TLR4 activation by lipopolysaccharide and Streptococcus mutans induces differential regulation of proliferation and migration in human dental pulp stem cells.

INTRODUCTION

Dental pulp stem cells (DPSCs) are suspected to be an important part of the innate immune response of dental pulp, which is triggered by microorganisms that progressively invade the human tooth during the formation of caries. This study was performed to elucidate the expression of toll-like receptor 4 (TLR4) in dental pulp of deep caries and to determine whether TLR4 modulates the proliferation and migration of DPSCs.

METHODS

Pulp tissue samples were collected from freshly extracted human wisdom tooth. Immunohistochemistry and immunofluorescence were performed to determine the distribution of TLR4 in healthy dental pulp and dental pulp in deep caries. DPSCs were cultured and purified by collecting multiple colonies. The proliferation and migration of DPSCs were examined by 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyl-tetrazolium bromide, scratch test, and transwell migration assay after stimulation with lipopolysaccharide and extracts from Streptococcus mutans. TLR4 messenger RNA (mRNA) and cytokine mRNA were evaluated with real-time polymerase chain reaction; TLR4 protein was examined with Western blot and immunocytochemistry.

RESULTS

TLR4 is expressed in the odontoblast layer and areas that colocalize with blood vessels to different levels in healthy teeth and teeth affected by caries. TLR4 mRNA, TLR4 protein, and mRNA of cytokine levels can be elevated with stimulations of LPS and extracts from S. mutans. Lipopolysaccharide and extracts from S. mutans treatment inhibited the proliferation of DPSCs but promoted migration; however, these changes were abolished when TLR4 was blocked by anti-TLR4 antibody.

CONCLUSIONS

These results suggest that TLR4 will be activated and regulate the proliferation and migration of DPSCs in deep caries. TLR4 may play an important role in the immune response by DPSCs.