This review provides an update on cartilage tissue engineering with particular focus on the head and neck. It is aimed at scientists and clinicians who are interested in tissue engineering and its clinical applicability. Principal tissue engineering strategies are summarized in the first part of this review. In the second part, current clinical approaches to auricular, nasal and tracheal reconstruction are discussed from a surgical perspective. By this approach, the requirements for clinical applicability are outlined and new insight into relevant aims of research is given to accelerate the transfer from bench to bedside.

Ideal tissue engineering frameworks should be both an optimal biological microenvironment and a shape and stability providing framework. In this study we tried to combine the advantages of cell-derived artificial extracellular matrix (ECM) with those of 3D printed polycaprolactone (PCL) scaffolds. In Part A, both chondrogenic and osteogenic ECMs were produced by human adipose derived stem cells (hASCs) on 3D-printed PCL scaffolds and then decellularized to create cell free functionalized PCL scaffolds, named acPCL and aoPCL respectively. The decellularization resulted in a significant reduction of the DNA content as well as the removal of nuclei while the ECM was largely preserved. In Part B the bioactivation and the effect of the ac/aoPCL scaffolds on the proliferation, differentiation, and gene expression of hASCs was investigated. The ac/aoPCL scaffolds were found to be non-toxic and allow good adhesion, but do not affect proliferation. In the in vitro investigation of cartilage regeneration, biochemical analysis showed that acPCL scaffolds have an additional effect on chondrogenic differentiation as gene expression analysis showed markers of cartilage hypertrophy. The aoPCL showed a large influence on the differentiation of hASCs. In control medium they were able to stimulate hASCs to produce calcium alone and all genes relevant investigated for osteogenesis were significantly higher expressed on aoPCL than on unmodified PCL. Therefore, we believe that ac/aoPCL scaffolds have a high potential to improve regenerative capacity of unmodified PCL scaffolds and should be further investigated.

Background: Cartilage shortage is a major problem in facial reconstructive surgery. Prior studies have shown that decellularized porcine nasal septal cartilage (DPNC) seeded with primary human nasal chondrocytes enabled cartilage regeneration and showed potential as a replacement material for nasal cartilage. Since adipose tissue‐derived stem cells (ASCs) are easily accessible and almost abundantly available, they appear to be a promising alternative to limited chondrocytes making the combination of DPNC and ASCs a feasible approach towards clinical translation. Thus, this study was intended to investigate the interactions between ASCs and DPNC in an in vitro model.Methods: DPNCs were seeded and 3D‐cultured with primary human ASCs that were priorly characterized with trilineage differentiation and flow cytometry. Cell vitality and proliferation were evaluated by Live‐Dead, alamarBlue, and PicoGreen assays. Chondrogenic differentiation was examined by DMMB assay and cryosectioning‐based histology. Cell invasion within DPNC was visualized and quantified by fluorescent histology (DAPI, Phalloidin).Results: ASCs showed good adherence to DPNC and Live‐Dead assay proved their viability over 2 weeks. AlamarBlueassay showed an increase in metabolic activity compared to 2D cultures, and PicoGreen assay demonstrated an increase of cell number within DPNC over time. Biochemical assays and histology added evidence of chondrogenic differentiation of 3D‐cultured ASCs under the influence of chondrogenic induction medium. Fluorescent image analysis showed a significant increase of cell‐occupied areas of scaffolds over time (P < .05).Conclusions: DPNC scaffolds provided a suitable environment for ASCs that allowed good cell vitality, high proliferation, and chondrogenic differentiation. Thus, the use of ASCs and DPNC yields a promising alternative to the use of primary human chondrocytes. For facial cartilage tissue engineering, we regard ASCs as an attractive alternative to human nasal chondrocytes due to their better accessibility and availability. Further research will be necessary to determine long‐term effects and in vivo outcomes of ASCs and DPNC in cartilage regeneration of the face.

The use of alloplastic materials instead of autologous cartilage grafts offers a new perspective in craniofacial reconstructive surgery. Particularly for regenerative approaches, customized implants enable the surgeon to restore the cartilaginous framework of the ear without donor site morbidity. However, high development and production costs of commercially available implants impede clinical translation. For this reason, the usability of a low-cost 3D printer (Ultimaker 2+) as an inhouse-production tool for cheap surgical implants was investigated. The open software architecture of the 3D printer was modified in order to enable printing of biocompatible and biologically degradable polycaprolactone (PCL). Firstly, the printing accuracy and limitations of a PCL implant were compared to reference materials acrylonitrile butadiene styrene (ABS) and polylactic acid (PLA). Then the self-made PCL-scaffold was seeded with adipose-tissue derived stem cells (ASCs), and biocompatibility was compared to a commercially available PCL-scaffold using a cell viability staining (FDA/PI) and a dsDNA quantification assay (PicoGreen). Secondly, porous and solid patient-customized ear constructs were manufactured from mirrored CT-imagining data using a computer-assisted design (CAD) and computer-assisted manufacturing (CAM) approach to evaluate printing accuracy and reproducibility. The results show that printing of a porous PCL scaffolds was possible, with an accuracy equivalent to the reference materials at an edge length of 10 mm and a pore size of 0.67 mm. Cell viability, adhesion, and proliferation of the ASCs were equivalent on self-made and the commercially available PCL-scaffolds. Patient-customized ear constructs could be produced well in solid form and with limited accuracy in porous form from all three thermoplastic materials. Printing dimensions and quality of the modified low-cost 3D printer are sufficient for selected tissue engineering applications, and the manufacturing of personalized ear models for surgical simulation at manufacturing costs of EUR 0.04 per cell culture scaffold and EUR 0.90 (0.56) per solid (porous) ear construct made from PCL. Therefore, in-house production of PCL-based tissue engineering scaffolds and surgical implants should be further investigated to facilitate the use of new materials and 3D printing in daily clinical routine.

Cartilage regeneration remains a significant challenge in the field of regenerative medicine. Acellular matrix-based cartilage tissue regeneration offers an innovative approach to repairing cartilage defects by providing a scaffold for new tissue growth. Its significance lies in its potential to restore joint function, mitigate pain, and improve the quality of life for patients suffering from cartilage-related injuries and conditions. Recent advances in acellular matrix-based cartilage regeneration have focused on enhancing scaffold properties for improved cell adhesion, proliferation, and differentiation. Moreover, several scaffold techniques such as combining ADM and ACM with cartilage tissue, as well as biphasic scaffolding enjoy rising research activity. Incorporating bioactive factors and advanced manufacturing techniques holds promise for producing more biomimetic scaffolds, advancing efficient cartilage repair and regeneration. Obstacles in acellular matrix-based cartilage regeneration include achieving proper integration with surrounding tissue and ensuring long-term durability of the regenerated cartilage. Further, issues such as high costs and limited availability of suitable cells for scaffold seeding must be considered. The heterogeneity and limited regenerative capabilities of cartilage need to be addressed for successful clinical translation. Research should focus on exploring advanced biomaterials and developing new techniques, regarding easily reproductible scaffolds, ideally constructed from clinically validated and readily available commercial products. Findings underline the potential of AM-based approaches, especially rising exploration of tissue-derived ADM and ACM. In future, the primary objective should not only be the regeneration of small cartilage defects, but rather focus on fully regenerating a joint or larger cartilage defect.