E7 oncoprotein of human papillomavirus type 16 expressed constitutively in the epidermis has no effect on E7-specific B- or Th-repertoires or on the immune response induced or sustained after immunization with E7 protein

A line of FVB (H-2(q)) mice transgenic for the E6/E7 open reading frames of Human Papillomavirus type 16 driven from the alpha-A crystallin promoter expresses E7 mRNA in lens and skin epithelium. E7 protein is detectable in adult skin, coinciding with the development or inflammatory skin disease, which progresses to papillomata and squamous carcinomata in some mice. By examining the outcome of parenteral immunization with E7 protein, we sought to determine whether endogenous expression of E7 in skin had induced a preexisting immune outcome, i.e., specific immunity or tolerance, or whether the mice remain naive (''ignorant'') to E7. Our data show that the antibody response to defined E7 B-epitopes, the proliferative response to Th epitopes, and the delayed-type hypersensitivity (DTH) response to whole E7 did not differ between groups or young and old E6/E7 transgenic mice (likely having different degrees of lifetime exposure to E7 protein) or between E6/E7-transgenic and nontransgenic parental strain control mice. Although an E7-specific CTL response could not be induced in the H-2(q) background of these mice, incorporation of a D-b allele into the genome allowed comparison of D-b-restricted CTL responses in E6/E7 transgenic and nontransgenic mice. Experiments indicated that the E7-immunization-induced CTL response did not differ significantly between E6/E7 transgenic and nontransgenic mice. We interpret these results to indicate that in spite of expression of E7 protein in adult skin, E6/E7 transgenic mice remain immunologically naive (ignorant) of E7 epitopes presented by immunization. (C) 1997 Academic Press.

Histopathology, humoral and cellular immune response in the murine model of Leishmania (Viannia) shawi

Leishmania (Viannia) shawl was recently characterized and few studies concerning modifications in cellular and humoral immune responses in experimental leishmaniasis have been conducted. In this work, immunopathological changes induced by L. shawl in chronically infected BALB/c mice were investigated. Infected BALB/c mice developed increased lesion size associated with strong inflammatory infiltrate diffusely distributed in the dermis, with highly infected macrophages. The humoral immune response was predominantly directed toward the IgG1 isotype. The functional activity of CD4(+) and CD8(+) T cells showed significantly increased TNF-alpha mRNA levels associated with reduced IFN-gamma expression by CD4(+) T cells and the double negative (dn) CD4CD8 cell subset. High IL-4 levels expressed by CD8(+) T cells and dnCD4CD8 and TGF-beta by CD4(+) and CD8(+) T cells were detected, while IL-10 was highly expressed by all three cell subpopulations. Taken together, these results show an evident imbalance between TNF-alpha and IFN-gamma that is unfavorable to amastigote replication control. Furthermore, L. shawi seems to regulate different cell populations to express deactivating cytokines to avoid its own destruction. This study indicates BALB/c mice as a potentially good experimental model for further studies on American cutaneous leishmaniosis caused by L. shawi. (C) 2010 Elsevier Ireland Ltd. All rights reserved.

Characterization of cytotoxic immune response in skin and mucosal lesions of paracoccidioidomycosis

Background: CD8+ T cells and natural killer (NK) cells are involved in the immune response against some pathogens. For this purpose, we investigated the in situ paracoccidioidomycosis (PCM) immune response addressing the participation of NK cells, CD8+ T cells, perforin and granzyme B expression. Methods: Sixty biopsies of PCM skin and mucosa were classified according to the presence of compact granulomas (G1), poorly organized granulomas (G2) and both kinds in the same lesion (G3). CD8+ T cells, NK cells, perforin and granzyme B were showed by immunohistochemistry. Results: CD8+ T cells were increased over NK cells in cutaneous G1 and G2 lesions. There was no difference regarding such cells in G3 lesions, although they were abundant in such lesions. In mucosa, CD8+ T cells were increased in number over NK cells in all groups. Granzyme B in skin increased in G2 and G3. The number of granzyme did not differ in mucosal lesions in the three groups. Conclusions: CD8+ T cells and NK cells play a role in PCM cutaneous and mucosal lesions. The predominance of CD8+ T cells over NK cells may represent an effective response against the fungi. Moreover, the high number of granzyme B expressing cells corroborates this possibility.

