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.

Immune responses to ISCOM (R) formulations in animal and primate models

ISCOMs(R) are typically 40 nm cage-like structures comprising antigen, saponin, cholesterol and phospholipid. ISCOMs(R) have been shown to induce antibody responses and activate T helper cells and cyrolytic T lymphocytes in a number of animal species, including non-human primates. Recent clinical studies have demonstrated that ISCOMs(R) are also able to induce antibody and cellular immune responses in humans. This review describes the current understanding of the ability of ISCOMs(R) to induce immune responses and the mechanisms underlying this property. Recent progress in the characterisation and manufacture of ISCOMs(R) will also be discussed. (C) 2001 Elsevier Science Ltd. All rights reserved.

Immune responses in hookworm infections

Hookworms infect perhaps one-fifth of the entire human population, yet little is known about their interaction with our immune system. The two major species are Necator americanus, which is adapted to tropical conditions, and Ancylostoma duodenale, which predominates in more temperate zones. While having many common features, they also differ in several key aspects of their biology. Host immune responses are triggered by larval invasion of the skin, larval migration through the circulation and lungs, and worm establishment in the intestine, where adult worms feed on blood and mucosa while injecting various molecules that facilitate feeding and modulate host protective responses. Despite repeated exposure, protective immunity does not seem to develop in humans, so that infections occur in all age groups (depending on exposure patterns) and tend to be prolonged. Responses to both larval and adult worms have a characteristic T-helper type 2 profile, with activated mast cells in the gut mucosa, elevated levels of circulating immunoglobulin E, and eosinoophilia in the peripheral blood and local tissues, features also characteristic of type I hypersensitivity reactions. The longevity of adult hookworms is determined probably more by parasite genetics than by host immunity. However, many of the proteins released by the parasites seem to have immunomodulatory activity, presumably for self-protection. Advances in molecular biotechnology enable the identification and characterization of increasing numbers of these parasite molecules and should enhance our detailed understanding of the protective and pathogenetic mechanisms in hookworm infections.

Antibody levels to the class I and II epitopes of the M protein and myosin are related to group A streptococcal exposure in endemic populations

Rheumatic fever (RF)/rheumatic heart disease (RHD) and post-streptococcal glomerulonephritis are thought to be autoimmune diseases, and follow group A streptococcal (GAS) infection. Different GAS M types have been associated with rheumatogenicity or nephritogenicity and categorized into either of two distinct classes (I or II) based on amino acid sequences present within the repeat region ('C' repeats) of the M protein. Sera from ARF patients have previously been shown to contain elevated levels of antibodies to the class I-specific epitope and myosin with the class I-specific antibodies also being cross-reactive to myosin, suggesting a disease association. This study shows that immunoreactivity of the class I-specific peptide and myosin does not differ between controls and acute RF (ARF)/RHD in populations that are highly endemic for GAS, raising the possibility that the association is related to GAS exposure, not the presence of ARF/RHD. Peptide inhibition studies suggest that the class I epitope may be conformational and residue 10 of the peptide is critical for antibody binding. We demonstrate that correlation of antibody levels between the class I and II epitope is due to class II-specific antibodies recognizing a common epitope with class I which is contained within the sequence RDL-ASRE. Our results suggest that antibody prevalence to class I and II epitopes and myosin is associated with GAS exposure, and that antibodies to these epitopes are not an indicator of disease nor a pathogenic factor in endemic populations.

Post-antibiotic growth suppression of linezolid against Gram-positive bacteria

The in vitro post-antibiotic effects (PAEs) of eight different concentrations of linezolid against Gram-positive cocci were investigated and the results analysed using the sigmoid E-max model for mathematically modelling the PAE. Mean maximal linezolid PAEs against strains of Staphylococcus aureus, Staphylococcus epidermidis, Enterococcus faecalis, Enterococcus faecium and Streptococcus pneumoniae were 2.2, 1.8, 2.8, 2.0 and 3.0 h, respectively. Resistance to methicillin (for the staphylococci), vancomycin (for the enterococci) and penicillin (for the pneumococci) had no effect on the duration of the PAE. Results of PAE testing support twice-daily dosing of linezolid in humans.

