Can axial loading restore in vivo disc geometry, opening pressure, and T2 relaxation time?

Background: Cadaveric intervertebral discs are often studied for a variety of research questions, and outcomes are interpreted in the in vivo context. Unfortunately, the cadaveric disc does not inherently represent the LIVE condition, such that the disc structure (geometry), composition (T2 relaxation time), and mechanical function (opening pressure, OP) measured in the cadaver do not necessarily represent the in vivo disc. Methods: We conducted serial evaluations in the Yucatan minipig of disc geometry, T2 relaxation time, and OP to quantify the changes that occur with progressive dissection and used axial loading to restore the in vivo condition. Results: We found no difference in any parameter from LIVE to TORSO; thus, within 2 h of sacrifice, the TORSO disc can represent the LIVE condition. With serial dissection and sample preparation the disc height increased (SEGMENT height 18% higher than TORSO), OP decreased (POTTED was 67% lower than TORSO), and T2 time was unchanged. With axial loading, an imposed stress of 0.20-0.33 MPa returned the disc to in vivo, LIVE disc geometry and OP, although T2 time was decreased. There was a linear correlation between applied stress and OP, and this was conserved across multiple studies and species. Conclusion: To restore the LIVE disc state in human studies or other animal models, we recommend measuring the OP/stress relationship and using this relationship to select the applied stress necessary to recover the in vivo condition.

Injectable Radiopaque Hyaluronic Acid Granular Hydrogels for Intervertebral Disc Repair.

Injectable hydrogels offer minimally-invasive treatment options for degenerative disc disease, a prevalent condition affecting millions annually. Many hydrogels explored for intervertebral disc (IVD) repair suffer from weak mechanical integrity, migration issues, and expulsion. To overcome these limitations, an injectable and radiopaque hyaluronic acid granular hydrogel is developed. The granular structure provides easy injectability and low extrusion forces, while the radiopacity enables direct visualization during injection into the disc and non-invasive monitoring after injection. The radiopaque granular hydrogel is injected into rabbit disc explants to investigate restoration of healthy disc mechanics following needle puncture injury ex vivo and then delivered in a minimally-invasive manner into the intradiscal space in a clinically-relevant in vivo large animal goat model of IVD degeneration initiated through degradation by chondroitinase. The radiopaque granular hydrogel successfully halted loss of disc height due to degeneration. Further, the hydrogel not only enhanced proteoglycan content and reduced collagen content in the nucleus pulposus (NP) region compared to degenerative discs, but also helped to maintain the structural integrity of the disc and promote healthy segregation of the NP and annulus fibrosus regions. Overall, this study demonstrates the great potential of an injectable radiopaque granular hydrogel for treatment of degenerative disc disease.

Tension-activated nanofiber patches delivering an anti-inflammatory drug improve repair in a goat intervertebral disc herniation model.

Conventional microdiscectomy treatment for intervertebral disc herniation alleviates pain but does not repair the annulus fibrosus, resulting in a high incidence of recurrent herniation and persistent dysfunction. The lack of repair and the acute inflammation that arise after injury can further compromise the disc and result in disc-wide degeneration in the long term. To address this clinical need, we developed tension-activated repair patches (TARPs) for annulus fibrosus repair and local delivery of the anti-inflammatory factor anakinra (a recombinant interleukin-1 receptor antagonist). TARPs transmit physiologic strain to mechanically activated microcapsules embedded within the patch, which release encapsulated bioactive molecules in direct response to spinal loading. Mechanically activated microcapsules carrying anakinra were loaded into TARPs, and the effects of TARP-mediated annular repair and anakinra delivery were evaluated in a goat model of annular injury in the cervical spine. TARPs integrated with native tissue and provided structural reinforcement at the injury site that prevented aberrant disc-wide remodeling resulting from detensioning of the annular fibrosus. The delivery of anakinra by TARP implantation increased matrix deposition and retention at the injury site and improved maintenance of disc extracellular matrix. Anakinra delivery additionally attenuated the inflammatory response associated with TARP implantation, decreasing osteolysis in adjacent vertebrae and preserving disc cellularity and matrix organization throughout the annulus fibrosus. These results demonstrate the therapeutic potential of TARPs for the treatment of intervertebral disc herniation.

EXPERIMENTAL CHOLECALCIFEROL SUPPLEMENTATION IN A HERD OF MANAGED ASIAN ELEPHANTS (ELEPHAS MAXIMUS).

Vitamin D supplementation may pose a significant health risk in species where levels of deficiency, sufficiency, and toxicity have not been clearly established, and species-specific research on vitamin D supplementation should be performed. This study documented the effect of vitamin D supplementation on serum vitamin D metabolites and other analytes of Ca homeostasis in Asian elephants (Elephas maximus). Six adult Asian elephants received PO supplementation with cholecalciferol at 300 IU/kg of body weight (BW) once a week for 24 wk. Serum was analyzed every 4 wk for 25-hydroxyvitamin D2/D3 [25(OH)D]; 24,25-dihydroxyvitamin D2/D3 [24,25(OH)2D]; 1,25-dihydroxyvitamin D [1,25(OH)2D]; parathyroid hormone (PTH); total Ca; ionized Ca (iCa); P; and Mg. After the supplement was discontinued, serum 25(OH)D2/D3 was measured every 4 wk until levels returned to baseline. At the start of the study, the average serum 25(OH)D3 was nondetectable (<1.5 ng/ml). With cholecalciferol supplementation, 25(OH)D3 increased at an average rate of 2.26 ng/ml per month and reached an average concentration of 12.9 ± 3.46 ng/ml at 24 wk. Both 24,25(OH)2D3 and 1,25(OH)2D increased over time with supplementation from an average of <1.5 to 12.9 ng/ml and from 9.67 to 36.4 pg/ml, respectively. PTH, iCa, Ca, P, and Mg remained within reported normal ranges throughout supplementation. After the supplement was discontinued, serum 25(OH)D3 demonstrated a slow decline to baseline, taking an average of 48 wk. Elephants demonstrated significant individual variation in response to supplementation and subsequent return to baseline. Supplementation of Asian elephants with a weekly dose of 300 IU/kg BW cholecalciferol for 24 wk appears to be effective and safe. Additional clinical studies would be necessary to investigate the safety of other routes of administration, dosages, and duration of vitamin D supplementation, as well as associated health benefits.

