progress in the pathogenesis of ibodownloads.hindawi.com/journals/cjgh/1993/658920.pdf · progress...

PROGRESS IN THE PATHOGENESIS OF IBO

The role of macrophages in the pathogenesis of inflammatory

bowel disease

YR M AI 11 nA ML), MRCP

YR MAHIDA, The role of macrophages in the pathogenesis of inflammatory bowel disease. Can J Gastroenterol 1993;7(2):83-86. Normal intestinal macrophages are a he terogenous population of cells with phenotype possibly related todbtinct funct1ons. TI1e increase in the mucosa! populauon of macrophages in inflammatory howcl disease is derived from circulating monocytes which may have an enhanced capacity to perform a number of functions such as respiratory burst aet1v1ty, production of cytokines such as interleukin (IL) -1, IL-8 and po~ibly also antigen presenting activity. These activities (as well as others which remain to he characterized) may play an important role in the pathogenesis of Crohn's disease and ulcerative colitis.

Key Words: Crohn' s disease. Cycocynes, Dendritic cells , lncerleukins , Macrophages, Ulceratwe colitis

Role des macrophages dans la pathogenese de la maladie intestinale inflammatoire

RESUME: Les macrophages mtestinaux normaux sont une population hetcrogcnc de cellules dont le phenotype est possiblemcnt lie a des fonctions di~tinctes. L'augmentation de la population de macrophages au sem de la muqueusc dans la malad1e intestmale inflarrunatoire est dcrivee des monocytes circulants qui peuvent eue dotes d 'une capac1te accrue a accomplir certaines fonctions au plan de la respiration cellulaire, de la production de cytokincs comme l'mterleukine (IL)-1, IL-8 et possiblement egalement J'une activite anugemque. Ces activ1tes (et d'autres qu'il reste a decouvrir) peuvent jouer un role important clans la pathogcnese de la maladic de C rohn et de la colite ulcereuse.

Gastrormestinal Unit, Massachuseus General Hospital, Boston, USA Correspondence and reprints: Dr YR Mahula, Dwisron of Therapeurics, Univl'rnty Hmp,ral,

Queen's Medical Cencre, Nomngham NG7 2UH, U111tecl Kingdom

c~, J GASTROENTEROL VOL 7 No 2 FEI\RUAR) 1993

M AlROPHAl,b ARE [)ERJVE!) PRL

dominantly from c1 rcu laung monocytes which migrate into t issues. The functions of the e cells can he d ivided broadly m to th ree categories. First , they have a large capacity to produce a variety of mole~ules w1ch a broad range of bio logical activi ties. Second ly, they can process and present anugem, to T cells to m mate 1mmunolog1cal respomes. Thirdly, they are also highly phagocyttc. cells capable of 111-

gcstmg microbes and their products as well as other molecules.

T here is an increase in the mucosa! populanon of macrophages m mflammatory bowel d isease (IBO) which, m the form of granulomata, arc part icularly pro mment in Crohn's disease. There 1s much less mformation on human mtest inal macrophages than other intestinal cell popula tions an<l some of the studies on these cells m normal md1-viduals and IBO patients arc rev iewed. ln the intestmc, these cells arc a heterogenous populatton which have been studied in situ in ttssuc secttons; subsequently functional studies have been performed on isolated cells.

83

MAIIIDA

STUDIES ON MACROPHAGES IN SITU IN NORMAL BOWEL

Peyer's patches: Immune responses in the gut arc thuughl to he initiated in Peycr's patches and macrophages arc likely to be involved in this proces~. Studies using monoclonal antibodies and hiswchem1rnl stains h;ivc identified suhpl)pu lations of these cells 111

the~c regiom ( I ). A disuncl suhpopulat ion is present in rhe dome region, close to the surface epithelium, in a sire where rhey arc likely to encounter antigens taken up hy m1crofold ce lls. Other subsets arc present in the germinal centre and imerfollicular region. The latter cells arc phenotypically similar to intcrdigitaling cells of the paracorticnl T-Jependent regions of lymph nodes. Lamina propria: Macrophage hc tcrogcnei ty is also seen in the lamina propria of normal small and large intestine (2,3 ). Macrophages below the surface epithelium tend to be large and round, with strong acid phosphata~c activity, whereas those in deeper lamina propri.'l arc smaller with more processes and weak acid phosphatase activity. There arc also differences in phenotypes of macrophages between the small and large mtcstine; in the former, smaller cells with weaker acid phosphatase activity hut stronger A TPase activity prevail, compared to the latter (2). It is likely that these phcnotypic differences arc related tll their different functional requ irements which may be related to differences in the luminal contents.

