Heat shock proteins in immunopathology

Heat shock proteins in immunopathology Pieter C.M. Res, Jelle E.R. Thole and Rend R.P. de Vries University Hospital, Leiden, The Netherlands In recent years, studies have suggested that autoimmunity and/or immunopathology may sometimes result from the immune response to heat shock proteins of autologous cells and microorganisms. Focusing on the T-cell mediated responses, we review the lastest literature on this issue with regard to three hypothetical concepts of immunopathology in which heat shock proteins might play a role.

Current #Opinion in Immunology 1991, 3:924-929

Introduction

parasites,

Heat shock proteins (hsps) are among the most conserved mokzcules in nature, and in recent years have received much attention because their functions were shown to be vital in the eukaryotic and prokaryotic cell. Another important finding was that hsps were recognized as major targets in the immune response against many pathogens (for a review on hsps and immune responses

see [L-31.

Hsps may be divided into a number of families on the basis of their approximate molecular weight, and members from the extremely conserved hsp65, hsp70 and hsp90 families have been identified in all the prokaryotes and eulcaryotes studied so far. For example, the mycobacterial hsp65 displays 50% homology with the human hsp65

141.

Under physiological conditions these three families function as so called ‘molecular chaperones’ for polypeptides that, by transient binding to these polypeptides, guide their assembly into oligomeric structures, their folding and unfolding during transport into various compartments, or their degradation by proteolytic enzymes. Other hsp families are proteases that degrade r&folded or ‘foreign’ proteins. Under a variety of stress stimuli such as heat-hence the term heat shock proteins-but also nutrient deprivation, oxygen radicals, viral, bacterial, and parasitic infections, cells respond by increasing the production of specific hsp and by changing the intracellular compartment of some hsp. Some hsps may even become expressed at the ceU surface. The heat shock response probably protects the cell during adverse conditions by maintaining a functional conformation of essential proteins, and by assisting in the removal of denatured proteins. Studies of the immune response to hsps, in patients and healthy individuals infected with pathogenic bacteria or

have indicated that 11~~65, hsp70 and hsp90 expressed by these microorganisms are major targets for antibodies and T cells. However, in healthy individuals without any apparent infection, antibodies and T cells specific to, for example, the mycobacterial hsp65 could be detected also. Some of these studies showed that part of the T-cell responses were directed to self hsp65, indicating the presence of autoreactive T cells in both healthy individuals and patients [ 5,6,7*-9*,1O**,ll]. Because of the presence of hsp in microorganisms and autologous cells it is difficult to define the origin of the specific hsp that induced these responses. T-cell epitopes specific for bacterial hsp65 and epitopes conserved between bacteria and self hsp65 have been defined [7*,10**,11]. However, T-ceU epitopes specific for self hsps have not been identified yet. Hsps may be immunodominant in both patients and healthy individuals because these proteins are abundant in both pathogenic and non-pathogenic microorganisms, especially under the adverse conditions imposed upon them by the host environment. In addition, the presence of hsps in these microorganisms but also in stressed or transformed autologous cells ensures the almost continuous restimulation of immunologic memory for hsps in the host. Finally, some hsps may be intrinsically antigenic, because they are easily processed and presented to the immune system, and the fact that hsp65, hsp70 and hsp90 are functionally ‘sticky’ may be of significance in this respect. These data point at a twofold function of the immune response to hsp. Firstly, it may be used to eliminate pathogenic microorganisms. The rapidity of this response provides a first line of defense before immunity to pathogen-specific antigens is mounted. Secondly, this response may function in the elimination of stressed or autologous cells that express a sufficient concentration of hsp on their ceU surface, whereas normal cells do not

Abbreviations BCG-hacillus Calmette-CuPrin; HPLC-high performance liquid chromatography; hsp-heat shock protein; IDDM-insulin-dependent diabetes mellitus; IL-interleukin; IFN-interferon; NOD-non-obese diabetic; PBMNC-peripheral blood mononuclear cell; RA-rheumatoid arthritis; ReA-reactive arthritis; SFMNC-synovial fluid mononuclear cell.

