The IgA/IgM receptor expressed on a murine B cell lymphoma is poly-Ig receptor.

T560, a mouse B lymphoma that originated in gut-associated lymphoid tissue, expresses receptors that bind dimeric IgA and IgM in a mutually inhibitory manner but have little affinity for monomeric IgA. Evidence presented in this paper indicates that the receptor is poly-Ig receptor (pIgR) known in humans and domestic cattle to bind both IgA and IgM. The evidence includes the demonstration that binding of IgM is J chain dependent, and that pIg-precipitated receptor has an appropriate Mr of 116-120 kDa and can be detected on immunoblots with specific rabbit anti-mouse pIgR. Overlapping RT-PCR performed using template mRNA from T560 cells and oligonucleotide primer pairs designed from the published sequence of mouse liver pIgR indicate that T560 cells express mRNA virtually identical with that of the epithelial cell pIgR throughout its external, transmembrane, and intracytoplasmic coding regions. Studies using mutant IgAs suggest that the Calpha2 domain of dimeric IgA is not involved in high-affinity binding to the T560 pIgR. Inasmuch as this mouse B cell pIgR binds IgM better than IgA, it is similar to human pIgR and differs from rat, mouse, and rabbit epithelial cell pIgRs that bind IgA but not IgM. Possible explanations for this difference are discussed. All clones of T560 contain some cells that spontaneously secrete both IgG2a and IgA, but all of the IgA recoverable from the medium and from cell lysates is monomeric; it cannot be converted to secretory IgA by T560 cells.

IgA production in MHC class II-deficient mice is primarily a function of B-1a cells.

Mice deficient in MHC class II expression (C2d mice) do not make antibody to protein antigens administered systemically, but their ability to produce IgA antibody to antigen administered at mucosal sites has not been described. We investigated IgA production by C2d mice and their IgA antibody response to antigen given orally. Young C2d mice had normal amounts of serum IgA, intestinal-secreted IgA and normal numbers of intestinal IgA plasma cells, compared to control C57BL/6 mice. IgA production by C2d mice increased with age. Following oral immunization with cholera toxin, C57BL/6 mice responded with IgA and IgG antibody, and had increased numbers of IgA plasma cells, but C2d mice gave no response. The Peyer's patch and mesenteric lymph node tissues of C2d mice contained very few CD4-expressing T cells. Thus, C2d mice have no typical mucosal CD4 Th cells and cannot respond to a strong oral immunogen, yet they still produced and secreted IgA. We hypothesized that B-1 lymphocytes could provide a source of IgA independent of antigen-specific T cell help. Young C2d mice had normal numbers of peritoneal B-1a cells and their frequency increased with age. To test the role of these B-1a cells, we bred C2d mice to obtain mice that had no MHC class II expression and expressed the Xid gene that confers deficiency in B-1a cells. These double-deficient mice had 10-fold less serum and secreted IgA than all other F2 littermates. We conclude that B-1a cells are essential for the majority of IgA production in C2d mice. Thus, the C2d mouse may provide a useful tool for analysis of the role of intestinal IgA provided by B-1a cells.

Oral administration of polymer-grafted starch microparticles activates gut-associated lymphocytes and primes mice for a subsequent systemic antigen challenge.

