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Mechanisms and Role of HLA and non-HLA Alloantibodies

Transplantation Research Center, Brigham and Women’s Hospital, Boston, Massachusetts

Address correspondence to: Dr. Kathryn J. Tinckam, Brigham and Women’s Hospital, Tissue Typing Lab, PBB 161, 75 Francis Street, Boston, MA 02115. Phone: 617-732-8562; Fax: 617-264-5104; E-mail: ktinckam{at}rics.bwh.harvard.edu

	Abstract

Top Abstract Introduction Antigen Targets T Cell-B Cell Interactions... Antibody-Mediated Damage Detection of Antibodies in... Antibody-Mediated Syndromes Accommodation Conclusion References

 
The role of alloantibodies against HLA and non-HLA targets is becoming increasingly recognized as critical in the pathogenesis of acute and chronic renal allograft outcomes. This review discusses the antigenic targets, the mechanisms of T and B cell activation that result in the production of antibody, the complement cascade, methods of antibody detection, and the evidence that alloantibody-mediated mechanisms are active in acute and chronic rejection.

	Introduction

Top Abstract Introduction Antigen Targets T Cell-B Cell Interactions... Antibody-Mediated Damage Detection of Antibodies in... Antibody-Mediated Syndromes Accommodation Conclusion References

 
For the vast majority of the 51 yr since the first kidney transplant, T cell–mediated inflammation was believed to be the central process in allograft rejection. Therapies to prevent and treat allograft rejection consequently have been directed primarily against T cells. Improvements in these drugs have led to greatly improved rates of acute cellular rejection and 1-yr graft survival; however, acute rejection does still occur, as does long-term chronic rejection. It was the development of the immunohistochemical process for visualization of complement split product C4d in graft tissue that first provided concrete evidence linking antibody binding and complement activation in renal allografts to the mechanism by which damage occurs in this setting (1). We now recognize that alloantibodies play a role in rejections that do not respond to T cell therapies and, indeed, require targeted therapies that address the various mechanisms by which they exert their effects. Newer, more sensitive technologies for serum antibody screening are allowing for clearer delineation of the relationship between antibodies and acute and chronic allograft pathologies and their attendant clinical outcomes. This review discusses the antigenic targets of the humoral alloimmune response, the mechanism of antibody generation, the pathophysiology of antibody-mediated cell damage, the phenomenon of accommodation, and overview of the current understanding and classification of antibody-mediated syndromes, including the evidence that antibodies are active in these clinical syndromes and the presently available therapies.

	Antigen Targets

Top Abstract Introduction Antigen Targets T Cell-B Cell Interactions... Antibody-Mediated Damage Detection of Antibodies in... Antibody-Mediated Syndromes Accommodation Conclusion References

 
Alloantibodies that are of key interest in transplantation are those that principally are directed against the MHC molecules (also known as HLA), which are responsible for presentation of foreign antigen to T cells. Donor MHC may act as a direct antigenic target (direct allorecognition) or first be processed by recipient antigen-presenting cells (APC) for subsequent presentation to recipient T cells (indirect allorecognition). In addition, autoantibodies and antibodies to minor histocompatibility antigens increasingly are acknowledged as potential targets of the humoral alloimmune response. The most ubiquitous antigens to which the population is sensitized are the ABO blood group antigens (2,3); on the basis of population frequencies in the United States, the chance that any two individuals will be ABO incompatible is 35%.

MHC class I molecules (known as A, B, and C antigens) are found on the surface of all nucleated cells in the body, of which endothelial cells are of particular significance in transplantation. MHC class II molecule (DR and DQ antigen) expression is limited to the surface of B cells, APC, and microvascular endothelial cells. MHC molecules are extremely polymorphic, with more than 1600 different alleles presently documented in humans. This property increases the chances of sensitization (development of alloantibodies), which can happen upon exposure to nonself MHC or other nonself antigens (commonly through blood transfusion, pregnancy, or previous transplantation).

In addition to these major histocompatibility antigens, a large number of minor histocompatibility antigens have been recognized. These minor histocompatibility antigens first were defined as significant during rapid rejection in mouse skin graft models and have been shown further to cause endothelial cell apoptosis in other animal studies (4,5). Although the significance of minor histocompatibility antigens in humans is less well defined, antibodies against nonclassical MHC molecules, such as MHC class I polypeptide-related sequences A (MICA) and B (MICB) have been implicated with acute renal allograft rejection and loss (6,7).

