Medial Collateral Ligament Healing in Macrophage Metalloelastase (MMP-12)-Deficient Mice Rick W. Wright, Tracy Allen, Hossam B. El-Zawawy, Michael D. Brodt, Matthew J. Silva, Corey S. Gill, Linda J. Sandell Department of Orthopaedic Surgery, Washington University School of Medicine at Barnes-Jewish Hospital, St. Louis, Missouri Received 17 October 2005; accepted 2 March 2006 Published online 31 August 2006 in Wiley InterScience (www.interscience.wiley.com). DOI 10.1002/jor.20222 ABSTRACT: Medial collateral ligament (MCL) injuries heal by a wound repair scar response controlled by a complex cellular and cytokine environment. Many enzymes participate in wound repair, particularly the matrix metalloproteinases. We hypothesize macrophage metalloelastase (MME/MMP-12) deficiency results in impaired healing of MCL injury. One hundred fifty MME- deficient and 150 WT (MMEþ/þ) mice underwent knee MCL transection with the opposite knee as a sham operated control. Mice were sacrificed at 3, 7, 28, 42, and 56 days. At each of the five time points, 15 mice were utilized for biological and 15 were utilized for biomechanical testing. Outcome measures were the presence of macrophages to represent the inflammatory phase of wound healing, collagen synthesis to assay for matrix repair, and biomechanical testing for repair strength. Immunohistochemistry demonstrated significantly fewer macrophages in cut MCLs from MME- deficient mice versus wild-type (WT) mice at 3, 7, 28, and 42 days (all p  0.04). In situ hybridization to Col1a1 mRNA in the MME-deficient cut MCLs at 7, 28, and 42 day time points showed a decreased level of type I pro-collagen mRNA compared to the WT cut MCLs ( p < 0.05). Biomechanical testing revealed cut ligaments from MME-deficient mice had significantly lower ultimate force and stiffness compared to cut ligaments from WT mice ( p < 0.001), with maximal differences of 40% at 7 days for ultimate force and 28 days for stiffness ( p < 0.05 by Tukey post hoc test). We conclude MME is important in the multifactorial cascade of knee MCL injury healing, showing significant differences in both the early inflammatory and in the matrix tissue synthesis phases. ß 2006 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 24:2106–2113, 2006 Keywords: mouse; medial collateral ligament; knee; macrophage metalloelastase INTRODUCTION Ligamentous injuries account for a significant number of musculoskeletal disorders treated in the United States, where it has been estimated there are greater than 20 million reported injuries involving joints and ligaments each year. 1 A large number of these injuries involve the knee medial collateral ligament (MCL). Currently, due to a lack of understanding of the molecular events surrounding the healing process, we are limited in our ability to improve the patient’s short- and long-term functional outcome after an MCL injury. Previous studies in animal models have concentrated on the biological response and bio- mechanical function with respect to healing. Healing as assessed by ultimate load to failure typically plateaus at 70% compared to the normal MCL. 2–8 Investigators have tried to manipulate the healing process by addition of a variety of growth factors at the healing site. 9–11 This approach to healing has shown some biological effects, however, biomechanical function has not dramatically improved or accelerated. Therefore, further information on the role of macrophage metalloelastase (MME) in the early cascade of ligament repair of the knee MCL is needed. The predominant extracellular matrix compo- nents of the knee MCL are collagen, elastin, fibronectin, glycoproteins, and proteoglycans. Collagen is the major structural macromolecule, representing 75% to 80% of the dry weight. Microscopically, the collagen fibers are organized in parallel arrays. Type I collagen is the predomi- nant genetic type of collagen found in normal ligaments in a ratio of 3:1 with type III collagen. In healing ligaments, type I and type III collagens are produced in a 1:1 ratio. 12 Elastin, an insoluble protein found distributed with collagen in ligament tissue, makes up 5% to 7% of the dry weight in normal ligamentous tissue. Elastin provides the elasticity or recoil capabilities found throughout the ligament. Soft-tissue wound healing is known to occur via an organized, temporal sequence of events consist- ing of four overlapping phases: hemorrhage (initial 2106 JOURNAL OF ORTHOPAEDIC RESEARCH NOVEMBER 2006 Correspondence to: Rick W. Wright (Telephone: 314-747- 2813; Fax: 314-747-2599; E-mail: RWWright1@aol.com) ß 2006 Orthopaedic Research Society. Published by Wiley Periodicals, Inc.