title: Five percent oxygen tension is not beneficial for neocartilage formation in scaffold-free cell cultures.
abstract: We have investigated whether 5% oxygen tension (O(2)) is beneficial for neocartilage formation when chondrocytes are cultured in transwell-COL inserts. Six million bovine primary chondrocytes were cultured in an insert with DMEM supplemented with 10% fetal bovine serum and antibiotics, with or without glucosamine sulphate (GS) in a 5% or 20% O(2) environment for 2, 4, or 6 weeks. The samples were collected for the histological staining of proteoglycans (PGs) and type II collagen, quantitative reverse transcription with the polymerase chain reaction (RT-PCR) analyses of the mRNA expression of aggrecan and procollagen α(1)(II), procollagen α(2)(I) and hyaluronan synthase 2, quantitation of PGs, and agarose gel electrophoresis. Neocartilage produced at 20% O(2) appeared larger than that at 5% O(2). Histological staining showed that more PGs and type II collagen and better native cartilage structure were produced at 20% than at 5% O(2). The thickness of neocartilage increased during the culture period. Quantitative RT-PCR showed that the procollagen α(1)(II) mRNA expression level was significantly higher at 20% than at 5% O(2). However, no significant difference in gene expression and PG content was found between control and GS-treated cultures at either 20% or 5% O(2). Thus, in contrast to monolayer cultures, engineered cartilage from scaffold-free cultured chondrocytes at 20% O(2) produced better extracellular matrix (ECM) than that at 5% O(2). PGs were mainly large. Exogenous GS was not beneficial for the ECM in scaffold-free chondrocyte cultures.
authors.affiliation: Department of Biosciences, Biocenter Kuopio, University of Eastern Finland, Kuopio, Finland. chengjuan.qu@uef.fi
authors.forename: Chengjuan
authors.initials: C
authors.lastname: Qu
authors.forename: Heli
authors.initials: H
authors.lastname: Lindeberg
authors.forename: Janne H
authors.initials: JH
authors.lastname: Ylärinne
authors.forename: Mikko J
authors.initials: MJ
authors.lastname: Lammi
mesh_terms: D000818:Animals; D002356:Cartilage / Q000187:drug effects* / Q000254:growth & development*; D002417:Cattle; D018929:Cell Culture Techniques / Q000379:methods*; D019902:Chondrocytes / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D024042:Collagen Type I / Q000235:genetics / Q000378:metabolism; D024043:Collagen Type II / Q000378:metabolism; D005109:Extracellular Matrix / Q000187:drug effects / Q000235:genetics / Q000378:metabolism; D005786:Gene Expression Regulation / Q000187:drug effects; D009928:Organ Specificity / Q000187:drug effects / Q000235:genetics; D010100:Oxygen / Q000494:pharmacology*; D011509:Proteoglycans / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D054457:Tissue Scaffolds / Q000737:chemistry*
chemical_list: D024042:Collagen Type I; D024043:Collagen Type II; D011509:Proteoglycans; D012333:RNA, Messenger; D010100:Oxygen