Search

To clarify the mechanism of production of des-gamma-carboxy (abnormal) prothrombin (DCP) by hepatocellular carcinoma (HCC), we measured the levels of vitamin K, DCP, immunoreactive prothrombin and the activity of gamma-glutamyl carboxylase in liver tissues from HCC patients and in the medium of cultured human hepatoma cells. There was no significant difference in vitamin K (K1, MK-4) contents between HCC and non-HCC cirrhotic liver tissues. The activity of gamma-glutamyl carboxylase per unit amount of endogenous microsomal prothrombin precursor was decreased in HCC tissue compared with non-HCC liver tissue (positive plasma DCP: 335 +/- 72 vs 372 +/- 67, negative plasma DCP: 370 +/- 84 vs 393 +/- 56 nmol/min per mg prothrombin precursor, P > 0.05), although the total incorporation of 14COOH into microsomal precursor protein was higher in the former. By contrast, levels of DCP and immunoreactive prothrombin in HCC tissue were greater (P < 0.05) than those in non-HCC cirrhotic liver tissue. Furthermore, production of large amounts of immunoreactive prothrombin was observed in human hepatoma cells huH-1 and huH-2, which produced large amounts of DCP. These results suggest that there was excessive synthesis of prothrombin precursors by human HCC tissue and hepatoma cell lines huH-1 and huH-2. Thus, excessive synthesis of prothrombin precursors seems to be the main mechanism of DCP production by HCC.

Proliferating cell nuclear antigen (PCNA), also known as cyclin, is an auxiliary protein of DNA polymerase-delta and is found only in the nuclei of proliferating cells in the late G1 and S phases. The proliferation of hepatocellular carcinoma (HCC) by immunohistochemical staining for PCNA using paraffin sections of 20 surgically resected HCC specimens was analysed. The mean percentage of PCNA-positive nuclei in the HCC tissue was 10.3% in grade I of Edmondson and Steiner's classification, 25.5% in grade II, 28.4% in grade III and 41.5% in grade IV. In early HCC, we observed only a few PCNA-positive tumour cells. However, PCNA-positive nuclei were numerous in the tumour thrombi found in portal vein branches, in regions of extracapsular tumour growth, and in the inner nodules of tumours with a nodule-in-nodule formation. Proliferating cell nuclear antigen positivity was correlated with an increase of the nucleocytoplasmic ratio of tumour cells as determined by image analysis. Our findings showed that PCNA positivity was correlated with the histological grade and invasiveness of HCC, suggesting that this antigen may be used as an indicator to predict tumour invasion in patients with HCC.

We measured plasma abnormal prothrombin (des-gamma-carboxy prothrombin; DCP) levels in normal subjects and in patients with hepatocellular carcinoma and other various diseases using the enzyme-linked immunosorbent assay developed by Motohara et al. (Pediatr Res 1985; 19: 354-357). Fifty-eight percent of 52 patients with hepatocellular carcinoma had elevated DCP levels; 24 of 28 patients with advanced or moderately advanced hepatocellular carcinoma were positive. By contrast, 50 normal controls, 13 pregnant women and 10 patients with acute hepatitis had normal levels. Three of 55 patients with chronic liver disease, and 6 of 32 patients with other malignancies, showed a slight increase. Thus, increased plasma DCP appears useful for the diagnosis of hepatocellular carcinoma. To elucidate the mechanism for the increase of DCP in hepatocellular carcinoma, we cultured a human hepatoma cell line, huH-2, and measured the levels of this abnormal prothrombin in the medium. The huH-2 cells produced large amounts of DCP in the medium without added vitamin K. It increased in a cell concentration- and time-dependent fashion. These cells produced no detectable amount of DCP in the medium with added vitamin K. Thus, human hepatoma cell line huH-2 produces DCP, and its production is dependent on the amount of vitamin K available in the medium. Des-gamma-carboxy prothrombin may be a useful tumor marker for the diagnosis of hepatocellular carcinoma.

