Invited author

The role of immune cytokines in the pathogenesis of periapical lesions Stashenko P. The role of immune cytokines in the pathogenesis of periapical lesions. Endod Dent Traumatol 1990; 6: 89-96. Abstract - Bacterial infection of the dental pulp results in pulpal destruction, and subsequently stimulates an inflammatory cell response and destruction of bone in the periapex. Bacterial components, including lipopolysaccharides, induce the production of many polypeptide mediators, or cytokines, by inflammatory cells. These cytokines, which include macrophage-derived interleukin-1 beta, interleukin-1 alpha and tumor necrosis factor, and lymphocyte-derived lymphotoxin, have been show^n to potently stimulate bone resorption and to inhibit reparative bone formation in vitro and in vivo. This review presents the hypothesis that immune cytokines play a major role in the pathogenesis of periapical lesions.

Bacterial infection of the dental pulp commonly results in the formation of a dental periapical lesion with the concomitant resorption of bone. Bone resorption is also a prominent pathogenic feature of other chronic inflammatory diseases, including periodontitis, rheumatoid arthritis and osteomyelitis, as well as of certain malignances, notably multiple myeloma (1). Although bacteria are the etiologic agents of periapical lesion formation (2), and bone resorption is an active process carried out by osteoclasts (3), the intermediate pathway(s) linking the infection with osteoclastic resorption are poorly understood at present. Bacterial components, especially hpopolysaccharides (LPS), can themselves stimulate osteoclastic resorption, but are of relatively low potency (4, 5). Products of arachidonic acid metabolism, most notably prostaglandin E2, have also been implicated in the process of inflammatory resorption (6-9). Recently, mounting evidence has indicated that various mediators derived from immune cells, termed cytokines, may also be of importance. In this regard, the chronic inflammatory cells present in large numbers in periapical lesions produce several highly potent cytokines that cause bone resorption. These include macrophagederived interleukin-1 (IL-1) (10, 11), and tumor necrosis factor (TNFa) (12, 13), and the lymphocyte product lymphotoxin (LT, also termed TNF|3) U2, 13). This review discusses the immunology of periapical lesions, with special emphasis on the

Philip Stasiieni(o Department of Immunology Forsyth Dental Center, Boston, Massachusetts, USA

Key words: cytokines; bone; periapical; resorption. Dr. P, Stashenko, Department of Immunology, Forsyth Dental Center, 140 Fenway, Boston, MA 02115, USA. Accepted for publication January 22, 1990.

properties and mechanism of action of the boneresorptive cytokines.

Bacteria and their cemponents stimuiate periapicai iesien development The classic studies of Kakehashi et al. (2) elegantly demonstrate the causal relationship between bacterial infection and periapical lesion formation. In these studies, exposure of the pulp and infection from the oral cavity resulted in the development of periapical lesions in rats maintained in a conventional environment, whereas germ-free animals with pulp exposures failed to develop lesions. Moreover, germ-free animals formed large dentin bridges at exposure sites, indicating the potential for hard tissue regeneration in the absence of infection. The relationship between infection and periapical lesion development has been further illustrated by the work of Sundqvist (14), who found that, in traumatized, necrotic teeth with intact crowns, bacteria could be isolated only from teeth which had developed periapical lesions. Intact bacteria and soluble bacterial components, including cell wall structures, LPS and toxins, are highly antigenic, resulting in the stimulation of specific immune responses by the host (15). Speciflc responses are characterized by the activation only of the clones of T- and B-lymphocytes that bear

Stasheni(o complementary antigen-binding receptors for a foreign, or "non-self structures, in this case, for bacterial antigens. The antigen receptors on B-lymphocytes are membrane-bound versions of antibody molecules, whereas the T-lymphocyte receptor is a unique molecule that is evolutionarily related to, but distinct from antibody (16). Examples of speciflc responses include proliferation and mediator production by T-lymphocytes, and T-cell-regulated macrophage activation and antibody production by B-lymphocytes. In addition, nonspecific responses also undoubtedly contribute to periapical lesion pathogenesis. Nonspecific responses are stimulated via pathways that bypass antigen receptors in lymphocytes or do not involve lymphocytes at all. For example, LPS can activate B-lymphocytes polyclonally to proliferate and secrete antibody of diverse speciflcity (17). LPS is also a potent stimulator of macrophages, inducing them to produce several bone-resorptive mediators, including IL-1 and TNF (18). Polymorphonuclear leukocytes (PMNL) are attracted to sites of infection by a number of bacteria-derived chemoattractants, including the peptide F-MetLeu-Phe, which constitutes the amino terminus of many bacterial proteins. PMNL also undergo migration in response to the complement components C3b and C5a, which are generated following complement activation by antigen-antibody complexes. Antigen-antibody complexes placed within the root canal space have been shown to induce bone resorption, probably via a PMNL-mediated mechanism (19).

