Gingival cell IL-2 and IL-4 in early-onset periodontitis

The purpose of this study was to compare, using cell blot analysis, the association of gingival tissue mononuclear cells (GTMC) isolated from lesions displaying histories of early-onset periodontitis (EOP; typically B-lymphocyte dominated) and gingivitis (typically T-lymphocyte dominated) with the B-cell stimulating cytokine, interleukin (IL)-4, and the T-cell stimulating cytokine, IL-2. Eleven EOP patients and 11 age- and gender-similar gingivitis control (GC) subjects participated. Gingival tissue adjacent to the alveolar crest normally removed during surgery was digested in collagenase-containing media and GTMC were isolated by density gradient centrifugation. Cells were separated into four aliquots. One was left unstimulated; the remainder were stimulated for 2 hours with Porphyromonas gingivalis outer membrane protein, mitogen Concanavalin A, or common antigen tetanus toxoid. Cells then were centrifuged onto transfer membranes and incubated in RPMI 1640 media for 6 hours to allow adsorption of secreted cytokine. Membranes were treated with monoclonal anti-IL-2 or anti-IL-4, followed by a biotin-conjugated second layer, streptavidin-alkaline phosphatase and nitro blue tetrazolium/5-bromo-4-chloro-indolyl-phosphate (NBT/BCIP) color development. A higher percentage of GTMC from EOP patients were IL-2⁺ when stimulated with P. gingivalis compared with GTMC from GC patients (20 ± 2% vs. 12 ± 2%, P < 0.003). A higher percentage of non-stimulated GTMC from EOP patients produced IL-4 than from GC (22 ± 4% vs. 6 ± 3%, P < 0.00007), as well as when stimulated with P. gingivalis (22 ± 3% vs. 13 ± 2%, P < 0.01). Analysis of IL-2/IL-4 positive GTMC ratios revealed significant differences between EOP and GC for non-stimulated cultures (0.5 ± 0.2 vs. 2.9 ± 1.0, P < 0.02). Digital analysis indicated significantly greater areas of IL-4 staining for EOP than GC GTMC in non-stimulated cultures (P < 0.006). These data would support the hypothesis that altered IL-2/IL-4 activities are associated with the periodontitis lesion.