The mucosal and systemic humoral immune systems can function essentially independent of each other, responding to mucosal and parenteral antigens, respectively. Nevertheless, antigen administered by one route can modify responsiveness to subsequent immunization by an alternate route. Here we demonstrated, in mice, in addition to stimulating rapid and robust sera antibody responses, intragastric (i.g.) immunization with human serum albumin (HSA)-containing starch microparticles (MP) grafted with 3-(triethoxysilyl)-propyl-terminated polydimethylsiloxane (TS-PDMS) primed for enhanced specific sera IgG following a parenteral antigen boost. After as little as one i.g. immunization with microentrapped, but not with soluble, HSA antigen-specific proliferation and antibody secretion were detected in Peyer's patches (PP); this activity peaked after three i.g. MP immunizations. We observed a progressive dissemination of antigen-specific lymphocyte reactivity from PP to splenic tissue following oral MP immunization. Similarly, we observed a shift in HSA-specific antibody-secreting cells from PP and mesenteric lymph nodes to splenic tissue following i.g. MP immunization. We also demonstrated that oral immunization with microentrapped, but not with soluble HSA, resulted in enhanced numbers of spontaneous Th2-cytokine secreting lymphocytes which disseminated from mucosal to systemic lymphoid compartments. This observation coincided with our findings that HSA-specific sera IgG1 responses in animals given HSA in MP were significantly higher than those detected in the sera of mice given soluble HSA i.g., both before and after parenteral antigen challenge. These findings suggest that orally-administered TS-PDMS-grafted MP, by stimulating elements of the mucosal immune system, are a valuable addition to mucosal and systemic vaccine delivery systems.

Intranasal immunization with polymer-grafted microparticles activates the nasal-associated lymphoid tissue and draining lymph nodes.

Waldeyer's ring is located at the juncture of the respiratory and alimentary tracts, where it is bombarded by inhaled and ingested antigens. However, knowledge of its exact function or consequences of its removal is incomplete. Recently, the murine nasal-associated lymphoid tissue (NALT) has been reported to have functional similarities to Waldeyer's ring and, thus, might be a suitable model to examine the function of oronasopharyngeal lymphoid tissues. To explore the capability of NALT to incite local mucosal and systemic immunity, we immunized mice intranasally (i.n.) with 3-(triethoxysilyl)-propyl-terminated polydimethylsiloxane (TS-PDMS)-grafted microparticles (MP), an inoculant previously shown to induce robust systemic and mucosal humoral immunity following intragastric (i.g.) administration. We demonstrated that i.n. immunization with low doses of microentrapped, but not soluble, human serum albumin (HSA) evoked robust circulating IgG responses (P < 0.05). Additionally, NALT cells isolated from MP-treated mice proliferated in vitro when restimulated with HSA (P < 0.05), suggesting that i.n. immunization with HSA-containing MP incited specific immunity via NALT cell activation. Coinciding with these observations, after i.n. MP administration HSA-specific spot-forming cells (SFC) were observed in NALT, and later posterior cervical lymph nodes (pCLN) and spleen (SPL), suggesting that the observed MP-induced specific systemic antibody responses emanated from the NALT. We also showed that i.n. immunization with HSA-containing TS-PDMS-grafted MP stimulated interleukin-4 (IL-4)-secreting lymphocytes in the NALT. This cytokine microenvironment was probably responsible for driving the IgG1 sera response observed after i.n. MP administration, via the migration of NALT-derived IgG1-committed B cells. Interestingly, unlike i.g. MP administration, i.n. immunization with HSA-containing MP did not evoke detectable specific IgA in any lymphoid tissue examined, or in nasal secretions, probably reflecting differences between NALT and other mucosae-associated lymphoid tissues (MALT).

Nasopharyngeal colonization with nontypeable Haemophilus influenzae in chinchillas.

Colonization of the nasopharynx by a middle ear pathogen is the first step in the development of otitis media in humans. The establishment of an animal model of nasopharyngeal colonization would therefore be of great utility in assessing the potential protective ability of candidate vaccine antigens (especially adhesins) against otitis media. A chinchilla nasopharyngeal colonization model for nontypeable Haemophilus influenzae (NTHI) was developed with antibiotic-resistant strains. This model does not require coinfection with a virus. There was no significant difference in the efficiency of NTHI colonization between adult (1- to 2-year-old) and young (2- to 3-month-old) animals. However, the incidence of middle ear infection following nasopharyngeal colonization was significantly higher in young animals (83 to 89%) than in adult chinchillas (10 to 30%). Chinchillas that had recovered either from a previous middle ear infection caused by NTHI or from an infection by intranasal inoculation with NTHI were completely protected against nasopharyngeal colonization with a homologous strain and were found to be the best positive controls in protection studies. Systemic immunization of chinchillas with inactivated whole-cell preparations significantly protected animals not only against homologous NTHI colonization but also partially against heterologous NTHI infection. In all protected animals, significant serum anti-P6 and anti-HMW antibody responses were observed. The outer membrane P6 and high-molecular-weight (HMW) proteins appear to be promising candidate vaccine antigens to prevent nasopharyngeal colonization and middle ear infection caused by NTHI.

