MHC antigens: new methods in tissue typing

MHC antigens: new methods in tissue typing K.I. Department

of Molecular Current

Welsh

Immunogenetics,

Opinion

in Immunology

Guys Hospital, 1989,

London, UK

1:117&1183

Introduction

a person responds duced responses,

The magnitude of the influence of the major histo compatibility complex (MHC) on allograft survival has example been hotly disputed over the years. .4 god is provided by the question of the influence of MHC matching on cadaver renal allograft survival figures. Here. even within single groups, sequential publications have not always agreed (Pate1 et al, N En@ J Med 1968, 2?9:501-502; Mickey et al, TissueAntigens 1971, 1:57-60; Opelz et al, Trawpkznt Proc 1974, 17:371-382). A vet) sophisticated summary of the relevant literature up until 1986 is presented in the major review of the influence of the MHC on all forms of biology, including transplantation, in the book by Klein (Natural History of the MHC, John Wiley, 1986). In the last 2 or 3 years, the techniques used in tissue typing laboratories, which had remained essentially unchanged since the introduction of the Terasaki plate (Terasaki and McClelland, Nature 1964, 204998) have been revolutionized by the appli cation of DNA and magnetic bead technologies and by the advent of user-friendly ‘flow cytometers’. My purpose here is to outline the effects of the application of these and other new techniques on current thinking about the intluence of the MHC on allograft selection, function and survival. In recent months. over 1000 papers from the world’s leading laboratories involved in transplant-related research have appeared in a single volume (Proceed ings of the XII International Congress of the Transplan tation Society, Trarqblmzt Proc 1981, vol 21). A further 500 appear in the conference abstracts (Abstracts of the XII International Congress of the Transplantation Societ);. 1988). These are well worth reading for a state-of-the-art

bath form of transplantation is diIlerent as regards the relevance of each of the above. It is also important to remember that studies of rodent models, especially rat, mainly consider the cellular side of rejection, due to the limited damage which can be mediated by antibody in these systems (Fig. 1).

U@W

Tissue typing has four connected functions relating tc) the MHC and transplantation: ( 1 ) it defines the human leukocyte antigen (HIA) antigen type for the matching of donor and recipient organs--MHC matching; (2 ) it de fines which antigens a recipient cannot be given because of previous responses induced by pregnancies, transfu sions, transplants, etc. and hence indicates which mis matches may safely be given-windows; (3) it checks at the time of transplantation that no humoral response is present against a donor MHC antigen-ross-match ing and. (4 ) it defines when and to which MHC antigens

to after a transplant-transplant-in

Renal allografting MHC

matching

For sibling transplants, short and long-term graft survival is best obtained by using MHC-identical siblings. There is no clear-cut distinction between the use of donors which share no or one haplotype with the recipient (Cicciarelli et al, Tran.@antution 1987, 43:636-640). Such transplants have long- and short-term survival times, better, though not significantly so, than cadaver allografts (Cicciarelli et al., 1987). For cadaver renal allografts, a clear distinction is now emerging between transplants matched fully for B and DR and for all other groups, at least in transfused recipients receiving cyclosporin A as the major immunosuppressive agent (Cicciarelli et al., 1987). There is no con sensus hierarchy outside this definition. It is important to remember, in the field of HLA matching. especially in work from ‘vested interest’ centres, that there may be five or six papers, each with a slightly different emphasis, but all based on the same multicentre data and emanating from the same group. In reaching this conclusion, I have pooled data relating to kidney allografts into transfused patients where cyclosporin A is used as the major im munosuppressive agent For grafts into first-time recipients most groups are rt’ porting around 85% l- and 2-year graft survival figures and matching (even on the above definition) is less important in the best centres [ 1 ] For grafts into sensitized recipients, the best groups are reporting around 80% land 2-year graft survival figures and matching is less imp portant in these centres. A simple definition of ‘optimal matching’ which applies to both live-related and cadaver allografts as well as to grafts

Abbreviations HLA

1178

human

leukocvtt

@

antigen; MHC

Current

Science

major

hlstocompatibillty

Ltd ISSN 0952-7915

complex

MHC

antigens:

?? Sensitization (previous graft) Blood transfusions HLA-B and -DR identity Lack of drug side effects

?? Multiple transfusions leading to a high titre of anti-HLA antibodies ?? Age

new methods

in tissue

typing

Welsh

Fig. 1. Recipient (a) and donor (b) factors affecting success in kidney transplantation. Donor and recipient factors are connected. For example, human leukocyte antigen (HLA) antibodies and long cold ischaemic times carry synergistic weight (Terasaki, !mmuno/ Lett 1989, 21:33-39). Courtesy of Drs Welsh, Roitt, Brostoff and Male Cower Medical Publishing.

and

?? Other diseases Compliance

?? Anatomical abnormality

?? No anatomical or pathological abnormalities

?? Warm ischaemia time > 45 min

e

?? Cold ischaemia time >24h

Clinical evidence of good renal function before donation

?? Donors aged S-50 years

?? Evidence of infection

into sensitized recipients appears to be ‘donor-recipient combinations matched fully for HA-B and DR antigens’.

Windows These

apply to grafts into sensitized patients only. In a patient with antibodies to many antigens, it is often diEicult to define the specificity of the response. Definition does aliow successful transplantation with any antigen to which there has not been a response [Z]. This applies both to antigens putatively exposed to, but not rtlsponded to, from previous untyped blood transfusions and to antigens definitety exposed to, but not responded to, from previous tmnsplants [3]. It also allows successful transplantation with any antigen to which a response has been made-providing: ( 1) that the response arose from transfusion and not pregnancy or transplant, and (2) that the response had abated at least 6 months previous@

Cross-matching

The introduction of Bow cytometry has made a large impact here. It is good practice to use a cross-match technique which is more sensitive than the standard NIH or Kissmeyer-Neilsson technique for transplants into sensitized patients (Cook it d, C(in T~rz.@a?zt. 1987, 3:253-256). For transplants into non-sensitized patients, false positives can occur (Cook et aL, 1987). Flow cytometry; and the cheaper and more widely used (at least in the US) anti-immunoglobulin technique both detect anti-MHC Tantibodies with greater sensitivity than the colder techniques. A more rapid technique has recently been described (Vartdal et al., Tissue Antigens 1986, 28:301-3321, which is more sensitive than rhe Kissmeyer-Neilsson technique and which involves using monoclonal antibody-coated beads to separate target cells from donor blood The technique involves the coupfing of a monoclonal antibody directed against a defined

1179

1180

Transplantation

cell population (e.g. B cells) to a magnetic bead. These cells can then be separated out within minutes directly from whole blood. Beads with cells attached can be used directly in complement-mediated tests with tissue typing or recipient sera, provided that a two-colour fluorescence test is used as measure of cell death (Vartdal et al, 19861.

Cross-matching

is less relevant to bone marrow transplantation simply because donor and recipient are so well matched. There is, however, one clear report of a cross-match-positive transplant doing badly [8].

This

Transplant-induced Transplant-induced

responses

In the pact, the literature generally reported that posttransplant antibody responses directed against donor MHC antigens had little influence on graft function or survival, but two recent reports have questioned this (Mar tin et al, Tran.splantation 1987, 44:5&57; Palmer et al., Tramphnt Proc 1989, 21:76&?6~). This change may be due to the relative inefficiency of cyclosporin A at blocking antibody responses (especially secondary ones> compared with its potency in suppressing cell-mediated rejection. A greater proportion of rejection episodes may now be considered to have an antibody involvement. New techniques for determining the number of T cells which can be specifically activated to target class I and class II donor MHC antigens (precursor T cell frequencies; Sharrock et aC, Tran.pkzntution 1987, 43:69!+703) [4], and for determining the number of cells which can be speciIically activated by donor MHC antigens (precursor helper T cell frequencies; Orosz et aL, Transpkmtation 1987, 43:71%724), and the number of specific suppressor cells [5] are available and are beginning to be applied both to pre- (see bone marrow and heart sections) and post-transplant monitoring.

