Each HLA allele has a unique four-, six-, or eight-letter or digit name (Table 31-1). The length of the allele designation depends on the sequence of the allele and that of its nearest relative. All alleles receive a four-letter or digit name; six- and eight-digit names are only assigned when necessary.

Table 31-1

HLA Naming System ^∗

Nomenclature	Indicates
HLA	Human leukocyte antigen (HLA) region and prefix for an HLA gene
HLA-DRB1	Particular HLA locus (e.g., DRB1)
HLA-DRB1^∗13	Group of alleles that encode the DR13 antigen
HLA-DRB1^∗1301	Specific HLA allele
HLA-DRB1^∗1301N	Null allele
HLA-DRB1^∗130102	Allele that differs by a synonymous mutation
HLA-DRB1^∗13010102	Allele that contains a mutation outside the coding region
HLA-A^∗2409N	Null allele
HLA-A^∗3014L	Allele encoding a protein with significantly reduced or low cell surface expression
HLA-A^∗24020102L	Allele encoding a protein with significantly reduced or low cell surface expression, where the mutation is found outside the coding region
HLA-B^∗44020102S	Allele encoding a protein expressed as a secreted molecule only
HLA-A^∗3211Q	Allele that has a mutation previously shown to have a significant effect on cell surface expression, but where this has not been confirmed and its expression remains questionable

^∗As of June 2007, no alleles have been named with the “C” or “A” suffixes.

The first two digits describe the type, which often corresponds to the serologic antigen carried by an allotype. The third and fourth digits are used to list the subtypes, with numbers assigned in the order in which DNA sequences have been determined.

Alleles whose numbers differ in the first four digits must differ in one or more nucleotide substitutions that change the amino acid sequence of the encoded protein. Alleles that differ only by synonymous nucleotide substitutions (also called silent or noncoding substitutions) within the coding sequence are distinguished by the use of fifth and sixth digits. Alleles that only differ by sequence polymorphisms in the introns or in the 5′ or 3′ untranslated regions that flank the exons and introns are distinguished by the use of seventh and eighth digits.

In addition to the unique allele designation, optional suffixes may be added to an allele to indicate its expression status. Alleles shown not to be expressed, termed null alleles, have been given the suffix N. Alleles shown to be alternatively expressed may have the suffix L, S, C, A, or Q.

The suffix L is used to indicate an allele shown to have low cell surface expression compared with normal levels. The S suffix is used to denote an allele specifying a protein that is expressed as a soluble secreted molecule but that is not present on the cell surface. A C suffix indicates an allele product that is present in the cytoplasm but not on the cell surface. An A suffix indicates aberrant expression, where there is some doubt as to whether a protein is expressed. A Q suffix is used when the expression of an allele is questionable, given that the mutation seen in the allele has previously been shown to affect normal expression levels.

Major Histocompatibility Complex Regions

The MHC is divided into four major regions (Table 31-2)—D, B, C, and A. The A, B, and C regions are the classic or class Ia genes that code for class I molecules. The D region codes for class II molecules. Class I includes HLA-A, B, and C. The three principal loci (A, B, and C) and their respective antigens are numbered 1, 2, 3, and so on. The class II gene region antigens are encoded in the HLA-D region and can be subdivided into three families, HLA-DR, HLA-DC (DQ), and HLA-SB (DP).

Table 31-2

Examples of Nomenclature of HLA Alleles

Allele (New Nomenclature)	Frequently Used Shorthand
Class I
HLA-A^∗0101	HLA-A1
HLA-B^∗0801	HLA-B8
Class II
HLA-DRB1^∗0101	HLA-DR1
HLA-DRB1^∗0301	HLA-DR3

Adapted from Peakman M, Vergani D: Basic and clinical immunology, ed 2, New York, 2009, Churchill Livingstone.

Classes of Human Leukocyte Antigen Molecules

Structurally, there are two classes of HLA molecules, class I and class II (Table 31-3). Both classes are cell surface heterodimeric structures. Class I HLA molecules consist of an alpha chain, a highly polymorphic glycoprotein, encoded within the MHC on chromosome 6. This alpha chain noncovalently associates with beta-2 microglobulin, a nonpolymorphic glycoprotein, encoded by a non-HLA gene on chromosome 15. Class II HLA molecules are composed of alpha chains and beta chains encoded within the MHC. The conformation of class I and class II HLA molecules provides each with a groove in which linear peptides, consisting of 8 to 25 peptides, are displayed for recognition by the cell surface expression on lymphocytes of a transmembrane heterodimeric receptor. All nucleated cells of the body display transmembrane class I HLA molecules in association with the non–transmembrane beta-2 microglobulin molecule.

Table 31-3

Comparison of Major Histocompatibility Complex Class I and Class II

Parameter	Class I	Class II
Loci	HLA-A, B, and C	HLA-DN, DO, DP, DQ, and DR
Distribution	Most nucleated cells	B lymphocytes, macrophages, other antigen-presenting cells, activated T lymphocytes
Function	To present endogenous antigen to cytotoxic T lymphocytes	To present endogenous antigen to helper T lymphocytes

MHC, Major histocompatibility complex.

Class I and class II antigens can be found on body cells and in body fluids. Class I and class II molecules are surface membrane proteins. Class I molecules are transmembrane glycoproteins, but the class II dimer molecule differs from class I in that both dimers span the cell membrane. Class I and class II gene products are biochemically distinct, although they appear to be distantly related through evolution. Class III gene products such as C2, C4A, C4B, and Bf complement components are incomplete but these structures are defined by genes lying between or very near the HLA-B and HLA-DR loci.

Multiple alleles occur at each locus. Genes of class I, II, and III antigens at each locus are inherited as codominant alleles. Inheritance within families closely follows simple mendelian dominant characteristics. Conservation of entire haplotypes through generation after generation is the general rule. Very strong linkage disequilibrium is displayed between several HLA loci, creating super or extended haplotypes that may differ from race to race. For example, the most frequent Caucasoid superextended haplotype, AL, Xw7, BB, BfS, C2-1, C4AQOB1, DR3, is almost absent in Asians.

Role of Major Histocompatibility Complex and Human Leukocyte Antigens

The histocompatibility complex that encodes cell surface antigens was first discovered in graft rejection experiments with mice. When the antigens were matched between donor and recipient, the ability of a graft to survive was remarkably improved. A comparable genetic system of alloantigens was subsequently identified in human beings.

The presence of HLA was first recognized when multiply transfused patients experienced transfusion reactions despite proper crossmatching. It was discovered that these reactions were caused by leukocyte antibodies rather than by antibodies directed against erythrocyte antigens. These same antibodies were subsequently discovered in the sera of multiparous women.

The MHC gene products have an important role in clinical immunology. For example, transplants are rejected if performed against MHC barriers; thus, immunosuppressive therapy is required. These antigens are of primary importance and are second only to the ABO antigens in influencing the genetic basis of survival or rejection of transplanted organs.

Although HLA was originally identified by its role in transplant rejection, it is now recognized that the products of HLA genes play a crucial role in our immune system. T cells do not recognize antigens directly but do so when the antigen is presented on the surface of an antigen-presenting cell (APC), the macrophage. In addition to presentation of the antigen, the macrophage must present another molecule for this response to occur. This molecule is a cell surface glycoprotein coded in each species by the MHC. T cells are able to interact with the histocompatibility molecules only if they are genetically identical (MHC restriction).

Both class I and class II antigens function as targets of T lymphocytes that regulate the immune response. Class I molecules regulate interactions between cytolytic T cells and target cells and class II molecules restrict the activity of regulatory T cells (helper, suppressor, and amplifier subsets). Thus, class II molecules regulate the interaction between helper T cells and APCs. Cytotoxic T cells directed against class I antigens are inhibited by CD8 cells; cytotoxic T cells directed against class II antigens are inhibited by CD4 cells. Many genes in both class I and class II gene families have no known functions.

The class I and class II molecules can also bind to self antigens produced in the normal process of cellular protein degradation. Usually, these are not recognized by the T cell receptor (TCR; tolerance). In transplant patients, most immune responses are generated not from bacterial antigens, viral antigens, or self antigens, but from the presentation of alloepitopes derived from the transplanted tissue to circulating T lymphocytes. Two types of alloepitopes are present on transplanted tissue, private and public. Cross-reactive groups have been defined that categorize the cross-reactive alleles of HLA-A and HLA-B.

Class III molecules bear no clear relationship to class I and II molecules aside from their genetic linkage (presence of the gene in or near the MHC complex). Class III molecules are involved in immunologic phenomenon because they represent components of the complement pathways.

Human Leukocyte Antigen Applications

HLA matching is of value in organ transplantation, as well as in the transplantation of bone marrow. The most important HLA antigens are HLA-A, and HLA-B. Everyone has two types of each of these major HLA antigens; there are many different subtypes of HLA-A and of the others. The best possible match is 6/6; the worst possible match is 0/6.

In kidney allografts, the method of organ preservation, the time elapsed between harvesting and transplanting, the number of pretransplantation blood transfusions, the recipient’s age, and the primary cause for kidney failure are all important determinants of early transplantation success or failure. HLA compatibility, however, exerts the strongest influence on long-term kidney survival. The 1-year survival for kidneys transplanted from an HLA-identical sibling approaches 95%. Approximately 50% to 65% of cadaveric kidneys mismatched for all four HLA-A and -B antigens function for 6 months but deteriorate thereafter with time. Only 15% to 25% of these mismatched cadaveric kidneys remain functioning 4 years after transplantation.

It is obligatory to select HLA-identical donors for bone marrow transplantation to reduce the frequency of graft-versus-host disease (GVHD; see later). A method, that depletes donor marrow T cells capable of recognizing foreign host antigens has greatly reduced the incidence of GVHD.

HLA-matched platelets are useful for patients who are refractory to treatment with random donor platelets. In paternity testing, HLA typing is used, along with the determination of ABO, Rh, MNS, Kell, Duffy, and Kidd erythrocyte antigen. In the past, most laboratories involved in testing individuals in disputed parentage cases used only the ABO, Rh, and MNS systems. The chances of identifying a falsely accused man with these tests were 58%. Additional testing for Kell, Duffy, and Kidd erythrocyte antigens and for HLA typing has an exclusion rate estimated at 92%.

