Pathology of Kidney Transplantation

Cynthia C. Nast

Arthur H. Cohen

The gold standard for assessing structural abnormalities in the transplanted kidney is standard tissue histopathology of a biopsy or transplant nephrectomy. Immunofluorescence also is necessary for identification of certain types of acute rejection, and electron microscopy may be required to evaluate glomerular lesions. Fine-needle aspiration with cytologic evaluation of cells aspirated from the graft using a thin needle has been used in the past to determine the cause of acute allograft dysfunction. However, the advent of thinner core needles with relative safety of the core biopsy procedure has minimized the use of aspiration cytology in the clinical setting.

CORE-NEEDLE BIOPSY

Indications and Technique

Kidney transplant biopsies are most frequently performed at times of graft dysfunction when the etiology cannot be accurately elucidated by clinical or noninvasive means. Protocol biopsies are performed at predetermined intervals after transplantation at some centers in an attempt to recognize so-called subclinical rejection (see Chapter 9); they also may be required as part of clinical trials for the evaluation of new immunosuppressive drugs (see Chapter 5). More precise clinical indications for biopsy are reviewed in Chapters 9 and 10. Transplantation programs vary in their reliance on biopsies and the clinical setting in which biopsies are performed.

Preparations for transplant biopsy are similar to those for biopsy of the native kidney. Informed consent is required from patients, who should be specifically warned of the risk for bleeding and occasional damage to the graft (see “Complications,” later). Before biopsy, coagulation studies are usually performed, although in the absence of liver disease, use of anticoagulants, thrombocytopenia, or a clinical history of bleeding, these may not be necessary. The blood pressure should be controlled at a level of less than 160/100 mm Hg.

The locations of the graft and biopsy site can be determined by palpation or by ultrasound guidance. A small pillow or towel rolled in the small of the patient’s back may facilitate palpation. Ultrasound offers the advantage of more precise localization of the graft and its depth and may reduce the frequency of inadequate specimens. Ultrasound may detect perinephric fluid collections or hydronephrosis. It is unwise to perform biopsy through a fluid collection because of the inability to tamponade the biopsy site adequately. Significant hydronephrosis should be relieved before the biopsy is performed because it may be the cause of the graft dysfunction; a small blood clot after the biopsy may exaggerate the degree of obstruction. Generally, the upper or lower pole of the transplant is sought, depending on which is more easily palpated or is nearer the surface. If the location of the biopsy site is difficult to ascertain or if the kidney is deep, it is wise to use real-time ultrasound with visual guidance or a fixed biopsy guide device (see Chapter 13).

Disposable automatic spring-loaded needles (18-gauge needles are usually adequate) have largely replaced the traditional modified 14-gauge Vim-Silverman needle and may be less traumatic to the kidney. The site chosen for the biopsy is locally anesthetized with 1% lidocaine, and a small stab wound in the skin is made to facilitate the passage of the needle. Precise instructions for use of the newer needles are provided in the package inserts. The needles are advanced up to the depth determined by ultrasound or until an increase in resistance is felt as the needle makes contact with the kidney. When the automatic needles are used, it may be advisable to withdraw the needle slightly before taking the sample to avoid excessive depth and ensure a cortical sample.

Two biopsy cores should be adequate. It is advisable to inspect the specimen immediately with a stereomicroscope to ensure adequacy. As soon as the needle is withdrawn, hemostasis should be augmented by manual compression or with a sandbag. Postbiopsy orders should include observation of the patient’s vital signs every 15 minutes for at least 2 hours and then hourly for several hours. Patients initially should be immobile; in the absence of macroscopic hematuria, ambulation can begin after 6 to 8 hours. Many transplantation centers permit outpatients to go home the same day the biopsy is performed.

Complications

Core needle biopsy is an invasive technique and is not risk free; these risks must be weighed against the benefit gained from the information obtained from the procedure. Careful assessment of potential risks and benefits must precede every decision to subject a patient to a biopsy.

All major complications after needle biopsy manifest as perinephric or urinary bleeding. Transient macroscopic hematuria is common and is of little clinical significance. Macroscopic hematuria follows about 3% of biopsies and may prolong hospitalization or lead to blood transfusion or placement of a bladder catheter for clot drainage. Ureteral obstruction occasionally occurs, requiring placement of a percutaneous nephrostomy; massive hemorrhage necessitating surgical exploration, graft nephrectomy, or angiographic embolization is rare. Postbiopsy arteriovenous fistulas sometimes may be detected by Doppler ultrasound and usually can be treated expectantly. Angiographic embolization may occasionally be required, and graft loss has been reported.

