Annals of the Rheumatic Diseases, 1988; 47, 463-467

P component in the synovium in rheumatoid andosteoarthritisM G BUTLER, A J D'ARDENNE. AND D L SCOTT

Fromii the Departmzents of Rhelnzwatology amid Histopatlologv, St Bartholomew's Hospit(ll Climiical ResearchCenitre, West Smizitlhfield, Londloni

SUMMARY P component is present in amyloid deposits, normal serum, and normal tissues inrelation to elastic fibres. Its pathological role in inflammatory synovitis was investigated. Itsdistribution was determined immunohistologically in 33 synovia: 15 rheumatoid; sevenosteoarthritic; seven traumatic controls; and four infected biopsy specimens. P component waspresent in two circumscribed distributions: extracellular fibrils in dense fibroelastic tissue of themore fibrotic synovia; and in the arterial wall, where it was confined to a single elastic lamina insome cases and in others showed reduplication and fragmentation. These were not related toamyloid material. It shows no disease specificity, but P component categorises the nature of thepathological reaction and is typically in biopsy specimens showing the development of chronicfibrosis. There was close codistribution of P component with elastic tissue, though this was notabsolute. P component had a different distribution from C reactive protein (in synovial lining celllayer), and fibronectin, which was absent from fibrotic areas. Understanding the pathologicalinteractions of P component may help elucidate why some synovial reactions remaininflammatory and others progress to chronic fibrosis.

Structural glycoproteins are a heterogeneous groupof extracellular connective tissue matrix proteins.'They include fibronectin, laminin, P component,and related glycoproteins. The distribution of fibro-nectin and laminin has been described in thesynovium in rheumatoid arthritis and relateddisorders.2-7 P component has not been previouslyexamined in normal or diseased synovium. It is aserum protein with marked homology to C reactiveprotein.8' P component is not an acute phasereactant in humans, though it is in mice."' Tissue Pcomponent was originally described as an invariablecomponent of amyloid deposits.'" Its distribution isnow known to be more extensive, with a significantstructural role in glomerular basement mem-branes,'2 and more widely in tissues in relation toelastic fibres.'-3To investigate whether P component has a

pathogenic role we examined its distribution in theinflamed synovia from patients with rheumatoidarthritis and also studied osteoarthritic and non-inflammatory synovial biopsy specimens. For com-

Accepted for publication 19 Noscmber 1987.Corrcspondence to Dr D L Scott. Dcpartmcnt of Rhcumatology.St Bartholomew's Hospitill. Wcst Smithficid. London ECIA 7BE.

parative purposes we evaluated its relation tofibronectin, laminin. and C reactive protein.

Materials and methods

Thirty three operative synovial biopsy specimenswere obtained from 15 patients with rheumatoidarthritis, seven with osteoarthritis, four with tuber-culosis of the synovium, and seven with torn menisciand other non-inflammatory traumatic conditions.The distributions of P component, laminin, fib-ronectin, and C reactive protein were studied byindirect immunohistochemistry on 6 [sm sections offresh frozen tissue, on ethanol fixed, paraffinembedded tissue by the Saint Marie method,'4 andformal fixed, paraffin embedded tissue after enzymedigestion. '5 Both immunofluorescence and im-munoperoxidase techniques were employed. Tissuesections were also stained by haematoxylin andeosin and by Weigert's stain for elastic tissue.Further paraffin sections were stained by alcoholicCongo red and amyloid deposits sought by bothpolarised and fluorescent light.

Monospecific antisera employed were as follows:goat antirabbit P component (Atlantic Antibodies053-03); goat antihuman fibronectin (Sigma F-

463

copyright. on D

ecember 29, 2019 by guest. P

rotected byhttp://ard.bm

j.com/

Ann R

heum D

is: first published as 10.1136/ard.47.6.463 on 1 June 1988. Dow

nloaded from

464 Butler, D'Ardenne, Scott

1509); rabbit anti-C reactive protein (Dako A073);rabbit anti-laminin (BRL 6265SA). Appropriatelylabelled second antisera for indirect immuno-fluorescence and immunoperoxidase techniqueswere obtained from Dako. Control sections wereincubated with non-immune goat and rabbit sera inplace of the primary antisera. All control sectionswere negative. Sections were counterstained withhaematoxylin.