Histepathology of periapical lesions Few inflammatory cells are present in the normal dental pulp, with the exception of relatively small numbers of dendritic macrophages, which can function as antigen-presenting cells, and T-lymphocytes, which are probably recirculating rather than resident cells (20). At early stages foflowing pulpal infection, however, an acute, nonspeciflc inflammatory response dominated by PMNL and macrophages is observed (21). Subsequently, a specific antibacterial immune response is also generated, consisting of lymphocytes, macrophages, and plasma cells (22). The presence of both acute and chronic cellular components is typical of the mixed cell responses seen in many longstanding bacterial infections. If the infectious process overwhelms pulpal defenses and destroys pulp tissues, including its blood supply, inflammatory cells are subsequently unable to enter the necrotic root canal and eliminate the bacteria. A second line of local immune defense, i.e., the periapical lesion, consisting of an accumu-

lation of inflammatory cells, then occurs, which functions to limit the infection to the root canal system. It should be emphasized that although this response is usually referred to as a lesion, it actually represents a protective host cellular response. Most of our information about the histopathology of periapical lesions derives from analyses of longstanding, chronic human lesions of undetermined activity status. These studies have demonstrated that periapical lesions consist predominantly of granulomas and cysts, although cysts almost always are surrounded by granulomatous tissue (22). Periapical granulomas contain approximately equal numbers of inflammatory cells and various connective tissue elements (23). Conflicting results concerning the predominant inflammatory cell types present in granulomas have been reported. Nevertheless, it is clear that diverse inflammatory cell types including PMNL, T- and B-lymphocytes, macrophages, and plasma cells synthesizing all immunoglobulin classes are present (23-31). These cells can thus potentially mediate the entire spectrum of immunologic phenomena, including immediate (anaphylactic) hypersensitivity (IgE), delayed-type hypersensitivity, including mediator production (by T-cells and macrophages), and immune complex reactivity (IgM, IgG, PMNL). However, it is not yet known which of these mechanisms are operative in localizing bacteria to the root canal system, and in preventing direct invasion of bone with the development of osteomyelitis. Of the lymphocytes present, T-cells have generally been reported to be more numerous than Bcells (25, 27, 29, 30), indicating the presence of an ongoing, specific antibacterial reactivity! Analysis of T-cell subsets has revealed that the 2 types of T-lymphocytes, T-helper (TH) and T-suppressor (TS) cells, are represented in approximately equal number in chronic lesions (TH/TS ratio < 1.0) (31, 33-35). This compares with a ratio of about 2.0 in blood. Of interest, our own studies with actively developing lesions in rats indicate that TH cells outnumber TS cells during the phase of greatest lesion expansion, whereas TS cells predominate at later time periods when lesion size stabilizes (34). Taken together, these data suggest that TH cells may play a key role in periapical lesion development, whereas TS cells may dampen excessive immune reactivity, leading to cessation of lesion growth. In fact, TH cells possess several mechanisms by which they can mediate bone lysis. Besides stimulating B-cells to produce antibody (and ultimately the formation of antigen-antibody complexes), activated TH cells elaborate the resorptive cytokine lymphotoxin (LT) (36). In addition, they produce macrophage mi-

Resorptive cytoiiines and periapical lesions gration-inhibitory factor (MIF) and gamma-interferon (37), which together recruit and activate macrophages. Activated macrophages in turn elaborate the bone resorptive monokines IL-1 and TNFa, as well as PGE2 (see below). Bone-resorptive mediators Cytokines The bone resorption in inflammatory diseases and in certain malignancies, particularly malignant myeloma, had until recently been attributed to the action of a poorly characterized entity termed osteoclast-activating factor (OAF). OAF was originally described by Horton et al. (38) in 1972 as a boneresorptive lymphokine produced by human peripheral blood mononuclear cells activated by mitogens or dental plaque antigens. The major component of OAF was recently purified and characterized in our laboratories, and was found to be identical to the monokine interleukin lp (IL-IP) (11). Based on recovery during biochemical purification, 60% or more of total bone-resorbing activity can be attributed to IL-1 (3 (11, 39). The remainder of OAF activity is caused by the action of IL-1 a, TNFa, LT, and by synergistic interactions between these mediators (11, 13) (Table 1). In terms of stimulating resorption in vitro, IL-ip is the most active of these cytokines, with half-maximal activity at a concentration of4.5 x 10"'' M (< 1 ng/ml). ILlP is about 15-fold more potent than IL-1 a, and 1000-fold more potent than LT or TNFa (12). Recently, IL-ip, IL-1 a, and TNFa have also been shown to stimulate bone resorption in vivo (40, 41) (Fig. 1). The finding that these mediators exert similar effects in vivo and in vitro indicates their potential importance in the pathogenesis of osteolytic diseases. The genes for the a and P forms of IL-1 have been cloned and sequenced, showing that they are distinct but related molecules of nearly identical molecular weight (17,400 Da), but with only about 35% amino acid homology (42). Despite their different structures, IL-ip and IL-1 a bind to the same receptor (43). IL-ip is the major IL-1 species secreted by macrophages in humans, in about 5 times

Table 1, Immune cytokines with bone-resorbing activity. Predominant Cytokine cell source IL-1 a TNFa LT

macrophages macrophages macrophages lymphocytes

Molecular weight Relative potency (Da) in bone resorption^ 17,400 17,400

34,000 (dimer) 70,000 (trimer)

1.000 0.064 0.001 0.001

'Assessed in vitro using the fetal rat long bone assay (13).

