Advanced Glycation End-Products Increase Calprotectin in Human Gingival Epithelial Cells

Objectives: Advanced glycation end-products (AGEs) in gingival tissues of diabetes patients aggravate periodontal disease, but their mechanisms are not well known. S100A8 has protective functions for epithelial cells and calprotectin (S100A8/S100A9) inhibits bacterial invasion into oral epithelium by antimicrobial activity and protecting epithelial cells. In this study, we investigated effects of AGE and Porphyromonas gingivalis lipopolysaccharide (PgLPS) on S100A8 and S100A9 expression and their translocation in human gingival epithelial cells.

Methods: OBA-9 cell, an immortalized human gingival epithelial cell line, was incubated with or without BSA, AGE and PgLPS. mRNA expression of S100A8 and S100A9 were examined by real-time PCR. Calprotectin were quantitated by ELISA. The localization of S100A8 and S100A9 in OBA-9 cells was investigated by immunohistochemical analysis. Western blotting for MAPK and NF-κB phosphorylation was performed, and the effects of their inhibitors and depletion of RAGE or TLR2 using their siRNAs on S100A8 and S100A9 expressions were investigated in AGE-stimulated epithelial cells.

Results: OBA-9 cells endogenously expressed S100A8 and S100A9. AGE increased S100A8 and S100A9 mRNA and calprotectin levels and PgLPS amplified this induction. S100A8 was located in the cellular nucleus of control cells treated with BSA in the presence or absence of PgLPS and most translocated to the cytoplasm by AGE stimulation. In contrast, S100A9 was located in the cytoplasm and was not altered following incubation with AGE or PgLPS. RAGE knockdown and inhibitors of p38, JNK and NF-κB counteracted AGE- and PgLPS-stimulated S100A8 and S100A9 expressions.
Conclusions: AGE and PgLPS increase S100A8 and S100A9 expressions through the p38 and JNK MAPK, and NF-κB and modulate localization of S100A8 and S100A9 in gingival epithelial cells and may play complex roles in the gingiva of diabetes-associated periodontitis.