Effects of Immobilization on the Extensibility of Periarticular Connective

01 96-601 1 /81/0302-0067$02.00/0 THE JOURNAL OF ORTHOPAEDIC AND SPORTS PHYSICAL THERAPY Copyright 0 1981 by The Orthopaedic and Sports Physical Therapy Sections of the American Physical Therapy Association Effects of Immobilization on the Extensibility of Periarticular Connective - Tissue ROBERT DONATELLI,* MS, PT, HELEN OWENS-BURKHART,t MS, PT Copyright 1981. All rights reserved. Connective tissue, one of the basic components of the human body, is a focus of many treatment procedures in orthopaedic physical therapy. As physical therapists, it is important for us to understand the nature of connective tissue and the histological changes that occur when connective tissue is immobilized. These changes are directly related to the causes of joint stiffness, leading to restricted movement. A basic knowledge of connective tissue is necessary to effectively restore normal joint mobility. In addition, this basic understanding provides a rationale for the treatment programs and assists in the development of realistic goals for our patients. This article discusses the biochemical and histological changes of periarticular connective tissue resulting from immobilization of synovial joints. Conclusions derived are confined to hypotheses only, due to lack of applied research in this area. CLASSIFICATION Muscle, nerve, epithelial, and connective tissues constitute the four basic tissues of the human body." The most abundant of these is connective tissue, which as the name implies serves to connect structures, such as ligaments and joint capsules." The general heading of connective tissue includes five main groups, each with a specific function based largely on the composition and orientation of intercellular substance^.'^ The groups are: ordinary connective tissue, including both dense and loose types; adipose tissue; blood cells and blood cell-form- * Codirector of Orthopaedic Physical Therapy. Clayton General Hospital, and Assistant Professor, lnstitute of Graduate Health Sciences. Atlanta. GA. t Ms. Owens-Burkhart completed this paper while finishing her Master's degree in Orthopaedic Physical Therapy at The Institute of Graduate Health Sciences. She is currently at Palos Heights Therapy Center. Palos Heights, IL 60463. 67 ing tissues; cartilage; and bone." The emphasis of this paper is the effects of immobilization on dense ordinary periarticular connective tissue structures, which include ligaments, tendons, synovial membrane, fascia, and fibrous joint capsules. BIOCHEMISTRY AND HISTOLOGY An understanding of connective tissue ultrastructure is essential to appreciate periarticular connective tissue responses to immobilization. The two main components of connective tissue are the cells and the extracellular matrix.'' The matrix consists primarily of collagen and elastin fibers and a nonfibrous ground substance, both products of the secretory fibroblastic cell of connective tissue.''^ I 3 Cells in dense connective tissue are mainly fibrocytes located between the collagen fibers. It is within the cell that procollagen, the precursor to collagen, is formed." An initial step in procollagen formation is the assembly of amino acids to form a chains (polypeptide chains). Each a chain consists of repetitive sequences of amino acids along its course. The first and second amino acids may be any of the numerous amino acids such as proline, lysine, or glycine. However, the third is always glycine." Collagen is unique because a large amount of the proline and lysine is hydroxylated by the addition of an OH group. The function of hydroxyproline is not known. However, the hydroxylysine accounts for the cross-links of adjacent collagen molecules which give collagen fibers its mechanical strength.'*, I4 The three a chains are united by intramolecular hydrogen bonds to form a helix which is transported outside the cell as a tropocollagen molecule. Outside the cell, the tropocollagen molecules align via ionic and hydrophobic interactions to form collagen fibrils. Further intermolecular bonding or cross-links between the fibrils