responses

Here the anti-MHC response post-grafting can be either way round, i.e. it may be host-versus-graft or graftversushost, and many consider the latter to be the more important. It has long been known that a proportion of the graft-versus-host response is mediated by host cells. In patients treated with cyclosporin A, at least, host cell reactions with self-class II appear to be part of the graft versus-host reaction [ 91.

liver MHC

transplantation matching

kidney transplants, early reports were contra dictory, but two recent reports have shown that HI-4 matching is associated with shorter graft survival times (Markus et al In Transplantation Society Abstracts, 1988, p 174; Goldstein et al In Transplantation Society Abstracts, 1988, p 175). The locus responsible for this phenomenon appears to be the DQ locus. DNA technology is necessary for accurate typing of the DQ (and indeed the DP locus) so earlier reports carry less weight.

As with

Windows

Not applied at present. Bone marrow MHC

transplantation

matching

It is recognized by most groups that HLA matching is very much more critical for bone marrow transplantation than it is for renal allografting, with trials of mismatched recipients often showing mediocre results (Cahn et al., Br J Haemutoll988, 69:345-350). New restriction fragment length polymorphism techniques, especially those for class II MHC typing, have therefore been widely applied [6,7] to obtain better matches outside families. The problem with the application of these techniques, especially when DP typing is included, is that identity is only found in identical twins or a minute proportion of ran dom donors, Drawing the line of what constitutes ‘optimal matching’ is still under discussion.

Windows

Precursor T cell frequency analysis has been applied (mainly retrospectively so far) to determine how many recipient cells can be activated to kill cells from the donor. Initial results are encouraging and the technique will almost certainly be applied to other forms of transplanta tion [4,5],

Cross-matching

Positive anti-class I or anti-class II cross-matches do not result in hyperacute rejection of liver allografts. This led to many groups ignoring the cross-match, but sera were collected and analysed retrospectively with respect to the outcome. A consensus indicates that transplantation against a positive class I cross-match leads to poorer graft survival figures. A possible reason why livers do not suffer from hyperacute rejection has been suggested (Pollard et al., Trarwplant Proc 1989, 21:425-426). Although a soluble antigen will complex with antibody, and could prevent hyperacute rejection, it is unlikely to af feet T cells. Furthermore, a significant amount of blood is given ‘on the table’ at the time of transplant and recipients with a high panel reactivity require most blood [lo]. Serum from liver-grafted animals is, however, known to mediate a tolerogenic effect in a strain-specilic manner (Ramada, Experimental Liver Transplantation, CRC Press, 1988) and the major candidate is circulating MHC (Summoto et al, Tramphzt Proc 1989, 21r431-432). One would suspect that aggregation or complex formation is responsible for this effect. Transfected cells have been used to tackle this problem once more in the kidney [ 111.

MHC antigens: new methods in tissue typing Welsh Post-transplant monitoring

Cross-matching

In liver transplantation, HIA antibodies produced post-

Anti-HLA antibodies can cause hyperacute rejection of heart allografts although higher titres are required for hearts than for kidneys. For reasons of speed, most transplants are carried out without using the cross-match test but the advent of the faster magnetic bead tests may change this (Vartdal et al., 1986).

transplant are also a poor prognostic indicator (Hemade et al. In Transplantation Society Abstracts, 1988, p 377) and may be linked to the occurrence of vanishing bile duct syndrome. In animal models, donor-specific effector and suppressor cell numbers are beginning to be monitored [5]. One report (121 suggests that there are two types of rejection, one linked to MHC compatibility and one to incompatibility This is a particularly fascinating possibility in the light of the studies which claim that matching is a contraindication for liver allograft function.

Post-transplant monitoring As with livers and kidneys, the production of anti-HLA

antibodies post-transplantation is associated with poor prognosis.

Hearts

Pancreas

MHC matching

There is no defined pattern emerging for pancreas-only transplants. For the majority, which go into renal allograft recipients, the rules are the same as those which apply to renal allografts. The problem of whether depletion of certain cell types from the donor pancreas or donor islet preparation will aid engraftment is still open [ 151.

