Type 2 diabetes and the clinically normal pulp: An in vitro study

Abstract

Aim: The aim of this study was to investigate the effect of type 2 diabetes (T2D) on clinically normal dental pulp tissue by using special stains and immunohistochemistry (IHC) to determine the morphology of the coronal pulp and distribution of immune markers in non-T2D and T2D groups.

Methodology: Ethics approval for this in vitro pilot study was obtained from the University of Otago Human Ethics Committee (16/069). Twenty extracted permanent molar teeth diagnosed as having clinically normal pulp status were collected. Ten teeth were from participants with well-controlled T2D and ten from participants without diabetes (non-T2D). Each tooth was sectioned transversely at the cemento-enamel junction before the crowns were decalcified and embedded in paraffin. Sections were stained with haematoxylin and eosin, Massons trichrome, and van Gieson stains for histological and morphological evaluation. IHC using anti-CD4, anti-CD68 and anti-CD83 and anti-IL1β, anti-IL6, anti-IL17, anti-TNF-α, anti-TLR2, anti-TLR4 and anti-FOXP3 identified proteins of interest. Qualitative and semi-quantitative analyses evaluated the morphology of the dental pulp and protein expression. Data analyses were performed with GraphPad Prism, using Student's t-test and multiple regression using SPSS at p < .05.

Results: Special stains demonstrated morphological differences in the T2D dental pulp compared with non-T2D. Qualitative analysis indicated that the pulp in the T2D samples was consistently less cellular, less vascular, showed evidence of thickened blood vessel walls, increased pulp calcification and collagen deposition. Semi-quantitative analysis of IHC samples showed the T2D pulp had significantly increased expression of macrophage and dendritic cell markers CD68 (p < .001) and CD83 (p = .04), and there was significantly greater expression of inflammatory cytokines IL1β (p = .01), IL6 (p < .0001), IL17 (p < .0001) and TNF-α (p = .01). T2D samples showed a significant increase in markers of innate inflammation, TLR2 (p < .001) and TLR4 (p < .001) and decreased expression of regulatory T-cell marker, FOXP3 (p = .01). Multiple regression showed that age-corrected differences were statistically significant.

Conclusion: Preliminary findings suggest that T2D may exert a similar response in the pulp to complications in other body sites. Hyperglycaemia is associated with changes in the morphology of the clinically normal dental pulp with altered immune cell and cytokine expression.

Keywords: dental pulp; histomorphology; immune response; immunohistochemistry; type 2 diabetes.