Ref. 11,13,42 10,13,42 12,13,47,48

12,13,46

the quantity of IL-1 a (44). IL-1 a may function predominantly as a cell-bound rather than soluble mediator (44). TNFa and LT are both primarily mediators of cytotoxicity for transformed and malignant cells, and are produced by activated macrophages and lymphocytes respectively (36, 45). Native LT is a trimer of 70 kDa (46), and native TNFa a dimer of 34 kDa (47). Similar to the IL-1 species, TNFa and LT are approximately 30% homologous (48). Macrophages become activated to produce ILla, IL-ip and TNFa by phagocytosis of bacteria during invasion of tissues, or by stimulation with bacterial components, particularly LPS (18). Of interest, elevated IL-1 activity has been found in gingival crevicular fluid adjacent to sites of inflammation (49), as well as in rheumatic joints (50). Both IL-ip and TNFa, but not IL-1 a, have been localized by immunochemistry in inflamed periodontal tissues (51). In addition, certain specialized cells, including keratinocytes and gingival epithelial cells (52), haye been reported to produce IL-1. This finding may have relevance in infrabony cyst expansion, whose mechanism has received much speculation but about which we still have little definitive information (53). IL-1 exerts its effects on a wide spectrum of cell types. In addition to its bone-resorptive property, IL-1 is an immune stimulator of T-cells and B-cells (54) and mediates a multiplicity of biological effects that comprise the so-called acute phase response. These include the induction of fever (55), increased numbers of circulating neutrophils (56), synthesis of acute phase proteins (57), muscle catabolism (58), fibroblast proliferation (59), stimulation of collagenase production by fibroblasts and synovial cells (60), and prostaglandin production by many cell types, including flbroblasts and osteoblasts (61). Prostaglandins Products of arachidonic acid metabolism have long been implicated in the pathogenesis of inflammatory bone resorption. PGE2 resorbs bone in vitro (6), and has been found in elevated concentration in inflamed periodontal sites (7) and in crevicular fluid adjacent to these sites (8). The potential importance of PGE2 in periodontal disease is indicated by the finding that long-term administration of the cyclooxygenase inhibitor flurbiprofen inhibits naturally occurring periodontal destruction in the beagle dog (9). PGE2 is present in elevated concentration in pulps that are symptomatic compared with asymptomatic teeth (62). Paradoxically however, local infusions of PGE2 in vivo have actually been found to increase bone formation rather than bone resorption (63), and inhibitors of PGE production

II

Stasheni(o such as ibuprofen inhibit fracture repair (64). The net effect of PGE on bone mass therefore remains somewhat controversial. Regulatory interactions occur between PGE2 and cytokines, especially IL-1. IL-1 stimulates the production of PGE2 by various connective tissue cell types, including flbroblasts, chondrocytes, and synovial cefls (60, 61). PGE2 can inhibit T-cell proliferation and mediator production (such as LT), and has also been reported to inhibit IL-1 production by macrophages, suggesting a possible negative feedback pathway (65). These interactions are probably complex and their relative importance in bone destructive diseases in vivo is presently unknown. Bacterial components Bacterial components, in particular metabolic products such as short-chain fatty acids and LPS, may act to stimulate the bone destruction associated with periapical and periodontal diseases. LPS has been detected in infected root canals (66) and periapical lesions (67). However, it has not been established whether the levels of LPS present are capable of resorbing bone, since the limulus lysate test used to detect LPS is only semiquantitative and quite nonspeciflc (68). In general, LPS exerts half-maximal bone resorbing activity at concentrations of > 1 l^g/ml (4, 5), compared with cytokines such as IL1, which are equally active at < 1 ng/ml. Moreover, not all LPS have bone-resorbing activity. Fractionation studies of inflamed periodontal tissues have shown that few sites contained resorbing activity of > 100 kDa (LPS size range). In contrast, many sites had resorbing activity in the range of 10-100 kDa and < 10 kDa, consistent with the presence of cytokines and PGE2 respectively (5). Thus, although LPS and other bacterial components may act as activators of periapical lesion inflammatory cells, which in turn produce cytokines and prostaglandins, the role of LPS as a direct effector of osteolysis may be minor. It should be noted that, although cytokines are attractive candidates as stimulators of bone resorption in periapical lesion development, at present we have little if any conclusive information on this point. Gertainly all 3 groups of mediators - cytokines, prostaglandins, and bacterial components are often present together at sites of inflammatory resorption. This problem is further complicated by the fact that most inflammatory resorption, including periapical destruction, does not occur continuously, but rather is sporadic, with bursts of activity (69). The relative importance of these substances will be more definitively established when elevations in a given mediator can be correlated with periods of active bone destruction, for example, in a wefl92