STUDIES ON MACROPHAGES IN SITU IN ACTIVE IBO

In active lBO, macrophages with strong acid phosphatase activity arc present throughout the lam ina propria, and also in the submucosa in Crohn's disease. Using a panel of monoclonal antibodies, three suhpopulations of macrophages have been identified that are present 111 the laminc1 propria of the mucosa with act ive IBO but absent or only rarely present in the normal mucosa. These subpopulations were iJentified with the monoclonal antibodies RFD9 to FcRTII (C016) and to intcrleukin-2 (lL-2) receptor.

84

RFD9 pos1t1ve macrophages: In the normal intestine, thi s antibody only labelled macrophages in the germinal centres of Peycr's parches. In active IBO, RFD9 positive macrophages were seen in the lamina propria {3-5) and were present in clusters, especially in Crnhn \ disease. RFD9 positive cells h,Wl' abn ht'en seen in the inflamed mucnsa tif pouches (constructed after colcctnmy for ulcerative colitis) (6) but not in ccliac disease (7) or bacterial col II is. Thus, studies to date suggest that RFD9 pnsttivc macrophages arc seen nnly 111 the lamina propria of the mucosa with active IBO. It can be postulated that the cl usters of RFD9 posit ive macrophages in Crohn's disease may go on to form granulomata. Macrophages expressing FcRm: Surface labelling of macrophages with the anti-FcR111 antibody (CD16) was seen in the lamma propria of mucosa with active IBO (but not normal mucosa). Animal scudies suggest that cells expressing these low affinity lgG receptors may be involved in taking up and clearmg immune complexes (8), and the mucosa! macrophages in lBD may be performing a similar function. Macrophages expressing receptors for IL-2: Using an enhanced a lka line phosphatase amialkaline phosphatase technique, many lL-2 receptor positive cells have been demonstrated in the mucosa with active IBO (9). T hese cells had morphological appearances of lymphocytes and macrophages. Identity of the latter was confirmed by studies on isolated cells ( which also showed that these cells are in an enhanced state of activation) . Using a similar technique, Choy et al have subsequently also reported the presence of lL-2 receptor positive cells in the mucosa with active ulcerative colitis and Crohn's disease and have confirmed the identity of the other population of cells co be T cells ( 10). Studies on Crohn's granulomata: Heterogeneity of macrophages in and around granu lomata h as also been demonstrated (11 ). Epithclioid cells and giant cells were labelled with the antiboJy RFD9 but not RFD7, whereas macrophages in the surrounding lam ma propria were usual ly RFD7-positive but

RFD9 negative. The epithcl irnd cclb expressed I L-2 receptor~ and were strongly acid phosphate activny positive. Double staining studies showed that the epithclinid cel ls were closely associated with CD4 positive (hel1~er) T cells whereas CD8 positive cells were much fewer m number and located mainly at the periphery. Thus, Crohn\ grnnulomata comprise collccttons of cells of d1stmct phenotype~ which are likely to be constantly u1tcractmg with each other.

FUNCTIONAL STUDIES ON ISOLATED MACROPHAGES

Respiratory burst activity: Compared to a resting cel l, an activated macrophage has an enhanced capacity to perform a number of funct ions which include secretion of oxygen radicals and enzymes (such as neutral proteases and acid hydrolases). There is an associated increased expression of major histocompatibility complex (MHC) class ll molecules and Jownregulation of mannose receptors on the cell surface. Activation of macrophages may occur in response to microorganisms and their products or interferon gamma (secreted by T cells) and allows the cell to mediate antimicrobial and cytotox ic effects (12).

In order to determine the activation phenotype of intestinal macrophages, the capacity of the isolated cells to undergo a respiratory burst (as assessed by conversion of the dye nitroblue tetrazolium to a blue brown reaction product) was scudied ( 13).

Opsonised zyrnosan and phorbol myristate acetate were used to trigger the release of oxygen radicals. The majority (80% or more) of isolated normal intestinal macrophages did not demonstrate release of oxygen radicals. In contrast, a significantly increased proportion of macrophages from the intestine of patients with IBO were able to undergo a respiratory burst, suggesting that they were activated.