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Heat shock proteins in immunopathology Res, Thole, de Vries

generate this density of self epitopes at their outer membranes. Thus, immunity to hsp, in general, may be beneficial to the human host in the elimination of both foreign microorganisms and aberrant autologous cells. However, in recent years it has become clear that depending upon factors such as genetic background, hormones, and environmental stimuli, an improperly regulated immune response to hsp at certain sites in the host may result in immunopathology, and even autoimmune disease. This may be caused by hsp-reactive T cells and/or antibodies. An example of a disease in which immunopathology may be induced by antibodies is lepromatous leprosy. Lepromatous leprosy patients suffer from numerous granuloma with a high dose of live M~~6acterium leprue bacilli. The peripheral blood mononuclear cells of lepromatous leprosy patients display no proliferation to a sonicate of the inciting pathogen, whereas high antibody titers to M leprueare present in the sera of these patients, suggesting that the latter components of the immune system are probably responsible for the immunopathology observed in lepromatous leprosy patients. Systemic lupus erythematosus is an example of an autoimmune disease in which antibodies to hsps namely, hsp70, hsp90 and ubiquitin have been implicated in the disease process [ 12-141. However, in this review we will focus on three different, but not mutually exclusive, concepts of immunopathology concerning disregulation at the level of T-cell-mediated immunity and w-ill briefly discuss a few human and animal (autoimmune) diseases to which these concepts might apply.

lmmunopathology resulting from the T-cell response to exogenous hsp The first concept concerns the possibility of immunopathology as a direct consequence of a poorly controlled T-cell-mediated immunity to hsps of invading microorganisms. If the imrnunopathology caused by the host defense response against exogenous hsps specifically affects certain sites or organs of the body, one may mistake the clinical features as being the result of autoimmunity (which is actually the reactivity of T cells or antibodies to self components). This may well account for reactive arthritis (ReA), an acute form of arthritis initiated by an infection of the gastrointestinal or the genitourinary tract by certain bacteria, such as Chhmydia, Shigelh, Salmonella and Yersinia. Several experimental findings suggest that a local response of joint T cells to proteins (hsps) of the inciting bacteria deposed in the joint is perhaps involved in the initiation or perpetuation of the arthritis. These findings are the following: products of the speciUc bacteria initiating the ReA have been demonstrated in the joint, although attempts to isolate and culture these bacteria from the synovial fluid have failed [ 15=,16~] ; the synovial fluid mononuclear cells (SFMNCs) display strong proliferative responses to extracts of the whole bacteria, and synovial fluid T-cell clones responding to these bacteria have been generated [ 10=*,17,18*,19]. Specific recognition of bacterial hsp65 by T cells of joints has been

demonstrated [ 10**,18*,20**]. In the absence of added Chhmydia antigens, Ch’umydia-reactive T-cell lines from a CMurn~~dia-infected ReA paitent responded to autologus synovial fluid denditic cells, but not to autologous peripheral blood dendritic cells . T-cell lines made against control antigens were not stimulated. This suggests that those synovial fluid dendritic cells specifically picked up Chhmydia antigens in vivo [21-l. Another possibility is that the T-cell lines responding to these synovial fluid dendritic cells were actually recognizing host epitopes, for instance, of the human hsp65 (see below). In addition to ReA, the concept described in this section may also be relevant for trachoma, an inflammatory disease of the eye aRlict.ing individuals infected with Chlumydiu truchomutis. Similar to the joints of ReA patients, Chlamydial antigens and DNA are detected in the conjunctivae of individuals suffering from trachoma and viable Chlamydiu are not demonstrable. In particular, studies on experimentally induced trachoma in animal models provide convincing support for the hypothesis that the host immune response to CbLamydiu can be deleterious. It was found that Wamydia hsp65, after application onto the conjunctivae, elicits ocular intlammation in guinea pigs and non-human primates infected with Chhmydia bacteria. In contrast, inoculation of Cbhmy din hsp70 had no effect, suggesting that only the T-cell response to Chhmydia hsp65 is critical in inducing eye inUammation [ 22*,23**]. In this disease too, the possibility of cross-reactivity with self hsp65 cannot be excluded (see below).

lmmunopathology resulting from T-cell cross-reactivity between exogenous and endogenous proteins The second concept is one of the most popular among immunologists today. Certain T cells, activated by hsps of the invading microorganisms, can cause autoimmunity by a cross-reaction with molecules in autologous tissues. These molecules may be either hsp or non-hsp molecules. A clear example of an animal autoimmune disease that may fit this concept is adjuvant arthritis, a disease inducible in Lewis rats by immunization with dead M tuberculosis in oil. T cells able to transfer the disease have been isolated from lymph node cells of tiected animals. These T cells recognize an epitope on the 65 kD of M tuberculos&nd cross-react with proteoglycans [ 24,251. After injecting rats with (FITC)-labeled, M-tiercul&presensitized T cells (and anticollagen IgG), the transferred presensitized T cells could be detected in the inflamed joints [26]. This tiding suggests that hsp65-reactive T cells, activated after immunization with M. tuberculo5iq migrate into the joints and cause joint destruction as a result of a proliferative response to proteoglycans. The adjuvant arthritis model initiated the extensive research of recent years into a possible role of M. tuberculasis and especially its hsp65 in the pathogenesis

