Inflammation and Growth Factors

0~%2-5347/97~1.571-0303$0LI.00/0 TIIE JOI'HSAI. OF lJw1.oi;y Copyright 0 1997 by AAIMEHICAN LJHCILCMX.AL A.SSO(:IATION, IN^.

Vol 157. 303-905.January 1997 PnntPd i n l!.S.A.

INFLAMMATION AND GROWTH FACTORS SHARON M. WAHL From the C'ellitlur. 1tirriruriolog.y Section, National Institute of Dental Research, Nutioncil Institutes of Health, Bethesdn, Maryland

ABSTRACT

Purpose: Pathways of inflammation culminating in tissue repair and scarring are defined. Materials and Methods: Inflammatory responses were induced in vitro a n d in vivo to monitor t h e mechanisms of leukocyte activation, t h e inflammatory products generated, t h e resolution of t h e response and tissue repair. Results: Recruitment and activation of circulating leukocytes at sites of t r a u m a a n d inflammation are essential to debridement, clearance of infectious organisms and repair of tissue injury. Once activated, infiltrating leukocytes generate a plethora of inflammatory mediators important in host defense. However, failure to clear t h e original stimulus and resolve t h e inflammatory response results i n persistent release of these mediators, including cytokines, enzymes, eicosanoids, and reactive nitrogen and oxygen intermediates, which can cause tissue damage. Overabundant matrix generation during t h e chronic repair process may cause fibroobstructive pathology. Conclusions: Disruption of these pathways, including leukocyte recruitment, may have therapeutic efficacy. KEY WORDS:inflammation, growth substances, penile induration, fibrosis In adults most inflammatory responses resolve with the development of scar tissue.' However, in some tissues the deposition of matrix components may impair tissue function. Peyronie's disease represents an example of impaired function associated with scar tissue that develops in the tunica albuginea.2 Peyronie's disease may occur in response to acute or repetitive trauma,3 in which microvascular injury and fibrin deposition lead to the recruitment and activation of inflammatory cells. The potential sequence of events in Peyronie's disease is acute or repetitive trauma, microvascular injury, fibrin deposition, inflammatory cell infiltration, chronic inflammation, fibroblast activation and proliferation, and pathological fibrosis (plaque). The inflammatory cells regulate fibroblast activation and proliferation, with the subsequent deposition of matrix leading to the development of fibrotic plaques.

wall through adhesion receptor interactions. Adherence of the white blood cells to the vessel wall precedes the extravasation of these cells into the tissues where they participate in the inflammatory response. The recruitment of leukocytes a t sites of inflammation involves a series of events initiated by a rolling phenomenon in which leukocyte selectins, carbohydrate-binding lectin receptors, interact with low af€inity ligands on the endothelium to slow the movement of the white blood cells.7~8Exposure to chemoattractants or cytokines induces up regulation andor activation of adhesion molecules on the leukocytes and endothelial cells to promote a firmer attachment. As these cells interact the leukocytes commence migration between the endothelial cells towards the site of tissue inflammation. Much of this interaction is mediated by the cellular adhesion molecules, selectins, members of the immunoglobulin (Ig) superfamily and integrins.9-11 An expanding family of integrins, each composed of 1 a and 1 /? subunit, includes the INFLAMMATORY RESPONSE P2 heterodimers primarily involved in cell-to-cell interacInflammation is a complex tissue response to injury in tions and the Pl integrins that promote cell-to-matrix andor which a network of chemical signals initiates and maintains cell-to-cell interactions. The a5pl receptor is specific for the the host response by stimulating endothelial cells, inducing extracellular matrix molecule, fibronectin, and interacts with leukocyte recruitment and activating phagocytic leukocyte its arginyl-glycyl-aspartic acid domain.9.1" Other Pl intefunctions.1.4 The series of events that occurs subsequent to grins may interact with arginyl-glycyl-aspartic acid or with trauma or antigen deposition are similar, being relatively different domains in the extracellular matrix substrates in independent of the causative agent. Persistent injury or in- addition to vascular ligands. Interaction of leukocytes with matrix proteins or endotheflammatory stimulation may result in pathological inflammation and fibrotic repair as occurs in Peyronie's d i ~ e a s e . ~ lium is regulated by inflammatory molecules.11 For example, Initially, platelets aggregate and release inflammatory me- exposure of peripheral blood monocytes to TGF-P induced diators, including transforming growth factor+ (TGF-P), increased adhesion of the monocytes to laminin, type IV that have a central role in the inflammatory process. The role collagen and fibronectin.12 The increased adhesion was assoof TGF-P in leukocyte recruitment and activation is that it ciated with synthesis of new receptors, and messenger riboincreases integrin expression and adhesion, is chemotactic, nucleic acid (mRNA)isolated from TGF-/?-treatedmonocytes induces type IV collagenase, auto-induces TGF-/? and acti- contained augmented levels of a5 and Pl subunit mRNA. The vates growth factor/cytokine prod~ction.5.~ The platelet prod- monocytes exhibited a TGF-/3 dose and time dependent inucts, in concert with additional inflammatory mediators, in- crease in expression of mRNA for the fibronectin receptor teract with vascular leukocytes and endothelial cells to bring subunits compared to control populations. Whereas inabout slowing and adhesion of leukocytes at the inflamma- creased a5 mRNA levels were evident within 90 minutes tory site. Signaling molecules, including cytokines and after exposure to TGF-P, the P l integrin subunit was differgrowth factors, mediate localized adherence to the vascular entially regulated. Constitutive expression of P1 mRNA was 303