defined animal model of periapical lesion development. Ceils involved in hone resorption Osteoclasts are multinucleated cells specialized for the resorption of bone (3). Osteoclasts possess a ruffled membrane that is apposed to the bone surface at areas of active resorption. The ruffled membrane is surrounded by a clear zone that seals ofl' the area of resorption, thus allowing local acidification and dissolution of mineral. Osteoclasts are also rich in proteolytic and other enzymes, notably tartrate-resistant acid phosphatase (TRAP), which is a widely used phenotypic marker of this cefl type (70). Osteoclasts thus have the capacity to remove both the mineral and proteinaceous components of bone. Osteoclasts are derived from stem cells present in the bone marrow (71). Following commitment to this differentiation pathway, osteoclast precursors first proliferate, and subsequently fuse to form typical multinucleated mature osteoclasts. The proliferation and fusion of the precursor cells is promoted by bone-resorptive mediators such as IL-1, TNFa, and LT and by the calcium-mobilizing parathyroid hormone (72). Osteoclast production is inhibited by calcitonin and by certain bone growth factors, including transforming growth factor P (3). There has been considerable controversy in the literature concerning the ability of macrophages to resorb bone and the relationship between osteoclasts and cells of the macrophage series. Osteoclasts and macrophage-monocytes do share a common stem cell precursor; however, at an early stage in development the 2 cell lineages diverge and it is generally accepted that they represent distinct, noncbnverting

Fig. 1. Osteoclastic bone resorption induced by IL-lp in vivo. Adult Sprague-Dawley rats were given daily injections of IL-1 P (1 Hg/kg) subcutaneously for 7 d. Paraffin sections were stained for the osteoclast marker tartrate-resistant acid phosphatase (TRAP). Arrows indicate stained cells. Magnification x 100.

Resorptive cytokines and periapicai iesions cell types (73). Like osteoclasts, macrophages can also fuse to form multinucleated giant cells. However, giant cells are antigenically distinct from osteoclasts, do not possess TRAP, and most importantly, are incapable of forming typical resorption lacunae on bone surfaces (74). The contention that macrophages and giant cells resorb bone is based primarily on the observations that these cells can phagocytize and degrade bone particles in vitro (75), and that they participate in removing dead bone (sequellum), such as occurs in dysbaric osteonecrosis (diver's disease) or advanced osteomyehtis (76). Unhke osteoclasts, macrophages do not have the capacity to resorb living bone or to initiate surface resorption. Although all resorption is an active cell process dependent on osteoclasts, studies with fractionated bone cells have revealed that the osteoblast, the bone-forming cell type, rather than the osteoclast, possesses receptors for resorptive hormones and cytokines (77-79). Chambers et al. (80, 81) have demonstrated that, in the functional absence of osteoblasts, osteoclasts are unable to respond to the resorptive mediators PTH, PGE, 1,25-DHCC, IL-1, TNFa or LT. The mechanism of interaction between osteoblasts and osteoclasts is not well understood, although several models have been proposed. One hypothesis suggests that, following stimulation with bone-resorptive mediators, osteoblasts that normally line bone surfaces move aside to expose bone to resorption by osteoclasts (82). Osteoblast-derived collagenase or plasminogen activator (83) may participate in this process by predisposing surfaces to osteoclastic attack. In the Chambers model, a lowmolecular-weight factor distinct from prostaglandins (Factor X) produced by stimulated osteoblasts is proposed to provide the final common pathway resorptive signal to osteoclasts (84). The nature of Factor X is unknown but is under active investigation.

resorption rate (88), an observation that has led to the use of PTH in the treatment of osteoporosis. Although the mechanism (s) of the coupling phenomenon are incompletely understood, the release of chemotactic and/or growth factors by resorbing osteoclasts from bone matrix may play a key role in the recruitment and stimulation of osteoblasts, which then repair the resorbed volume of bone. A number of bone growth factors have been implicated as coupling factors, including transforming growth factor P, insulin-like growth factor I, skeletal growth factor, platelet-derived growth factor, and others (89). In contrast to physiologic turnover, complete bone repair occurs infrequently following resorption at chronic inflammatory sites such as periapicai lesions or marginal periodontitis, at least in the presence of ongoing infection. Resorption without compensatory formation, or an uncoupled state, is also seen in certain malignancies such as myeloma (90). Recent results from this and other laboratories (12, 91, 92) have shown that cytokines, in addition to stimulating osteoblasts to provide a resorptive signal for osteoclasts, also inhibit osteoblastic bone formation. Bone formation is exquisitely sensitive to the presence of cytokines, since inhibition occurs at 50fold lower concentration than is required to stimulate resorption. For IL-ip this translates to inhi-

Bacterial Components Antigens

LPS

Cytokines as uncoupling factors Bone is not a static tissue, but undergoes continual turnover or remodeling. It has been estimated that 7% ofthe adult skeleton is resorbed and subsequently replaced by new bone formation each year (85). During this physiologic remodeling process, the amount of bone resorbed at each site is precisely and completely replaced within several months by new bone formation. This highly regulated process is referred to as the coupling of bone resorption and bone formation (86). Similarly, in metabolic diseases such as hyperparathyroidism (87), or with experimental infusion of PTH in animals (88), both a generalized resorptive response as well as compensatory bone formation are elicited. PTH may in fact stimulate a bone formation rate higher than the

Reparath/e Bone Formation

Factor 'X'I ^__JL._^^

'Coupling Factors' (TGFp. I G F - 1 , PDGF)

( Osteoclast

Bone resorption

Fig. 2. Hypothesized role of resorptive cytokines in periapicai lesion pathogenesis. || indicates inhibition of reparative bone formation by cytokines.