68 DONATELLI AND OWENS-BURKHART JOSPT Vol. 3, NO. 2 Copyright 1981. All rights reserved. occurs creating the extracellular collagen fiber.5. " Intra- and intermolecular bonds are seen in Figure 1. As collagen fibers mature, intra- and intermolecular bonds or cross-links increase in number, thereby providing tensile strength to the fibers.'. 93 l5 Three major cross-links reported in human collagen are dihydroxylysinonorleucine, hydroxylysinonorleucine, and histidonohydroxymerodesmo~ine.'~ Based on the arrangement of its collagen fibers, dense connective tissue is commonly classified into two types, irregular and regular." The irregular type has collagen fibers running in different directions in the same plane. This is quite functional for capsules, aponeuroses, and sheaths which physiologically are stressed in many directions." In the regularly arranged tissue, the fibers run more or less in the same plane and same linear direction. This arrangement affords great tensile strength to ligaments and tendons, which physiologically receive primarily unidirectional stress. The nonfibrous portion of the connective tis- sue matrix, often referred to as the ground substance, is composed of acid mucopolysaccharides or glycoaminoglycans (GAGs) and water.lo." It is important to be familiar with GAG and its function to understand the changes that occur with immobilization and its effect on connective tissue extensibility. The four major GAGs found in connective tissue are hyaluronic acid, chondroitin-4-sulfate, chondroitin-6-sulfate, and dermatan sulfate. Generally, GAGs are bound to a protein and collectively referred to as a proteoglycan. In connective tissue, proteoglycans combine with water to form a proteoglycan aggregate." Water makes up 60-70% of the total connective tissue content. GAG has an enormous waterbinding capacity and is responsible for this large water ~ontent.'~~ " Together the GAG and water form a semifluid viscous gel within which the collagen and fibrocytes are embedded. Specifically, hyaluronic acid with water is thought to serve as a lubricant between the collagen fibers.4. I I. 18 The lubricant maintains a distance between the fibers, thereby permitting free glid- Fig. 1. Intra- and intermolecular cross-links.

JOSPT Fall 1 98 1 CONNECTIVE TISSUE IMMOBILIZATION 69 ing of the fibers past each other and perhaps preventing excessive cross-linking. Free gliding is essential for normal connective tissue mobilit^.^ This fiber orientation and mobility can best be exemplified by the fiber weave in the Chinese finger trap seen in Figure 2A. When placed under tension, as during stretching, the small cables move on one another, causing the diameter of the trap to be reduced as the system lengthens (Fig. 2B). In summary, fibrocytes produce the components of connective tissue extracellular matrix. The nonfibrous ground substance comprised of GAG and water along with collagen fibers inter- Copyright 1981. All rights reserved. Fig. 2. A, before tension is applied to the cables (Chinese finger trap); B, when placed under tension, the small cables move on one another causing the diameter of the trap to be reduced as the system lengthens4

70 DONATELLI AND OWENS-BURKHART JOSPT Vol. 3, No. 2 Copyright 1981. All rights reserved. Fig. 3. Model of collagen cross-link at the molecular level. A and 6, preexisting fibers; C, newly synthesized fibril; 0, cross-link as the fibril joins the fiber; X, nodal point where the fibers normally slide past one another freely.4 acts to maintain the structural integrity and allow free gliding between fibers, enhancing the tissue extensibility. EFFECTS OF IMMOBILIZATION Knowledge of the effects of immobilization is largely derived from animal studies in controlled laboratory settings. This creates obvious limitations to the applicability of these studies to humans. The experiments involve the knees of primates, dogs, rats, and rabbits immobilized at various ranges with various internal fixation devices. Woo and Akeson immobilized the experimental animals for 9 weeks.'