Effect of Age on Wnt/β-catenin Signalling in Dental Pulp Cells

Objectives: Pharmacological use of small molecule GSK-3β inhibitor delivered on a collagen sponge (CS) proved to stimulate the Wnt/β-catenin signalling pathway, leading to natural dentine repair in-vivo. The objective of the present study was to assess the Wnt/β-catenin signalling induction of primary rat pulp cells exposed to various GSK-3β inhibitor and to analyse the influence of age on its induction efficiency.

Methods: Primary pulp cell cultures were established from explants of maxillary and mandibular first molars of different aged Wistar rats: 5-day-old, 3-week-old and 3-month old. MTS-toxicity assay was performed to assess the optimal concentration for each test compound. Cells were treated with various GSK-3β inhibitors for 6h and 24h. After respective treatment, RNA was extracted and whole cell lysates were prepared. Relative Axin2 gene expression (qPCR analysis) and active β-catenin levels (Western Blotting) were assessed.
7-week-old adult Wistar rats (192.2±10.5g) were assigned to 5 experimental and 1 control groups (n=4) and pulp damage was performed under general anaesthesia on the first upper molars. Pulps were covered with respective GSK-3β inhibitors + CS and sealed with glass ionomer cement. Pulp tissue was collected 24h after treatment and relative Axin2 expression was assessed.

Results: The fold change induction of relative Axin2 expression and active β-catenin levels in cells obtained within different aged rats was similar among the tested GSK-3β inhibitors. Similar response were also reproduced in-vivo.

Conclusions: Considering future treatment strategies in reparative dentinogenesis the current findings indicated, that the tested GSK-3β inhibitors stimulated Wnt/β-catenin signalling in an age-independent manner and therefore might provide a broad application spectrum for further translational research. In addition, primary cell culture of the target tissue should be considered for investigating drug-response-testing prior to in-vivo experiments.