TGFβ Inhibition Stimulates Collagen Maturation to Enhance Bone Repair and Fracture Resistance in a Murine Myeloma Model

Multiple myeloma is a plasma cell malignancy that causes debilitating bone disease and fractures, in which TGFβ plays a central role. Current treatments do not repair existing damage and fractures remain a common occurrence. We developed a novel low tumor phase murine model mimicking the plateau phase in patients as we hypothesized this would be an ideal time to treat with a bone anabolic. Using in vivo μCT we show substantial and rapid bone lesion repair (and prevention) driven by SD-208 (TGFβ receptor I kinase inhibitor) and chemotherapy (bortezomib and lenalidomide) in mice with human U266-GFP-luc myeloma. We discovered that lesion repair occurred via an intramembranous fracture repair-like mechanism and that SD-208 enhanced collagen matrix maturation to significantly improve fracture resistance. Lesion healing was associated with VEGFA expression in woven bone, reduced osteocyte-derived PTHrP, increased osteoblasts, decreased osteoclasts, and lower serum tartrate-resistant acid phosphatase 5b (TRACP-5b). SD-208 also completely prevented bone lesion development in mice with aggressive JJN3 tumors, and was more effective than an anti-TGFβ neutralizing antibody (1D11). We also discovered that SD-208 promoted osteoblastic differentiation (and overcame the TGFβ-induced block in osteoblastogenesis) in myeloma patient bone marrow stromal cells in vitro, comparable to normal donors. The improved bone quality and fracture-resistance with SD-208 provides incentive for clinical translation to improve myeloma patient quality of life by reducing fracture risk and fatality. © 2019 American Society for Bone and Mineral Research.     

Novel Microhydroxyapatite Particles in a Collagen Scaffold: A Bioactive Bone Void Filler?

Background

Treatment of segmental bone loss remains a major challenge in orthopaedic surgery. Traditional techniques (eg, autograft) and newer techniques (eg, recombinant human bone morphogenetic protein-2 [rhBMP-2]) have well-established performance limitations and safety concerns respectively. Consequently there is an unmet need for osteoinductive bone graft substitutes that may eliminate or reduce the use of rhBMP-2.

Questions/purposes

Using an established rabbit radius osteotomy defect model with positive (autogenous bone graft) and negative (empty sham) control groups, we asked: (1) whether a collagen-glycosaminoglycan scaffold alone can heal the defect, (2) whether the addition of hydroxyapatite particles to the collagen scaffold promote faster healing, and (3) whether the collagen-glycosaminoglycan and collagen-hydroxyapatite scaffolds are able to promote faster healing (by carrying a low dose rhBMP-2).

Methods

A 15-mm transosseous radius defect in 4-month-old skeletally mature New Zealand White rabbits were treated with either collagen-hydroxyapatite or collagen-glycosaminoglycan scaffolds with and without rhBMP-2. Autogenous bone graft served as a positive control. Time-series radiographs at four intervals and postmortem micro-CT and histological analysis at 16 weeks were performed. Qualitative histological analysis of postmortem explants, and qualitative and volumetric 3-D analysis of standard radiographs and micro-CT scans enabled direct comparison of healing between test groups.

Results

Six weeks after implantation the collagen-glycosaminoglycan group had callus occupying greater than ½ the defect, whereas the sham (empty) control defect was still empty and the autogenous bone graft defect was completely filled with unremodeled bone. At 6 weeks, the collagen-hydroxyapatite scaffold groups showed greater defect filling with dense callus compared with the collagen-glycosaminoglycan controls. At 16 weeks, the autogenous bone graft groups showed evidence of early-stage medullary canal formation beginning at the proximal and distal defect borders. The collagen-glycosaminoglycan and collagen-glycosaminoglycan-rhBMP-2 groups had nearly complete medullary canal formation and anatomic healing at 16 weeks. However, collagen-hydroxyapatite-rhBMP-2 scaffolds showed the best levels of healing, exhibiting a dense callus which completely filled the defect.

Conclusions

The collagen-hydroxyapatite scaffold showed comparable healing to the current gold standard of autogenous bone graft. It also performed comparably to collagen-glycosaminoglycan-rhBMP-2, a representative commercial device in current clinical use, but without the cost and safety concerns.