Available evidence suggests that the situation for hearts may be very sirnllar to that described for kidneys, i.e. organs matched for recipient B and DR do better than less well-matched groups [ 131 (Opelz et al. In Transplanta tion Society Abstracts, 1988, p 228). In animal models, recipient MHC expression and genetic background are as important as the number of mismatches [ 141.

Corneas Early suggestions of the importance of HLA-A,-B and -C matching are in doubt, especially in view of the sparcity

Windows

Not applied at present.

Table 1. Human

organ transplantation:

the influence

Kidney

Optimal

Liver

of the major

Heart

histocompatibility

complex.

Bone marrow

Pancreas

Cornea

*(-)

‘(“)

*(‘j

*(***I

?? *(*)

“(“‘I

‘(‘)

*(**)

***(***I

*(‘I

‘(“)

matching

*(,**I

A,B,C B and DR

‘(-1 . ..(_)

“‘(“)

A,B,C and DR

*Y**)

‘(‘I

*(*)

n.a.

na.

n.a.

‘(‘I

na.

n.a.

n.a.

“(n.a.r

na.

na.

+

***I*-~

c)

n.a.

***(**i

(~ I

n.a.

.i**,

Windows “*!‘)

B cell-defined T cell-defined

n.a.

Cross-matchrng

)

**j

“‘(“‘)

“(‘I

Antibody “( -)

Post-transplant

Increased

MHC

expression

+Cross-matching

for bone

as the tieid moves for example, low matching complex; -~

away

**(-)

?? !“*\, marrow

*(‘*j

matches,

etc. *, Indicates

n.a.. not applied:

*(‘*j

transplantation

irom perfect

groups who do not HLA-match protocols

“(-1

not thought

is not necessary antibodies

need

relative

I I

in cases where

WIII become

more sophisticated

importance,

too important

more

donor

Important

‘windowing’

with previous

‘(‘1

and recipient

i81. Thus table

procedures.

values in brackets.

at the time point indicated.

are absolutely IS meant

Different MHC,

drug regimes

major

identical,

but

as a sample summary. counteract

histocompatibility

1181

1182

Transplantation

of MHC expression on this organ [ 161. Arguments that vascularized beds might cause up-regulation of the MHC are probably true, but it has recently been shown that even such high-risk recipients may have transplants with out regard to MHC matching [ 171.

with extracted donor histocompatibilty antigen and a brief course of cyclosporine. Trun.pkznC Pm 1989, 21:2&251. AUoantigen-speciiic T e&&or cell, and T suppressor cell frequencies were measured before and after heart aUo*g in rats. Basically, effector celI frequencies decrease and suppressor ceU frequencies increase. A new technique, simple and weU used

Table 1 lists the relative importance of many of the points discussed above and indicates whether or not they have increased or decreased in consensus import over the past few years. In liver transplantation, it is the good match which does badly..

6. 0

Increased MHC expression post-grafting (e.g. on biopsy or fine needle aspirates) has not been discussed here. This is because, after a plethera of early papers, there have been few recent reports (Leszczynski et al, Kidney Int 1987, 31:1311-1316), perhaps suggesting that better ways are evolving to assess rejection episodes or that the persistence of the expression after the event confuses the picture. Currently, there does appear to be much interest in the link between class II expression and grafwersushost disease [ 91.

NOREENHJ, DAMDSONML, MCCUUOUGH J, BACH FH, SEGW M: HL4 class-II antigen typing by restriction went length polymorphism (RFLP) in unrelated bone marrow transplantation patients. Tyan.pkznt Pm 1989, 21:296%2970. This is not the Iirst attempt to define a restriction fragment length poly morphism test with the correct degree of discrimination for bone marrow matching, but enough of the early references and data are cited to justify the enzyme and probes used. The definition of an ‘optimum match’ for a bone marrow transplant will depend on this type of approach. 7

SHIMOYAMAH, HORI~~I T, OKABAYA~HIT, CHONO S, TANAKA I, MryAKE N, HAYASHID, ORITA K, INOKO H, ANDO A, TSUJI in K: DNA typing of HLA class II antigens is informative kidney transplantation. Tyan.pkznt Pm 1989, 21:64%648. Contains little proof, but describes the interesting idea that one simply uses specilic enzyme probe combinations and defines incompatibility at each locus as the number of bands which are ditferent. The delinition of an ‘optimum match’ for a bone marrow transplant will depend on this type of approach. 0

8.