Chlamydia trachomatis infection in the female reproductive tract of the rat: influence of progesterone on infectivity and immune response.

As the most common cause of sexually transmitted disease in women, chlamydial infections can lead to pelvic inflammatory disease, infertility, and ectopic pregnancy. To better understand the role played by sex hormones in modulating the immune response of the genital tract to microbial infections, we have developed a rat model to study Chlamydia trachomatis infection. Inbred female Lewis rats were primed with progesterone and inoculated by intrauterine instillation of C. trachomatis (mouse pneumonitis strain MoPn) into each uterine horn. When infected animals were examined for the presence of chlamydial antigens 14 days postinfection, both the uterus and vagina were found to be positive compared to those of saline-treated animals, which did not show specific staining. The involvement of local and systemic immune systems following chlamydial infection was determined by analyzing major histocompatibility complex (MHC) class II expression in the reproductive tract and lymphocyte proliferation in response to mitogenic and chlamydia-specific stimulation of cells from the spleen and lymph nodes (LN) draining the reproductive tract. Enhanced proliferation was observed in LN following mitogenic but not antigenic (MOMP [major outer membrane protein]) stimulation. In contrast, spleen cell proliferation was lower in chlamydia-infected rats than in saline-treated controls. MHC class II expression, an indicator of inflammatory responses, was upregulated in the uterus, on glandular epithelial cells, and adjacent to glands in response to chlamydial infection. In other experiments, when rats were infected at estrus and diestrus without prior progesterone priming, chlamydial inclusions were not detected in either the uterus or vagina. However, enhanced lymphocyte proliferation was observed in response to mitogenic and MOMP stimulation in the reproductive tract-draining LN from estrous and diestrous animals. These findings indicate that under appropriate endocrine conditions, the rat uterus is susceptible to C. trachomatis infection and that immune responses to this pathogen can be detected locally and systemically. Further, they suggest that clearance of the infection from the reproductive tract involves immune cells from the LN draining the reproductive tract.

Comparison of murine nasal-associated lymphoid tissue and Peyer's patches.

The nasal mucosal is the first site of contact with inhaled antigens. However, the nature of local immune responses and the role of nasal-associated lymphoid tissue (NALT) in those responses have rarely been studied. To characterize the cells involved in mucosally derived immune responses, NALT and Peyer's patch (PP) cells from normal mice, and mice immunized intragastrically or intranasally with cholera toxin (CT), were isolated and analyzed. Compared with PP cells, unstimulated NALT cells contained a higher proportion of T-cells. The CD4:CD8 ratio in NALT cell preparations was less than that observed in PP and more closely resembled that seen in spleen. Additionally, the total B-cell frequency in NALT cell isolates was 20% lower than that observed in PP cell preparations. Although NALT and PP cell isolates contained both mature B-cells and cells undergoing activation to express surface IgA, unlike PP, NALT showed no significant frequency of IgA-switched cells. After intranasal immunization with CT, toxin-specific IgA antibody-forming cells (AFCs) were detected in NALT cell preparations. The numbers of these cells correlated with CT-specific IgA in nasal, but not in gut washes or sera, thus suggesting local nasal production of antigen-specific mucosal antibodies. There was no evidence of anti-CT AFCs in NALT or CT-specific antibody in nasal washes after intragastric CT administration. These results support the notion that nasal mucosal antibody production is best achieved via direct stimulation of IgA-committed, NALT-derived B-cells.