Clinical Relevance

The collagen-glycosaminoglycan scaffold may be suitable for a low load-bearing defect. The collagen-hydroxyapatite scaffold may be suitable for a load-bearing defect. The rhBMP-2 containing collagen-glycosaminoglycan and collagen-hydroxyapatite scaffolds may be suitable for established nonunion defects.     

Does Demineralized Bone Matrix Enhance Tendon-to-Bone Healing after Rotator Cuff Repair in a Rabbit Model?

BackgroudThe purpose of this study was to compare the histologic outcomes of rotator cuff (RC) repair with demineralized bone matrix (DBM) augmentation and those without DBM augmentation and to evaluate the role of DBM for tendon-to-bone (TB) healing in a rabbit model.MethodsTwenty-six adult male New Zealand white rabbits were randomly allocated to the control group (n = 13) or the DBM group (n = 13). Repair was performed 8 weeks after complete transection of the right supraspinatus tendon of all rabbits. In the control group, RC repair was achieved by a standard transosseous technique. In the DBM group, RC repair was achieved using the same technique, and DBM was interposed between the cuff and bone. After 8 weeks, the RC tendon entheses from all rabbits were processed for gross and histologic examination.ResultsOn gross TB healing, 2 of 11 specimens in the control group were unhealed and no specimen was grossly unhealed in the DBM group (p = 0.421). In the control group, the tendon midsubstance was disorganized with randomly and loosely arranged collagen fibers and rounded fibroblastic nuclei. The TB interface was predominantly fibrous with small regions of fibrocartilage, especially mineralized fibrocartilage. In the DBM group, the tendon midsubstance appeared normal and comprised densely arranged collagen fibers, with orientated crimped collagen fibers running in the longitudinal direction of the tendon. These fibers were interspersed with elongated fibroblast nuclei. The TB interface consisted of organized collagen fibers with large quantities of fibrocartilage and mineralized fibrocartilage.ConclusionsThe use of DBM for TB interface healing in rabbit experiments showed good results in gross and histologic analysis. However, it is difficult to draw a solid conclusion because the sample size is small. Further evaluation in the in vivo setting is necessary to determine clinical recommendations.     

Is Nitric Oxide a Mediator of the Effects of Low-Intensity Electrical Stimulation on Bone in Ovariectomized Rats?

Low-intensity electrical stimulation (LIES) may counteract the effects of ovariectomy (OVX) on nitric oxide synthase (NOS) expression, osteocyte viability, bone structure, and microarchitecture in rats (Lirani-Galvão et al., Calcif Tissue Int 84:502–509, 2009). The aim of the present study was to investigate if these effects of LIES could be mediated by NO. We analyzed the effects of NO blockage (by l-NAME) in the response to LIES on osteocyte viability, bone structure, and microarchitecture in OVX rats. Sixty rats (200–220 g) were divided into six groups: sham, sham-l-NAME (6 mg/kg/day), OVX, OVX-l-NAME, OVX-LIES, and OVX-LIES-l-NAME. After 12 weeks, rats were killed and tibiae collected for histomorphometric analysis and immunohistochemical detection of endothelial NOS (eNOS), inducible NOS (iNOS), and osteocyte apoptosis (caspase-3 and TUNEL). In the presence of l-NAME, LIES did not counteract the OVX-induced effects on bone volume and trabecular number (as on OVX-LIES). l-NAME blocked the stimulatory effects of LIES on iNOS and eNOS expression of OVX rats. Both l-NAME and LIES decreased osteocyte apoptosis. Our results showed that in OVX rats l-NAME partially blocks the effects of LIES on bone structure, turnover, and expression of iNOS and eNOS, suggesting that NO may be a mediator of some positive effects of LIES on bone.     

Is Collagen a Good Banding Material for Outlet Control of Vertical Gastroplasty? Preliminary Study in Pigs

