**2. Benign epithelial liver tumours**

Liver cell adenoma and bile duct adenoma will be discussed here. The regenerative process‐ es with the emphasis on focal nodular hyperplasia are described considering the differential diagnosis.

### **2.1. Liver cell adenoma and its differential diagnosis with focal nodular hyperplasia**

Liver cell adenoma or hepatic adenoma is defined as benign tumour arising from hepato‐ cytes. The epidemiology is characterised by female predominance (90%) and strong associa‐ tion with oral contraceptive use [26-27] as 85% of affected persons have such history. Liver cell adenoma was rare before the era of oral contraceptives [27]. At present, the incidence has increased but is still low: 3-4 /100 000 per year in long-term users of oral contraception [27-29]. The patients mostly are 20-39 years old. The other risk factors of hepatic adenoma include androgen burden. The tumours can also arise spontaneously or occasionally can be related toglycogen storage diseases or diabetes mellitus. Clinically, the patients mostly are symptomatic. Abdominal fullness can be attributed to the presence of mass lesion; pain can be caused by necrosis [27]. Rupture and bleeding (40%) represent dangerous complications [27,29-31]; the risk of these events is increased in pregnant ladies affected by liver cell adeno‐ ma due to prior use of hormonal contraceptives. Risk of malignant transformation also is recognised [29,32]. By literature analysis, Farges and Dokmak concluded that 5% of resected hepatic adenomas bear HCC foci [32]. The risk of malignant transformation is higher in ade‐ nomas exceeding the size of 5 cm irrespectively of the number of adenomas as well as in males. Grossly, liver cell adenomas are mostly unifocal (80%) and subcapsular. The tumours can be quite large (5-20 cm). In most cases (75%) adenomas are encapsulated [27]. However, the capsule can be thin or absent [10]. In contrast to HCC, adenomas usually are not associ‐ ated with cirrhosis [31]. Otherwise, radiological similarities exist between adenoma and HCC as both can be large, have rich vascularity and can undergo necrosis [31]. Microscopi‐ cally, the tumour is composed by hepatocytes lacking anaplasia and arranged in thin (1-2 cells) trabeculae [27,29]. Cellular atypia and macrotrabeculae must be absent. Single arterio‐ les, a pair of arteriole and venule or isolated biliary ducts are scattered throughout the le‐ sion. However, well-formed triads enveloped in connective tissue are absent within the lesion. The tumour can be distinguished from normal liver by larger size of neoplastic cells, presence of capsule and lack of triad-containing portal tracts. Steatosis, hydropic degenera‐ tion or Mallory hyaline can be observed. Fibrous tissue, haemosiderin and calcifications can develop in the consequence of haemorrhage. The immunophenotype is characterised by ex‐ pression of Hep Par 1 and other antigens that confirms the hepatic origin and by lower pro‐ liferation than in HCC. Molecular typing is emerging for liver cell adenoma as well. At present, up to 4 molecular types are identified:


cal process and radiologically detected changes [15]. Further, the obtained tissue must be subjected to adequate technological process. Innovations here allow shortening the turnover time significantly. Next, the evaluation of morphology must be done searching for the char‐ acteristic traits of the above noticed tumours. However, due to the limited tissue amount in the biopsy, the tumour architecture sometimes is difficult to identify embarrassing the dis‐ tinction between nodular hyperplastic process, benign tumour or low-grade malignancy. In contrast, high-grade malignancies can show significant cytological atypia by few signs of differentiation embarrassing the detection of histogenesis [6] and the distinction between

Immunohistochemical markers as glypican-3 [1], Hep Par 1 [3,6], CD10 [3], alpha-fetopro‐ tein [6] and TTF-1 [16] are useful in the HCC diagnostics. Alterations of CD31 and CD34 positive endothelial cell network reflect vascular remodelling during hepatic carcinogenesis [7]. Cytokeratin (CK) 19 and 7 are characteristic for cholangiocellular carcinoma [3]. In meta‐ stases, organospecific markers including CDX2, mammaglobin, nuclear expression of TTF-1 or presence of neuroendocrine markers can confirm extra-hepatic origin [17]. As colorectal, breast, lung and neuroendocrine cancers are frequent cause of metastatic liver damage [2] high diagnostic value of immunohistochemistry (IHC) can be expected. However, the exact detec‐ tion of histogenesis can be difficult with metastatic pancreatic or gastric tumours and highgrade malignancies. IHC is mandatory for the diagnostics of haematological neoplasms and epithelioid haemangioendothelioma. Assessment of tumour biological potential can be done by IHC, evaluating Ki-67, Cyclin D1, FOXJ1, stem cell markers, matrix metalloproteinases and other markers [7-8,18-22]. Novel markers appear continuously as heat-shock protein 70 [23].

