CD28-deficient mice generate an impaired Th2 response to Schistosoma mansoni infection

Engagement of CD28 on T cells provides a co-stimulatory signal necessary for T cell activation and differentiation. Recent findings suggest that priming of T helper (Th)2 cells is more dependent on CD28 activation than Th1 cells. The present study examines whether mice that lack expression of CD28 as a result of gene targeting are capable of generating a Th2 response characteristic during infection with the intravascular trematode parasite Schistosoma mansoni. Mutant and control mice were either inoculated in the footpad with S. mansoni eggs (a potent inducer of a Th2 response) or infected percutaneously with the parasite. Draining lymph nodes (after footpad injection) or spleen cells (after natural infection) were harvested at 12 days and 8 weeks, respectively, and examined for cytokine responses to egg antigens. CD28-deficient mice (−/−) generated diminished egg antigen-driven interleukin (IL)-4 and IL-5 production (by 5- to 17-fold, respectively) compared to CD28-expressing (+/+) littermates. In contrast, lymphocyte proliferation and interferon (IFN)-γ production to egg antigens were equivalent for mutant and control mice. Infected CD28−/− mice also had reduced immunoglobulin secretion. Serum levels of parasite antigen-specific IgG1 and polyclonal IgE were significantly diminished in CD28−/− compared to CD28+/+ mice. Lack of CD28 expression had no effect on granuloma formation around eggs trapped in the liver, but increased susceptibility of mice to primary schistosomiasis infection. These studies indicate that CD28 activation contributes to T cell priming required for generation of a Th2 response to an intravascular dwelling helminth parasite.     

Comparison of thymocyte development and cytokine production in CD7-deficient, CD28-deficient and CD7/CD28 double-deficient mice

CD7 and CD28 are Ig superfamily molecules expressed on thymocytes and mature T cells that share common signaling 0mechanisms and are co-mitogens for T cell activation. CD7-deficient mice are resistant to lipopolysaccharide (LPS)-induced shock syndrome, and have diminished in vivo LPS-triggered IFN-gamma and tumor necrosis factor (TNF)-alpha production. CD28-deficient mice have decreased serum Ig levels, defective IgG isotype switching, decreased T cell IL-2 production and are resistant to Staphylococcus aureus enterotoxin-induced shock. To determine synergistic roles CD7 and CD28 might play in thymocyte development and function, we have generated and characterized CD7/CD28 double-deficient mice. CD7/CD28-deficient mice were healthy, reproduced normally, had normal numbers of thymocyte subsets and had normal thymus histology. Anti-CD3 mAb induced similar levels of apoptosis in CD7-deficient, CD28-deficient and CD7/CD28 double-deficient thymocytes as in control C57BL/6 mice (P = NS). Similarly, thymocyte viability, apoptosis and necrosis following ionomycin or dexamethasone treatment were the same in control, CD7-deficient, CD28-deficient and CD7/CD28-deficient mice. CD28-deficient and CD7/CD28-deficient thymocytes had decreased [3H]thymidine incorporation responses to concanavalin A (Con A) stimulation compared to control mice (P < or = 0.01 and P < or = 0.05 respectively). CD7/CD28 double-deficient mice had significantly reduced numbers of B7-1/B7-2 double-positive cells compared to freshly isolated wild-type, CD7-deficient and CD28-deficient thymocytes. Con A-stimulated CD4/CD8 double-negative (DN) thymocytes from CD7/CD28 double-deficient mice expressed significantly lower levels of CD25 when compared to CD4/CD8 DN thymocytes from wild-type, CD7-deficient and CD28-deficient mice (P < 0.05). Anti-CD3-triggered CD7/CD28-deficient thymocytes also had decreased IFN-gamma and TNF-alpha production compared to C57BL/6 control, CD7-deficient and CD28-deficient mice (P < or = 0.05). Thus, CD7 and CD28 deficiencies combined to produce abnormalities in the absolute number of B7-1/B7-2-expressing cells in the thymus, thymocyte IL-2 receptor expression and CD3-triggered cytokine production.     

Simvastatin does not prevent acute cardiac allograft rejection in CD28-deficient mice

Background  

Heterotopic cardiac transplantation in mice was used to determine whether complementary immunomodulation by statins prevents lymphocyte function-associated antigen (LFA)-1-dependent acute allograft rejection in vivo.     