Results

All synovial biopsy specimens contained someimmunoreactive P component, though in certaininstances only small amounts were present. Immuno-histological staining for P component showed a simi-lar pattern with formal fixed paraffin embeddedtissues, cold ethanol fixed tissue, or fresh frozenmaterial. Two patterns of immunoreactivity for Pcomponent were discerned: firstly, fibrillar immuno-reactivity in dense fibroelastic tissue beneath thesynovial lining cells (Fig. 1) and secondly, vascularrelated immunoreactivity. There was perivascularpositivity around most vessels larger than capillar-ies. Some arteries showed increased immunoreac-tive P component in the vessel walls (Fig. 2). Inothers it was confined to a single internal lamina.Vascular basement membranes were negative for Pcomponent.

There was a close relation between the distribu-tion of P component and that of elastic fibres.Vessels with abundant immunoreactive P compo-

nent in their wall corresponded with those showingconspicuous reduplication of their internal elasticlaminae on conventional histological staining (Fig.2). Fibrillar P component was in a similar distribu-tion to that of stromal elastic fibres. The codistribu-tion was not complete, however, and there wasfrequently more immunoreactivity for P conmponentthan elastic fibre staining. At no time was it possibleto demonstrate any staining for amyloid.Comparisons of immunoreactive P component, C

reactive protein, fibronectin, and laminin showed noevidence to suggest they were codistributed. Creactive protein was confined to the synovial liningcell layer, which showed only weak positivity.Fibronectin was far more widely distributed withreactivity within the synovial lining cell layer, inarticular fibres in the underlying subsynovial con-nective tissue, and in close relationship to smallsynovial blood vessels. Laminin was present in allvascular basement membranes but not in internalelastic laminae. P component contrasted with thesein being more closely associated with fibrosis,whereas the others were more prominent in areas ofsynovial proliferation.There was no clear relation between diagnostic

category and the amount of immunoreactive Pcomponent. Its distribution, however, showed adistinct pattern in relation to the pathologicalchanges in the synovium. Control biopsy specimensshowed only very small amounts of P component. Inthe inflamed synovia, principally in some cases ofrheumatoid arthritis and the patients with tubercu-

I ...

/.. Fig. I Immunoreactive Pcomponent in a fibrillardistribution among densefibroelastic tissues.

copyright. on D

ecember 29, 2019 by guest. P

rotected byhttp://ard.bm

j.com/

Ann R

heum D

is: first published as 10.1136/ard.47.6.463 on 1 June 1988. Dow

nloaded from

P component in the synovium in RA and OA 465

_ p----trrS #)

k-I*IS

I1 K

;>

_f 4

t

.1

.I

Fia. 2(ai1)

A

Figz. 2(b)Fig. 2 Consecutive sections of a small artery showing similar distribution of positive staining. (a) Immunoreactive Pcomponent; (b) Weigert's elastic stain.

r A~~~~~~~~~~~~~~~~~~~1

#1~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~1

Fig. 3 P component in the wallsofsmall blood vessels showingevidence of reduplication.

jI

It.

24

..4It.I . -

- .e I

I

/I

- I

*.b.

copyright. on D

ecember 29, 2019 by guest. P

rotected byhttp://ard.bm

j.com/

Ann R

heum D

is: first published as 10.1136/ard.47.6.463 on 1 June 1988. Dow

nloaded from

466 Butler, D'Ardenne, Scott

lous synovitis, P component was conspicuouslyabsent from areas showing inflammatory cell infil-tration. In contrast, biopsy specimens with earlyfibrotic changes showed more prominent P compo-nent (Fig. 3). It was especially prominent in theblood vessel wall at these sites, where it frequentlysuggested evidence of reduplication of the elasticlaminae and related structures.

