Tornier Medical Education

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1 Tornier Medical Education D E C E M B E R 7, Orthopaedic Uses of Tissue Matrices in Tendon Repair: Why All Tissue Matrices are Not Created Equal John R. Harper, Ph.D, Research Department, LifeCell Corporation, a KCI Company, Branchburg, New Jersey Kevin L. Ohashi, Ph.D & Dale R. Peterson, Ph.D, Clinical and Research Department, Tornier, Inc., Edina, Minnesota

2 Orthopaedic Uses of Tissue Matrices in Tendon Repair: Why All Tissue Matrices are Not Created Equal By John R. Harper*, Ph.D,, Kevin L. Ohashi*, Ph.D & Dale R. Peterson, Ph.D Introduction Significant advancements in tendon repair using improved surgical techniques and development of better anchors, sutures, and surgical instrumentation for aiding fixation have improved the overall clinical outcome of procedures such as rotator cuff or Achilles tendon repair. Unfortunately, these improvements have not addressed the fundamental problem with many of the tendon repairs the native tendon often is significantly compromised (thin or friable) at the repair site. As a result, we continue to see reports of large failure rates (34-94%) 1-3 or unsatisfactory functional improvements 4, 5 in the repair of the rotator cuff tears or Achilles tendon ruptures. Biological matrices i were developed for use in orthopaedic soft tissue applications with the hope of improving the clinical outcomes of these tendon repairs. 6 It was hypothesized that if one could reinforce the repair site and encourage tissue remodeling through incorporation of the matrix to form new tendon tissue, the repair would have a better clinical result. Extensive in vitro (bench top biochemical and mechanical) and in vivo studies were undertaken to demonstrate that this was a safe and efficacious approach. 7 Although these early biological matrices showed great promise in preclinical studies, a puzzlingly high clinical failure rate with aseptic cysts, edema, heat, immunologic reactions, and, in rare cases, expulsions of the matrix was observed. 5, 8 Reasons for failure of matrices: The human biological response to a matrix derived from human, animal or synthetic sources is more complex than early testing could capture. Regardless of the basic source of the biological matrix there are critical factors such as processing techniques, sterilization method, surgical technique, repair application, and rehabilitation programs that contribute to the clinical success of the procedure. This review of orthopaedic biological matrices describes and explains the fundamental elements that may influence the clinical success or failure of these devices or tissues ii and introduces a novel biological matrix device ( reconstructive tissue matrix, Tornier, Inc., Edina, MN USA)., developed in partnership between Tornier and LifeCell Corporation, a KCI company, addresses some of the concerns of earlier devices and tissue and satisfies the clinical need for a device that biologically augments and mechanically reinforces orthopaedic soft tissue repairs. Goals of this Review of Biological Matrices 1. Describe the ideal characteristics of a biological matrix and how previously developed matrices attempted to meet some of these characteristics. 2. Provide a fundamental understanding of why all matrices are not created equal. Unlike some may suggest, material source (e.g. human or animal) is not as relevant as the processing of the matrix. Depending on the processing used to clean, stabilize, sterilize, and package the matrix, the biological mechanism of action of the matrix will greatly differ. 3. Introduce reconstructive tissue matrix and describe how it approaches the characteristics of an ideal matrix for orthopaedic soft tissue repair in terms of sterility, tissue reinforcement, and ease of use. Mechanism of Action Ideal biological reinforcement of orthopaedic soft tissue repair should support biological incorporation and remodeling of the matrix into host tissue during the healing process. Unfortunately, most matrices used in orthopaedic soft tissue applications do not provide the proper environment to support regeneration but rather elicit a rapid response from the host. In such a repair response, an active inflammatory response directly attributable to the biological matrix occurs and may lead to scar tissue formation or encapsulation (if the matrix cannot be resorbed) by host tissue. The Tornier Medical Education *LifeCell Corporation, a KCI Company, Branchburg, NJ *Clinical and Research Department, Tornier, Inc., Edina, MN 1

