Regeneration Of Segmental Defects Research Articles

253. ESOPHAGEAL REGENERATION FOLLOWING IMPLANTATION OF A TISSUE ENGINEERED ESOPHAGEAL IMPLANT: UNDERSTANDING THE REGENERATION TIME COURSE AND TRANSLATION TO THE CLINIC

Abstract Diseases of the esophagus, damage of the esophagus due to injury or congenital defects during fetal esophageal development, i.e., Esophageal Atresia (EA), typically require surgical intervention to restore esophageal continuity. The development of novel tissue engineered tubular structures would improve the treatment options for these conditions by providing an alternative that is organ sparing and can be manufactured to fit the exact dimensions of the defect. Biostage has developed an autologous tissue engineered Esophageal Implant, composed of an electrospun cellular scaffold seeded with autologous mesenchymal stromal cells, that can stimulate repair and regeneration of segmental defects in the esophagus following surgical implantation. Biostage has received approval from the FDA on an Investigational New Drg (IND) application for a clinical trial in adults requiring esophageal reconstruction. A discussion of the regeneration time course, in adults and in a pediatric model as well as the critical features of the IND, the Regulatory process for treating adults with the intention to treat Esophageal Atresia and the clinical study parameters highlight the novelty of this tissue engineered combination product.

The effect of different coatings on bone response and degradation behavior of porous magnesium-strontium devices in segmental defect regeneration

Regeneration of long-bone segmental defects remains a challenge for orthopedic surgery. Current treatment options often require several revision procedures to maintain acceptable alignment and achieve osseous healing. A novel hollow tubular system utilizing magnesium-strontium (Mg–Sr) alloy with autogenous morselized bone filled inside to repair segmental defects was developed. To improve the corrosion and biocompatible properties, two coatings, Ca–P and Sr–P coatings, were prepared on surface of the implants. Feasibility of applying these coated implants was systematically evaluated in vitro and in vivo, and simultaneously to have a better understanding on the relationship of degradation and bone regeneration on the healing process. According to the in vitro corrosion study by electrochemical measurements, greater corrosion resistance was obtained for Ca–P coated sample, and attributed to the double-layer protective structure. The cytotoxicity and alkaline phosphatase (ALP) assays demonstrated enhanced bioactivity for Sr–P coated group because of the long-lasting release of beneficial Sr2+. At 12 weeks post-implantation with Mg–Sr alloy porous device, the segmental defects were effectively repaired with respect to both integrity and continuity. In addition, compared with the Ca–P coated implant, the Sr–P coated implant was more proficient at promoting bone formation and mineralization. In summary, the Sr–P coated implants have bioactive properties and exceptional durability, and promote bone healing that is close to the natural rate, implying their potential application for the regeneration of segmental defects.

Regeneration of segmental defects in metatarsus of sheep with vascularized and customized 3D-printed calcium phosphate scaffolds

Although autografts are considered to be the gold standard treatment for reconstruction of large bone defects resulting from trauma or diseases, donor site morbidity and limited availability restrict their use. Successful bone repair also depends on sufficient vascularization and to address this challenge, novel strategies focus on the development of vascularized biomaterial scaffolds. This pilot study aimed to investigate the feasibility of regenerating large bone defects in sheep using 3D-printed customized calcium phosphate scaffolds with or without surgical vascularization. Pre-operative computed tomography scans were performed to visualize the metatarsus and vasculature and to fabricate customized scaffolds and surgical guides by 3D printing. Critical-sized segmental defects created in the mid-diaphyseal region of the metatarsus were either left empty or treated with the 3D scaffold alone or in combination with an axial vascular pedicle. Bone regeneration was evaluated 1, 2 and 3 months post-implantation. After 3 months, the untreated defect remained non-bridged while the 3D scaffold guided bone regeneration. The presence of the vascular pedicle further enhanced bone formation. Histology confirmed bone growth inside the porous 3D scaffolds with or without vascular pedicle inclusion. Taken together, this pilot study demonstrated the feasibility of precised pre-surgical planning and reconstruction of large bone defects with 3D-printed personalized scaffolds.