Nowadays, pathology is not any more purely descriptive but it is becoming more functional and clinically relevant. The classic morphologic characteristics must be combined with inte‐ grated evaluation of neoplastic process in the liver, including histogenesis, grading, clonal changes, type and extent of vascularisation, immunophenotype, heterogeneity, prediction of treatment sensitivity and the clinical behaviour [7]. New technologies as proteomic profiling and genomic marker analysis should be applied in the evaluation of liver tumours [4]. Mi‐ croRNA studies can lead to new findings in cancer pathogenesis and prediction of treatment

The aim of the following chapter is to describe morphological and immunohistochemical characteristics of primary and secondary liver tumours in order to develop logistic basis for differential diagnosis of these processes in biopsy materials. Short discussion about the gen‐

Liver cell adenoma and bile duct adenoma will be discussed here. The regenerative process‐ es with the emphasis on focal nodular hyperplasia are described considering the differential

esis and clinical course of each tumour will be included as well.

**2. Benign epithelial liver tumours**

primary and metastatic tumour.

116 Liver Biopsy - Indications, Procedures, Results

efficacy [24,25].

diagnosis.


The hepatic adenomas with *TCF1* gene mutation comprise 35-40% of liver cell adenomas. The patients are female. The tumour loses the expression and functions of hepatocyte nu‐ clear factor 1 (HNF1) encoded by *TCF1* gene. Inactivation of the gene can be caused by mutation in both alleles or by combination of a mutation and 12q deletion leading to loss of heterozygosity in the corresponding region [33]. Germ-line mutation of *HNF1* gene man‐ ifests as maturity-onset diabetes of the young (MODY), type 3, in association with liver adenomatosis [34]. However, the spectrum of *HNF1A* somatic mutations in liver cell adeno‐ ma differs from that in patients with MODY3 and suggests genotoxic damage [35]. By IHC, loss of liver fatty acid binding protein can be observed. Not surprisingly, the adenomas show steatosis [29].

nostics between FNH and hepatic adenoma. As described by Walther and Jain, CK19 and CD56 detect rich network of proliferating biliary ducts in the fibrous septa of FNH but re‐ veal only few isolated ducts within the parenchyma of hepatic adenoma. Expression of CK7 is remarkable for the focal presence in parenchyma of liver cell adenoma in contrast to FNH while both lesions show expression of CK7 in biliary ducts. Thus, panel of CK19, CD56 and CK7 can be advised to solve the differential diagnosis in core biopsy [29]. Immunohisto‐ chemical expression pattern of glutamine synthetase differs between normal liver tissue, FNH and liver cell tumours as well. In healthy tissue, glutamine synthetase is present in per‐ ivenular hepatocytes. These positive areas are expanded in FNH [39]. In hepatic adenomas, glutamine synthetase expression is either diffuse of negative. In the last situation, the nega‐ tivity in the tumour can be incomplete, with focally preserved expression in the tumour pe‐ riphery [29] and thus difficult to interpret, especially in small biopsies where the preserved

Primary and Metastatic Tumours of the Liver: Expanding Scope of Morphological and Immunohistochemical Details...

http://dx.doi.org/10.5772/52838

119

In nodular regenerative hyperplasia, the liver contains many small regenerative nodules. Partial nodular transformation affects hilar area and is characterised by group of regenera‐

Considering the differential diagnosis with HCC, thick trabecular cords, cytologic anaplasia and invasive growth reveal the malignant biological potential. The thickening of trabeculae is defined as presence of more than 2 cell layers in the trabeculae. The anaplasia is recog‐ nised by nuclear hyperchromasia, prominent nucleoli and increase in the nucleo: cytoplas‐ mic ratio. Presence of mitoses practically excludes the diagnosis of hepatic adenoma. Atypical mitoses are absolute evidence of malignancy. The invasive growth can manifest as invasion through the capsule, infiltration into liver parenchyma and true invasion into

Bile duct adenoma is defined as a benign neoplasm of portal bile ducts. The epidemiologic data suggest rare occurrence. However, as the tumours mostly are small and asymptomatic [27], the true incidence and prevalence is unknown. Grossly, bile duct adenomas are mostly solitary (83%), subcapsular (95%) and small (below 1 cm). By light microscopy, the lesion is characterised by demarcated proliferation of bile ducts lacking atypia. The immunopheno‐ type repeats the staining characteristics of biliary ducts exhibiting expression of cytokeratins 7 and 19 [27]. The differential diagnosis can include small foci of low-grade cholangiocarci‐ noma or metastatic low-grade adenocarcinoma, but the benign cytological appearance is helpful. Von Meyenburg hamartoma differs from bile duct adenoma, as the hamartomas would be multiple and show traits of cholestasis. However, the exact separation might not be of crucial importance due to benign course of biliary adenoma and pathogenetic sugges‐

tion that biliary adenoma represent a reactive process rather than true neoplasm.

positive focus seems to be dominant.

blood vessels [27].