3 disadvantage of replacing host native tissue with scar tissue is the significantly compromised biomechanical properties of the scar tissue as compared to native host tissue. The disadvantage of encapsulating the matrix is the potential for a chronic inflammatory response, reliance on nonphysiological tissue for mechanical strength, and potential for long-term infection due to the acellular environment. Using extensive studies in primates, the biological mechanism of action of matrices can be grouped into three general categories: (a) a mechanism of action that supports regeneration during which the matrix is being remodeled into native host tissue; (b) a resorptive mechanism of action characterized by a rapid repair response leading to resorption of the matrix and scar formation; and, (c) an encapsulative mechanism of action in which an inflammatory response leads to encapsulation of the matrix. Table 1 provides an overview of the events that lead to a positive (supports regeneration) or negative (resorption or encapsulation) recognition of the biological matrices. The biological response to different matrices is categorized by dividing these responses into positive or negative recognition and their mechanistic responses. Table 1: Summary of Matrix Mechanisms of Action Recognition Type Mechanism of Action Key Advantages Key Disadvantages R E G E N E R AT I V E Positive Recognition Body recognizes as self + Regenerative Body accepts and integrates the intact tissue matrix as part of the host through rapid revascularization and cell repopulation Supports regeneration /remodeling of matrix into native host tissue Remodeled tissue has better biomechanical properties as compared to scar tissue Difficult to process tissue properly to encourage a response that supports regeneration Negative Recognition Body recognizes as foreign - Resorption (scar formation) Body attacks the damaged tissue to break it down and eliminate it Rapid matrix removal and filling in of defect with scar tissue Biomechanical properties of scar tissue are inferior to normal tissue Active inflammatory response can lead clinically to pain and swelling Some matrices require re-intervention and removal of matrix R E PA R AT I V E Negative Recognition Body recognizes as foreign - Encapsulation Body attacks the cross-linked tissue to extrude or wall it off from the host Prevents catastrophic resorption Inhibits native cellular and vascular in-growth Active inflammatory response can lead clinically to pain and swelling Some matrices require re-intervention and removal of matrix 2

4 Different commercially available matrices were evaluated in a primate animal model. The matrices were implanted into an interpositional model created by removing a 4 x 7 cm rectangular section of the abdominal fascia of African green monkeys and suturing the matrices into the defect. 9 The response was evaluated at time points up to 6 months after implantation. Examples of positive and negative responses are shown in Figure 1, which illustrates the incorporation and remodeling associated with a matrix supporting regeneration (), scarring and contracture observed with a matrix derived from small intestine submucosa (SIS), and tissue encapsulation of a highly cross-linked matrix. R E G E N E R AT I V E R E PA R AT I V E Supports Regeneration Resorption/Scar Formation Encapsulation (a) (b) (c) Figure 1: Six-month macroscopic observations of biological matrices in primates: (a) porcine tissue matrix showing support of regeneration (); (b) porcine SIS residual inflammation and tissue contraction (scarring/fibrosis); and, (c) cross-linked porcine dermis showing limited vascularization and cellular repopulation of graft material and encapsulated matrix. Orthopaedic Soft Tissue Biological Matrix The goal of matrix development is to support this host regenerative response while adequately reinforcing the repair site. Therefore, the processing should accomplish several objectives: 1. Significantly reduce cellular and antigenic components 2. Retain critical biochemical components without damaging them 3. Minimize the inflammatory response 4. Support early cellular repopulation and revascularization 5. Incorporate mechanical properties sufficient to provide reinforcement of the repair 6. Terminally sterilize the matrix to medical device standards 7. Stabilize and package the matrix so it can be stored at room temperature and used immediately without hydration Based on the clinical outcomes we can divide the existing biological matrices into two principal categories: (a) reparative matrices; and, (b) matrices that support regeneration. The reparative matrices achieve some of the attributes listed above; GraftJacket is a matrix that supports regeneration but does not have all these desired attributes. Reconstructive Tissue Matrix achieves all seven goals to support a regenerative response with the benefits of sterility, handling, and ease of use. Table 2: Commercial Reparative Matrix Products Reparative Matrix Technologies Tissue Source Cross-linked Reduction of Hyperacute Rejection Antigens Sterilized to 10-6 SAL Supports Regeneration Restore (DePuy/ J&J) Porcine Small Intestinal Submucosa (SIS) No No Yes No R E PA R AT I V E CuffPatch (Biomet and Organogenesis) ZCR (Zimmer and TSL) TissueMend (Stryker/ TEI Biosciences) Porcine Small Intestinal Submucosa (SIS) Yes No Yes No Porcine Dermis Yes No Yes No Fetal Bovine Dermis Not Intentionally No Yes No OrthAdapt (Pegasus Biologics) Equine Pericardium Yes No Unknown No 3

