The effect of implantation on scaffoldless three-dimensional engineered bone constructs

Abstract

Our laboratory has previously developed scaffoldless engineered bone constructs (EBC). Bone marrow stromal cells (BMSC) were harvested from rat femur and cultured in medium that induced osteogenic differentiation. After reaching confluence, the monolayer of cells contracted around two constraint points forming a cylinder. EBCs were placed in small diameter (0.5905 x 0.0625 in.) or large diameter (0.5905 x 0.125 in.) silicone tubing and implanted intramuscularly in the hind limb of a rat. Bone mineral content (BMC) of the EBC was analyzed before implantation and at 1 and 2 mo following implantation and compared to that of native femur bone at different stages of development. Negligible BMC was observed in E-20 femur or EBCs prior to implantation. One-month implantation in both small and large tubing increased BMC in the EBC. BMC of EBC from large tubing was greater than in 14 d rat neonatal femurs, but was 2% and 3% of BMC content in adult bone after 1 and 2 mo of implantation, respectively. Alizarine Red and osteopontin staining of the EBCs before and after implantation confirmed increased bone mineralization in the implanted EBCs. Implanted EBCs also had extensive vascularization. Our data suggest that BMSC can be successfully used for the generation of scaffoldless EBC, and this model can be potentially used for the generation of autologous bone transplants in humans.

Isolation, characterization and osteogenic differentiation of adipose-derived stem cells: from small to large animal models

“…In particular, rabbits are useful for screening several experimental conditions because of their low cost of maintenance, whereas pigs represent a more predictive model because of their greater similarity to human beings with regard to diet, weight and working load. The femoral defect in the rat also represents a valid model in the orthopaedic field (Cuomo et al 2009;Smietana et al 2009). However, we are interested in an autologous cell-implant model, which we believe will not be easy to obtain in rat.…”

Tissue-Engineered Tendon for Enthesis Regeneration in a Rat Rotator Cuff Model

“…15,17 Growth medium (GM) consisted of Dulbecco's modified Eagle's medium (DMEM; Gibco, Cat# 10565-042) supplemented with 20% Fetal Bovine Serum (FBS; Gibco, Cat# 10437-028), 2% antibiotic-antimycotic (Sigma; Cat# A9909), 600 ng/mL bFGF, and 40 ng/mL dexamethasone. Differentiation medium (DM) consisted of DMEM supplemented with 7% Horse Serum Albumin (HS; Gibco, Cat# 16050-122) and 2% antibiotic-antimycotic, plus the addition of 0.13 mg/mL ascorbic acid-2 phosphate, 0.05 mg/mL L-proline, 400 ng/mL dexamethasone, and 2 ng/mL transforming growth factor-beta (TGF-β; Peprotech, Cat# 100-21).…”

“…15,17 Briefly, rat femora were removed from the animal and cut at the proximal and distal metaphysis. The marrow contents were extracted through centrifugation (900 g for 10 min) and cultured on 100 mm tissue-culture-treated polystyrene plates (BD Falcon).…”

“…14,15,17 Briefly, mesenchymal stem cells (MSCs) were plated onto 100 mm cell culture dishes at 21 × 10 3 cells/cm 2 and suspended in 8 mL of GM supplemented with 0.13 mg/mL ascorbic acid-2 phosphate and 0.05 mg/mL L-proline. The dishes were placed in a 37°C, 5% CO 2 incubator and medium was changed every 2–3 days.…”

Engineering Muscle Constructs for the Creation of Functional Engineered Musculoskeletal Tissue

“…Therefore, scaffold-free constructs may be more appropriate for chronic reconstruction rather than acute repair of traumatic injury. Despite these current limitations, scaffold-free technologies have been used to fabricate multiphasic constructs of muscle–tendon–bone and bone–ligament used successfully as skeletal tissue replacements tissues for VML in small rodent models and ligament in large animal models [18,21,26,79]. …”