Repair of femoral fractures in calves via external skeletal fixator with intramedullary pin tie-in.

OBJECTIVE: To describe the success rate and associated complications of external skeletal fixator/intramedullary pin tie-in for calf femoral fracture fixation. STUDY DESIGN: Clinical retrospective. ANIMALS: Ten calves, less than 30 days old, with diaphyseal/metaphyseal femoral fractures. METHODS: Medical records were reviewed from the University of Wisconsin from 2000 to 2020. Fractures were repaired using open reduction and fixation. An intramedullary Steinman pin was placed, exiting near the greater trochanter, and utilized for fracture reduction. Bicortical transfixation pins were placed distal and proximal to the fracture site. Poly(methyl methacrylate) (PMMA)-filled tubing connected the transfixation pins and proximal intramedullary pin, creating a Type 1a external skeletal fixator (ESF) tie-in. Follow up was obtained via medical records and phone interviews. RESULTS: Short-term survival rate was 7/10 (70%). Postanesthetic death occurred once. Postoperative complications occurred in all remaining cases. The most common findings were transfixation pin tract lucency and lameness (6/9 cases), implant dysfunction (5/9 cases), and infection (4/9 cases). Three of five cases with long-term follow up survived; all went on to productive careers. CONCLUSION: Although patients were prone to postoperative complications, short-term survival was comparable to previous reports. CLINICAL SIGNIFICANCE: The external skeletal fixator/intramedullary (ESF/IM) pin tie-in is less expensive and offers comparable success rates to other methods, providing a lower cost option for calf femoral fracture repair.

In Vitro and In Vivo Evaluation of Ultrasound-Triggered Release From Novel Spinal Device.

OBJECTIVES: Bacterial infection following spinal fusion is a major clinical concern with up to 20% incidence. An ultrasound-triggered bulk-release system to combat postsurgical bacterial survival was designed and evaluated. METHODS: Polylactic acid (PLA) clips were loaded with vancomycin (VAN) and microbubbles (Sonazoid, GE HealthCare) in vitro. Stability was determined over 14 days. VAN-loaded clips were submerged in water and insonated using a Logiq E10 scanner (GE HealthCare) with a curvilinear C6 probe. Doppler-induced VAN release was quantified using spectrophotometry. For in vivo testing, clips were loaded with methylene blue (MeB) solution and Sonazoid. These clips were implanted into a rabbit along the spine at L2 and L5, as well as a pig at L1 and L3, then insonated in Doppler mode using the C6 probe. RESULTS: Sonazoid microbubbles were better preserved when incubated in VAN compared with distilled water at 4°C, 25°C, and 37°C incubation temperatures (P = .0131). Contrast enhancement was observed from both solutions when incubated at 4°C storage conditions. Insonated clips achieved average cumulative VAN release of 101.8 ± 2.8% (81.4 ± 2.8 mg) after 72 hours. Uninsonated clips had only 0.3 ± 0.1% (0.3 ± 0.1 mg) average cumulative VAN release (P < .0001). Clips retrieved from the rabbit did not rupture with insonation nor produce MeB staining of surrounding tissues. In the pig, the PLA film was visibly ruptured and MeB tissue was observed following insonation, whereas the uninsonated clip was intact. CONCLUSION: These results demonstrate ultrasound-triggered release of an encapsulated prophylactic solution and provide an important proof-of-concept for continuing large animal evaluations for translational merit.

Microbubble cavitation restores Staphylococcus aureus antibiotic susceptibility in vitro and in a septic arthritis model.

Treatment failure in joint infections is associated with fibrinous, antibiotic-resistant, floating and tissue-associated Staphylococcus aureus aggregates formed in synovial fluid (SynF). We explore whether antibiotic activity could be increased against Staphylococcus aureus aggregates using ultrasound-triggered microbubble destruction (UTMD), in vitro and in a porcine model of septic arthritis. In vitro, when bacterially laden SynF is diluted, akin to the dilution achieved clinically with lavage and local injection of antibiotics, amikacin and ultrasound application result in increased bacterial metabolism, aggregate permeabilization, and a 4-5 log decrease in colony forming units, independent of microbubble destruction. Without SynF dilution, amikacin + UTMD does not increase antibiotic activity. Importantly, in the porcine model of septic arthritis, no bacteria are recovered from the SynF after treatment with amikacin and UTMD-ultrasound without UTMD is insufficient. Our data suggest that UTMD + antibiotics may serve as an important adjunct for the treatment of septic arthritis.