Immunization with pVAXhsp65 Decreases Inflammation and Modulates Immune Response in Experimental Encephalomyelitis

Background: A DNA vaccine (pVAXhsp65) containing the gene of a heat-shock protein (hsp65) from Mycobacterium leprae showed high immunogenicity and protective efficacy against tuberculosis in BALB/c mice. A possible deleterious effect related to autoimmunity needed to be tested because hsp65 is highly homologous to the correspondent mammalian protein. In this investigation we tested the effect of a previous immunization with DNAhsp65 in the development of experimental autoimmune encephalomyelitis (EAE), a rat model of multiple sclerosis. Methods: Female Lewis rats were immunized with 3 pVAXhsp65 doses by intramuscular route. Fifteen days after the last DNA dose the animals were evaluated for specific immunity or submitted to induction of EAE. Animals were evaluated daily for weight loss and clinical score, and euthanized during the recovery phase to assess the immune response and inflammatory infiltration at the central nervous system. Results: Immunization with pVAXhsp65 induced a specific immune response characterized by production of IgG(2b) anti-hsp65 antibodies and IFN-gamma secretion. Previous immunization with pVAXhsp65 did not change EAE clinical manifestations (weight and clinical score). However, the vaccine clearly decreased brain and lumbar spinal cord inflammation. In addition, it downmodulated IFN-gamma and IL-10 production by peripheral lymphoid organs. Conclusion: Our data demonstrated that this vaccine does not trigger a deleterious effect on EAE development and also points to a potential protective effect. Copyright (C) 2010 S. Karger AG, Basel

Reappraisal of the immunopathogenesis of disseminated leishmaniasis: in situ and systemic immune response

Disseminated leishmaniasis (DL) is an emerging form of Leishmania braziliensis infection characterised by multiple cutaneous lesions on different parts of the body and a high rate of mucosal involvement. Systemic production of TNF alpha and IFN-gamma in DL patients is lower than in cutaneous leishmaniasis (CL) caused by L braziliensis, which may account for parasite dissemination due to the decreased ability to control parasite growth. In this study, the systemic and in situ immune response of DL and CL patients was characterised through evaluation of chemokine and cytokine production. In situ evaluation showed similar production of IFN gamma, TNF alpha, IL-10, transforming growth factor-beta (TGF beta), chemokine (C-C motif) ligand 2 (CCL2), CCL3, CCL11 and chemokine (C-X-C motif) ligand 10 (CXCL10) in papular and ulcerative lesions from DL as well as in ulcerated lesions from CL. Serum levels of CXCL9, a chemokine that attracts 1-cells, was higher in serum from DL than from CL These data indicate that a decrease in the type 1 immune response in peripheral blood of DL patients is due to attraction of Leishmania antigen-activated T-cells to the multiple cutaneous lesions. This may account for the absence of or few parasites in the lesions and for the development of ulcers similar to those observed in CL (C) 2011 Royal Society of Tropical Medicine and Hygiene. Published by Elsevier Ltd. All rights reserved.

Host genetic background affects regulatory T-cell activity that influences the magnitude of cellular immune response against Mycobacterium tuberculosis

Using two mouse strains with different abilities to generate interferon (IFN)-gamma production after Mycobacterium tuberculosis infection, we tested the hypothesis that the frequency and activity of regulatory T (Treg) cells are influenced by genetic background. Our results demonstrated that the suppressive activity of spleen Treg cells from infected or uninfected BALB/c mice was enhanced, inhibiting IFN-gamma and interleukin (IL)-2 production. Infected C57BL/6 mice exhibited a decrease in the frequency of lung Treg cells and an increased ratio CD4(+):CD4(+)Foxp3(+) cells compared with infected BALB/c mice and uninfected C57BL/6 mice. Moreover, infected C57BL/6 mice also had a decrease in the immunosuppressive capacity of spleen Treg cells, higher lung IFN-gamma and IL-17 production, and restricted the infection better than BALB/c mice. Adoptive transfer of BALB/c Treg cells into BALB/c mice induced an increase in bacterial colony-forming unit (CFU) counts. Furthermore, BALB/c mice treated with anti-CD25 antibody exhibited lung CFU counts significantly lower than mice treated with irrelevant antibody. Our results show that in BALB/c mice, the Treg cells have a stronger influence than that in C57BL/6 mice. These data suggest that BALB/c and C57BL/6 mice may use some different mechanisms to control M. tuberculosis infection. Therefore, the role of Treg cells should be explored during the development of immune modulators, both from the perspective of the pathogen and the host. Immunology and Cell Biology (2011) 89, 526-534; doi:10.1038/icb.2010.116; published online 19 October 2010

