Patrick W. Whitlock

An Acellular, Allograft-Derived Meniscus Scaffold in an Ovine Model

PURPOSE The purpose of this study was to develop a meniscus scaffold that has increased porosity and maintains the native meniscus extracellular matrix in an ovine model. METHODS The medial menisci of skeletally mature ovine (n = 16) were harvested; half were made into meniscus scaffolds (n = 8), and half remained intact (n = 8). Intact and scaffold meniscus tissues were compared by use of histology, DNA content analysis, in vitro cellular biocompatibility assays, and ultrastructural analysis. An additional 16 knees were used to investigate the biomechanics of the intact meniscus compared with the meniscus scaffold. RESULTS DNA content and histology showed a significant decrease in cellular and nuclear content in the meniscus scaffold (P < .003). Biocompatibility was supported through in vitro cellular assays. Scanning electron microscopy and micro-computed tomography showed a substantial increase in porosity and pore connectivity in the meniscus scaffold compared with the intact meniscus (P < .01). There was no statistical difference between the ultimate load or elastic modulus of the intact and meniscus scaffolds. CONCLUSIONS In this study a meniscus scaffold was evaluated for potential clinical application as a meniscus transplant construct in an ovine model. The data showed that a decellularized meniscus scaffold with increased porosity was comparable to the intact meniscus, with an absence of in vitro cellular toxicity. Although some compositional alterations of the extracellular matrix are to be expected during processing, it is evident that many of the essential structural components remained functional with maintenance of biomechanical properties. CLINICAL RELEVANCE This meniscus scaffold has potential for future clinical application as a meniscus transplant construct.

Controlled formation of biosilica structures in vitro

Herein we describe the controlled formation of biosilica structures by manipulation of the physical reaction environment; we were able to synthesize arched and elongated silica structures using a synthetic peptide; the results presented here are evidence that in vitro biocatalysis may be controlled in order to form desired silica structures.

Evaluation of in vivo rotator cuff muscle function after acute and chronic detachment of the supraspinatus tendon: an experimental study in an animal model.

BACKGROUND Surgical repair of large chronic rotator cuff tears can be technically demanding because it requires manipulation of a muscle-tendon unit that is scarred, retracted, and stiffer than normal, all of which contribute to increased tension at the repair site. The purpose of the present study was to characterize the in vivo rotator cuff muscle-tendon unit function after acute and chronic injury at surgically relevant preload tensions. METHODS Sixty-two Sprague-Dawley rats were divided into a healthy, uninjured (control) group (n = 22), an acute injury group (n = 20), and a chronic injury group (n = 20) and underwent in vivo muscle force testing and electromyographic testing of the supraspinatus muscle-tendon unit at various preload tensions. RESULTS Preload tension affected the maximum supraspinatus muscle contractile force in all groups (p < 0.05). At the peak tension required to repair an acute tear, there was a 28% to 30% reduction in maximum tetanic contraction amplitude in all groups (p < 0.05). At the peak tension required to repair a chronic tear, there was a 40% to 53% reduction in maximal tetanic contraction amplitude in all groups (p < 0.05). The uninjured (control) group showed increased muscle endurance (p < 0.05) in comparison with the acute injury and chronic injury groups at all preload tensions. The chronic injury group showed reduced compound motor action potential amplitude (p < 0.05). CONCLUSIONS Both the acute and chronic injury groups demonstrated functional impairment related to increasing preload tensions. Higher repair tensions, associated with the chronic injury setting, resulted in greater functional impairment. The present study also demonstrates an association between increased time from rotator cuff tendon injury and impaired in vivo rotator cuff muscle electromyographic findings.

A novel process for optimizing musculoskeletal allograft tissue to improve safety, ultrastructural properties, and cell infiltration.