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Autoimmunity

of rheumatoid arthritis (RA). Many investigators favor a model in which a cross-reactivity of T cells between the mycobacterial and the human hsp65 is central to the induction of joint inflammation. The data, which support a role of the mycobacterial hsp65, are briefly summarized. First, T-cell clones reactive to the mycobacterial hsp65 have been isolated from the synotial fluid of inflamed joints demonstrating the presence of these cells at the site of in.Uammation [ 27*,28,29*]. Second, in many studies it has been demonstrated that SFMNCs of most RA patients-in comparison with their PBMNCs-show strong responses to mycobacterial hsp65 and also to an acetone precipitate of M. tuberculosis (containing the hsp65) [30,31]. This suggests a homing to or an expansion of the hsp65-reactive T cells in the joints. Third, it was argued that a cross-reactivity with self hsp65 may be the cause of such a local expansion, especially because signilicant self expression of this molecule was demonstrated in the cartilage-pannus junction in rheumatoid joints and in rheumatoid nodules, but not in normal joints or normal or inflamed kidney or liver [32*]. Fourth, the set-a of RA patients contain high levels of IgG and IgA antibody to the recombinant mycobacterial hsp65 compared with the sera of control individuals [33]. Finally, the hsp65 can stimulate T cells to produce factors involved in joint destruction; it has been shown, in an in vitroco-culture system, that the mycobacterial hsp65 antigens can stimulate the mononuclear cells of RA patients to produce interleukins that inhibit the synthesis of human cartilage proteoglycan [34]. The data concerning a specific role for the mycobacterial hsp65 in RA are far from conclusive. Data that contradict a role for the hsp65 are substantial. The mere presence of T cells reactive to the hsp65 in the synovial fluid does not imply that these T cells pIay any role in joint inflammation. Indeed, as stated in the introduction, T cells reactive to the mycobacterial hsp and cross-reactive with self hsp65 are also present In the T-cell repertoire of individuals without any clinical signs of infection or autoim munity, and may even be beneficial to the host. An important indication of an involvement of a T-cell response to the mycobacterial hsp65 in RA would be an increased number of these T cells in the synovial fluid. However, a limiting dilution analysis to address this question has not yet been performed and we have found that the elevated responses of SFMNCs to the recombinant mycobacterial hsp65 are in most cases induced by contaminating antigens of the E. wli strain used to produce the recombimant hsp65 [35*]. Furthermore, no hsp65reactive clones were present among 30 M. tubercul&reactive TceU clones non-specifically derived from M. tubercul& stimulated synovial fluid T-cell lines of four RA patients and also no evidence was obtained for an imrnunodominant recognition of another ~4. tubercular&protein [36*]. Thus, unless suppression of hsp65 responses within the SFMNCs occurs, these results do not favor a selective expansion of hsp65reactive T cells in the synovial fluid. The hypothesis that a cross-reactivity of T cells between the mycobacterkrl and human hsp65 is crucial in RA is controversial, therefore, at least at the moment. Moreover, all mycobacterial hsp65-reactive T-cell clones obtained from

RA patients thus far are not restricted by DRl ad/or DR4 [27*,29*], the HIA antigens associated with an increased risk of developing RA, Of course, as in ReA the first concept may also play a role in RA. Particular M. tuberculosis proteins dilfused away from the site of infection may end up in the joint causing the local stimulation of M. tuberculostireactive T cells. Data suggesting that T-cell cross-reactivity between the mycobacterial and human hsp65 might play a role in juvenile RA are more firm. SFMNC of juvenile RA patients have been shown to respond to high performance liquid chromatography (HPLC) purified fractions of a human recombinant hsp65 preparation, whereas similar HPLC fractions of the E. coli vector did not induce proliferation. A significant correlation was found between the presence of reactivity to the human and mycobacterial hsp65, suggesting the possible recognition of cross-reactive epitopes on these molecules [37**].

