Chantelle C. Bozynski

The effect of female presence on spermiation, and of male sexual activity on ‘ready’ sperm in the male guppy

Abstract This study examines the effect of female presence on the amount of ‘strippable’ sperm, and the relationship between male sexual activity and the amount of ‘ready’ sperm in the male guppy Poecilia reticulata . The amount of strippable sperm was greater in males paired for one week with females than in isolated males. We separated these ‘paired’ males from females by a clear partition and thus exposed males to visual and possibly olfactory stimuli. In the second experiment, control males spent more time attending females and performed more sexual acts during a 15-min test period than males previously stripped of spermatophores. These results indicate that females play an important role in stimulating sperm availability in males and that the amount of ready sperm influences male sexual behaviour. Copyright 2003 The Association for the Study of Animal Behaviour. Published by Elsevier Science Ltd. All rights reserved.

Multiple injections of leukoreduced platelet rich plasma reduce pain and functional impairment in a canine model of ACL and meniscal deficiency

Platelet rich plasma (PRP) is used to treat many musculoskeletal disorders. We used a canine model to determine the effects of multiple intra‐articular injections of leukoreduced PRP (ACP) on anterior cruciate ligament healing, meniscal healing, and progression of osteoarthritis (OA). With Animal Care and Use Committee (ACUC) approval, 12 dogs underwent partial ACL transection and meniscal release in one knee. At weeks 1, 2, 3, 6, and 8 after insult, dogs were treated with intra‐articular injections (2 ml) of either ACP (n = 6) or saline (n = 6). Dogs were assessed over 6 months to determine comfortable range of motion (CROM), lameness, pain, effusion, kinetics, and radiographic and arthroscopic assessments. At 6‐month endpoint, dogs were assessed for ACL material properties and histopathology. Saline‐treated dogs had significantly (p < 0.04) more CROM loss, significantly (p < 0.01) more pain, significantly (p < 0.05) more severe lameness, significantly (p < 0.05) lower function, and significantly (p < 0.05) lower %Total Pressure Index in affected hindlimbs compared to ACP‐treated dogs. Radiographic OA increased significantly (p < 0.01) over time within each group. Arthroscopically, saline‐treated knees showed moderate to severe synovitis, further ACL disruption, and medial compartment cartilage loss, and ACP‐treated knees showed evidence of ACL repair and less severe synovitis. ACL material properties in ACP‐treated knees were closer to normal than in saline‐treated knees, however, the differences were not statistically significant. ACL histopathology was significantly (p< 0.05) less severe in ACP‐treated knees compared to saline‐treated knees. Five intra‐articular injections of leukoreduced PRP had beneficial effects for ACL healing, improved range of motion, decreased pain, and improved limb function for up to 6 months in this model.

Importance Of Donor Chondrocyte Viability in Osteochondral Allografting

Objectives: Osteochondral allografting (OCA) provides a biologic treatment option for functional restoration of large articular cartilage defects in multiple joints. While successful outcomes after OCA have been linked to viable donor chondrocytes, the importance of donor cell viability has not been comprehensively validated. The purpose of this study was to use a canine model to determine the importance of donor chondrocyte viability at the time of implantation with respect to functional success of femoral condylar OCAs based on gross, cell viability, histologic, biochemical, and biomechanical outcome measures. Methods: With IACUC approval, adult female dogs (n = 16) were implanted with 8-mm cylindrical osteochondral allografts from male dogs in the lateral and medial femoral condyles of one knee. Osteochondral allografts were preserved for 28 or 60 days after procurement and chondrocyte viability was quantified prior to implantation. Two different storage media, temperatures and time points were used in order to obtain a spectrum of %chondrocyte viability at the time of implantation. A successful outcome was defined as an osteochondral allograft that was associated with graft integration, maintenance of hyaline cartilage, lack of associated cartilage disorder, and lack of fibrillation, fissuring, or fibrous tissue infiltration of the allograft based on subjective radiographic, arthroscopic, gross, and histologic assessments at 6 months after implantation. Results: Chondrocyte viability ranged from 23% to 99% at the time of implantation. All successful grafts had greater than 70% chondrocyte viability at the time of implantation and no graft with chondrocyte viability <70% was associated with a successful outcome. Live-dead stained sections and histologic findings with respect to cell morphology suggested that successful grafts were consistently comprised of viable chondrocytes in lacunae, while grafts that were not successful were comprised of non-viable chondrocytes with infiltration of fibroblasts from the surrounding recipient tissues. In situ PCR (FISH) assays were performed in an attempt to distinguish donor (male) cells from recipient (female) cells. Unfortunately, this technique was exceptionally difficult to perform on intact articular cartilage sections, and consistent, repeatable data could not be obtained from this testing. However, the data did support histologic and live-dead data, which strongly suggested that successful grafts retained viable donor (male) chondrocytes and unsuccessful grafts degraded and were replaced by fibrous tissue populated with recipient (female) fibroblasts. Conclusion: Viable chondrocytes in osteochondral allografts at the time of transplantation are primarily responsible for maintenance of donor articular cartilage health long term. Optimizing chondrocyte viability in all aspects of osteochondral allografting - including procurement, processing, storage, transportation, and surgical implantation - needs to be a primary focus for clinical use of OCA.

