Functional Analysis of Interleukin 8 Gene Haplotypes Associated With Periodontitis

Objectives: Patients carrying the ATC/TTC haplotype in the Interleukin 8 (IL8) gene were demonstrated as twice more susceptible to Chronic Periodontitis (CP), then named (IL8+). However, the functional role of IL8 haplotypes in immune response have been little investigated. The aim of this study was to investigate the functionality of IL8 haplotypes to evaluate the influence of this genetic variation in the regulation of immune response to CP.
Methods: Peripheral blood of twelve patients carrying the haplotype ATC/TTC (IL8+) or ATT/TTC (IL8-) were collected and separated in neutrophils, monocytes and T cells. The respective cultured cells were stimulated for 4 hours with PMA+Ionomycin, Interleukin-1β, Porphyromonas gingivalis (P.g.) or Aggregatibacter actinomycetemcomitans (A.a.). Total RNA was extracted from each cell type to investigate the mRNA levels of IL4, IL8, IL12 and Tumor Necrosis Factor alpha (TNFA) genes by RT-qPCR. For protein levels analysis by the multiplex assay, the total blood of the same patients was plated and stimulated for 12 hours with the same stimuli.
Results: Neutrophils and T cells stimulated with A.a. and P.g. showed increased TNFA and IL8 mRNA levels in patients carrying the (IL8+) haplotype compared to the (IL8-). Regarding protein analyses, higher levels of the IL-1β, IL-6, IL-8, IFNα, IL-10, IFN-γ, IL-2R, MIP-1a and MIP-1b were observed in the (IL8+) patients after the stimulus with A.a. and P.g.
Conclusions: In conclusion, the patients carrying the ATC/TCC (IL8+) haplotype associated with the susceptibility to CP showed increased gene expression and protein levels of important cytokines related with immune response to CP. This genetic carriage was biologically functional since it was associated with the immune cells response to periodontopathogens.