Search

Background: Liver cancer stem cells (LCSCs) play key roles in the metastasis, recurrence, and chemotherapeutic resistance of hepatocellular carcinoma (HCC). Our previous research showed that the POSTN gene is closely related to the malignant progression and poor prognosis of HCC. This study aimed to elucidate the role of POSTN in generating LCSCs and maintaining their stemness as well as the underlying mechanisms., Methods: Human HCC tissues and matched adjacent normal tissues were obtained from 110 patients. Immunohistochemistry, western blotting (WB), and RT-PCR were performed to detect the expression of POSTN and stemness factors. The roles of transforming growth factor (TGF)-β1 and AP-2α in the POSTN-induced stemness transformation of HCC cells were explored in vitro and in vivo using LCSCs obtained by CD133 + cell sorting., Results: The high expression of POSTN was correlated with the expression of various stemness factors, particularly CD133, in our HCC patient cohort and in TCGA and ICGC datasets. Knockdown of POSTN expression decreased the abilities of HCC cell lines to form tumours in xenograft mouse models. Knockdown of POSTN expression also suppressed cell viability and clone formation, invasion, and sphere formation abilities in vitro. Knockdown of AP-2α attenuated the generation of CD133 + LCSCs and their malignant behaviours, indicating that AP-2α was a critical factor that mediated the POSTN-induced stemness transformation and maintenance of HCC cells. The role of AP-2α was verified by using a specific αvβ3 antagonist, cilengitide, in vitro and in vivo. Activation of POSTN could release TGFβ1 from the extracellular matrix and initiated POSTN/TGFβ1 positive feedback signalling. Furthermore, we found that the combined use of cilengitide and lenvatinib suppressed the growth of HCC cells with high POSTN expression more effectively than the use of lenvatinib alone in the patient-derived xenograft (PDX) mouse model., Conclusions: The POSTN/TGFβ1 positive feedback pathway regulates the expression of stemness factors and the malignant progression of HCC cells by regulating the transcriptional activation of AP-2α. This pathway may serve as a new target for targeted gene therapy in HCC.

Objective: To investigate the efficacy of celastrol treatment of hepatocellular carcinoma (HCC) cells in vitro and in vivo and to propose a mechanism of action., Methods: A human HepG2 liver cancer cell line and a xenograft tumor model were used to investigate the effects of celastrol on HCC in vitro and in vivo. A CCK-8 kit was used to detect cell viability. Flow cytometry and terminal-deoxynucleoitidyl transferase mediated nick end labeling staining were used to detect apoptosis. Western blotting and immunohistochemistry were used to detect the expression of cleaved-caspase-3, cleaved-caspase-8, cleaved-caspase-9, cleaved-PARP, mammalian target of rapamycin (mTOR), and p-mTOR. Hematoxylin-eosin staining was used to observe the tissue morphology., Results: Celastrol decreased the viability of HepG2 cells and induced apoptosis. Western blot assays indicated that celastrol up-regulated cleaved-caspase-3, cleaved-caspase-8, cleaved-caspase-9, and cleaved-PARP by inhibiting the phosphorylation of mTOR in HepG2 cells. Moreover, celastrol inhibited the tumor growth in a xenograft model. Celastrol also induced caspase-dependent apoptosis (up-regulation of cleaved-caspase- 3, -8, -9, and cleaved-PARP) and inhibited the activation of mTOR in vivo., Conclusion: Celastrol induces caspase-dependent apoptosis in HCC cells by inhibiting the activation of mTOR.

Background: Serum Golgi protein 73 (GP73) as a novel and potential marker for diagnosing hepatocellular carcinoma (HCC) have been found to be elevated in HCC patients and associated with clinical variables representing tumor growth and invasiveness. The aim of this study was to prepare a pair of monoclonal antibodys (mAbs) against GP73 and develop a newly designed double-antibody sandwich enzyme-linked immunosorbent assay (s-ELISA), which would be used in the detection of serum GP73 (sGP73) as well as in the diagnosis of HCC., Materials and Methods: Produced by prokaryotic expression, the purified recombinant GP73 (rGP73), produced by prokaryotic expression, was used to immunize the Balb/c mice. Two hybridoma cell lines against GP73 were obtained by fusing mouse Sp2/0 myeloma cells with spleen cells from the immunized mice. The titers of anti-GP73 mAb reached 1:243,000. Western blotting analysis and Immunohistochemistry staining revealed that anti-GP73 mAb could recognize GP73 protein. The double-antibody s-ELISA was successfully established and validated by 119 HCC and 103 normal serum samples., Results: showed that the detection limit of this method could reach 1.56 ng/ml, and sGP73 levels in HCC group (mean=190.6 ng/ml) were much higher than those of in healthy controls (mean=70.92 ng/ml)., Conclusions: Results of our study not only showed that sGP73 levels of HCC patients were significantly higher than those of healthy controls, but also indicated that the laboratory homemade anti-GP73 mAbs could be the optimal tool used in evaluating sGP73 levels, which would provide a solid foundation for subsequent clinical applications.