HLA typing is also useful in forensic medicine, anthropology, and basic research in immunology. In studies of racial ancestry and migration, some antigens are almost excluded or confined to a race (e.g., A1 and B8 are rarely detected in peoples indigenous to central and eastern Asia, and Bw57 is uncommon in whites and African Americans). These distinctions allow for precise conclusions to be drawn regarding origin and ancestry.

HLA testing has increasingly been used as a diagnostic and genetic counseling tool. Knowledge of HLA antigens and their linkage has become important because of the recognized association of certain antigens (Box 31-1) with distinct immunologic-mediated reactions, autoimmune diseases, some neoplasms, and other disorders; these disorders, although nonimmunologic, are influenced by non-HLA genes also located within the major MHC region.

Box 31-1 Relationship of Certain Human Leukocyte Antigens and Diseases

Ankylosing spondylitis	B27
Reiter’s syndrome	B27
Psoriasis vulgaris	Cw6
Rheumatoid arthritis	DR4
Behçet’s disease	B5 (Bw51)
Type 1 diabetes	DR3
Gold-induced nephropathy	DR5
Congenital adrenal hyperplasia	B47
Chronic lymphatic leukemia	DR5
Kaposi’s sarcoma (Mediterranean)	DR5

The estimated relative risks or chances of developing a disease if a given antigen is present may be elevated in individuals bearing certain HLA antigens compared to individuals who lack the antigen (Table 31-4). The HLA-B27 antigen is the only HLA antigen with a disease association strong enough to be useful in differential diagnosis. Although the degree of association between HLA antigens and other diseases may be statistically significant, it is not strong enough to be of diagnostic or prognostic value.

Table 31-4

Relationship of Human Leukocyte Antigens to Risk of Disease

Antigen Present	Related Disease	Risk^∗
B27	Ankylosing spondylitis Reiter’s syndrome Anterior uveitis Arthritic infection with Yersinia or Salmonella Psoriatic arthritis with spinal involvement Spondylitis associated with inflammatory bowel disease Juvenile chronic arthritis with spinal involvement	100×^† 40× 25× 20× 11× 9× 5×
B8	Celiac disease Addison’s disease Myasthenia gravis Dermatitis herpetiformis Chronic active hepatitis Sjögren’s syndrome Diabetes mellitus (insulin dependent) Thyrotoxicosis	9× 6× 5× 4× 4× 3× 2× 2×
B5	Behçet’s syndrome	6×
BW38	Psoriatic arthritis	7×
BW15	Diabetes mellitus (insulin-dependent)	3×
DR2	Goodpasture’s syndrome Multiple sclerosis	16× 4×
DR3	Gluten-sensitive enteropathy Dermatitis herpetiformis Subacute cutaneous lupus erythematosus Addison’s disease Sjögren’s syndrome (primary)	21× 14× 12× 11× 10×
DR4	Pemphigus^‡ Giant cell arthritis Rheumatoid arthritis Juvenile (insulin-dependent) diabetes mellitus	32× 8× 6× 5×
DR5	Pauciarticular juvenile arthritis Scleroderma Hashimoto’s thyroiditis	5× 5× 3×

^∗Increased risk of developing the disease over a lifetime.

^†Varies with ethnic group (e.g., 3× for Pima Indians and 300× for Japanese).

^‡Jewish persons.

Adapted from Ashman RF: Rheumatic diseases. In Lawlor GJ, Fischer TJ, editors: Manual of allergy and immunology, ed 2, Boston, 1998, Little, Brown.

Although only 8% of normal whites carry the HLA-B27 antigen, 90% of patients with ankylosing spondylitis (AS) or spondylitis in association with Reiter’s syndrome are positive for the antigen. An elevated percentage of HLA-B27–positive patients is also observed in juvenile chronic arthritis with spinal involvement. Therefore, the major indication for screening for HLA-B27 test is to rule out AS when back pain develops in relatives of patients with the disease and to help distinguish incomplete Reiter’s syndrome from gonococcal arthritis, or chronic or atypical Reiter’s syndrome from rheumatoid arthritis. A negative test for HLA-B27, however, does not exclude the diagnosis of AS or Reiter’s syndrome.

Laboratory Evaluation of Potential Transplant Recipients and Donors

Systems developed to ascertain compatibility between donor and recipient include HLA typing and screening of the potential recipient’s serum for the presence of antibodies associated with rejection. Graft success is generally correlated with the presence of a compatible crossmatch, although some transplantation teams have proceeded despite a positive (incompatible) crossmatch.

Human Leukocyte Antigen Typing

A potential recipient needs to have HLA typing (Fig. 31-1, A). A family search may be conducted for a suitable donor. If a suitable match is not found, the patient is placed on a waiting list (see Fig. 31-1, B). When an organ becomes available, the donor is HLA-typed and a computerized search is made for a suitable recipient (see Fig. 31-1, C).

Figure 31-1 Patients (recipients) requiring a solid organ transplant such as a kidney are human leukocyte antigen (HLA)–typed **(A)** and then placed on a transplant registry waiting list **(B).**

C, When a potential organ becomes available, the donor is HLA-typed. If the donor and recipient demonstrate a suitable match by computer, a blood sample is procured from the donor and crossmatched with the recipient’s blood to determine compatibility. D, If no HLA antibodies are detected against the donor cells, the organ is harvested and transplanted to the recipient. (From Nairn R, Helbert M: Immunology for medical students, ed 2, St Louis, 2007, Mosby.)

A newer method of HLA typing is polymerase chain reaction (PCR) amplification of DNA, followed by probing with sequence-specific oligonucleotide probes (SSOPs) and PCR amplification of alleles at loci using allele-specific primers. A simple computer program has been developed to assign the alleles and genotypes based on the probe hybridization pattern.

The difference between HLA-genotyped and zero mismatches reflects the imperfection of the HLA-typing process. HLA genotype–matched means an HLA-identical sibling when all alleles are truly identical. A zero-mismatched unrelated donor may be mismatched because typing does not distinguish between very closely related alleles. In addition, there may be some effect of non-HLA loci because HLA-identical siblings will share only 50% of their minor histocompatibility loci. The degree of donor-recipient mismatch is somewhat obvious, even in the first year.

Flow Cytometry

Single-cell analysis by flow cytometry is the most sensitive method for crossmatching and antibody identification (see Chapter 13). Tagged T or B lymphocytes are incubated with the patient’s serum to allow the formation of antigen-antibody complexes on the cell surface. Unbound proteins are washed away and the bound antibodies are detected with a second antibody, anti–human immunoglobulin G (IgG) labeled with a chromophore. An alternative flow cytometry format uses microparticles coated with HLA antigens of known specificity (obtained through recombinant techniques) instead of lymphocytes.

Facts About Solid Organ Transplantation

Organ transplantation is widely viewed as the preferred treatment for end-stage organ failure because of the quality of life that the treatment offers for patients and because of the long-term cost benefits. The increased demand for organ transplantation is fueled by the transplantation success rate. Worldwide, the demand for transplantation procedures is increasing by about 15%/year, but the number of donated organs has remained static.

The waiting time for allergenic organ transplantation varies widely for many reasons. Each patient’s situation is different. Some patients are more ill than others when they are put on the transplant waiting list. Some patients become sick more quickly than others or respond differently to treatments. Patients may have medical conditions that make finding a good match more difficult.

How long a patient waits for a transplant depends on the following factors:

• Blood type (some rarer than others)

• Tissue type

• Height and weight of transplant candidate

• Size of donated organ

• Medical urgency

• Time on the waiting list

• Distance between donor’s hospital and potential donor organ

• Number of donors in local area over time

• Transplantation center’s criteria for accepting organ offers

Depending on the type of organ needed, some factors are more important than others.

In 1984, the U.S. Congress passed the National Organ Transplant Act. The goal of this legislation was to match a low supply of organs with the most critically ill patients, regardless of where they reside. About 79 patients receive an organ transplant every day in the United States. In 2009, more than 28,000 patients received an organ transplant.

On March 22, 2012, the United Network for Organ Sharing patient waiting list contained 113,612 names. The list continues to grow because of the scarcity of organs (Box 31-2). Most of these registrants are waiting for a kidney transplant, followed by those waiting for a liver transplant and heart transplant. Other transplant registrants are waiting for lung, kidney and pancreas, pancreas, pancreatic islet cell, heart and lung, and intestine. Approximately 25% of patients waiting for a liver transplant are children younger than 10 years.

Box 31-2 Factors Contributing to the Scarcity of Organs

Demand for transplantation has increased because more patients are considered eligible.

Age limits for heart and liver transplants eligibility have increased.

Diabetes is no longer an absolute contraindication for transplant eligibility.

The number of donors has shown little growth.

Adapted from Caplan A: Organ procurement and transplantation: ethical and practical issues, 1995 (www.upenn.edu/ldi/issuebrief2_5.html).

The most common reasons for needing a transplant vary by the type of organ. Kidney recipients usually have diabetes, glomerulonephritis, hypertensive nephrosclerosis, or polycystic kidneys. Liver recipient patients typically have noncholestatic cirrhosis, cholestatic liver disease, biliary atresia, acute hepatic necrosis, or hepatitis C infection. Patients with cardiomyopathy, congenital heart disease, valvular heart disease, or coronary artery disease are the most frequent heart transplant recipients.

The number of patients living with a function graft has generally increased over the last decade. Graft survival time depends on many factors, including the type of organ transplanted (Fig. 31-2 and Table 31-5).

Table 31-5

Selected Examples of Single Organ Graft Transplant Survival (Unadjusted Graft Survival Expressed in Percentage (%))

Follow–up Period
	3 Months	1 Year	3 Years	5 Years	10 Years
	TX 2007-2008	TX 2007-2008	2005-2008	2003-2008	1998-2008
Kidney: Deceased Donor Kidney: Living Donor	95.6 98.0	91.7 96.5	81.8 90.5	70.8 82.8	44.9 61.2
Pancreas	85.0	74.8	62.6	52.0	35.1
Liver: Deceased Donor	91.7	85.2	75.1	68.5	54.8
Liver: Living Donor	91.3	88.2	80.1	74.6	59.6
Intestine	91.3	81.7	55.2	41.5	24.0
Heart	93.1	88.3	80.7	73.9	54.7
Lung	91.6	81.5	63.3	51.2	26.1

Tx = year or inclusive years of tranplantation.