BACKGROUND This study evaluated the properties of scaffold derived from freeze-dried human Achilles tendon allograft for use in anterior cruciate ligament (ACL) reconstruction. Our hypothesis was that such an allograft could be processed using a method to remove cellular and infectious material, producing a cytocompatible, architecturally modified scaffold possessing tensile properties suitable for ACL reconstruction. METHODS Fifty-two allografts were provided by a tissue bank. Twenty-one were used as controls to assess cellularity, DNA content, microarchitecture, porosity, cytocompatibility, and tensile properties in vitro (n = 13) and in vivo (n = 8). Thirty-one were processed to produce scaffolds that were similarly assessed for these properties in vitro (n = 23) and in vivo (n = 8). The elimination of added enveloped and nonenveloped viruses was also determined in vitro after each processing step. RESULTS A subjective decrease in cellularity and a significant decrease in DNA content were observed in the scaffolds compared with the allografts from which they had been derived. The porosity was increased significantly, and the scaffolds were cytocompatible in vitro. Processing resulted in significantly increased elongation of the scaffolds (138% of the elongation of the unprocessed allograft) during tensile testing. No other significant differences in tensile properties were observed in vitro or in vivo. The number of infiltrating host cells and the depth to which those cells infiltrated were significantly greater in the scaffolds. No enveloped viruses and only two of 10(8) nonenveloped viruses were detected in the scaffolds after processing, corresponding to a sterility assurance level of 0.2 × 10(-7). CONCLUSIONS Allografts were processed using a method that removed cellular and infectious material to produce a decellularized, cytocompatible, architecturally modified scaffold with tensile properties that differed minimally from those of human allograft tissue both in vitro and in vivo. The scaffold production process also resulted in an increase in porosity that led to increased cell infiltration in vivo.

Fibrinogen adsorbs from aqueous media to microscopic droplets of poly(dimethylsiloxane) and remains coagulable

Fibrinogen binds from aqueous media containing it to droplets of linear trimethylsilyl-terminated poly(dimethylsiloxane) (PDMS) dispersed in those same media. Once bound, fibrinogen elutes from emulsified droplets of PDMS only very slowly, even when incubated in buffer that contains a physiologic concentration of the protein. The bound fibrinogen is coagulable, as indicated by the thrombin-dependent agglutination of droplets. Thus fibrinogen bound to droplets of PDMS renders an adhesive potential to the surface of the droplets, a potential that may have relevance to the biologic processing of the polymer in vivo.

The science of rotator cuff tears: translating animal models to clinical recommendations using simulation analysis

PurposeThe purpose of this article is to review basic science studies using various animal models for rotator cuff research and to describe structural, biomechanical, and functional changes to muscle following rotator cuff tears. The use of computational simulations to translate the findings from animal models to human scale is further detailed.MethodsA comprehensive review was performed of the basic science literature describing the use of animal models and simulation analysis to examine muscle function following rotator cuff injury and repair in the ageing population.ResultsThe findings from various studies of rotator cuff pathology emphasize the importance of preventing permanent muscular changes with detrimental results. In vivo muscle function, electromyography, and passive muscle–tendon unit properties were studied before and after supraspinatus tenotomy in a rodent rotator cuff injury model (acute vs chronic). Then, a series of simulation experiments were conducted using a validated computational human musculoskeletal shoulder model to assess both passive and active tension of rotator cuff repairs based on surgical positioning.ConclusionOutcomes of rotator cuff repair may be improved by earlier surgical intervention, with lower surgical repair tensions and fewer electromyographic neuromuscular changes. An integrated approach of animal experiments, computer simulation analyses, and clinical studies may allow us to gain a fundamental understanding of the underlying pathology and interpret the results for clinical translation.

Effect of cyclic strain on tensile properties of a naturally derived, decellularized tendon scaffold seeded with allogeneic tenocytes and associated messenger RNA expression.

Naturally derived tendon scaffolds have the potential to improve the treatment of flexor tendon injuries. Seeded and unseeded tendon scaffolds were maintained in the presence or absence of physiologic strain for 7 days. After 7 days, the tensile properties and associated messenger RNA expression were compared. Seeded scaffolds maintained in the absence of strain had significantly lower tensile properties than unseeded tendons and fresh-frozen tendons. The loss of tensile properties was associated with elevated matrix metalloproteinase-2 and collagen III expression. Tensile properties of seeded scaffolds maintained in the presence of strain for 7 days after seeding did not differ from those of fresh-frozen tendons. This study demonstrates that the tensile properties of seeded, naturally derived tendon scaffolds will degrade rapidly in the absence of cyclic strain. Seeded scaffolds used for tendon reconstruction should be maintained under cyclic strain to maintain essential tensile properties.