93

Stashenko bition exerted at 8xlO"'^M (0.01 ng/ml). Inhibition of formation involves down-regulation of bone matrix protein synthesis and alkaline phosphatase expression, without effects on osteoblast proliferation (91). Of interest, IL-1 (3 has recently been shown to be produced in large quantity by malignant myeloma cells, which may account for the uncoupling that occurs in this disease (93). Our present working hypothesis is that IL-ip and perhaps other cytokines are also responsible for uncoupling in periapicai lesions (Fig. 2). A possible protective role for inflammatory bone resorption Since bone resorption and inhibition of reparative bone formation are generally thought of as undesirable consequences of infection, the question logically arises as to why these mechanisms have evolved, and what survival value they confer on the organism. It might be speculated that resorption may permit bone to retreat from areas of active infection, thus preventing direct hard tissue invasion and the development of osteomyelitis, a serious and possibly lifethreatening condition. A concomitant result of resorption is that space is created for the infiltration of protective inflammatory cells, allowing formation of an immunologically active buffer zone that isolates the infection from the hard tissue. Whether these or other explanations eventually prove to be correct, it is clear, nevertheless, that the local regulation of bone mass by cytokines is in all likelihood an important element in the development of pulpal and periapicai pathology.

10. GowEN M, WOOD D D , EHRIE EJ, MCGUIRE M K B , RUSSELL RGG. An interleukin 1-like factor stimulates bone resorption in vitro. Nature 1983; 306: 378-80. 11. DEW^HIRST F E , STASHENKO P, MOLE JE, TSURUMACHI T. Purification and partial sequence of human osteoclast-activating factor: identity with interleukin 1(3. J Immunol 1985; 1362562-8. 12. BERTOLINI D R ,

BRINGMAN TS,

SMITH

13. STASHENKO P, DEWHIRST FE,

14.

15.

16.

17. 18.

19.

20.

21.

PEROS WJ,

KENT RL,

8. OFFENBACHER S, ODLE B M , VAN DYKE TE.

The

use

of

crevicular fluid prostaglandin E2 levels as a predictor of periodontal attachment loss. J Periodont Res 1986; 21: 101-12. 9. JEFFCOAT M K , WILLIAMS R C , WECHTER W J et al. Flurbiprofen treatment of periodontal disease in beagles. J Periodont Res 1986; 21: 624-33.

94

AGO

J M . Synergistic interactions between interleukin 1, tumor necrosis factor, and lymphotoxin in bone resorption. J Immunol 1987; 138: 1464-8. SuNDqviST G. Bacteriological studies ofnecrotic dental pulps (PhD thesis). Umea: Umea University Odontol Dissertation no. 7, 1976; 1-94. STASHENKO P. Regulatory effect of monocytes on T cell proliferative responses to oral microbial antigens. Infect Immun 1982; 38: 938-47. KRONENBERG M , SIU G , HOOD LE, SHASTRI N . The molecular genetics ofthe T-cell antigen receptor and T-cell antigen recognition. Ann Rev Immunol 1986; 4: 529-91. CouTiNHo A, MoLLER G. Thymus-independent B-cell induction and paralysis. Adv Immunol 1975; 21: 113-236. BuRGHETT SK, WEAVER W M , WESTALL J A , LARSEN A, KRONHEIM S, WILSON CB. Regulation of tumor necrosis factor/cachectin and IL-1 secretion in human mononuclear phagocytes. J Immunol 1988; 140: 3473-81. ToRABiNEjAD M, CLAGETT J, ENGEL D. A cat model for evaluation of mechanisms of bone resorption: induction of bone loss by simulated immune complexes and inhibition by indomethacin. Calcif Tissue Int 1979; 29: 207-14. JoNTELL M, GuNRAj MN, BERGENHOLTZ G . Immunocompetent cells in the normal dental pulp. J Dent Res 1987; 66: 1149-53. BERGENHOLTZ G . Effect of bacterial products on inflammatory reactions in the dental pulp. Scand J Dent Res 1977; 85: 122-9. LANGELAND K , BLOCK RM, GROSSMAN LL A histopathologic and histobacteriologic study of 35 periapical endodontic lesions in root-filled teeth. J Endod 1977; 3: 8-22.

23. STERN M H , DREIZEN S, MACKLER BF, SELBST AG,

1. MuNDY GR, MARTIN TJ. The hypercalcemia of malignancy: pathogenesis and treatment. Metabolism 1982; 31: 765-71. 2. KAKEHASHI S, STANLEY HR, FITZGERALD RJ. The effects of surgical exposures of dental pulps in germ-free and conventional laboratory rats. Oral Surg Oral Med Oral Pathol 1965; 20: 340-9. 3. VAES G . Cellular biology and biochemical mechanism of bone resorption. Clin Orthop 1988; 231: 239-71. 4. HAUSMANN E, WEINFELD N , MILLER WA. Effects of lipopolysaccharides on bone resorption in tissue culture. Cal Tissue Res 1972; 9: 272-82. 5. OFFENBACHER S . Mediators of bone resorption in periodontal disease (MS thesis). Boston, MA: Harvard School of Dental Medicine, 1981. 6. KLEIN DC, RAISZ L G . Prostaglandins: stimulation of bone resorption in tissue culture. Endocrinology 1970; 86: 1436-42. 7. GooDsoN M, DEWHIRST F E , BRUNETTI A. Prostaglandin E2 levels in human periodontal disease. Prostaglandins 1974; 6: 81-5.