Tubular open‐porous β‐tricalcium phosphate polycaprolactone scaffolds as guiding structure for segmental bone defect regeneration in a novel sheep model

Large segmental bone defect reconstruction with sufficient functional restoration is one of the most demanding challenges in orthopaedic surgery. Available regenerative treatment options, as the vascularized bone graft transfer, the Masquelet technique or the Ilizarov distraction osteogenesis, are associated with specific indications and distinct limitations. As an alternative, a hollow cylindrical ceramic-polymer composite scaffold (β-tricalcium phosphate and poly-lactid co-ε- caprolactone), facilitating a strong surface guiding effect for tissue ingrowth (group 1; n = 6) was investigated here. In combination with an additional autologous, cancellous bone graft filling, the scaffold's ability to work as an open-porous membrane to improve the defect healing process was analysed (group 2; n = 6). A novel model of a critical size (40 mm) tibia osteotomy defect stabilized with an external hybrid-ring fixator, was established in sheep. Segmental defect regeneration and tissue organization in relation to the scaffold were analysed radiologically, (immune-) histologically, and with second-harmonic generation imaging 12 weeks after surgery. The scaffold's tubular shape and open-porous structure controlled the collagen fibre orientation within the bone defect and guided the following mineralization process along the scaffold surface. In combination with the osteoinductive stimulus, a unilateral bony bridging of the critically sized defect was achieved in one third of the animals. The external hybrid-ring fixator was appropriate for large segmental defect stabilization in sheep.

Electrospun Yarn Reinforced NanoHA Composite Matrix as a Potential Bone Substitute for Enhanced Regeneration of Segmental Defects.

Nanohydroxyapatite (nanoHA) is a well-established synthetic bone substitute with excellent osteoconduction and osteointegration. However, brittleness coupled with slow degradation curtails its load-bearing and bone regeneration potential, respectively. To address these limitations, nanoHA composite matrix reinforced with electrospun fibrous yarns was fabricated and tested in vitro and in vivo. Different weight percentages (5, 10, 15 wt%) and varying lengths (short and continuous) of poly(l-lactic acid) yarns were randomly dispersed in a gelatinous matrix containing nanoHA. This significantly improved the compressive strength as well as work of fracture, especially for continuous yarns at high weight percentages (10 and 15 wt%). Incorporation of yarns did not adversely affect the pore size (50-350 μm) or porosity of the scaffolds as well as the in vitro cellular response. Finally, when tested in a critical-sized femoral segmental defect in rat, the nanocomposite scaffolds induced osteoblast cell infiltration at 2 months that subsequently underwent increased mature lamellar bone formation at 4 months, in both the mid and peripheral defect regions. Histomorphometric analysis demonstrated that new bone formation and biomaterial degradation were significantly enhanced in the composite scaffold when compared to commercially available HA. Overall, the composite matrix reinforced with electrospun yarns proved to be a potential bone substitute having an appropriate balance between mechanical strength, porosity, biodegradation, and bone regeneration ability.

A Tissue Engineering Solution for Segmental Defect Regeneration in Load-Bearing Long Bones

The reconstruction of large defects (>10 mm) in humans usually relies on bone graft transplantation. Limiting factors include availability of graft material, comorbidity, and insufficient integration into the damaged bone. We compare the gold standard autograft with biodegradable composite scaffolds consisting of medical-grade polycaprolactone and tricalcium phosphate combined with autologous bone marrow-derived mesenchymal stem cells (MSCs) or recombinant human bone morphogenetic protein 7 (rhBMP-7). Critical-sized defects in sheep--a model closely resembling human bone formation and structure--were treated with autograft, rhBMP-7, or MSCs. Bridging was observed within 3 months for both the autograft and the rhBMP-7 treatment. After 12 months, biomechanical analysis and microcomputed tomography imaging showed significantly greater bone formation and superior strength for the biomaterial scaffolds loaded with rhBMP-7 compared to the autograft. Axial bone distribution was greater at the interfaces. With rhBMP-7, at 3 months, the radial bone distribution within the scaffolds was homogeneous. At 12 months, however, significantly more bone was found in the scaffold architecture, indicating bone remodeling. Scaffolds alone or with MSC inclusion did not induce levels of bone formation comparable to those of the autograft and rhBMP-7 groups. Applied clinically, this approach using rhBMP-7 could overcome autograft-associated limitations.

Calcium Alkaline Phosphate Scaffolds for Bone Regeneration 3D-Fabricated by Additive Manufacturing

Calcium alkaline phosphate granulates can be used for substitution of several bone defects but for the reconstruction of large skeletal parts in the maxillofacial and orthopaedic fields fitted scaffolds are preferable. Within the additive manufacturing methods, the 3D printing process offers exciting opportunities to generate defined porous scaffolds. We used a R1 printer from ProMetal Company, USA, for producing scaffolds directly from a ceramic powder. For this direct free form fabrication technology the powder has to possess a lot of specific properties both for the generation of a stable green body and also for the subsequent sintering preparation. For this printing process we prepared different granules in a fluidized bed process containing Ca2KNa(PO4)2as main crystalline phase. Granules were characterized by different methods and several sieve fractions were used for preparing disc like and cylindrical parts. The suitability of granules for this printing process was determined by porosity and strength of produced bodies. Next to granules’ performance both of these properties can be directly influenced by 3D printing process parameters. With knowledge of suitable process parameters scaffolds with different porosity in a respective desired design can be created. In this study, cylindrical scaffolds with graded porosity were produced for bone regeneration of segmental defects in maxillofacial surgery and dental implantology by tissue engineering.

