Thomas Engstrand

Injectable cell-free template for bone-tissue formation

Here we present a novel injectable hydrogel which forms a template for de novo formation of bone tissue. Hydrogel formation takes place in situ in less than 1 min by the cross-linking of multifunctional hyaluronic acid and polyvinyl alcohol derivatives. Endogenous cells are recruited in vivo by incorporating bone morphogenetic protein-2 (BMP-2), a powerful promoter for osteogenic differentiation. The hydrogel was evaluated in vitro by performing a cell viability test and a release study and in vivo by a rat ectopic model. Examination by X-ray, microcomputed tomography, and histology revealed a significant bone formation at the target site for gels containing BMP-2, and a complete degradation was observed for gels without BMP-2 four weeks after injection. There were no signs of inflammation or foreign body response in either group and we believe that this system has the potential as an off-the-shelf injectable to be used where bone tissue is needed.

Bone morphogenetic protein-2 delivered by hyaluronan-based hydrogel induces massive bone formation and healing of cranial defects in minipigs.

BACKGROUND Reconstruction of large craniofacial bone defects is a challenge using bone transplants or alloplastic materials. The use of bone morphogenetic protein (BMP)-2 together with a suitable carrier is an attractive option that may facilitate new bone formation. The authors have developed a hydrogel that is formed in situ by the cross-linking of multifunctional hyaluronic acid and polyvinyl alcohol derivatives mixed with hydroxyapatite nanoparticles, in the presence of BMP-2. The aim of this study was to evaluate the suitability of the hydrogel as a carrier for BMP-2 in repairing critical size cranial defects in a minipig model. METHODS Cranial defects (2 x 4 cm) were created in 14 minipigs. The experimental groups were as follows: group 1, craniotomy and application of 5 ml of hydrogel with 1.25 mg of BMP-2 (n = 6); group 2, craniotomy and application of 5 ml of hydrogel without BMP-2 (n = 6); and group 3, craniotomy with no further treatment (n = 2). RESULTS After 3 months, computed tomographic and histologic examinations were performed. There was spontaneous ossification in the untreated group, but the healing was incomplete. The hydrogel alone demonstrated no further effects. The addition of 1.25 mg of BMP-2 to the hydrogel induced a greater than 100 percent increase in bone volume (p = 0.003) and complete healing of the defects. Histologic examination revealed compact lamellar bone in the BMP group without intertrabecular fibrous tissue, as was seen in the other groups. The hydrogel was resorbed completely within 3 months and, importantly, caused no inflammatory reaction. CONCLUSION The injectable hydrogel may be favorable as a BMP-2 carrier for bone reconstruction.

Three-dimensional technology and bone morphogenetic protein in frontal bone reconstruction.

Osteoinductive bone morphogenetic proteins (BMPs) may be used in humans to facilitate healing of bony defects. The effect of different BMPs is, as with many other growth factors, highly dependent on the delivery vehicle. Bovine type I collagen is currently used in the clinical setting as a carrier and has been approved in several countries for human use. Here, we report the reconstruction of a frontal bone defect using heparin together with bovine type I collagen, hyaluronic acid, and fibrin as vehicles for BMP-2. A bony structure was created on the back of the patient by treating the latissimus dorsi muscle with the growth factor. A polyamide mold was used as a template to achieve the desired shape. The bone structure was transplanted into the defect site via microsurgical techniques. Although the prefabricated bone was not large enough tocover the entire frontal defect, the reconstruction was completed by using an additional cranial implant.

