91731_ch03 12/8/06 7:33 PM Page 71 3 Repair, Regeneration, and Fibrosis Gregory C. Sephel Stephen C. Woodward The Basic Processes of Healing Regeneration Migration of Cells Stem cells Extracellular Matrix Cell Proliferation Remodeling Conditions That Modify Repair Cell Proliferation Local Factors Repair Repair Patterns Repair and Regeneration Suboptimal Wound Repair Wound Healing bservations regarding the repair of wounds (i.e., wound architecture are unaltered. Thus, wounds that do not heal may re- healing) date to physicians in ancient Egypt and battle flect excess proteinase activity, decreased matrix accumulation, Osurgeons in classic Greece. The liver’s ability to regenerate or altered matrix assembly. Conversely, fibrosis and scarring forms the basis of the Greek myth involving Prometheus. The may result from reduced proteinase activity or increased matrix clotting of blood to prevent exsanguination was recognized as accumulation. Whereas the formation of new collagen during the first necessary event in wound healing. At the time of the repair is required for increased strength of the healing site, American Civil War, the development of “laudable pus” in chronic fibrosis is a major component of diseases that involve wounds was thought to be necessary, and its emergence was not chronic injury. appreciated as a symptom of infection but considered a positive sign in the healing process. Later studies of wound infection led The Basic Processes of Healing to the discovery that inflammatory cells are primary actors in the repair process. Although scurvy (see Chapter 8) was described in Many of the basic cellular and molecular mechanisms necessary the 16th century by the British navy, it was not until the 20th for wound healing are found in other processes involving dynamic century that vitamin C (ascorbic acid) was found to be necessary tissue changes, such as development and tumor growth. Three for the function of prolyl hydroxylase, an enzyme required for key cellular mechanisms are necessary for wound healing: proper folding and stabilization of collagen into a triple helix. • Cellular migration The study of wound healing now encompasses a complex • Extracellular matrix organization and remodeling environment containing many cell types, matrix proteins, growth factors, and cytokines, which regulate and modulate the • Cell proliferation repair process. Nearly every stage in the repair process is redundantly controlled, and there is no single rate-limiting step, Migration of Cells Initiates Repair with the possible exception of the ingress of inflammatory cells. Successful healing maintains tissue function and repairs tissue barriers, Cells That Migrate to the Wound preventing blood loss and infection, but is usually accomplished Ingress of cells into a wound and activation of local cells are ini- through collagen deposition or scarring (fibrosis). Advances in our tiated by mediators that are either released de novo by resident understanding of growth factors, extracellular matrix and stem cells or from reserves stored in the granules of platelets and cell biology are improving healing, and offer the possibility of basophils. The contents of these granules include cytokines, regenerating tissues to their normal architecture and of chemoattractants, proteases, and mediators of inflammation, engineering replacement tissues. which together (1) control vascular delivery, (2) degrade dam- Successful repair relies upon a crucial balance between the aged tissue, and (3) initiate the repair cascade. Platelets are ac- yin of matrix deposition and the yang of matrix degradation. tivated when bound to collagen exposed at sites of endothelial Regenerative processes are favored when the matrix composition and damage, and their ensuing aggregation, in combination with 71 91731_ch03 12/8/06 7:33 PM Page 72 72 Pathology fibrin cross-linking, limits blood loss. Activated platelets release fingerlike membrane protrusions labeled filopodia. Cell polar- platelet-derived growth factor (PDGF) and other molecules ization and membrane extensions are initiated by growth factors that facilitate adhesion, coagulation, vasoconstriction, repair, or chemokines, which trigger a response by binding to their and clot resorption. Mast cells are bone marrow-derived cells specific receptors on the cell surface. Actin fibrils polymerize whose granules contain high concentrations of heparin. They and form a network at the membrane’s leading edge, thereby reside in connective tissue near small blood vessels and respond propelling lamellipodia and filopodia forward, with traction to foreign antigens by releasing the contents of their granules, provided via attachments to the extracellular matrix substrate. many of which are angiogenic. Resident