DD,

MuNDY GR. Stimulation of bone resorption and inhibition of bone formation in vitro by human tumor necrosis factors. Nature 1986; 319: 516-8.

22.

References

NEDWIN GE,

LEVY

BM. Quantitative analysis of cellular composition of human periapicai granulomas. J Endod 1981; 7: 117-22. 24. STERN MH, DREIZEN S, MACKLER BF, LEVY BM. Isolation and characterization of inflammatory cells from the human periapicai granuloma. J Dent Res 1982; 61: 1408-12. 25. BERGENHOLTZ G , LEKHOLM U , LILJENBERG B , LINDHE J.

Morphometric analysis of chronic inflammatory periapicai lesions in root-filled teeth. Oral Surg Oral Med Oral Pathol 1983; 55: 295-301. 26. ToRABiNEjAD M, KETTERING J D . Identification and relative concentration of B and T lymphocytes in human chronic periapicai lesions. J Endod 1985; 11: 122-5. 27. PuLVER WH, TAUBMAN M A . SMITH D J . Immune components in human dental periapicai lesions. J Endod 1978; 23: 435-43. 28. MATTHEWS JB, MASON G I . Immunoglobulin producing cells in human periapicai granulomas. Br J Oral Surg 1983; 21:

192-7. 29. NiLSEN R, JoHANNEssEN AC, SKAUG N , MATRE R . In situ characterization of mononuclear cells in human dental periapicai inflammatory lesions using monoclonal antibodies. Oral Surg Oral Med Oral Pathol 1984; 58: 160-5. 30. Yu SM, STASHENKO P. Identification of inflammatory cells in developing rat periapicai lesions. J Endod 1987; 13: 53540.

Resorptive cytokines and periapicai iesions 31. BABAL P, SOLER P, BROZMAN M , JAKUBOVSKY J, BEYLY M , BASSET F. In situ characterization of cells in periapicai granu-

52.

lomas by monoclonal antibodies. Oral Surg Oral Med Oral Pathol 1989; 64: 48-52. 32. TORABINEJAD M, EBY W C , NAIDORF I J . Inflammatory and immunological aspects of the pathogenesis of human periapicai lesions. J Endod 1985; 11: 479-88.

53.

33. CYMERMAN JJ, CYMERMAN D H , WALTERS J, NEVINS AJ.

54.

34.

35.

36. 37.

Hu-

man T lymphocyte subpopulations in chronic periapicai lesions. J Endod 1984; 10: 9-11. STASHENKO P, YU SM. T helper and T suppressor cell reversal during the development of induced rat periapicai lesions. J Dent Res 1989; 68: 830-4. BARKHORDER RA, DESOUZA Y G . Human T lymphocyte subpopulations in periapicai lesions. Oral Surg Oral Med Oral Pathol 1988; 65: 763-6. LEOPARDI E , ROSENAU W. Production of a-lymphotoxin by human T cell subsets. Cell Immunol 1984; 83: 73-82. ScHULTZ RM, KLEINSCHMIDT WJ. Functional identity between murine gamma interferon and macrophage activating factor. Nature 1983; 305: 239-40.

38. HORTON JE, RAISZ LG, SIMMONS HA, OPPENHEIM JJ,

MER-

GENHAGEN SE. Bone resorbing activity in supernatant fluid from cultured human peripheral blood leukocytes. Science 1972; 177: 793-5. 39. LORENZO JA, SOUSA SL, ALANDER C , RAISZ LG, DINARELLO

CA. Comparison of the bone resorbing activity in the supernatants from phytohemagglutinin-stimulated human peripheral blood mononuclear cells with that of cytokines through use of an antiserum to interleukin 1. Endocrinology 1987; 121: 1164-70. 40. KoNiG A, MuHLBAUER RC, FLEISCH H . Tumor necrosis factor a and interleukin 1 stimulate bone resorption in vivo as measured by urinary (3H) tetracycline excretion from prelabeled mice. J Bone Min Res 1988; 6: 621-7.

55. 56.

57.

58. CLOWES G H A ,

42.

43.

44. 45.

46.

47. SHIRAI T, YAMAGUCHI H , ITO H , TODD C W , WALLACE R B .

Cloning and expression in Escherichia coli of the gene for human tumour necrosis factor. Nature 1985; 313: 803-6. 48. PENNICA D, NEDWIN GE, HAYFLICK JS et al. Human tumour necrosis factor: precursor structure, expression and homology to lymphotoxin. Nature 1984; 312: 724-9.

GEORGE BC,

VILLEE CA,

SARAVIS

CA.

