Hydrogel-based Delivery of rhBMP-2 Improves Healing of Large Bone Defects Compared With Autograft

Abstract

Background Autologous bone grafting remains the gold standard in the treatment of large bone defects but is limited by tissue availability and donor site morbidity. Recombinant human bone morphogenetic protein-2 (rhBMP-2), delivered with a collagen sponge, is clinically used to treat large bone defects and complications such as delayed healing or nonunion. For the same dose of rhBMP-2, we have shown that a hybrid nanofiber mesh-alginate (NMA-rhBMP-2) delivery system provides longer-term release and increases functional bone regeneration in critically sized rat femoral bone defects compared with a collagen sponge. However, no comparisons of healing efficiencies have been made thus far between this hybrid delivery system and the gold standard of using autograft. Questions/purposes We compared the efficacy of the NMA-rhBMP-2 hybrid delivery system to morselized autograft and hypothesized that the functional regeneration of large bone defects observed with sustained BMP delivery would be at least comparable to autograft treatment as measured by total bone volume and ex vivo mechanical properties. Methods Bilateral critically sized femoral bone defects in rats were treated with either live autograft or with the NMA-rhBMP-2 hybrid delivery system such that each animal received one treatment per leg. Healing was monitored by radiography and histology at 2, 4, 8, and 12 weeks. Defects were evaluated for bone formation by longitudinal micro-CT scans over 12 weeks (n = 14 per group). The bone volume, bone density, and the total new bone formed beyond 2 weeks within the defect were calculated from micro-CT reconstructions and values compared for the 2-, 4-, 8-, and 12-week scans within and across the two treatment groups. Two animals were used for bone labeling with subcutaneously injected dyes at 4, 8, and 12 weeks followed by histology at 12 weeks to

Delivery vehicle effects on bone regeneration and heterotopic ossification induced by high dose BMP-2

“…6B, bright red areas with Safranin-O stain). Most bone in the defect appeared to be mature bone, based on the dark red staining with Mallory's modified aniline blue stain (Fig 6C) as described previously [38]. In contrast, the alginate group showed areas of bone interrupted by large pieces of alginate (Fig.…”

“…Here we evaluated the influence of two delivery scaffolds loaded with the same BMP-2 dose of 120 μg/kg (30 μg per average 250 g rat), which falls in the typical range of doses used clinically: 50–800 μg/kg (based on the assumption of an 80 kg individual) [51, 56]. More importantly, previous studies in this rat segmental defect model [40, 57, 58] have established 2–2.5 μg BMP-2 as the healing dose (12–15 times lower than the dose used here), with superior retention and healing in alginate scaffolds compared to ACS [11] and autograft (5 μg BMP-2 dose) [38]. …”

“…Mid-sagittal 7 μm non-decalcified sections were obtained from the defect center by a tape transfer technique [38, 46] (Section Lab, Hiroshima, Japan). Mineralization was identified by von Kossa staining with Fast Red counterstain.…”

“…Samples were decalcified, paraffin embedded, and sectioned using a tape transfer technique (Section Labs, Hiroshima, Japan). Sagittal sections obtained from the mid-defect region were deparaffinized and stained with H&E to characterize tissue morphology, Safranin-O and Fast Green to identify alginate and cartilage, and Mallory's modified aniline blue stain for identification of mature and immature bone/mineralized tissue, as described extensively for this experimental model [11, 36-38, 40]. …”

“…Thus, the aim of this study was to provide a direct comparison of the effects of the delivery system on bone regeneration and heterotopic ossification, resulting from a supraphysiological dose of recombinant human BMP-2 (rhBMP-2) - delivered in the clinically common ACS or an alginate hydrogel based system. While our group has investigated the hybrid alginate hydrogel system in a pre-clinical animal model of critical sized segmental bone defect [11, 33-41] and demonstrated improved functional bone regeneration compared to autograft [38], and ACS [11, 36, 37] at lower rhBMP-2 doses, the performance of this hybrid system as a carrier for supraphysiological doses of rhBMP-2 has not been evaluated. Moreover, though exuberant ectopic bone formation has been shown as a part of earlier studies using high dose rhBMP-2, its quantitative characterization and temporal evolution have not been adequately described.…”