Autoantibodies also may be important but are similarly incompletely described at this time. An intriguing recent study demonstrated an association between an autoantibody specific for angiotensin II type 1 receptor and hypertension, fibrinoid necrosis, and acute renal allograft dysfunction (8). Antivimentin and antimyosin antibodies have been shown in cardiac transplantation to relate to long-term allograft survival (9,10), and antibodies against collagen and percalan are associated with chronic renal rejection in animal studies (11).

The ABO blood group antigens are carbohydrate moieties on glycolipids that are present on the surface of most tissues, including endothelium and erythrocytes. Specific antibodies against the nonexpressed antigens arise within the first few months of life in response to environmental antigens.

	T Cell–B Cell Interactions and Antibody Production

Top Abstract Introduction Antigen Targets T Cell-B Cell Interactions... Antibody-Mediated Damage Detection of Antibodies in... Antibody-Mediated Syndromes Accommodation Conclusion References

 
High-affinity antibody production by B cells to a particular target is dependent on sufficient help from antigen specific T cells. Furthermore, the T and B cells that respond to a new antigen must be in close proximity to each other within the lymphoid organs and must be activated from their naïve state before they can interact to produce an effector response. When an APC (usually a dendritic cell) moves to the lymph node (or spleen) to interact with these effector cells, a series of events ensue to meet this geographic requirement. Upon antigen presentation to a T cell, a change in the T cell shape occurs, consistent with impending diapedesis (12), and the chemokine receptor CXCR5 is upregulated (13). Subsequently, in response to the chemokine CXC13, located in the primary follicles where B cells reside, the T cells migrate toward the B cell location (14). Reciprocal activity occurs with the B cell (15), which upon its activation acquires chemokine receptor CCR7 that responds to T cell zone chemokines CCL19 and CCL21, ensuring interaction of T and B cells at the junction of their respective locations. Once activated, B cells then process antigen in a similar manner to the dendritic cells, displaying the peptide in the groove of class II MHC (16). In the case of transplantation, this antigen is either donor MHC itself or donor MHC processed and subsequently presented by recipient B cells.

Upon B cell antigen presentation to T cell receptor, several hours of interaction occur. Both cells change shape and maximize their contact surface area, permitting efficient and sustained interaction between their respective surface receptors and counter-receptors (17). In this way, soluble factors also may exchange between these cells.

The molecular mechanism that triggers the ultimate antibody response is the recognition by T cell receptors of the specific peptide in the MHC class II groove on the B cell. This initial bridging is augmented quickly by a series of additional links to facilitate the first signal. First, CD4 on the T cell binds a nonpolymorphic (ubiquitous) region on the MHC class II molecule, and then the adhesion molecule CD11a/CD18 (also called LFA-1) T cell integrin expresses high affinity for its intracellular adhesion molecule (ICAM; CD54) ligand upon B cell activation, mechanically stabilizing the bridge. At this time, the CD4–class II links that are located at the peripheral part of the synapse with LFA-1–ICAM adhesion complexes locate centrally (18). As the binding is stabilized, the antigen bridge moves centrally to initiate signaling in both cells (19). The activation of T cells forms an independent complex discussion and is not considered further here.

Additional molecular interactions (or accessory signals) between T and B cells work in concert, leading to their successive induction and recruitment. Four distinct pathways must be functional to mediate the second signal, which facilitates class switching and formation of the germinal center (20–23). The T and B cell molecules involved are, respectively, (1) CD100 to CD72, enhancing B cell survival and class II molecule upregulation; (2) CD154 to CD40, increasing CD80 and CD86, class II and CD95 expression, isotype switching, germinal center formation, and formation of B memory cells, as well as inducing IL-6, IL-10, IL-12, Lymphotoxin, TNF, and other chemokines; (3) CD28 to CD86 transducing activation signals in the T cell, with the subsequent indirect effect on B cells; and (4) Icos to B7 resulting in B cell proliferation and differentiation and germinal center formation (20–23).