-4*" Upon gross inspection of the synovial joints postimmobilization, the authors noted excessive connective tissue deposition in the joint and joint recesses. With time, this excessive fatty fibrous connective tissue formed mature scar and created intraarticular adhesion^.^ Biochemical changes within the matrix of the periarticular structures result from lack of movement. Movement is essential for the prevention of contracture and adhesion formation within the joint.4 Movement is also necessary for proper orientation of collagen fibers as they develop.3s The matrix changes reported with immobilization are relatively uniform in ligament, capsule, tendon, and fascia. These changes involve extracellular water loss and GAG depletion, along with collagen cross-linking changes. With immobilization, there is a significant water loss. Akeson et al.' report a 4.4% loss of water following immobilization, with the tissue appearing "dry and gli~tening,"~ especially in the interfascia1 planes. The most significant change is a reduction in GAG content within the matrix. Akeson's studies', 3.4. 19 report a 40% decrease in hya- luronic acid and a 30% decrease in chondroitin- 4- and chondroitin-6-sulfate, with little or no change in total collagen content. Although the synthesis of new collagen during immobilization is reduced, there is a significant increase in the three major cross-links, especially dihydroxylysinon~rleucine.~ A reduction in the semifluid gel due to loss of

JOSPT Fall 198 1 CONNECTIVE TISSUE IMMOBILIZATION 71 Copyright 1981. All rights reserved. GAG and water may result in a reduction of the critical fiber distance between collagen fibers. Friction is created between fibers at the nodal points. However, development of cross-links at the nodal points is minimal because the distance between the fibers, although reduced, is still too large to permit cross-link formation. Instead, cross-link formation hampering collagen mobility occurs between the newly synthesized fibrils and preexisting collagen fibers (Fig. 3). Clinically, this restriction is perceived as a mechanical constraint to movement. Furthermore, when the joint is immobilized, the lack of movement perpetuates the random orientation of the newly synthesized collagen fibrils and facilitates the development of irregular cross-links in strategic regions of the collagen weave pattern (Fig. 4h4 Water and GAG losses are consistent with the affinity of GAG for water. The fact that the GAG was preferentially lost over collagen is explained by the rapid turnover rate of GAG (half life of 1.7-7 l7 as compared to collagen (half life of 300-500 days).i3 In summary, animal studies demonstrate that with immobilization of synovial joints biochemical and histological changes occur within the periarticular connective tissue structures. There is significant reduction in GAG content with subsequent water loss, contributing to abnormal crosslink formation and joint restriction. In addition, within the joint space and its recesses, there is excessive connective tissue deposition in the form of fatty fibrous infiltrate, which later matures to form scar tissue adhering to intraarticular surfaces further restricting motion. EFFECTS OF MOBILIZATION There is no documentation of the effects of mobilization, following immobilization, on crosslink formation. There is, however, documentation that movement maintains lubrication and critical fiber distance within the matrix and assures an orderly deposition of new collagen fibrils, thereby preventing abnormal cross-link formation 3. 4, 19 Enneking and Horowitz6 and Evans et al7 document that forceful manipulation breaks intracapsular fibrofatty adhesions that may have Fig. 4. Diagram showing the idealized weave pattern of collagen fibers. It can be demonstrated that the fixed contact at strategic sites (e.g., points d and e) can severely restrict the extension of this collagen weave.4 A, collagen fiber arrangement; 8, collagen fiber cross-links; C, normal stretch; D, restricted stretch due to cross-link.