Specimen Handling

Detailed methods for handling tissue specimens are beyond the scope of this chapter. For all specimens, portions are obtained for each of the three traditional methods of evaluating renal parenchyma: light microscopy, immunofluorescence, and electron microscopy. For the initial biopsy, all methods should be used; for subsequent biopsies, electron microscopy is performed only if indicated. This approach allows the pathologist to obtain maximal diagnostic and prognostic information. In selected instances, rapid processing or frozen sections can be performed on the tissue placed in fixative for light microscopy when an immediate assessment of the changes in the graft is necessary for initiating or modifying therapy.

TRANSPLANT REJECTION

Traditionally, three major forms of rejection are recognized: hyperacute, acute, and chronic. Each has reasonably distinctive changes, although acute and chronic rejection may be present simultaneously, resulting in a mixture of histopathologic features. Table 14.1 lists the pathologic findings in the major lesions responsible for functional impairment of the graft.

TABLE 14.1 Histopathologic Findings in the Major Causes of Allograft Dysfunction

Type	Interstitium	Tubules	Glomeruli	Arteries
Acute T-cell mediated rejection	Edema, lymphocytes	Lymphocytes, cell degeneration	Capillary lymphocytes	Swollen endothelium, intimal lymphocytes, foam cells
Arterial acute antibody-mediated rejection	Hemorrhage, zonal infarction, PTC C4d	Necrosis	Neutrophils, thrombosis	Necrosis, neutrphils, thrombosis
Microvascular (C4d+) acute antibody-mediated rejection	PTC C4d ± PTC neutrophils, monocytes	± Necrosis	Neutrophils, monocytes	Normal
Acute tubular necrosis	Edema	Cell degeneration, necrosis, mitoses	Normal	Normal
Acute calcineurin inhibitor toxicity	Edema	Isometric vacuoles, cell degeneration	Normal	Normal
Chronic rejection	Fibrosis, lymphocytes	Atrophy, dropout	Chronic transplant glomerulopathy	Fibrosis, lymphocytes narrowed lumina
Chronic calcineurin inhibitor toxicity	“Striped” fibrosis	Atrophy	Ischemic collapse	Arteriolopathy, hyalinization
PTC, peritubular capillary.

Hyperacute Rejection

Hyperacute rejection is produced by preformed cytotoxic antibodies and is an infrequent event so long as the pretransplantation crossmatch is negative (see Chapters 3 and 7). It may manifest shortly after vascular anastomoses are established, or it may be delayed up to 3 days. It is characterized by rapid and widespread vascular thrombosis, predominantly affecting arteries, arterioles, and glomeruli, often with polymorphonuclear leukocytes incorporated in the thrombi. The kidney is usually cyanotic, slightly edematous, and flaccid, and urine production suddenly ceases or does not begin at all. If the kidney is not removed immediately, extensive cellular necrosis ensues, followed after 24 hours by numerous cortical and medullary infarcts. Immunofluorescence may disclose capillary and arterial wall immunoglobulin G (IgG) or IgM, C3, and fibrin, with fibrin also in the thrombi. Peritubular capillary C4d deposition occurs after 48 to 72 hours if the kidney remains viable during this time. Electron microscopy in the early lesions indicates degeneration and early necrosis of vascular endothelium.

Hyperacute rejection needs to be differentiated from other circumstances in which extensive vascular thrombi occur. The differential diagnosis includes physical perfusion-related injury to vascular endothelium and injury caused by cold-reacting IgM antibodies against blood cells. Both of these conditions
rarely may manifest in the immediate post-transplantation period and may produce entrapment of leukocytes in thrombi. It is only in hyperacute rejection, however, that polymorphonuclear leukocytes are typically and regularly incorporated in the thrombi. Recurrent hemolytic uremic syndrome and a thrombotic microangiopathy associated with administration of the calcineurin inhibitors (discussed later under “Calcineurin Inhibitor Nephrotoxicity”) are characterized by thrombi, usually without leukocytes, and are generally lateroccurring lesions.

Acute Rejection

When the term acute rejection is used, it typically refers to acute cell-mediated rejection. However, two distinct immunopathologic mechanisms are responsible for acute rejection: cell-mediated immunity and antibody-mediated (humoral) immunity. It is critical to differentiate the processes.