Source: Organ Procurement and Transplantation Network (OPTN) and Scientific Registry of Transplant Recipients (SRTR). OPTN / SRTR 2010 Annual Data Report. Rockville, MD: Department of Health and Human Services, Health Resources and Services Administration, Healthcare Systems Bureau, Division of Transplantation; 2011. www.srtr.org/annual_reports/2010

Figure 31-2 Number of surviving patients after kidney or liver transplants from deceased or living donors. (From Organ Procurement and Transplantation Network (OPTN) and Scientific Registry of Transplant Recipients (SRTR). OPTN / SRTR 2010 Annual Data Report. Rockville, MD: Department of Health and Human Services, Health Resources and Services Administration, Healthcare Systems Bureau, Division of Transplantation; 2011, www.srtr.org/annual_reports/2010.)

Transplantation Terminology

The transplanting or grafting of an organ or tissue ranges from self-transplantation, such as skin grafts from one part of the body to another to correct burn injuries, or hair transplants from one area of the scalp to another to correct pattern baldness, to the grafting of a body component from one species to another, such as transplanting a pig’s heart valve to a human. Table 31-6 defines the most recent terms used in transplantation.

Table 31-6

Transplantation Terms

Term	Definition
Autograft	Graft transferred from one position to another in the same individual (e.g., skin, hair, bone)
Syngraft	Graft transplanted between different but identical recipient and donor (e.g., kidney transplant between monozygous twins)
Allograft (homograft)	Graft between genetically different recipient and donor of the same species; grafted donor tissue or organ contains antigens not present in recipient
Xenograft (heterograft)	Graft between individuals of different species (e.g., pig heart valve to a human heart)

Types of Transplants

Eleven different organs or human body parts can be transplanted—blood vessels, bone, bone marrow or stem cells (see Chapter 32), cornea, heart, kidneys, liver, lung, middle ear, pancreas, and skin. Successful organ transplants have increased since the advent of the immunosuppressive drug cyclosporine (cyclosporin A).

Living donor transplants have attracted significant media attention. According to the United Network for Organ Sharing and the Health Resources and Services Administration of the U.S. Department of Health and Human Services, a living donor may donate a single kidney, segment of the liver, portion of the pancreas, or the lobe of a lung.

Bone

Bone matrix autografts or allografts are common. Transplantation of bone matrix is used after certain limb-sparing tumor resections and to correct congenital bone abnormalities. The major criteria for bone donation are a lack of infection, no history of IV drug use, and no history of prolonged steroid therapy or human growth hormone treatment. Bone can be easily harvested and frozen. Freezing not only preserves the bone but offers the additional benefit of concomitant diminution of histocompatibility antigens.

The major technical requirement for allograft transplantation is maintaining the periosteal sheath of the recipient bone to strip the donor bone completely of all periosteal elements. Transplantation of bone is an easy procedure. Processed bone lacks significant quantities of immunogenic substances; therefore, the need for immunosuppression is almost completely eliminated.

Cornea

Corneal transplants have been a common form of therapy for many years. The first human corneal eye bank was established in New York City in 1944. This type of transplantation has an extremely high success rate because of the ease in obtaining and storing viable corneas.

Corneal grafts are generally performed to replace nonhealing corneal ulcerations. Graft rejection is minimal because of the following: (1) the avascularity (lack of blood vessels) of this tissue; (2) a reasonably low concentration of class I transplantation antigens; and (3) an essential absence of class II antigens. To prevent rejection, grafts are made as small as possible and are placed centrally to avoid contact with the highly vascularized limbic region. Eccentrically placed grafts are subject to a high rate of immunologic failure because vascularity will allow for lymphocyte contact. Immunosuppressive agents are not routinely administered.

Heart

The first successful allograft cardiac transplantation was performed in 1967 by Dr. Christian Barnard in Cape Town, South Africa. The criteria for selecting the donor and recipient combination for cardiac transplantation are essentially the same as those used for cadaveric renal transplantation. The most significant exclusion for cardiac transplantation, however, is the presence of an active infection. Cardiac transplant donors must have sustained irreversible brain death, but near-normal cardiac function must be maintained. Prophylactic antibiotics and cytotoxic drugs are given to the donor just before harvesting of the heart. Because of the urgency of most situations, most grafts are performed despite multiple HLA incompatibilities. Transplant recipients are maintained on immunosuppressive therapy, anticoagulants, and antithrombotic agents, as well as on a low-lipid diet.

Due to advances in immunosuppression following heart transplantation, there has been an increase in the rate of 1-year survival among recipients to almost 90%, but acute cellular rejection is still observed during the first year after transplantation and at lower rates after the first year. Endomyocardial biopsy remains the primary method for monitoring organ rejection for heart transplants. An alternative method for detecting the rejection of a heart transplant, aside from endomyocardial biopsy, is quantitative assessment of mononuclear cell gene expression in peripheral blood specimens. A study conducted with 602 patients to compare the two methods for monitoring patients for rejection has shown that the overall rate of survival does not differ significantly according to the method of monitoring.

Heart Valves

Xenogenic valve replacement is a standard modality for the treatment of aortic and mitral valve defects. Sources of these xenogenic valves are bovine (cow) or porcine (pig); the valves are chemically or physically modified to reduce antigenicity.

Patients receiving xenoallografts of heart valves are not immunosuppressed after surgery because only minimal or nonexistent graft rejection reactions take place in these modified valves.

Intestine

The first successful intestine transplantation was performed at the University of Toronto in 1986, although the patient only survived for 10 days. The first intestinal transplant recipient to survive for an extended amount of time was a 3½-year-old girl who lived for 192 days in 1987. Intestinal transplantation has improved over the past decade along with the number of intestinal transplantations performed in North America. In 2008, 185 intestinal transplantations were performed. With recent surgical advances, control of acute cellular rejection, and decrease in lethal infections, the rate of patient survival for the first year now exceeds 90%.

When the small intestine is transplanted alone it is referred to as an isolated intestinal transplant, but intestinal transplantations are usually performed with other organs with a composite allograft or with organs implanted separately from the same donor. Suitable intestinal organ donors have stable cardiopulmonary status and liver function. Potential organ transplant recipients with systemic infection and malignancy are excluded.

After a donor is accepted, selective decontamination of the gastrointestinal tract is begun through a nasogastric tube using polyethylene glycol. Generally, the recipient of the transplant is a person suffering from short gut syndrome, in which the intestine had been resected for a variety of reasons.

Kidney

The first successful human kidney transplantation was performed in 1954 between monozygotic twins. Induction of tolerance (see later) was attempted through the use of sublethal total body irradiation and allogeneic bone marrow transplantation, followed by renal transplantation. By 1960, renal transplantation was firmly established as a viable treatment for end-stage renal disease. Because of the continuing problems associated with total-body irradiation, chemical immunosuppression became the mode of treatment. The criteria for recipients of renal allografts generally exclude older patients and patients with a history of malignancy. In addition, patients with active sepsis or patients in whom chronic infection may be reactivated by treatment with steroids or immunosuppressive therapy are also not considered transplantation candidates.

Traditionally, kidney donations are not accepted from individuals older than 65 years because of a decreased likelihood of recipient survival. Donors are excluded if chronic renal disease or sepsis is present. Transplant donations are usually not accepted from those with generalized or systemic diseases such as diabetes mellitus, hypertension, and tuberculosis. Because of the severe shortage of donor kidneys, organs from donors older than 55 years or from donors with a history of hypertension or diabetes mellitus have been used with increasing frequency. Young trauma victims are the most desirable source of cadaveric organ transplants, including the kidneys. Cadaveric organs are not accepted from donors with a history of any malignancy other than that involving the central nervous system.

In addition to tissue compatibility, newer methods of harvesting kidneys have reduced the sensitizing effect related to passenger leukocytes against transplantation antigens borne on these cells. HLA-A and HLA-B loci matches have the best chance for long-term survival of the graft and recipient. The increased survival rate with HLA-A and HLA-B matches is determined not as much by class I compatibility as by the HLA-D region–related antigens associated with these regions. The strongest association between transplantation survival and tissue antigens is with the D region–related antigens (DR, MB, MT). Lewis antigens on the erythrocytes and H-Y antigens associated with X and Y chromosomes are among the other antigen systems that demonstrate a reasonably significant association with graft survival.

Liver

Potential liver transplant recipients must have no extrahepatic disease or infection present. The largest group of transplant recipients has been those with congenital biliary atresia. Patients with cirrhosis may also be good candidates. HLA crossmatching appears to increase the rate of graft survival, but the influence of tissue typing is somewhat unclear. Immunosuppressive regimens such as azathioprine and corticosteroids or cyclosporin A increase survival. Major complications of this procedure have been biliary tract fistulae or leaks, which have occurred in 30% to 50% of patients.

Lung

Successful lung transplantation has been difficult to achieve because of technical, logistic, and immunologic problems. Technically, the lung donor and recipient must have essentially identical bronchial circumferences to obtain a good match. An additional technical problem is that the lungs are extremely sensitive to ischemic damage, and successful preservation after harvesting has been unsuccessful. Occasionally, lung-heart combination transplantation has been attempted. The combined procedure is less difficult than single-organ transplantation.

The lungs are susceptible to infection; sepsis is very common among potential donors. Severe rejection is common because of the high density of Ia-positive cells in the vasculature and the high concentration of passenger leukocytes trapped in the alveoli and blood vessels. Intensive immunosuppressive therapy is needed to maintain the graft. Many lung recipients have died from massive infection and sepsis.

Pancreas

Newer modes of transplantation include full pancreatic or isolated islet cell transplantation. Pancreatic grafts have been successful for only a short period because of a high rate of technical failure or irreversible rejection. Transplantation of small quantities of isolated islet cells into the retroperitoneal space, however, has demonstrated a reasonably good success rate.

Pancreatic islet transplants are risky and experimental, with about 50% of patients achieving insulin dependency after 1 year. From December 16, 1966 to December 31, 2008, more than 30,000 pancreas transplants were reported to the International Pancreas Transplant Registry (IPTR). There are three types of pancreatic transplantations that can be done: pancreas-kidney transplantation (SPK, the most common; 73%); pancreas transplantation after kidney transplantation (PAK; 18%); and pancreas transplantation alone (PTA; 9%).

