title: Growth differentiation factor 5 modulation of chondrogenesis of self-assembled constructs involves gap junction-mediated intercellular communication.
abstract: A novel scaffold-free self-assembled cartilage construct has been generated and used to repair particular chondral defects effectively. However, the mechanisms related to the construction of these self-assembled cartilages have not yet been fully elucidated. We hypothesize that gap junction intercellular communication (GJIC) plays a critical role in the development of self-assembled constructs upon GDF-5 induction. In this study, we investigated the effect of connexin 43 (C×43) mediated GJIC on GDF-5 modulation of chondrogenesis from two aspects, cell monolayer culture and 3-D self-assembly culture. We induced cells or self-assembled constructs with chondrogenic media (CM), growth differentiation factor 5 (GDF-5) or 1-heptanol for 3 weeks. At the end of that time, the results of quantitative fluorescence redistribution after photobleaching (FRAP) assay and immunofluorescence demonstrated that GDF-5 improved both GJIC and chondrogenic differentiation to a significant degree while 1-heptanol nearly offset the expected improvements in chondrogenesis. Biochemical assay and histology showed that GDF-5 can obviously enhance GAG, C×43 and type II collagen expressions. Conversely, we also showed that while 1-heptanol weakened GAG and type II collagen expression in self-assembled constructs, it had no effect on C×43 expression. Furthermore, real-time polymerase chain reaction showed that GDF-5 enhanced GAG and type II collagen transcription while 1-heptanol reduced them, but was affectless on C×43 transcription. This suggests that the generation of scaffold-free self-assembled cartilage from human mesenchymal stem cells upon GDF-5 induction may be mediated, at least in part, via the modulation of GJIC.
authors.affiliation: Department of Orthopedics, Union Hospital, Tongji Medical College, Huazhong University of Science & Technology, Wuhan, Hubei, China.
authors.forename: Zhibo
authors.initials: Z
authors.lastname: Sun
authors.forename: Yukun
authors.initials: Y
authors.lastname: Zhang
authors.forename: Shuhua
authors.initials: S
authors.lastname: Yang
authors.forename: Jie
authors.initials: J
authors.lastname: Jia
authors.forename: Shunan
authors.initials: S
authors.lastname: Ye
authors.forename: Dong
authors.initials: D
authors.lastname: Chen
authors.forename: Fengbo
authors.initials: F
authors.lastname: Mo
mesh_terms: D000704:Analysis of Variance; D002450:Cell Communication / Q000502:physiology*; D018929:Cell Culture Techniques; D020219:Chondrogenesis / Q000502:physiology*; D018031:Connexin 43 / Q000378:metabolism*; D017931:DNA Primers / Q000235:genetics; D036681:Fluorescence Recovery After Photobleaching; D017629:Gap Junctions / Q000502:physiology*; D055428:Growth Differentiation Factor 5 / Q000235:genetics / Q000378:metabolism*; D019850:Heptanol / Q000378:metabolism; D006801:Humans; D007150:Immunohistochemistry; D059630:Mesenchymal Stem Cells / Q000502:physiology*; D060888:Real-Time Polymerase Chain Reaction; D015533:Transcriptional Activation / Q000502:physiology
chemical_list: D018031:Connexin 43; D017931:DNA Primers; C525893:GDF5 protein, human; C500216:GJA1 protein, human; D055428:Growth Differentiation Factor 5; D019850:Heptanol