When the normal intestinal macrophages were stimulated with interferon-gamma (normally a very potent activator of macrophages), significant proportions of the cells were still unahlc to release oxygen radicals. This

CAN J GASTROENTEROI V~)L 7 Nt1 2 FEBRUARY 1993

suggests that the majority of the normal intestinal tissue macrophages arc JownregulateJ with respect to this function and that the cells with 111-creased capacity to release oxygen radicals arc derived prc<lom1mmtly from monocytes which hnve recently migrated into the muco~a. Oxygen radicals produccJ by activateJ macrophages may also contribute to tissue Jamagc m the mucosa. Antigen presen ting activity: ScuJics in mice have shown that Jen<lrittc cells are the most porent antigen present111g cells and that these cells may be J1st111ct from macrophages. In order to characterize cells with potent antigen presenting activity m the human intc~tinal mucosa, isolated mononuc.lcar cells were studied (14 ). Their capacity to

induce proliferation of highly purified, resting, allogene1c, peripheral hlooJ T cells wa~ useJ to assess antigen presenting awviry. This activity 111 isolated mononuclear cells was rcduccJ upon Jcplet1on of macrophages (by aJhercnce co fibroncctin and pannmg using a macrophage spec1f1c. monoclonal ancibody). In contrast, the fibronectin adherent cells ( which are enriched for macrophages) had enhanced antigen presenting activity. During the mixed lymphocyte rcacuons, clusters were ,hown to contain cclb with a dendritic morphology which were closely associated with proli feratingT cells. The cells w1thdendritic morphology were stnmgly positive for human lymphocyte an-

REFERENCES l Mah1da YR, Patel S, Jewell DP.

Mononuclear phagocyte system nf human Peyer's patches: An 1mmunoh1swchem1cal Mudy using monoclonal antibodies. Clin Exp lmmunol 1989;75:82-6. Selby WS, Poulter L W, Hobhs S, Jewell DP, Janossy G. Heterogeneity of HLA-DR pos1civc histiocytl's in human intestinal lamina proprn1: ;i combined hi!.tochemical and immunohistulog1cal analysis. J Clm Pacho!. 1983; 36: 379-84.

J. Mah1da YR, Pacel S, Gionc.hecti P, Vaux D, Jewell DP. Macrophage subpopulations in lamin.1 pmpria nf normal anJ mfiamcd colon ,mJ tem1mal ileum. Gut l 989;10:826-34.

4 G1onchem P, Mah1d.1 YR. Pacel S, Jewell DP. Macrophage and

tigen DR ( HLADR) and also cxpn.:sseJ antigens specific for macrophages. It was concluded therefore that cells with potent antigen prescnung activity 111 the intest ina l mucosa have charn<-tenst1cs of both dendrittc. cells anJ macrophages.

Cells isolated from mucosa wtth active IBO were shown to have enhanced antigen presenting activity compared to mononuclear cells from normal mucosa. IL-IP production : IL-I is a cyrok111e with a wide range of biological properties which may play an nnportant role in the pathogenesis of IBD ( I 5). Monocytcs and macrophages arc the main source of rh1s cytokine of wh ich IL- Ip 1s the prcdommant secreted from.

In vitro culture nf mononuclear cells isolated from intestinal mucosa with active IBD have been shown to produce s1g111ficantly greater amounts of IL- l p than eel ls from nnrma I mucosa ( 16). Depletion studies (by pann111g) showed that macrophages were the predom111ant pnxlucers nf this cytok111e in the intestine. Culture of mononuclear cells from normal mucosa in the presence of l1popolysacchamlc (a very potent stimulant of IL-I proJuccion by monocytes) did not result 111 any increase in the pnxluctton of IL- Ip, whereas there was an mcrease 111 the amount of the cytokine produced by cells from the inflamed mucosa. As for studies on respiratory burst activity,

lymphocyte subpopulauons 111

magnifying enJoscoptC lesmns nf Crohn\ disea~e. Clm Exp lmmunol 1988;72:373-6.

5. Allison MC, Cnmwall :,, Poulter LW, Dhillon AP, Pounder RE. Macrophage heterogeneity m normal colonic mucosa and 111 mfiamma[<)ry bowel d1sea,e Gut I 988;29: l 531-8.

6. De Silv,1 HT, Jones M, Prince C, Kettlewell M, Mortensen NJ, Jewell DP. Lymphn..:yte and macrophage subpopul.irions m pelvic ileal pouches. Gm 199l;32:l 160.

7. Mahida YR, G.1llagher A, Kurlak L, Hawkey CJ. M;1emphages in nonnal and cneliac dundenal mucosa. uasmienterolngy I 990;98:A46 I.

8. Clarhon SB, K11nberley RP, Valimky et al Blockagl' of cle,uance nf immune n1mple,e, hy .11111-Fc ·gamma receptor

CAN] 0ASTR0ENTER0l VOL 7 No 2 rERRUARY 1991

Mac rophages In IBD

chis suggests that the normal mtcstinal tissue macrophages arc downregulated and that the enhanced production of IL-IP 111 lBD 1s denved from monocytes which haw recendy migrated 111to the mucosa.