Skin

The development of nonimmunogenic skin replacement materials has lowered the demand for skin allografts. Skin allografts elicit the rejection phenomenon because skin has an extremely high density of MHC class I antigens. Therefore, sensitization and recognition of antigenic differences are likely, with resultant rejection of the grafted skin. If done, skin allografts are performed and supported with immunosuppressive therapy.

Graft-Versus-Host Disease

Graft-versus-host disease (GVHD) can be an unintentional consequence of blood transfusion or transplantation in severely immunocompromised or immunosuppressed patients. The degree of immunodeficiency in the host, rather than the number of transfused immunocompetent lymphocytes, determines whether GVHD will occur.

Etiology

When immunocompetent T lymphocytes are transfused from a donor to an immunodeficient or immunosuppressed recipient, the transfused or grafted lymphocytes recognize that the antigens of the host are foreign and react immunologically against them (Table 31-7). Instead of the usual transplantation reaction of host against graft, the reverse graft-versus-host reaction occurs and produces an inflammatory response.

Table 31-7

Requirements for Potential Graft-versus-Host Disease

Factor	Comments
1. Source of immunocompetent lymphocytes	Blood products, bone marrow transplant, organ transplant
2. Human leukocyte antigen differences between patient and recipient	The stronger the antigen difference, the more severe the reaction.
3. Inability to reject donor cells	Patients are severely immunocompromised or immunosuppressed.

In a normal lymphocyte transfer reaction, the results of a GVHD are usually not serious because the recipient is capable of destroying the foreign lymphocytes. However, engraftment and multiplication of donor lymphocytes in an immunosuppressed recipient are a real possibility because lymphocytes capable of mitosis can be found in stored blood products. If the recipient cannot reject the transfused lymphocytes, the grafted lymphocytes may cause uncontrolled destruction of the host’s tissues and eventually death. A patient can develop chronic or acute GVHD. The stronger the antigen difference, the more severe is the reaction.

Epidemiology

It is now accepted that GVHD can occur whenever immunologically competent allogeneic lymphocytes are transfused into a severely immunocompromised host. Patients at risk include those who are immunodeficient or immunosuppressed with severe lymphocytopenia and bone marrow suppression. Despite chemotherapy at the time of bone marrow transplantation, patients are highly likely to develop acute GVHD and some of these immunocompromised patients will die of GVHD or associated infections.

Chronic GVHD affects 20% to 40% of patients within 6 months after transplantation. Two factors closely associated with the development of chronic GVHD are increasing age and a preceding episode of acute GVHD.

Cases of transfusion-related GVHD have increased significantly in the past 2 decades. This reaction has been reported subsequent to blood transfusion in bone marrow transplant recipients after total-body irradiation and in adults receiving intensive chemotherapy for hematologic malig-nancies. GVHD has also occurred in infants with severe congenital immunodeficiency and in those who received intrauterine transfusions followed by exchange transfusion. Almost 90% of patients with posttransfusion GVHD will die of acute complications of the disease. The usual cause of death is generalized infection.

Signs and Symptoms

GVHD causes an inflammatory response. Posttransfusion symptoms begin within 3 to 30 days after transfusion. Because of lymphocytic infiltration of the intestine, skin, and liver, mucosal destruction results, including ulcerative skin and mouth lesions, diarrhea, and liver destruction. Other clinical symptoms include jaundice, fever, anemia, weight loss, skin rash, and splenomegaly.

In bone marrow transplant patients, acute GVHD develops within the first 3 months of transplantation. The initial manifestations are lesions of the skin, liver, and gastrointestinal tract. An erythematous maculopapular skin rash, particularly on the palms and soles, is usually the first sign of GVHD. Disease progression is characterized by diarrhea, often with abdominal pain, and liver disease. Other signs and symptoms of complications related to therapy include fever, granulocytopenia, and bacteremia. Interstitial pneumonia, frequently associated with cytomegalovirus (CMV), can also occur.

Chronic GVHD resembles a collagen vascular disease, with skin changes such as erythema and cutaneous ulcers, and a liver dysfunction characterized by bile duct degeneration and cholestasis. Patients with chronic GVHD are susceptible to bacterial infections. For example, increasing age and preexisting lung disease increase the incidence of interstitial pneumonia.

Immunologic Manifestation

In immunocompromised patients, the transfused or grafted lymphocytes recognize the antigens of the host as foreign and react immunologically against them. Instead of the usual transplantation reaction of host against graft, the reverse GVHD occurs.

Diagnostic Evaluation

Laboratory evidence of immunosuppression or immunodeficiency, such as a decreased total lymphocyte concentration, suggests that a patient may develop GVHD. Evidence of inflammation, such as an increased C-reactive protein (CRP) level, elevated leukocyte count with granulocytosis, and increased erythrocyte sedimentation rate (ESR), may suggest that GVHD has developed in GVHD candidates. Complications of anemia and liver disease, characterized by increased levels of bilirubin and blood enzymes (e.g., transaminases, alkaline phosphatase), and the presence of opportunistic pathogens (e.g., CMV) can further support the diagnosis.

Pathologic features include lymphocytic and monocytic infiltration into perivascular spaces in the dermis and dermoepidermal junction of the skin and into the epithelium of the oropharynx, tongue, and esophagus. Infiltration can also be observed into the base of the intestinal crypts of the small and large bowels and into the periportal area of the liver, with secondary necrosis of cells in infiltrated tissues.

Prevention

The incidence of GVHD can be minimized by depletion of mature lymphocytes from the marrow by using monoclonal antibodies or physical methods. The risk of GVHD can be minimized, if not eliminated, by irradiation of the marrow transplant or blood products. Blood product irradiation is believed to be the most efficient and probably the most economical method available for the prevention of posttransfusion GVHD.

No cases of posttransfusion GVHD have been reported after the administration of irradiated blood products irradiated with an effective and appropriate radiation dose. Several categories of patients possess the clinical indications for the use of irradiated products.

High-Risk Patients

Patients at the highest risk with an absolute need for irradiated blood products include the following:

• Recipients of autologous or allogeneic bone marrow grafts. Recipients of autologous bone marrow may be expected to have the same risk of posttransfusion GVHD as patients receiving allogeneic bone marrow.

• Children with severe congenital immunodeficiency syndromes involving T lymphocytes. The degree of immunodeficiency in the host, rather than the number of transfused immunocompetent cells, determines whether GVHD will occur.

Intermediate-Risk Patients

Patients considered to be at a lower risk of developing GVHD include the following:

• Infants receiving intrauterine transfusions, followed by exchange transfusions, and possibly infants receiving only exchange transfusions. The immune mechanism of the fetus and newborn may not be sufficiently mature to reject foreign lymphocytes and prior transfusions may induce a state of immune tolerance in the newborn. Transfused lymphocytes may continue to circulate for a prolonged period in some immunologically tolerant hosts without the development of GVHD. There is insufficient evidence to recommend irradiation of blood given to all premature infants.

• Patients receiving total-body radiation or immunosuppressive therapy for disorders such as lymphoma and acute leukemia. Although routine irradiation of blood products given to these patients can be justified, it cannot be regarded as absolutely indicated because the risk of developing GVHD is so low. Blood product irradiation, however, is advised for selected patients with hematologic malignancies, especially when transfusions are given during or near the time of sustained and severe therapy-induced immunosuppression.

Low-Risk Patients

Patients also at risk but considered the least susceptible include the following:

• Patients with solid tumors. The incidence of the development of GVHD in these patients is difficult to determine. However, it has developed in nonhematologic malignancies such as neuroblastoma. In one case, GVHD developed after infusion of a single unit of packed red blood cells (RBCs).

• Patients with aplastic anemia receiving antithymocyte globulin theoretically may be at increased risk of posttransfusion GVHD during therapy-induced periods of lymphocytopenia.

• Although a theoretical risk of posttransfusion GVHD may exist in patients with acquired immunodeficiency syndrome (AIDS), the disease has not actually been observed in this disorder. The routine use of irradiated blood is not recommended.

Effects of Radiation on Specific Cellular Components

Lymphocytes

Ionizing radiation is known to inhibit lymphocyte mitotic activity and blast transformation. Irradiation of normal donor lymphocytes with 1500 rad from a cesium-137 source results in a 90% reduction in mitogen-stimulated ¹⁴C-thymidine incorporation. An 85% reduction in mitogen-induced blast transformation after exposure to 1500 rad and a 97% to 98.5% reduction in mitogenic response have been noted after an appropriate exposure to radiation.

Granulocytes

Ionizing radiation may impair granulocyte function in a dose-dependent manner. The degree of actual damage to granulocytes is controversial. Chemotactic activity decreased linearly with increasing doses of irradiation from 500 to 120,000 rad, but the reduction only reached statistical significance at 10,000 rad. A linear dose-response curve demonstrates that granulocyte locomotion is affected by very small doses of irradiation. An appropriate dose of radiation is likely to eliminate lymphocytic mitotic activity and prevent GVHD without causing significant damage to granulocytes or altering their chemotactic or bactericidal ability. Irradiation before transfusion has been demonstrated to contribute to defective oxidative metabolism, but this effect is highly variable.

Mature Red Blood Cells

Mature RBCs appear to be highly resistant to radiation damage. After RBCs were exposed to 10,000 rad, ⁵²Cr-labeled in vivo RBC survival was the same as that of untreated controls. Stored erythrocytes can be treated with up to 20,000 rad without changing their viability or in vitro properties, including adenosine triphosphate (ATP) and 2,3-diphosphoglycerate (2,3-DPG) levels, plasma hemoglobin (Hb), and potassium ions (K⁺).

Platelets

Ionizing radiation may impair platelet function. Although this impairment is dose-dependent, the effects of irradiation on platelets have been difficult to characterize. Several studies have demonstrated unchanged in vivo platelet survival after exposure to 5000 to 75,000 rad. A 33% decrease in the expected platelet count increase was noted after transfusion of platelets exposed to 5000 rad, and similarly irradiated autologous platelets had a diminished ability to correct the bleeding times in a small number of volunteers who had consumed aspirin.