Experimental autoimmune meningitis: a novel neurological disease in CD28-deficient mice

C57BL/6 mice develop T-cell-mediated experimental autoimmune encephalomyelitis (EAE) after immunization with the neuroantigen myelin oligodendrocyte glycoprotein. (MOG). We immunized CD28-deficient C57BL/6 mice to determine the role of T cell costimulation in the immune response to MOG. CD28-/- mice developed experimental autoimmune meningitis (EAM). EAM is a fatal, acute disease characterized by simultaneous weakness in all limbs, photophobia, irritability, and spatial disorientation. Histologically, EAM consisted of an infiltrate of myeloid, monocytic, and lymphocytic leukocytes within the leptomeninges. In contrast, the brain parenchyma was unaffected. EAM was mediated by CD4+ T cells since CD4 depletion prevented the disease. Upon rechallenge, mice in which EAM was prevented by CD4+ cell depletion developed EAE not EAM. Therefore, the presence or absence of CD28 determines the initial phenotype of the immune response to MOG. EAM, which develops in the absence of CD28, is a unique experimental model for immune-mediated aseptic meningitis.     

Staphylococcal enterotoxin B-induced activation and concomitant resistance to cell death in CD28-deficient HLA-DQ8 transgenic mice

HLA class II molecules present superantigens more efficiently than their murine counterpart. Therefore, transgenic mice expressing HLA-DQ8 with and without CD28 were used to address the role of CD28 in staphylococcal enterotoxin B (SEB)-driven immune responses. SEB-induced in vitro proliferation of naive DQ8.CD28(-/-) splenocytes was comparable to DQ8.CD28(+/+) cells, and was several fold higher than that of C57BL/10 and BALB/c splenocytes. SEB-activated, naive DQ8.CD28(-/-) cells in vitro produced significantly less IL-2, IL-4 and IL-10 than DQ8.CD28(+/+) cells, while IFN-gamma and IL-6 production was comparable. SEB-induced in vivo expansion of CD4(+) T cells and, to a greater extent, CD8(+) T cells was compromised in DQ8.CD28(-/-) mice, indicating that SEB-induced proliferation of CD8(+) T cells is more dependent on CD28 co-stimulation. Upon re-stimulation, SEB-primed CD28(+/+) T cells failed to proliferate but were capable of producing cytokines. Conversely, CD28(-/-) T cells were capable of proliferation, but not cytokine production. SEB-primed CD28-deficient cells produced significantly less nitric oxide when compared to CD28-sufficient cells following re-stimulation with SEB. CD28(+/+) and not CD28(-/-) mice were highly susceptible to SEB-induced lethal shock characterized by significantly elevated serum IFN-gamma. Thus, (i) efficient presentation of SEB by HLA-DQ8 circumvents co-stimulation through CD28, (ii) unique CD28-derived signals are mandatory for generation of certain effector functions, and (iii) absence of CD28-derived signals confers resistance to activation-induced cell death and protects mice from SEB-induced shock.     

Increased susceptibility to airway responses in CD40-deficient mice

The interaction between CD40 and its ligand (CD154) is crucial for IL-12 production and effective humoral immunity such as IgE production. Although the interaction seems to play a crucial role in asthmatic inflammation, previous studies investigating the role of the CD40 and CD154 interaction in experimental animal models of asthma are complicated due to multistep reactions in developing asthma. Here, in order to investigate the role of CD40 in the effector phase in the development of airway responses, we used CD40-deficient mice backcrossed with mice transgenic for an ovalbumin (OVA)-specific TCR (TCRtg). Using intranasal OVA administration followed by aerosol inhalation of OVA, greater airway hyperreactivity and eosinophilia in bronchoalveolar lavage fluid (BALF) were observed in CD40-deficient mice backcrossed with TCRtg mice (CD40-/-/ TCRtg mice), compared with control littermates (CD40+/+/ TCRtg mice). CD4+ helper T cell subset analysis of lung draining lymph nodes revealed that the Th1 component was significantly decreased in CD40-/-/ TCRtg mice. Airway hyperreactivity and airway eosinophilia significantly correlated with the predomination of Th2 cells. Cytokine measurements in BALF also showed decreased IL-12 and the predominance of Th2 cells in CD40-/-/ TCRtg mice. These results suggest that CD40 may play a protective role in developing asthma in the phase after establishing specific memory T cells through the regulation of the balance between Th1 and Th2 cells presumably via induction of IL-12.     