lmmunopathology resulting from the T-cell response to endogenous hsp The third concept relates to autoreactivity of T cells to autologous hsp without the presensitization of those T cells by hsp of microorganisms. This concerns T cells responding to either conserved or non-conserved epitopes of self hsp. In the former case it is difficult to exclude the presensitization of such T cells by exogenous hsp as is shown by the development of insulin-dependent diabetes mellitus (IDDM) in non-obese diabetic (NOD) mice. This is a spontaneously occurring disease that becomes manifest at 4-6 months of age and is associated with T-cell reactivity to the mouse’s own hsp65 [38-l. A T-cell done reactive to an epitope on the human hsp65 is able to induce diabetes upon transfer to recipient mice [39=-l. Because this epitope differs in one amino acid from the mouse hsp65, this clone can probably also respond to the self hsp65. However, the epitope is on a conserved part of the hsp65 and the clone also responds to the mycobacterial hsp65, although to a lesser extent [ 390*]. The possibility remains, therefore, that an infection with mycobacteria triggers the response to self hsp65. Whether autoimmunity can be caused by responses of T cells to non-conserved parts of self hsp is still uncertain, because such T cells have never been demonstrated, although this has never been thoroughly investigated. Cohen and Young (40.1 have proposed that hsps may be immunodominant because they belong to a limited set of conserved proteins for which the immune response is already encoded by preformed networks of regulator cells. Furthermore, they suggest that antibodies speci& for conserved epitopes on hsps facilitate the uptake and presentation in B cells of both self and non-self hsps to T cells. In this case the bias of the immune response towards recognition of conserved self proteins also causes the immunodominance of exogenous hsp. If the immune response is focused upon recognition of self hsps one should also expect the existence of T cells recognizing self speciIic epitopes on hsps.

Heat shock proteins in immunopathology Res, Thole, de Vries

Conclusions Normally, the immune response to hsps probably serves to protect the host against infectious microorganisms (exogenous hsp) or against aberrant host cells (endogenous hsp). However, depending on factors, thus far unknown, but probably concerning HLA background and environmental stimuli, the host immune response may result in immunopathology or autoimmunity. Three hypothetical concepts are presented in this review with regard to the possible role of T-cell responses to hsps, in particular the hsp65 in these disease processes. Although in animal autoimmune diseases T-cell responses to exogenous or endogenous hsps have been shown to play a role in the pathogenesis of these diseases, the demonstration of such a role in human disorders still awaits fimher investigation.

Acknowledgements We are especially grateful to many investigators who shared with us unpublished data and insights, including I Cohen, H Gaston, S Knight, A Quayle and B Wilbrink. This work was supported by the Dutch League against Rheumatism (PR) and the Royal Netherlands Academy of AWLS and Science UT).

References and recommended reading Papers of special interest, published within the annual period of review. have been highlighted as: . of interest .. of outstanding interest 1.

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4.

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w B. KIE~SIJNC R , VAN E M B D E N JDA, THOU J E R , KUMARATNE DS, PISA P, WONDIMLI A, OI-I’ENHOFF THM: Induction of Antigen-Speci%c CD4 + HLA-DR-restricted Cytotoxic T Ceils as welI as NonspeciIic Nonrestricted KiIIer CeIIs by the Recombinant Mycobacterial 65-kDa Heat-shock Protein. Eur J Immunol 199Q, 20:369-377.

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KOGA T. WAND-WURTIFNBERGER A, DEBRUYN J, MUNK ME, SCHOEL B, KAUFMANN SHE: T Cells Against a Bacterial Heat Shock Rotein Recognize Stressed Macrophages. Science 1989, 245:1112-1115.

7. .

MUNK ME, SCHOEL B, MODROW S, KARR RW, YOUNG R& KAUFMU~M SHE: T Lymphocytes from Healthy InclIvIduals with Specifxcity to Self-Epitopes Shared by the Mycobacterial and Human 65XiIodaIton Heat Shock Rotein. J Immunol 1989, 143~2844-2849. Reports that in eight ouf of nine cases M. Mzrcrtlosis preactimted PBMNCs from healthy individuals displayed class II-restricted cytotoxic D

T lymphocyte activity against autologous targets primed with peptides of human hsp65. This is clear evidence that T cells which recognize a shared epitope between M trrbercufos& and human hsp65 are part of the T.ceU repertoire of normal individuals. 8. .

