**Anne Bernhardt \*, Birgit Paul and Michael Gelinsky**

Centre for Translational Bone, Joint and Soft Tissue Research, University Hospital Carl Gustav Carus and Faculty of Medicine of Technische Universität Dresden, Fetscherstraße 74, 01307 Dresden, Germany; birgit.paul@tu-dresden.de (B.P.); michael.gelinsky@tu-dresden.de (M.G.)

**\*** Correspondence: anne.bernhardt@tu-dresden.de; Tel.: +49-351-458-6692

Received: 10 January 2018; Accepted: 10 March 2018; Published: 13 March 2018

**Abstract:** Background: Collagens of marine origin are applied increasingly as alternatives to mammalian collagens in tissue engineering. The aim of the present study was to develop a biphasic scaffold from exclusively marine collagens supporting both osteogenic and chondrogenic differentiation and to find a suitable setup for in vitro chondrogenic and osteogenic differentiation of human mesenchymal stroma cells (hMSC). Methods: Biphasic scaffolds from biomimetically mineralized salmon collagen and fibrillized jellyfish collagen were fabricated by joint freeze-drying and crosslinking. Different experiments were performed to analyze the influence of cell density and TGF-β on osteogenic differentiation of the cells in the scaffolds. Gene expression analysis and analysis of cartilage extracellular matrix components were performed and activity of alkaline phosphatase was determined. Furthermore, histological sections of differentiated cells in the biphasic scaffolds were analyzed. Results: Stable biphasic scaffolds from two different marine collagens were prepared. An in vitro setup for osteochondral differentiation was developed involving (1) different seeding densities in the phases; (2) additional application of alginate hydrogel in the chondral part; (3) pre-differentiation and sequential seeding of the scaffolds and (4) osteochondral medium. Spatially separated osteogenic and chondrogenic differentiation of hMSC was achieved in this setup, while osteochondral medium in combination with the biphasic scaffolds alone was not sufficient to reach this ambition. Conclusions: Biphasic, but monolithic scaffolds from exclusively marine collagens are suitable for the development of osteochondral constructs.

**Keywords:** jellyfish collagen; mineralized salmon collagen; osteochondral tissue engineering; biphasic scaffold; osteochondral medium; alginate
