Dendritic cells, T-cell infiltration, and Grp94 expression in cholangiocellular carcinoma

Although dendritic cells (DCs) play an important role in tumor immunity, there have been no reports on their role in cholangiocellular carcinoma (CCC). In 26 formalin-fixed, paraffin-embedded tissue sections from patients with CCC, cells positive for CD83 (a marker of mature DCs), CD1a (a marker of immature DCs), and CD8 and CD4 (T cell markers) were counted, and expression of glucose-regulated protein (grp) 94, which is considered to participate in the maturation of DCs, was evaluated by immunohistochemistry and Western blot analysis to study the relationship between their expression and patients' disease outcome. The number of CD83-positive DCs at the invasive margin of CCCs correlated significantly with the number of CD8-positive or CD4-positive T cells in the cancerous region and was significantly higher in grp94-positive cancer than in grp94-negative cancer (P = 0.0006). CD83-positive patients (positive cells in invasive margin > 12.4/field) had both a significantly lower incidence of lymph node metastasis (23.1% vs 69.2%; P = 0.0206) and a better outcome than CD83-negative patients (P <0.001). We conclude that mature DCs are distributed predominantly at the invasive margin of cancers, and a significantly higher number of mature DCs at the invasive margin are observed in patients with grp94-positive cancer cells. Mature DCs may enhance CD8- and CD4-positive cell infiltration into cancers and improve prognosis in patients with CCC, due in part to abatement of lymph node metastasis.     

Expression of glycoconjugates in intrahepatic cholangiocellular carcinoma

Summary Twenty cases of intrahepatic cholangiocellular carcinoma (IHCCC) were studied by lectin histochemistry for their glycoconjugate expression. Combined alcian blue-peroxidic acid Schiff (ABPS) staining was also made for mucins in these tissues. Results showed that epithelial cells of intrahepatic bile ducts contained varied quantity of DBA, WGA, LCA, Con A, PHA, RCA_I, BLA_I, SJA, SBA, PSA and UEA_I receptors but no PNA receptors. Lectin receptors were present at varying rates; 100% (Con A), 95% (PSA), 90% (LCA), 95% (WGA), 30% (BSA_I), 35% (DBA), 65% (PHA), 74% (PNA), 85% (RCA_I), 35% (SBA), 25% (SJA) and 55% (UEA0, respectively. The positive rates in well-differentiated IHCCC were significantly higher than those in either moderately or poorly-differentiated IHCCC. The distribution of lectin receptors in IHCCC was obviously different from that in peri-carcinomatous, cirrhotic and normal liver, and also differed from that in epithelial cells of normal intrahepatic bile ducts. All larger ducts were stained by ABPS and were mainly blue colour, while only 80% of IHCCC were positive for ABPS staining. Most of them were blue, others were red or purple. The results suggest that the expression of glycoconjugates had changed after the neoplastic transformation of bile duct cells.     

HLA expression in hepatocellular carcinoma cell lines

The present study undertook to investigate the biological significance of human leucocyte antigen expression in hepatocellular carcinoma and to elucidate the role of potential modulating agents on human leucocyte antigen expression. These studies used several hepatic tumour-derived cell lines as in vitro model systems. The cell lines included PLC/PRF/5 (Alexander cell line), Hep3B. HepG2, TONG PHC, HA22T/VGH, HA59T/VGH and Mahlavu, The cell lines K562 and Raji were used as negative and positive controls, respectively. K562, a B tymphoid-derived cell line, was shown to express negligible amounts of human leucocyte anligens, while Raji, an erythromyeloid-derived cell line, expressed both class I and class II human leucocyte anligens as well as their respective invariant chains, β₂-niicroglobulin and Ii. Using an ELISA, experiments performed on these cell lines confirmed the natural expression of class I and class II antigens by the HA22T/VGH and HA59T/VGH cell lines, whereas PLC/PRF/5 displayed class II surface antigens only. The effects of modulating agents such as interferon-gamma sodium bulyrate and clofazimine on human leucocyte antigen expression were investigated using the HA22T/VGH, HA59T/VGH and TONG PHC cell lines. These agents increased class 1 and class II human leucocyte antigen expression on HA22T/VGH and TONG PHC cells, but had no effect on the HA59T/VGH cell line. The results suggest a potential use for these agents as modulators of human leucocyte antigen expression by human heptocellular cell lines.     