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INFLAMMATION AND GROWTH FACTORS

not elevated at 90 minutes but it was substantially increased within 12 hours.12 The P l subunit may not require such rapid synthesis following activation because it is constitutively expressed. Following adhesion, the blood cells emigrate into the tissues aided by secretion of tissue degrading enzymes such as collagenase.12 Migration from the vessels at the site of adherence then proceeds by the process of chemotaxis in response to chemotactically active secreted factors. Among the factors responsible for inducing directed migration are complement fragments (C5a), products of the coagulation and kinin systems, platelet products and cytokines, including the chemokines and TGF-P.6 If an infection occurs bacteria release formyl-methionyl peptides, which are potent chemotactic factors.13 Products of inflammatory cells, released once the cells are activated at the site of inflammation, also trigger recruitment of additional leukocytes by a paracrine pathway.I4 Responding to chemotactic signals, the leukocytes accumulate at lesion sites where they are subjected to a barrage of inflammatory mediators.1.4 Cytokines and growth factors bind to their corresponding receptors on neutrophils, lymphocytes and monocytes, to initiate signal transduction with enhanced biochemical, endocytic and synthetic functions. In the absence of activating factors monocytes undergo prol6 grammed cell death (apoptosis) and are rapidly ~leared.1~. However, activated macrophages release a network of biologically active molecules, including prostaglandins, reactive oxygen metabolites and nitrogen intermediates, enzymes and A cytokines, which orchestrate an inflammatory re~ponse.1~ number of these secreted products, such as reactive oxygen metabolites and hydrogen peroxide, may also contribute to tissue damage's as do nitrogen intermediates.'* Central in this process is the generation of cytokines and growth factors that are also preparatory for the next stage of the inflammatory response, the healing process.5.20.21 Whereas interleukin-1, tumor necrosis factor-a, platelet derived growth factor and TGF-p promote the inflammatory response, these same cytokines contribute to tissue repair as the inciting agent or injury is resolved. Tissue repair is dependent on the accumulation and activation of fibroblasts, which are responsible for the deposition of extracellular matrix molecules and the generation of scar tissue.14 Cytokines may directly or indirectly influence fibroblast recruitment and proliferation by stimulating the fibroblasts to produce platelet derived growth factor, which then acts in an autocrine or paracrine fashion to regulate cell growth. TGF-6 indirectly regulates fibroblast proliferation but directly stimulates matrix production.6 Not only is collagen synthesis up regulated by TGF-6, but fibronectin and other matrix molecules are also increased in vitr0.5 After direct injection of TGF-0 into a tissue there is a n increased inflammatory cell recruitment, followed by production of an extracellular matrk6.12In addition, TGF-p regulates matrix accumulation by inhibiting the production of metalloproteinases and increasing the production of protease inhibitors.6

Under normal circumstances, once the inflammatory response begins to subside, cytokine regulation of fibroblast activation and proliferation progresses to tissue repair, which occurs with minimal scarring. If, however, antigens persist or repetitive injuries occur,prolonged inflammation becomes a chronic inflammatory lesion with the potential for substantial tissue damage. As the host continues its attempts at tissue repair, overzealous repair and scarring may evolve into fibro-obstructive pathology. Consequently, considerable effort ha8 been expended in identifying cellular or molecular targets for interrupting and regulating these events.

INTERRUPTING THE INFLAMMATORY RESPONSE

One approach to interrupting chronic inflammation to inhibit the accompanying pathology has focused on blocking leukocyte recruitment to sites of inflammation. Inhibitors of leukocyte adhesion as well as cytokine antagonists could potentially minimize damaging inflammatory consequences. In recent studies synthetic fibronectin peptides have been identified that have the capability of blocking inflammatory cell adhesion.22 When arginyl-glycyl-aspartic acid containing peptides coupled to a protein carrier were injected into an experimental animal in which synovial inflammation was induced by bacterial cell walls, the peptides inhibited the development of arthritis. Treatment of arthritic animals with the protein camer alone had no significant effect, whereas the carrier conjugated peptide effectively suppressed acute synovitis. Histological analysis revealed inhibition of inflammatory cell recruitment into the s y n o v i ~ m .An even more dramatic suppression of the chronic inflammatory lesions was apparent, and the arginyl-glycyl-aspartic acid peptide maintained suppression of arthritis even a h r treatment wag discontinued.22 Studies are in progress to characterize further this powerful anti-inflammatory action and by pursuing such an approach, it may be possible to manipulate selectively inflammatory processes, with the intention of minimizing fibrotic pathology. CONCLUSION

I n summary, as we learn more about normal and pathological responses to tissue injury, new concepts of therapy may emerge. Since leukocyte adhesion is a mandatory requirement in the evolution of an inflammatory response, interfering with adhesion can interrupt the recruitment of inflammatory cells and the subsequent cell-mediated events. Peptide antagonists represent but 1new mechanism for modulating the course of persistent inflammation. By disrupting the chronic recruitment of inflammatory cells, tissue degradation and fibrotic repair may be minimized. If Peyronie's disease is associated with inflammation dependent pathology, this andor other anti-inflammatory approaches may be beneficial. REFERENCES

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