5 Current Biological Matrix Products Reparative matrix technologies: Reparative matrix technologies utilize a wide range of processes and treatments to provide a collagen matrix. However, the critical role of matrix structure and biochemistry in supporting orthopaedic soft tissue regeneration was not widely appreciated during the development of the reparative matrices. Commercially available reparative products are shown in Table 2. Consequence of damaged matrices: Patients immune systems can detect and react to matrices which have been damaged by the manufacturing process. Preparing a good matrix requires careful balancing of the conditions to remove cells and antigens without removing or damaging the many desirable components in the extracellular matrix. Damage may be caused by ph excursions, solvents, elevated temperature, dehydration, reactive chemicals, or ionizing sterilization methods. Damaged matrices provoke an inflammatory response which in extreme cases can be so severe that the matrices are resorbed in just a few weeks following implantation resulting in excessive scarring. Figure 2 shows two examples of damaged matrices. Even after six months the damaged matrices are filled with lymphocytes indicating immune rejection of the matrices. Macrophages Lymphocytes Foreign Body Giant Cell (a) Figure 2: (a) Porcine SIS matrix; and, (b) fetal bovine dermis after 6 months in the primate abdominal interpositional model showing a significant number of inflammatory cells present. Cross-linking of matrices: Several matrix technologies utilize tissue cross-linking methods to provide longer term mechanical strength to the matrix, minimize rapid resorption or compensate for damage during sterilization by irradiation. However, cross-linking also damages the natural matrix and extracellular matrix components. Cellular repopulation has been shown to be hindered by such matrices (two examples are given in Figure 3). Without active cellular repopulation, revascularization of the tissue does not occur. As a result, the matrix remains prone to infection, but more importantly the matrix has limited potential for remodeling or supporting regeneration of new host tissue. Histological and gross examination of specimens at six month shows a chronic foreign body reaction along with encapsulation of the matrix. (b) Lymphocytes Poor Cell Infiltration Macrophages No Cellular or Vascular Ingrowth Macrophages Lymphocytes Foreign Body Giant Cell No Cellular or Vascular Ingrowth Inflamation (a) (b) Figure 3: Six-month explants of (a) cross-linked porcine dermis; and, (b) cross-linked equine pericardium in the primate interpositional model (H&E) showing acellular matrix regions and significant number of inflammatory cells present. Immune reaction to xenogeneic matrices: If a matrix originates from an animal source (xenogeneic material) significant reduction of specific antigenic components should be performed. Antigenic components seen by the human host can result in an inflammatory reaction to the matrix. The human body has an inflammatory response to xenogeneic tissues. Specifically, tissue from lower species (such as pig, cow, horse) sources have a macromolecule known as the α-gal antigen that is recognized by the human and primate immune system as foreign. In fact, higher primates such as humans have antibodies that readily recognize the α-gal antigen. Accordingly, reduction of this antigen during processing is critical to prevent a hyperacute response to the device matrix. 14 Recognition of xenogeneic reaction to animal sourced matrices: Clinical use of the Restore matrix in rotator cuff repair reinforcement showed high rates of rotator cuff repair failure in clinical studies. 5, 8 Feedback on the Restore matrix appears to also show high incidence of postoperative edema, pain, and swelling 5, 8 unrelated to infection. Up to 20-30% of patients have shown a hyperacute sensitivity to xenogeneic tissues 5 which generally leads to reoperation. Zheng et al 2005 have demonstrated that the Restore matrix processing, in fact, did not remove all the porcine cellular DNA material. 10 In addition, the processing of the matrix did not include reduction of the hyperacute rejection antigens (α-gal antigen). Consequently, the high immunological response to the Restore matrix is likely associated with the insufficient processing of the material to remove the antigenic components from the matrix. Figure 4 is representative data showing the fold increase in IgG antibody titer to the α-gal antigen as a function of time for various products in a primate model. These data are in contrast to the advanced tissue matrix () described on the following page. 4