Prolongation of activation is achieved by accessory signals, including CD134/CD134L, which assists in proliferation (likely via its effect on T cell activation [24,25]), and isotype switching and CD70/CD27, which stimulates production of memory B cells and plasma cells (26). Negative accessory signals also exist to modulate the T–B cell response. T cell CTLA-4 expression is upregulated on activated T cells, and this in turn interacts with CD80/CD86 and then downregulates T cell activity after a few days (27). T cell CD153 and B cell CD32 interact late in activation to prevent isotype switching and additional B cell maturation (27). Finally, to distinguish an immune response, the CD95-CD95L (Fas-FasL) modulators induce apoptosis in activated T and B cells (28).

The result of initial T and B cell interaction is class-switch recombination (29), with B cells then making all Ig types in addition to their constitutive expression of IgM. Initially, short-lived plasma cells and memory B cells locate in the T zone, and a small percentage produce the first peak of specific (IgM) antibodies (30), which are of only low to medium affinity for the antigen. Subsequent B cells form the germinal center, where they proliferate with numerous mutations in the variable regions of the antibodies. Each "new and different" B cell’s receptor is tested by dendritic-like cells in the follicle (31), and those with insufficient affinity for the antigen are deleted by apoptosis (32). Multiple generations of mutation occur, and, eventually, only a small number remain, forming long-term memory B cells and plasma cells that produce high-affinity antibody of all isotypes (33). These long-term memory B cells can survive for many years and upon re-exposure to the same or similar antigens can be almost immediately reactivated to produce copious highly specific antibody.

	Antibody-Mediated Damage

Top Abstract Introduction Antigen Targets T Cell-B Cell Interactions... Antibody-Mediated Damage Detection of Antibodies in... Antibody-Mediated Syndromes Accommodation Conclusion References

 
Complement Activation
Complement fixing IgG or IgM antibody (irrespective of target) on the vascular endothelium is the predominant way by which antibodies exert their effect on the target organ, classically characterized as hyperacute or acute rejection. The two major pathways of complement activation are the classical pathway, in which certain isotypes of antibodies bind to antigens, initiating the complement cascade, and the alternative pathway, which is activated on microbial cell surfaces in the absence of antibody. We discuss only the former antibody-dependent pathway here (34).

IgG or IgM antibody bound to antigen on the allograft endothelium activates C1 (composed of C1q, C1r, and C1s components) via direct interaction with the C1q globular domain. A conformational change in C1q follows, with subsequent cleavage of C1r, which in turn cleaves and activates C1s, which then activates C2 and C4.

When C1s cleaves C4, C4a (small) and C4b (large) fragments are formed, exposing a sulfhydryl group on C4b that rapidly inactivates by binding to nearby molecules as esters or amides and after inactivation by factor I to C4d remains covalently bound in tissue, thereby easily detectable as a marker of complement activation and, by inference, previous recent antibody–antigen interaction (35–37). There is no evidence that C4d has any functional activity; however, it co-distributes with type IV collagen along the capillary basement membrane and along endothelium (36) and is cleared from tissue after antibody activity has ceased. The presence of C4d does not guarantee that the final common pathway and attendant tissue damage will occur. If activation stops at the C4 level (where C4d would be present in the graft) but activation of C3 did not occur (and no C3d would be detectable in the graft), then graft injury may not occur.

For graft injury to occur, C4b combines with the enzymatically active fragment C2a to form a C4b/C2a complex known as C3-convertase. After C3-convertase has formed, C3 cleaves into C3a and C3b. When the C3b product is present along with C3-convertase, it covalently binds to form C4b/C2a/C3b (the C5-convertase). This cleaves C5, forming C5a and C5b, with the latter initiation formation of the membrane attack complex (MAC) composed of membrane-bound C5b and subsequent complement proteins C6, 7, 8, and 9. The MAC causes lysis of endothelial cells and graft rejection, dependent on C6. Furthermore, C3a and C5a are chemoattractant to neutrophils and macrophages, which express surface receptors for these fragments. C3a also releases prostaglandin E2 from macrophages, and C5a results in edema via histamine release from mast cells (38).

Activation of endothelial cells also is an effect of complement; C3a and C5a activity on their receptors results in increased adhesion molecule expression from endothelial cells (39–41). Exposure to soluble (as opposed to membrane bound) C5b-C9 also increases expression of endothelial adhesion molecules (E-selectin, ICAM-1, and vascular cellular adhesion molecule-1) via IL-1a (42).