Prognostic impact of cholangiocellular and sarcomatous components in combined hepatocellular and cholangiocarcinoma

Combined hepatocellular and cholangiocarcinoma (cHC-CC) is a rare type of liver cancer displaying both hepatocellular and cholangiocellular components. The cholangiocellular carcinoma (CC) in these tumors ranges from focal to prominent. Those cHC-CCs with sarcomatous features are reported to have a poor prognosis. To clarify whether the CC and sarcomatous component affects the prognosis, we classified 40 patients with cHC-CCs into 4 groups according to the presence of a sarcomatous component and the extent of the CC component. Seven (17.5%) tumors showed areas with a sarcomatous component. The remaining tumors were divided into a low-CC group (CC occupying <30% of the tumor, n = 12), a middle-CC group (30%-60%, n = 15), and a high-CC group (>60%, n = 6). Vascular invasion was more frequently present in the high-CC and sarcomatous group than in the other groups (P = .0007). No lymph node metastasis occurred in either the low- or the middle-CC groups, but it was detected in 3 (50%) cases of the high-CC group and in 2 (29%) cases of the sarcomatous group (P < .0001). There was a tendency for tumor size to increase from the low- to the middle- to the high-CC group. The Ki-67 labeling index values for the hepatocellular carcinoma, CC, and sarcomatous components were 11.4% +/- 12.9%, 25.4% +/- 18.3%, and 46.0% +/- 23.6%, respectively. The overall survival of patients in the high-CC and sarcomatous group was significantly poorer than that of patients in the low- and middle-CC groups (P = .0048). By multivariate analysis of overall survival, lymph node metastasis, histological subgroup, and vascular invasion were significant independent prognostic factors. A cHC-CC with a large CC component is as aggressive as cHC-CC with sarcomatous features.     

肝癌组织中KLF17的表达及临床意义

目的 检测KLF17在肝癌组织的表达并评价其对肝癌预后的影响.方法 免疫组化法检测KLF17在肿瘤组织中的表达,以及KLF17表达对患者无进展生存和总生存期的影响.比较原代培养的Hep11和Hep12细胞侵袭和迁移的差异,以及KLF表达的差异.干扰KLF17表达后Transwell法检测Hep11迁移和侵袭的能力.结果 36名有随访结果并可评估的患者中,KLF17 0 ～2分19人,3～5分的17人.KLF17 3～5分的患者无进展生存期较0～2分的患者长,有统计学差异,分别为(39.3±4.2)个月和(23.4±4.9)个月(P＜0.05).KLF17 3～5分的患者总生存为(46.9±3.2)个月,KLF17 0 ～2分的患者为(35.2±4.2)个月(P＜0.05).在原代培养肝癌细胞中,来自原发灶的Hep11比来自转移复发灶的Hep12的KLF17表达高,Hep11的KLF17干扰后,迁移和侵袭能力增强(P＜0.05).结论 KLF17表达可能是肝癌患者预后的独立预测因素.     

P-糖蛋白在肝细胞癌中的表达及临床研究

目的：探讨Ｐ－糖蛋白在细胞癌组织中表达及临床意义。方法：采用免疫组化法对７２例肝细胞癌，９例正常肝组织进行Ｐ－糖蛋白测定，作相关临床因素分析。结果：肝细胞癌组织中Ｐ－糖蛋白的阳性表达率为４４．０％，正常肝组织表达率为２２．０％（Ｐ〈０．０５）。Ｐ－糖蛋白的表达与肝癌的组织分型、分级及预后无明显相关性，并且化疗并不影响Ｐ－糖蛋白阳性表达病人的预后。结论：Ｐ－糖蛋白在肝细胞癌中的过表达，提示其在肝细胞     

Potential role of leptin expression in hepatocellular carcinoma

BACKGROUND: Obesity is associated with hepatocellular carcinoma (HCC). The association may result from the aberrant expression of adipokines. AIM: To explore the potential biological effect and prognostic value of leptin, one of the adipokines, in HCC. METHODS: Immunohistochemistry was used to evaluate the expression of leptin in 68 patients with HCC. The expression of Ki-67 and microvessel density (MVD) of tumorous lesions in HCC were also analysed. The result of leptin expression was further correlated with Ki-67 expression, intratumour MVD, clinicopathological characteristics, overall survival and the postoperative use of medroxyprogesterone acetate (MPA). RESULTS: High leptin expression was seen in 60.3% of patients with HCC and was significantly correlated with intratumour MVD (high v low; 59.2 (standard deviation 3.2) v 44.2 (19.5), p = 0.004), but not with Ki-67 expression. No marked correlation was seen between leptin expression and clinicopathological characteristics. However, using a multivariate Cox's proportional hazards model, leptin expression was a predictor for improved overall survival of patients with HCC (odds ratio 0.16; 95% confidence interval 0.03 to 0.87; p = 0.033). In addition, the Kaplan-Meier survival curve showed that high leptin expression was associated with a better survival in patients with HCC, treated postoperatively with MPA (p = 0.008, log rank test). CONCLUSION: High leptin expression was associated with an increased intratumour MVD and thus may be associated with HCC development. In addition, high leptin expression was a predictor for improved survival of patients with HCC, treated postoperatively with MPA.     

