b2MICROGLOBULIN AND AMYLOIDOSIS IN CHRONIC DIALYSIS PATIENTS

The Pathogenesis of b2-Microglobulin–Induced Bone Lesionsin Dialysis-Related Amyloidosis

Martin Tran,* Gregory W. Rutecki,*† and Stuart M. Sprague*†*Department of Medicine, Division of Nephrology, Evanston Northwestern Healthcare and †NorthwesternUniversity Medical School, Evanston, Illinois

ABSTRACT

Dialysis-related amyloidosis (DRA), also referred to as b2-microglobulin amyloidosis (Ab2M), is an important cause ofmorbidity in patients with chronic renal failure and in those whoare on dialysis. Although DRA deposits from affected jointshave been characterized as a unique amyloid fibril protein, b2M,less is known about the pathologic role of b2M as a mediator ofbone and joint disease. Potential mechanisms for b2M patholog-

ic interaction in bone include bone growth factors, cytokines,and advanced glycation end products (AGEs). It appears thatDRA is the result of a complex interaction between bone resorp-tion and surrounding tissue destruction culminating in b2M de-position and amyloid formation. More work is required to eluci-date the relationship between b2M accumulation and progres-sive tissue destruction.

Dialysis-related amyloidosis (DRA), also referred toasb2-microglobulin amyloidosis (Ab2M), is most com-monly seen in patients with end-stage renal disease(ESRD) undergoing long-term hemodialysis therapy(13). However, it also occurs among patients undergoingchronic peritoneal dialysis and prior to initiation of dia-lytic therapy (1, 4). The duration of dialytic therapy is animportant factor associated with the development ofDRA. Although DRA may be observed within 3–5 yearsof the initiation of dialysis, it is almost universal in pa-tients treated by hemodialysis for 15 years or more (1, 5).

DRA is characterized by amyloid deposition mainly inbone and surrounding joints, with clinical presentationsincluding carpal tunnel syndrome, destructive arthropa-thy, and pathologic subchondral bone erosions and cysts(1). Deposits removed from DRA-affected joints andperiarticular bone have demonstrated a unique amyloidfibril protein that was found to beb2M (2, 6). However,the pathologic role ofb2M as a mediator of bone diseaseis not completely understood. Essential issues regardingAb2M as well as its association with DRA remain unre-solved. The mechanism by whichb2M transforms intobone and joint amyloid deposits is unknown. The methodby whichb2M interferes with the normal bone remodel-ing process is also undetermined. This article reviewsinformation relevant tob2M as a mediator of bone ab-normalities in patients with renal disease.

b2M as a Bone Growth Factor

The physiologic effects ofb2M on bone remodelinginvolve complex regulatory pathways. Bone cells pro-duce b2M (7). It has been proposed that bone-derivedgrowth factor (BGDF), which has been isolated from ratcalvaria and bovine bone matrices, isb2M, as verified byBDGF and murineb2M amino acid composition to haveidentical amino-terminal sequences (8, 9). Humanb2Mand isolated BDGF similarly stimulate cell proliferationand protein synthesis in isolated osteoblast cultures, sug-gesting thatb2M has a mitogenic effect on human mu-rine bone cells. In several studies, variable concentra-tions of b2M have been required to stimulate osteoblastproliferation as measured by [3H]-thymidine uptake (7,8, 10). However, other investigators were not able todemonstrate similar results using isolated chicken osteo-blasts or murine calvarial cultures. In these studies, hu-man b2M did not induce a mitogenic effect, and themitogenic effect of bovine bone matrix-derivedb2M wasattributable to contamination with transforming growthfactor (TGF)-b or other unexplained factors (8, 9, 11).The incongruity observed with [3H]-thymidine incorpo-ration between isolated osteoblasts and calvaria suggestthat additional factors were present. Other studies havedemonstrated that although osteoblasts respond tob2M,b2M did not induce an increased production of alkalinephosphatase or osteocalcin by isolated osteoblasts (7,11). Thus it is uncertain if theb2M mitogenic effect onosteoblasts is direct, a result of contamination, or aninteraction with other local factors.