Evaluation of Partial Transection versus Synovial Debridement of the ACL as Novel Canine Models for Management of ACL Injuries.

A major hurdle in investigating important clinical questions in knee ligament treatment is a lack of valid translational animal models. This study characterizes the effects of partial transection versus synovial debridement of the anterior (cranial) cruciate ligament (ACL) in dogs. A total of 27 adult purpose-bred research hounds underwent surgery and were assessed over the following 8 weeks. Dogs were randomized into the following three ACL status groups: sham control (n = 9), intact ACL with synovial debridement (exposed ACL) (n = 9), and partial transection of the ACL (partial tear ACL) (n = 9). Dogs in the exposed ACL group and partial tear ACL group had significantly (p < 0.05) more severe lameness, pain, effusion, reduced function, and reduced comfortable range of motion compared with controls, with the partial tear ACL group being most severely affected. More severe ACL and whole-joint pathology, and radiographic scores for osteoarthritis were present in the partial tear ACL group compared with exposed and/or sham control group. On the basis of these findings, biologic components of ACL injury (exposed ACL) played a role in whole-joint inflammation, but the clinical and pathological effects were more severe when both biologic and biomechanical components were present (i.e., partial tear ACL). These novel canine models were successfully developed to evaluate partial transection versus synovial debridement of the ACL and these models will be used to evaluate treatment options for acute management of ACL injuries.

Evaluation of synthetic osteochondral implants.

This translational animal model study was designed to assess function, bone ingrowth and integration, and joint pathology associated with two different synthetic, bilayered osteochondral implants over a 3-month period after implantation into the femoral condyles of dogs. SynACart-Titanium (n = 6) and SynACart-PEEK (n = 6) (Arthrex, Naples, FL, and Sites Medical, Columbia City, IN) implants were press-fit into the lateral or medial femoral condyle (alternating location) of purpose-bred adult research dogs. Dogs were humanely euthanized 3 months after surgery and the operated knees were assessed radiographically, arthroscopically, grossly, and histologically. Based on all assessments, both types of implants were well tolerated and safe with no evidence for infection, migration, or rejection. Half of the SynACart-PEEK implants showed radiographic and histologic evidence of poor incorporation with all of these being in the lateral femoral condyle. SynACart-Titanium implants were considered effective in terms of integration into bone, lack of damage to surrounding and apposing articular cartilage, and maintenance of implant integrity and architecture for the duration of the study.

A Canine Arthroscopic Anterior Cruciate Ligament Reconstruction Model for Study of Synthetic Augmentation of Tendon Allografts.

Abstract Novel graft types, fixation methods, and means for augmenting anterior cruciate ligament (ACL) reconstructions require preclinical validation prior to safe and effective clinical application. The objective of this study was to describe and validate a translational canine model for all‐inside arthroscopic complete ACL reconstruction using a quadriceps tendon allograft with internal brace (QTIB). With institutional approval, adult research hounds underwent complete transection of the native ACL followed by all‐inside ACL reconstruction using the novel QTIB construct with suspensory fixation (n = 10). Contralateral knees were used as nonoperated controls (n = 10). Dogs were assessed over a 6‐month period using functional, diagnostic imaging, gross, biomechanical, and histologic outcome measures required for preclinical animal models. Study results suggest that the novel QTIB construct used for complete ACL reconstruction can provide sustained knee stability and function without the development of premature osteoarthritis in a rigorous and valid preclinical model. The unique configuration of the QTIB construct—the combination of a tendon allograft with a synthetic suture tape internal brace—allowed for an effective biologic‐synthetic load‐sharing ACL construct. It prevented early failure, allowed for direct, four‐zone graft‐to‐bone healing, and functional graft remodeling while avoiding problems noted with use of all‐synthetic grafts.

Does Anterior Cruciate Ligament Innervation Matter for Joint Function and Development of Osteoarthritis

&NA; Complications following anterior cruciate ligament (ACL) injury and reconstruction that include chronic dysfunction, second ACL injury, and posttraumatic osteoarthritis (OA) may be interrelated and stem from the inability to fully restore native ACL integrity and function. The loss of ACL sensory input following injury may significantly contribute to joint dysfunction. We developed a novel preclinical animal model to assess the contributions of ACL sensory afferents to knee joint function and health. We hypothesized that ACL sensory denervation would manifest in knee joint dysfunction and development of early OA. Purpose‐bred, adult research dogs (n = 9 dogs, 18 knees) underwent arthroscopic surgery to create three treatment groups: (1) partial ACL tear, (2) ACL denervated, and (3) whole‐joint denervated. A neurotoxin injected directly into the ACL or into the joint space was used to induce sensory denervation, and sham procedures were done on contralateral knees. After intervention, dogs participated in a regimented exercise program. Gait analysis and clinically relevant functional assessments were performed. At week 12, the animals were humanely euthanatized for arthroscopic, gross, and histologic assessments. ACL partial tear group demonstrated the greatest overall knee dysfunction. Clinical measures of function revealed a significant difference between the ACL partial tear and ACL denervated group (p < 0.05), but these differences were not observed between the ACL and whole‐joint denervated groups (p > 0.05). A significant reduction in limb loading was experienced by the ACL partial tear group compared with controls (p < 0.05) but not between other groups. Arthroscopic evaluation found no evidence for overt articular cartilage damage, meniscal pathology, or osteophyte formation was noted in any group. No significant differences (p > 0.05) were observed in ACL pathology and OA severity scores between the ACL partial tear and the ACL denervated groups. The results of our study indicate that ACL sensory loss may contribute to joint dysfunction and subsequent OA changes. Further investigation and development of this model are important to improve clinical therapies and optimize patient outcomes following ACL injury.