Intranasal antigen targeting to MHC class II molecules primes local IgA and serum IgG antibody responses in mice.

Covalent conjugates of hen egg lysozyme (HEL) and anti-major histocompatibility complex (MHC) class II monoclonal antibodies (mAb) were used to immunize mice intranasally. Three weeks after intranasal (IN) priming, mice responded rapidly to IN challenge with a mixture of HEL and cholera toxin (CT), by producing large titres of anti-HEL IgA and IgG1 antibody in serum, and IgA antibody in nasal secretions. No secondary response to HEL plus CT occurred in mice that received no priming or mice primed with HEL alone. The secondary serum IgG antibody response was dominated by the IgG1 subclass. HEL combined with CT adjuvant worked much better than HEL alone in producing the secondary response. Control conjugates, containing an IgG isotype-matched mAb without specificity for mouse tissues, provided poor priming. Mice expressing MHC class II molecules, to which the anti-MHC II mAb could not bind, produced a weak antibody response compared with those that expressed the appropriate. MHC class II molecule. Our results demonstrate that immunotargeting to MHC class II molecules via the IN route allows priming of the local IgA and circulating IgG antibody, and indicate that this technique is a feasible approach for delivery of subunit vaccines in the upper respiratory tract.

Novel polymer-grafted starch microparticles for mucosal delivery of vaccines.

Recent studies have demonstrated that systemic and mucosal administration of soluble antigens in biodegradable microparticles can potentiate antigen-specific humoral and cellular immune responses. However, current microparticle formulations are not adequate for all vaccine antigens, necessitating the further development of microparticle carrier systems. In this study, we developed a novel microparticle fabrication technique in which human serum albumin (HSA) was entrapped in starch microparticles grafted with 3-(triethoxysilyl)-propyl-terminated polydimethylsiloxane (TS-PDMS), a biocompatible silicone polymer. The immunogenicity of HSA was preserved during the microparticle fabrication process. Following intraperitoneal immunization of mice, TS-PDMS-grafted microparticles (MP) dramatically enhanced serum IgG responses compared with ungrafted MP and soluble HSA alone (P < 0.001). When delivered orally, both TS-PDMS-grafted and ungrafted microparticles elicited HSA-specific IgA responses in gut secretions, in contrast to orally administered soluble antigen. Indeed, TS-PDMS-grafted microparticles stimulated significantly stronger serum IgG (P < 0.005) and IgA (P < 0.001) responses compared with those elicited by ungrafted microparticles. These findings indicate that TS-PDMS-grafted starch microparticles have potential as systemic and mucosal vaccine delivery vehicles.

Intestinal immunization of mice with antigen conjugated to anti-MHC class II antibodies.

We have explored a new technique for immunization of the intestinal tract of mice, using protein antigens bound to antibodies with specificity for murine MHC class II molecules (MHC-II). Either of two protein antigens, hen avidin (AV) or hen egg lysozyme (HEL) were covalently conjugated to anti-MHC-II antibodies and the purified conjugates were given orally (p.o.) or by direct intraduodenal (i.d.) injection into the intestinal lumen of mice. A secondary immunization p.o. with the same conjugate or with the non-conjugated antigen in the presence of cholera toxin (CTX) resulted in production of both intestinal secretory IgA and serum IgA antibody by those mice. In addition, serum IgG antibodies were produced. Conjugates with appropriate MHC-II specificity targeted the antigen because they induced more IgA and IgG antibody than conjugates with irrelevant antibody specificity or antigen alone, and because they induced antibody in mice that were genetic low responders to antigen. The results indicate the feasibility of oral subunit type vaccines with antibody targeting technology.

Murine intestinal disaccharidases: identification of structural variants of sucrase-isomaltase complex.

