Biopsy Interpretation of the Liver, 2nd ed

3. Technical Considerations

 

Liver biopsy samples needed for study can be obtained percutaneously as a core with a biopsy needle, as a wedge or core either during open laparotomy or with the laparoscope, or with a biopsy forceps inserted through the jugular vein. Alternatively, fine-needle aspiration can be used. Each method has limitations; therefore, to avoid an unnecessary procedure or forestall the preparation of a sample that cannot provide the needed answer, the purpose of the biopsy should be determined before the method is chosen (1).

Percutaneous needle biopsy technique has proven to be the most useful method for obtaining liver tissue representative of diffuse liver disease and has been the standard for almost 50 years. Core needle biopsy does not provide the sought-after answer in all cases. Although the special utility of liver biopsy is based on the fact that the liver tends to react uniformly to a great variety of injuries and stimuli, it must be remembered that sampling error can occur in various hepatic disorders. For example, the changes of primary sclerosing cholangitis are not uniformly seen throughout the liver. Primary sclerosing cholangitis can usually be diagnosed with imaging studies, whereas a series of biopsies may fail to demonstrate typical changes. Similarly, characteristic inclusions of cytomegalovirus, in either the posttransplantation patient or the acquired immune deficiency syndrome patient, may not be seen in every section of liver prepared. Several serial sections may be required or a more sensitive technique, such as DNA in situ hybridization, may be required. Of course, tumor nodules, either deep or superficial, can be missed by the relatively blind percutaneous technique, even when performed with the aid of computed tomography.

SIZE MATTERS

An adequate specimen is necessary for the evaluation of diffuse liver disease. Hepatitis C patient biopsies are the most common. Grading and staging, whether using English language diagnosis or scoring system or both, is the standard of practice, used to determine the need for therapy as well as to evaluate prognosis. Ideally, the liver core sample should be obtained with a 16- or 14-gauge needle. The core should be at least 2.0 cm in total length and should include at least 10 complete portal tracts (e-Fig. 3.1).

Even with diffuse disease, percutaneous biopsy not meeting these criteria can prove insufficient. There is considerable difference in the samples obtained with biopsy needles because the needles themselves differ in many aspects. Biopsy needles vary in terms of the length of the sample obtained, the diameter of the sample obtained, and the degree of compression artifact. For example, smaller biopsy needles may not obtain sufficient numbers of portal tracts to appropriately evaluate the immune biliary disorders. In cirrhosis, a small needle may not obtain any septal structures and may yield only fragments of parenchyma, contributing to difficulty in establishing the correct diagnosis.

In our experience wedge biopsy, in the absence of a grossly recognizable lesion, and prompted by what the surgeon perceives to be an abnormal-appearing liver, is often disappointing. Wedge biopsy is the technique of choice for a focal lesion presenting at or immediately below the capsule. If a primary, generalized liver disorder is suspected, a needle biopsy should be performed, even in the intraoperative setting. In wedge biopsy samples, (a) the capsule may be thickened and relatively little diagnostically useful parenchyma obtained; (2) (b) capsule fibrosis extending into the liver may mimic cirrhosis (documentation of features clearly indicative of regeneration of hepatocytes is required); (c) there may be clusters of chronic inflammatory cells immediately below the capsule that can be misinterpreted as evidence of chronic hepatitis; and (d) acute inflammatory cells may be prominent throughout the lobule as a nonspecific sequel to laparotomy. Bile duct hamartomas (von Meyenburg complex) are relatively common and often multiple. They may attract the attention of the surgeon who will biopsy them suspecting metastasis. The lesions can then be misinterpreted by the unwary pathologist as metastatic carcinoma.

The principal indication for laparoscopic biopsy is to diagnose a focal lesion not easily obtained, because of location or size, with standard percutaneous liver biopsy or with computed tomography-guided aspiration. In the absence of a distinct nodule or mass, core needle biopsy should be obtained rather than wedge biopsy.

Transjugular biopsy is generally used for patients with generalized liver disease who have marked ascites or for those with severe thrombocytopenia or another coagulation disorder. The specimen obtained with transjugular biopsy forceps is often markedly fragmented and portal tracts may be absent, but the technique for obtaining core biopsies is available and the samples can be excellent (3,4,5). It is an excellent technique for the confirmation of a predominantly hepatocytic disorder.

Aspiration biopsy samples obviously do not demonstrate architectural changes, which are key in the recognition of many disorders. They have also proven to be of limited use in the diagnosis of diffuse inflammatory disorders of the liver and are most often obtained for the evaluation of a space-occupying lesion.

CARE AND HANDLING OF THE LIVER BIOPSY SPECIMEN

Although prompt fixation of the liver biopsy is vital to ensure optimal quality of the histologic section, forethought about the goals of the biopsy, including consultation by the clinician with the pathologist, can help to guarantee that all studies needed for diagnosis can be performed without the necessity of a second biopsy performed solely for the purpose of obtaining appropriately prepared tissue.

Formalin, the most common fixative used, is exceedingly stable, penetrates and adequately fixes tissues well, and is the most economical of fixatives. Furthermore, formalin allows for the subsequent application of many histochemical, immunohistochemical, and molecular procedures. The characteristics of tissues fixed in formalin are well known throughout the world, and the cytologic alterations subsequent to fixation are familiar in most settings.