Immunologic Tolerance

The importance of tolerance to self antigens was recognized early in the study of immunology. Immunologic tolerance is the acquisition of nonreactivity toward particular antigens. Self-recognition (tolerance) is a critical process, and the failure to recognize self antigens can result in autoimmune disease (see Chapter 28).

Various pathways to immunologic tolerance have been recognized. It has been suggested that T and B cells are affected independently and differently and may be tolerated under certain circumstances. Several mechanisms may operate simultaneously in a single host. During fetal development of the immune system and during the first few weeks of neonatal life, none of the cells of the immune system has reached maturity. For this reason, the entire immune system is particularly susceptible to tolerance induction at this stage of development.

T Cell Tolerance

T cells do not show a marked difference in tolerance at different stages of maturation. The antigen required to produce tolerance and the circumstance of its presentation are specific for each individual T cell subset. At least three pathways have been recognized for T cell tolerance:

1. Clonal abortion. Immature T cell clones may be aborted in a manner similar to that of B cells.

2. Functional deletion. The subsets of a mature T cell may be individually deleted, leading to the loss of only one of the functions of the T cell group.

3. T cell suppression. T cell suppressors actively suppress the actions of other T cell subsets or B cells.

B Cell Tolerance

As a B cell matures, it becomes less susceptible to tolerization. In addition, during B cell maturation, the forms of antigen presentation that will produce tolerance also vary. Four pathways have been established for the induction of B cell tolerance. Therefore, the mode of tolerance depends on the maturity of the cell, antigen, and manner of antigen presentation to the immune system.

The pathways of B cell tolerance are as follows:

1. Clonal abortion. A low concentration of multivalent antigen may cause the immature clone to abort. Tolerance of immature B cells by this mechanism is high.

2. Clonal exhaustion. Repeated antigen challenge with a T-independent antigen may remove all mature functional B cell clones. Tolerance of mature B cells is moderate.

3. Functional deletion. The combined absence of the helper T subset and presence of T-dependent antigen (or with T suppressor cells), or an excess of T-independent antigen, prevents mature B cells from functioning normally. The ability to tolerize B cells by this mechanism is moderate.

4. Antibody-forming cell blockade. An excess of T-independent antigen interferes with the secretion of antibody by antibody-forming cells. B cell tolerance by this mechanism is low.

Immune Response Gene–Associated Antigens

The specific immune responses to a variety of antigenic substances are now known to be regulated by an immune response (Ir) gene. Ir gene control is considered genetically dominant. The homology of the HLA-D region with the animal I region suggests that the human Ir gene might be linked to the HLA complex. Evidence for the existence of the Ir gene has been obtained from family and population studies. Additional evidence for the presence of Ir genes comes from HLA-linked disease susceptibility genes and HLA-disease associations. It is believed that individuals who lack this gene are unresponsive.

The generally accepted concept is that the Ir gene is responsible for the interaction of T cells with B cells and macrophages, which are necessary for T cell activation. Activation of T cells is required for the following:

• Conversion to active helper function

• Production of lymphokines

Mediation of delayed and contact hypersensitivity, as the proliferative response to antigen, depends on the interaction of a T cell with an APC, usually macrophage-monocytes. Helper function also depends on T cell interaction with precursors of antibody-secreting cells. T cells interact with these cells by recognizing specific antigen bound to macrophages or to B cells and the I region gene products expressed on the surface of these cells. T cells are able to recognize the precise details of antigen structure and distinguish between two closely related Ir gene–associated molecules expressed on the surface of these APCs or on the B cell.

Graft Rejection

Organs vary with respect to their susceptibility to rejection based on inherent immunogenicity (Box 31-3), which is influenced by factors such as vascularity.

The role of sensitized lymphocytes and antibodies in graft rejection differs and is influenced by the type of organ transplanted. Lymphocytes, particularly recirculating small lymphocytes, are effective in shortening graft survival. Cell-mediated immunity is responsible for the rejection of skin and solid tumors. However, humoral antibodies can also be involved in the rejection process. The complexity of the action and interaction of cellular and humoral factors in grafts is considerable. Five possible categories of graft rejection have been demonstrated in human kidney transplant rejection—hyperacute, accelerated, acute, chronic, and immunopathologic (Table 31-8; Color Plate 17).

Table 31-8

Categories and Characteristics of Graft Rejection Based on Immune Destruction of Kidney Grafts

Type	Time of Tissue Damage	Predominant Mechanism	Cause
Hyperacute	Within minutes	Humoral	Preformed cytotoxic antibodies to donor antigens
Accelerated	2-5 days	Cell-mediated	Previous sensitization to donor antigens
Acute	7-21 days	Cell-mediated (possibly antibody cell-mediated cytotoxicity)	Development of allogeneic reaction to donor antigens
Chronic	Later than 3 mo	Cell-mediated	Disturbance of host-graft tolerance
Immunopathologic damage to the new organ	Later than 3 mo	1. Immune complex disorder 2. Complex formation with soluble antigens	Immunopathologic mechanisms related to circumstances necessitating transplantation

First-Set and Second-Set Rejections

Skin transplantation is the most common experimental model for transplantation research (Fig. 31-3). Rejection of skin and solid tumors can be divided into first-set and second-set rejections. Activation of cellular immunity by T cells is the predominant cause of the first-set allograft rejection. Lymphocytes can directly attack cellular antigens to which they are sensitized by previous exposure or by cytotoxic lymphokines. The primary role of lymphocytes in first-set rejection is consistent with the histology of early reaction and shows infiltration by mononuclear cells, with very few polymorphonuclear leukocytes or plasma cells. Sensitization occurs within the first few days of transplantation, and the tissue is lost in 10 to 20 days.

When sensitized lymphocytes are already present because of prior graft rejection, an accelerated rejection of tissue results from regrafting, called second-set rejection. Lymphocytes from a sensitized animal transferred to a first-graft recipient will accelerate rejection of the graft. Graft rejection is primarily a T cell function, with some assistance from antibodies.

Hyperacute Rejection

Hyperacute reactions are caused entirely by the presence of preformed humoral antibodies in the host, which react with donor tissue cellular antigens. These antibodies are usually anti-A–related or anti-B–related antibodies to the ABO blood group systems or antibodies to class I MHC antigens (hypersensitivity type II). Potential recipients harboring antibodies to HLA-A, HLA-B, and HLA-C (class I) but not HLA-DR (class II) antigens are at high risk for this process.

The interaction of cellular antigens with antibodies activates the complement system and leads to grafted cell lysis and clotting in the grafted tissue. Kidney allografts can be rejected by the hyperacute rejection process within minutes of transplantation. The irreversible kidney damage of hyperacute rejection is characterized by sludging of erythrocytes, development of microthrombi in the small arterioles and glomerular capillaries, and infiltration of phagocytic cells.

Genetically altered pig organs could be available for transplantation into human beings within 2 years, but it is likely to be at least 5 years before full-scale studies can begin. Future xenotransplantation will depend on overcoming problems of hyperacute rejection. In hyperacute rejection, the recipient of the organ produces xenoreactive antibodies, which lodge on the cells lining the blood vessels of the new organ and trigger the release of complement. This release triggers inflammation, swelling, and ultimately blockage of the blood vessels, leading to death of the organ.

Accelerated Rejection

Accelerated rejection is comparable to the second-set rejection phenomenon observed in animal models. In these cases, retransplantation is less severe than hyperacute rejection and is considered to be accelerated rejection. Accelerated rejection is caused by activation of the T cell–mediated response.

Acute Rejection

Acute rejection can result after the first exposure to alloantigens. In this reaction, donor antigens select reactive T cell clones and initiate visible manifestation of rejection within 6 to 14 days. The early processes in acute rejection appear to be T cell–mediated; however, later aspects may involve antibodies and complement.

Acute rejection is equivalent to a first-set allograft rejection in experimental animals and is primarily mediated by cells, as in accelerated rejection. Immunopathologic changes include the presence of immune complex deposition and other hypersensitivity reactions already present in the recipient.

Acute rejection takes place when there is HLA incompatibility. Recipient T cells can respond to donor peptides presented by a recipient MHC or to donor MHC molecules themselves. The better the HLA match, the more successful are the prospects for nonrejection. Because of the shortage of organs and the huge demand for organs, partially mismatched organs (e.g., kidneys) may be used. The survival of the kidney is related to the degree of mismatching, especially at the HLA-DR loci. Despite mismatching, 1-year survival with five mismatches was almost 80% because of the effect of potent immunosuppressive drugs.

A recipient may respond to minor histocompatibility antigens. Minor antigens are encoded by genes outside the HLA. These minor histocompatibility antigen mismatches are not detected by standard tissue typing techniques but may cause rejection despite a good HLA match. Up to one third of transplants can be rejected because of minor antigens.

Acute early rejection, which occurs up to about 10 days after transplantation, is histologically characterized by dense cellular infiltration and rupture of peritubular capillaries. It appears to be a cell-mediated hypersensitivity reaction involving T cells. In comparison, acute late rejection occurs 11 days or more after transplantation in patients suppressed with prednisone and azathioprine. In kidney allografts, acute late rejection is probably caused by the binding of immunoglobulin, presumably antibody and complement, to the arterioles and glomerular capillaries, where they can be visualized by immunofluorescent techniques. These immunoglobulin deposits on the vessel walls include platelet aggregates in glomerular capillaries, which cause acute renal shutdown. The possibility of damage to antibody-coated cells through antibody-dependent, cell-mediated cytotoxicity (ADCC) may also take place.

Chronic Rejection

Chronic rejection occurs in most graft recipients. The process results in a slow but continual loss of organ function over months or years. However, chronic rejection is often responsive to various immunosuppressive therapies.

In kidney allografts, this insidious rejection is associated with subendothelial deposits of immunoglobulin and the C3 component of complement on the glomerular basement membranes. This may occasionally be an expression of an underlying immune complex disorder that may have originally necessitated the transplantation, or it may result from complex formation with soluble antigens derived from the grafted kidney.