This study was directed to determine the extent of variability in structure or expression of intestinal disaccharidase [gamma-glucoamylase (gamma-GA), sucrase-isomaltase (SI), and lactase] between different strains of mice. Reduced levels of sucrase activity (approximately 20 U/g of protein) were observed in three strains of mice belonging to the CBA/Ca lineage. Four other strains of mice analyzed exhibited higher levels of sucrase activity (approximately 50 U/g of protein). Decreased levels of sucrase in CBA/Ca mice were not associated with decreased levels of activity associated with the isomaltase subunit or with decreased levels of SI mRNA expression. High-performance liquid chromatographic gel filtration, heat inactivation, and kinetic analysis indicated that the differences between strains in sucrase activity might be attributed to structural differences in the sucrase subunit of the SI complex, thus rendering it more susceptible to cleavage and inactivation. However, no differences in kinetic properties of the sucrase subunit were observed between strains. Murine gamma-GA was found to account for a greater proportion of maltase activity (approximately 70%) than that observed in other species (i.e., approximately 20%). In addition, CBA/Ca mice were found to be deficient in intestinal maltase activity (approximately 60 U/g) compared with the other strains studied (approximately 300 U/g).

Characterization of intestinal gamma-glucoamylase deficiency in CBA/Ca mice.

In previous work, we found that CBA/Ca mice display only 20% of the maltase activity present in other mouse strains. In this study, we characterized more fully the maltase deficiency in CBA/Ca mice. Virtually all of the intestinal maltase activity of CBA/Ca mice was inactivated at 50 degrees C, indicating that it was due only to the sucrase-isomaltase complex. High-performance liquid chromatographic analysis revealed that CBA/Ca mice had undetectable maltase activity displaying the molecular mass characteristic of murine gamma-glucoamylase (gamma-GA) (530 kDa). Gel electrophoretic analysis confirmed that CBA/Ca mice lacked maltase activity with molecular mass of 530 kDa corresponding to gamma-GA. Two-dimensional electrophoretic analysis revealed that the gamma-GA deficiency in CBA/Ca mice was due to the failure to synthesize the enzyme and not to the synthesis of an inactive protein. gamma-GA maltase activity could not be induced in CBA/Ca mice by a diet rich in starch, whereas the activity of other disaccharidases were readily increased. gamma-GA-deficient CBA/Ca mice appear to lack any gross metabolic abnormality resulting from this defect.

Vaccination by cholera toxin conjugated to a herpes simplex virus type 2 glycoprotein D peptide.

Immunization of BALB/cJ mice with a peptide corresponding to residues 1 to 23 of glycoprotein D [gD(1-23)] from herpes simplex virus type 2 (HSV-2) elicits antibody responses which correlate with protection against lethal HSV-2 infection. In the present study, we examined the ability of cholera toxin (CTX) to act as an immunogenic carrier for gD(1-23). The number of gD(1-23) residues conjugated to CTX affected its binding to GM1 ganglioside and physiological toxicity, both of which are factors affecting oral immunogenicity. The antibody response elicited after intraperitoneal (i.p.) immunization with the CTX-gD(1-23) conjugate was protective against a lethal i.p. challenge with HSV-2. In other experiments, mice were immunized i.p. on day 0 and subsequent immunizations conducted on days 14 and 28 were administered either intragastrically or intravaginally (i.vag.). Intraperitoneal priming followed by either i.p or intragastric boosting resulted in anti-HSV-2 antibodies in vaginal washings and in protection against a lethal i.vag. challenge with HSV-2. Intraperitoneal priming followed by i.vag. boosting did not elicit anti-HSV-2 antibodies in vaginal washings and did not protect mice against a lethal i.vag. challenge with HSV-2. These results suggest that CTX can act as a systemic and an oral delivery molecule for the covalently linked gD(1-23) peptide and that such conjugates can elicit protective immune responses against systemic and genital HSV-2 infection.

Rat secretory component binds poorly to rodent IgM.