Enhanced healing of goat femur‐defect using BMP7 gene‐modified BMSCs and load‐bearing tissue‐engineered bone

Segmental defect regeneration is still a clinical challenge. In this study, we investigated the feasibility of bone marrow stromal cells (BMSCs) infected with adenoviral vector containing the bone morphogenetic protein 7 gene (AdBMP7) and load-bearing to enhance bone regeneration in a critically sized femoral defect in the goat model. The defects were implanted with AdBMP7-infected BMSCs/coral (BMP7 group) or noninfected BMSCs/coral (control group), respectively, stabilized with an internal fixation rod and interlocking nails. Bridging of the segmental defects was evaluated by radiographs monthly, and confirmed by biomechanical tests. Much callus was found in the BMP7 group, and nails were taken off after 3 months of implantation, indicating that regenerated bone in the defect can be remodeled by load-bearing, whereas after 6 months in control group. After load-bearing, it is about 5 months; the mechanical property of newly formed bone in the BMP7 group was restored, but 8 months in control group. Our data suggested that the BMP7 gene-modified BMSCs and load-bearing can promote bone regeneration in segmental defects.

Segmental bone regeneration using a load-bearing biodegradable carrier of bone morphogenetic protein-2

Segmental defect regeneration has been a clinical challenge. Current tissue-engineering approach using porous biodegradable scaffolds to delivery osteogenic cells and growth factors demonstrated success in facilitating bone regeneration in these cases. However, due to the lack of mechanical property, the porous scaffolds were evaluated in non-load bearing area or were stabilized with stress-shielding devices (bone plate or external fixation). In this paper, we tested a scaffold that does not require a bone plate because it has sufficient biomechanical strength. The tube-shaped scaffolds were manufactured from poly(propylene) fumarate/tricalcium phosphate (PPF/TCP) composites. Dicalcium phosphate dehydrate (DCPD) were used as bone morphogenetic protein-2 (BMP-2) carrier. Twenty-two scaffolds were implanted in 5mm segmental defects in rat femurs stabilized with K-wire for 6 and 15 weeks with and without 10 microg of rhBMP-2. Bridging of the segmental defect was evaluated first radiographically and was confirmed by histology and micro-computer tomography (microCT) imaging. The scaffolds in the BMP group maintained the bone length throughout the duration of the study and allow for bridging. The scaffolds in the control group failed to induce bridging and collapsed at 15 weeks. Peripheral computed tomography (pQCT) showed that BMP-2 does not increase the bone mineral density in the callus. Finally, the scaffold in BMP group was found to restore the mechanical property of the rat femur after 15 weeks. Our results demonstrated that the load-bearing BMP-2 scaffold can maintain bone length and allow successfully regeneration in segmental defects.

Activation of extracellular signal-regulated kinase (ERK) and p38 kinase in shock wave-promoted bone formation of segmental defect in rats

Extracorporeal shock waves (ESW) have recently been used in bone repair. Extracellular signal-regulated kinase (ERK) and p38 kinase are found to act as important mediators for osteogenic factor and mechanical-stimulated proliferation and differentiation of bone-forming cells. A previous study reported that ESW promoted healing of segmental defects in rats by inducing bone morophogenetic proteins (Bone 32 (2003) 387–396) and stimulating osteogenic differentiation of mesenchymal stem cells. In this study, we found that ERK and p38 activation was involved in ESW-augmented bone regeneration of segmental defects. ESW treatment (0.16 mJ/mm 2, 1 Hz, 500 impulses) rapidly promoted [ 3H]-thymidine uptake in 1 day and progressively increased alkaline phosphatase activity, collagen I, II, and osteocalcin synthesis in callus organ culture within 14 days after treatment. Results of [γ- 32P]-phosphotransferase activity assay showed that ERK and p38 in calluses were rapidly activated 1 day and 7 days after ESW treatment, respectively. Histological observation showed that segmental defects subjected to ESW treatment underwent typical bone formation (mesenchymal cell aggregation, hypertrophic cartilage, and endochondral/intramembrane ossification). Intensive bone formation coincided with evident expression of phosphorylated ERK and p38. Moreover, expression of phosphorylated ERK persisted in mesenchymal, chondral, and osteoblastic cells at newly developed bone and cartilage, and the expression of activated p38 was evident on chondral cells located at hypertrophic cartilage. Our findings suggest that mitogen-activated protein kinases (MAPK) regulate the stimulation of biophysical ESW, triggering mitogenic and osteogenic responses in the defects. ERK phosphorylation is active throughout the period of ESW-induced bone regeneration. p38 activation most likely plays an important role in signaling cartilage formation in callus.