Anti-Inflammatory Activity of a Demineralized Bone Matrix: An In Vitro Pilot Study

Demineralized bone matrix (DBM) is commonly used for the reconstruction of bone defects. Early graft consolidation involves a transient inflammatory process. It is, however, unclear whether DBM can modulate this process. To test this possibility, we prepared acid lysates of demineralized ground cortical (DGC) and moldable demineralized fibers (MDF). Murine RAW 264.7 and primary bone marrow macrophages were exposed to acid lysates of DGC and MFD prior to provoking an inflammatory response with lipopolysaccharide (LPS). Similarly, murine ST2 mesenchymal cells were exposed to DGC and MFD with and without interleukin 1β (IL1) and TNFα. We show here that acid lysates of DGC and MFD reduced the expression of IL1 and IL6 in RAW 264.7 macrophages, as determined by RT-PCR and, for IL6, by immunoassay. This response was confirmed with primary macrophages. Likewise, desalted acid lysates exert anti-inflammatory properties on RAW 264.7 cells and in ST2 cells, the forced expression of IL6, inducible nitric oxide synthase (iNOS) and chemokine ligand 5 (CCL5) was reduced. These in vitro findings suggest that DGC and MFD lower the inflammation-induced expression of inflammatory mediators in murine cell-based bioassays.

Novel Fracture Site Targeting Drug Improves Fracture Healing with Pain Relief: Preclinical Evaluation of MAK123

Non-union bone fracture occurs in 5-10% of fracture injuries. Interventions include surgery with local implantation of autograft, allograft, demineralized bone matrix, and/or bone morphogenetic proteins. These types of fracture injuries are also accompanied by acute and chronic pain states. In most instances, opioids are provided to injured patients during and after surgery. With the opioid crisis, identifying new analgesic therapies that could reduce or eliminate opioid use, while also improving bone healing is important. Here we show the ability of a novel compound, MAK123, to both enhance bone healing and reduce pain behavior in a surgically induced femoral fracture mouse model. Briefly, 20 male C57BL/6 mice underwent a surgically induced femoral fracture and then were treated with 0, 2, 6, or 20 mg/kg, 3X/week for the 3 week study duration. Weekly X-rays were used to examine healing progression. Prior to euthanasia, mice underwent behavioral testing to measure evoked pain behaviors. Upon euthanasia, ex vivo µCT imaging and analysis was completed to assess fracture callus size and composition. While all doses of MAK123 tested resulted in improved healing, the 6mg/kg dose resulted in accelerated bone healing and a significant increase in mineralized callus volume (p<0.05). Similarly, while all doses of MAK123 reduced evoked responses to tactile stimulus as demonstrated by increased paw withdrawal thresholds, 6 mg/kg of MAK123 resulted in a more robust and significant improvement (p<0.05). We postulate that optimization of the dosing schedule/concentration could further improve both bone healing and behavioral measures thought to represent pain in rodents. That said, these promising pre-clinical data warrant further evaluation as MAK123 may prove to be a unique tool for orthopaedic surgery usage whereby it could both improve bone healing and reduce clinical pain, improving overall patient outcomes.

Clinical and Radio Graphical Evaluation of Implants Placed in Ridge Preserved Sites versus Naturally Healed Sites

Introduction: Dental implant has proved to be a pioneer in prosthetic rehabilitation overcoming the disadvantage of a removable or a fixed partial denture. The aim of the study was to clinically and radiographically evaluate and compare changes of hard and soft tissues around the implants which will be placed in ridge preserved sites with implant placed in naturally healed extraction sites. Materials and Methods: In this study 10 Patients having tooth with hopeless prognosis requiring tooth extraction followed by implant placement were enrolled. These patients were randomly divided in to two groups. Group I: Socket preservation with demineralized bone matrix putty group. Group II: Naturally healed socket after extraction, followed by implant placement in both groups. Results: The results of the current study indicated that demineralisedbone matrix putty, when used in patients for dental augmentation in either mandibular or maxillary sites, resulted in replacement of the graft material with bone by as early as 4-6 months, there upon enabling implant placement and subsequent prosthetic reconstruction. Conclusion: Bucco lingual ridge width and width of keratinized gingiva can be preserved by ridge preservation after extraction. Short-term survival rates and clinical outcomes of both groups were similar and were comparable. Further clinical trials with longer duration follow up with larger sample size should be done to get more affirmative and conclusive results.

Preliminary outcomes of the combination of demineralized bone matrix and platelet Rich plasma in the treatment of long bone non-unions

Background A variety of bone graft substitutes have been introduced into the treatment of bone non-unions. However, clinical outcomes from current evidences are various and conflicting. This study aimed to present the preliminary outcomes of a treatment protocol in which the combination of demineralized bone matrix (DBM) and platelet rich plasma (PRP) was used as a bone graft substitute for long bone non-unions. Methods Data of this retrospective study were reviewed and collected from a consecutive case series involving 43 patients who presented with a long bone non-union and were treated in our department from October 2018 to May 2019. The combination of DMB and PRP was applied as a bone defect filler in 16 patients, whilst the other 27 patients were treated with iliac bone autografting. Patients’ demographics, postoperative complications and the result of bone union were compared and evaluated. Results The demographic data between the two groups were comparable. No significant difference was found with regard to the incidence of postoperative complications. No graft rejection, heterotopic ossification or other complications were noted. The distribution of bony healing time was rather scattered but did not differ significantly between the groups (7.533 ± 3.357 months vs. 6.625 ± 2.516 months; P=0.341). Union was identified radiographically in 15 of 16 patients in the DBM+PRP group and in 24 of 27 patients in autograft group. Conclusions The present study identified that low incidence of postoperative complications and satisfactory bony healing rate could be achieved in the treatment of long bone non-unions augmented with the combination of DBM and PRP. Although these findings might indicate the promising future of this treatment protocol, larger and higher quality studies should also be executed to assess its routine use.

