Journal of Cytology
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Year : 2013  |  Volume : 30  |  Issue : 2  |  Page : 116-120
Cytomorphological variables of hepatic malignancies in fine needle aspiration smears with special reference to grading of hepatocellular carcinoma

1 Department of Pathology, Gandhi Medical College, Bhopal, Madhya Pradesh, India
2 Department of Radiodiagnosis, Gandhi Medical College, Bhopal, Madhya Pradesh, India

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Date of Web Publication29-May-2013


Background: The evaluation and management of discrete hepatic masses is a clinical problem. Ultrasound guided fine needle aspiration cytology (FNAC) is a rapid, accurate and safe diagnostic procedure that can be used in various neoplastic and non-neoplastic diseases of the liver.
Aim: To evaluate the cytomorphological features of hepatocellular carcinoma.
Materials and Methods: Ultrasound-guided fine needle aspiration was performed on 52 patients clinically suspected of having hepatic lesion.
Results: Malignancy was detected in 50 cases. The primary malignancies consisted of 15 cases of hepatocellular carcinoma (HCC), 4 cases of cholangiocarcinoma and 2 of hepatoblastoma. There were 29 metastatic lesions, which included 26 cases of adenocarcinoma, 2 cases of squamous cell carcinoma and 1 of lymphoma. The key diagnostic features for HCC were trabecular arrangement, polygonal cells with eosinophilic and granular cytoplasm, hyperchromatic nuclei with macronucleoli. Atypical naked hepatocytic nuclei and malignant cells separated by sinusoidal capillaries were also commonly seen. The sensitivity and specificity of FNA for malignancies was found to be 96% and 100% respectively, yielding a positive predictive value of 100% and negative predictive value of 50%.
Conclusions: There was a very good correlation of cytological criteria and histopathological diagnosis in hepatocellular carcinoma of various grades as well as in differentiating primary from metastatic tumors. It is recommended that image directed FNAC should be the primary diagnostic modality for assessing potential malignancy in any patient with a localized hepatic mass.

Keywords: Fine needle aspiration cytology; grading; hepatocellular carcinoma; metastatic; ultrasonography guided.

How to cite this article:
Balani S, Malik R, Malik R, Kapoor N. Cytomorphological variables of hepatic malignancies in fine needle aspiration smears with special reference to grading of hepatocellular carcinoma. J Cytol 2013;30:116-20

How to cite this URL:
Balani S, Malik R, Malik R, Kapoor N. Cytomorphological variables of hepatic malignancies in fine needle aspiration smears with special reference to grading of hepatocellular carcinoma. J Cytol [serial online] 2013 [cited 2023 Mar 28];30:116-20. Available from:

   Introduction Top

The evaluation and management of discrete hepatic masses is a common clinical problem. [1] The differential diagnosis of hepatic mass lesions includes primary liver tumors, metastatic deposits, congenital and acquired cyst, abscesses and granulomas. [2] Ultrasound-guided fine needle aspiration cytology (FNAC) is a rapid, accurate and safe diagnostic procedure that can be used in diagnosing various neoplastic and non-neoplastic diseases of the liver. [3] The advantages of this technique are its high diagnostic accuracy and low cost, thereby rendering the older technique of blind percutaneous biopsy using a coarse needle obsolete. [4]

The aims and objectives of the study were grading of primary hepatocellular carcinoma (HCC) on the basis of cytomorphological variables, differentiation of primary hepatocellular carcinoma from metastatic carcinoma on the basis of cytological criteria, evaluating the technique and inadequacy rate of aspirated smearsand comparison of cytomorphological features of HCC with metastatic lesions.

   Materials and Methods Top

Ultrasound-guided (USG) percutaneous fine needle aspiration was performed on 52 patients (28 males and 24 females) with ultrasonographically suspected focal or diffuse neoplastic lesion of the liver.