Discussion

P component is a minor structural glycoprotein ofnormal and diseased synovia. It is closely related toelastic fibres in the synovium, in keeping withprevious observations of other normal and patho-logically involved tissues. 16 Its distribution is distinctfrom that of C reactive protein, laminin, andfibronectin. These differences are noteworthybecause there is marked structural homology bet-ween P component and C reactive protein, and invitro P component dimers specifically bind fibro-nectin in a calcium dependent manner. 17 Laminin isconfined to blood vessel basement membranes.7Fibronectin has a wider distribution in the synoviallining cell layer, in relation to fibrillar extracellularmatrix proteins, and around blood vessels.2-4 Creactive protein is located in the synovial lining celllayer.'8 This coincides with its relatively low syn-ovial fluid level,'9 suggesting deposition in the liningcells.

Tissue amyloid deposits always contain Pcomponent. " Indeed it is only recently that Pcomponent has been recognised as a component ofnormal tissue. None of the patients we studied wasconsidered to have systemic reactive amyloidosisnor was there evidence of amyloid deposits in thesynovium. Goffin et alP'" and Ladefoged2' have bothreported material with some histological features ofamyloid deposits in the synovium; though theevidence that they are true amyloidotic material isnot strong. The distribution of P component is alsodifferent from that of the non-collagenous reticulincomponent which is present in many tissues,22including the inflamed synovium.23 This tissue com-ponent is extracted by distilled water purification, asare amyloid fibrils. At the ultrastructural levelrheumatoid and osteoarthritic synovia contain largeamounts of fibrillar protein material, thoughits exact nature is uncertain (C J Morris and CHollywell, unpublished observations). Although byconventional standards synovial P component is notrelated to amyloid deposits, the exact nature anddistribution of extracellular fibrillar proteins inarthritis is a complex area of which relatively little isknown.P component is related to early fibrotic change

and is a weak marker of this process in thesynovium. It is not associated with inflammatory cellinfiltrates. The absence of any specific relation witha single diagnostic category is similar to findingswith other connective tissue proteins, and is typicalof many synovial histological changes. Nevertheless,the suggestion that P component may characteriseearly fibrosis is theoretically interesting, though atthis stage it is of limited diagnostic value.

Vascular changes are common in synovial biopsyspecimens, and vasculitis is a frequent extra-articular feature of rheumatoid arthritis. Inflamedsmall blood vessels in the synovium contain relativelylittle P component, but larger arteries have far moreP component in their walls, frequently suggestiveof reduplicative changes in the arterial wall. Com-pletely or partially occluded vessels often containlarge amounts of P component. The presence ofvessel wall P component may be particularly signifi-cant in the pathogenesis of rheumatoid vasculitis,and further studies are needed in this area.

Wc thank the Arthritis and Rheumatism Council. thc North EastThamcs Rcgional Hcalth Authority, and the Joint Rcscarch Boardof St Bartholomew's Hospital for their support.

References1 Hay E D. Cell biology of extracellular matrix. New York:Plenum Press, 1981.

2 Scott D L. Wainwright A C. Walton K W. Williamson N.Significancc of fibronectin in rhcumatoid arthritis and ostcoar-thritis. Anni Rheu,n Dis 1981; 40: 142-53.

3 Scott D L. Dclaimcre J P. Walton K W. The distribution offibronectin in the pannus in rhcumatoid arthritis. Br J ExpPathol 1981: 62: 362-8.

4 Vartio T. Vaheri A. Van Essen R. Isomaki H. Stenman S.Fibronectin in svnovial fluid and tissue in rheumatoid arthritis.Eur J Cliii Invest 1981: 11: 207-12.

5 Clemmcnscn 1. Holund B, Andcrsen R B. Fibrin aind fib-ronectin in rheumatoid synovial mcmbranc and rheumatoidsynovial fluid. Arthlritis Rheu,n 1983; 26: 479-85.

6 Mapp P 1. Revell P A. Fibronectin production by synovialintimatl cells. Rheu,natol 1hit 1985; 5: 229-37.

7 Scott D L, Salmon M. Morris C J, Wainwright A C. WaltonK W. Laminin and vascular proliferation in rheumatoid arthri-tis. Annii Rheuwn Dis 1984; 43: 551-5.