6 Fold Increase in α-gal Antibody Titer (a) 10 1 Orthadapt TissueMend Time (days) Time (days) Time (days) (b) (c) 10 1 ZCR Figure 4: Representative curves from (a) OrthAdapt; (b) TissueMend; and, (c) ZCR showing the fold increase in α-gal antibody titer to α-gal antigens in the matrix using a primate model. Two important observations can be made from these studies: (a) cross-linking of matrices does not hide the α-gal antigen; and, (b) even purified collagen material maintains significant antigenic components. GraftJacket matrix: Wright Medical introduced a matrix that supports regeneration based on human dermis with the objective of supporting regeneration of host tissue (see Table 3). Unlike other commercial products, this biological matrix is not a device but human allograft tissue. As such this matrix is not regulated under FDA device regulations. Significant advancements in matrix process development were made to limit damage associated with removal of donor cellular components. Therefore, allograft rejection is not an issue with this matrix. Furthermore, because the matrix is derived from human dermis there is no need to address antigenic response to α-gal antigen. To encourage remodeling of the matrix and generation of new host tissue no cross-linking technology has been utilized with this allograft product. It is noteworthy to mention that sterilization of this biological matrix is not performed the matrix is processed under aseptic conditions. Good clinical results indicating a mechanism of action that supports regeneration with GraftJacket have been reported. However, limitations associated with handling characteristics, such as lengthy hydration time and specific orientation requirements are inconvenient during the procedure. In addition, this matrix is not sterilized and also may be subject to institutional concerns associated with traditional allograft tissue such as variations in properties Table 3: Allograft Reconstructive Matrix Technology Technology GraftJacket (Wright Medical) Tissue Source Human Dermis Crosslinked No Sterilized to 10-6 SAL and quality of matrix material based on inconsistent properties of donor tissues (i.e. age, history). Tornier introduced the next generation reconstructive matrix technology (Table 4) to address some of the inherent limitations of the reconstructive allograft-based matrix (Table 3). The development goals were: 1. To use stable, consistent, well controlled and mechanically sufficient source tissue (porcine dermis) 2. To significantly reduce the α-gal antigen responsible for xenogeneic response 3. To have an intact (non-damaged) matrix to support regeneration 4. To have a packaged sterile matrix, ready to use, and stored at room temperature 5. To have a non-cross-linked matrix to allow rapid cell population and vascular in-growth to encourage remodeling and support tissue regeneration 6. To tailor the device as appropriate for orthopaedic soft tissue use Evidence to Demonstrate s Ability to Support Regeneration was developed through a series of preclinical tests with the goal of achieving the key characteristics of GraftJacket while introducing more consistent handling, simpler storage, and terminal sterilization. No Supports Regeneration Yes R E G E N E R AT I V E 5

7 Table 4: Next Generation Reconstructive Matrix Technology Technology Tissue Source Cross-linked (Tornier) Porcine Dermis No Sterilized to 10-6 SAL Yes Supports Regeneration Yes R E G E N E R AT I V E processing: Through five years of advanced research and development in the primate model, a process was developed which successfully reduces antigenic material while preserving the remainder of the matrix even after terminal sterilization. Primates were used in these studies to mimic the human response to the α-gal antigen. Study results have allowed prediction of the expected biological reaction and mechanism of action because the primates have a very high homology to humans. The processing removes the cells and specifically reduces the residual α-gal remaining