Mucin in primary liver carcinomas : Combined hepatocellular-cholangiocarcino ma or variant hepatocellular carcinoma

OBJECTIVE: To investigate whether the presence of mucin defines a combined hepatocellular-cholangiocarcinoma or merely a variant of usual hepatocellular carcinoma (HCC). METHODS: From 1979-92, 124 cases of usual HCC were studied at Niigata University Hospital, Niigata City, Japan, and several affiliated hospitals. Histological diagnoses were determined according to World Health Organization (WHO) criteria. Hematoxylin and eosin stain, periodic acid-Schiff (PAS) stain, PAS stain after diastase digestion (D-PAS) and silver stain tests were performed. Cases containing D-PAS-positive substances were also stained by Alcian blue (AB) stain, high iron diamine (HID) stain and concanavaline A paradox-3 type (ConA3) stain. The classification of mucin was determined by AB, HID and ConA3 stains. RESULTS: Mucin was recognized in the area of HCC by mucin stains in 25 of 124 cases. Two forms of mucin existence were classified: extracellular and intracellular. Mucins were classified by histochemical stains into three types: sulfomucin, sialomucin and neutral mucin. CONCLUSIONS: According to the WHO histological classification of primary carcinoma of the liver, mucin existence is characteristic of intrahepatic cholangiocarcinoma. But if mucin exists in morphologically usual HCC, it is better to diagnose it as a variant of HCC (with mucin) rather than as a combined hepatocellular-cholangiocarcinoma.

P rimary carcinoma of the liver can be classified histologi- cally as hepatocellular carcinoma (HCC), intrahepatic cholangiocarcinoma (CC) and combined hepatocellularcholangiocarcinoma (HCC-CC).HCC is characterized by trabecular patterns of hepatocyte-like cells with stroma of sinusoid-like capillaries and sometimes bile production.CC is characterized by glandular structures with abundant fibrous stroma.Mucin production is thought to characterize CC.Combined HCC-CC is a tumour containing unequivocal elements of both HCC and CC (1).In this study, we classified HCC into two groups by mucin stains and compared the two groups from clinical and histopathological points of view.

RESULTS
Twenty-five of the 124 cases were HCC-mucin-positive (Table 1).The other 99 cases were HCC-mucin-negative.Clinical data and background liver disease are shown in Tables 2 and 3.
In a review of the 25 HCC-mucin-positive cases, two types of mucin were classified: extracellular (n=22, 88%)mucin was recognized inside the extracellular space surrounded by carcinoma cells of HCC (Figures 1,2); and intracellular (n=3, 12%) -mucin was recognized inside the carcinoma cells of HCC (Figure 3).
There was no correlation between mucin characteristics and histopathological differentiation; similarly, there was no correlation between histopathological differentiation and mucin form (c 2 , P<0.05).
No difference was found between group A (25 cases of HCC-mucin-positive) and group B (99 cases of HCCmucin-negative) in terms of age, sex, positive ratio of hepatitis B surface antigen, positive ratio of antihepatitis C virus, serum alpha-fetoprotein level, serum carcinoembryonic antigen level and background liver diseases (Student's t test, P<0.05).

DISCUSSION
According to the usual histopathological classification of primary carcinoma of the liver, mucin production can be recognized in CC, CC areas of combined HCC-CC7 and bile duct cystadenocarcinoma, but not in HCC.Mucin production is one characteristic of CC (1,4).There is a lot of evidence that these two types of liver carcinoma (HCC and CC) differ in their etiological, epidemiological, clinical and histopathological features (5-7).Combined HCC-CC7 presents both characteristics histopathologically.
Allen and Lisa (8) reported five cases of combined HCC-CC and reviewed 11 others from the literature.They defined combined HCC-CC based on the three types of tumours: separate masses of HCC and CC; contiguous but independent masses of HCC and CC; and an intimate intermingling of hepatocellular and glandular elements.Also Goodman and Ishak (9) classified combined HCC-CC into three types: collision tumours, apparently a coincidental occurrence of both HCC and CC in the same patient; transitional tumours, in which there are areas of intermediate differentiation and an identifiable transition between HCC and CC; and fibrolamellar tumours, which resemble the fibrolamellar variant of HCC but also contain mucinproducing pseudoglands.In their classification, one common criterion of CC was the production of mucin.
Wada et al (10) reported that intrahepatic metastatic lesions of HCC showing a trabecular growth pattern produced both bile and mucin.They diagnosed the metastatic lesion as combined HCC-CC because of the presence of mucin.Robert and colleagues (11) concluded that mucin can be found within the pseudogland-like structures of HCC metastatic to the ovary.In combined HCC-CC, Uekusa et al (12) recognized mucin inside the pseudogland-like structures and inside the carcinoma cells in the area of HCC, but these workers did not find the stains helpful for discrimination.These three reports show a similar contradiction: histopathologically, the diagnosis was HCC, but mucin was recognized.Mucin presence changed the diagnoses in these reports.

Hirota et al
Twenty-five of the 124 HCC cases in this study were HCC-mucin-positive (group A) and the other 99 cases were HCC-mucin-negative (group B).There was no difference between the groups concerning clinical features and histopathological backgrounds.If mucin stains had not been per-formed, the two groups would have been regarded as usual HCC.According to WHO criteria (1), the 25 HCCmucin-positive cases must be diagnosed as combined HCC-CC rather than HCC because they contain mucin.Mucin existence is important enough to change diagnoses.Goodman and Ishak (9), and Popper and Shaffner (13) have questioned this criterion about mucin production.
Apparent in this study were some problems concerning mucin; for example, mucin characteristics were not fixed.Also, it was unclear whether the mucin was derived from carcinoma cells (14), stroma or mucinous degeneration (15).In addition, bile duct cystadenocarcinoma and CC contain sialomucin predominantly (16).
If the origin of mucin is not clear, it is inappropriate to change the morphological diagnoses.

TABLE 3 Comparison of histopathological background
Extracellular type of mucin existence.A Large spaces can be seen in hepatocellular carcinoma, which is surrounded by sinusoid-like capillaries.Hematoxylin and eosin stain.B Mucin is stained with red inside the large spaces.Diastase digestion periodic acid-Schiff stain.C Mucin, composed of sialomucin, is stained blue.Alcian blue stain (xE 330) Mucin is stained with red in the small spaces between carcinoma cells.Diastase digestion periodic acid-Schiff stain.B Mucin is stained with brown inside the small spaces between carcinoma cells of hepatocellular carcinoma.
Figure 3) Intracellular type of mucin existence.A Hepatocellular carcinoma surrounded by sinusoid-like capillaries can be seen.Hematoxylin and eosin stain.B Part of carcinoma cells of hepatocellular carcinoma is stained with blue.Mucin is composed of sialomucin.Alcian blue stain (xE 409) Figure 2) Left Extracellular type of mucin existence.A