Local profile of cytokines and nitric oxide in patients with bacterial vaginosis and cervical intraepithelial neoplasia

Objective: To evaluate the local immune response in patients with bacterial vaginosis (BV) and cervical intraepithelial neoplasia (CIN), as assessed by cytokine and nitric oxide (NO) concentrations. Study design: Patients attending for routine gynaecological examination were prospectively enrolled in groups: BV (n = 25) diagnosed by clinical criteria, CIN graded I to III (n = 35, 6 CIN 1, 8 CIN 11 and 21 CIN 111) by histological analysis, and controls (n = 15) without clinical and cytological findings. Randomly selected patients within CIN group at grades 11 or III (n = 15) were re-evaluated at 60 days after surgical treatment. Endocervical (EC) and vaginal secretion samples were collected by cytobrush and the levels of cytokines (ELISA) and NO metabolite (Griess reaction) were assayed. Results: NO was assessed in all subjects, and cytokines in all controls, 15 BV and 30 CIN patients. Interleukin-6 (IL-6), interleukin-8 (IL-8), interleukin-10 (IL-10) and nitrite levels were higher in EC than in vaginal secretions in BV and CIN groups. In CIN group, IL-8, IL-10 and nitrite concentrations were greater in EC and/or vaginal secretions than in BV or controls. Surgical treatment reduced IL-8 levels in EC and vaginal secretions. Conclusion: A similar local immune profile was found in BV and CIN groups. The increased local production of IL-8, IL-10 and NO in CIN suggests a role for these mediators in the immune response against tumour or tumour development. (c) 2007 Elsevier Ireland Ltd. All rights reserved.

Mutations involved in Aicardi-Goutieres syndrome implicate SAMHD1 as regulator of the innate immune response

Aicardi-Goutieres syndrome is a mendelian mimic of congenital infection and also shows overlap with systemic lupus erythematosus at both a clinical and biochemical level. The recent identification of mutations in TREX1 and genes encoding the RNASEH2 complex and studies of the function of TREX1 in DNA metabolism have defined a previously unknown mechanism for the initiation of autoimmunity by interferon-stimulatory nucleic acid. Here we describe mutations in SAMHD1 as the cause of AGS at the AGS5 locus and present data to show that SAMHD1 may act as a negative regulator of the cell-intrinsic antiviral response.

Oleic acid modulation of the immune response in wound healing: A new approach for skin repair

Injury triggers inflammatory responses and tissue repair. Several treatments are currently in use to accelerate healing: however, more efficient formulations are still needed for specific injuries. Since unsaturated fatty acids modulate immune responses, we aimed to evaluate their therapeutic effects on wound healing. Skin wounds were induced in BALB/c mice and treated for 5 days with n-3, n-9 fatty acids or vehicle (control). n-9 treated mice presented smaller wounds than control and n-3 at 120 h post-surgery (p.s.). Collagen III mRNA,TIMP1 and MMP9 were significantly elevated in n-9 group compared to n-3 or vehicle at 120 h p.s. Among the inflammatory mediators studied we found that IL-10, TNF-alpha and IL-17 were also higher in n-9 treated group compared to n-3 or vehicle at 120 h p.s. Interestingly, COX2 had decreased expression on wound tissue treated with n-9. Inflammatory infiltrate analysis revealed diminished frequency of CD4(+), CD8(+) and CD11b(+) cells in n-9 wounds at 24 and 120 h p.s., which was not related to cell death, since in vitro apoptosis experiments did not show any cell damage after fatty acids administration. These results suggested that unsaturated fatty acids, specifically n-9, modulate the inflammation in the wound and enhance reparative response in vivo. n-9 may be a useful tool in the treatment of cutaneous wounds. (C) 2010 Elsevier GmbH. All rights reserved.

A DNA vaccine candidate encoding the structural prM/E proteins elicits a strong immune response and protects mice against dengue-4 virus infection

A DNA vaccine expressing dengue-4 virus premembrane (prM) and envelope (E) genes was produced by inserting these genes into a mammalian expression plasmid (pCI). Following a thorough screening, including confirmation of protein expression in vitro, a recombinant clone expressing these genes was selected and used to immunize BALB/c mice. After 3 immunizations all the animals produced detectable levels of neutralizing antibodies against dengue-4 virus. The cytokines levels and T cell proliferation, detected ex vivo from the spleen of the immunized mice, showed that our construction induced substantial immune stimulation after three doses. Even though the antibody levels, induced by our DNA vaccine, were lower than those obtained in mice immunized with dengue-4 virus the levels of protection were high with this vaccine. This observation is further supported by the fact that 80% of the vaccine immunized group was protected against lethal challenge. In conclusion, we developed a DNA vaccine employing the genes of the prM and E proteins from dengue-4 virus that protects mice against this virus. (C) 2010 Elsevier Ltd. All rights reserved.