健常者及び肝細胞癌(HCC)を含む各種疾患例において血漿中の異常プロトロンビンPIVKA-II (P-II)をEIA法を用いて測定し,HCC例の57.7%が高値を示した.他の肝疾患,悪性腫瘍疾患例では陽性率は極めて低く,そのうち肝硬変例では全例低値であり,HCCに高い特異性が認められた.HCCにおけるP-IIの上昇機序を検索する目的でAFP産生株であるヒト肝癌由来細胞株(huH-2)を用いて培養上清中のP-IIを測定した結果,細胞濃度に比例してP-II量は増加し,vitamin K添加によりP-IIは検出されなかった.以上からヒト肝癌培養細胞huH-2はP-IIを産生し,それはvitamin Kの存在に左右される事が示唆され,P-IIは臨床的に有意義な腫瘍マーカーとみなされる.

HBワクチン接種例の長期観察と,follow-up後にHBs抗体の力価(PHA法)が23以下の例にHBワクチンのBooster injectionを施行し,そのHBs抗体responseを検討した.HBsAg, anti-HBs陰性の116例に1981年8月から82年11月の期間にHBワクチン(HBsAg 20μg)を,初回,4週,24週後に接種した.そして,84年5月,85年8月にfollow-up後の検査を施行した.HBs抗体(PHA法)陽性率は,ワクチン接種終了時の28週では,79.8%で,経過とともに(平均156週)25例陰性化し,57.7%と低下し,力価も28週をピークとして低下した.又,力価上昇の1例はHBウイルスの感染と考えられた. Booster injectionでは,Responderでは全例,すみやかに十分なHBs抗体responseがみられ,HBs抗体産生メモリは保たれていると考えられ,Low responderでは70%, No responderでは23.1%であった.経過とともに,HBワクチンによって得られたHBs抗体の力価は低下し,力価が感染予防に不十分な例には,Booster injection(1回)が必要であるので,少なくとも1年毎の定期的なfollow-upを要すると思われた.

A sero-epidemiological study was made of 6863 apparently healthy randomly selected volunteers from various parts of Taiwan. It revealed an 18% incidence of hepatitis B surface antigen (HBsAg) carriers amongst the general population, the prevalence being significantly higher in males than females (21.7% vs 14.1%). The Takasago natives showed the highest HBsAg carrier rate (27.0%). Those who are primarily derived from Ming Nan had a significantly higher prevalence (19.0%) than those from Hakka (14.7%) and recent immigrants from Mainland China (14.6%). About one-third of female carriers of child-bearing age in southern Taiwan were positive for e antigen (HBeAg) in a limited study; they may be an important source of HB virus (HBV) infection. The positivity rate for HBeAg was significantly higher in subjects below 40 years of age (36.7%) than those above 40 years (7.5%). There was a significant association between HBsAg positivity and elevated serum transaminase levels, especially in males; 20.3% of carriers had elevated transaminase in comparison with 11.1% of female carriers. Among HBsAg seronegative individuals, the frequency of abnormal transaminase increased with age with an overall abnormality rate of 10.6% in males and 5.0% in females, suggesting that factors other than HBV may also contribute to liver disease in Taiwan. Altogether, 9.3% had elevated transaminase. Out of 82 randomly selected HBsAg carriers who were evenly distributed in geography, serum antidelta antibody was positive in only five men (6.1%), of whom four had a mild elevation of transaminase levels (< 100 KU). The overall positivity rate for anti-hepatitis A (HA) antibody was 93.0% in southern Taiwan, indicating poor sanitation. These findings indicate that HB and HA virus infections are highly endemic in Taiwan and that delta infection is infrequent despite a high prevalence of HB virus infection.

Hepatocellular carcinoma (HCC) associated with cirrhosis and HCC developing in a noncirrhotic liver may have differing pathogeneses. To study this possibility, 425 autopsied cases of HCC were investigated. Of these, 45 livers were not cirrhotic, 50 were highly cirrhotic (liver weight less than 99 g), and the remaining 331 were cirrhotic but not so highly. The average age was significantly older in the highly cirrhotic group, suggesting a longer premalignant period of chronic liver disease. The liver weight in the noncirrhotic group was about 3.5 times that in the highly cirrhotic group. Hepatitis B surface antigen was positive in serum in only 9.3% and in liver tissue in 10% in the noncirrhotic cases, the positivity rate being much lower compared with other groups (P less than 0.005--0.01), yet antibody to HB core was positive in 90%. The antibody titers were low, however, indicating that these noncirrhotic patients had in the past had HB virus (HBV) infection with no residual chronic B hepatitis. Analysis of the grades of anaplasia of cancer tissue demonstrated an inverse correlation between the degree of fibrosis and grade of anaplasia, i.e., the more advanced the fibrosis, the less anaplastic the cancer. These data suggest that HCC arising in highly cirrhotic liver and in noncirrhotic livers have different pathogenetic backgrounds, and that HBV infection, even though transient, has a certain role in hepatocarcinogenesis. The generally held conjecture that HCC in a noncirrhotic liver is caused by nonviral carcinogens and HCC arising on the ground of cirrhosis is due to HBV seems untenable in such a simple concept.