in the porcine extracellular matrix. The intact extracellular matrix supports regeneration. Accordingly, the goal is to remove all the cellular components and reduce the α-gal, but not alter or damage the structure or biochemistry of the patch. It is easy to inadvertently extract biochemical elements if one is not careful. Through a complex series of gentle treatments, the process takes great care to not damage or denature the matrix. The resulting matrix supports tissue regeneration. model shows cellular repopulation without a non-specific inflammatory response as observed in the reparative matrix devices. The matrix is already infiltrated with host fibroblasts and new capillaries. After six months is well integrated into the abdominal fascia. The cell density is returning to normal levels; the fibroblasts are mature; and the collagen matrix is being remodeled to a more linear orientation like that in normal fascia versus the amorphous woven collagen structure of dermis. Thus, the matrix has supported regeneration across a defect which would otherwise fill with scar tissue. These findings are consistent with the early clinical experience in which more than patients have received implants with follow-up for up to 5 months in orthopaedic application, and more than 5,000 patients with follow-up over one year in general surgery applications. The tests used in the development of included: 1. Preclinical histological evidence from primate models to demonstrate structurally intact matrix post-processing and evidence to support regeneration 2. Reduction of Antigenic Component Study to demonstrate significant reduction in antigen (α-gal) is achieved through matrix processing 3. Use of Collagenase Susceptibility Assay to independently demonstrate that the matrix is not denatured, cleaved, or cross-linked after processing 4. Use of Differential Scanning Calorimetry (DSC) Study to demonstrate that the collagen matrix is not denatured or cross-linked after processing and sterilization 5. Biomechanical study to demonstrate mechanical properties of intact matrix post-processing Histological evidence: Evidence that supports tissue regeneration is shown in Figure 5 where the device after just two weeks in the primate abdominal interpositional (a) Figure 5: after (a) two weeks; and, (b) six months in primate abdominal interpositional model (H&E stain) shows early cellular repopulation and revascularization and later evidence of matrix remodeling with fibroblasts distributed throughout the repair region.. To contrast the different mechanisms of action, histological sections of different matrices were taken at six months in a primate abdominal implantation model and are shown in Figure 6. The left-hand image (a) demonstrates that supports a regenerative mechanism of action whereas images (b) and (c) illustrate resorption and encapsulation, respectively, seen in other types of matrices. (b) 6

8 R E G E N E R AT I V E R E PA R AT I V E (a) (b) (c) Figure 6: (a) histology images look normal, showing capillaries, fibroblasts, and red staining of aligned collagen; (b) SIS samples show scarring and a large number of inflammatory cells; and, (c) cross-linked porcine dermis shows a capsule forming around the matrix with chronic inflammation. Xenogeneic response: The role of xenogeneic antigens in matrix performance has been debated over the years. Since most antigens are found on or in the cells in the tissue, all companies try to remove cells from their matrix products. However the α-gal antigen is found in the matrices derived from xenogeneic tissue. To demonstrate that the α-gal antigen has been reduced during the matrix processing, samples of the matrix were taken and histological sections were prepared (Figure 7) using α-gal antigen specific immunohistochemical staining. 14 Staining an acellular xenograft matrix prior to processing results in a uniform brown color under histological examination throughout the matrix (7a) indicating the matrix is fully loaded with α-gal antigen. However, by reducing the α-gal load in the tissue through careful processing, one can see the change in the degree of staining in the histology image (7b). In the graph shown (7c), an untreated xenograft material was implanted into old world primates (African green monkeys) and the antibody titer to the α-gal was shown to dramatically increase. Both primates and humans have pre-existing antibodies to the α-gal antigen. GraftJacket (which is derived from human dermis and does not have α-gal antigens) and did not show changes in the antibody titer level. This demonstrates that reduction of the α-gal antigen minimizes the primate immune response to xenograft tissues. (Source: Data