title: Culture temperature affects human chondrocyte messenger RNA expression in monolayer and pellet culture systems.
abstract: Cell-based therapy has been explored for articular cartilage regeneration. Autologous chondrocyte implantation is a promising cell-based technique for repairing articular cartilage defects. However, there are several issues such as chondrocyte de-differentiation. While numerous studies have been designed to overcome some of these issues, only a few have focused on the thermal environment that can affect chondrocyte metabolism and phenotype. In this study, the effects of different culture temperatures on human chondrocyte metabolism- and phenotype-related gene expression were investigated in 2D and 3D environments. Human chondrocytes were cultured in a monolayer or in a pellet culture system at three different culture temperatures (32°C, 37°C, and 41°C) for 3 days. The results showed that the total RNA level, normalized to the threshold cycle value of internal reference genes, was higher at lower temperatures in both culture systems. Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and citrate synthase (CS), which are involved in glycolysis and the citric acid cycle, respectively, were expressed at similar levels at 32°C and 37°C in pellet cultures, but the levels were significantly lower at 41°C. Expression of the chondrogenic markers, collagen type IIA1 (COL2A1) and aggrecan (ACAN), was higher at 37°C than at 32°C and 41°C in both culture systems. However, this phenomenon did not coincide with SRY (sex-determining region Y)-box 9 (SOX9), which is a fundamental transcription factor for chondrogenesis, indicating that a SOX9-independent pathway might be involved in this phenomenon. In conclusion, the expression of chondrocyte metabolism-related genes at 32°C was maintained or enhanced compared to that at 37°C. However, chondrogenesis-related genes were further induced at 37°C in both culture systems. Therefore, manipulating the culture temperature may be an advantageous approach for regulating human chondrocyte metabolic activity and chondrogenesis.
authors.affiliation: Department of Motor Function Analysis, Human Health Sciences, Graduate School of Medicine, Kyoto University, Kyoto, Japan; Japan Society for the Promotion of Science, Tokyo, Japan.
authors.forename: Akira
authors.initials: A
authors.lastname: Ito
authors.affiliation: Department of Motor Function Analysis, Human Health Sciences, Graduate School of Medicine, Kyoto University, Kyoto, Japan.
authors.forename: Momoko
authors.initials: M
authors.lastname: Nagai
authors.affiliation: Department of Motor Function Analysis, Human Health Sciences, Graduate School of Medicine, Kyoto University, Kyoto, Japan.
authors.forename: Junichi
authors.initials: J
authors.lastname: Tajino
authors.affiliation: Department of Motor Function Analysis, Human Health Sciences, Graduate School of Medicine, Kyoto University, Kyoto, Japan; Japan Society for the Promotion of Science, Tokyo, Japan.
authors.forename: Shoki
authors.initials: S
authors.lastname: Yamaguchi
authors.affiliation: Department of Motor Function Analysis, Human Health Sciences, Graduate School of Medicine, Kyoto University, Kyoto, Japan.
authors.forename: Hirotaka
authors.initials: H
authors.lastname: Iijima
authors.affiliation: Department of Motor Function Analysis, Human Health Sciences, Graduate School of Medicine, Kyoto University, Kyoto, Japan.
authors.forename: Xiangkai
authors.initials: X
authors.lastname: Zhang
authors.affiliation: Department of Development and Rehabilitation of Motor Function, Human Health Sciences, Graduate School of Medicine, Kyoto University, Kyoto, Japan.
authors.forename: Tomoki
authors.initials: T
authors.lastname: Aoyama
authors.affiliation: Department of Motor Function Analysis, Human Health Sciences, Graduate School of Medicine, Kyoto University, Kyoto, Japan.
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Kuroki
mesh_terms: D053674:Aggrecans / Q000235:genetics; D002358:Cartilage, Articular / Q000166:cytology*; D018929:Cell Culture Techniques / Q000379:methods*; D002478:Cells, Cultured; D019902:Chondrocytes / Q000166:cytology* / Q000378:metabolism; D002950:Citrate (si)-Synthase / Q000235:genetics; D024043:Collagen Type II / Q000235:genetics; D005260:Female; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases / Q000235:genetics; D006801:Humans; D008875:Middle Aged; D012333:RNA, Messenger / Q000378:metabolism*; D013696:Temperature
chemical_list: C508319:ACAN protein, human; D053674:Aggrecans; C499177:COL2A1 protein, human; D024043:Collagen Type II; D012333:RNA, Messenger; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases; D002950:Citrate (si)-Synthase