72 DONATELLI AND OWENS-BURKHART JOSPT Vol. 3, NO. 2 Copyright 1981. All rights reserved. formed within the joint during immobilization. These authors speculate that manipulation may have caused some peeling of the fibrofatty tissue from the femoral condyles of the experimental animals. Although motion was restored following the manipulation, the ragged edges of adhesions remained in the joint. Research on movement and its effect on connective tissue will prove invaluable to the therapist. It will help to substantiate the role of mobilization in reversing biochemical and histological changes secondary to immobilization. If movement is the major stimulus for biological activity, then the amount, the duration, the frequency, the rate, and the time of initiation of the movement are all important in producing the desired therapeutic effects on connective tissue structures. These factors must be determined before we can comprehend the optimal benefits of mobilization. SUMMARY This paper has provided the reader with a basic review of the biochemical and histological effects of immobilization on periarticular connective tissue. Much of the mechanism underlying the contracture process remains speculative and many questions remain unanswered. Connective tissue research is a relatively new field with sparse scientific documentation performed with human subjects. Research directed towards the understanding of the effects of mobilization may provide the therapist with better direction in treating stiff restricted joints. The authors would like to thank Ms. Mary Beth Glasheen-Wray and Mr. Scot Irwin for their assistance during the preparation of this paper; Dr. Wayne Akeson for the kind permission to reproduce Figures 3 and 4 from his work; the 1981 class of the Institute of Graduate Health Sciences; and the initial motivation from Dr. John Merryman. REFERENCES 1. Akeson WH. Amiel D. LaViolette D: The connective tissue response to immobility: a study of the chondroitin 4- and 6-sulfate and dermatan sulfate changes in periarticular connective tissue of control and immobilized knees of dogs. Clin Orthop 51 :183-197. 1967 2. Akeson WH, Amiel D, LaViolette D, Secrist D: The connective tissue response to immobility: an accelerated aging response. Exp Gerontol 3:289-301. 1968 3. Akeson WH, Amiel D, Mechanic GL, Woo S, Harwood FL, Hamer ML: Collagen crosslinking alterations in joint contractures: changes in reducible crosslinks in periarticular connective tissue collagen after nine weeks of immobilization. Connect Tissue Res 5:5-19. 1977 4. Akeson WH, Amiel D, Woo S: Immobility effects of synovial joints: the pathomechanics of joint contracture. Biorheology 17:95-110, 1980 5. Bryant M: Wound healing. Clinical Symposia. Summit, NJ: Ciba Pharmaceutical Co, 1977 6. Enneking W, Horowitz M: The intra-articular effects of immobilization on the human knee. J Bone Joint Surg 54-A:973-985, 1972 7. Evans E, Eggers G, Butler J, Blumel J: Immobilization and remobilization of rats knee joints. J Bone Joint Surg 42-A:737-758, 1960 8. Freeman MAR: Adult Articular Cartilage, Ed 2. pp 183-1 96. Tunbridge Wells, England, Pitman Medical, 1979 9. Fujimoto D, Moriguchi T, lshida T, Hayashi H: The structure of pyridinoline, a collagen crosslink. Biochem Biophys Res Commun 84:52-57. 1978 10. Gray's Anatomy. Ed 35. Warwick R. Williams P (eds). Philadelphia: WB Saunders Co, 1974 11. Ham A, Cormack D: Histology. Ed 8. Philadelphia: JB Lippincott Co. 1979 12. LaVigne A, Watkins R: Preliminary Results on Immobilization: Induced Stiffness of Monkey Knee Joints and Posterior Capsule. Perspectives in Biomedical Engineering. Proceedings of a Symposium of Biological Engineering Society, University of Strathclyde. Glasgow, June 1972. Baltimore: University Park Press, 1973 13. Neuberger A. Slack H: The metabolism of collagen from liver, bones. skin, and tendon in normal rat. Biochem J 53:47-52, 1953 14. Nimni ME, Marcel E: The molecular organization of collagen and its role in determining the biophysical properties of the connective tissues. Biorheology 17:51-82, 1980 15. Peacock E. Van Winkle W: Wound Repair. Philadelphia: WB Saunders Co, 1976 16. Schiller S, Matthew M, Cifonelli J, Dorfman A: The metabolism of mucopolysaccharides in animals: further studies on skin utilizing Ci4 glucose, C14 acetate, and S35 sodium sulfate. J Biol Chem 21 8:139-145. 1956 17. Schiller S, Matthews M, Golfaber L, Ludoweig J. Dorfman A: The metabolism of mucopolysaccharides in animal: studies in skin utilizing labeled acetate. J Biol Chem 21 2:531-535. 1955 18. Swann D, Radin E, Nazimiec M: Role of hyaluronic acid in joint lubrication. Ann Rheum Dis 33:318-326, 1974 19. Woo S. Matthews JV, Akeson WH, Amiel D, Convery R: Connective tissue response to immobility: correlative study of biomechanical and biochemical measurements of normal and immobilized rabbit knees. Arthritis Rheum 18:257-264, 1975