Minimally invasive mandibular bone augmentation using injectable hydrogels

Hyaluronic acid‐based hydrogels are proven biocompatible materials and excellent carriers of bone morphogenetic protein‐2 (BMP‐2) that have been successfully tested for bone generation in vivo. Different formulations, with or without nanohydroxyapatite, have shown promise for craniofacial applications. In this study, 28 rats were used to investigate whether it is possible to achieve mandibular bone augmentation upon injection of novel hyaluronic acid‐based hydrogels containing nanohydroxyapatite and different concentrations of BMP‐2 (0, 5 and 150 µg/ml). The biomaterials were injected subperiosteally through fine needles into the innate mandibular diastema, imitating a clinical procedure for resorbed mandibles. No incisions, flaps or sutures were necessary. After 8 weeks the mandibles were evaluated by peripheral quantitative computed tomography (pQCT), micro‐computed tomography (μCT), histology, immunohistochemistry and fluorochrome labelling. As a result, engineered bone was observed in all treated mandibles, with a statistically significant increase in mandibular bone volume correlated with the amount of BMP‐2 loaded in the hydrogel formula. We therefore demonstrated that minimally invasive mandibular bone augmentation is possible upon injection in rats, when using the appropriate injectable scaffolds. This represents an attractive clinical alternative for oral implantology patients. Copyright

A Novel Biodegradable Delivery System for Bone Morphogenetic Protein-2

Background: The efficacy of recombinant growth factors in vivo is highly dependent on the delivery vehicle. The authors investigated the osteoinductive effects of recombinant human bone morphogenetic proteins (BMP)-2 implanted together with a complex of heparin and chitosan. Methods: Sixty rats were used. Three different carriers in gel formulation (type I collagen, heparin/type I collagen, and heparin/chitosan) were mixed with either 0, 10, or 50 &mgr;g of BMP-2, making the number of groups nine. The gels were injected into the quadriceps muscles of both legs in 45 rats (n = 10 per group). Freeze-dried formulations of the carriers were also tested with the same amounts of BMP-2 using 15 rats (n = 5 per group). Four weeks after implantation, the quality and amount of newly formed bone were assessed. Results: Chitosan was shown to protect the heparinase-mediated degradation of heparin in vitro. The osteoinductive effects of BMP-2 in combination with heparin/chitosan were superior as compared with BMP-2 implanted together with type I collagen. Interestingly, the heparin/chitosan complex induced a small amount of bone also without BMP-2 added. The heparin/chitosan was completely absorbed after 4 weeks as determined by histologic evaluation, and a normal active bone formation was present. The freeze-dried formulations of the carriers demonstrated similar osteoinductive effects as the gels. Conclusions: An osteoinductive formula for clinical use is needed for general bone reconstruction. Heparin in complex with chitosan has the ability to stabilize or activate the growth factor in vivo and induce the generation of new bone in good yields.

In situ cross-linkable hyaluronan hydrogel enhances chondrogenesis

The present work describes the feasibility of a cross‐linkable injectable hyaluronan hydrogel for cartilage repair. The hydrogel used is a two‐component system based on aldehyde‐modified hyaluronan and hydrazide‐modified polyvinyl alcohol, which are rapidly cross‐linked in situ upon mixing. The in vitro study showed that chondrocytes and mesenchymal cells cultured in the gel form cartilage‐like tissue, rich in glycosaminoglycans, collagen type II and aggrecan. In a rabbit animal model the injection of the hydrogel improved the healing of a full‐thickness cartilage defect created in the knee as compared to non‐treated controls. This rabbit study showed that the regenerated cartilage defects stained more intensely for type II collagen upon treatment with the hydrogel. The hyaluronan‐based hydrogel may be used as a delivery vehicle for both growth factors and/or cells for cartilage repair. The in vivo study also indicated that the hydrogel alone has a beneficial effect on cartilage regeneration. Copyright

Development of a bioactive implant for repair and potential healing of cranial defects

The repair of complex craniofacial bone defects is challenging and a successful result is dependent on the size of the defect, quality of the soft tissue covering the defect, and choice of reconstruction method. The objective of this study was to develop a bioactive cranial implant that could provide a permanent reconstructive solution to the patient by stimulating bone healing of the defect. In this paper the authors report on the feasibility and clinical results of using such a newly developed device for the repair of a large traumatic and therapy-resistant cranial bone defect. The patient had undergone numerous attempts at repair, in which established methods had been tried without success. A mosaic-designed device was manufactured and implanted, comprising interconnected ceramic tiles with a defined calcium phosphate composition. The clinical outcome 30 months after surgery revealed a restored cranial vault without postoperative complications. Computed tomography demonstrated signs of bone ingrowth. Examination with combined (18)F-fluoride PET and CT provided further evidence of bone healing of the cranial defect.