The immune response to Plasmodium chabaudi malaria in interleukin-4-deficient mice

Interleukin(IL)-4 promotes the development of T helper (T_H)2 cells, induces immunoglobulin class switching to IgG1 and is thought to be essential for switching to IgE. During a primary infection with the erythrocytic stages of Plasmodium chabaudi chabaudi, T_H1 and T_H2 cells specific for the parasite appear sequentially as infection progresses. To dissect the possible role of T_H2 responses at the later stages of infection, mice with a targetted disruption of the IL-4 gene were infected with P. chabaudi. IL-4-deficient mice were able to control and clear a primary infection, although recrudescent parasitemias were significantly higher in these mice compared with wild-type littermates, demonstrating that IL-4 per se is not required for parasite elimination. To evaluate the actual impairment of T_H2 functions in the absence of IL-4 in vivo during an infection with P. chabaudi, the cellular and humoral responses to the parasite generated in vitro and in vivo were compared in the two types of mice. Our data indicate that in vitro T_H1 responses and ex vivo IL-12 mRNA levels were sustained in the IL-4-deficient mice compared with wild-type littermates. Correspondingly, T_H2-associated cytokine mRNA such as IL-5 and IL-6, but not IL-10, were reduced early in infection in the deficient animals. However, these cytokines were expressed at comparable levels at the later stages of infection in both types of mice. Reflecting these differences in T_H function, IgG1 responses were decreased in vitro and delayed in vivo, whereas IgG2a and IgG2b responses appeared earlier in vivo in the deficient mice. Strikingly, IgE secretion was not blocked in vivo in the deficient mice; the onset of the synthesis of IgE mRNA was delayed during infection and the amount of circulating IgE was five times lower than in the wild-type littermates after 5 weeks of infection. All these impairments of T_H-related activities were insufficient to affect parasite clearance in the deficient mice, probably due to the fact that such activities were only delayed and could take place normally at the later stages of infection.     

Immunity to infection in IL-17-deficient mice and humans

Mice with defective IL-17 immunity display a broad vulnerability to various infectious agents at diverse mucocutaneous surfaces. In humans, the study of patients with various primary immunodeficiencies, including autosomal dominant hyper-IgE syndrome caused by dominant-negative STAT3 mutations and autosomal recessive autoimmune polyendocrinopathy syndrome type 1 caused by null mutations in AIRE, has suggested that IL-17A, IL-17F and/or IL-22 are essential for mucocutaneous immunity to Candida albicans. This hypothesis was confirmed by the identification of rare patients with chronic mucocutaneous candidiasis (CMC) due to autosomal recessive IL-17RA deficiency and autosomal dominant IL-17F deficiency. Heterozygosity for gain-of-function mutations in STAT1 in additional patients with CMC was recently shown to inhibit the development of IL-17 T cells. Although the infectious phenotype of patients with CMC and inborn errors of IL-17 immunity remains to be finely delineated, it appears that human IL-17A and IL-17F display redundancy for protective immunity in natural conditions that is not seen in their mouse orthologs in experimental conditions.     

Toll-like receptor 2-deficient mice succumb to Mycobacterium tuberculosis infection

Recognition of Mycobacterium tuberculosis by the innate immune system is essential in the development of an adaptive immune response. Mycobacterial cell wall components activate macrophages through Toll-like receptor (TLR) 2, suggesting that this innate immune receptor plays a role in the host response to M. tuberculosis infection. After aerosol infection with either 100 or 500 live mycobacteria, TLR2-deficient mice display reduced bacterial clearance, a defective granulomatous response, and develop chronic pneumonia. Analysis of pulmonary immune responses in TLR2-deficient mice after 500 mycobacterial aerosol challenge showed increased levels of interferon-gamma, tumor necrosis factor-alpha, and interleukin-12p40 as well as increased numbers of CD4(+) and CD8(+) cells. Furthermore, TLR2-deficient mice mounted elevated Ag-specific type 1 T-cell responses that were not protective because all deficient mice succumb to infection within 5 months. Taken together, the data suggests that TLR2 may function as a regulator of inflammation, and in its absence an exaggerated immune inflammatory response develops.     