Background: To maintain the long-term effects of a gastric bariatric operation, bands are often placed to control the restriction. Erosion into the gastric wall by these devices remains a problem. A soft resiliant prosthesis of animal origin, constituted by a network of non-absorbable collagen fibres, may be a solution to this problem. This study assessed, in a porcine model, the histological reaction of the gastric wall following apposition of a band of porcine collagen (Pelvicol^?, Bard). Methods: 15 female pigs weighing on average 21 kg underwent vertical banded gastroplasty (VBG). Stoma control was achieved with a band of porcine collagen (2 cm wide, 7 cm long and 2 mm thick). The pigs were sacrificed 1 month after VBG, and histological analysis was performed at a macroscopic and microscopic level. Results: There was no peri-operative death, although 2 pigs died in the postoperative period (the first case developed a bowel fistula and sepsis, and the second pig died of unrelated causes). There were 2 additional morbidities (gastric fistula on the linear staple-line away from the Pelvicol^? band) that led to an early euthanasia of 2 pigs. Post-mortem macroscopic analyses in the remaining 11 pigs did not reveal migration of the device, and there was no tissue reaction on postoperative microscopic analyses. 10 of the pigs had lost weight at 1 month, averaging 3.42 kg. Conclusion: Porcine collagen appears to be an effective and safe alternative to the current methods of control of pouch outlet. The flexibility and homogeneity of this prosthesis may be useful to limit the risk of erosion of the gastric wall. Although these properties have been assessed in pelvic operations in humans, this work needs to be studied in a prospective long-term study in humans.     

Does Exercise Modify the Effects of Zoledronic Acid on Bone Mass,Microarchitecture, Biomechanics,and Turnover in Ovariectomized Rats?

Regular activity has effects on bone size, shape, and density, resulting in an increase in mechanical strength. The mechanism of action that underlies this improvement in bone strength is mainly linked to an increase in bone formation. Zoledronic acid (Z), in contrast, may prevent bone strength changes in ovariectomized (OVX) rodents by its potent antiresorptive effects. Based on these assumptions we hypothesized that combined effects of exercise (E) and Z may produce higher benefits on bone changes resulting from estrogen deficiency than either intervention alone. At 6 months of age, 60 female Wistar rats were OVX or sham operated (SH) and divided into five groups: SH, OVX, OVX-E, OVX-Z, and OVX-ZE. OVX rats were treated with a single IV injection of Z (20 μg/kg) or vehicle and submitted or not to treadmill exercise (15 m/min, 60 min/day, 5 days/week) for 12 weeks. Whole-body BMD and bone turnover markers were analyzed longitudinally. At sacrifice, femurs were removed. BMD by DXA, three-point bending test, and μCT were performed to study biomechanical and trabecular structure parameters, respectively. After 12 weeks, bone volume fraction decreased in OVX rats, whereas bone turnover rate, trabecular spacing, and structure model index increased compared with those in the SH group (P < 0.05). Zoledronic acid prevented the ovariectomy-induced trabecular bone loss and its subsequent trabecular microarchitectural deterioration. Treadmill exercise running was shown to preserve the bone strength and to induce bone turnover changes in favor of bone formation. However, the combined effects of zoledronic acid and running exercise applied simultaneously did not produce any synergetic or additive effects.     

Complete Repair of Bladder Exstrophy in Boys: Can Hypospadias Be Avoided?

INTRODUCTION: Complete repair of classic bladder exstrophy in male newborns has been successful with minimal morbidity. However, the technique may create hypospadias in some cases. We have recently adopted a modification to obtain an orthotopic meatus in bladder exstrophy boys. MATERIALS AND METHODS: Between November 1998 and December 2002 complete repair of classic bladder exstrophy was carried out in 27 boys. Complete penile disassembly was performed in 22 boys including 4 newborns and 18 older children; mean age 3+/-2 years old. Modified disassembly was used in the last 5 boys including 4 newborns and a 9-month-old boy. During repair of epispadias, the dissection starts on the ventral aspect of the penis as usual. The urethral plate is separated from both corpora cavernosa to allow ventral transposition of the plate. In the complete disassembly technique, the urethral plate is completely separated from both hemiglans. In this modification, while the urethral plate is completely separated from both corpora cavernosa, the extreme distal end of the urethral plate remains attached to the distal ends of both hemiglans. Thus, when the plate is tubularized with fine interrupted sutures the meatus ends up at the tip of the glans penis. The symphysis pubis is re-approximated as usual and corpora cavernosa are approximated dorsally in the midline. RESULTS: There was no major complication. Mean follow-up was 43+/-7 and 7+/-4 months for complete and modified disassembly groups respectively. Of the 22 boys, who underwent complete disassembly, 15 (68%) ended up with hypospadias and 7 (32%) had an orthotopic meatus. Modified disassembly has resulted in an excellent cosmetic appearance and orthotopic meatus in the 5 boys. Penile length was not shorter than boys who underwent complete disassembly. Parents of the 5 boys noticed normal straight morning erection. CONCLUSION: Although complete penile disassembly allows ventral placement of the urethra, hypospadias is created in approximately two thirds of the cases. When the extreme distal end of the urethral plate remains attached to the distal ends of both hemiglans (modified disassembly), orthotopic meatus can be obtained in all boys. With modified disassembly, posterior mobilization of the bladder and urethra does not result in corporal angulations or shortening because of the proportionate inward movement of the corpora that accompanies symphyseal approximation. The modification is feasible in newborns and infants. These short-term results may obviate the need for later penile reconstructive procedure.     