In general, the pathologist decides on the fixative with which he or she is most comfortable; most hepatopathologists prefer fixation with 4% neutral buffered formalin. The core needle biopsy requires at least 2 to 3 hours for adequate fixation, although microwave processing can be used to reduce fixation time. The wedge biopsy can require as long as 12 hours for fixation, unless it is immediately sectioned into 1- to 2-mm-thick portions, in which case satisfactory fixation can be obtained in shorter periods.

The hepatologist or surgeon should be encouraged to ask certain questions prior to immersion of the tissue in fixative: Do I suspect a lymphoma? If so, I may want to obtain a fixative favored for the study of hematologic disorders. Will it also be useful to freeze tissues for immunohistochemical study of lymphocyte markers? Do I suspect Wilson disease? I may want to prepare tissue for electron microscopy. Do I suspect a bacterial disease? I may want to submit a small portion of tissue for microbiologic studies. Do I want to confirm the presence of hepatitis C, Epstein-Barr virus, or other viruses? I may want to save fresh tissue, or use formalin-fixed tissue, for polymerase chain reaction. Do I suspect a metabolic disorder? Which metabolic disorder? I may want to use more than one type of fixative. I may want to fresh-freeze some tissue for specialized histochemical studies as well as fix tissue for electron microscopy. Do I suspect a neoplasm? I may want a “touch print” for cytopathology studies. What kind of neoplasm? Do I need specialized histochemistry? Do I need electron microscopy?

Do I need to talk to the pathologist before I perform the biopsy? The correct answer to this question is often “yes.”

Special Handling of Liver Biopsies and Aspirates

Too often the pathologist is asked to provide a definitive diagnosis on tissue that has been inappropriately obtained and prepared.

First, don't let the specimen dry or actively contribute to its drying! Biopsy samples should not be placed on dry gauze, paper towel, or cloth towel, all of which will dehydrate the individual hepatocytes (Fig. 3.1, e-Figs. 3.2-3.5). The sample should be kept on saline-moisturized Telfa gauze. However, biopsy samples should never be immersed in saline; this leads to artifactual hepatocyte swelling and disruption of architecture (Fig. 3.2, e-Figs. 3.6, 3.7). Samples for microbiologic studies should be obtained without drying the specimen; dry culture swabs also dehydrate cells and affect morphology. A portion of the biopsy core should be submitted separately for this purpose. Exposure to room temperature for more than a few minutes causes specimen drying. If fixation cannot be done immediately, the tissue should be kept cool either by placing it in a closed container in a refrigerator, not a freezer, or by placing a jar containing the tissue in a larger container with ice-cold water or ice cubes, not dry ice. A variant of drying artifact is sometimes seen in wedge biopsy because of cautery effect in which the sinusoids become irregularly dilated (Fig. 3.3).

FIGURE 3.1 Drying artifact. Both samples are from the same case. A was promptly immersed in formalin. B was placed on a dry non-Telfa gauze for approximately 1 minute. Note the artifactually increased eosinophilia in B, as well as what appears to be increased nuclear cytoplasmic ratio (hematoxylin-eosin, original ×100).

FIGURE 3.2 Image showing dissolution of liver architecture because of immersion of the specimen in saline for more than 15 minutes (hematoxylin-eosin, original ×200.)

Second, prompt fixation is vital! If special biochemical studies, such as those for metabolic disorders, are needed, a portion of the biopsy should be immediately placed in the -70°C freezer or in an isopentane holding container. Tissue can also be placed in an appropriate support medium, such as Zeus medium, and then frozen.

Third, various artifactual changes can be seen in the tissue section, most of which are easily recognized (e-Figs. 3.8-3.11).

Fourth, fine-needle aspiration specimens must also be handled promptly. Delay causes artifactual changes because of drying and can prevent accurate evaluation. The most useful portion often is in the needle hub, which should be flushed after preparation of the smear. A clot in the needle hub, where it joins the syringe, can be recovered by mechanically scraping it free with either a new needle or an orangewood stick so as to prepare a “cell button” for paraffin embedding and sectioning (1).

FIGURE 3.3 Cautery artifact. Sinusoidal spaces are irregularly and haphazardly dilated, and hepatocytes have irregular forms with loss of cytoplasm and relatively nuclear enlargement (hematoxylin-eosin, original ×100).

REFERENCES

1. Geller SA, Pitman MD, Nichols WS. Cellular and molecular diagnostic techniques. In: Burt AD, Portmann BC, Ferrell LD, et al., eds. MacSween's Pathology of the Liver. 5th Ed. London: Churchill Livingstone, 2007:119-146.

2. Petrelli M, Scheuer PJ. Variation in subcapsular liver structure and its significance in the interpretation of wedge biopsies. J Clin Pathol 1967;20:743-748.

3. De Hoyos A, Loredo ML, Martinez-Rios MA, et al. Transjugular biopsy in 52 patients with an automated Tru-cut-type needle. Dig Dis Sci 1999;44:177-180.

4. Mammen T, Keshava SN, Eapen CE, et al. Transjugular liver biopsy: a retrospective analysis of 601 cases. J Vasc Interv Radiol 2008;19:351-358.

5. Soyer P, Fargeaudou Y, Boudiaf M, et al. Transjugular liver biopsy using ultrasonographic guidance for jugular vein puncture and an automated device for hepatic tissue sampling: a retrospective analysis of 200 consecutive cases. Abdom Imaging 2008;33:627-632.



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