Laser perforation and cell seeding improve bacterial nanocellulose as a potential cartilage implant in the in vitro cartilage punch model
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Bacterial nanocellulose (BNC) shows high biocompatibility as wound dressing or dura mater, blood vessel, and cartilage implant. Three-dimensional perforation (3-D-∅) favors migration of chondrocytes into the BNC and cartilage matrix formation. Thus, the regenerative capacity of 3-D-∅ BNC implants was tested in a standardized bovine cartilage punch model. Cartilage rings containing a central defect with an outer diameter of 6 mm and an inner diameter of 2 mm were prepared from the trochlear groove (femur-patellar articulation site). Three-D-∅ BNC implants (cell-free or cell-loaded) were cultured inside the cartilage rings for up to 12 weeks. Cartilage-BNC-constructs were then investigated by histology (hematoxylin/eosin; safranin O) and immunohistology (aggrecan, collagens 1 and 2), as well as for protein content, RNA expression, and implant push-out force. Cartilage-BNC-constructs remained vital with preserved matrix integrity during culture and almost no loss of matrix-bound proteoglycan (aggrecan) or collagen 2 from ‘host’ cartilage (with very limited quantities of collagen 1). Interestingly, 3-D-∅ BNC implants displayed: (1) significantly increased superficial, but also 3-D cell seeding over time (cell-loaded significantly earlier than cell-free); (2) progressively increased aggrecan/collagen 1 and collagen 2/collagen 1 mRNA ratios, as well as aggrecan and collagen 2 protein levels; and (3) significantly increased push-out forces over time (cell-loaded). Progressively increasing cell seeding and chondrogenic differentiation suggest beginning cartilage regeneration of the 3-D-∅ BNC in this model system, and indicate an excellent potential of 3-D-∅ BNC as a cartilage replacement material. Cell-loading may favor implant performance by accelerating cell colonization.
KeywordsBovine cartilage punch model Three-dimensional perforation Bacterial nanocellulose Articular cartilage Implant push-out force
The authors are grateful to Cordula Müller, Bärbel Ukena, Ulrike Körner, and Tobias Fiedler (Experimental Rheumatology Unit, Jena University Hospital) for expert technical assistance and qRT-PCR analyses, Maren Siedentop, Daniela Warnecke, and Fabian Holzner (Biomechanics Laboratory, Klinikum rechts der Isar, Technische Universität München) for expert biomechanical testing of implant push-out forces, Hannes Ahrem (Jenpolymer Materials Ltd & Co. KG, Jena, Germany) for producing the BNC hydrogels, and Daniel Conrad and Hartmut Müller (Günter-Köhler-Institut für Fügetechnik und Werkstoffprüfung GmbH, Jena) for laser structuring of the BNC hydrogels.
We gratefully acknowledge the partial financial support of the Bundesministerium für Bildung und Forschung (BMBF), Grant References 13N12601 and 0315577C.
Compliance with ethical standards
Conflicts of interest
The author(s) declared no potential conflicts of interest with respect to the research, authorship, and/or publication of this article.
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