Abstract: Objective To compare a new thermosensitive recombinant human amelogenin (rhAm) carrier and traditional propylene glycol alginate (PGA) carrier for their characteristics, antibacterial activity, and biocompatibility with human periodontal membrane fibroblasts. Methods PGA- rhAm was prepared by mixing 3.3% PGA and rhAm, and CS-βGP-rhAm was prepared by mixing 2% chitosan (CS) with rhAm and then with 60% β- sodium glycerophosphate solution (βGP) as the crosslinking agent. The biophysical properties of the prepared carriers were characterized, and their antibacterial activity was assessed by observing Staphylococcus aureus growth. The biocompatibility of the carriers was evaluated in human periodontal membrane fibroblasts (hPDLFs) using CCK8 assay and scratch test, and mRNA and protein expressions of osteogenic genes of the cells incubated with the carriers were detected using RT-qPCR and Western blotting; osteogenic differentiation of the cells was detected using alkaline phosphatase staining. Results PGA-rhAm had a viscosity value of 3.262±0.055 Pa.s. CS-βGP-rhAm had a solidification capacity of 6 min at 37 ℃ with a pH value close to that of the oral cavity and a swelling rate of about 90%. CS-β GP-rhAm maintained sustained release of rhAm for over 2 weeks with a self-degradation time over 3 weeks. CS-βGP-rhAm more effectively inhibited the growth of S. aureus than rhAm- loaded PGA. While PGA did not obviously affect the proliferation of hPDLFs, both CS-βGP and CS-βGP- rhAm significantly promoted the cell proliferation（P<0.001). Scratch test showed that after rhAm loading, both CS-βGP and PGA promoted cell migration (P<0.01). CS-βGP-rhAm significantlyenhanced the mRNA expressions of RUNX2 and OCN mRNA level and the protein expressions of Ki67, RUNX2, collagen I, and β-catenin (P<0.05); PGA-rhAm only enhanced RUNX2 (P<0.05) and OCN (P<0.01) mRNA expressions without significant effects on the protein expressions. Alkaline phosphatase staining results showed that CS-βGP, but not PGA, promoted osteogenic differentiation of hPDLFs. Conclusion CS-βGP carrier is capable of sustained release of rhAm, inhibiting the growth of S. aureus, and improving the biological activity of hPDLFs without affecting the bioactivity of rhAm after drug loading.

Key words: recombinant human amelogenin; chitosan; propylene glycol alginate; human periodontal ligament fibroblast