Interleukin-10 (-592 C/A) and interleukin-12B (+16974 A/C) gene polymorphisms and the interleukin-10 ATA haplotype are associated with periodontitis in a Taiwanese population.

BACKGROUND AND OBJECTIVE: Single nucleotide polymorphisms are assumed to be associated with the differential production of cytokines. We evaluated gene polymorphisms of interleukin-10 (-592C>A, -819C>T and -1082G>A) and interleukin-12B (+16974) in patients with chronic periodontitis (n = 145) and generalized aggressive periodontitis (n = 65) in comparison with healthy controls (n = 126). MATERIAL AND METHODS: Gene promoter polymorphisms were analyzed by polymerase chain reaction with sequence-specific primers. Genotype and allele frequencies were analyzed using the chi-square test and logistic regression analysis. RESULTS: The interleukin-10 -592 polymorphism showed significant differences among the three groups (p = 0.0330). The genotype frequencies of the -592 locus between the chronic periodontitis and healthy control groups were significantly different (AC vs. AA: odds ratio = 0.33). The combination ATA/ATA seemed to be associated with susceptibility to generalized aggressive periodontitis (p = 0.0276). Patients with the composite ATA/ACC were less likely to develop chronic periodontitis (p = 0.0248). The CC genotype of interleukin-12B (+16974) was related to chronic periodontitis (CC vs. AA, p = 0.0211; CC vs. AA+AC, p = 0.0187). The AC heterozygosity of interleukin-12B was significantly lower in chronic periodontitis vs. healthy controls (p = 0.0500). CONCLUSION: The interleukin-10 gene polymorphism at position -592C>A may be associated with a lower risk for development of chronic periodontitis. The interleukin-10 haplotype ATA is associated with generalized aggressive periodontitis. On the other hand, interleukin-12B genetic variants at position +16974 are associated with susceptibility to chronic periodontitis.

Immunostimulating complexes (ISCOMs) for nasal vaccination.

The immunostimulating complex (ISCOM) is documented as a strong adjuvant and delivery system for parenteral immunization. Its effectiveness for mucosal immunization has also been proven with various incorporated antigens. Lövgren et al. were the first to demonstrate the capacity of influenza virus ISCOMs to induce mucosal immune response and protection after one comparatively low nasal dose. Further studies show that similar to Cholera toxin (CT) and Escherichia coli heat-labile toxin (LT), ISCOMs break immunological tolerance and exert strong mucosal adjuvant activity, resulting in secretory IgA and systemic immune responses. Striking is the capacity of ISCOMs to induce CTL response also after nasal administration. In contrast to CT, ISCOMs initiate mucosal as well as systemic immune responses in an IL-12 dependent manner but independently of IL-4. The recombinant B subunit of cholera toxin (rCTB) was incorporated in the same ISCOM particle to explore symbiotic effects. The IgA response to rCTB in lungs was increased 100-fold when rCTB was administered nasally in ISCOMs and more than 10-fold in the remote mucosa of the genital tract. An enhanced IgA response to a passenger antigen OVA was recorded in the remote genital tract. After i.n. administration of the envelope proteins of respiratory syncytial virus in ISCOMs, high serum antibodies were induced, almost at the same levels as those following parenteral immunization and potent IgA responses were also evoked both at the local respiratory mucosa, and in the cases tested at the distant mucosae of the genital and intestinal tracts. Similar results have also been recorded with ISCOMs containing envelope proteins from Herpes simplex virus, Influenza virus and Mycoplasma mycoides. The mucosal targeting property of envelope proteins of RSV was utilized in an HIV-gp120 RSV ISCOM formulation. After nasal administration an enhanced mucosal IgA response to gp120 was observed in the female reproductive tract. In general, antigens derived from envelope viruses or cell membranes incorporated into ISCOMs retain their biological activity and conformation, encompassing the mucosal targeting and virus neutralizing properties.

Different respiratory syncytial virus and Quillaja saponin formulations induce murine peritoneal cells to express different proinflammatory cytokine profiles.

The recognition of a pathogen or a vaccine antigen formulation by cells in the innate immune system leads to production of proinflammatory cytokines, which will determine the ensuing acquired immune response quantitatively and qualitatively. Tumour necrosis factor (TNF)-alpha, interleukin (IL)-1 and IL-6 are the first set of cytokines produced upon such an encounter, which have roles both in protective immunity and immunopathogenesis evident with respiratory syncytial virus (RSV). RSV antigens in different physical adjuvant-vaccine formulations were analysed for their capacity to provoke cultured murine peritoneal cells to produce these three proinflammatory cytokines. RSV immunostimulating complex (ISCOM), i.e. both antigen and adjuvant are incorporated in the same particle, induced high levels of IL-1alpha being of the same magnitude or higher than those of live RSV and lipopolysaccharide (LPS). Live virus and LPS induced higher levels of IL-6 and TNF-alpha than ISCOM and so did non-adjuvanted UV-inactivated RSV but only at high doses. ISCOM-Matrix, i.e. ISCOM without antigens, admixed as a separate entity to inactivated RSV, downregulated or blocked the cytokine response to the inactivated RSV in contrast to ISCOM. Kinetic studies showed that ISCOM induced cytokine production first detected at hours 1, 2, 4 for TNF-alpha, IL-6 and IL-1alpha respectively, which was earlier than for the other antigen formulations containing corresponding doses of antigen and/or Quillaja adjuvant. Peak values for production of TNF-alpha and IL-6 were at 8 h and for IL-1alpha at 72 h following stimulation with ISCOM. The delayed appearance of IL-1alpha may reflect the cell-bound nature of this cytokine.