The MAC can trigger proliferation of endothelial cells via release of growth factors (platelet-derived growth factor, ß-FGF) (43) and chemokines (CCL2, CCL5, and CXCL8) via IL-1a. Similarly, soluble C5b-C9 promotes secretion of CCL2 and CXCL8 via NF-B pathways (44). Both C5a and C5b-C9 also can trigger synthesis of tissue factor (45,46), which may be responsible in part for the thrombotic injury that dominates severe humoral rejection.

The complement system also is involved in maintaining the normal immune response. Both B and dendritic cells express complement receptor 1 (which binds C3b-C4b) and complement receptor 2 (which binds C3d). CR2 activation lowers the threshold for B cell activation, and complement deficiencies in animal models have been associated with prolonged graft survival and reduced chronic rejection (47–49).

Antibody Action without Complement
The action of antibody on endothelial cells in the absence of complement activation also may have a role in allograft rejection, particularly chronic allograft rejection (50). Even in the absence of complement, endothelial cells demonstrate activation and proliferation in the presence of MHC class I antibodies in vitro (51,52). This activation may be at least partly causative of arterial intimal proliferation that is characteristic of chronic humoral rejection. Noncomplement mechanisms also may stem from direct antibody cell lysis through an Fc receptor on the surface of natural killer cells and macrophages; however, there is only limited evidence that this mechanism is related to acute rejection (53).

	Detection of Antibodies in Serum

Top Abstract Introduction Antigen Targets T Cell-B Cell Interactions... Antibody-Mediated Damage Detection of Antibodies in... Antibody-Mediated Syndromes Accommodation Conclusion References

 
Lymphocytotoxicity assays, as first described in 1969 by Patel and Terasaki (54), have formed the basis of antibody detection through the present. More recently, high-throughput methods of increasing sensitivity (for low-level antibodies) and specificity (for anti-HLA antibodies) have been developed, which has greatly facilitated the increased interest in antibody-mediated processes (Figure 1). The differences in sensitivity of these tests can be dramatic, with flow cytometric assays being more sensitive than either ELISA or cytotoxic methods (55). This difference in sensitivity must be considered when interpreting studies of antibodies and subsequent clinicopathologic outcomes.

View larger version (40K): [in this window] [in a new window] [as a PowerPoint slide]   Figure 1. Methods of antibody detection methods of increasing sensitivity are shown from left to right. Complement-dependent cytotoxicity (CDC): (A) Serum from a patient is added to donor (panel or individual) lymphocytes. If antibodies are present, specific for donor antigen, then they will bind. (B) After washing, nonbound antibodies are removed. (C) Complement is added, which binds to antibodies that are present in sufficient amount and forms the membrane attack complex, killing the cell. (D) In the presence of a vital dye, the dead cells appear red, indicating that donor-specific antibody indeed was present. Anti-human globulin–enhanced CDC (AHG-CDC): (A) Serum from patients is added to donor (panel or individual) lymphocytes and binds to cells with target antigen present. In this example, there are fewer donor-specific antibodies present. (B) After washing, the unbound antibodies are removed. (C) AHG is added, binding to any remaining cell-bound antibody, increasing total Ig amount for complement activation. (D) In the presence of a vital dye, the dead cells appear red, indicating that donor-specific antibody indeed was present. Flow cytometry: (A) Patient serum is added to donor cells. (B) After washing, only bound antibody remains. (C) Anti-human Ig tagged with a fluorescent dye is added and binds to any remaining cell-bound antibody. (D) When cells are run through a flow cytometer, those with fluorescent antibody bound (and thus donor-specific antibody bound) will be detected as increased fluorescence (green peak, right). If no cells have fluorescent antibody bound, then the peak will indicate negative fluorescence (white peak, left). Flow solid phase assay: Purified HLA antigen is bound to inert beads. (A) Patient serum is added to beads. (B) After washing, antibodies that were bound to HLA antigen remain. (C). Anti-human Ig tagged with a fluorescent dye is added and binds to any remaining antibody. (D) When beads are run through a flow cytometer, those with antibody bound are counted with increased fluorescence.

In the case of panel reactive antibody (PRA) testing, the fraction of wells that contain a majority of dead cells compared with the total number of wells examined forms the percentage of PRA. Depending on the nature of the cells used in the panel, it also may be possible to determine the specificity of the antibody (i.e., to which antigen[s] it is binding). For cross-matching, the donor lymphocytes are B and T cells from a single potential donor, such that any antibodies detected are, by definition, donor specific. A positive T cell cross-match suggests class I donor-specific antibodies and is a contraindication to transplantation. Positive B cell cross-matches with negative T cell reactions may indicate low titer class I antibody, class II antibody, or autoantibody/non-HLA antibody, and their effect on subsequent transplantation is determined on an individual basis.