**2.2. Bile duct adenoma**

tive nodules surrounded by fibrous tissue [27].

Inflammatory hepatic adenomas constitute 50% of liver cell adenomas and can be associated with obesity, smoking and alcohol use. Pathogenetically, inflammatory hepatic adenoma is characterised by IL-6 pathway activation centred on gp130 protein in IL-6 receptor. The recep‐ tor can be subjected to ligand-independent activation due to mutation in *IL6ST* gene, or the levels of gp130 can be elevated. The IL-6 receptor activation leads to recruitment of inflamma‐ tory cells through gp130-mediated production of chemokine CCL20. The mutation was found in 60% of inflammatory adenomas [36]. However, the IL-6 pathway activation is universal in the inflammatory hepatic adenoma. Microscopically, inflammatory infiltrates are observed in addition to the architecture and cytologic details of adenoma. Occasional bile ductules, dilat‐ ed sinusoids and arterioles can be present. Haemorrhage is frequent. By IHC, expression of acute phase reactants serum amyloid A and C-reactive protein is marked [29].

A group of hepatic adenomas is associated with *beta-catenin* mutation [37-38].The beta-cate‐ nin pathway is not affected in *TCF1* inactivated group [29,38]. Beta-catenin activation can be assayed by immunohistochemical over-expression of glutamine synthetase or by aberrant nuclear localisation of beta-catenin. However, the tumours can show dysplastic changes more characteristic for HCC thus possibly this group will be reclassified into well-differenti‐ ated HCC [29,36].

The last group of hepatic adenomas (5%) lacking *TCF1* inactivation, inflammatory signature and beta-catenin mutation [29] could represent distinct group with peculiar pathway of mo‐ lecular pathogenesis or result of technological shortcomings.

The differential diagnosis of hepatic adenoma in biopsy includes low-grade HCC and hy‐ perplastic lesions like focal nodular hyperplasia, nodular regenerative hyperplasia and par‐ tial nodular transformation [27].

Focal nodular hyperplasia (FNH) is a comparatively frequent differential diagnosis of hepat‐ ic adenoma. The FNH incidence is estimated as 3% [29-30,39]. FNH is characterised by pres‐ ence of hypervascular stellate scar in liver parenchymal nodule. The blood vessels are located in the middle of star-like fibrous tissue while the periphery is occupied by proliferat‐ ing bile ductules. The morphologically remarkable abundant vascularity is in accordance with the hypothesis of the FNH origin due to microscopic arterial malformation [40-42]. The crucial difference between FNH and adenoma is pathogenetic as the former is thought to be hyperplastic lesion, while adenoma is a neoplasm. The presence of stellate scar and lack of peripheral capsule in FNH contrasts with presence of peripheral capsule and almost com‐ plete lack of connective tissue or portal triads within adenoma. If the architecture is incom‐ pletely represented in the biopsy, molecular characteristics should be able to discriminate between the two inherently different processes, the hyperplasia and tumour. The immuno‐ histochemical markers of biliary differentiation have been employed in the differential diag‐ nostics between FNH and hepatic adenoma. As described by Walther and Jain, CK19 and CD56 detect rich network of proliferating biliary ducts in the fibrous septa of FNH but re‐ veal only few isolated ducts within the parenchyma of hepatic adenoma. Expression of CK7 is remarkable for the focal presence in parenchyma of liver cell adenoma in contrast to FNH while both lesions show expression of CK7 in biliary ducts. Thus, panel of CK19, CD56 and CK7 can be advised to solve the differential diagnosis in core biopsy [29]. Immunohisto‐ chemical expression pattern of glutamine synthetase differs between normal liver tissue, FNH and liver cell tumours as well. In healthy tissue, glutamine synthetase is present in per‐ ivenular hepatocytes. These positive areas are expanded in FNH [39]. In hepatic adenomas, glutamine synthetase expression is either diffuse of negative. In the last situation, the nega‐ tivity in the tumour can be incomplete, with focally preserved expression in the tumour pe‐ riphery [29] and thus difficult to interpret, especially in small biopsies where the preserved positive focus seems to be dominant.

In nodular regenerative hyperplasia, the liver contains many small regenerative nodules. Partial nodular transformation affects hilar area and is characterised by group of regenera‐ tive nodules surrounded by fibrous tissue [27].