LLMB JR, BAL V, ROTHBARD JB, MJXHLERT & MENDEZ-SAMPERIO P, YOLING DB: The MycobacteriaI GroEI Stress Protein: a Common Target of T-cell Recognition in Infection and Autoimmunity. J Attfoimmrrnol 1989, 2 (suppf):93100. Documents that four T.cell lines from T cells of the ascitic effusion of a tuberculosis patient, were generated against diierent peptides of M. trrhcrctt/osis hsp65 and tested for their cross-reactivity with E. coli and human hsp65. Two T.cell lines responded to M. tuberculosis hsp65 specifically, two cross-reacted with E. coli hsp65, and one of these latter lines also responded to human hsp65. In addition to these T cells from disease sites, a T.cell clone isolated from the peripheral blood of a healthy individual was shown to be able to recognize a shared epitope on the M ~rrhercultis. E coli and human hsp65. 9. .

STEINHOF U, SCHOEL B. KAUF~&NN SHE: Lysis of Interferon-a Activated Schwann Cells by Cross-reactive CD8+ ap T CeIIs with Specificity for the Mycobactedal 65 kD Heat Shock Protein. In1 Immunol 1990, 3:27%284. This study demonstrates cross-reactivity of murine T cells between mycobacterial hsp65 ant. a shared autologous epitope. CD8+ class I-restricted cyrotoxic T lymphocytes, preactimted with fragments of mycobacterial hsp65, were shown to be able to lyse IFN-a-activated Schmnn cells. The presensitization by M. reprae hsp65 of T cells autoreactive fo nerve cells might be a phenomenon involved in tuberculoid leprosy. 10. . .

GASTON JSH, LIFE PJ, JENNER PJ, COISTON M J . BACON PA Recognition of a Mycobacterial-specific Epitope In the 65 Kilodalton Heat-Shock Protein by SynoviaI Fluid Derived T CeU Clones. / fip Med 1990, 171:831-834. Reports that T-cell clones reactive with an epitope contained within a 1-15 amino acid stretch of the mycobacterial hsp65 were isolated from .synovial Auid of a patient with an acute arthritis. This region shows no homology to the human hsp65, indicating that T cells reactive with a mycobacteria-specific epitope were present at the site of in%ammatIon in this patient. 11.

THOLE JER, VAN DER ZEE R: The 65 kD Antigen: Molecular Studies on a Ubiquitous Antigen. In hJo&cular Biology of My cobacleti [book] edited by McFadden D. London: Academic Press Ltd, 1990, pp 37-62.

12.

MINOTA S, CA~~ERON B, WEKH WJ, WLNF~ID JB: Autoantibodies to the Constitutive 73-kD Member of the hsp70 Family of Heat Shock Proteins in Systemic Lupus Etytbematosus. J Erp bJed 1988, 168:1475-1480.

13.

MJNOTA S, KOYASU S, YAHARA I, WINFIEIJJ J: AutoantibodIes to the Heat-Shock Protein hsp90 in Systemic Lupus EryXhematosus. J Clin IntIes/ 1988, 81:106-109.

14.

MLJLLER S, BREAMS JP. VAN REGENMOK~FL MH: Resence of Antibodies to Ubiquitin During the Autoimmune Response Associated with Systemic Lupus Erythematosus. Proc Nat1 Acad Sci US4 1988, 85:817&3180.

GRANFOW K, JALKANEN S. LUNDBERG AA, MARI-IKOIA 0, vON E~SEN R, LAHESMAA-RANTALA R, ISOMAKI 0, PEKKOLA-HEINO K, MERIM-PALO R, SAAIUO R, ET AL: Yerslniu Antigens in the SynoviaI Fluid from Patients with Reactive Arthritis. N Engl Med 1989, 320:21&221. Describes that synovial Huid cells ot patients with ReA induced by Ye&n&z infection stain positive with rabbit antisera and monoclonal antibodies to Yersinia en&nxo~oticu antigens. Synovlal Ruid ceUs of RA patients stain negative. 15. .

GRANSFOR~ K, JAUVWEN S, LUNDBERG & MAKI.IKoLA 0, vON ESSEN R, LUIESMAA.~AL~ Ft, SMIUO R, IsoMAla H , ARNOLD W. TONANEN k Salmonella LIpopoIysacharide in Synwial CeUs from Patients with Reactive Arthritis. Lancet 1990, i&5-688. Synotial cells from patients with ReA following Salmonella enh?riw&or Salmonella wimurium infection stained posit& with rabbit antisera against heat-killed S. enk?ritidLi and .S. &&bimuriuna, and with mono16. .

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Autoimmunity clonal antibodies to SafmoneNa lipopolysaccharide. The specificity of the presence of Sulmonel~ antigens in the joints of patients with a tifmoneffa~reacrive arthritis was demonstrated by the absence of staining in synotial cells from patients with RA or )‘ersi?lia reactive arthritis. 17.