Studies on ussue homogenates have confirmed enhanced 111 vivo producuon of IL-113 111 the mucosa of patients with act1ve IBO ( 17,18). Stud ies on IL-6 and IL-8 : Recent studies on lL-6 and IL-8 ( which can be secreted macrophages) 111 IBO suggest that macrophage~ may have different functional capac1t1es 1n ukcranve col1us and Crohn\ disease. Higher c1rculacmg levels of IL-6 have heen found in act ive Cmhn's disease compared tn

ulcerative cnlttis hy three groups 111dcpendently ( 19-2 1 ).

Higher tissue levels of IL-8 have been demon,trated m the mucosa of patients with acm·e ulcernt1ve colitis compared to acnvc Crohn \ d1se,t~c. Patients with active ulcerative colitis also had high urc.ulat111g levels of ,mnhod1es co IL-8, hut this was not rhe ca~c in patient, with active Crohn's disease where the level~ were similar co normal controls (22). IL-8 1s a very potent chemoattractant for ncutroph1ls and the high mucosa! levels in active ulcerative colm, may explam the characteristic predom111ance nf neutrophils 111 this wnd1uon. The role of macrophag~ 111 the differential production of these cytokines 111 ulcerative colms and Crohn's d1sca\C rcma111s to be determined.

monuclunal ,mu1',><.ly. J Exp Med l 986; 164:474-89.

9. Mahida YR, Wu K, Pntd S,Jcwcll DP. lL-2 recep(l)r exprc'-'>mn hy macro, phages 111 mfiamm.irory howel d1se;1se. Clin Exp lmmunol 1988;74: 382-6.

lO. Ch,i} MY, Walker-Smith JA, W11l1ams CB, MacDnnalJ TI. Difforenual expression of CD2 5 ( inccrleukin 2 receptor) on lamina propna T cells and macrophages in the mte,rinal le,mns m Cmhn\ d1se,1sl' .mJ ukcrarn·e collfls. Gut 1990; 3 l: 1365-70

1 I. M,1hida YR, Pacd S, Jewell DP Macrnphage an,I lymphnq re subpnpulnwms 111 the grnnuloma nf Cwhn's disease. In: Macl'lcrmo11 RP. ed. lnflammarory Bowel Disease. Currenc Status and Future Approach. A111-1erdanr Ebev1er Suence Puhh,her, BV, 1988:137-41.

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MAll!l)A

12. Adams DO, I lamilron TA. The cell biology of macrophage activation. Ann Rev lmmunol l 984;2:283-318.

13. Mahida YR, Wu K. Patel S, Jewell DP. Respiratory burst activity of inccstinal macrophages in nom1al and inflammatory howcl dtsease. Gm 1989;30: 1362-70.

14. Mah,da YR, Wu K, Jewell DP. Ch:micterizauon of antigen presenting activity of mononuclear cell, isolated from normal and inflammatory bowel disease colon and ileum. Immunology l 988;65:543-9.

15. Cominelh F, Nast CC, Clark BD, Schindler R, Llerena R, Eysselcin VE, Thompson RC, Dinarcllo CA. lnterleukin-1 (IL 1) gene expression, synthesis and effect t1f specific receptor

86

hlockade in rabbit immune complex colit1,. J Clin Invest l 990;86:972-80.

16. Mahida YR, Wu K,Jewell DP. Enhanced production of interleukin IP hy mononuclear cells 1sol,1ted from mucosa with active ulcerative colms or Crohn\ disease. Gut l 989;30:835-8.

I 7. Mahida YR, Lamming CED, Gallagher A, Hawthorne AB, Hawkey CJ. 5-aminosalicyhc acid is a potent inhibitor of !LIP production in organ culture of colonic biopsies frl1m patients with inflammatory bowel disease. Gut 1991 ;32:50-4.

18. Ugumsky M, Simon PL, Karmeli FC, Rachmilewitz D. Role of interleukin- I m mflammatory bowel disease -enhanced producuon durmg active disease.Gut 1990;31:686-9.

19. Mahida YR, Kurlak L, Gallagher A, Hawkey CJ. High circulating levels of interleukin 6 in act1ve Crohn's disease but not ulcerative colms. Gue 1991 ;32: 1531-4.

20. Gross V, Andus T, Caesar 1, Roth M, Scholmench J. Evidence for continuous stimulation of intcrleukin-6 production in Crohn \ disease. Gastroenrerology 1992;102:514-9.

21. Lobo AJ, Jones SC, Evans SW, Banks R, Rathbone BJ, Axon A TR. Plasma interleukm-6 in inflammatory bowel disease. Gut 1990;3 l :All94.

22. Mah1da YR, Ceska M, Effenherger F, Kurlac L, Lindley I, Hawkey CJ Enhanced synthesis ofNAPI/IL8 in active ulcerative colitis. Clin Set 1992;82:273-5.

CAN J GASTRl)ENTERt)l Vt)L 7 No 2 FEBRUARY 199~

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