EGCG diminishes CCL2 expression in human osteoblasts

Objective: To assess the effects of epigallocatechin-3-gallate (EGCG) on oncostatin M (OSM)-induced CCL2 synthesis and the associated signaling pathways in human osteoblastic cells. The therapeutic effect of EGCG on periapical lesion in rat models was also studied.

Methods: CCL2 and c-Fos mRNA expression was analyzed by Northern blotting. Modulating effects of EGCG on the activation of Raf-1, Akt and PI3K were examined by co-immunoprecipitation, Western blot and PI3-kinase activity assay. Interactions between c-Fos and CCL2 promoter were evaluated by electrophoretic mobility shift assay (EMSA) and chromatin immunoprecipitation (ChIP). The effect of EGCG on rat periapical lesions was examined radiographically and immunohistochemically.

Results: EGCG inhibited OSM-stimulated CCL2 expression in primary human osteoblasts and MG-63 (a human osteoblastic cell line). In MG-63, EGCG alleviated the OSM-induced phosphorylation of Raf-1 (Ser338) but restored the de-phosphorylation of Raf-1S259. EGCG increased the activity of PI3K, the level of phospho-AktS473 and binding between Raf-1 and active Akt. EMSA and ChIP revealed that EGCG attenuated AP-1/ CCL2 promoter interaction, possibly through reducing c-Fos synthesis. Co-distribution of CD-68+ macrophages and CCL2+ osteoblasts in osteolytic areas was obvious in periapical lesions. Administration of EGCG markedly diminished the extent of bone loss, macrophage infiltration and the amount of CCL2-synthesizing osteoblasts.

Conclusion: By stimulating PI3K activity, EGCG promoted Akt/Raf-1 cross-talk, resulting in decreased AP-1 binding to CCL2 promoter and finally reduced CCL2 production in osteoblastic cells. EGCG alleviated the severity of periapical osteolysis, probably by suppressing CCL2 synthesis in osteoblasts to diminish macrophage infiltration. Our data support the therapeutic potential of EGCG on periapical periodontitis.