The antibodies that are detected by cytotoxicity are usually against HLA antigen but occasionally may be against non-HLA antigen also. They may be IgG or IgM, the latter of which is not usually of concern in transplantation unless the recipient has experienced a sensitizing event (e.g., blood transfusion) in the preceding few weeks. Heat treatment of the serum or treatment with DTE breaks the IgM pentamer, rendering them nonreactive, such that IgG antibodies may be reliably identified.

Anti-Human Globulin–Enhanced Complement-Dependent Cytotoxicity
If antibodies are of lower titer, then they may not be present in sufficient amount to activate the complement cascade. When anti-human globulin (AHG) is added to the reaction well of a cytotoxicity assay, it binds to antidonor antibody that is already present and bound to the lymphocytes. Unbound antibody along with AHG will be removed in the wash step. The remainder of the assay is performed as described above, but lower titer antibody than standard complement-dependent cytotoxicity (CDC) methods is detectable. AHG enhancement of the T cell CDC cross-match is routine.

Flow Cytometry
Cells.  Even lower titer antibodies and those that do not bind complement may be detected using the most sensitive method of flow cytometry. Donor cells (either panel for PRA or single specific donor for a cross-match) are mixed with recipient serum and washed to remove unbound antibody. Instead of addition of complement, however, antibody to human Ig that has been conjugated with a fluorescent dye is added. This secondary antibody will bind to lymphocyte-bound antibody. When passed through a flow cytometer, cells with primary (antidonor) and secondary (colored) antibody are counted as having higher fluorescence, when the flow cytometer laser excites the color tag. If a threshold of fluorescence is reached, then the test is considered to be positive for the detection of antibody.

Neither complement activation nor high-titer antibody is required to render this test positive. As such, it is possible that a donor–recipient pair may have a negative CDC cross-match but a positive flow cytometry cross-match. Although not a cause of hyperacute rejection per se, these antibodies do have important clinical consequences, with higher rates of acute rejection, worse rejections, and higher rates of graft loss than in patients without these low-level antibodies (56–62). Furthermore, because the secondary antibody is usually specific to IgG, there is no false positivity from IgM antibody. The decision to transplant across a positive flow cross-match is currently center specific.

Solid Phase.  In these assays, which are used for antibody screening before transplantation and confirmation of antibody specificity both before and after transplantation, recipient serum is mixed with inert beads (or an ELISA platform) that bear purified recombinant HLA antigen. As such, only anti-HLA antibody, if present, will bind. Addition of a secondary fluorescence antibody permits for quantification of how many beads have anti-HLA antibody bound. The degree of fluorescence measured represents the amount of anti-HLA antibody present in the original serum sample. The ELISA solid-phase platform has similar sensitivity. This particular method can be used for screening and also to determine reliably specificity of any antibodies found; however, clinical interpretation is necessary to determine the significance of these results (63). The assay is specific for IgG, and non-HLA antibodies are not detected.

Non-HLA Antibody Detection
Currently, there are no commercially available methods for the reliable and reproducible detection of non-HLA antibodies. Methods that are used within research laboratories include flow cytometry against panels of endothelial cells and monocytes and noncommercial ELISA/solid-phase assays of purified antigens. As it becomes increasingly recognized that non-HLA antibodies contribute significantly to antibody-mediated processes, it will be important to find reproducible assays for these antibodies such that their prevalence and impact may be quantified better.

	Antibody-Mediated Syndromes

Top Abstract Introduction Antigen Targets T Cell-B Cell Interactions... Antibody-Mediated Damage Detection of Antibodies in... Antibody-Mediated Syndromes Accommodation Conclusion References

 
The first clinically recognized antibody-mediated syndrome in the modern era of transplantation was described in 1968 in the landmark paper of Terasaki and Patel (54). In a study of 225 renal transplant patients, in which 32 had primary nonfunction of the graft, 24 of 32 had evidence of a circulating factor in recipient serum that caused CDC of donor lymphocytes, compared with only six of 193 with primary graft function who had this factor demonstrable. The primary nonfunction in this case now is recognized as hyperacute rejection in which catastrophic intravascular thrombosis and necrosis are almost immediate after graft reperfusion. The circulating factor described now is known to be antidonor antibody, in most cases, anti-HLA antibody. The CDC assay used in this study, with relatively little modification in the past 37 yr, has formed the basis for the T cell cross-match. Recognition of this antibody-mediated syndrome and the ability of the T cell cross-match to predict its occurrence if positive have virtually eliminated the entity of hyperacute rejection in modern transplantation.