Altered airway responsiveness in CD38-deficient mice

Cyclic ADP-ribose (cADPR) mobilizes calcium from intracellular stores and contributes to agonist-induced intracellular calcium elevation in airway smooth muscle (ASM). In this study we determined the functional role of CD38/cADPR signaling in the regulation of airway tone using CD38 deficient (cd38(-/-)) mice. The responsiveness to different doses of methacholine, as determined by changes in lung resistance and dynamic compliance, was significantly (P < or = 0.05) lower in cd38(-/-) mice compared with wild-type controls. To determine the mechanism responsible for the reduced responsiveness, we measured the intracellular calcium responses to contractile agonists in ASM cells. In ASM cells isolated from cd38(-/-) mice, the intracellular calcium responses to acetylcholine and endothelin-1 were significantly lower than in controls. Pretreatment of ASM cells with a cADPR antagonist resulted in attenuated intracellular calcium responses to endothelin-1 in cells isolated from wild-type mice, but not in those isolated from the cd38(-/-) mice. Very low cADPR levels and no detectable ADP-ribosyl cyclase activity were observed in lung tissue from cd38(-/-) mice, suggesting that CD38 is a critical source for cADPR synthesis. The results of the present study demonstrate that CD38/cADPR contributes to airway smooth muscle tone and responsiveness through its effects on agonist-induced elevation of intracellular calcium in ASM cells.     

Role of CD44 in activation-induced cell death: CD44-deficient mice exhibit enhanced T cell response to conventional and superantigens

T cells upon activation are known to up-regulate CD44 expression. However, the precise function of CD44 on activated T cells is not clear. In this report, we demonstrate that signaling through CD44 plays an important role in activation-induced cell death (AICD). CD44 knockout (KO) mice had an elevated in vivo primary and in vitro secondary response to challenge with conalbumin, anti-CD3 mAb and staphylococcal enterotoxin A (SEA), which correlated with reduced AICD when compared to CD44 wild-type mice. In addition, CD44 KO mice exhibited increased delayed-type hypersensitivity response to dinitrofluorobenzene. In a model examining in vitro AICD, splenocytes from CD44 KO mice showed resistance to TCR-mediated apoptosis when compared to splenocytes from CD44 wild-type mice. In addition, signaling through CD44 led to increased apoptosis in TCR-activated but not resting T cells from CD44 wild-type mice without affecting Fas expression. Injection of SEA into mice deficient in CD44 and Fas (CD44 KO/lpr) led to an increased primary response when compared to mice that expressed CD44 but not Fas (CD44 WT/lpr), suggesting that the enhanced response to SEA was dependent on CD44 but not Fas expression. Administration of anti-CD44 mAb into CD44 wild-type mice caused a significant decrease in antigen-specific T cell response. Together, these data implicate CD44 as an important regulator of AICD in T cells. Furthermore, targeting CD44 in vivo may constitute a novel approach to induce apoptosis in activated T cells, and therefore to treat autoimmune diseases, allograft rejection and graft versus host disease.     

Contribution of interleukin-12 (IL-12) and the CD28/B7 and CD40/CD40 ligand pathways to the development of a pathological T-cell response in IL-10-deficient mice

The ability of interleukin-10 (IL-10) to suppress accessory cell functions required for optimal T-cell activation makes it an important inhibitor of cell-mediated immunity. Thus, after infection with the protozoan parasite Toxoplasma gondii, IL-10 knockout (KO) mice develop a CD4(+)-T-cell-dependent shock-like reaction with high levels of IL-12 and gamma interferon (IFN-gamma) in serum, leading to death of mice during the acute phase of infection. Previous studies from this laboratory have shown that simultaneous blockade of CD28 and CD40 can prevent this lethal reaction by inhibiting the production of IFN-gamma. However, the blockade of costimulation did not affect systemic levels of IL-12. To better understand the relationship between IL-12 and the CD28 and CD40 pathways in mediating immune hyperactivity, antagonists of these factors were used to determine their effects on the development of a pathological T-cell response in IL-10 KO mice. Blockade of IL-12 or the CD28/B7 interaction alone did not affect survival; however, the combined blockade of both pathways resulted in decreased production of IFN-gamma and the survival of IL-10 KO mice. To assess the role of the two ligands for CD28, B7.1 and B7.2, IL-10 KO mice were treated with alphaIL-12 plus alphaB7.1 or alphaB7.2 or the combination of all three antibodies. These studies revealed that blockade of both B7 molecules is required for decreased production of IFN-gamma and survival of infected IL-10 KO mice, suggesting that B7.1 and B7.2 can contribute to the lethal shock-like reaction in IL-10 KO mice. In contrast, neutralization of IL-12 and blockade of the CD40/CD40 ligand (CD40L) interaction in vivo did not alter the production of IFN-gamma and only resulted in a small delay in time to death of mice. Together, these data suggest that the CD28/B7 interaction has a central role in the development of a pathological T-cell response in IL-10 KO mice, which is distinct from the role of the CD40/CD40L and IL-12 pathways.     