Can We Confidently Study VO2 Kinetics in Young People?

The study of VO₂ kinetics offers the potential to non-invasively examine the cardiorespiratory and metabolic response to dynamic exercise and limitations to every day physical activity. Its non-invasive nature makes it hugely attractive for use with young people, both healthy and those with disease, and yet the literature, whilst growing with respect to adults, remains confined to a cluster of studies with these special populations. It is most likely that this is partly due to the methodological difficulties involved in studying VO₂ kinetics in young people which are not present, or present to a lesser degree, with adults. This article reviews these methodological issues, and explains the main procedures that might be used to overcome them.

Key points

• The VO₂ kinetic response to exercise represents the combined efficiency of the cardiovascular, pulmonary and metabolic systems, and an accurate assessment of the response potentially provides a great deal of useful information via non-invasive methodology.
• An accurate assessment of the VO₂ kinetic response is however inherently difficult with children and especially those with reduced exercise tolerance, due primarily to the apparent breath-by-breath noise which masks the true underlying physiological response, and the small amplitudes of the response signal.
• Despite this, it is possible to assess and quantify the VO₂ kinetic response with children if appropriate steps are taken to apply carefully selected methodologies and report response variables with confidence intervals. In this way, both the researcher and the reader can be confident that the data reported is meaningful.

Key words: Children, oxygen kinetics, methodology     

Dose-Dependent Resorption of Allograft by rhBMP-2 Uncompensated by New Bone Formation—A Canine Study With Implants and Zoledronate

Background

Impacted bone allograft is used to restore lost bone in total joint arthroplasties. Bone morphogenetic proteins (BMPs) can induce new bone formation to improve allograft incorporation, but they simultaneously invoke a seemingly dose-dependent allograft resorption mediated by osteoclasts. Bisphosphonates effectively inhibit osteoclast activity. Predicting allograft resorption when augmented with bone morphogenetic protein 2 (BMP-2), we intended to investigate whether a balanced bone metabolism was achievable within a range of BMP-2 doses with systemic zoledronate treatment.

Second-generation Highly Cross-linked X3? Polyethylene Wear: A Preliminary Radiostereometric Analysis Study

Background  

First-generation highly cross-linked polyethylene liners have reduced the incidence of wear particle-induced osteolysis. However, failed acetabular liners have shown evidence of surface cracking, mechanical failure, and oxidative damage. This has led to the development of second-generation highly cross-linked polyethylene, which has improved wear and mechanical properties and resistance to oxidation in vitro. Owing to its recent introduction, there are no publications describing its clinical performance.     

Evaluation of the Potential Use of Trabecular Bone Score to Complement Bone Mineral Density in the Diagnosis of Osteoporosis: A Preliminary Spine BMD–Matched,Case-Control Study

The trabecular bone score (TBS) is a new parameter that is determined from gray-level analysis of dual-energy X-ray absorptiometry (DXA) images. It relies on the mean thickness and volume fraction of trabecular bone microarchitecture. This was a preliminary case-control study to evaluate the potential diagnostic value of TBS as a complement to bone mineral density (BMD), by comparing postmenopausal women with and without fractures. The sample consisted of 45 women with osteoporotic fractures (5 hip fractures, 20 vertebral fractures, and 20 other types of fracture) and 155 women without a fracture. Stratification was performed, taking into account each type of fracture (except hip), and women with and without fractures were matched for age and spine BMD. BMD and TBS were measured at the total spine. TBS measured at the total spine revealed a significant difference between the fracture and age– and spine BMD–matched nonfracture group, when considering all types of fractures and vertebral fractures. In these cases, the diagnostic value of the combination of BMD and TBS likely will be higher compared with that of BMD alone. TBS, as evaluated from standard DXA scans directly, potentially complements BMD in the detection of osteoporotic fractures. Prospective studies are necessary to fully evaluate the potential role of TBS as a complementary risk factor for fracture.     

