title: Comparison of short-run cell seeding methods for poly(L-lactide-co-1,5-dioxepan-2-one) scaffold intended for bone tissue engineering.
abstract: Constructs intended for bone tissue engineering are influenced by the initial cell seeding procedure. The seeding method should be rapid, convenient, improve cell spatial distribution, and have no negative effects on cellular viability and differentiation. This study aimed to compare the effect of short-run seeding methods (centrifuge and vortex) with a static method on the scaffolds prepared from poly(L-lactide-co-1,5-dioxepan-2-one) by solvent-casting particulate-leaching (SCPL) technique. Human osteoblast-like cells (HOB) were seeded by the three methods described above. The seeding efficiency was determined by attached cell numbers. Cellular proliferation was analyzed by WST-1 and dsDNA assay. Cell distribution was examined by scanning electron (SEM) and fluorescence microscopy. Expression of alkaline phosphatase (ALP), collagen type I (Col I), osteocalcin (OC) and proliferating cell nuclear antigen (PCNA) were determined by real time RT-PCR. Results indicated that centrifuge and vortex increased seeding efficiency and had no negative effects on cellular viability. The data obtained by the fluorescence microscope confirmed the SEM results that the vortex method improved cell distribution through the scaffolds more than the other two methods (p<0.05). The RT-PCR results showed no significant differences on the expression of mRNA between the three methods of the above markers. The vortex method was found to be a simple and feasible seeding method for the poly(L-lactide-co-1,5-dioxepan-2-one) scaffolds.
authors.affiliation: Dept. of Clinical Dentistry-Center for Clinical Dental Research, Faculty of Medicine and Dentistry, University of Bergen, Norway. zhe.xing@uib.no
authors.forename: Zhe
authors.initials: Z
authors.lastname: Xing
authors.forename: Ying
authors.initials: Y
authors.lastname: Xue
authors.forename: Staffan
authors.initials: S
authors.lastname: Dånmark
authors.forename: Anna
authors.initials: A
authors.lastname: Finne-Wistrand
authors.forename: Kristina
authors.initials: K
authors.lastname: Arvidson
authors.forename: Sølve
authors.initials: S
authors.lastname: Hellem
authors.forename: Zhuang-Qun
authors.initials: ZQ
authors.lastname: Yang
authors.forename: Kamal
authors.initials: K
authors.lastname: Mustafa
mesh_terms: D000469:Alkaline Phosphatase / Q000235:genetics; D002448:Cell Adhesion; D018929:Cell Culture Techniques; D002454:Cell Differentiation; D049109:Cell Proliferation; D002470:Cell Survival; D002478:Cells, Cultured; D024042:Collagen Type I / Q000235:genetics; D006801:Humans; D008855:Microscopy, Electron, Scanning; D008856:Microscopy, Fluorescence; D010006:Osteoblasts / Q000502:physiology* / Q000648:ultrastructure; D015675:Osteocalcin / Q000235:genetics; D010012:Osteogenesis; D011091:Polyesters / Q000737:chemistry*; D018809:Proliferating Cell Nuclear Antigen / Q000235:genetics; D012333:RNA, Messenger / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013997:Time Factors; D023822:Tissue Engineering / Q000379:methods*; D054457:Tissue Scaffolds
chemical_list: D024042:Collagen Type I; D011091:Polyesters; D018809:Proliferating Cell Nuclear Antigen; D012333:RNA, Messenger; C556909:poly(L-lactide)-co-(1,5-dioxepan-2-one); D015675:Osteocalcin; D000469:Alkaline Phosphatase