Fine needle aspirations were performed by the radiologist with 22 gauge needle for superficial lesions and 18 gauge spinal needle for deep-seated lesions, attached to 20 mL disposable plastic syringe. Taking all aseptic precautions, area was cleaned, and during suspended respiration, needle was introduced percutaneously into the lesion evaluated by ultrasound. Four to five smears were made and immediately fixed in 95% alcohol for 15-20 minutes and then stained with Papanicolaou stain.

The final diagnosis was made by histological confirmation of percutaneous liver biopsies and surgical biopsies and clinical correlation with laboratory (liver function tests specially serum bilirubin and alkaline phosphatase, hepatitis B and hepatitis C viral markers and tumor markers like alfa-fetoprotein, carcino-embryonic antigen (CEA) etc.) and ultrasound findings.

   Results Top

There were 52 patients with a mean age of 48 years. More than 50% of cases were between 30 and 60 years of age. Twenty eight cases (53.8%) were males and 24 (46.2%) were females.

Out of the 52 aspirates, 46 (88.5%) were malignant, 1 (1.9%) was benign, 3 (5.8%) were non-representative, as it contained blood with few scattered hepatocytes, and 2 (3.8%) were suspicious of malignancy. In smears typed as suspicious, very few atypical cells were seen [Table 1].
Table 1: Distribution of liver aspirates

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On FNAC, in 26 cases (56.5%), metastatic tumors were diagnosed. The most common metastatic tumor was adenocarcinoma found in 24 cases (92.3%) with one case (3.8%) each of squamous cell carcinoma and lymphoma. Nine cases were metastases from gall bladder, eight from colon, one each from breast and prostate. In seven cases, diagnosis of metastatic adenocarcinoma was given, but primary site could not be ascertained.

The commonest primary tumor was HCC diagnosed in 14 cases (30.4%). Based on cytological features, HCC was differentiated into 3 grades. All the cases of HCC were confirmed histologically.

In grade I/well differentiated HCC, tumor cells resembled hepatocytes with abundant and granular cytoplasm, central round nuclei and slightly increased nucleocytoplasmic ratio. Chromatin was coarse granular, and prominent nucleoli (usually single) were present [Figure 1]a.
Figure 1: FNAC of liver: (a) Tumor cells from grade I HCC arranged in trabecular pattern as well as scattered singly. Cell cords are 3-4 cell thick. Cells show bile, hyperchromasia, moderate granular cytoplasm, slightly increase neucleocytoplasmic ratio and 1 - 2 prominent nucleoli. (Pap, ×400). (b) Grade II HCC - central and peripherally placed nuclei, coarse granular chromatin (Pap, ×400). (c) Grade III HCC - pleomorphism, giant cells, abnormal mitoses and naked nuclei (Pap, ×400). (d) Cholangiocarcinoma - small cells with mild anisonucleosis with micro acinar pattern (Pap, ×400)

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Grade II/moderately differentiated HCC showed tumor cells with less abundant cytoplasm, high nucleocytoplasmic ratio and round to ovoid nuclei located peripherally. Chromatin was coarsely granular and unevenly distributed, and nucleoli were prominent. Binucleation was common, and naked hepatocytic nuclei were also seen [Figure 1]b.

Grade III HCC showed two distinct patterns - poorly differentiated and pleomorphic. Poorly differentiated cells showed no resemblance to hepatocytes, having relatively scanty cytoplasm. Nuclei showed variation in size and shape, having granular chromatin. Nucleoli were irregular and sometimes several. Atypical naked hepatocytic nuclei were also seen.

Pleomorphic type showed large cells with abundant cytoplasm, irregular and eccentric nuclei and prominent nucleoli. Multinucleated tumor giant cells were frequently observed. Rare or no trabeculae or bile was seen [Figure 1]c.

Six cases (42.8%) of HCC were grade III followed by 4 cases (28.6%) each of grade I and grade II HCC.

The other primary liver tumors were cholangiocarcinoma 4 cases (8.7%) and 02 cases (4.3%) of hepatoblastoma. Cytological features in cholangiocarcinoma were small cells having scant cytoplasm and ovoid hyperchromatic nuclei arranged in loose clusters as well as scattered singly. Prominent nucleolus was not a constant feature [Figure 1]d.