on file at LifeCell Corporation. Correlation of this data to results in humans has not been established). (a) (c) Antibody Concentration (µg/ml) Figure 7: (a) Immunohistochemical staining demonstrating remaining α-gal antigen in porcine dermis extracellular matrix prior to α-gal reduction process; (b) immunohistochemical staining demonstrating significant reduction of α-gal antigen in ; and, (c) primate antibody response to GraftJacket and to before and after the proprietary process to remove α-gal. Matrix remodeling assessment: Collagenase is one of the enzymes secreted by cells to remove damaged collagen and to remodel tissues. The rate at which biological matrices are dissolved in collagenase solutions can be used to determine if the matrices have been altered by the manufacturing process. For example, matrices which have been exposed to harsh chemicals or high doses of ionizing radiation will be partially denatured. When exposed to collagenase damaged matrices will dissolve faster than normal dermis. On the other hand, matrices that have been cross-linked deliberately with chemicals or unintentionally by heat and/or dehydration are more resistant to collagenase. This inhibits the primary mechanism by which cellular and vascular repopulation occurs. Figure 8 shows the collagenase digestion times of, normal dermis, and several other matrices. Only and GraftJacket are digested at the same rate as normal dermis. (b) Primate Interposition Model Anti α-gal Antibody Graftjacket Acellular porcine dermis containing αα-gal antigen Time (days post-implant) 7

9 Figure 8: Collagenase digestion rates of and other matrices. Matrix damage assessment: Differential scanning calorimetry (DSC) is a method for detecting changes in biological matrices after processing. The instrument measures the temperature at which the collagen in the matrix denatures or melts and also measures how much collagen is being denatured at that temperature. Normal dermis,, and GraftJacket denature at approximately 60 C. Damaged or partially denatured matrices will melt at lower temperatures while cross-linked matrices will melt at higher temperatures. Figure 9 demonstrates the DSC curves of several commercial matrices in triplicates. Heat Flow OrthAdapt ZCR % Tissue Remaining Graft Jacket TissueMend Collamend OrthAdapt Fresh Graft Jacket Dermis TissueMend Time of Collagenase Digestion (Hrs) Temperature (ºC) Figure 9: DSC melting temperatures of and other matrices showing normal curves for GraftJacket and, cross-linking in OrthAdapt, a small amount of native collagen in cross-linked ZCR, and variable denaturation and cross-linking in TissueMend. Matrix biomechanical properties: Mechanical testing of matrices can also show the effects of different processing conditions. Suture retention strength is a critically important mechanical property of a biological matrix. Figure 10 shows the suture retention strength of and other matrices on the market. 11 These matrices differ in tissue source, thickness of the matrix (e.g. GraftJacket Maxforce (GJ MF) Extreme is significantly thicker than the other matrices shown) and cross-linking amongst many other variables. Cross-linking is often employed to strengthen a matrix, but in the case of suture retention strength cross-linking can make the matrix more brittle and actually reduce the strength. Figure 10: Suture retention strength of and other matrices Suture Retention (N) 0 GJ MF Extreme 200 Sterilization GJ MF Tissue- Mend GJ RM ZCR Restore Cuff- Patch Ortho- Adapt There are significant differences between aseptic processing and sterilization. There are also differences in the degree to which medical devices are sterilized. Aseptic processing is how many biological materials are processed (i.e. antibodies, biological tissues) since normal sterilization methods often significantly damage biologics. During aseptic processing, the material is procured and processed following specific special handling techniques to prevent introduction of additional bacteria. Allograft materials, such as GraftJacket, are required to screen and test donors and to follow regulations in accordance with the FDA s requirements for banked human tissue and Standards for Tissue Banking of the American Association of Tissue Banks (AATB). Additionally, states such as California, Florida, New York, Maryland, Illinois, and others have state specific guidelines for tissue banking. However, aseptic products cannot be labeled sterile. Sterilization on the other hand is treatment of a biologic or medical device to kill organisms remaining on the product. The degree of sterilization is a Sterility Assurance Level (SAL). The FDA considers a sterility assurance level (SAL) of 10-3 as an adequate sterility level for implantation of biological medical devices due to the risk of damage to biologics with traditional sterilization methods. This means there is a one in 1,000 probability that a viable microbe exists in or on the implantable device. 