Our previous studies and those of others indicated that human secretory component (SC), the five domain extracellular portion of the poly Ig receptor, binds avidly to both pIgA and IgM. In this study we report that in rodents, SC binds primarily to pIgA. Rat secretory component was isolated from bile and radiolabeled to known specific activity with 125I. Radiolabeled rat SC was incubated with rat and mouse monoclonal proteins for 1 h at room temperature and overnight at 4 degrees D. Binding of 125I-rat SC to Ig was determined in two ways: 1) immunoprecipitation of putative 125I-rat SC-Ig complexes with anti-L chain antibodies; 2) HPLC gel filtration on an analytical TSK 4000 column that separated free 125I-rat SC from 125I-rat SC bound to Ig. Both methods of analysis yielded similar results. Rat and mouse polymeric (p) IgA bound rat SC with high avidity, although the binding activity of the IgM from either species was virtually nil. The number of SC-binding sites on rat polymeric Ig was determined by immunoprecipitation of mixtures of rat pIg with saturating concentrations of 125I-rat SC and yielded values of 1.0 and 0.05 for rat pIgA and IgM, respectively. The significance of these findings with respect to the biologic function of the pIg R in rodents and the nature of the pIg R-binding site on pIg is discussed.

Further evidence that hepatic sources confer biliary antibody in the rat.

The notion that bile-dedicated antibody is made within the liver by migratory antibody-forming cells (AFC) was examined further in rats. Livers from immunized animals were removed to perfusion in isolation so that plasma influences on bile antibody would be obviated. Antibody was secreted for at least 5 hr by the livers of rats that had received intravenous (i.v.) or intra-Peyer's patch (IPP) immunization with horse erythrocytes. After initially declining, the titres stabilized at 5-8% of the starting value for IPP-immunized rats and at 0.8% for i.v.-immunized animals, levels that were then sustained. In other experiments, the biliary antibody output was measured in immunized rats in the period immediately following splenectomy, an expedient that would deny the liver any newly formed AFC. Splenectomy during spleen-based, IgM antibody responses led to bile titres falling, over about 12 hr, to 21% of initial values. This level was then maintained for at least another 12 hr. Serum titres over this period remained static. Lastly, the bile ducts of immunized rats were ligated to test whether locally made antibody that was destined for bile could be forced instead to reflux to blood. Biliary obstruction during IgM responses to horse erythrocytes and pneumococcal polysaccharide, type 3, was found to raise significantly serum antibody titres. For pneumococcal polysaccharide, the serum response was also noticeably prolonged. These findings are consistent with the biliary antibody of immunized rats being constituted, in part, from local sources and not from plasma alone.

Cholera toxin conjugates for intragastric vaccination against herpes simplex virus type 2.

In this study, we tested the hypothesis that enteric immunization with cholera toxin (CTX) conjugated to glycoprotein D (gD) of herpes simplex virus type 2 (HSV-2) or a peptide corresponding to residues (1-23) of gD (gD(1-23)) would induce relevant antiviral immunity. Intraperitoneal (IP) immunization of mice with CTX-gD(1-23) conjugate induced anti-HSV-2 sera antibody responses which correlated with protection from a lethal IP challenge with HSV-2. Intragastric (IG) immunization of mice with the same conjugate or a CTX-gD conjugate did not result in measurable anti-HSV-2 responses in sera or vaginal washings and only small numbers of anti-HSV-2 antibody-secreting cells (ASC) were found in intestinal lamina propria cell and splenocyte preparations. In comparison, anti-CTX responses were detected in sera and vaginal washings after IG immunization with CTX and anti-CTX ASC in lamina propria cell preparations accounted for 5-10% of total ASC detected at this site. No significant differences in the survival of mice immunized with the conjugates were noted after a lethal intravaginal (IVAG) challenge with HSV-2. The poor enteric immunogenicity of gD(1-23) and gD conjugated to CTX was attributable to proteolysis in the gastrointestinal tract. These results indicate that although peptide-CTX conjugates can induce protective immune responses when administered parenterally, it may not be feasible to use peptides as the basis of an oral vaccine unless methods are developed to protect these antigens from degradation in the gastrointestinal tract.