Muscle proteolysis induced by a circulating peptide in patients with sepsis or trauma. N Engl J Med 1983; 308: 545-52. 59. SCHMIDT JA, MIZEL SB, COHEN D, GREEN I. Interleukin 1: a potential regulator of fibroblast proliferation. J Immunol 1982; 128: 2177-82. 60. MIZEL SB,

DAYER J M ,

KRANE SM,

MERGENHAGEN

SE.

Stimulation of rheumatoid synovial cell collagenase and prostaglandin production by partially purified lymphocyte activating factor (interleukin 1). Proc Nat Acad Sci USA 1981; 78: 2474-7. 61. TATAKIS DN, ScHNEEBERGER G, DziAK R. Recombinant interleukin-1 stimulates prostaglandin E2 production by osteoblastic cells: synergy with parathyroid hormone. Calcif Tissue Int 1988; 42: 358-62. 62. COHEN JS, READER A, FERTEL R , BECK M , MEYERS WJ.

41. SABATINI M , BOYCE B, AUFDEMORTE T, BONEWALD L , MUN-

DY G. Infusions of recombinant interleukins la and lP cause hypercalcemia in normal mice. Proc Natl Acad Sci USA 1988; 85: 5232-9. MARCH CJ, MOSLEY B, LARSEN A et al. Cloning, sequence and expression of two distinct human interleukin-1 complementary DNAs. Nature 1985; 315: 641-5. KiLiAN PL, KAFFKA K L , STERN AS et al. Interleukin 1 alpha and interleukin 1 beta bind to the same receptor on T cells. J Immunol 1986; 136: 4509-14. DINARELLO CA. Interleukin-1 and its biologically related cytokines. Adv Immunol 1989; 44: 153-205. MANNEL D N , MOORE RN, MERGENHAGEN SE. Macrophages as a source of tumoricidal activity (tumor necrotizing factor). Infect Immun 1980; 30: 523-30. GRAY P W , AGGARWAL BB, BENTON C V et al. Cloning and expression of cDNA for human lymphotoxin, a lymphokine with tumour necrosis activity. Nature 1984; 312: 721-4.

(Special Issue): Abstract #526. TA, ST ADLER BM, KATZ SI, OPPENHEIN JJ. Epidermal cell (keratinocyte) derived thymocyte activating factor {ETAY).J Immunol 1981; 127: 1493-8. MEGHJI S, HARVEY W, HARRIS M . Interleukin 1-like activity in cystic lesions of the jaw. Br J Oral Maxillofac Surg 1989; 27: 1-11. WOOD DD. Purification and properties of human B cell activating factor. I. Comparison ofthe plaque-stimulating activity with thymocyte-stimulating activity. J Immunol 1979; 123: 2395-401. DINARELLO CA, WOLFF SM. Molecular basis of fever in humans. Am J Med 1982; 72: 799-819. KAMPSCHMIDT R F , LONG RD, UPCHURCH H F . Neutrophil releasing activity in rats injected with endogenous pyrogen. Proc Soc Exp Biol Med 1972; 139: 1224-6. DINARELLO CA. Interleukin 1 and the pathogenesis of the acute phase response. N Engl J Med 1984; 311: 1413-8. LUGER

63. 64. 65.

66. 67.

68.

69.

70.

49. CHARON JA, LUGER T A , MERGENHAGEN SE, OPPENHEIM JJ.

A

radioimmunoassay determination of the concentrations of prostaglandins E2 and F2a in painful and asymptomatic human dental pulps. J Endod 1985; //.• 330-5. CHYUN Y S , RAISZ LG. Stimulation of bone formation by prostaglandin E2. Prostaglandins 1984; 27: 97-103. Ro S, SuDMANN E, MARTON P F . Effect of indomethacin on fracture healing in rats. Acta Orthop Scand 1976; 47: 588-99. KNUDSEN PJ, DINARELLO CA, STROM TB. Prostaglandins inhibit post-transcriptional interleukin 1 synthesis by increasing intracellular cyclic adenosine monophosphate. J Immunol 1986; 137: 3189-94. SCHEIN B, SCHILDER H . Endotoxin content in endodontically involved teeth. J Endod 1975; /; 19-21. ScHONFELD SE, GREENING AB, GLICK DH, FRANK AL, SIMON JH, HERLES SM. Endotoxic activity in periapicai lesions. Oral Surg Oral Med Oral Pathol 1982; 53: 82-7. ELIN R J , WOLFF SM. Nonspecificity of the limulus amebocyte lysate test: positive reactions with polynucleotides and proteins. J Infect Dis 1973; 128: 349-52. GooDSON JM, TANNER A C R , HAFFAJEE AD, SORNBERGER GC, SocRANSKY SS. Patterns of progression and regression of advanced periodontal disease. J Clin Periodontol 1982; 9: 472-81. VAES G . Lysosomes and the cellular physiology of bone resorption. In: DINGLE JT, FELL HB, eds. Lysosomes in biology and pathology, vol 1. Amsterdam: North-Hofland, 1969: 217. FiscHMAN DA, HAY ED. Origin of osteoclasts from mononuclear leukocytes in regenerating new limbs. Anat Rec 1962; 143: 329-38. TAKAHASHI N , MUNDY GR, ROODMAN G D . Recombinant human interferon-y inhibits formation of human osteoclastlike cells. J Immunol 1986; 137: 3544-9.