The host response in secondary hydatidosis of mice

Summary The infection of young albino mice by the intraperitoneal injection of viable hydatid protoscolices provided a suitable model for the investigation of the factors controlling the subsequent host immune response. Antibodies detectable by haemagglutination and complement fixation appeared around the 14th week after infection. Increases in antibody titres were related to the potency of the antigenic impulse(s) produced by the healthy germinal membrane and viable protoscolices. Our results suggest that qualitative and quantitative variations in excretory, secretory and somatic antigens, occurring during the different phases of parasite development, incite the production of a multitude of antibodies, only a few of which are detectable by conventional methods.     

Pneumocystis infection in an immunocompetent host can promote collateral sensitization to respiratory antigens

Infection with the opportunistic fungal pathogen Pneumocystis is assumed to pass without persistent pathology in immunocompetent hosts. However, when immunocompetent BALB/c mice were inoculated with Pneumocystis, a vigorous Th2-like pulmonary inflammation ensued and peaked at 14 days postinfection. This coincided with a 10-fold increase in the number of antigen-presenting cells (APCs) in the lung, and these cells were capable of presenting antigen in vitro, as well as greater uptake of antigen in vivo. When mice were presented with exogenous antigen at the 14-day time point of the infection, they developed respiratory sensitization to that antigen, in the form of increased airway hyperresponsiveness upon a later challenge, whereas mice not infected but presented with antigen did not. Like other forms of collateral sensitization, this response was dependent on interleukin-4 receptor signaling. This ability to facilitate sensitization to exogenous antigen has been previously reported for other infectious disease agents; however, Pneumocystis appears to be uniquely capable in this respect, as a single intranasal dose without added adjuvant, when it was administered at the appropriate time, was sufficient to initiate sensitization. Pneumocystis infection probably occurs in most humans during the first few years of life, and in the vast majority of cases, it fails to cause any overt direct pathology. However, as we show here, Pneumocystis can be an agent of comorbidity at this time by facilitating respiratory sensitization that may relate to the later development or exacerbation of obstructive airway disease.     

Initiation of the host response against murine gammaherpesvirus infection in immunocompetent mice

Murine gammaherpesvirus 68 (gammaHV-68) provides a useful model for understanding the initiation of the host response against the gammaherpesviruses. Its value as a model for such studies lies in large part with the inherent difficulties in investigating human responses against EBV and HHV-8 during the first few days following infection. While studies aimed at defining the initiation of gammaHV-68 infection are far from complete, an unexpected trend in this early host response has already emerged. Despite viral replication and the beginnings of viral latency at the site of infection during the first few days following infection, the early host response seems surprisingly inadequate. For example, the pro-inflammatory response is quite limited, and with the exception of the type I interferons, it is not at all clear what innate responses are necessary to provide protection from acute infection. This confusion results from the lack of any significant effect on acute viral replication in several strains of mice which have been made genetically deficient in the expression of particular pro-inflammatory molecules. It is likely that these unexpected results reflect the ability of gammaherpesviruses to carefully control the initial response so that they are efficacious pathogens even in immunocompetent hosts.     