b2M may play a role in a complex interaction withhormonal receptors or as a regulator of the growth-promoting effects of other growth factors. Studies haveshown that the mitogenic effects of insulin-like growth

Address correspondence to: Stuart M. Sprague, DO, Divi-sion of Nephrology, Evanston Northwestern Healthcare,2650 Ridge Ave., Evanston, IL 60201. E-mail: ssprague@northwestern.edu.Seminars in Dialysis—Vol 14, No 2 (March–April) 2001 pp.131–133

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factor (IGF-I) on bone are augmented byb2M in upregu-lating IGF-I receptors, increasing IGF-I transcripts andpolypeptide levels. These effects ofb2M are selectivesinceb2M does not enhance TGF-b receptors or tran-scripts (10). Although not fully explained, it is plausiblethat b2M may have a physiologic role in bone remodel-ing.

b2M as a Bone-Resorbing Agent

Although the effect ofb2M on osteoblast proliferationis controversial, multiple other effects ofb2M on bonecell metabolism have been observed. Subcutaneous in-jection of b2M induces histologic evidence of bone re-sorption in neonatal mice (12), and purified humanb2Minduces a dose- and time-dependent net calcium efflux incultured murine calvaria (11, 13). This calcium efflux ismediated, in part, by interleukin (IL)-1b (14). b2M alsostimulates the synthesis of IL-6, a potent bone-resorbingcytokine, leading to an increase in mRNA and proteinlevels in osteoblasts (15).b2M has also been shown tostimulate synovial fibroblasts to produce stromelysin, aneutral matrix metalloproteinase (MMP), which is be-lieved to be a key enzyme causing articular destruction ininflammatory joint diseases (16). The finding thatb2Minduces the synthesis of collagenase-1 from rabbit syno-vial fibroblasts and the preferential collagen binding ca-pacity ofb2M also supports the hypothesis thatb2M hasa principal role in modulating connective tissue break-down (17, 18). Migita et al. (19) demonstrated thatb2Mincreases COX-2 protein and mRNA expression in adose-dependent manner from human synovial cells;however, utilizing the mouse calvarial resorption model,we were unable to demonstrate an effect ofb2M effecton prostaglandin E2 production (14).

Theoretically both hormonal and local regulatory fac-tors can interact withb2M either as a result of directstimulation byb2M or by aggravatingb2M effects onbone resorption. Various studies, however, have beenunable to demonstrate an effect of parathyroid hormone(PTH) on b2M transcription (20) or an effect of 1,25-dihydroxyvitamin D3 on b2M immunoreactivity in os-teoblast cultures (7). It has been demonstrated that sub-maximal and pharmacologic concentrations of PTH didnot augmentb2M calcium efflux from incubated calvarialcells (11). These preliminary data suggest thatb2M maynot interact with calciotropic hormones (7, 11, 20).

Cytokines, including IL-1, IL-6, and TNF-a, haveknown stimulatory effects on cell-mediated bone resorp-tion (21, 22). Patients undergoing chronic dialysis haveenhanced production and elevated basal concentrationsof cytokines (23, 24). With elevated levels it is possiblethat cytokines increase the rate of bone resorption andpredispose osteoarticular structures to the deposition ofb2M. The bone resorbing effects ofb2M, on the otherhand, may be mediated by interaction with cytokines.This result is consistent with the observation thatb2M-induced calcium release from neonatal calvaria can becompletely prevented by IL-1b antibody (14). Both hor-monal and local regulatory factors participate in a com-plex regulation of bone mineralization;b2M may act viaany of these pathways.