Cell Kinetic and morphological studies of human cholangiocellular carcinoma

We investigated the cell kinetics and morphologies of cholangiocellular carcinoma (CCC) using 48 autopsied or surgically resected cases (47 were adenocarcinoma and the remaining adenosquamous cell carcinoma), all of which were formalin-fixed and paraffin-embedded. Cell kinetics were analyzed by counting the number of argyrophilic nucleolar organizer regions (AgNOR) using immunostaining of proliferating cell nuclear antigens (PCNA) and flow cytometric DNA analysis. Dedifferentiation of CCC was positively correlated with AgNOR number (2.22 ± 0.21 in well differentiated, 3.66 ± 0.85 in moderately differentiated and 4.17 ± 0.49 in poorly differentiated adenocarcinomas, respectively). In 22 cases, the labeling index (LI) of PCNA was higher in moderately and poorly differentiated adenocarcinomas (24.0 ± 2.35 and 26.0 ± 4.89, respectively) than in well differentiated ones (10.8 ± 2.14). A majority of well differentiated ones were diploid, while aneuploidy prevailed in moderately to poorly differentiated ones. These data suggest that cell proliferative indices and nuclear DNA analysis of CCC accurately reflect their histological grading. The anatomical location of CCC along the biliary tree had no relation to either of the cell kinetic data. In autopsy cases, the patients with organ and lymph node metastases tended to show a higher DNA index and aneuploidy. This study implies that a combination of several cell kinetic data is valuable for the evaluation of the biological behaviors of CCC, and also supports further studies of cell kinetics of CCC using small-sized biopsy specimens, as a prognostic indicator.     

Downregulation of P-cadherin expression in hepatocellular carcinoma induces tumorigenicity

P-cadherin is a major contributor to cell-cell adhesion in epithelial tissues, playing pivotal roles in important morphogenetic and differentiation processes and in maintaining tissue integrity and homeostasis. Alterations of P-cadherin expression have been observed during the progression of several carcinomas where it appears to act as tumor suppressive or oncogenic in a context-dependent manner. Here, we found a significant downregulation of P-cadherin in hepatocellular carcinoma (HCC) cell lines and tissues compared to primary human hepatocytes and non-malignant liver tissues. Combined immunohistochemical analysis of a tissue microarray containing matched pairs of HCC tissue and corresponding non-tumorous liver tissue of 69 patients confirmed reduced P-cadherin expression in more than half of the cases. In 35 human HCC tissues, the P-cadherin immunosignal was completely lost which correlated with tumor staging and proliferation. Also in vitro, P-cadherin suppression in HCC cells via siRNA induced proliferation compared to cells transfected with control-siRNA. In summary, downregulation of P-cadherin expression appears to induce tumorigenicity in HCC. Therefore, P-cadherin expression may serve as a prognostic marker and therapeutic target of this highly aggressive tumor.     

Cytokeratin expression in hepatocellular carcinoma: an immunohistochemical study

Normal human hepatocytes express cytokeratins no. 8 and 18, whereas bile duct cells contain the same cytokeratins and, in addition, cytokeratins no. 7 and 19. This cytokeratin pattern is believed to be preserved during neoplastic transformation. Thirty-four cases of hepatocellular carcinoma (11 well differentiated, 16 moderately differentiated, 7 poorly differentiated) were studied on frozen sections using monoclonal antisera directed against individual cytokeratins no. 7, 8, 18, and 19 in an immunoperoxidase procedure. In 17 of 34 cases, tumor cells showed only reactivity with monoclonals anticytokeratin no. 8 and 18. However, 17 of 34 cases showed an aberrant pattern in that a variable number of tumor cells were stained with anticytokeratins no. 7 and/or 19 in addition to no. 8 and 18. Only three of 11 well-differentiated cases displayed an unexpected cytokeratin pattern, whereas an aberrant pattern was present in all seven of seven poorly differentiated cases. These results are in conflict with previously published data obtained by two-dimensional gel electrophoresis and immunohistochemistry. They indicate that the cytokeratin pattern might not always be preserved during neoplastic transformation. The implication of this finding for the differential diagnosis of metastatic gastrointestinal carcinomas is discussed.     