Increased thymocyte-activating factor in human gingival fluid during gingival inflammation. Infect Immun 1982; 38: 1190-4. 50. WOOD DD, IHRIE E , DINARELLO C , COHEN P. Isolation of an interleukin-1-like factor from human joint effusions. Arthr Rheum 1983; 26: 975-81.

71.

51. JANDINSKI JJ, RYNAR JE, STEINLE M , LEUNG CE, DEASEY MJ, STASHENKO PP. Localization of Interleukin la and tu-

73. BURGER EH, VAN DER MEER J W M , VAN DE GEVEL JS, GRIBNAU JC, THESINGH CW, VAN FURTH R . In vitro formation

mor necrosis factor a in human gingiva. J Dent Res 1989; 68

of osteoclasts from long-term cultures of bone marrow mono-

72.

95

Stasbenito nuclear phagocytes. J Exp Med 1982; 156: 1604-14. 74. CHAMBERS TJ, HORTON MA. Failure of cells ofthe mononuclear phagocyte series to resorb bone. Calcif Tissue Int 1984; 36: 556-8. 75. KAHN AJ, STEWART CC, TEITELBAUM SL. Contact-mediated bone resorption by human monocytes in vitro. Science 1978; 199: 988-90. 76. DAVIDSON J K . Dysbaric disorders: aseptic bone necrosis in tunnel workers and divers. J Clin Rheum 1989; 3: 1-23. 77. RoDAN GA, MARTIN TJ. Role of osteoblasts in hormonal control of bone resorption. A hypothesis. Calcif Tissue Int 1981; 33: 349-50. 78. ROULEAU MF, WARSHAWSKY H , GOLTZMAN D. Parathyroid hormone binding in vivo to renal, hepatic and skeletal tissues of the rat using a radioautographic approach. Endocrinology 1986; 118: 919-31. 79. NARBAITZ R , STUMPF WE,

SAR M , HUANG S, DELUCA HF.

Autoradiographic localization of target cells for la,25-dihydroxyvitamin D3 in bones from fetal rats. Calcif Tissue Int 1983; 35: 177-81. 80. THOMSON BM, SAKLATVALA J, CHAMBERS TJ. Osteoblasts mediate interleukin 1 stimulation of bone resorption to rat osteoclasts. J Exp Med 1986; 164: 104-12. 81. THOMSON BM, MUNDY GR, CHAMBERS TJ. Tumor necrosis factors a and P induce osteoblastic cells to stimulate osteoclastic bone resorption. J Immunol 1987; 138: 775-9. 82. CHAMBERS TJ, FULLER K . Bone cells predispose bone surfaces to resorption by exposure of mineral to osteoclastic contact. J Cell Sci 1985; 76: 155-65. 83. HAMILTON JA,

LINGELBACH S, PARTRIDGE N C , MARTIN

TJ.

Regulation of plasminogen activator production by boneresorbing hormones in normal and malignant osteoblasts. Endocrinology 1985; 116: 2186-92.

96

84. McsHEEHY PMJ, CHAMBERS TJ. Osteoblast-like cells in the presence of parathyroid hormone release a soluble factor that stimulates osteoclastic bone resorption. Endocrinology 1986; 119: 1654-9. 85. M U N D Y GR, ROODMAN GD. Osteoclast ontogeny and function. In: PECK W, ed. Bone and mineral research V. Elsevier: Amsterdam, 1987: 209-80. 86. HARRIS WH, HEANEY RR Skeletal renewal and metabolic bone disease. N Engl J Med 1969; 280: 253-5. 87. AiTKEN RE, KERR JL, LLOYD HM. Primary hyperparathyroidism with osteosclerosis and calcification in articular cartilage. Am J Med 1964; 37: 813-24. 88. WALKER R G . The induction of osteopetrotic changes in hypophysectomized, thyroparathyoidectomized, and intact rats of various ages. Endocrinology 1971; 89: 1389-1406. 89. MUNDY GR. Identifying mechanisms for increasing bone mass. J Natl Inst Health Res 1989; 1: 65-9. 90. STEWART AF, VIGNERY A, SILVERGLATE A, RAVIN ND,

Li-

voLSi V, BROADUS A . BARON R . Quantitative bone histomorphometry in humoral hypercalcemia of malignancy: uncoupling of bone cell activity. J Clin Endocrinol Metah 1982; 55: 219-27. 91. STASHENKO P, DEWHIRST FE, ROONEY ML,

DESJARDINS LA,

JD. Interleukin lp is a potent inhibitor of bone resorption in vitro. J Bone Min Res 1987; 2: 559-65. 92. STASHENKO P, OBERNESSER MS, DEWHIRST FE. Effect of immune cytokines on bone. Immunol Invest 1989; 18: 23949. HEELEY

93. KAWANO M , YAMAMOTO I, IWATO K , TANAKA H , AOSUKA

H. Interleukin 1 rather than lymphotoxin is the major bone resorbing activity in human multiple myeloma. Blood 1989; 73: 1646-9.