Advanced glycation end product (AGE) modificationof b2M appears to further increase theb2M-inducedbone resorption and cytokine production (25, 26). Uti-lizing an in vitro bone resorption assay, the number ofresorption pits formed by isolated osteoclasts are signifi-cantly increased by AGE-modifiedb2M compared tonative b2M (26). AGE modification ofb2M seems toalter bone metabolism in a variety of ways: it not onlyincreases bone resorption, but also decreases fibroblasticcollagen deposition. AGE modification ofb2M, com-pared to unmodifiedb2M, decreases fibroblastic synthe-sis of type I collagen (27). Of interest, the amyloid de-posits and surrounding macrophages from patients withDRA react with a monoclonal anti-AGE antibody (28).AGE-modifiedb2M stimulates chemotaxis of monocytesand macrophages and enhances the secretion of cyto-kines (29). Studies have demonstrated that purifiedb2Mmodified by AGE interacts with AGE receptor (RAGE)molecules on monocytes and macrophages, thus stimu-lating the release of cytokines such as platelet-derivedgrowth factor (PDGF), IL-6, TNF-a, and IL-1b. TheAGE-modifiedb2M stimulation of monocyte chemotax-is may be blocked by anti-AGE receptor antibodies. Ofinterest, RAGE has also been identified on osteoblasts(30).

Excess soluble RAGE inhibits the induction of TNF-afrom macrophages, demonstrating that the biologic ef-fects of b2M on monocyte/macrophages is RAGE de-pendent (29). In addition, it has been demonstrated thatAGE-modifiedb2M collected from patients, or producedin vitro, significantly increased in vitro bone resorptioncompared to normalb2M (25). Furthermore, it has beendemonstrated that sufficient secretions of TNF-a and IL-1b stimulate collagenase synthesis in cultured synovialcells, which may subsequently lead to collagen degrada-tion and connective tissue breakdown, a potential nidusfor further b2M deposition (31). Among the variousforms of acidicb2M, only AGE-modifiedb2M appearsto have these biologic properties. Thus the biologic effectof AGE-modifiedb2M on monocytes and macrophagesis thought to be mediated by RAGE. The finding thatAGE-modifiedb2M further induces bone resorption andosteoblastic cytokine release, as well as reduces type Icollagen synthesis by fibroblasts compared to unmodi-fied b2M, suggests a major role for AGE modification inthe bony destruction observed in DRA. The clinicalmanifestations of DRA might thus result from local in-flammatory reactions induced by monocytes and macro-phages, synovial cells, osteoclasts/osteoblasts in re-sponse to an AGE-modifiedb2M after progressive AGEtransformation of long-lived amyloid deposits.

Conclusion

In summary, the exact pathogenesis ofb2M as a me-diator of bone disease is unknown. DRA is typically seenamong long-term chronic hemodialysis patients. The in-creased duration of dialytic therapy directly correlateswith the incidence ofb2M amyloid accumulation in os-teoarticular tissue. Elevated serum levels ofb2M inchronic hemodialysis patients with or without clinical or

132 Tran et al.

histologic amyloidosis are similar, arguing againstsimple precipitation. Therefore various metabolic pro-cesses that might include proteolysis and/or AGE modi-fication may play a role inb2M amyloid deposition. Themechanism by which asymptomatic amyloid depositiontransforms into an inflammatory state with bone destruc-tion needs to be further elucidated. Experimental datasuggest thatb2M is directly involved in the process ofbone resorption via several complex regulatory pathwaysthat are probably mediated by both hormonal and localbone remodeling factors. Recent research with AGEs hasdemonstrated uremia as a state of increased oxidativestress, which stimulates the auto-oxidation of a variety ofunstable precursor substances, leading to augmentedAGE production. AGE-modifiedb2M has the ability toattract monocytes and stimulate macrophages to releaseproinflammatory cytokines, increase osteoclast-inducedbone resorption, and synthesize synovial cell collage-nase. Inflammatory local bone destruction precipitatesthe migration of further inflammatory cells, and the en-suing release of more cytokines, leading to further tissuedestruction. Local bone resorption and surrounding tis-sue become the site for continued deposition ofb2M andamyloid formation. Clearly more work is required to fur-ther elucidate the relationship between the accumulationfor b2M and evolution of progressive tissue destruction.

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133b2M-INDUCED BONE DISEASE