In vitro periodontal ligament cell expansion by co-culture method and formation of multi-layered periodontal ligament-derived cell sheets

Abstract

ObjectivePer-implantitis is one of the implant treatment complications. Dentists have failed to restore damaged periodontium by using conventional therapies. Tissue engineering (stem cells, scaffold and growth factors) aims to reconstruct natural tissues. The paper aimed to isolate both periodontal ligament stem cells (PDLSCs) and bone marrow mesenchymal stem cells (BMMSCs) and use them in a co-culture method to create three-layered cell sheets for reconstructing natural periodontal ligament (PDL) tissue.Materials and methodsBMMSCs were isolated from rabbit tibia and femur, and PDLSC culture was established from the lower right incisor. The cells were co-cultured to induce BMMSC differentiation into PDL cells. Cell morphology, stem cells and PDL-specific markers (CD90, CD34, and periostin) were also detected using immunofluorescent assay. Co-cultured cell monolayers were detached using temperature-responsive tissue culture dishes and collagen graft to create the three-layer construct. The 3D-engineered tissue was examined histologically and by field emission scanning electron microscopy (FESEM).ResultsBMMSCs co-cultured with PDLSCs successfully induced more PDL cells. The newly induced PDL cells exhibited periostin and CD90 expression. Fluorescence green intensity was measured for the co-cultured cells that were stained with periostin, the mean fluorescence green intensity (periostin expression) was significantly higher for the newly induced PDL cells after 1, 2, and 3 weeks when compared with control (BM-MSCs), at 21 days non-significant difference was measured when compared with control (PDLSCs).The results showed the successful formation of 3D multilayer PDL tissue. Histological cross-section showed cell sheets and the stable adhesion between them. FESEM examination was conducted for the cross-section, showing three-layered cell sheets with stable adhesion between cells.ConclusionsThe results of this paper report that the three layered-cell sheets were successfully constructed by the novel use of collagen graft as a scaffold to be used in treatment of periodontitis and to envelop the dental implants to create biohybrid implant.

Preparing polycaprolactone scaffolds using electrospinning technique for construction of artificial periodontal ligament tissue

“…Colonies and confluency of periodental ligament stem cells (PDLSCs) have also been described in another study. 19 The enzymatic dissociation method was used to obtain PDLSCs with greater proliferation rates, better colony-forming efficiency, and stronger differentiation capacity in culture than those obtained using the outgrowth method. Furthermore, the success rate of primary culture was higher with type I collagenase than with trypsin, which is in agreement with previously reported results.…”

“…Coverslips were fixed on microscopic slides, and the slides were examined in the dark using a fluorescence microscope (Leica, Germany). 19 …”

Enhanced VEGF/VEGF-R and RUNX2 Expression in Human Periodontal Ligament Stem Cells Cultured on Sandblasted/Etched Titanium Disk

“…In dentistry, human periodontal ligament stem cells (hPDLSCs) have been the mostly utilized MSCs population. They have been collected from the periodontal ligament, a specialized connective tissue that linked alveolar bone socket with the tooth root surface, distinguished by various cell populations as osteoblasts, fibroblasts, epithelial cells, endothelial cells, and stem cells (De Colli et al, 2015;Safi et al, 2019). The hPDLSCs show multipotent and proliferative properties (Winning et al, 2017).…”

Effects of Long Durations of RF–Magnetron Sputtering Deposition of Hydroxyapatite on Titanium Dental Implants

“…38,39 An extended osseointegration will result from slow resorption of implant coating and bone remodeling. Coated Ti dental implants will be covered with three-layered cell sheets that successfully form using collagen grafts as a scaffold 40 to construct biohybrid implants. Abbreviations: HA, hydroxyapatite; S a , roughness average; S dq , slop root mean square; S dr , increment of the interfacial surface area relative to a flat plane baseline; S q , height root mean square of the surface; Ti, titanium.…”