Rejection refers to the activation of the recipient immune system against the allograft and, depending on the time course and clinical presentation, can be classified as subclinical, acute, or chronic. Subclinical rejection occurs when renal biopsy shows the presence of histologic findings of acute rejection without accompanying clinical deterioration (64). Acute rejection develops over days and results in a sudden decline in renal function in association with specific pathologic findings that demonstrate acute inflammation. Chronic rejection, however, is characterized by tubular atrophy and interstitial fibrosis in the clinical setting of a slow decline in renal function over months to years (65).

Despite awareness of the importance of antibodies at the time of transplantation, there remained for many years considerable skepticism regarding any role of antibodies in any of these other clinical presentations after transplantation. The breakthrough came with the use of immunoperoxidase staining for C4d as "proof" of antibody activity in a graft. This technique has allowed for renewed interest in and definition of more specific antibody-mediated syndromes in both the early and late posttransplantation periods. We discuss the evidence supporting the role of antibodies in these clinical syndromes next.

Role of Antibodies in Acute Rejection
Strong suggestion that circulating antibodies may be present and exerting a role in the pathogenesis of acute rejection in addition to the cellular cytotoxicity that was already well described and recognized (65) was reported in 1990 by Halloran et al. (66). In a series of 64 patients, anti-HLA class I antibodies were present in the sera of 100% of patients with acute rejection, demonstrating peritubular capillaritis and vascular lesions, compared with only 41% of patients without similar histology. Subsequent reporting of C4d deposits in peritubular capillaries in patients who demonstrated cellular rejection (1) allowed for further definition and refinement of the histologic findings that are associated with acute antibody-mediated rejection (AAMR).

C4d staining as indicative of AAMR is present in up to 50% of patients who undergo biopsy because of renal dysfunction and up to 32% of biopsies that demonstrate acute rejection (1,37,67–74) (Table 1). Furthermore, C4d staining can be present subclinically, as can cellular rejection, in up to 25% of protocol biopsies, without (76) or with subclinical cellular rejection (67). The presence of C4d in AAMR is highly correlated with circulating donor-specific antibody detected in recipient serum, with sensitivity and specificity >95%, and is superior to histology alone, with sensitivity and specificity of 68 and 96%, respectively (67).

Just as peritubular capillary C4d staining is associated with circulating alloantibodies in biopsies that demonstrate acute rejection histology, it is similarly associated with circulating alloantibodies in up to 21 to 85% of biopsies that show chronic rejection changes, in comparison with 0 to 22% of biopsies that demonstrate nonimmune chronic injury (72,79–85) (Table 3). Furthermore, acute rejections with C4d-positive staining are more likely to lead to chronic rejection (32 to 44%) compared with those that are C4d negative (8 to 14%) (87,88).

C4d positivity not only is associated with chronic rejection and transplant glomerulopathy but also predicts it. Regele et al. (72) demonstrated that in first-year biopsies, C4d staining was a strong predictor of subsequent glomerulopathy after 12 mo (46 versus only 6% in the control group).

Anti-HLA antibody also has been eluted from needle biopsies of functional grafts with chronic allograft nephropathy (89), and a significant correlation between their presence and the presence of C4d staining and plasma cell infiltrate was found (90). Furthermore, anti-HLA antibodies were not found in the biopsy eluates of well-tolerated transplants, strongly supporting a pathogenic role of donor-specific anti-HLA antibody in chronic rejection and chronic allograft nephropathy. Therefore, analogous to the new criteria approved for acute humoral rejection, chronic antibody-mediated damage is increasingly recognized as a distinct entity (91) and is currently pending addition to the Banff Criteria (Table 4).