Biomaterials and biologics in craniofacial reconstruction.

Abstract Complications related to surgery, including infection, wound dehiscence, and implant protrusion, are costly and may cause severe morbidity to patients. The choice of implants materials is critical for a successful outcome, particularly in craniofacial reconstructions. This review discusses the potential benefits and drawbacks of biologically active materials used for craniofacial bone repair as alternatives to inert implant prostheses.

Cartilage repair of experimentally 11 induced osteochondral defects in New Zealand White rabbits

Articular cartilage has a limited capacity for self-repair in adult humans, and methods used to stimulate regeneration often result in re-growth of fibrous cartilage, which has lower durability. No current treatment option can provide complete repair. The possibility of growth factor delivery into the joint for cartilage regeneration after injury would be an attractive treatment option. A full thickness osteochondral defect of 4 mm in diameter and 2 mm deep was created by mechanical drilling in the medial femoral condyle in 20 female adult New Zealand White rabbits. In an attempt to improve regeneration a hyaluronic hydrogel system, with or without bone morphogenetic protein-2 (BMP-2) was delivered intraarticularly. The contralateral joint defect was treated with saline as control. Throughout the study, rabbits were clinically examined and after 12 (n = 6) or 24 (n = 9) weeks, the rabbits were euthanized and the joints evaluated by histology. The defects healed with fibrocartilage like tissue, and the filling of the defects ranged from less than 25% to complete. The healing of the defects varied both inter- and intra-group wise. Treatment with hyaluronan gel with or without BMP-2 had no effect on cartilage regeneration compared with controls. Instead, severe ectopic bone formation was found in seven joints treated with BMP-2. In conclusion, the present study shows that neither treatment with hyaluronic gel alone, nor in combination with BMP-2, improves the healing of an induced cartilage defect in rabbits. It further shows that BMP-2 can induce ectopic bone formation, which severely affects the functionality of the joint.

Variation in calvarial bone healing capacity: a clinical study on the effects of BMP-2-hydrogel or bone autograft treatments at different cranial locations.

BackgroundBone morphogenetic protein-2 (BMP-2) together with a suitable carrier is an attractive option that may be used for craniofacial bone reconstruction. In this prospective randomized study, a hyaluronan-based hydrogel with BMP-2 was used to achieve bone healing in standardized critical-size cranial defects in humans after neurosurgery. MethodsTwelve patients were randomized into the treatment group (N = 6) or control group (N = 6). In the treatment group, holes made during craniotomy were treated with hydrogel with BMP-2, 250 µg/mL, or hydrogel without BMP-2. In the remaining hole/s in the same patient, Spongostan (Ethicon) alone or Tisseel (Baxter) mixed with autologous bone matrix were used as negative and positive controls, respectively. In the control group, the holes were treated with Spongostan or Tisseel mixed with bone autograft. Bone healing was assessed with CT scans after 3 and 6 months. Bone areas in treated defects were measured and statistical analysis was performed. ResultsIndependent of location, bone healing in defects treated with Tisseel with autograft, hydrogel alone, or hydrogel with BMP-2 was significantly increased compared to negative control (P < 0.001, P = 0.002, and P = 0.005, respectively). In general, all defects healed significantly better in the frontal bone as compared to parietal-temporal location, except for defects treated with Tisseel and autograft, which healed well independently of location. No local or systemic side effects, including excessive bone overgrowth or inflammatory reaction, were seen in treated patients. ConclusionsTissue engineering of bone with hyaluronan-based hydrogel shows good healing of cranial defects, comparable with bone autografts. The hydrogel itself may represent a novel alternative to autologous bone transplants in craniofacial bone repair. The study also reveals a general superior healing capacity in the frontal bone as compared to parietal/temporal bones.