GOON JSH, LIFE PF, GRANFORS K. MEIUHALTO-PALO R, BAIIEY L, CONSALVEY S, TONANEN A, BACON PA Synovial T Lymphocyte Recognition of Organisms which Trigger Reactive Arthritis. Cfin Exp Immunot 1989, 76:348-353.

GASTON JSH, LIFE PF, BAILEY LC, BACON PA: In vitro Responses to a 65XIIodalton MycobacteriaI Protein by Synovial T Cells thorn Inllammatory Arthritis Patients. J Immunol 1989, 143:2494-2500. Mononuclear cells from the synotial fluid and peripheral blood of patients with tierent arthropathies were tested for their proliferation in the presence of a recombinant M @rue hsp65. SFMNCS but not PBMNCs of most patients showed responses to this antigen and a role of hsp65 in inducing joint inflammation was proposed. 18.

.

19.

HERMANN E, FLEI~CHER B, MAYET WJ, Pow T, MEYER ZLJM, BUSCHE~IDE KH: Response of SynoviaI Fluid T CeU Clones to Yeniniu Entercolitica Antigens in Patients with Reactive Arthritis. Clin Exp Immunol 1989, 75:365-370.

20. . .

LIFE PF, BASSEY EOE, GASTON JSH: T Cell Recognition of Bacterial Heat Shock Proteins In Inflammatory Arthritis. Inr. munof Rev 1991, 121:113135. An excellent review, focusing mainly on reactive arthritis, and also containing data unpublished previously. One of the most important conclusions is that hsp65 is one of the antigens from the triggering bacteria recognized by synovial T cells from reactive Rek This is based on rhe recognition by SFMNCs of immunoblot fractions of the bacteria implicated in ReA and on the cross-reaction by synovial fluid T-cell clones responsive to the triggering bacteria with E coli hsp 65. STAGG AJ, HARDING B, HUGHES R, KEAT A , KNIGHT SC: Antigen Presenting Ceils Indicate the Causative Antigen in Reactive ArtIuitIs. Artbrih3 Rbeum 1990, 33:S106. Reports that cell lines responsive to ~~&m?y&a and purified protein derivative (PPD) were generated from SFMNCs and PBMNCs from patients with a Chhmydia-induced ReA Antigen-presenting cells from the synovial Euid but not from peripheral blood induced proliferation of the C&my&-specific ceU lines in the absence of added antigen. The PPD-speclIic cell lines were not stimulated by synovial fluid dendritic cells or peripheral blood dendritic cells, indicating that the SFDC have picked up chlamydti antigens in the joint.

27. .

MORRISON RP. BELLAND RJ, LYNC K, CAUIWELL

HD: ChIamydial Disease Pathogenesis; the 57-kD ChIamydIaI Hvoersensitivity Antigen isa Stress Response Antigen. / E$ Med 1989, 170:1271-1283. Documents the gene coding for C%rnydb hsp65. which was cloned and sequenced. T AYLOR HR. MACIEAN IW, BRUNHAM RC, PAL S, WHIIIUMHUDSON J: ChIamydial Heat Shock Proteins and Trachoma. Infixf Immun 1990, 58:3061-3063. cbhmydia hsp65 but nor hsp70 is an important antigen in inducing trachoma, which starts after an infection with chlamydiu Inoculation of hsp65 onto the eyes of cynomolgus monkeys, which had been sensitized by previous ocular chlamydial infection, resulted in the induction of an ocular delayed-type hypersensitivity reaction. The application of hsp70 had no effect. 23.

28.

VAN EDEN W, HOLOSHE! J, NEVO 2, FRENKEL 4 KIAJMAN A, C OHEN IR: Arthritis Induced by a T CelI Clone that Responds to Mycobacterium tuberculosis and to Proteoglycans. Proc Natl Acad Sci USA 1985, 82:5117-5120.

25.

VAN EDEN W, THOU? JER, VAN DER ZEE R, N~~RLXIJ A, VAN EML%DEN JDA, HENSEN EJ, COHEN IR: Cloning of the Mycobacterial Epitope Recognized by T Lymphocytes In idjuvant Arthritis. Nature 1988, 331:171-173.

26.

Q U I N N DEJOY S, FERGUSON-CHANOWKIZ K , O R O N S K Y AI, ZABRISKIE JJ3, KERWAR SS: Studies on the Horning of Mycobacterium-sensirized T Lymphocytes to the Synoviurn During Passiie Adjuvant Arthritis. Cell I m m u n o l 1990, 130:195-203.