12 However, the FDA requires a SAL of 10-6 (a one in a million chance of a viable microbe) to be able to label the product as sterile. 13 Terminal sterilization means that after the final sterilization step the material is not touched anymore and the package remains sealed. is sterilized without damaging the matrix through a series of gentle proprietary processes. The processes used and the low dose irradiation are non-damaging to the matrix and the sterilization process has been validated to achieve an SAL of

10 Summary Other studies of biological matrices have demonstrated three distinct mechanisms of action, namely supporting regeneration, resorption/scarring, and encapsulation. This paper summarizes how matrix processing produces the mechanisms observed in vivo. Historically, tissue matrices have attempted to incorporate the ideal attributes of a biological matrix, but have fallen short, leading primarily to resorption/scarring or encapsulation mechanisms of action. While the GraftJacket tissue matrix demonstrated a mechanism of action that supports regeneration, it did not achieve the handling, storage, and sterility requirements ideal for clinical use., a next generation tissue matrix that supports regeneration, incorporates the ideal attributes through an extensive and iterative development and testing program. In summary, tissue matrix offers: An intact (non-damaged) matrix to support regeneration; A sterility assurance with an SAL of 10-6 ; A non-cross-linked matrix to allow rapid cell population and vascular in-growth; A product that can be stored at room temperature; A product that can be immediately used upon a two minute rinse; A product with significantly reduced levels of α-gal antigen responsible for xenogeneic response; A product with a stable, consistent well controlled, mechanically sufficient source tissue; A range of tailored sizes appropriate for orthopaedic soft tissue use; and A product with full mechanical properties right out of the package. References 1. Galatz, L.M., et al., The outcome and repair integrity of completely arthroscopically repaired large and massive rotator cuff tears. J Bone Joint Surg Am, A(2): p Gerber, C., B. Fuchs, and J. Hodler, The results of repair of massive tears of the rotator cuff. J Bone Joint Surg Am, (4): p Mellado, J.M., et al., Surgically repaired massive rotator cuff tears: MRI of tendon integrity, muscle fatty degeneration, and muscle atrophy correlated with intraoperative and clinical findings. AJR Am J Roentgenol, (5): p Chiodo, C.P. and M.G. Wilson, Current concepts review: acute ruptures of the achilles tendon. Foot Ankle Int, (4): p Iannotti, J.P., et al., Porcine small intestine submucosa augmentation of surgical repair of chronic two-tendon rotator cuff tears. A randomized, controlled trial. J Bone Joint Surg Am, (6): p Dejardin, L.M., et al., Tissue-engineered rotator cuff tendon using porcine small intestine submucosa. Histologic and mechanical evaluation in dogs. Am J Sports Med, (2): p Dejardin, L.M., S.P. Arnoczky, and R.B. Clarke, Use of small intestinal submucosal implants for regeneration of large fascial defects: an experimental study in dogs. J Biomed Mater Res, (2): p Sclamberg, S.G., et al., Six-month magnetic resonance imaging follow-up of large and massive rotator cuff repairs reinforced with porcine small intestinal submucosa. J Shoulder Elbow Surg, (5): p Sandor, M., et al., Host response to implanted porcinederived biologic materials in a primate model of abdominal wall repair. Tissue Eng Part A, (12): p Zheng, M.H., et al., Porcine small intestine submucosa (SIS) is not an acellular collagenous matrix and contains porcine DNA: possible implications in human implantation. J Biomed Mater Res B Appl Biomater, (1): p Barber, F.A., M.A. Herbert, and D.A. Coons, Tendon augmentation grafts: biomechanical failure loads and failure patterns. Arthroscopy, (5): p McAllister, D.R., et al., Allograft update: the current status of tissue regulation, procurement, processing, and sterilization. Am J Sports Med, (12): p Updated 510(k) Sterility Review Guidance K90-1: Final Guidance for Industry and FDA. 2002, Food and Drug Administration. 14. Xu H, Wan H, Zuo W, et al. A porcine-derived acellular dermal scaffold that supports soft tissue regeneration: removal of terminal galactose-alpha-(1,3)-galactose and retention of matrix structure. Tissue engineering. Part A. 2009;15(7): BAW-0739 Rev.D 9