macrophages, tissue- Actin-related proteins stimulate actin assembly, and numerous fixed mesenchymal cells, and epithelial cells release mediators actin-binding proteins act like molecular tinker toys, rapidly that not only contribute to the early response, but also perpet- constructing, stabilizing and destabilizing actin networks. uate it. Their numbers are increased through proliferation and The leading edge of the cell membrane impinges upon the extracel- recruitment to the site of injury (Fig. 3-1). The following are char- lular matrix and adheres to it through transmembrane adhesion acteristic of skin wounds: receptors termed integrins (see Chapter 2). These molecules are • Leukocytes arrive at the wound site early and migrate highly redundant, and many different integrin heterodimer rapidly by forming small focal adhesions (focal contacts). A combinations recognize the same matrix components (e.g., family of small peptide chemoattractants (chemokines), are collagen, laminin, fibronectin). Focal contacts develop through capable of restricted or broad recruitment of particular the adherence of the integrin extracellular domain to the leukocytes (see Chapter 2). connective tissue matrix. Focal adhesions form under the cell body, whereas smaller focal contacts form at the leading edge of • Polymorphonuclear leukocytes are rapidly recruited from migrating cells. The focal contact anchors the actin stress fibers, the bone marrow and invade the wound site within the first against which myosins pull to extend or contract the cell body. day. They degrade and destroy nonviable tissue by releasing As the cell moves forward, older adhesions at the rear are their granular contents. weakened or destabilized, allowing the trailing edge to retract. • Macrophages arrive shortly after neutrophils but persist for More than 50 proteins have been associated with the forma- days or longer. They phagocytose debris and orchestrate the tion of adhesion plaques. The cytoplasmic domain of integrins is developing granulation tissue by the release of cytokines and the foundation of a protein cascade that acts to anchor actin stress chemoattractants. fibers. The Rho-family of GTPases (Rho, Rac, and Cdc42) act as • Fibroblasts, myofibroblasts, pericytes, and smooth muscle molecular switches that interact with surface receptors to regulate cells are recruited by growth factors and matrix degradation matrix assembly, generate focal adhesions, and organize the actin products, arriving in a skin wound by day 3 or 4. These cells are cytoskeleton. responsible for fibroplasia, synthesis of connective tissue Importantly, the integrins transmit intracellular signals to cells matrix, tissue remodeling, wound contraction, and wound that also regulate cellular survival, proliferation, and differentiation. strength. These functional properties are also affected by other cell • Endothelial cells form nascent capillaries by responding to activators (e.g., cytokines) by signaling through cytoplasmic tails growth factors and are visible in a skin wound beyond day 3. of integrins from inside the cell to the outside matrix. In this The development of capillaries is necessary for the exchange manner, cytokines can also influence the organization and of gases, the delivery of nutrients, and the influx of inflam- tension in matrix and tissue. Thus, growth factors and integrins matory cells. share several common signaling pathways, but integrins are unique in their ability to organize and anchor the cytoskeleton. • Epithelial cells in the epidermis move across the surface of a Cytoskeletal connections are involved in cell-cell and cell-matrix skin wound, penetrate the provisional matrix (see below), connections and determine the shape and differentiation of ep- and migrate upon stromal collagen, which is coated with ithelial, endothelial, and other cells. plasma glycoproteins, fibrinogen, and fibronectin. The pro- cess of reepithelialization is delayed if the migrating epithelial cells must reconstitute a damaged basement membrane. In Extracellular Matrix Sustains the Repair Process addition, the phenotype of the epithelial layer is altered in the Three types of extracellular matrix contribute to the organization, absence of basement membrane. physical properties, and function of tissue: • Stem cells from bone marrow, the bulb of the hair follicle, • Basement membrane and the basal epidermal layer provide a renewable source of • Provisional matrix epidermal and dermal cells capable of differentiation, prolif- eration, and migration. Under appropriate conditions, these • Connective tissue (interstitial matrix or stroma) cells form new blood vessels and new epithelium and regen- erate skin structures, such as hair follicles and sebaceous Basement Membranes glands.