HOLOSHI=

J, KON~NG F , COUGAN

JE. D

E

BR~I~N J, STROBER S:

Isolation of CD4-CD8-Mycobacteria-Reactive T Lymphocyte Clones from Rheumatoid Arthritis Synovial Fluid. Numre 1989, 339:226-229. 29. .

QCAME AJ, KJEU)SON-K~~GH JK, SHU.GUANG LI, OFIXING F, SHINMCK T. FOKKE 0. NATVIG JB: Rheumatoid Synovial Fluid Derived T’ CeU Clones Responsive to MycobacteriaI Antigens Including the 65 kD Heat Shock Protein. / Cei/ &c&em 1991, 15AS320. Clones were raised from synovial T cells of a DR;i.S+ RA patient. In the first cloning bacillus CalmenrCuerin (BCG) wz used as a srlecting stimulus. None of the BCG-reactive clones responded to hsp65. In the second cloning, hsp65 was used and T-cell clones reactive to this antigen were isolated. One of these clones MS tested in more detail and was found to recognize amino acids 241-255 of the mycobacrerial hsp65 in combination with DQ, 30.

BuR!.~Es~~R GR, ALTSIIDI. U, KAIDEN J R , EhlhlRlCH F : Stimulaforv Resoonse Towards the 65 kD Heat Shock Protein and Other Micobacterial Antigens in Patients with Rheumatoid Arthritis. / %eum 1991, 18:171-176.

31.

RES PCM, SCHAAR CG. BREED\‘EID FC. V A N E D E N W , FAN EMFIOEN JDA, COHEN IR, DE VIUES RRP: Synovial Fluid T CeU Reactivity Against the 65kD Heat Shock Protein of Mycohacteda in Early Chronic Arthritis. Lnncet 1988, ii:47%480.

32.

KARISSON-PARIU

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A, SODEPSIROM

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R, KL4R!ZSKOG L: Presence of Human 65 kD Heat Shock Protein (hsp) in Inflamed Joints and Subcutaneous Nodules of RA Patients. Stand I Immtcnol 1990, 31:283-288. A strong reactivity of a monoclonal antibody to human hsp65 was ohserved with the cartilage-pannus junction in rheumatoid joints and in rheumatoid nodules, but not in normal joints or in normal or inflamed kidney or liver. The increased expression of hsp65 in disease sires in RA suggests that this molecule may be involved in inducing T-cell reactivity at these sites. 33.

TSOULFA G, ROOK GA, BAHR GM, SATTAR MA, BEHBEHANI K, YOUNG DB, MEHLERT A , VAN EMBOEN JD, HAY FC, ISENBERG DA: Elevated IgG Antibody LcveIs fo the Mycobacterial 65. kDa Heat Shock Protien are Characteristic of Patients with Rheumatoid Arthritis. Stand J Immunol 1989, 30:519-527.

34.

WIIBRJNK B, VAN R OY JIA~~, HOLEWJN M, DEN O?TER W, BlJLsha WI. -. VAN EDEN W: The 65kD Heat Shock Protein can Ac&ate Rheumatoid Synovial FIuid Mononuclear Cells fo Inhibit Human Cartilage Roteoglycan Synthesis. Brj fieumatol 1991, in press.

. .

24.

MJ, BACON PA: Fine

Specificity of MycobacreriaI 65 kD Stress Protein (M65kD)Specific SynovIal T C e l l C l o n e s f r o m P a t i e n t s w i t h a Self Limiting and Persistent Arthritis. At’fhri(b /&urn 1990, 33:X6. T-cell clones reactive with the mycobacterial hsp65 were derived from the synovial fluid of a DR4+ RA patienr. AU clones responded to an epitope on the carboxy-terminal region of hsp65 and, unexpectedly. were DP-restricted.

21. .

22. .

GASTON JSH, LIFE PF, JENNER PJ. COLSTON

35. .

R ES PCM, TELGT D, VAN LANl JM, OUDKERK P OOL, BREEOVEID FC. DE VRIFS RRP: High Antigen Reactivity ln Mononculear CeIIs from Sites of Chronic Inflammation. tincei 1990, i:1406-1409. Reports that SFMNCs of most RA patients display enhanced proliferative resoonses to M. tuberculosis comoared with the PBMNCs of the same patients. This study demonstrates’the following: elevated responses of SFMNCs are not speci6r~Uy directed fo M. tuberculti but are also observed when E coli is used; enhanced antigen reactivity may be a geneml feature of cells from sites of in&rnmation, because mononuclear cells from patients with a pleural effusion resulting from nmplastic involvement also display stronger responses to M. tiercufosisand E coli than the corresponding PBMNC. 36. .