Route of administration of chimeric BPV1VLP determines the character of the induced immune responses

To examine the mucosal immune response to papillomavirus virus-like particles (PV-VLP), mice were immunized with VLP intrarectally (i.r.), intravaginally (i.va.) or intramuscularly (i.m.) without adjuvant. PV-VLP were assembled with chimeric BPV-1 L1 proteins incorporating sequence from HIV-1 gp 120, either the V3 loop or a shorter peptide incorporating a known CTL epitope (HIVP18I10). Antibody specific for BPV-1 VLP and P18 peptide was detected in serum following i.m., but not i.r. or i.va. immunization. Denatured VLP induced a much reduced immune response when compared with native VLP, Immune responses following mucosal administration of VLP were generally weaker than following systemic administration. VLP specific IgA was higher in intestine washes following i.r. than i.va. immunization, and higher in vaginal washes following i.m. than i.r. or i.va. immunization. No differences in specific antibody responses were seen between animals immunized with BPV-1 P18 VLP or with BPV-1 V3 VLP. Cytotoxic T lymphocyte precursors specific for the P18 CTL epitope were recovered from the spleen following i.m., i.va. or i.r. immunization with P18 VLP, and were similarly detected in Peyer's patches following i.m. or i.r. immunization. Thus, mucosal or systemic immunization with PV VLP induces mucosal CTL responses and this may be important for vaccines for mucosal infection with human papillomaviruses and for other viruses.

Induction of immune tolerance by dendritic cells: implications for preventative and therapeutic immunotherapy of autoimmune disease

Dendritic cells (DC) have a key role in controlling the immune response, by determining the outcome of antigen presentation to T cells. Through costimulatory molecules and other factors, DC are involved in the maintenance of peripheral tolerance through modulation of the immune response. This modulation occurs both constitutively, and in inflammation, in order to prevent autoimmunity and to control established immune responses. Dendritic cell control of immune responses may be mediated through cytokine or cell-contact dependent mechanisms. The molecular and cellular basis of these controls is being understood at an increasingly more complex level. This understanding is reaching a level at which DC-based therapies for the induction of immune regulation in autoimmunity can be tested in vivo. This review outlines the current state of knowledge of DC in immune tolerance, and proposes how DC might control both T cell responses, and themselves, to prevent autoimmunity and maintain peripheral tolerance.

Oral disease in animals: The Australian perspective. Isolation and characterisation of black-pigmented bacteria from the oral cavity of marsupials

A wide range of animals suffer from periodontal disease. However, there is very little reported on disease and oral micro-biota of Australian animals. Therefore, the oral cavity of 90 marsupials was examined for oral health status. Plaque samples were collected from the subgingival margins using curettes; or swabs. Plaque samples were plated onto. non-selective trypticase soy agar plates, selective trypticase soy agar, non-selective and selective Wilkens Chalgrens, Agar. Plates were incubated in an anaerobic atmosphere and examined after 7-14 days for the presence of black-brown-pigmented colonies. A combination of morphological and biochemical tests were used (colonial morphology, pigmentation, aerobic growth, Gram reaction, fluorescence under long-wave UV light (360 nm), production of catalase, enzymatic activity with fluorogenic substrates and haemagglutination of sheep red cells) to identify these organisms. Black-pigmented bacteria were cultivated from the plaque of 32 animals including six eastern grey kangaroos, a musky rat kangaroo, a whiptail and a red-necked wallaby, 18 koalas, a bandicoot and five brushtail possums. No black-pigmented colonies were cultivated from squirrel or sugar gliders or quokkas or from marsupial mice. The majority of isolates were identified as Porphyromonas gingivalis-like species with the higher prevalence of isolation from the oral cavity of macropods (the kangaroos and wallabies). Oral diseases, such as gingivitis can be found in native Australian animals with older koalas having an increase in disease indicators and black-pigmented bacteria. Non-selective Wilkens Chalgren Agar was the medium of choice for the isolation of black-pigmented bacteria. (C) 2002 Elsevier Science Ltd. All rights reserved.

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.

Isolation of Gemmata-like and Isosphaera-like planctomycete bacteria from soil and freshwater

New cultured strains of the planctomycete division (order Planctomycetales) of the domain Bacteria related to species in the genera Gemmata and Isosphaera were isolated from soil, freshwater, and a laboratory ampicillin solution. Phylogenetic analysis of the 16S rRNA gene from eight representative isolates showed that all the isolates were members of the planctomycete division. Six isolates clustered with Gemmata obscuriglobus and related strains, while two isolates clustered with Isosphaera pallida. A double-membrane-bounded nucleoid was observed in Gemmata-related isolates but not in Isosphaera-related isolates, consistent with the ultrastructures of existing species of each genus. Two isolates from this study represent the first planctomycetes successfully cultivated from soil.