Impaired Bone Health in Inflammatory Bowel Disease: A Case–Control Study in 80 Pediatric Patients

Previous studies have indicated that children with inflammatory bowel disease (IBD) may not achieve optimal bone mass. We evaluated the skeletal characteristics in children and adolescents with IBD. This cross-sectional cohort study comprised 80 IBD patients (median age 14.9?years, range 5-20) with a median disease duration of 3.4?years; 51 had ulcerative colitis, 26 Crohn disease, and 3 unspecified colitis. Eighty age- and gender-matched healthy subjects served as controls. Areal bone mineral density (aBMD), body composition, and vertebral fractures (VFs) were assessed by DXA. Bone age (BA) was determined for IBD patients. Findings were correlated with disease- and treatment-related parameters and biochemistry. IBD patients had lower BA-adjusted lumbar spine and whole-body aBMD (p?150?mg/kg for the preceding 3?years increased the risk for low whole-body aBMD (OR?=?5.5, 95?% CI 1.3-23.3, p?=?0.02). VFs were found in 11?% of patients and in 3?% of controls (p?=?0.02). IBD in childhood was associated with low aBMD and reduced bone mass accrual relative to muscle mass; the risk for subclinical VFs may be increased. These observations warrant careful follow-up and active preventive measures.     

Cancellous Bone Properties and Matrix Content of TGF-β2 and IGF-I in Human Tibia: A Pilot Study

Transforming and insulin-like growth factors are important in regulating bone mass. Thus, one would anticipate correlations between matrix concentrations of growth factors and functional properties of bone. We therefore investigated the relationships of (1) TGF-β2 and (2) IGF-I matrix concentrations with the trabecular microstructure, stress distribution, and mechanical properties of tibial cancellous bone from six male human cadavers. Trabecular stress amplification (VMExp/σ_app) and variability (VMCOV) were calculated using microcomputed tomography (μCT)-based finite element simulations. Bone volume fraction (BV/TV), surface/volume ratio (BS/BV), trabecular thickness (Tb.Th), number (Tb.N) and separation (Tb.Sp), connectivity (Eu.N), and anisotropy (DA) were measured using 3-D morphometry. Bone stiffness and strength were measured by mechanical testing. Matrix concentrations of TGF-β2 and IGF-I were measured by ELISA. We found higher matrix concentrations of TGF-β2 were associated with higher Tb.Sp and VMExp/σ_app for pooled data and within subjects. Similarly, a higher matrix concentration of IGF-I was associated with lower stiffness, strength, BV/TV and Tb.Th and with higher BS/BV, Tb.Sp, VMExp/σ_app and VMCOV for pooled data and within subjects. IGF-I and Tb.N were negatively associated within subjects. It appears variations of the stress distribution in cancellous bone correlate with the variation of the concentrations of TGF-β2 and IGF-I in bone matrix: increased local matrix concentrations of growth factors are associated with poor biomechanical and architectural properties of tibial cancellous bone.     

Intermediate Follow-Up after Endovascular Aneurysm Repair: Can We Forgo CT Scanning in Certain Patients?

Current recommendations for follow-up after endovascular repair of abdominal aortic aneurysms (EVAR) include yearly computed tomographic (CT) scans after the first year. We hypothesize that this is unnecessary for patients who have aneurysm sacs that are stable or shrinking at 1 year and no evidence of endoleak. To explore this hypothesis, we reviewed the records of all patients undergoing EVAR at our institution who were implanted with grafts that are currently commercially available and had a minimum of 18 months' follow-up. Of 415 patients who underwent EVAR over an 8-year period, 93 met the entry criteria. At a mean follow-up of approximately 3 years, secondary interventions were required in 13%, 39%, and 25% of patients undergoing EVAR with Zenith, AneuRx, and Excluder devices, respectively, and secondary interventions after the first year were required in 3%, 22%, and 8% of such grafts, respectively. Seventy-one patients (76%) had aneurysm sacs that were stable or shrinking at 1 year and no endoleak. Only two of these patients subsequently required reintervention. Both patients had AneuRx grafts, and both problems could have easily been identified without CT scanning. Our data support the hypothesis that patients who meet these criteria at 1 year are unlikely to have problems that cannot be identified by ultrasound and/or clinical evaluation alone and, thus, that CT scans are not necessary after this point, especially in patients with Zenith or reengineered Excluder devices.