Cartilage resurfacing potential of PLGA scaffolds loaded with autologous cells from cartilage, fat, and bone marrow in an ovine model of osteochondral focal defect

Abstract

Current developments in tissue engineering strategies for articular cartilage regeneration focus on the design of supportive three-dimensional scaffolds and their use in combination with cells from different sources. The challenge of translating initial successes in small laboratory animals into the clinics involves pilot studies in large animal models, where safety and efficacy should be investigated during prolonged follow-up periods. Here we present, in a single study, the long-term (up to 1 year) effect of biocompatible porous scaffolds non-seeded and seeded with fresh ex vivo expanded autologous progenitor cells that were derived from three different cell sources [cartilage, fat and bone marrow (BM)] in order to evaluate their advantages as cartilage resurfacing agents. An ovine model of critical size osteochondral focal defect was used and the test items were implanted arthroscopically into the knees. Evidence of regeneration of hyaline quality tissue was observed at 6 and 12 months post-treatment with variable success depending on the cell source. Cartilage and BM-derived mesenchymal stromal cells (MSC), but not those derived from fat, resulted in the best quality of new cartilage, as judged qualitatively by magnetic resonance imaging and macroscopic assessment, and by histological quantitative scores. Given the limitations in sourcing cartilage tissue and the risk of donor site morbidity, BM emerges as a preferential source of MSC for novel cartilage resurfacing therapies of osteochondral defects using copolymeric poly-D,L-lactide-co-glycolide scaffolds.

A reproducible method for the isolation and expansion of ovine mesenchymal stromal cells from bone marrow for use in regenerative medicine preclinical studies

“…Conditions for the differentiation assays were adapted to ovine samples using StemPro differentiation media (Gibco) (Table ). Safranin O (Sigma‐Aldrich), Oil Red O (Sigma‐Aldrich), alkaline phosphatase (Takara Bio Inc., Shiga, Japan) and Von Kossa silver nitrate (Sigma‐Aldrich) stainings were performed for the determination of the outcome of the differentiation assays as described elsewhere (Caminal et al, ).…”

“…This is not only because of the multilineage progenitor potential of MSCs, but also because of their capacity to modulate the host microenvironment by secreting a variety of factors, such as cytokines, chemokines or growth factors, which can mediate diverse functions via crosstalk between different cell types (Friedenstein et al, ; Doorn et al, ; Murphy et al, ). Examples of using sheep as a model system include spinal fusion (Goldschlager et al, ), segmental tibial defects (Niemeyer et al, ), chondral/osteochondral repair (Ahern et al, ; Caminal et al, ,b, ; Vives et al, ), tendon and ligament regeneration (Lacitignola et al, ) and osteonecrosis of the femoral head (Velez et al, ), among others.…”

Management of knee osteoarthritis. Current status and future trends

“…As a result, it has improved mechanical strength, biodegradability, biocompatibility, minimal toxicity, and resistance to hydrolysis (Gentile et al, ). PLGA was investigated in AC regeneration, either alone or in combination with other polymers (Caminal et al, ; Chang et al, ; Hsu et al, ; Qi et al, ). Several formations of PLGA materials were proposed, such as hydrogels, scaffolds, microspheres, membranes, and films (Gentile et al, ; Makadia and Siegel, ).…”

Clinical translation of a mesenchymal stromal cell-based therapy developed in a large animal model and two case studies of the treatment of atrophic pseudoarthrosis

“…For the production of ovine MSCs, BM samples were collected from the sternum of the animals and subsequently used for the isolation of mononuclear cells (MNCs) and the establishment of BM‐MSC lines, as described previously (Caminal et al ., ). For the manufacture of a TEP consisting of cell–scaffold composites, 6 × 10 6 BM‐MSCs were incubated with 1 cm 3 of trabecular bone particles (Osteoplant; Bioteck) in a single‐use bioreactor (Hexascreen Culture Technologies S.L., Barcelona, Spain) stirred at 200 rpm overnight in 12 ml of Dulbecco's modified Eagle's medium (DMEM; Gibco, Carlsbad, CA, USA) supplemented with 10% pooled sheep serum (Caminal et al ., ; Soley et al ., ). The number of cells loaded onto the scaffold and the colonization rate (%) were calculated indirectly by counting the non‐seeded cells present in the supernatant.…”