RES PCM, ORSINI DLM, VAN IAAR JM, JAN~ON AAM, ABOU-ZEUI C, DE VRlEs RRP: Diversity in Antigen Recognition by Mycobacterium Tuberculosis Reactive T Cell Clones from the Synovial Fluid of Rheumatoid Arthritis Patients. Eur J Immunol 1991, 21:1297-1302.

Heat shock proteins in immunopathology Res, Thole, de Vries T cells were non.specificaUy cloned from T-cell Lines of four RA patients indu& by a single stimulation of SFMNCs with &f. I~herc~tlwis and subsequent outgrowth in 1L.2. The M. tuberculosis-rextivc T-cell clones were tested for their response to hsp65 and immunohlot sep arated fractions of I%I lr&ercr~Iac-is None of the clones respondcui to hsp65. In addition, on the basis of the recognition pattern of the immunohlot fractions no evidence w z found for a dominant recognition of another M rr~ercularis protein. This argues ag.Gnst a selective homing or expansion of certain types of AI. cLlbrrcftln~is.reactivc T cells in the joints of RA patiems, ix. after a crossrt-dction with an autoantigen. 37. . .

De Gtbum-Mt:rmia DE, VAN DI~R ZEE R, RlJKkXS GT. SCHLIlIHhtAN H.J. BIJIS~IA JWJ, ZEGERS BJM. Vrw EDEN W: Recognition of

Human 6OkD Hear Shock Protein by Mononuclear Cells from Patients with Juvenile Chronic Arthritis. Lnnccl 1991. 337:1369-1373. The SFMNCs and PBMNCs of juvenile RA patients display prolifentive responses to HPLC-purified frdctions of rcx~mhinant human hsp65, mycobacrrrial hsp65, and mycohacterial hsp70. A correlation between the response to human and mycobacterial hsp65 wa,, reportcif. No response to the human hsp65 fractions in SFMNCs from RA patients and PBMNCs from healthy children and young adults was measured. 38.

EIJA~

D, Mm~ovrrz D. Rlisl-IEI:

T, VAN Dm ZEE R, COHEN IR: In-

in the pathogenesis of IDDM wax demonstrated by the finding that antihsp65 T cells can accelerate the induction of IDDM in young NOD mice. 39.

ELLS

D , RESHEF T. BIRK OS, VAN D ER ZEE R, WAUCER MD: Vaccination Against Autoimmune Mouse Diabetes with a T Cell Epitope of the Human 6%kDa Heat Shock Rotein. Proc Nail Acud Sci 1M 1990, 87:30~3031. The diabetescausing Txell clones described in the previous study, which were raised against ,Cf. izrherculasis hsp65, respond even better to the human hsp65. The epitope recognized on the human hsp65 is contained within amino acids 4371160 (peptide ~277). This epitope differs in only one amino acid from the mouse’s own hsp65, indicating the autoreactive nature of these clones. Both the anti-hsp65 T-cell clones as the epitope containing peptide ~277 can be used as vaccines to induce resistance to IDDM. . .

40.

COHEN lR, YOUNG DB: The Immune System’s View of Invad-

ing Microorganisms, Autoimmunity and the Immunological Homunculus. /nm~rrt?ol Tc&j~ 1991, 12:105-110. In this review article the authors propose the immunological homunculus tls a model to explain the apparent immunodominance of certain self antigens and its consequence: the uniformity of autoimmune disease. The homonculus is a picture of the immunological self and con. sists of a sysrem of preformed cellular networks that regulate the response to a limited set of conserved proteins. l

. .

ducrion and Therapy of Autoimmune Diabetes in the Nonobese Diabetic (NOD/Lt) Mouse by a 65.kDa Heat Shock Prorein. &ix Null Acad SC; [ISA 1970. 87:157615BO. This study demonstrates the crucial rote of hsp65 in the pathogenesis of spontaneously occuring IDDM in NOD mice. The onset of this disease is associated with T-cell and antibody responses to a self antigen crossreactive with mycobacterial hsp65. In addition, the crucial role of hsp65

PCM Res, JER Thole. RRP de Vries. Department of lmmunohematology and Blood bank, Building 1, E3-Q, University Hospital, PO Box 9600. NL.2300, Leiden, The Netherlands.

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