Mechanisms of Rejection

General Characteristics

Variations in the expression of class II histocompatibility antigens by different tissues and the presence of APCs in some tissues greatly influence the success of a transplant. APCs that enter the graft through the donor’s circulation are likely to elicit graft rejection. If these so-called passenger lymphocytes leave the graft after transplantation and enter the draining lymphatic system, they are particularly effective in sensitizing the host.

Rejection of a graft displays the following two key features of adaptive immunity:

• Memory

• Specificity

Only sites accessible to the immune system in the recipient are susceptible to graft rejection. Certain privileged sites in the body allow allogeneic grafts to survive indefinitely.

Role of T Cells

Graft rejection is primarily regulated by the interaction of the host’s T cells with the antigens of the graft. Unmodified rejection, however, results from the destructive effects of cytotoxic T (Tc) cells, activated macrophages, and antibody.

In tissue transplants, the graft consists of tissue cells that carry class I antigens (HLA-A, HLA-B, and HLA-C) and of lymphocytes that carry class I and class II antigens (HLA-D and related antigens of an associated Ir gene). Activated T cells specific for class I antigens have the potential to express cytotoxic activity, which damages the endothelium and parenchymal cells of the graft. Binding of these cells to the class I antigens on target cells of the donor organ triggers the release of lymphokines and subsequently activates a nonspecific inflammatory response in the allograft.

T cells specific for class II antigens of the donor tissue cannot react directly with the parenchymal cells of the graft not expressing class II antigens. However, these cells can activate lymphocytes in the transplant through lymphocyte release. Therefore, damage to the graft can result from a cytotoxic reaction directed against cells of the transplanted organ, a severe nonspecific inflammatory response, or both.

Activation of helper T (Th) cells by class II antigens such as HLA-DR probably stimulates the release of interleukin-1 (IL-1). IL-1 subsequently stimulates the release of various lymphokines from Th cells, which in turn activate macrophages, Tc cells, and antibody-releasing B cells, as well as increase the immunogenicity of the graft. In addition, macrophages and other accessory cells are subsequently stimulated by T cell products and release IL-1, which in turn stimulates the formation of IL-2 receptors and the release of IL-2 by Th cells. IL-2 interacts with specific IL-2 receptors expressed on activated Th and Tc cells. This interaction stimulates the initiation of DNA synthesis and the eventual clonal proliferation of IL-2 receptor–bearing cells. IL-2 also causes the release of interferon-γ (IFN-γ), which activates macrophages and stimulates the release of B cell differentiation factors required for the proliferation of antigen-activated B cells. The release of IL-2–dependent IFN-γ by activated T cells may initiate a vicious circle, because IFN-γ induces the expression of class II molecules on endothelial cells, as well as the expression of certain class II–negative macrophages.

Histologic examination of an allogenic skin graft during the process of rejection demonstrates that the dermis becomes infiltrated by mononuclear cells, many of which are small lymphocytes. This accumulation of lymphocytes precedes the destruction of the graft by several days. Although this graft rejection process is caused by Tc cells, in some cases Th cells are also elicited by MHC gene differences. Graft rejection may be a special form of response related to delayed hypersensitivity reactions, in which case the ultimate effectors of graft destruction are the monocyte-macrophages recruited to the site. It is debatable whether the macrophages seen in grafts are effectors of graft destruction or arrive only as a consequence of the inflammatory process and cell damage.

Antibody Effects

Cell-mediated immunity is the major effector mechanism in graft rejection. Antibodies, however, can also be involved in graft rejection. Antibodies can cause rapid (hyperacute) graft rejection, but they are usually less significant than cell-mediated immunity. Exceptions include cases in which the recipient has been previously sensitized to a particular antigen, reactions occur to hematopoietic cells, or the graft is directly connected to the host’s blood circulation (e.g., kidney allograft).

In dispersed cellular grafts, such as infusion of erythrocytes, leukocytes, and platelets, antibodies (humoral immunity) may dominate the rejection process because antigens are fully exposed to a preexisting or developing antibody response. Cells are highly susceptible to complement-activated membrane damage. If cytolysis does not occur immediately, antibodies may function as opsonins to encourage phagocytic destruction of transfused cells.

Humoral immunity is suspected of playing a major role in the rejection of xenografts. Xenografts possess a large number of antigens shared between donor and recipient. One species can possess agglutinins for cells of distantly related species, which can attack the xenogenic tissue as soon as it is transplanted.

Immunosuppression

For most patients who receive a donated organ, immunosuppressant drug therapy and monitoring of the concentration of immunosuppressants play a critical role in the success of the transplant. Laboratory methods for measuring immunosuppressant drug concentrations in blood include immunoassay, high-performance liquid chromatography (HPLC), and liquid chromatography with mass spectrometry (LC-MS). Clinical laboratories are increasingly using LC-MS for routine measurement of immunosuppressants.

Immunosuppression is used for the following:

• Induction (intense immunosuppression in the initial days after transplantation)

• Maintenance of transplant

• Reversal of established rejection

Forms of immunosuppression include chemical (Box 31-4), biologic, and irradiation of the lymphoid system or the donated organ. The immunosuppressive activities of therapeutic agents used in transplantation directly interfere with the allograft rejection response. The problem arising from all immunosuppressive techniques is that the person is more susceptible to infection. If infection occurs, immunosuppression must be suspended, at which time allogeneic reactions frequently develop.

Box 31-4 Immunosuppression in Human Organ Transplantation

1945-1955	Research on antimetabolites, including 6-mercaptopurine, azathioprine, and corticosteroids, used to improve kidney graft survival.
Late 1960s	Antilymphocyte globulin proved successful.
1976	Cyclosporine developed.
1983	Cyclosporine approved by FDA.
1984	OKT3 (muromonab-CD3) approved by FDA.
1994	Tacrolimus (FK-506) approved by FDA.
1995	Mycophenolic acid approved by FDA (almost 30 years after development).
1996	Cyclosporine microemulsion (Neoral) approved by FDA.
1997	Antithymocyte globulin approved by FDA.
1997	Dacliximab (Zenapax) approved by FDA.
1999	Sirolimus (Rapamune) approved by FDA.
2004 2008-2010 2011	Enteric-coated mycophenolic acid approved by FDA, generic formulations of both tacrolimus and mycophenolate mofetil approved by FDA. Nulojix (belatacept) approved by FDA.

FDA, U.S. Food and Drug Administration.

Immunosuppressive measures may be antigen-specific or antigen-nonspecific (Table 31-9). Antigen-nonspecific immunosuppression includes drugs and other methods of specifically altering T cell function. Many cytotoxic drugs are primarily active against dividing cells and therefore have some functional specificity for any cells activated to divide by donor antigens. The use of these drugs is limited by the toxic effects that they may have on other dividing cells or on the physiologic functioning of organs such as the liver.

Table 31-9

Types of Immunosuppressive Treatment

Drug	Mechanism of Action
Corticosteroids	Reduce inflammation by inhibiting macrophage cytokine secretion
Cyclosporine and FK506	Blocks T cell cytokine production by inhibiting the phosphatase calcineurin and then blocking activation of the NFAT transcription factor
Mycophenolate mofetil	Blocks lymphocyte proliferation by inhibiting guanine nucleotide synthesis in lymphocytes
Rapamycin	Blocks lymphocyte proliferation by inhibiting IL-2 signaling
Anti-CD3 monoclonal antibody	Depletes T cells by binding to CD3 and promoting phagocytosis or complement-mediated lysis (used to treat acute rejection)
Anti–IL-2 receptor antibody	Inhibits T cell proliferation by blocking IL-2 binding; may also opsonize and help eliminate activated IL-2R-expressing T cells
CTLA4-Ig Nulojix (belatacept)	Inhibits T cell activation by blocking B7 costimulator binding to T cell CD28 (clinical trials) A selective T cell costimulation blocker

NFAT, Nuclear factor of activated T cells. IL, interleukin, CTLA4-Ig, cytotoxic T lymphocyte-associated protein-4-immunoglobulin (fusion protein).

FDA approves Nulojix for kidney transplant patients www.fda.gov/NewsEvents

Adapted from Abbas AK, Lichtman AH: Basic immunology, ed 3, St Louis, 2011, Saunders.

Antigen-specific immunosuppression is an ideal form of immunosuppression. Antigen-specific tolerance is that induced by the infusion of donor cells. This is generally impractical in transplantation, but may be useful in the phenomenon of immunologic enhancement. Enhancement of tolerance has been attempted in renal allograft patients. In a donor-specific blood transfusion program, the patient is transfused several times before elective transplantation with blood from the prospective kidney donor. The overall effect of these transfusions appears to be a tolerance of the recipient to donor transplantation antigens other than those in the HLA-linked regions, such as minor histocompatibility loci, RBC loci, and leukocyte surface antigens. This treatment has greatly prolonged graft survival in these patients.

Cytotoxic Drugs

Cytotoxic drugs are the most common form of therapy and usually include alkylating agents, purine and pyrimidine analogues (Fig. 31-4), folic acid analogues, or the alkaloids. The drugs of choice, excluding alkylating drugs, are azathioprine, 6-mercaptopurine, 6-thioguanine, 5-fluorouracil, cytosine arabinoside, methotrexate and aminopterin, and vinblastine and vincristine.

Figure 31-4 Pyrimidine analogues *(upper row)* and purine analogues *(lower row)* with B and T cell–suppressing activity. A large number of similar compounds are available for human use. (Adapted from Barrett JT: Textbook of immunology, ed 5, St Louis, 1988, Mosby.)

Most immunosuppressive drugs administered alone cannot produce antigen-specific tolerance because they act equally on all susceptible clones. Except for certain drugs (e.g., cyclosporin A), most immunosuppressive agents can be rendered antigen-specific only by including an antigen-specific element in the tolerizing regimen. In these cases, the drugs act as cofactors in tolerogenesis. Experimental evidence has suggested that these regimens may act as follows:

• Lowering the threshold for tolerance induction

• Blocking the differentiation sequence in cells triggered by antigen

Azathioprine

Since its introduction in 1961, azathioprine, an oral purine analogue that is an antimetabolite with multiple activities, has been the mainstay of antirejection therapy. Azathioprine requires activation to 6-mercaptopurine, which is further metabolized to active 6-thioguanine nucleotides. Metabolites of azathioprine, such as the in vivo metabolite 6-mercaptopurine, are incorporated into cellular DNA. This inhibits purine nucleotide synthesis and metabolism and alters the synthesis and function of ribonucleic acid (RNA). Therefore, azathioprine acts at an early stage in T cell or B cell activation during the proliferative cycle of effector lymphocyte clones. Azathioprine is useful in preventing acute rejection because it inhibits the primary immune response; however, it has little or no effect on secondary responses. Adverse effects include bone marrow suppression, myopathy, alopecia, pancreatitis, and hepatitis. A drug interaction can occur with allopurinol.

