Treatment of osteochondral joint surface defects: towards engineering a bio-artificial joint.

Current clinical treatments for large long bone and osteochondral defects are limited by either tissue availability or incapacity to regenerate tissues to their original functionality. Regenerative medicine, and more specifically tissue engineering (TE), shows great potential to meet this clinical need by creating in vitro implants able to heal large bone and osteochondral defects. The current PhD project aimed to develop a developmentally inspired microspheroid-based process for the regeneration of bone and cartilage. 

Mimicking the developmental stages of long bones (endochondral ossification) is expected to generate in vitro intermediate tissues that remodel into functional tissues upon implantation; an approach called “developmental engineering”. Many aspects of the endochondral ossification process are recapitulated during fracture healing by the formation of a cartilage intermediate, termed “soft callus”. Generation of hypertrophic cartilaginous in vitro pellets (1-2 mm) have previously been shown to form bone ossicles upon ectopic implantation. However, diffusion and assembly limitations are prone to hamper translation towards scalable production. In this thesis, the developmental engineering concept was taken to the next level through the assembly of smaller building-blocks (100-200 µm) to create living implants.

Human periosteum derived cells (hPDCs) were seeded in microwells to generate microspheroids followed by chondrogenic differentiation into “callus organoids”. The organoids attained autonomy and exhibited the capacity to form ectopic bone microorgans in vivo. This potency was linked to specific gene signatures mimicking those found in developing and healing long bones. Furthermore, “callus organoids” spontaneously bioassembled in vitro into engineered tissues which were able to heal murine critical‐sized long bone defects. The “callus organoids” were subsequently assembled with “cartilage microtissues” derived from human induced pluripotent stem cells (iPSCs). The dual constructs formed osteochondral-like tissue units upon ectopic implantation, hence revealing their distinct biological potency and maintained zonal patterning. These data demonstrate the possibility to generate complex in vitro tissues using living building blocks for regeneration of bone and cartilage. However, certain considerations are required regarding building block fusion and size. Fusion and spreading capacity of hPDC-derived microtissues decreased with prolonged chondrogenic differentiation. Additionally, assembly of microtissues at an early time-point in vitro resulted in the formation of non-differentiated central regions, if the final size/thickness was not limited to ≤1mm.

The developmental engineering approach described in this thesis was proven successful in a small animal model (mouse) and demonstrated a proof-of-concept for the possibility of scalable spheroid-based constructs for the regeneration of bone and cartilage. Further actions with regard to scale-up and building-block characterization should be taken to enable large animal studies and future clinical translation.