View larger version (69K): [in this window] [in a new window] [as a PowerPoint slide]   Figure 2. Proposed stages of humoral alloimmunity. The first stage of an alloimmune response is circulating alloantibody. If antibody deposits and activates complement, then C4d will be detectable in the allograft. Subsequently, if accommodation does not occur, then pathologic changes that are consistent with tissue damage will occur; if extensive enough, then graft dysfunction will result. The pathologic changes seen and time to development of graft dysfunction can be variable. Adapted from reference (91).

This presence of antibody within the graft is the basis of stage 2 of the proposed model in which C4d is detectable in the microvasculature of the graft but no evidence of graft dysfunction is present. This finding and that of circulating antibodies do not necessarily terminate in rejection, and the graft at this stage actually may be demonstrating accommodation (discussed later). By stage 3, in addition to the C4d staining, there are pathologic changes consistent with antibody-mediated damage, and by stage 4, graft dysfunction is present. Although each stage is a prerequisite for the next, the progression through all stages is not known to be inevitable. Nonetheless, it is an important model that facilitates more precise definitions of antibody-mediated processes and allow for the development of stage-specific interventions and treatment strategies.

Role of Non-HLA Antibodies
A plethora of non-HLA antibodies have been shown in a variety of small studies to be associated with acute and chronic humoral consequences. The extent to which these have been explored is related to the lack of commercially available and reproducibly validated assays for the infinite number of potential antibodies to a myriad of targets in the renal allograft. Certainly, the observation that >10% of cases with C4d positivity fail to show circulating anti-HLA antibody is suggestive that non-HLA antibodies also are to be considered (67). Correlations between anti-endothelial antibodies and chronic allograft rejection have been documented (96,97). Antivimentin (a cytosolic protein derived from endothelial cells and expressed in the intima and media of arteries) antibodies are associated with early transplantation coronary artery disease (chronic rejection) in cardiac allografts (9). It has been shown that posttransplantation development of alloantibody specific to MICA is correlated with chronic rejection and poor allograft survival (6,98).

	Accommodation

Top Abstract Introduction Antigen Targets T Cell-B Cell Interactions... Antibody-Mediated Damage Detection of Antibodies in... Antibody-Mediated Syndromes Accommodation Conclusion References

 
Certain alloantibodies are not associated with acute graft injury and in fact may correlate with good graft survival. The best example of this is in long-term outcomes after ABO-incompatible transplantation. Despite that the anti-A/B antibodies return, the graft is not rejected (99). The transient depletion of graft-specific antibodies at the time of transplantation prevents hyperacute rejection. If antibodies rebound within 10 d, then there is a high rate of acute rejection; however, if the antibody rebound is delayed until 3 wk or later, then there is no correlation between titer and rejection (100). Such accommodation is formally defined as the resistance of an allograft to the acute pathologic effects of graft-specific antibodies and complement fixation (101).

In HLA-mismatched grafts, the presence of alloantibodies in the absence of graft dysfunction fulfills this criterion. However; because these patients have a greater prevalence of late graft loss, it suggests that if this is in fact accommodation, then it may be only transient or at best incomplete.

The graft may resist the effects of antibody and complement activation by developing proteins that are protective of endothelial cells (102). In rodent models, antiapoptotic proteins have increased expression in accommodated xenografts (103). Direct complement regulation also may be involved. Cell surface inhibitors of the classical and alternative complement pathways have been identified.

	Conclusion

Top Abstract Introduction Antigen Targets T Cell-B Cell Interactions... Antibody-Mediated Damage Detection of Antibodies in... Antibody-Mediated Syndromes Accommodation Conclusion References

 
In the past few years, there has been an increasing interest in antibody-mediated injury in renal transplant recipients. This interest has been spurred by the improved ability to detect antibody activity through C4d staining as well as the development of increasingly sensitive methods for detecting circulating antibodies. Antibody-associated injury has been found to be associated with both acute and chronic types of injury, although it is unclear whether some of the humorally mediated injury that is currently being reported has always been present but undetected or it represents a heightened humoral response that is brought on by changes in immunosuppressive medications. Undoubtedly, this will continue to be an area of great interest in terms of fully understanding the mechanisms of antibody-mediated injury as well as the potential for clinical intervention.

	Footnotes

 
Published online ahead of print. Publication date available at www.cjasn.org.

	References

Top Abstract Introduction Antigen Targets T Cell-B Cell Interactions... Antibody-Mediated Damage Detection of Antibodies in... Antibody-Mediated Syndromes Accommodation Conclusion References

 

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