Corticosteroids

Corticosteroids can be used in conjunction with azathioprine or other immunosuppressants such as cyclosporine. Corticosteroids directly inhibit antigen-driven T cell proliferation, but steroids do not act directly on the IL-2–producing T cell. They do, however, inhibit production of lymphokines by preventing monocytes from releasing IL-1, thereby blocking IL-1–dependent release of IL-2 from antigen-activated T cells. Other activities of monocytes, such as inhibition of chemotaxis, are also likely to be important in the immunosuppressive process.

High doses of corticosteroids are used to treat acute rejection. In addition, steroids probably reverse in vivo rejection episodes by preventing the production of IL-2, which would inhibit activated T cells as an essential trophic factor.

Cyclosporine (Cyclosporin A)

Cyclosporine, isolated in 1971 from the fungus Tolypocladium inflatum, has become the mainstay of immunosuppressive therapy in transplantation. Cyclosporine affects T cells preferentially by inhibiting the induction of cytotoxic T cells. Unlike corticosteroids, cyclosporine does not inhibit the capacity of all accessory cells to release IL-1. Cyclosporine blocks calcineurin to the IL-2 gene transcription pathway and the release of certain other lymphokines (e.g., IFN-γ). Cyclosporine binds to cyclophilin and the complex binds to and inhibits calcineurin (a protein phosphatase). This prevents activation of the IL-2 transcription factor.

The secretion of B cell growth and differentiation factors by activated T cells is also inhibited by cyclosporin A. Therefore, under the influence of cyclosporin A, Th cell–dependent B cells are not fully activated because of a lack of necessary Th cell stimulation. In pharmacologic doses, however, cyclosporin A does not grossly interfere with the activation and proliferation of suppressor T cells. Studies have shown prolonged renal allograft survival with cyclosporin A, despite potential mismatches of the HLA system. Adverse effects of corticosteroids include fluid retention, electrolyte abnormalities, hyperglycemia, hypertension, peptic ulcer disease, osteoporosis, and adrenal insufficiency. Hepatotoxicity has been observed in 4% to 7% of patients. Drug interactions can occur with grapefruit juice, erythromycin, oral contraceptives, and a variety of other drugs. Drug monitoring is critical because of the narrow therapeutic range.

A newer cyclosporine microemulsion offers the advantage of improved trough measurement correlation with the actual patient circulating concentration.

Tacrolimus

Tacrolimus (FK-506), a macrolide with mechanisms similar to that of cyclosporine, is derived from a fungus, Streptomyces tsukubaensis, found in soil samples in Japan. FK-506 is 50 to 100 times more powerful than cyclosporine. Its primary target appears to be the Th lymphocytes, with little effect on other aspects of the immune response. FK-506 acts early in the process of T cell activation and inhibits the production of IL-2. As a result, T lymphocytes do not proliferate, secretion of IFN-γ is inhibited, MHC class II antigens are not induced, and further activation of macrophages does not occur.

Because FK-506 is a more potent immunosuppressant than cyclosporine, patient recovery time is faster. FK-506 has higher toxicity compared with cyclosporine. Nephrotoxicity, hyperkalemia, hypokalemia, hypomagnesemia, hypertension, and other side effects may occur, but FK-506 causes no serious side effects (e.g., kidney damage, elevated blood pressure, mood swings). Patients receiving FK-506 have increased susceptibility to infections (e.g., CMV) and an increased risk of developing lymphoma or posttransplantation lymphoproliferative diseases. Inhibitors and inducers of P-450 3A4 may demonstrate an altered rate of metabolism that requires an adjustment in drug dose.

Sirolimus

Sirolimus (Rapamune), previously referred to as rapamycin, was under development for more than 20 years before gaining approval by the U.S. Food and Drug Administration (FDA). Sirolimus is derived from the fungus Streptomyces hygroscopicus from the soil of Easter Island. Structurally, sirolimus resembles tacrolimus and has the same intracellular binding protein or immunophilin, known as FKBP-12, but sirolimus has a novel mechanism of action. Sirolimus is a substrate for P-450 3A4 and inhibits the activation and proliferation of T lymphocytes and subsequent production of IL-2, IL-4, and IL-15. Sirolimus also inhibits antibody production. Sirolimus has been approved as an adjunctive agent (in combination with steroids) for the prevention of acute renal allograft rejection. The main side effects include increased risk of infections and lymphoma, hypercholesterolemia, hypertriglyceridemia, interstitial pneumonitis, insomnia and tremor, and thrombocytopenia.

Mycophenolate Mofetil

Mycophenolate mofetil (RS-61443) inhibits de novo guanosine synthesis by inhibiting inosine monophosphate dehydrogenase. This drug inhibits T and B lymphocyte proliferation and antibody formation by B lymphocytes and has been efficacious as prophylactic and rescue therapy in refractory renal allograft rejection in clinical trials.

Mycophenolate mofetil (MMF; CellCept), is a drug that is now being used more frequently in treatment plans as a substitute for azathioprine. MMF prevents the production of cells such as azathioprine but is believed to be more effective for preventing rejection in patients. Studies have suggested that mycophenolate is effective in preventing acute rejection and may also slow the progression to chronic rejection. Adverse side effects include a lowering in blood cell development, which can cause abdominal pain, vomiting, and diarrhea, but generally it is a well-tolerated drug.

Antilymphocyte (Antithymocyte) Globulin

Other immunosuppressive measures directed at T cells include the use of antilymphocyte (antithymocyte) globulin (ATG), an IgG polyclonal antibody, at the time of transplantation and the use of lymphoid irradiation before transplantation. The usefulness of ATG for preventing or reversing rejection in renal allograft recipients has been well established. Adverse side effects can include complement-mediated lysis of lymphocytes, serum sickness, leukopenia, and thrombocytopenia.

Among patients at high risk for acute rejection or delayed graft function who have received a kidney transplant from a cadaveric donor, induction therapy consisting of a 5-day course of antithymocyte globulin, as compared with basiliximab, reduces the incidence and severity of acute rejection but not the incidence of delayed graft function.

A regimen of total lymphoid irradiation plus antithymocyte globulin decreases the incidence of acute GVHD and allows graft antitumor activity in patients with lymphoid malignant diseases or acute leukemia treated with hematopoietic cell transplantation.

Nulojix

One of the newest drugs is Nulojix (belatacept). This drug was approved by the FDA in 2011 to prevent acute rejection in adult patients who have had a kidney transplant. This drug is approved for use with other immunosuppressants, e.g., basiliximab, mycophenolate mofetil, and corticosteroids. This type of drug is called a selective T cell costimulation blocker.

Monoclonal Antibodies

Monoclonal antibody (muromonab-CD3, OKT2 [Orthoclone, OKT3]) is used because the CD3 surface membrane marker is found on all mature post-thymic T cells. Interaction between OKT3 and the surface of mature T lymphocytes causes T cell depletion. The use of OKT3 reverses almost all acute renal transplant rejection and is indicated for the treatment of steroid-resistant rejection. A side effect of this drug is cytokine-release syndrome, a condition of flulike symptoms, dyspnea, aseptic meningitis, and pulmonary edema.

Dacliximab (Zenapax) is a recently approved humanized monoclonal antibody to the alpha subunit of the IL-2 receptor. A decreased incidence of renal allograft rejection has been observed with triple- and/or double-immunosuppressive regimens.

Immunosuppressive Protocols

Protocols for immunosuppression of transplant recipients vary widely, depending on the transplantation center, type of organ transplanted, after transplantation, underlying cause of organ failure, and preexisting conditions (Box 31-5). Protocols are becoming more complex because of more immunosuppressive drug choices. In general, protocols include the following:

Box 31-5 Sample Protocol (Liver)

Intraoperative	Methylprednisolone
Day 0	Methylprednisolone,
Day 1	Prednisolone
Day 2	Taper
Days 0-5	Antilymphocyte globulin (ATG), IV; given until adequate cyclosporin A levels obtained
Days 0-5	Azathioprine IV
Day 6	Azathioprine, PO
Day ?	Cyclosporin A

From Tsunoda S: Update on immunosuppression, Boston, 2000, Tufts University School of Medicine Transplant Teleconference Series.

• Lymphokine synthesis inhibitors (e.g., cyclosporine, tacrolimus).

• Nucleoside synthetase inhibitors (e.g., azathioprine, mycophenolate mofetil).

• Steroids (e.g., prednisone).

• Induction or pretransplantation therapy—may include antithymocyte globulin, CD3 or CD25, or dacliximab

New Approaches in Immunosuppression

Survival after solid organ transplantation has increased in the era of tacrolimus and mycophenolate. These drugs have enhanced specificity and potency for T and B lymphocytes compared with their predecessors, cyclosporine and azathioprine. Between 2008 and 2010, the United States Food and Drug Administration approved several generic formulations of both tacrolimus and mycophenolate mofetil. Deciding whether generic products can be safely substituted for the innovator product is a clinical dilemma similar to that which occurred when generic formulations of cyclosporine became available.

• Cellular transplants

• Transgenic organs

• Development of chimerism

• Localized immunosuppression

• Prevention of chronic rejection

Post–Organ Transplantation Complications

Because complications are associated with transplantation, their early diagnosis and treatment are essential. The primary risks of transplantation are rejection and infection. Five other major complications of organ transplantation are cancer, osteoporosis, diabetes, hypertension, and hypercholesterolemia.