The cytological features of hepatoblastoma were different in two cases, as one was fetal type and other was anaplastic type. Fetal type or highly differentiated hepatoblastoma showed cells resembling normal hepatocytes with rare mitoses arranged in 2 or 3 cell thick tumor cords. Anaplastic type or small cell undifferentiated type showed small cells with scant cytoplasm and densely stained nuclei. Rosette formation was also seen [Figure 2].
Figure 2: FNAC smear with cluster of tumor cells from hepatoblastoma showing fetal hepatocytes on left and small undifferentiated tumor cells forming rosettes on right. (Pap, ×100)

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The cytological diagnosis correlated well with the histopathological diagnosis in all the 46 cases of malignancy. Cases which were suspicious of malignancy on cytology were confirmed by histopathology. One was diagnosed as metastatic squamous cell carcinoma, and other was diagnosed as metastatic adenocarcinoma. Out of three inadequate smears, 2 were later found to be malignant and were diagnosed as HCC and metastatic adenocarcinoma, respectively, and 1 was focal nodular hyperplasia. One case which was negative for malignancy on cytology was diagnosed as liver abscess on histopathology.

Out of 52 cases, 50 were confirmed as malignant cases. Metastatic tumors constituted 29 cases (58%). Eleven cases showed primary in gall bladder as confirmed by USG findings and histology. Eight cases were having primary in colon as confirmed by histology and elevated CEA levels. Three cases were known cases of breast carcinoma, prostatic adenocarcinoma and non-Hodgkin lymphoma. In 7 cases of metastatic adenocarcinoma, primary lesion could not be detected [Table 2].
Table 2: Typing of liver tumors on final diagnosis

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The sensitivity of cytological analysis was 96%, and the specificity was 100%. The positive and negative predictive value was 100% and 50%, respectively. Accuracy of FNAC was found to be 96.1% with no false positive diagnosis.

   Discussion Top

Fine needle aspiration under ultrasound guidance has been found to be very useful in diagnosing hepatic tumors. [5],[6]

The most important requirement for cytodiagnosis is to obtain a representative sample. The ultrasound-guided FNA of liver lesions increases the accuracy of sampling, even for a deeply seated lesion, and has been proved to be a safe and accurate method for diagnosing primary liver carcinomas and metastases.

Inadequacy rate in current study using 18 gauge spinal needle for deep-seated lesions was 5.7%. Bell et al.[6] and Talukdar [7] reported inadequacy rate of 13% and 6.5%, respectively, using 22 gauge needle technique. Lowest inadequacy rate of 1% is reported by Guo et al.[8] using 22 gauge chiba needle. In the present study, causes of inadequacy were hemorrhagic and non-representative smears.

An aspirate that obtains material only from the surrounding tissue of the tumor may show reactive and proliferative changes, whereas an aspirate from the center of a large tumor may sample only degenerative and necrotic material. Therefore, for aspiration to be representative, the needle should pass through the entire mass.

Tao et al.[9] and Edoute et al.[10] advocated multiple aspirations, as this may minimize diagnostic errors. Multiple liver aspirations under guidance can almost certainly yield representative material of neoplastic cells from at least one aspirate. The yield of cells can be further increased by heparinisation of fine needles and syringes.

In most of the studies, the youngest patient was 10 year old. The youngest patient, 20 days old with liver lesion, was reported by Das et al. [3] In our study, the youngest case was 1 year old.

It is very difficult to differentiate between primary and metastatic tumors on cytology. Difficulty in cytological diagnosis arises at the end of malignant spectrum i.e., distinguishing well differentiated from benign lesions and separating less differentiated from metastatic malignancies.