Clinicopathological features were studied in 41 patients with histology-proven hepatocellular carcinoma without cirrhosis. Of these, 13 (31.7%) were positive for hepatitis B virus surface antigen (HBsAg) and 28 were negative. Twenty-six of 28 (92.9%) HBsAg negative cases had anticore antibody (anti-HBc) of low titres. The age of patients at the time of diagnosis of hepatocellular carcinoma was significantly younger in the HBsAg positive cases compared with the negative. The initial diagnostic clue was either abdominal mass or abdominal pain in 18 (43.9%) patients. However, the number of patients in whom hepatocellular carcinoma was detected by imaging in the absence of such clinical signs has increased recently. The tumour was encapsulated in 31 cases (75.6%) and most solitary encapsulated tumours were growing expansively. Tumours were resectable in 29 (70.7%) cases, and the prognosis of resected cases was much better than that of 12 non-operated cases. Twenty-one out of 29 (72.4%) resected tumours were solitary and encapsulated. However, recurrence of tumour occurred in 12 out of 28 (42.9%) cases, and it was within 2 years of resection in 11 cases. These observations and data seem to indicate that early detection and resection of tumour is very important in management, but tumour recurrence is inevitable in a considerable proportion of the patients with non-cirrhotic hepatocellular carcinoma.

A survey of 2985 apparently healthy Taiwanese in southern Taiwan revealed a high prevalence (18.2%) of hepatitis B surface antigen (HBsAg). It was significantly higher in males (22.3%) than in females (13.6%), but no correlation with family origin, socioeconomic status or residence was established. About one-third of the breeding female HBsAg carriers were HBeAg-positive, and these may be an important source in the spread of HBV. Subtyping of HBsAg in 63 subjects showed adw to be dominant in this area, and 8 subjects with suspicious results had overlapping heterotypic HBV’s (7 adwr and 1 adyw) in addition to 2 subjects in which coexistence of HBsAg and anti-HBs was recognized. The association between HBsAg positivity and serum transaminase elevation was significant, especially in the older groups who had a higher abnormal rate. Hepatitis A virus infection was serious too, with nearly 100% of people above 20 years of age being anti-HA antibody positive.

The conjugates formed in vitro by bilirubin UDP-glucuronyl transferase were studied by examining reaction products as intact tetrapyrroles, rather than as dipyrrolic azoderivatives. Bile pigments were extracted from conventional microsomal enzyme reaction mixtures by affinity chromatography over albumin-agarose, eluted with 50% ethanol, and separated by a silica gel thin layer chromatographic system. In the presence of UDPGA, native and activated microsomal preparations all formed both bilirubin mono- and diglucuronides from unconjugated bilirubin, and bilirubin diglucuronide from bilirubin monoglucuronide. No significant non-enzymatic conversion of mono- to diglucuronide occurred without UDPGA, or in the presence of denatured enzyme. Hence, bilirubin diglucuronide is a major product of bilirubin-UDP-glucuronyl transferase.

A case of biliary cystadenocarcinoma is presented in which the lesion seemed to partially differentiate into hepatocellular carcinoma. This patient had a cystic tumor in the left lobe of the liver, the interior being filled with papillary tumors. Histologic examination disclosed papillary growth of columnar tumor cells, together with tumor cells akin to hepatocytes (they had a distinct cell boundary, nuclear membrane, and nucleolus) showing a cobblestone appearance in some areas. Electron microscopic examination of this area showed bile canalicular structures. Histochemically, the tissue stained positively with the Luna-Ishak canalicular (LIC) technique; periodic acid-Schiff (PAS) staining demonstrated coexisting glycogen granulo-positive cells and mucus-producing cells. Thus, in this biliary cystadenocarcinoma, some tumor cells demonstrated the morphologic and histochemical features of hepatocytes.