Chemokines and chemokine receptors coordinate the inflammatory immune response in human cutaneous leishmaniasis

Cutaneous leishmaniasis (CL) includes different clinical manifestations displaying diverse intensities of dermal Inflammatory infiltrate Diffuse CL (DCL) cases are hyporesponsive and lesions show very few lymphocytes and a predominance of macrophages In contrast localized CL (LCL) cases are responsive to leishmanial antigen and lesions exhibit granulocytes and mononuclear cell infiltration in the early phases changing to a pattern with numerous lymphocytes and macrophages later in the lesion Therefore different chemokines may affect the predominance of cell infiltration in distinct clinical manifestations In lesions from LCL patients we examined by flow cytometry the presence of different chemokines and their receptors in T cells and we verified a higher expression of CXCR3 in the early stages of LCL (less than 30 days of infection) and a higher expression of CCR4 in the late stages of disease (more than 60 days of infection) We also observed a higher frequency of T cells producing IL-10 in the late stage of LCL Using immunohistochemistry we observed a higher expression of CCL7 CCL17 in lesions from late LCL as well as CCR4 suggesting a preferential recruitment of regulatory T cells in the late LCL Comparing lesions from LCL and DCL patients we observed a higher frequency of CCL7 in DCL lesions These results point out the Importance of the chemokines defining the different types of cells recruited to the site of the infection which could be related to the outcome of infection as well as the clinical form observed (C) 2010 American Society for Histocompatibility and Immunogenetics Published by Elsevier Inc All rights reserved

Immune response to Bacteroides forsythus in a murine model

A murine skin abscess model was used to study the immune response to an acute infection with Bacteroides forsythus. BALB/c mice were given subcutaneous injections of either viable or heat-killed B. forsythus, while a third sham-immunized control group received phosphate-buffered saline. Weights and lesion sizes were measured. Blood was collected from the heart and specific antibodies to B. forsythus measured by an ELISA. Swabs taken from the lesions and also from pooled blood were cultured anaerobically for viable B. forsythus. Viable B. forsythus-induced lesions reached maximum size at day 7. B. forsythus cells were recovered from lesions up to day 4 although none were cultured from blood samples. Heat-killed bacteria induced much smaller lesions. Serum antibody levels increased during the 9-day study period, being significantly higher in mice injected with viable compared with heat-killed B. forsythus. Antibody levels in sham control mice were significantly lower than those seen in the other two groups. These results showed that a subcutaneous injection of viable cells of B. forsythus elicited a pronounced abscess formation and induce higher levels of specific antibodies compared with that produced by an injection of dead bacteria. This suggests that, as with other periodontopathic organisms, this mouse model can be used to study the immune response to B. forsythus.

The effects of interleukin-10 depletion in vivo on the immune response to Porphyromonas gingivalis in a murine model