Infectious Diseases

Infections can be viral, such as CMV (80%), Epstein-Barr virus (20% to 30%), hepatitis B, or hepatitis C. Even rabies has been associated with organ transplantation. Other pathogens include Pneumocystis jiroveci (formerly known as P. carinii). Organisms associated with central nervous system infection in renal transplant recipients, in decreasing order of frequency, are Listeria, Cryptococcus, Mycobacterium, Nocardia, Aspergillus, Mucor, Toxoplasma, and Strongyloides spp. Published guidelines advise transplant teams to do the following to minimize transplant risk:

1. Screen for infectious disease agents in the donor and recipient before transplantation.

2. Culture and identify known and novel pathogens in recipients after transplantation.

3. Archive serologic samples before transplantation for identification of new infections later.

Cancer

Organ transplant recipients have a 20% greater risk of the development of cancer. The incidence of non-Hodgkin’s lymphoma is increased by 40%. The greatest risk for lymphoma is within the first 6 to 12 months after transplantation. Transplant recipients also have a greater risk of skin cancer and a slightly increased risk of cervical cancer. An increased risk of the development of cancer may be the result of chemotherapy and radiation therapy.

Osteoporosis

In the general population, osteoporosis affects one in four women and one in eight men. The general risk factors are age, postmenopausal state, sedentary lifestyle, and inadequate calcium intake. Transplant recipients are at an increased risk of developing osteoporosis because of pretransplantation immobility and long-term effects of steroid therapy. Regular bone density scanning should be a routine component of posttransplantation care.

Diabetes

Diabetes mellitus is a concern in two risk groups, patients with preexisting diabetes (25%) and those who develop diabetes after transplantation (20%). Patients with preexisting diabetes may require increased doses of insulin until stabilized on medications. Posttransplantation steroid-induced hyperglycemia can produce physiologic conditions that negatively affect a graft. Steroid medication might aggravate a familial tendency toward diabetes. The use of steroids results in decreased use of insulin by peripheral tissues, eventual insulin resistance with decreasing receptor sites, reduction in insulin production, and accelerated glycogenolysis by the liver to assist in glucose availability. These metabolic activities perpetuate hyperglycemia. In addition to threatening graft survival, diabetes can have other negative health consequences, such as adult blindness, vasculopathy, neuropathy, retinopathy, bladder infections, and a shortened lifespan.

Hypertension

An abnormal increase in blood pressure is usually a preexisting medical condition in transplant recipients. This condition is often associated with renal failure. Hypertension can negatively affect the patient’s general health and graft survival.

Hypercholesterolemia

An increased blood cholesterol is a serious posttransplantation concern because of long-term vascular effects to the patient and engrafted organ. Hypercholesterolemia can result from the return of the patient’s appetite and the lifting of dietary restrictions.

Xenotransplantation

Xenotransplantation is any procedure that involves the transplantation, implantation or infusion into a human recipient of either (a) live cells, tissues, or organs from a nonhuman animal source, or (b) human body fluids, cells, tissues or organs that have had ex vivo contact with live nonhuman animal cells, tissues or organs. The development of xenotransplantation is, in part, driven by the fact that the demand for human organs for clinical transplantation far exceeds the supply (Box 31-6). There is a global shortage of organs for transplantation. Pig heart valves are already used to repair human hearts, and porcine pancreatic islet cells are used to treat diabetes, so it is not a big leap to envision trans-species, whole-organ transplantation. Pigs are considered the most likely organ transplant donors for human beings because their organs are similar in size to human organs, they are easy to breed, and the extensive biologic differences between pigs and human beings make it unlikely for porcine diseases to infect human beings.

Box 31-6 Milestones in Xenotransplantation

1963-1964	Chimpanzee to human renal transplants
1964	Pig heart valve transplant
1968	Sheep heart transplant
1984	“Baby Fae” transplanted with a baboon heart
1992	Baboon to human liver transplant
1994	Pig pancreatic islets transplanted to insulin-dependent patients
1995	Neuronal cells from fetal pig transplanted to patients with Parkinson’s disease
1996	Baboon bone marrow transplanted to AIDS patient

Adapted from Wilde M: Rejection, retroviruses: major barriers to xenotransplantation, Adv Med Lab Prof 9:14–19, 1997.

Another application of cross-species organ use was successfully demonstrated in a phase I clinical trial that used transgenic pig livers as an ex vivo (outside the body) support system for patients with acute liver failure. The pig liver was used to bridge the gap between organ failure and obtaining an appropriate human liver for transplantation in these patients. Protocols are being developed for a phase I in vivo (inside the body) clinical trial.

Other procedures, some in clinical trials, use cells or tissues from other species to treat life-threatening illnesses such as cancer, AIDS, diabetes, liver failure, and Parkinson’s disease. Even if whole organs are not transplanted, animal cells or tissues will likely be used to treat many diseases. In 1995, physicians in California transplanted bone marrow from a baboon into an AIDS patient in a highly controversial procedure that prompted the creation of strict guidelines for transplantation by the FDA, National Institutes of Health (NIH), and Centers for Disease Control and Prevention (CDC).

Ethical and medical concerns surround xenotransplantation. Ethical concerns relate to selling organs. Donors may be paid as little as $1000 for a donated kidney in countries such as Brazil, India, or Moldova. A serious medical concern is the risk that transplanted tissue may carry unknown latent infections that once introduced into the recipient, could be activated and give rise to infection.

Biomarkers for Rejection

Emerging technologies, such as gene expression profiling, proteomics, metabolomics, and genomics, are rapidly advancing the pace of discovery of new biomarkers for rejection. These approaches are expected to generate improved diagnostic tests and knowledge that will lead to more effective therapies.

One of the most promising areas of transplant research, especially kidney transplantation, has been the discovery of biomarkers for rejection that are detectable in blood and urine. Biopsy-confirmed rejection, the current gold standard for diagnosis of allograft rejection, is invasive and subject to sampling errors. Development of noninvasive assays that detect molecular biomarkers for rejection could revolutionize the management of transplant recipients by the following:

• Detecting a prerejection profile that will allow therapeutic interventions before rejection causes graft dysfunction

• Improving the sensitivity and specificity of rejection diagnosis

• Developing new classification systems for rejection that will improve prognosis

• Providing information for designing individualized immunosuppressive regimens that could prevent rejection while minimizing drug toxicity

• Performing a Longitudinal Assessment of Posttransplant Immune Status (see Procedural Protocol)

FOXP3 mRNA

By studying concentrations of particular messenger RNAs (mRNAs) or proteins associated with immune activation or tissue stress, several gene products with altered expression in blood, urine, and biopsy tissue during rejection episodes have been identified. Urine concentrations of FOXP3 mRNA, a member of the forkhead family of cell differentiation genes and a lineage-specific transcript for graft-protecting regulatory T cells, can predict reversal of acute renal allograft rejection with high sensitivity and specificity.

Measurement of the products of individual genes such as FOXP3 probably will not supplant conventional biopsies for the diagnosis of rejection, but the development of panels of informative gene products in blood and urine, coupled with renal function and immune response markers, ultimately should achieve the sensitivities and specificities required for diagnosis and clinical management of kidney rejection.

Analyses of more than 1300 genes that were differentially expressed in kidney allografts have revealed three distinct molecular signatures of acute rejection that were more predictive of allograft survival than traditional histologic analysis. These data have also generated new hypotheses for the molecular mechanisms of rejection. For example, B cell infiltration is characteristic of aggressive acute rejection.

A new gene expression test, AlloMap (XDx, San Francisco), is being tested to explore its ability to predict acute cardiac allograft rejection. The test appears to detect the absence of moderate to severe cellular rejection, which might reduce the need for frequent biopsies.

CASE STUDY

Forty-year-old CG was seen by her family physician after several episodes of painless hematuria. On direct questioning, she complained of worsening malaise and swelling of her legs and hands over the previous 2 weeks. She also reported that despite a high fluid intake, she was urinating much less frequently than normal. She had no significant medical history.

On examination, the patient was pale and had generalized swelling of her extremities. Her temperature was 38.5° C (101° F) and her blood pressure was 160/110 mm Hg. She had no palpable masses or hepatosplenomegaly.

A diagnosis of idiopathic and rapidly progressive glomerulonephritis was made. She was given antihypertensive agents, corticosteroids, and azathioprine for 2 weeks, but her renal function deteriorated and end-stage renal failure was diagnosed. Hemodialysis was initiated.

In preparation for a possible renal transplant, she was tissue-typed for MHC antigens using anti-HLA antibodies. She was found to be HLA-A10, A28, B7, Bw52, Cw2, Cw6, DR2, DRw10, and blood group B positive. A suitable cadaveric kidney was found from a donor of HLA-A9, A28, B7, B17, Cw2, Cw6, DR2, DR4, and blood group B positive. A crossmatch of the patient’s serum with donor lymphocytes was satisfactory.

She underwent successful kidney transplantation. Her posttransplantation treatment was a combined triple-immunosuppressive regimen of prednisolone, cyclosporin A, and azathioprine. She progressed well immediately after transplantation.

Twelve days after engraftment, the patient developed a fever and was noted to be lethargic. Physical examination revealed generalized edema. Her blood pressure was 165/110 mm Hg. Her urine output had dropped significantly. A renal biopsy was performed. Histologic examination demonstrated significant interstitial mononuclear cell infiltration. This finding was consistent with the diagnosis of acute graft rejection. She was immediately treated with parenteral methylprednisolone. This treatment failed to improve her renal function and an antilymphocyte monoclonal antibody was administered. Her renal function improved and she was eventually discharged receiving cyclosporin A therapy.

Questions

1. Class II major histocompatibility complex (MHC) are encoded for in the ____ region.

a. A

b. B

c. C

d. D

2. In a kidney transplant, antibody to ABO antigens of donor tissue will produce ___________ rejection.

See Appendix A for the answers to multiple choice questions.

Critical Thinking Group Discussion Questions

1. What factors are important in matching donor to recipient in renal transplantation?

2. How does this patient’s graft rejection compare with other types of graft rejection?

See instructor site for the discussion of the answers to these questions.

• This assay assists in identification of transplant patients who are at risk of developing an infection due to over-immunosuppression. Periodic monitoring (Table 31-10) guides clinicians in making therapeutic decisions that avoid overimmunosuppression and underimmunosuppression.

Table 31-10

Patient Immune System Monitoring ^∗

Time	Interval
Pretransplant	Test as needed
Months 1-6	Test every 2 weeks
Months 7-12	Test monthly
After year 1	Perform routine monitoring (at minimum, test quarterly)