Annotated references

and recommended

reading ??

00

Of interest Of outstanding

interest

1. 0

CARPENI-ERCB, GOGUEN JJ?, BRAIXY JW, TURKA & CHO Sl, MILFORDEL HLA-B/DR matching and cadaver renal aUograft survival in New England. Tyun.pkznt Pm 1989, 21:66%664. Describes kidney graft survival figures in a small area and relates them to MHC matching. A clear indication of the renal aUograft sutviti figures obtainable in good centres. 2. .

CL%S FHJ,

@BELT

PER~IJN GG,

VANROD JJ:

Y, VONVEZEN A, DEWAAL LP, DAMAROJ. Selecton of cross-match negative HIA-A and/or HWB mismatched donors for highly sensitized patients. Tran.@anf PYOC 1989, 21665-666. Describes a sophisticated method of defining antigen mismatches which are safe to transplant across in highly sensitized recipients. Too many grafts are lost through taking risks in highly sensitized patients. 3.

WEISH

??

RAFI-ERY MJ, RIGDEN S:

KI, VAN DAM M, BEWICK ME,

KOFFMAN GK, TAUBE DH, transplantation of kidneys bearing previously mismatched HLA A and B locus antigens. Trun.p&znt Int 1988, 1:19C-195. Describes a sophisticated screening policy which, with information from previous grafts, can deline antigen mismatches which are safe. Also gives data from individual patients for the expert reader to per \ur Successful

KAMINSKIE, HOWS J, HROOKES P. MACKINNON S, HI!GHES T. AVAKIAN0, SH,UR~K C, Ciulh~A~ J, WATCHEIDR JR: tioreactive cytotoxic T-cell frequency analysis and HLA matching for bone marrow transplants from HLA matched unrelated donors. Tranqkmt Proc 1989, 2 I :29X%2977. A very neat demonstration of the use of the technique first described for humans by Sharrock et al. ( Trun@kzntation 1987, 43:699-703) which allows cytotoxic T lymphocyte precursor frequencies to be determined Cytotoxic T ceU precursor frequency analysis appears to be better than the MHC test in assessing the suitability of HLA-mismatched bone marrow donors. The authors described a new and important technique and the possibility of assessing T cells in the way we titre antibodies no% exisb L1. 0

i .

1X1 ‘I. bn:I’KoWsKi b.jl. hztl..L\ B(L) Decreased frequent? of alloantigen-specific T-cells following immunosuppression

S~ORNIKJC,

ELFENBEIN G,

G~-PoLE

J, G~EDERT

T,

GROSS

Role of anti-donor antibodies in bone marrow transplant rejection evaluation by flow cytometry and effect of plasma exchanges. Tyan.pkznt Proc 1989, 2 1:2974-2975. This paper, taken together with previous reports, indicates that positive cross-matches of the kind carried out in renal transplantation should be applied to bone marrow transplantation if any evidence of sensitization exists Cross-matching for specific bone marrow transplants should be considered. ??

S, WEINER RS:

SLOANEJP, ELLIOTI CJ, POWLESR HLA-DR expression in epidermal keratinocytes after allogeneic bone marrow tram+ plantation: relationship to histology, rash, marrow purg ing, and systemic graft-versus-host disease. Tyan.pkmkation 1988, 46:M3. Immunohistology studies on skin biopsies from recipients (mainly acute myeloid leukaemia) of allogeneic bone marrow showed Kerr atinocyte class II expression correlated with the presence of subsequent development of systemic graft-versus-host disease Such expression was not observed in patients who received marrow depleted of T iympho cytes. Class II expression and graft-versus-host disease is a developing lield. 9.

??

10.

