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Enzymatic Isolation of Articular Chondrons: Is It Much Different Than That of Chondrocytes?
van Mourik, Marloes; Schuiringa, Gerke H; Varion-Verhagen, Liesbeth P; Vonk, Lucienne A; van Donkelaar, Corrinus C; Ito, Keita; Foolen, Jasper.
Afiliación
  • van Mourik M; Orthopaedic Biomechanics and Department of Biomedical Engineering, Eindhoven University of Technology, Eindhoven, The Netherlands.
  • Schuiringa GH; Orthopaedic Biomechanics and Department of Biomedical Engineering, Eindhoven University of Technology, Eindhoven, The Netherlands.
  • Varion-Verhagen LP; Laboratory of Immunoengineering, Department of Biomedical Engineering, Eindhoven University of Technology, Eindhoven, The Netherlands.
  • Vonk LA; Department of Orthopaedics, University Medical Center Utrecht, Utrecht University, Utrecht, The Netherlands.
  • van Donkelaar CC; Orthopaedic Biomechanics and Department of Biomedical Engineering, Eindhoven University of Technology, Eindhoven, The Netherlands.
  • Ito K; Orthopaedic Biomechanics and Department of Biomedical Engineering, Eindhoven University of Technology, Eindhoven, The Netherlands.
  • Foolen J; Orthopaedic Biomechanics and Department of Biomedical Engineering, Eindhoven University of Technology, Eindhoven, The Netherlands.
Tissue Eng Part C Methods ; 29(1): 30-40, 2023 01.
Article en En | MEDLINE | ID: mdl-36576016
In native articular cartilage, chondrocytes (Chy) are completely capsulated by a pericellular matrix (PCM), together called the chondron (Chn). Due to its unique properties (w.r.t. territorial matrix) and importance in mechanotransduction, the PCM and Chn may be important in regenerative strategies. The current gold standard for the isolation of Chns from cartilage dates from 1997. Although previous research already showed the low cell yield and the heterogeneity of the isolated populations, their compositions and properties have never been thoroughly characterized. This study aimed to compare enzymatic isolation methods for Chy and Chns and characterizes the isolation efficiency and quality of the PCM. Bovine articular cartilage was digested according to the 5-h (5H) gold standard Chn isolation method (0.3% dispase +0.2% collagenase II), an overnight (ON) Chn isolation (0.15% dispase +0.1% collagenase II), and an ON Chy isolation (0.15% collagenase II +0.01% hyaluronidase). Type VI collagen staining, fluorescence-activated cell sorting (FACS) analysis, specific cell sorting, and immunohistochemistry were performed using a type VI collagen staining, to study their isolation efficiency and quality of the PCM. These analyses showed a heterogeneous mixture of Chy and Chns for all three methods. Although the 5H Chn isolation resulted in the highest percentage of Chns, the cell yield was significantly lower compared to the other isolation methods. FACS, based on the type VI collagen staining, successfully sorted the three identified cell populations. To maximize Chn yield and homogeneity, the ON Chn enzymatic digestion method should be combined with type VI collagen staining and specific cell sorting. Impact statement Since chondrocytes are highly dependent on their microenvironment for maintaining phenotypic stability, it is hypothesized that using chondrons results in superior outcomes in cartilage tissue engineering. This study reveals the constitution of cell populations obtained after enzymatic digestion of articular cartilage tissue and presents an alternative method to obtain a homogeneous population of chondrons. These data can improve the impact of studies investigating the effect of the pericellular matrix on neocartilage formation.
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Texto completo: 1 Colección: 01-internacional Banco de datos: MEDLINE Asunto principal: Cartílago Articular / Colágeno Tipo VI Límite: Animals Idioma: En Revista: Tissue Eng Part C Methods Asunto de la revista: BIOTECNOLOGIA / HISTOLOGIA Año: 2023 Tipo del documento: Article País de afiliación: Países Bajos

Texto completo: 1 Colección: 01-internacional Banco de datos: MEDLINE Asunto principal: Cartílago Articular / Colágeno Tipo VI Límite: Animals Idioma: En Revista: Tissue Eng Part C Methods Asunto de la revista: BIOTECNOLOGIA / HISTOLOGIA Año: 2023 Tipo del documento: Article País de afiliación: Países Bajos