Abstract Background and Objective Periodontal ligament cells (PDLCs) are an important
source for periodontal tissue healing and regeneration. Proper cell adhesion is a
key for survival of anchorage-dependent cells and also initiates further intracellular
signals for essential cellular functions. We aimed to test 3 different synthetic conjugates
with integrin-binding RGD sequence (SAK-c[RGDfC], AK-c[RGDfC], and SAK-opn on the
adhesion of human PDLCs and subsequent events including proliferation, migration,
behavior of cell surface molecules, and osteogenic differentiation. Materials and
Methods Synthetic peptides were synthesized by solid-phase technique and attached
to branched chain polymeric polypeptides via thioether linkage. Simple adsorption
method was used to coat tissue culture plastic or electric arrays. PDLCs were isolated
from 24 surgically extracted human third molars. Cell adhesion and proliferation were
measured with real-time impedimetric xCELLigence SP system. Cell migration assay was
performed with Ibidi? Culture inserts. Cell surface antigens were detected using flow
cytometry analysis. Osteogenic differentiation was assessed with alkaline phosphatase
(ALP) assay and Alizarin Red S staining, and real-time qPCR was performed to analyze
the osteoblast-related gene expression. Osteogenic differentiation and adipogenic
differentiation of PDLCs were monitored by real-time Electrical Cell-Substrate Impedance
Spectroscopy (ECIS). Results Primary outcome of this study relies on that all three
synthetic RGD peptides improved PDLC adhesion (P < .05). When animal serum is absent
in culture medium, SAK-c[RGDfC] and AK-c[RGDfC] elevated cell adhesion (P < .05).
Cell migration was enhanced by SAK-c[RGDfC] and AK-c[RGDfC] (P < .05). After 1-week
treatment, all synthetic peptides elevated CD105 (1.7- to 2.2-fold) and CD146 (1.3-
to 1.5-fold) markers and caused different integrin patterns. ALP activity (1.4-fold)
and ARS (1.8- and 2.0-fold) were increased by SAK-c[RGDfC] and AK-c[RGDfC] in absence
of osteogenic supplements, and all the peptides supported the mineralization under
osteogenic condition (P < .05). RT-qPCR revealed the upregulation of bone sialoprotein
(5.0- to 7.8-fold), osteocalcin (2.3- to 2.7-fold), and ALP (1.9- to 2.3-fold) gene
expression in osteogenesis-induced PDLCs. ECIS monitoring showed that higher impedance
was generated by the osteogenic induction compared with the adipogenic or the non-induced
(P < .05). Conclusions Our study demonstrates that SAK-c[RGDfC] and AK-c[RGDfC] improved
adhesion and migration of PDLCs and supported osteogenic differentiation of PDLCs.
These cyclic RGD peptides proved to be applicable biocompatible material in regenerative
medicine.