MARINO IR, WEBER

0

DUQUESNOY

RI

T,

HLA

KANG

YG,

Esqun%~

CO,

STARZL TE,

alloimmunization and blood requireliver transplantation. Tyun@ant Proc

ments in orthotopic 1989, 21~789791. HIA antibodies present at the time of transplant requirement. Novel, but obvious once you know.

increase

the bkx>d

MADSENJC, Woon KJ. SUPERINA Kk MORRIS PJ: Induction of immunological unresponsiveness using recipient cells transfected with donaor class-II MHC genes. Trumpkmt Proc1989. 21:477. ‘The use of modem transfectlon technolo8y to tackle the antigen in duced unresponsiveness problem again. Interestingly narrow range of doses which mediate unresponsiveness. One of a new breed of papers using molecular techniques to re-appraise unanswered questions. 11 .

G~JBEKNATIS G, KEMNIIT J, TLXH G. RINGE B, BUNZENDAHL H, RIEDEL T, M~JLLERR, PICHLMAYRR: Different features of acute liver allograft rejecton: their outcome and possible relationship to HLA compatibility. Tran.@hnt Proc 1989. 21:2213-2216. Small numbers, and not conclusive (nor claimed to be by the authors), but nevertheless interesting data showing different types of rejection .tre linked to different types of MI~C class II mismatch Type I occurs in cabes of DK compaahility and type II occurs in astxclation bith class Ii 12

??

MHC antigens.: new methods in tissue typing Welsh matching. Can explain why hWC matching can be associated liver graft suti figures.

with poor

KERMANRH, VAN BURENCT, LEWISRM, F’RAZIER OH, COOL!ZY D, KAHAN BD: The impact of HL4 A, B, and DR blood transfusions and immune responder status on cardiac allograft recipients treated with cyclosporine. Tran.+ntatim 1988, 45:335337. Shows in a single-centre study that DR matching is not important for heart aliogmfting unless the recipient has not been transf&ed. Reinforces the idea that heart recipients should be transfused. 13.

0

14. 0

STEPKOWSKI SM, R&L&AHMAD A, DUNCANWR: The role of class

I and class II MHC antigens in the rejection of vascularized heart allografts in mice. Tran.g&antation 1987, 44:753-758. Compares rejection times between strains mismatched for single and multiple class I and class II antigens. Concludes, as have studies in other systems, that MHC disparities fail to predict either the type or rate of heart gratI rejection in any particular donor-recipient combination. They suggest that an important factor is that MHC expression on donor tissue is neither strain-independent nor static within a strain. Such differences can tiect both induction and effector arms of the immune response. Separate involvement of immune response genes are also postulated. A thorough demonstration that the influence of the MHC on allograft rejection is multifactorial. 15. 0

SPILUNAAF-BILGEN EJ, BO~WMANE, DEBRUIN RWF, MARQUET RL, JEEKEL J: The effect of reduction of class-II antigens on

the survival of pancreatic aUografts. Tranqkznt Proc 1989. 21:51-%516. There is good evidence in some animal transplant models that masking or removal of class II antigen-bearing cells causes alIograft prolongation. This paper describes a rat system invoking pancreatic transplantation where there is no effect or either masking or removal of class II. A dissenting voice showing that the consensus tiew is not applicable in alI strain combinations. 16. 0

AFGIANNA D, WAKEFIELD D, WATKINSS:

HLA antigens in ocular tissues: I. In viva expression in human eyes. Tramplantation 1988, 45:610-613. Human ocular tissues, with the exception of conjunctival epithelium, do not normally express class I or class II HL4 antigens. This finding suggests that the cornea is an ‘immunologically privileged site’ unlikely fo suffer rejection unless a separate inJ%unmatory process causes ups regulation of MHC products on additional cells.

17.

MILLER K, HUBERC, NIEDERWIESER D, G&-I’INGERw:

Successful

engraftment of high-risk comeal allografts with short-term immunosuppression with cyclosporine A Trampkzntatio?z 1988, 45:651652. Independent of MHC match, high-risk (i.e. strongly vascularized) host corneas can be replaced successfully using the technique and immunosuppressive protocol described. Shows MHC matching is not important for comeal transplants. 0

1183