AIM: The aim of this work was to determine the pathways implicated in the mechanosensing of chondrocytes. METHODS: Rat chondrocytes were cultured in collagen hydrogels of different stiffness (2-20 Pa) in normoxia and hypoxia, in monolayer and embedded inside hydrogels. First, chondrocyte were cultured on hydrogels in the presence of antibodies to block integrins. Second, custom RT-PCR array plates and western blot were used to detect changes in expression of genes implicated in downstream signalling pathways. RESULTS: The results allowed us to demonstrate the mechanosensing of chondrocytes for changes in stiffness in the range of Pascals. We also identified Non-Muscle Myosin II (NMMII) and integrins α1, β1 and β3 as participants in the mechanosensing, since their blockade inhibits the sensing of the stiffness, and they are up-regulated in the process. RT-PCR arrays and western blot detected up-regulation of Paxillin, RhoA, Fos, Jun and Sox9. We detected no expression of Src in the monolayer cultures, but we found a role for this protein in 3D. The expression of HIF-1α was not modified under normoxia but was found to participate under hypoxia. Focal Adhesion Kinase (FAK), showed a direct relationship with the expression of Aggrecan in hypoxia and an inverse one in normoxia. Finally, immunofluorescence analysis located the expression of factors AP-1, Sox-9 and HIF-1α inside the cell nuclei and RhoA, Src, Paxillin and FAK close to the cytoplasmic membrane. CONCLUSIONS: We determined here some of the genes that are up-regulated during the process of chondrocyte mechanosensing.
AIM: The aim of this work was to determine the pathways implicated in the mechanosensing of chondrocytes. METHODS:Rat chondrocytes were cultured in collagen hydrogels of different stiffness (2-20 Pa) in normoxia and hypoxia, in monolayer and embedded inside hydrogels. First, chondrocyte were cultured on hydrogels in the presence of antibodies to block integrins. Second, custom RT-PCR array plates and western blot were used to detect changes in expression of genes implicated in downstream signalling pathways. RESULTS: The results allowed us to demonstrate the mechanosensing of chondrocytes for changes in stiffness in the range of Pascals. We also identified Non-Muscle Myosin II (NMMII) and integrins α1, β1 and β3 as participants in the mechanosensing, since their blockade inhibits the sensing of the stiffness, and they are up-regulated in the process. RT-PCR arrays and western blot detected up-regulation of Paxillin, RhoA, Fos, Jun and Sox9. We detected no expression of Src in the monolayer cultures, but we found a role for this protein in 3D. The expression of HIF-1α was not modified under normoxia but was found to participate under hypoxia. Focal Adhesion Kinase (FAK), showed a direct relationship with the expression of Aggrecan in hypoxia and an inverse one in normoxia. Finally, immunofluorescence analysis located the expression of factors AP-1, Sox-9 and HIF-1α inside the cell nuclei and RhoA, Src, Paxillin and FAK close to the cytoplasmic membrane. CONCLUSIONS: We determined here some of the genes that are up-regulated during the process of chondrocyte mechanosensing.
Authors: Tommy S de Windt; Jeanine A A Hendriks; Xing Zhao; Lucienne A Vonk; Laura B Creemers; Wouter J A Dhert; Mark A Randolph; Daniel B F Saris Journal: Stem Cells Transl Med Date: 2014-04-24 Impact factor: 6.940
Authors: Rene Olivares-Navarrete; Erin M Lee; Kathryn Smith; Sharon L Hyzy; Maryam Doroudi; Joseph K Williams; Ken Gall; Barbara D Boyan; Zvi Schwartz Journal: PLoS One Date: 2017-01-17 Impact factor: 3.240
Authors: Elisabeth B Wondimu; Kirsty L Culley; Justin Quinn; Jun Chang; Cecilia L Dragomir; Darren A Plumb; Mary B Goldring; Miguel Otero Journal: Sci Rep Date: 2018-04-24 Impact factor: 4.379