferred maternal immunity. By exchanging F1 scid/+ litters at birth among scid/scid, non-immune or reovirus immune BALB/c mothers we examined the relative ability of placental or colostral/milk transfer of virus specific maternal antibodies to interfere with reovirus immunization of the neonatal gut associated lymphoid tissues (GALT). Our data demonstrate that the Peyer's patches (PP) in 10-day-old mice are competent to support thymus dependent responses to acute reovirus stimulation that include the rapid (within 3 days) development of specific IgA plasma cells and the subsequent initiation of PP germinal center reactions. These neonatal mucosal immune responses occur independently of coincident specific maternal immune responses as evidenced by the identity of the reovirus specific responses engendered in F1 scid/+ pups of scid/scid versus +/+ mothers. However, transfer of pre-existing reovirus specific maternal antibody in milk via nursing on a reovirus immune (foster) mother completely abrogated reovirus specific neonatal IgA responses; while placental transfer of specific maternal antibody alone did not interfere with the immunization of the neonatal GALT with enteric reovirus. Reovirus challenge of 10-day-old mice was associated with a substantial bystander IgA response. Possible mechanisms responsible for the induction of the observed bystander IgA responses are discussed.


Subject(s)
Animals, Suckling/immunology , Antibodies, Viral/biosynthesis , Immunity, Maternally-Acquired/physiology , Immunoglobulin A/biosynthesis , Peyer's Patches/immunology , Reoviridae/immunology , Animals , Antibody Specificity , Antibody-Producing Cells/virology , Female , Germ-Free Life/immunology , Germinal Center/immunology , Germinal Center/virology , Male , Mice , Mice, Inbred BALB C , Mice, SCID , Reoviridae Infections/immunology
20.
J Virol ; 69(6): 3910-4, 1995 Jun.
Article in English | MEDLINE | ID: mdl-7745745

ABSTRACT

Severe combined immune deficient (SCID) mice infected orally with reovirus type 1/L die of hepatitis. Leukocytes bearing the cell surface antigens Thy-1.2 and asialoGM-1 (AsGM1) accumulate in the livers of infected animals. These cells display lytic activity toward natural killer-sensitive (YAC-1) and -resistant (P815) cell lines and murine hepatoma line Hepa 1/A1. Although these cells have the capacity to lyse infected hepatoma targets, they cannot clear the virus.


Subject(s)
Killer Cells, Lymphokine-Activated/immunology , Killer Cells, Natural/immunology , Reoviridae Infections/immunology , Animals , Cytotoxicity, Immunologic , Liver Diseases/immunology , Liver Diseases/virology , Mice , Mice, Inbred BALB C , Mice, SCID
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