Histological study of PIVKA-II expression in hepatocellular carcinoma and adenomatous hyperplasia

Although serum concentration of protein induced vitamin K absence or antagonist II (PIVKA-II) has been widely used for diagnosing hepatocellular carcinoma (HCC), little information is available concerning tissue PIVKA-II as an immunohistochemical marker for liver histology. In this study, we examined the expression of PIVKA-II in precancerous nodules (adenomatous hyperplasia) and various differentiation grades of HCC by immunohistochemical study using the monoclonal anti-PIVKA-II antibody (MU-3). We examined the relationship between tissue PIVKA-II staining and serum PIVKA-II level, tumor histology and tumor size. PIVKA-II was mainly detected in the cytoplasm of the HCC cells. The positive rates of PIVKA-II were as follows: adenomatous hyperplasia (AH), 0% (0/9); well-differentiated HCC, 65% (15/23); moderately differentiated HCC, 85% (22/26); poorly differentiated HCC, 54% (7/13). The expression of tissue PIVKA-II staining in moderately differentiated HCC was significantly higher than in well- or poorly differentiated HCC, whereas the serum PIVKA-II level in poorly differentiated HCC was higher than well- or moderately differentiated HCC. There was no relationship between the expression of PIVKA-II in cancer tissues and serum levels of PIVKA-II. Immunohistochemical studies revealed that PIVKA-II was expressed even in small-sized or well-differentiated HCC cells, but expression was not detected in AH. It was concluded that PIVKA-II is a useful immunohistochemical marker, even in small-sized or well-differentiated HCC.     

The expression and clinical significance of TPM4 in hepatocellular carcinoma

Hepatocellular carcinoma (HCC) is known as the fifth most common cancer in the world for its poor prognosis. New diagnostic markers and treatments are urgent to discover. To evaluate the protein expression of Tropomyosin4 (TPM4) and investigate its prognostic value in HCC, we collected 110 patients with different degrees of HCC and 10 patients with normal hepatic tissues and performed immunohistochemistry. Western bot was used to evaluate the expression of TPM4 in three HCC cell lines (HepG2, Huh7, SMMC-7721) and normal liver cell line LO2, as well as 7 HCC tissues and 7 normal hepatic tissues. The results of TPM4 staining revealed that TPM4 expression in HCC was higher than that in normal hepatic tissues, which was positive in 51.8% (n=57) and negative in 48.2% (n=53) while in normal hepatic tissues positive staining was in 10% (n=1) and negative staining was in 90% (n=9) (P=0.011). And the expression of TPM4 was related to pT status, grade and stage (P<0.001, P=0.015 and P<0.001, respectively). Western blot results indicated that TPM4 was high expressed in HCC cell line and HCC tissues. In conclusion, we believe that TPM4 can be applied as a diagnostic and prognostic marker to assist the management of HCC.     

肝癌耐药细胞的形态学及微小RNA表达谱分析

目的 比较3株肝癌耐药细胞(Huh-7/ADM、Huh-7/CBP、Huh-7/MMC)与亲本肝癌细胞Huh-7的形态学差异;分析这3株肝癌耐药细胞及Huh-7的miRNA表达谱,筛选3株耐药细胞中异常表达的微小RNA(miRNA).方法 光学显微镜下观察各细胞株细胞形态学特点、透射电镜下观察各细胞株细胞内超微结构特点.miRNA芯片检测各细胞株miRNA表达谱,real-time定量PCR(qPCR)法验证芯片结果.结果 镜下见肝癌耐药细胞比亲本肝癌细胞Huh-7更具多形性,核异型性亦增加;细胞周边微绒毛减少;胞质增多,胞质内细胞器含量增加,并出现脂滴、糖原等成分.miRNA表达谱分析显示:与Huh-7相比,有32个miRNA在3种肝癌耐药细胞株中表达同时上调,有22个miRNA在3种耐药细胞株中表达同时下调.real-time qPCR检测证实miR-15a、miR-16、miR-27b、miR-30b、miR-146a、miR-146b-5p、miR-181a、miR-181d、miR-194在各肝癌耐药细胞中表达同时上调.结论 耐药细胞异型性增加,筛选出的肝癌耐药细胞中异常表达miRNA,可为研究肝癌多药耐药与miRNA的相关性提供依据.