How to Perform Liver Biopsies

When investigating chronic liver disease, a cytological or histopathological assessment of liver tissue is frequently required. Prior to invasive sampling, it is necessary to exclude extrahepatic causes of elevated serum enzyme activities (e.g., due to endocrinopathies), extrahepatic causes of bilirubin elevation (e.g., due to sepsis), and extrahepatic causes of structural changes of the liver (e.g., due to steroid administration). Portosystemic vascular anomalies should be ruled out as well. If structural focal liver changes are present, preference should be given to less invasive technique such as contrast-enhanced ultrasonographic examination over a liver biopsy to differentiate between benign and malignant nodes. Postoperative bleeding is the most common complication of liver biopsy. In order to minimize the risk of bleeding, biopsy can be performed when following hemostatic parameters are determined: platelet count (>100 000 platelets/µl), platelet function (mucosal bleeding time <4 min), fibrinogen concentration (>50% of the lower limit of the reference interval), and coagulation times (<1.5 fold prolonged).

Various techniques for invasive liver sampling can be employed:

1.  Fine needle aspiration (FNA) biopsy of the liver

2a. Ultrasound­ assisted needle biopsy (Tru-Cut biopsy) of the liver

2b. Laparoscopic or surgical biopsy of the liver

1. Fine Needle Aspiration (FNA) Biopsy of the Liver

FNA is a simple and inexpensive method for investigating liver diseases. Due to its low complication rate, it is often the first invasive diagnostic step.

The following changes are usually being detected.

A) Hepatic Lipidosis

Hepatic lipidosis, typically with vacuolization of >50% hepatocytes can be easily detected on cytology. Hepatic lipidosis occurs usually during the course of another disease due to anorexia. It is sometimes possible that hepatic lipidosis is diagnosed by the cytological examination of a liver aspirate, but the inciting cause, for example an infiltrative hepatic disease, is not being recognized.

B) Bacterial and Parasitic Infections of the Liver and the Bile Duct

In cats, neutrophilic cholangitis is a common cause of bile duct inflammation due to bacterial infection arising from the intestinal tract. Cytological examination and culture (including antibiogram) of the bile is necessary for an exact diagnosis and for the optimization of treatment. An ultrasound-guided gallbladder aspiration is a safe method when specific rules are being observed. Gallbladder should be well filled and should be located close to the ultrasound probe. Patient should be anesthetized, so that movement does not occur during the aspiration.

Additionally, a second person should aspirate the bile through an extension tube, so that the person performing the aspiration can fully concentrate on the procedure, and ensure that the needle moves as little as possible. Furthermore, gallbladder should be completely emptied, so that the intravesical pressure is low, and the risk of a bile leakage through the puncture site is minimized. Analgesic administration is recommended before aspiration, and an intravenous broad-spectrum antibiotic administration is indicated after the aspiration in case of suspected bacterial cholangitis. Bacteria or other infectious agents (e.g., Echinococcus sp.) can also be detected on liver cytology.

C) Neoplasia

Round cell neoplasms such as lymphoma and mast cell tumor are easily diagnosed on liver cytology. Primary hepatic epithelial neoplasms (i.e., hepatocellular and cholangiocellular carcinoma) can sometimes pose a diagnostic challenge. Hepatocytes from hepatocellular adenoma and some well-differentiated hepatocellular carcinomas cannot be differentiated from normal hepatocytes or hepatocytes in hyperplastic or regenerative nodules. Therefore, in cases of solid neoplasia, a histological examination is often necessary for the assessment of infiltration and for definitive diagnosis. Mesenchymal neoplasms (hepatic sarcomas) often exfoliate poorly, resulting in low numbers of cells available for cytological evaluation. If non-diagnostic FNA samples are obtained, liver disease cannot be completely ruled out. In these cases, it is recommended to obtain biopsy samples for histopathological assessment.

Histopathological examination is often necessary to diagnose inflammatory, neoplastic, fibrotic and degenerative changes in the liver tissue. Important parenchymal liver diseases such as chronic hepatitis in the dog and various cholangitis forms in the cat can only be recognized from an adequate liver biopsy.

2a: Ultrasound-Assisted Needle Biopsy (Tru-Cut Biopsy) of the Liver

This procedure can be performed in sedated patients, is considered a low-risk and is cheap. Tru-Cut biopsy is contraindicated when a small, ultrasonographically poorly recognizable liver or abdominal effusion are detected on ultrasonographic examination. High-spring shot biopsy should not be used in cats, as the strong impulses can lead to a vagal shock reaction with a fatal outcome. In case of vacuolar hepatopathy, inflammatory disease and in some neoplasms, diagnosis by needle biopsy can often be made. Ultrasound-assisted biopsy is helpful in intrahepatic focal changes that are not visible from the liver surface. These lesions can be selectively biopsied under ultrasonographic control.

Biopsy sampling should be done in several places and with a 14–16 gauge biopsy needle. It has been shown that the histopathological correlation between surgical biopsies (taken laparoscopically as the gold standard) and Tru-Cut biopsies can be less than 50%, some of the reasons being the size of the sample and lack of agreement between pathologists. The size of the obtained standard needle biopsy specimen is limited to approximately 0.002% of the total hepatic parenchyma.

2b: Laparoscopic or Surgical Biopsy of the Liver

The assessment of larger tissue sample removed surgically or laparoscopically is considered a gold standard. It is unavoidable in situations when information is required regarding:

A.  Type of liver change (inflammatory, neoplastic, vacuolar, and fibrotic)

B.  Severity of liver disease (mild, moderate, severe)

C.  Duration of the disease (acute, chronic)

D.  Growth pattern of neoplasia (well-defined, infiltrating into surrounding tissue)

E.  Special investigation (special stains, fluorescence in situ hybridization and PCR testing for infectious agents, culture of liver tissue).

Compared to the ultrasound-guided sampling, the advantage of laparoscopic or surgical biopsy is the possibility to assess the liver surface macroscopically and to obtain larger biopsy sections for the exact evaluation of the liver architecture. Tru-Cut biopsies are often too small to be able to assess, for example, the extent of fibrosis or the invasiveness of tumor growth (adenoma versus adenocarcinoma).

Laparoscopy is considered a less invasive method than the surgical biopsy. During the surgical biopsy, however, post-bleeding can be directly controlled and direct therapeutic measures such as tumor or abscess removal or release of posthepatic obstruction can be carried out. When a diffuse liver disease is present, biopsies should be obtained laparoscopically from approximately 7–8 or surgically from approximately 3–4 areas. Five biopsy samples should be placed in formalin, two samples should be frozen (-20°C for quantitative copper determination and special tests) and one sample should be placed in a culture medium for bacteriological examination. Bile should also be aspirated from the gallbladder during each procedure, and sent for cytological and microbiological examination. In summary, liver biopsy sampling under specified conditions is a safe method which provides important information about the liver. The most appropriate biopsy method should be decided on a case-by-case basis and the benefit must be weighed against the risks.