Development of a Novel Canine Model for Posttraumatic Osteoarthritis of the Knee.

Translational models of posttraumatic osteoarthritis (PTOA) that accurately represent clinical pathology need to be developed. This study assessed a novel canine model for PTOA using impact injury. Impacts were delivered to the medial femoral condyle of dogs using a custom-designed impactor at 20, 40, or 60 MPa. Functional assessments and magnetic resonance imaging (MRI) were performed at 2 and 12 weeks, and arthroscopic and histologic assessments were performed at 12 weeks after injury. At 2 and 12 weeks, dogs had observable lameness, knee pain, effusion, loss in range of motion (ROM) and dysfunction in both hindlimbs with severity correlated strongly (r > 0.77) to impact level. At 12 weeks, function, pain, effusion, and ROM were significantly (p < 0.049) worse in knees impacted at 40 and 60 MPa compared with 20 MPa. MRI showed consistent cartilage and subchondral bone marrow lesions, and arthroscopy revealed synovitis and cartilage destruction in impacted knees, with increased severity for 40 and 60 MPa impacts. Histopathology was significantly (p = 0.049) more severe in 40 and 60 MPa and strongly correlated (r = 0.93) to impact level. This novel translational model appears to be valid for investigation of PTOA, including determination of temporal mechanisms of disease and preclinical testing for preventative and therapeutic strategies.

A canine hybrid double‐bundle model for study of arthroscopic ACL reconstruction

Development and validation of a large animal model for pre‐clinical studies of intra‐articular anterior cruciate ligament (ACL) reconstruction that addresses current limitations is highly desirable. The objective of the present study was to investigate a translational canine model for ACL reconstruction. With institutional approval, adult research hounds underwent arthroscopic debridement of the anteromedial bundle (AMB) of the ACL, and then either received a tendon autograft for “hybrid double‐bundle” ACL reconstruction (n = 12) or no graft to remain ACL/AMB‐deficient (n = 6). Contralateral knees were used as non‐operated controls (n = 18) and matched canine cadaveric knees were used as biomechanical controls (n = 6). Dogs were assessed using functional, diagnostic imaging, gross, biomechanical, and histologic outcome measures required for pre‐clinical animal models. The data suggest that this canine model was able to overcome the major limitations of large animal models used for translational research in ACL reconstruction and closely follow clinical aspects of human ACL reconstruction. The “hybrid double‐bundle” ACL reconstruction allowed for sustained knee function without the development of osteoarthritis and for significantly improved functional, diagnostic imaging, gross, biomechanical, and histologic outcomes in grafted knees compared to ACL/AMB‐deficient knees.

Validation of the Missouri Osteochondral Allograft Preservation System for the Maintenance of Osteochondral Allograft Quality During Prolonged Storage

Background: Fresh osteochondral allografts (OCAs) are limited in availability. The Missouri Osteochondral Allograft Preservation System (MOPS) has been reported to effectively preserve OCAs twice as long as current tissue bank protocols in preclinical studies. Hypothesis: The viable chondrocyte density (VCD) in OCAs preserved for up to 70 days using the MOPS will not be significantly different from day 0, and the VCD in MOPS-preserved OCAs will be significantly higher than for standard tissue bank preservation. Media changes during preservation will significantly improve the VCD. Study Design: Controlled laboratory study. Methods: Femoral condyles harvested from qualified donors (n = 12) were quartered (n = 48), assigned to 1 of 4 treatment groups (tissue bank protocol at 4°C or MOPS at 25°C, with or without media changes), and preserved for 0, 28, 56, or 70 days and assessed for the VCD and histopathological characteristics. In addition, osteochondral explants were created from the femoral condyles of 12 donors (n = 36 explants), assigned to the same groups and time points, and tested for biomechanical properties. Results: MOPS-preserved OCAs maintained the day 0 VCD through 56 days. OCAs stored using current tissue bank protocols had a significantly lower VCD compared with day 0 and the MOPS by day 28. OCA histological and biomechanical properties did not significantly change from day 0 for any group. Conclusion: The MOPS preserved essential OCA viability and quality at significantly higher levels than current tissue bank protocols for at least 56 days after procurement. Clinical Relevance: Improving the viability and duration of OCA preservation provides potential benefits to tissue banks, donor families, surgeons, and patients with respect to tissue use, financial costs, and outcomes.