Background: The immune response to Porphyromonas gingivalis in the mouse abscess model is known to be dependent upon CD4 T-cell activation and the regulatory role of cytokines. The role of interleukin-10 (IL-10) in this mouse model was examined in vivo. Methods: One-week-old, female BALB/c mice were divided into 4 groups. Groups 1 and 2 were given intraperitoneal (ip) injections of phosphate buffered saline (PBS) weekly for 5 weeks. Group 3 was given an ip injection of rat immunoglobulin. Group 4 was injected with rat anti-IL-10 antibodies. At week 6, group 1 was sham-immunized with PBS, and groups 2, 3, and 4 were injected with P gingivalis lipopolysaccharide (Pg-LPS) weekly for 2 weeks. One week after the final immunization, delayed-type hypersensitivity (DTH) was assessed by footpad swelling to Pg-LPS. The level of serum antibodies to Pg-LPS and IFN-gamma (IFN-gamma) was determined by enzyme-linked immunosorbent assay. Dorsal abscess formation induced by the injection of viable P gingivalis was examined daily for 30 days. Results: The footpad swelling of the anti-IL-10-treated group (group 4) was significantly higher than that of groups 1 to 3. Similarly, the serum IFN-gamma level in group 4 was much higher than that of the other experimental groups. There was no significant difference in serum IgG antibodies to Pg-LPS in any of the experimental groups. However, the level of IgM antibodies in group 4 mice was significantly lower than that in groups 2 and 3. In addition, serum IgG1 was suppressed in group 4 mice, while IgG2a antibodies were raised. However, there was no difference observed between the levels of IgG2b and IgG3 antibodies in any group of mice. The lesions in sham-immunized mice (group 1) persisted for 30 days, and those in group 2 and 3 were undetected by day 18 and 20, respectively. In sharp contrast, lesions in group 4 had healed completely by day 13. Conclusions: This study has shown that IL-10 depletion in vivo in P gingivalis LPS-induced immune response in mice led to an elevated DTH response, an increase in serum IFN-gamma levels, and raised levels of IgG and IgG2a antibodies. Treatment with anti-IL-10 antibodies resulted in suppressed IgG I and IgM responses and a more rapid healing of abscesses than in non-IL-10-depleted mice. These results suggest that IL-10 depletion in Pg-LPS-induced immune response in mice may lead to a Th1-like immune response and provide strong protection against a subsequent challenge with live P gingivalis in an abscess model.

The role of CD4(+) cells in vivo on the induction of the immune response to Porphyromonas gingivalis in mice

Background: It has previously been suggested that CD4(+) T cells play a pivotal role in regulating the immune response to periodontal pathogens. The aim of the present study therefore was to determine delayed type hypersensitivity (DTH), spleen cell proliferation, serum and splenic anti-Porphyromonas gingivalis antibody levels, and lesion sizes following challenge with viable P. gingiualis in CD4-depleted BALB/c mice immunized with P. gingiualis outer membrane proteins (OMP). Methods: Four groups of BALB/c mice were used. Groups 1 and 2 were injected intraperitoneally (ip) with saline for 3 consecutive days and then weekly throughout the experiment. Groups 3 and 4 were injected ip with rat immunoglobulin and a monoclonal rat anti-mouse CD4 antibody, respectively. Two days later, group 1 mice were injected ip with saline only, while all the other groups were immunized ip with P. gingiualis OMP weekly for 3 weeks. One week later following the last immunization of OMP, 3 separate experiments were conducted to determine: 1) the DTH response to P. gingiualis OMP by measuring footpad swelling; 2) the levels of antibodies to P. gingiualis in serum samples and spleen cell cultures using an enzyme-linked immunosorbent assay, as well as spleen cell proliferation after stimulation with OMP; and 3) the lesion sizes after a subcutaneous challenge with viable P. gingiualis cells. Results: In CD4(+) T-cell-depleted mice (group 4), the DTH response and antigen-stimulated cell proliferation were significantly suppressed when compared to groups 2 and 3. Similarly, the levels of serum and splenic IgM, IgG, and all IgG subclass antibodies to P. gingiualis OMP were depressed. Delayed healing of P. gingivalis-induced lesions was also observed in the CD4(+) T-cell-depleted group. Conclusions: This study has shown that depletion of CD4(+) T cells prior to immunization with P. gingiualis OMP led to the suppression of both the humoral and cell-mediated immune response to this microorganism and that this was associated with delayed healing. These results suggest that the induction of the immune response to P. gingiualis is a CD4(+) T-cell-dependent mechanism and that CD4(+) T cells are important in the healing process.

Nature and specificity of the required protective immune response that develops postchallenge in mice vaccinated with the 19-kilodalton fragment of Plasmodium yoelii merozoite surface protein 1

Immunity induced by the 19-kDa fragment of Plasmodium yoelii merozoite surface protein 1 (MSP1(19)) is dependent on high titers of specific antibodies present at the time of challenge and a continuing active immune response postinfection. However, the specificity of the active immune response postinfection has not been defined. In particular, it is not known whether anti-MSP1(19) antibodies that arise following infection alone are sufficient for protection. We developed systems to investigate whether an MSP1(19)-specific antibody response alone both prechallenge and postchallenge is sufficient for protection. We were able to exclude antibodies with other specificities, as well as any contribution of MSP1(19)-specific CD4(+) T cells acting independent of antibody, and we concluded that an immune response focused solely on MSP1(19)-specific antibodies is sufficient for protection. The data imply that the ability of natural infection to boost an MSPI,g-specific antibody response should greatly improve vaccine efficacy.