Considering the differential diagnosis with HCC, thick trabecular cords, cytologic anaplasia and invasive growth reveal the malignant biological potential. The thickening of trabeculae is defined as presence of more than 2 cell layers in the trabeculae. The anaplasia is recog‐ nised by nuclear hyperchromasia, prominent nucleoli and increase in the nucleo: cytoplas‐ mic ratio. Presence of mitoses practically excludes the diagnosis of hepatic adenoma. Atypical mitoses are absolute evidence of malignancy. The invasive growth can manifest as invasion through the capsule, infiltration into liver parenchyma and true invasion into blood vessels [27].

#### **2.2. Bile duct adenoma**

ifests as maturity-onset diabetes of the young (MODY), type 3, in association with liver adenomatosis [34]. However, the spectrum of *HNF1A* somatic mutations in liver cell adeno‐ ma differs from that in patients with MODY3 and suggests genotoxic damage [35]. By IHC, loss of liver fatty acid binding protein can be observed. Not surprisingly, the adenomas

Inflammatory hepatic adenomas constitute 50% of liver cell adenomas and can be associated with obesity, smoking and alcohol use. Pathogenetically, inflammatory hepatic adenoma is characterised by IL-6 pathway activation centred on gp130 protein in IL-6 receptor. The recep‐ tor can be subjected to ligand-independent activation due to mutation in *IL6ST* gene, or the levels of gp130 can be elevated. The IL-6 receptor activation leads to recruitment of inflamma‐ tory cells through gp130-mediated production of chemokine CCL20. The mutation was found in 60% of inflammatory adenomas [36]. However, the IL-6 pathway activation is universal in the inflammatory hepatic adenoma. Microscopically, inflammatory infiltrates are observed in addition to the architecture and cytologic details of adenoma. Occasional bile ductules, dilat‐ ed sinusoids and arterioles can be present. Haemorrhage is frequent. By IHC, expression of

A group of hepatic adenomas is associated with *beta-catenin* mutation [37-38].The beta-cate‐ nin pathway is not affected in *TCF1* inactivated group [29,38]. Beta-catenin activation can be assayed by immunohistochemical over-expression of glutamine synthetase or by aberrant nuclear localisation of beta-catenin. However, the tumours can show dysplastic changes more characteristic for HCC thus possibly this group will be reclassified into well-differenti‐

The last group of hepatic adenomas (5%) lacking *TCF1* inactivation, inflammatory signature and beta-catenin mutation [29] could represent distinct group with peculiar pathway of mo‐

The differential diagnosis of hepatic adenoma in biopsy includes low-grade HCC and hy‐ perplastic lesions like focal nodular hyperplasia, nodular regenerative hyperplasia and par‐

Focal nodular hyperplasia (FNH) is a comparatively frequent differential diagnosis of hepat‐ ic adenoma. The FNH incidence is estimated as 3% [29-30,39]. FNH is characterised by pres‐ ence of hypervascular stellate scar in liver parenchymal nodule. The blood vessels are located in the middle of star-like fibrous tissue while the periphery is occupied by proliferat‐ ing bile ductules. The morphologically remarkable abundant vascularity is in accordance with the hypothesis of the FNH origin due to microscopic arterial malformation [40-42]. The crucial difference between FNH and adenoma is pathogenetic as the former is thought to be hyperplastic lesion, while adenoma is a neoplasm. The presence of stellate scar and lack of peripheral capsule in FNH contrasts with presence of peripheral capsule and almost com‐ plete lack of connective tissue or portal triads within adenoma. If the architecture is incom‐ pletely represented in the biopsy, molecular characteristics should be able to discriminate between the two inherently different processes, the hyperplasia and tumour. The immuno‐ histochemical markers of biliary differentiation have been employed in the differential diag‐

acute phase reactants serum amyloid A and C-reactive protein is marked [29].

lecular pathogenesis or result of technological shortcomings.

show steatosis [29].

118 Liver Biopsy - Indications, Procedures, Results

ated HCC [29,36].

tial nodular transformation [27].

Bile duct adenoma is defined as a benign neoplasm of portal bile ducts. The epidemiologic data suggest rare occurrence. However, as the tumours mostly are small and asymptomatic [27], the true incidence and prevalence is unknown. Grossly, bile duct adenomas are mostly solitary (83%), subcapsular (95%) and small (below 1 cm). By light microscopy, the lesion is characterised by demarcated proliferation of bile ducts lacking atypia. The immunopheno‐ type repeats the staining characteristics of biliary ducts exhibiting expression of cytokeratins 7 and 19 [27]. The differential diagnosis can include small foci of low-grade cholangiocarci‐ noma or metastatic low-grade adenocarcinoma, but the benign cytological appearance is helpful. Von Meyenburg hamartoma differs from bile duct adenoma, as the hamartomas would be multiple and show traits of cholestasis. However, the exact separation might not be of crucial importance due to benign course of biliary adenoma and pathogenetic sugges‐ tion that biliary adenoma represent a reactive process rather than true neoplasm.