Binding of secretory component to protein 511, a pIgA mouse protein lacking 36 amino acid residues of the C alpha 3 domain.

Protein 511, a murine IgA protein described previously by Robinson and Appella [Proc. natn. Acad. Sci. U.S.A. 77, 4909-4913 (1980)] which lacks 36 amino acids in the C alpha 3 domain, was tested for its ability to bind to radiolabelled secretory component (125I-rat SC) and to be transported from blood to bile in the rat, a function described previously to be mediated by the poly Ig receptor (pIg R). When compared to other mouse pIgA proteins, the naturally occurring mutant protein 511 bound 125I-rat SC and was transported from blood to bile in a manner indistinguishable from wild-type pIgA protein. We conclude that the region of Fc alpha which is missing in protein 511, is not involved in mediating the binding of pIgA to the pIg R.

Acquired resistance to Giardia muris in X-linked immunodeficient mice.

A previous study from this laboratory (D. P. Snider, D. Skea, and B. J. Underdown, Infect. Immun. 56:2838-2842, 1988) indicated that immunodeficient mice expressing the xid gene develop prolonged infections with Giardia muris, unlike immunocompetent mice, which eliminate the intestinal protozoan parasite in 8 to 10 weeks. In this study, CBA/N (xid) and CBA/Ca mice were infected with G. muris cysts and at various times following this primary infection were cured by treatment with metronidazole. In contrast to the marked differences in the ability of xid and normal mice to eliminate a primary infection, mice of both strains were resistant to a secondary challenge of G. muris cysts. These data imply that the mechanism(s) responsible for elimination of a primary infection is not identical to those required to resist a secondary challenge infection. Splenocytes from immunocompetent CBA/Ca mice (but not immunodeficient CBA/N mice) could transfer the ability to eliminate a primary G. muris infection to irradiated mice of either strain. In contrast, splenocytes from previously infected CBA/Ca mice could not transfer resistance to a challenge infection, further supporting the hypothesis that there are differences between mechanisms required to eliminate a primary infection and those necessary to resist a second challenge infection.

Cognate T-cell help in the induction of IgA responses in vivo.

Evidence from in vitro studies indicates that immunoglobulin A (IgA) responses are highly T dependent, yet investigations of the requirement for cognate help for IgA responses in vivo have not previously been undertaken. Experiments reported in this paper employ hapten-carrier immunization of individual Peyer's patches (PP), the site of generation of IgA antibody-containing cells (ACC) responding to lumenal antigenic challenge in the intestine, to determine the requirements for T-cell and B-cell priming under normal physiological conditions in vivo. These experiments demonstrate that both hapten-specific B-cell priming and carrier-specific T-cell priming in PP are required for an IgA-specific anti-hapten ACC response in the intestinal lamina propria to subsequent lumenal challenge with hapten-carrier conjugate. These results confirm that IgA B-cell induction in PP requires cognate T-cell help. An IgA ACC response can also be obtained when hapten and carrier priming occur in different PP, providing functional significance for our previous observations that PP-derived T-helper cells are able to migrate between PP after priming.

Early expression in mice of genetic resistance to intestinal amebiasis.

Some strains of mice are known to be relatively resistant to hepatic or intestinal amebic infections. In order to know if the intestinal resistance is expressed few hours after infection, we inoculated axenic amebae in three inbred strains of mice either by direct intracecal injection or by infection of a washed-closed cecal loop. We found that amebae do not survive in conventional animals but they colonize longer in animals with the cecal loop. However, the survival was low after 24 hours postinfection. Balb/c mice were more susceptible and CBA mice more resistant. Our results suggest that genetic resistance to intestinal amebiasis is expressed in mice in the early phases of infection.