Important cytological criteria useful in distinguishing HCC from non-neoplastic liver are increased nucleocytoplasmic ratio, arrangement of tumor cells in trabecular pattern and atypical naked hepatocytic nuclei. Other secondary criteria useful in differentiation are nuclear features i.e., irregularly granular chromatin, uniformly prominent and multiple nucleoli. [11]

Cellular patterns observed in aspirated material useful for the diagnosis of HCC include either trabecular or acinar arrangement of cells, high cellularity of smears, polygonal cells with central nuclei, abundant granular cytoplasm, coarse granular chromatin, increased nucleocytoplasmic ratio, malignant cells separated by sinusoidal endothelial cells, presence of bile, prominent nucleoli, multinucleated tumor giant cells, atypical stripped nuclei, large and small cytoplasmic vacuoles, intracytoplasmic and intranuclear inclusions [12],[13],[14] [Table 3].
Table 3: Frequency of various morphological features in aspirated biopsy samples of HCC in present study

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Trabecular pattern was observed in 65% cases. Individual trabecula was well-defined, long and broad and consisted of 4 to 10 layers of tumor cells. Acinar pattern was seen in 38% cases. On the basis of frequency of occurrence, most consistent features were high cellularity, high nucleocytoplasmic ratio, hyperchromatic nuclei and irregular granular chromatin. Second common features were pleomorphism and macronucleoli followed by polygonal cells with central nucleus and well-defined cytoplasm, atypical naked hepatocytic nuclei and malignant cells separated by sinusoidal capillaries.

However, the number of cytological features indicating malignancy is variable with the differentiation of the tumor.

The primary cytological features to differentiate HCC from metastatic tumors stated by Bottles et al.[15] include polygonal cells with centrally placed nuclei, malignant cells separated by sinusoidal capillaries and presence of bile. Other secondary criteria useful in differentiating HCC are intranuclear cytoplasmic inclusions and endothelial cells surrounding tumor cells clusters. The secondary deposits in the liver may reproduce the histology of primary lesions. The salient features separating HCC from metastatic adenocarcinoma given by Greene et al.[16] were tumor cells in HCC are polygonal or polyhedral, whereas cells are usually columnar or cuboidal in adenocarcinoma; cells in HCC have abundant eosinophilic and granular cytoplasm with one or two macronucleoli, whereas adenocarcinoma cells may show mucin secretion, and presence of more than two nucleoli is common. Trabecular arrangement is suggestive of HCC, whereas acinar or glandular arrangement is more favorable to adenocarcinoma. Inflammatory background is commonly seen in adenocarcinoma. In our study, metastatic deposits from colon carcinoma showed large columnar cells with eccentric large nuclei and prominent nucleoli. Acinar arrangement and intracellular and/or extracellular mucin was frequently observed.

Metastases from breast carcinoma showed tumor cells arranged in clusters as well as singly. The cells were round to oval with central hyperchromatic nuclei. Metastatic squamous cell carcinoma showed large tumor cells with marked variation in size and shape. Cells showed abundant cytoplasm with well demarcated cell borders, hyperchromatic nuclei and coarse chromatin. Intracellular keratinisation was observed in one case.

Various studies have reported sensitivity varying from 67% to 99.5% and specificity 87% to 100%. [2],[6],[8],[10],[17] In the present study, tumor type could be determined by FNAC with specificity up to 100%. FNAC was able to distinguish between neoplastic and non-neoplastic lesions with an overall sensitivity of 96%.

The overall accuracy of procedure in the present study was 96.4%, which was comparable to the rate of accuracy reported in literature (78% to 97.82%). [2],[6],[8],[10]

   Conclusions Top

FNAC is a safe diagnostic modality for the typing and grading of hepatic tumors. The smears obtained under ultrasound guidance are highly representative. The laden cytological criteria have proven to be successful to pinpoint the diagnosis in smears, grading them and differentiating the primary from metastatic tumors.