Using distilled water for the extraction of mucosal antibodies and the subsequent application in RSV neutralization test.

Virus neutralization (VN) is an important functional test for evaluating RSV vaccines, also encompassing in mucosal secretion of the respiratory tract considering the infection route. In our previous study, an immunoglobin extraction method described by Bergquist et al. was adopted for RSV ELISA, but it was not suitable for virus neutralization test due to the cell toxicity of the 2% saponin solution used for the antibody extraction. In order to overcome this problem, several solvents including distilled water were tested in the present study for the capacity to extract immunogloblins. Antibodies in the extracts were evaluated and compared by ELISA. Distilled water was as efficient as the 2% saponin solution for extraction of total IgA, RSV specific IgA and IgG. More importantly, the organ extracts obtained subsequently could be used for virus neutralization test without causing adverse effect on the cell culture. Therefore, distilled water was finally chosen as the solvent for immunoglobulin extraction from mucosal organs when both ELISA and virus neutralization test are required.

Iscom and iscom-matrix enhance by intranasal route the IgA responses to OVA and rCTB in local and remote mucosal secretions.

Iscoms, with rCTB incorporated via the GM1 receptor, enhanced in mice the mucosal immunogenicity of rCTB as antigen after intranasal (i.n.) administration both by inducing IgA response in the remote intestinal tract mucosa and by a 100-fold increase of the specific IgA locally in the lungs. Iscom-matrix as a separate entity mixed with rCTB enhanced the rCTB-IgA response similarly. While OVA in iscoms induced high mucosal IgA responses, iscom-matrix co-administered with OVA induced low or no mucosal IgA response to OVA. A synergism between iscoms and rCTB could only be seen as an adjuvant targeting effect enhancing the IgA response to OVA in the remote genital tract mucosa. In serum, the immunomodulatory effect of iscoms after i.n. administration was seen as an enhanced serum IgG2a response.

Induction of antibody responses in the common mucosal immune system by respiratory syncytical virus immunostimulating complexes.

Immunostimulating complexes (ISCOMs) containing envelope proteins of respiratory syncytial virus (RSV) were explored as a mucosal delivery system for the capacity of inducing a common mucosal antibody response. Two intranasal (i.n.) administrations of BALB/c mice with ISCOMs induced potent serum IgG, and strong IgA responses to RSV locally in the lungs and the upper respiratory, and remotely in the genital and the intestinal tracts. Virtually no measurable IgA response was found in these mucosal organs after two subcutaneous (s.c.) immunizations. Virus neutralizing (VN) antibodies were detected in serum and in all of the mucosal organ extracts after both s.c. and i.n. immunizations indicating that the neutralizing epitopes were preserved after both mucosal and parenteral modes of administration. While the mucosal IgA response appears to be of mucosal origin, the IgG antibodies to RSV detected in the mucosal organs were likely of serum origin. However, the mucosal VN antibodies correlated with the IgG rather than the IgA levels. An enhanced IgA response to gp120 in various mucosal organs was recorded after i.n. immunization with gp120 incorporated in RSV ISCOMs, indicating a role of RSV envelope proteins in enhancing and targeting mucosal responses to passenger antigens.

The immunostimulating complex (ISCOM) is an efficient mucosal delivery system for respiratory syncytial virus (RSV) envelope antigens inducing high local and systemic antibody responses.

ISCOM is an efficient mucosal delivery system for RSV envelope proteins as measured by antibody responses in respiratory tract secretions and in sera of mice following two intranasal (i.n.) administrations. Intranasally administered RSV ISCOMs induced high levels of IgA antibodies both in the upper respiratory tract and in the lungs. In the lungs, a prominent and long-lasting IgA response was recorded, which still persisted 22 weeks after the second i.n. immunization when the experiment ended. Subcutaneous (s.c.) immunization only induced low IgA titres in the upper respiratory tract and no measurable response to RSV was found in the lungs. Differences were also noticed in serum between the i.n. and s.c. modes of immunization. ISCOMs given intranasally induced earlier, higher and longer lasting IgM and IgG1 serum anti-RSV antibody responses than those induced by the s.c. mode of administration. A low serum IgE response was only detectable at 2 weeks after i.n. immunization with ISCOMs and after s.c. immunization with an inactivated virus, but no IgE response was detectable after s.c. injection of ISCOMs. The serum IgA response was more pronounced following s.c. injection of inactivated virus than after i.n. application of ISCOMs, and a clear-cut booster effect was obtained with a second immunization. Virtually no serum IgA response was detected after the s.c. administration of ISCOMs. In conclusion, the high immune responses induced by RSV ISCOMs in the respiratory tract and serum after i.n. administration indicate prominent mucosal delivery and adjuvant properties of the ISCOMs, warranting further studies.