Cell-Mediated Acute Rejection

Cell-mediated acute rejection is the most common form of early rejection and has tubulointerstitial and vascular forms. Light microscopy and immunofluorescence microscopy with C4d immunostaining (see “Antibody-Mediated Acute Rejection and the C4d Stain”) are the major procedures used in diagnosing these lesions and in all cases of graft dysfunction. At times, routine immunofluorescence and electron microscopic evaluation may be helpful for the differential diagnosis. In tubulointerstitial cell-mediated acute rejection, primary abnormalities are in the interstitium, which is diffusely edematous and infiltrated by numerous leukocytes, most of which are mature and transformed lymphocytes (CD4, CD8), with fewer monocytes and plasma cells (Fig. 14.1). Eosinophils are either absent or found focally in small numbers; polymorphonuclear leukocytes are not a regular feature. Peritubular capillaries are dilated and contain lymphocytes that may be seen migrating into the interstitium. A characteristic lesion, called tubulitis, occurs, whereby lymphocytes and monocytes
extend into the walls and lumina of tubules, with associated degenerative changes of tubular epithelial cells. The cells and basement membranes of tubular walls may be damaged and discontinuous. When this lesion affects cast-containing distal tubules, cast matrix (Tamm-Horsfall protein) may be found in the interstitium and occasionally in peritubular capillaries and small veins. For tubulitis to have diagnostic significance, the inflammation should be documented in normal (nonatrophied) tubules. The significance of tubulitis solely in atrophied tubules is not known.

FIGURE 14.1 Acute cell-mediated tubulo-interstitial rejection. There is interstitial edema with lymphocytes in both the interstitium and tubular walls in association with tubular cell degeneration. (Periodic acid-methenamine silver stain, ×200.)

In cell-mediated vascular rejection, lymphocytes, monocytes, and less often foam cells undermine arterial endothelium and are found in the vascular intima, but rarely extend into the muscularis (Fig. 14.2). Endothelial cells are swollen, often vacuolated, and detached from the vacular wall, but arterial wall necrosis is not a feature of this type of acute rejection. This form of vascular rejection often occurs in concert with tubulointerstitial rejection. Acute transplant glomerulopathy is a form of glomerular cell-mediated rejection in which lymphocytes and monocytes accumulate in glomerular capillary lumina and mesangial regions (Fig. 14.3). Endothelial and mesangial cells are swollen, and capillary walls display subendothelial lucencies, with occasional segmental peripheral mesangial migration and interposition on ultrastructural examination. In biopsies demonstrating cellular rejection, immunofluorescence may disclose fibrin in the interstitium; segmental linear or granular IgM, C3, and fibrin may be found in glomerular capillary walls in acute transplant glomerulopathy. C4d staining is negative within peritubular capillaries. Ultrastructural examination usually confirms the light
microscopic findings and provides additional diagnostic information only for the glomerular lesion. When acute cell-mediated rejection is treated successfully, the interstitial inflammatory infiltrate diminishes rapidly, whereas edema, tubular inflammation, and tubular cell damage may persist for some time.

FIGURE 14.2 Acute cell-mediated vascular rejection. A small artery contains lymphocytes in the lumen and in the intima beneath swollen endothelial cells. Note the interstitial edema and infiltration by lymphocytes, which are also in the walls of tubules. (Periodic acid-Schiff stain, ×220.)

FIGURE 14.3 Acute transplant glomerulopathy. Glomerular capillary lumens contain monocytes and lymphocytes. There is also tubulo-interstitial rejection with interstitial edema, and lymphocytes in the interstitium and tubular walls. (Periodic acid-methenamine silver stain, ×200.)

Antibody-Mediated Acute Rejection and the C4d Stain

There are two types of antibody-mediated acute rejection: the classic arterial type and a more common type that is also C4d positive but without vascular involvement. The vascular form is an uncommon type of rejection and is characterized primarily by necrotizing arteritis, with mural fibrinoid necrosis and variable inflammation in the artery wall, including lymphocytes, monocytes, and neutrophils (Fig. 14.4). Endothelial cells are severely damaged or absent, and luminal thrombosis is common. This lesion typically results in cortical infarction with focal interstitial hemorrhage. Although the hyperacute rejection described previously is also antibody mediated, it differs from antibody-mediated vascular rejection in that it does not have an inflammatory or fibrinoid component in the vessel walls at its outset.

In the arterial form of antibody-mediated rejection, immunofluorescence discloses IgG and sometimes IgM accompanied by C3 in the walls of arteries. In these structures, fibrin may be intramural and intraluminal and also may be in the interstitium when hemorrhage is present. In cell-mediated vascular rejection, there is no antibody component; therefore, vascular rejection is an imprecise term that signifies merely inflammation of arteries, which can result from either cell-mediated or antibody-mediated immunity. When arterial inflammation is present, it is important to further categorize the rejection process to indicate the etiologic mechanism because appropriate therapy and prognosis differ. The antibody-mediated form is characterized by arterial mural necrosis, neutrophilic infiltrate, and luminal thrombosis and represents a more severe lesion that is poorly responsive to therapy.