LAMA5‐inspired adhesive dodecapeptide facilitates efficient dentine regeneration: An in vitro and in vivo study

The primary goal of this study was to investigate the potential effects of A5G81 in inducing reparative dentine (RD) formation both in vitro and in vivo.Cell adhesion was observed by crystal violet staining and quantified by Sodium Dodecyl Sulphate (SDS) extraction. Cell proliferation was investigated using Cell Counting Kit-8 (CCK-8) assay. Spreading of cytoskeleton was visualized using immunofluorescence staining. Protein expression level of Akt signalling pathway was compared in a human Akt pathway phosphorylation array. Genes that were up or downregulated by A5G81 were identified by RNA sequencing. The mRNA expression of odontoblastic markers was detected by quantitative real-time polymerase chain reaction (qPCR). Moreover, mineralization of human dental pulp cells (hDPCs) was visualized by alizarin red staining and quantified using cetylpyridinium chloride (CPC). A direct pulp-capping model was established in SD rats and the RD formation at 2 weeks after operation was observed using HE staining.A5G81 (optimal coating concentration: 0.5 mg/mL) promoted hDPCs adhesion and proliferation to a level that was similar to Type I collagen (COL-1). Meanwhile, A5G81 activated Akt signalling pathway, albeit to a lesser extent than COL-1. An inhibition test indicated that A5G81 induced hDPCs adhesion by activating PI3K pathway. A5G81 induced the expression of ECM remodelling genes and odontoblastic genes, which were demonstrated by RNA-seq and qPCR, respectively. In addition, A5G81 efficiently accelerated the mineralization of hDPCs in both immobilized and soluble forms, a property that makes it more applicable in dental clinic. Finally, the pulp-capping study in rats suggested that use of A5G81 could successfully induce the formation of RD within 2 weeks.Coating of A5G81 to non-tissue culture-treated polystyrene facilitates spreading, proliferation and differentiation of hDPCs, resulting in rapid RD formation in artificially exposed pulp.