Purpose: To investigate the role and activation mechanism of TAZ in periodontal ligament stem cells (PDLSCs) perceiving hierarchical microgroove/nanopore topography.
Materials and Methods: Titanium surface with hierarchical microgroove/nanopore topography fabricated by selective laser melting combined with alkali heat treatment (SLM-AHT) was used as experimental group, smooth titanium surface (Ti) and sandblasted, large-grit, acid-etched (SLA) titanium surface were employed as control groups. Alkaline phosphatase (ALP) activity assays, qRT-PCR, Western blotting, and immunofluorescence were carried out to evaluate the effect of SLM-AHT surface on PDLSC differentiation. Moreover, TAZ activation was investigated from the perspective of nuclear localization to transcriptional activity. TAZ knockdown PDLSCs were seeded on three titanium surfaces to detect osteogenesis- and adipogenesis-related gene expression levels. Immunofluorescence and Western blotting were employed to investigate the effect of the SLM-AHT surface on actin cytoskeletal polymerization and MAPK signaling pathway. Cytochalasin D and MAPK signaling pathway inhibitors were used to determine whether actin cytoskeletal polymerization and the MAPK signaling pathway were indispensable for TAZ activation.
Results: Our results showed that SLM-AHT surface had a greater potential to promote PDLSC osteogenic differentiation while inhibiting adipogenic differentiation than the other two groups. The nuclear localization and transcriptional activity of TAZ were strongly enhanced on the SLM-AHT surface. Moreover, after TAZ knockdown, the enhanced osteogenesis and decreased adipogenesis in SLM-AHT group could not be observed. In addition, SLM-AHT surface could promote actin cytoskeletal polymerization and upregulate p-ERK and p-p38 protein levels. After treatment with cytochalasin D and MAPK signaling pathway inhibitors, differences in the TAZ subcellular localization and transcriptional activity were no longer observed among the different titanium surfaces.
Conclusion: Our results demonstrated that actin cytoskeletal polymerization and MAPK signaling pathway activation triggered by SLM-AHT surface were essential for TAZ activation, which played a dominant role in SLM-AHT surface-induced stem cell fate decision.
Keywords: TAZ, hierarchical micro/nanoscale topography, periodontal ligament stem cells, osteogenic differentiation, adipogenic differentiation