Management of pelvic discontinuity in revision total hip arthroplasty: a review of the literature

Pelvic discontinuity is a complex problem in revision total hip arthroplasty. Although rare, the incidence is likely to increase due to the ageing population and the increasing number of total hip arthroplasties being performed. The various surgical options available to solve this problem include plating, massive allografts, reconstruction rings, custom triflanged components and tantalum implants. However, the optimal solution remains controversial. None of the known methods completely solves the major obstacles associated with this problem, such as restoration of massive bone loss, implant failure in the short- and long-term and high complication rates. This review discusses the diagnosis, decision making, and treatment options of pelvic discontinuity in revision total hip arthroplasty.

A Tissue-Engineered Approach to Tendon and Ligament Reconstruction

A tissue-engineered scaffold for use in tendon and ligament reconstruction has the potential to reduce or prevent disease transmission from allografts and diminish any potential immunologic foreign-body response associated with allograft reconstruction. In addition, the possibility of “off-the-shelf” availability of such a scaffold would eliminate the donor site morbidity, increased surgical time, increased cost, and poor function associated with autologous tendon harvest (currently considered the “gold standard” in clinical applications). An ideal scaffold for tendon and ligament reconstruction would be: naturally derived from either allogeneic or xenogeneic material amenable to host-cell mediated remodeling in vivo; devoid of cellular material to minimize inflammatory potential, disease transmission, and host immune response; cytocompatible; of optimal micro-architecture to promote efficient cell seeding, infiltration, and attachment of the recipient’s own cells prior to or after implantation; and distinguished by sufficient biomechanical integrity to withstand rehabilitation until complete remodeling has occurred. Such a scaffold should serve as template for cell attachment, promote rapid remodeling, possess increased strength, demonstrate improved healing, and permit early rehabilitation and return to function after implantation. In summary, an optimized scaffold would have the potential to vastly improve the treatment of tendon and ligament injuries, especially those associated with tumor, trauma, and congenital deficiencies where autograft or allograft tissue might not be available in sufficient quantity for reconstruction.

Proving Osteoinductive Potential of a Decellularized Xenograft Bone Substitute

Background Large bone defects remain a major clinical challenge for orthopaedic surgeons. Tissue engineered bone grafts have garnered increased attention as a solution to this problem. One ideal property of any bone graft is osteoinductivity or the ability to stimulate progenitor cell differentiation into a bone forming lineage. Questions Is the osteoinductive potential of a porcine bone xenograft maintained in vitro after undergoing a novel decellularization and oxidation process? Are porcine bone scaffolds osteoinductive in an in vivo animal model? Methods In Vitro – C2C12 pre-osteoblasts were seeded on the scaffold or a commercial grade demineralized bone matrix (DBM) to study osteogenic differentiation and compare osteoinductive potential. MC3T3-E1 pre-osteoblasts were seeded on the scaffold and compared to a control monolayer to identify early markers of osteogenic differentiation. In Vivo – MC3T3-E1-seeded scaffolds were implanted subcutaneously in mice and assessed for markers of early osteogenic differentiation, new bone formation (micro-computed tomography and histological assessment), and vascular infiltration (histology). Results Osteoinductive potential was demonstrated in in vitro experiments by similar osteogenic marker expression compared to DBM and significantly greater expression than a control monolayer. Osteoinductivity was confirmed with in vivo experiments showing both new bone formation and vascular infiltration. Conclusion Porcine bone maintains osteoinductive properties after decellularization and oxidation. Clinical Relevance This construct could potentially serve as a bone graft substitute maintaining the osteoinductive potential of native bone. The unrestricted supply and controlled donor biology may satisfy a large clinical need for orthopaedic cases requiring bone grafting.