8 Pepys M B, Dash A C, Markham R E, Thomas H C, WilliamsB D, Petrie A. Comparative clinical study of protein SAP(amyloid P component) and C reactive protein in serum. ClinExp Immunol 1978; 32: 119-24.

9 Pepys M B, Dash A C. Fletcher T C. Richardson N. Munn E A,Fernstein A. Analogues in other mammals and fish of humanplasma proteins. C reactive protein and amyloid P component.Nature 1978; 273: 168-70.

10 Pepys M B, Baltz M, Gomer K, Davies A J S. Docnhoff M.Scrum amyloid P-component is an acute phase reactant in themouse. Nature 1979; 278: 259-61.

11 Skinner M, Pepys M B, Cohen A S, Heller L M, Lian J B.Studies of amyloid protcin A P. In: Glenner C G, Pinhoe CostaP, Falcao de Freitas A, eds. Amyloid and amyloidosis.Amsterdam: Excerpta Medica, 1986: 384-94.

12 Dyck R F. Lockwood C M, Kershaw M, et al. Amyloid Pcomponent is a constituent of normal glomerular basementmembrane. J Exp Med 198(0; 152: 1162-74.

copyright. on D

ecember 29, 2019 by guest. P

rotected byhttp://ard.bm

j.com/

Ann R

heum D

is: first published as 10.1136/ard.47.6.463 on 1 June 1988. Dow

nloaded from

P component in the synovium in RA and OA 467

13 Breathnack S M, Melrose S M, Bhogal B, et al. Amyloid Pcomponent is located on elastic fibre microfibrils in normalhuman tissue. Nature 1981; 293: 652-4.

14 Saint Marie G. A paraffin embedding technique of studiesemploying immunofluorescence. J Histochem 1962; 10: 25-60.

15 Kirkpatrick P. D'Ardenne A J. Effects of fixation and onzymicdigestion on the immunohistochemical demonstration of lami-nin and fibronectin in paraffin embedded tissue. J Clin Pathol1984; 34: 637-44.

16 Breathnack S M. Melrose S M, Bhogal B, De Beer F C, BlackM M, Pepys M B. Immunohistological studies of amyloid Pcomponent in disorders of cutaneous elastic tissue. Br JDermatol 1982; 107: 443-52.

17 De Beer F C. Baltz M L. Holford S. Feinstein A. Pepys M B.Fibronectin and C4-binding protein are selectively bound byaggregated amyloid P component. J Exp Med 1981; 154:1134-49.

18 Gitlin J D, Gitlin J I. Gitlin D. Localisation of C reactive

protein in synovium of paticnts with rheumatoid arthritis.Arthritis Rheu,n 1977; 20: 1491-9.

19 Bourne J T. Shine B. Bewgum Baig F. Doyle D V. C reactiveprotein in synovial fluid and scrum. Br J Rheitinatol 1985: 24:203.

20 Goffin Y A. Thona Y. Potvliege P R. Microdeposition ofamyloid in the joints. Anntt Rheu,n Dis 1981: 40: 27-33.

21 Ladcfoged C. Amyloid in osteoarthritic hip joints: deposits inrelation to chondromatosis. pyrophosphate. and inflammatorvcell infiltrate in the synovial membrane and fibrous capsulc.Ann Rheurn Dis 1983; 42: 659-64.

22 Scott D L, Salmon M. Walton K W. Reticulin and its relatedstructural connective tissue proteins in the rheumatoid svno-vium. Histopathology 1984; 8: 469-79.

23 Unsworth D J, Scott D L, Almond T J, Beard H K, HolborowE J, Walton K W. Studies on reticulin. I. Serological andimmunohistological investigation of the occurrence of collagentype III, fibronectin and the non-collagenous glycoproteins ofPras and Glynn. Br J Exp Pathol 1982; 63: 154-66.

copyright. on D

ecember 29, 2019 by guest. P

rotected byhttp://ard.bm

j.com/

Ann R

heum D

is: first published as 10.1136/ard.47.6.463 on 1 June 1988. Dow

nloaded from