Mycobacterium avium infection in CD14-deficient mice fails to substantiate a significant role for CD14 in antimycobacterial protection or granulomatous inflammation

CD14 is a pattern-recognition receptor implicated in the inflammatory response to microbial components such as lipopolysaccharide, peptidoglycan and lipoarabinomannan. In this work, we made use of CD14-deficient (CD14-/-) mice to evaluate the relative importance of CD14 in response to infection with viable, intact cells of Mycobacterium avium in vitro and in vivo. Following co-incubation of either bone marrow-derived macrophages (Mphi) or thioglycollate-elicited peritoneal Mphi from CD14-/- mice with viable M. avium, tumour necrosis factor (TNF) production was significantly reduced and delayed compared to TNF secretion by infected CD14+/+ Mphi. However, following intravenous infection with a M. avium strain of either high virulence (TMC724) or intermediate virulence (SE01), there was no difference in the bacterial loads of lungs, livers or spleens at 3, 5 and 8 weeks postinfection in CD14-/- mice when compared with syngeneic CD14+/+ mice. At these time-points, TNF and interferon-gamma (IFN-gamma) mRNA expression in the liver was similar in infected CD14+/+ and CD14-/- mice, and granuloma formation and expression of inducible nitric oxide synthase within granuloma Mphi was the same in both mouse groups. In conclusion, although the absence of CD14 results in significantly reduced and delayed TNF production in response to stimulation with M. avium in vitro, there is no evidence that CD14 plays a significant role in either the antibacterial defence or the chronic granulomatous reaction to M. avium infection in vivo.     

Listeria monocytogenes infection in caspase-11-deficient mice

Caspase-11 (Cas11) is a cysteine protease involved in programmed cell death and cytokine maturation. Through activation of Cas1 (interleukin-1beta [IL-1beta]-converting enzyme), Cas11 is directly involved in the maturation of IL-1beta and IL-18. Apoptosis is mediated through Cas3. Given the role of apoptosis and cytokine signaling during the innate immune response in intracellular infection, we examined Cas11-deficient (Cas11(-/-)) mice during infection with Listeria monocytogenes. Cas11(-/-) and wild-type C57BL/6 mice were equally susceptible to intravenous infection with L. monocytogenes, resulting in similar bacterial burdens in tissue and similar survival rates. By contrast, enhanced susceptibility was observed in control mice on a mixed genetic 129/C57BL/DBA2 background. Cas11(-/-) and wild-type mice infected with Listeria had similar hepatic microabscess formation in terms of histologic appearance, size, and number. Apoptosis of L. monocytogenes-infected hepatocytes in vivo and in vitro in primary culture was not altered by the absence of Cas11. Serum IL-18 and IL-1beta levels were similar in Cas11(-/-) mice and controls. Endotoxin (lipopolysaccharide [LPS])-challenged Cas11(-/-) mice were deficient in the production of gamma interferon. IL-1beta responses in Cas11(-/-) were normal with intravenous administration of LPS but decreased with intraperitoneal administration. Our findings suggest that Cas11 deficiency does not impair the immune response to infection with L. monocytogenes. Apoptosis and maturation of IL-18 and IL-1beta were normal despite Cas11 deficiency. LPS-induced proinflammatory pathways are altered by the absence of Cas11. While Cas11-mediated Cas1 and Cas3 activation is crucial for cytokine maturation and apoptosis during inflammation, alternative pathways allow normal inflammatory and apoptotic responses during infection with L. monocytogenes.     

Impaired IgE response in SWAP-70-deficient mice

Protein SWAP-70 was initially isolated from nuclei of activated B cells and was implicated in the immunoglobulin class switch process. After B cell activation the protein translocates from the cytoplasm to the nucleus, and may serve to signal nuclear processes. We have generated mice deficient in SWAP-70 and found three main differences when compared to wild-type mice: (i) their B lymphocytes are two- to threefold more sensitive to gamma-irradiation than B cells of wild type; (ii) SWAP-70-deficient mice developed autoantibodies at a much higher frequency; and (iii) the CD40 signaling pathway is compromised in the mutant mice. CD40-dependent switching to the IgE isotype is reduced five- to eightfold in vitro. In SWAP-70-deficient mice, IgE levels prior to immunization were six- to sevenfold lower than in wild-type mice, and after immunization three- to fourfold lower. CD40-induced proliferation was transiently increased in the mutant. LPS-induced switching to other isotypes, however, and LPS-induced proliferation were normal. We propose that SWAP-70 serves a specific role in the CD40 signaling pathway, in particular in the IgE response.