Biologic Augmentation of Tibiotalocalcaneal Arthrodesis With Allogeneic Bone Block Is Associated With High Rates of Fusion

Background: The orthopedic conditions and systemic comorbidities that occur in patients who require bone block tibiotalocalcaneal (TTC) arthrodesis have made this procedure associated with a higher-than-normal risk of nonunion, graft collapse, hardware failure, and amputation. Here, we present a novel approach to bone block TTC arthrodesis using adjunctive osteoinductive agents and a prolonged course of protected weightbearing to assess if we could improve on historical outcomes. We also evaluated the efficacy of a vascularized medial femoral condyle (MFC) free flap to augment TTC arthrodesis. Methods: Fourteen adult patients underwent bone block TTC arthrodesis biologically augmented with fresh-frozen femoral head allograft, bone marrow aspirate concentrate, and demineralized bone matrix cortical fibers. Three patients with soft tissue defects underwent vascularized reconstruction with an MFC free flap. Radiographic union, the Foot Function Index (FFI), and PROMIS pain interference (PI), and physical function (PF) scores were assessed at follow-up. Results: TTC fusion was documented on plain radiograph in 13 of 14 patients (92.9%) and CT in 10 of 11 patients (90.9%). Mean time to fusion was 183.2 ± 83.2 days. One patient (7.1%) experienced nonunion and persistent infection requiring amputation. Patients who underwent vascularized bone grafting had significantly shorter time to fusion (112.3 ± 31.7 days vs 204.4 ± 82.7 days, P = .05). Patient-reported outcomes revealed mild to moderate pain and dysfunction after 1 year (mean FFI = 41.0% ± 23.1%, PROMIS PI = 58.3 ± 1.8, PROMIS PF = 39.0 ± 2.2). Conclusion: In this relatively small series, the biologic augmentation of bone block TTC arthrodesis with osteoinductive agents and protective weightbearing resulted in excellent rates of fusion, modest pain, and preserved function of the lower extremity in almost all those treated. Osseous healing appears to be enhanced and accelerated with application of an MFC flap. We believe that this approach offers a viable salvage option for these challenging clinical problems. Level of Evidence: Level IV, case series.

The effect of biphasic calcium phosphate and demineralized bone matrix on tooth eruption in mongrel dogs

Background: Bone grafts can provide an optimal environment for permanent tooth to erupt and enhance the stability of the alveolar maxilla. Although autologous bone is an optimal source for osteogenesis, its inevitable donor site morbidity has led to active research on bone substitutes. This study was designed to evaluate the safety and feasibility of using biphasic calcium phosphate (BCP; Osteon) as a bone substitute in dogs. Methods: Bilateral third and fourth premolars of four 15-week-old mongrel dogs were used. All teeth were extracted except the third premolar of the right mandible, which was used as a control. After extraction of the premolars, each dog was administered BCP (Osteon), demineralized bone matrix (DBM; DBX), and no graft in the hollow sockets of the right fourth premolar, left fourth premolar, and left third premolar, respectively. Radiographs were taken at 2-week intervals to check for tooth eruption. After 8 weeks, each dog was sacrificed, and tooth and bone biopsies were performed to check for the presence of tooth and bone substitute particle remnants. Results: Four weeks after the operation, permanent tooth eruptions had started at all the extraction sites in each dog. Eight weeks after the operation, all teeth had normally erupted, and histological examination revealed BCP particles at the right fourth premolar. Conclusion: In all four dogs, no delay in the eruption of the teeth or shape disfigurement of permanent teeth was observed on gross inspection and radiologic evaluation. On histological examination, most of the BCP and DBM were replaced by new bone. Bone substitutes can be used as graft materials in patients with alveolar clefts.

Preparation and Characterization of Moldable Demineralized Bone Matrix/Calcium Sulfate Composite Bone Graft Materials

Demineralized bone matrix (DBM) is a decalcified allo/xenograft retaining collagen and noncollagenous proteins, which has been extensively used because of its osteoconductive and osteoinductive properties. Calcium sulfate (CaSO4, CS) is a synthetic bone substitute used in bone healing with biocompatible, nontoxic, bioabsorbable, osteoconductive, and good mechanical characteristics. This study aims to prepare a DBM/CS composite bone graft material in a moldable putty form without compromising the peculiar properties of DBM and CS. For this purpose, firstly, porcine femur was defatted using chloroform/methanol and extracted by acid for demineralization, then freeze-dried and milled/sieved to obtain DBM powder. Secondly, the α-form and β-form of calcium sulfate hemihydrate (CaSO4 •0.5H2O, CSH) were produced by heating gypsum (CaSO4 •2H2O). The morphology and particle sizes of α- and β-CSH were obtained by SEM, and their chemical properties were confirmed by EDS, FTIR and XRD. Furthermore, the DBM-based graft was mixed with α- or β-CSH at a ratio of 9:1, and glycerol/4% HPMC was added as a carrier to produce a putty. DBM/CSH putty possesses a low washout rate, good mechanical strength and biocompatibility. In conclusion, we believe that the moldable DBM/CSH composite putty developed in this study could be a promising substitute for the currently available bone grafts, and might have practical application in the orthopedics field as a potential bone void filler.