AGE Induces Calcification- and Inflammation-related Factors in Dental Pulp Cells

Objectives: Advanced glycation end-products (AGE) have been found to play a role in the progression of vascular complications and inflammation.  Amorphous calcification frequently appears in dental pulp tissues of diabetic patients.  We previously reported that pathologic pulp calcifications such as pulp stones and thickened predentin occurred in diabetic rats and that  osteopontin (OPN) might be a key molecule of the pathologic pulp calcifications (Inagaki et al, J.Endod2010).   In addition, S100A8, S100A9 and interleukin-6 (IL-6) are reported to express in atherosclerotic plaques with inflammation.  The aim of this study is to determine the association of AGE with pathologic calcification and inflammation in pulp tissues using rat dental pulp cell cultures.

Methods: Dental pulp cells were cultured using incisors of Wistar rats as described by Kasugai et al. (Arch Oral Biol 1993).  Rat gingival fibroblasts were also cultured as the control cells.  AGE was prepared by a modification method of Takeuchi et al. (Mol Med1999) and added to the culture medium.  Alkaline phosphatase (ALPase) activity and calcified-nodule formation were measured.  mRNA expressions of receptor for AGE (RAGE), OPN, osteocalcin (OCN), S100A8, S100A9, IL-6 and IL-1β were determined by quantitative real-time PCR.  Amount of OPN and OCN proteins were measured by ELISA.

Results: ALPase activity and nodule formation were increased by AGE treatment in dental pulp cells.  AGE did not affect such markers in gingival fibroblasts.  AGE increased mRNA levels of RAGE, OPN, OCN, S100A8, S100A9, IL-6 and IL-1β.  Protein levels of OPN and OCN in dental pulp cells were increased by AGE, whereas its effect was less in gingival fibroblasts.  

Conclusions: AGE may be a stimulatory factor of pathologic calcification and inflammation in diabetic dental pulp tissues and the effect is characteristic in dental pulp cells.