title: In vitro human periodontal ligament-like tissue formation with porous poly-L-lactide matrix.
abstract: This study aimed to establish an in vitro human periodontal ligament-like tissue (HPdLLT) by three-dimensional culturing of human periodontal ligament fibroblasts (HPdLFs) in a porous poly-L-lactide (PLLA) matrix modified hydrophilically with ammonia solution. After ammonia modification, the surface roughness and culture-medium-soaking-up ability of the PLLA matrix increased, whereas the contact angle of water drops decreased. The thickness, porosity, and pore size of the PLLA matrix were 400±50 μm, 83.3%, and 75-150 μm, respectively. HPdLFs (1×10(5) cells) were seeded on the modified PLLA matrix and centrifuged to facilitate seeding into its interior and cultured for 14 days. Scanning electron microscope (SEM) observation, proliferation assay, picrosirius-red staining, and real-time polymerase chain reaction (RT-PCR) for type-1 collagen (COL1), periodontal ligament associated protein-1 (PLAP-1), fibroblast growth factor-2 (FGF-2), and alkaline phosphatase (ALP) mRNA were conducted on days 1, 3, 7, and 14. HPdLFs were observed entirely from the surface to the rear side of the matrix. Cell proliferation analysis, SEM observation, and picrosirius-red staining showed both progressive growth of 3D-cultured HPdLFs and extracellular matrix maturation by the secretion of COL1 and type 3 collagen (COL3) from days 1 to 14. Expressions of COL1, PLAP-1, and FGF-2 mRNA suggested the formation of cellular components and supplementation of extracellular components. Expressions of ALP, COL1, and PLAP-1 mRNA suggested the osteogenic potential of the HPdLLT. The results indicated in vitro HPdLLT formation, and it could be used in future periodontal ligament tissue engineering to achieve optimal periodontal regeneration.
authors.affiliation: Graduate School of Dentistry, Department of Orthodontics, Osaka Dental University, Hirakata-shi, Osaka-fu, Japan.
authors.forename: Wen
authors.initials: W
authors.lastname: Liao
authors.forename: Masahiro
authors.initials: M
authors.lastname: Okada
authors.forename: Fumito
authors.initials: F
authors.lastname: Sakamoto
authors.forename: Naoya
authors.initials: N
authors.lastname: Okita
authors.forename: Kaoru
authors.initials: K
authors.lastname: Inami
authors.forename: Aki
authors.initials: A
authors.lastname: Nishiura
authors.forename: Yoshiya
authors.initials: Y
authors.lastname: Hashimoto
authors.forename: Naoyuki
authors.initials: N
authors.lastname: Matsumoto
mesh_terms: D000469:Alkaline Phosphatase / Q000235:genetics / Q000378:metabolism; D000641:Ammonia / Q000737:chemistry; D018929:Cell Culture Techniques; D002478:Cells, Cultured; D024042:Collagen Type I / Q000235:genetics / Q000378:metabolism; D016326:Extracellular Matrix Proteins / Q000235:genetics / Q000378:metabolism; D016222:Fibroblast Growth Factor 2 / Q000235:genetics / Q000378:metabolism; D005347:Fibroblasts / Q000166:cytology / Q000378:metabolism; D006801:Humans; D008855:Microscopy, Electron, Scanning; D010513:Periodontal Ligament / Q000166:cytology; D011091:Polyesters / Q000737:chemistry*; D016062:Porosity; D023822:Tissue Engineering
chemical_list: C439889:ASPN protein, human; D024042:Collagen Type I; D016326:Extracellular Matrix Proteins; D011091:Polyesters; D016222:Fibroblast Growth Factor 2; C033616:poly(lactide); D000641:Ammonia; D000469:Alkaline Phosphatase