   References Top

1.Kuo FY, Chen WJ, Lu SN, Wang JH, Eng HL. Fine needle aspiration cytodiagnosis of liver tumors. Acta Cytol 2004;48:142-8.  Back to cited text no. 1
2.Mallikarjuna Swamy MC, Arathi C, Kodandaswamy C. Value of ultrasonography-guided fine needle aspiration cytology in the investigative sequence of hepatic lesions with an emphasis on hepatocellular carcinoma. J Cytol 2011;28:178-84.  Back to cited text no. 2
3.Das DK, Tripathi RP, Kumar N, Chachra KL, Sodhani P, Prakash S, et al. Role of guided fine needle aspiration cytology in diagnosis and classification of liver malignancies. Trop Gastroenterol 1997;18:101-6.  Back to cited text no. 3
4.Naggada HA, Ahidjo A, Ajagi NA. Correlation between ultrasound findings and ultrasound guided FNAC in the diagnosis of hepatic lesions: A Nigerian tertiary hospital experience. Int J Gastroenterol 2007;5:2.  Back to cited text no. 4
5.Schwerk WB, Schmitz-Moormann P. Ultrasonically guided fine needle biopsies in neoplastic liver diseases, cytohistologic diagnoses and echo pattern of lesions. Cancer 1981;48:1469-77.  Back to cited text no. 5
6.Bell DA, Carr CP, Szyfelbein WM. Fine needle aspiration cytology of focal liver lesions: Results obtained with examination of both cytologic and histologic preparations. Acta Cytol 1986;30:397-402.  Back to cited text no. 6
7.Talukder SI, Huq MH, Haque MA, Rahman S, Islam SM, Hussain GA, et al. Ultrasound guided fine needle aspiration cytology for diagnosis of mass lesions of liver. Mymensingh Med J 2004;13:25-9.  Back to cited text no. 7
8.Guo Z, Kurtycz DF, Salem R, De Las Casas LE, Caya JG, Hoerl HD. Radiology guided percutaneous fine needle aspiration biopsy of the liver: Retrospective study of 119 cases evaluating diagnostic effectiveness and clinical complications. Diagn Cytopathol 2002;26:283-9.  Back to cited text no. 8
9.Tao LC, Donat EE, Ho CS, McLoughlin MJ. Percutaneous fine needle aspiration biopsy of the liver: Cytodiagnosis of hepatic cancer. Acta Cytol 1979;23:287-91.  Back to cited text no. 9
10.Edoute Y, Tibon-Fisher O, Haim SB, Malberger E. Ultrasound guided fine needle aspiration of liver lesions. Am J Gastroenterol 1992;87:1138-41.  Back to cited text no. 10
11.Cohen MB, Haber MM, Holly EA, Ahn DK, Bottles K, Stoloff AC. Cytologic criteria to distinguish hepatocellular carcinoma from nonneoplastic liver. Am J Clin Pathol 1991;95:125-30.  Back to cited text no. 11
12.Ali MA, Akhtar M, Mattingly RC. Morphologic spectrum of hepatocellular carcinoma as seen in fine needle aspiration biopsies. Acta Cytol 1986;30:294-302.  Back to cited text no. 12
13.Noguchi S, Yamamoto R, Tatsuta M, Kasugai H, Okuda S, Wada A, et al. Cell features and patterns in fine needle aspirates of hepatocellular carcinoma. Cancer 1986;58:321-8.  Back to cited text no. 13
14.Soyuer I, Ekinci C, Kaya M, Genc Y, Bahar K. Diagnosis of hepatocellular carcinoma by fine needle aspiration cytology: Cellular features. Acta Cytol 2003;47:581-9.  Back to cited text no. 14
15.Bottles K, Cohen MB, Holly EA, Chiu SH, Abele JS, Cello JP, et al. A stepwise logistic regression analysis of hepatocellular carcinoma: An aspiration biopsy study. Cancer 1988;62:558-63.  Back to cited text no. 15
16.Greene CA, Suen KC. Some cytologic features of hepatocellular carcinoma as seen in fine needle aspirates. Acta Cytol 1984;28:713-8.  Back to cited text no. 16
17.Samartunga H, Wright G. Value of fine needle aspiration biopsy cytology in the diagnosis of discrete hepatic lesions suspicious for malignancy. Aust N Z J Surg 1992;62:540-4.  Back to cited text no. 17

Correspondence Address:
Sharda Balani
A-68, Vijay Nagar, Lalghati, Bhopal - 462 001
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Source of Support: None, Conflict of Interest: None

DOI: 10.4103/0970-9371.112655

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