Only a few cases of pedunculated hepatocellular carcinoma (P-HCC) have been reported in the literature. The common sites of extrahepatic metastases in patients with HCC are the lungs, regional lymph nodes, kidney, bone marrow and adrenals. Metastasis to spleen is mostly via hematogenous metastasis, direct metastasis to spleen was very rare. We report a case of P-HCC presenting as a left upper abdominal lesions which involved the spleen that was actually a P-HCC with splenic metastasis. This case is unique as P-HCC directly involved the spleen which is not via hematogenous metastasis., (2009 The WJG Press and Baishideng. All rights reserved.)

Objective: To access the capability of 1H nuclear magnetic resonance (NMR) -based metabonomics in the evaluation of graft function in the perioperation period of liver transplantation., Methods: Plasma samples of 15 male primary hepatic carcinoma patients were collected for clinical biochemical analysis and 1H NMR spectroscopy 1 day before operation, 1 day and 1 week after the operation. The NMR data were analyzed using principal component analysis., Results: Metabonomic analysis indicated that, compared with those before operation, blood concentrations of valine, alanine, acetone, succinic acid, glutamine, choline, lactate, and glucose increased significantly 1 day after transplantation. One week later, the levels of lipids and choline increased notably, while those of glucose and amino acids decreased. Principal component analysis showed significant difference between metabolic profiles of plasma samples of variant periods of liver transplantation, due to the variation of the levels of glucose, lipids, lactate, and choline. A good agreement was observed between clinical chemistry and metabonomic data., Conclusions: Metabonomic analysis can clearly identify the difference between the plasma samples of primary hepatic carcinoma patients at different time during the perioperation period of liver transplantation. It therefore may be a promising new technology in predicting the outcomes of liver transplantation.

Aim: To develop a cancer vaccine of dendritic cells derived from human cord blood CD34+ cells and to investigate its cytotoxicity on human hepatocarcinoma cells in vitro and in severe combined immunodeficiency (SCID) mice., Methods: Lymphocytes from cord blood or peripheral blood were primed by DCs, which were derived from cord blood and pulsed with whole tumor cell lysates. Nonradiative neutral red uptake assay was adopted to detect the cytotoxicity of primed lymphocytes on human hepatocarcinoma cell line BEL-7402 in vitro. The anti-tumor effect of primed lymphocytes in vivo was detected in SCID mice, including therapeutic effect and vaccination effect., Results: The cytotoxicity of DC vaccine primed lymphocytes from cord blood or peripheral blood on human hepatocarcinoma cell line BEL-7402 was significantly higher than that of unprimed lymphocytes in vitro (44.09% vs 14.69%, 47.92% vs 19.44%, P<0.01). There was no significant difference between the cytotoxicity of primed lymphocytes from cord blood and peripheral blood (P>0.05). The tumor growth rate and tumor size were smaller in SCID mice treated or vaccinated with primed lymphocytes than those with unprimed lymphocytes. SCID mice vaccinated with primed lymphocytes had a lower tumor incidence (80% vs 100%, P<0.05) and delayed tumor latent period compared with mice vaccinated with unprimed lymphocytes (11 d vs 7 d, P<0.01)., Conclusion: Vaccine of cord blood derived-DCs has an inhibitory activity on growth of human hepatocarcinoma cells in vitro and in SCID mice. The results also implicate the potential role of cord blood derived-DC vaccine in clinical tumor immunotherapy.

Background: The resection rate of primary liver tumor in China is only about 20%. A lot of patients with moderate and advanced liver tumor may lose the chance of operation. The objective of present research was to study the efficacy of transcatheter arterial chemoembolization (TACE) combined with percutaneous injection of chemical agents and acetic acids in the treatment of patients with primary liver cancer (PLC)., Methods: Thirty-three patients with middle and advanced stage of PLC were divided into two groups: percutaneous injection of chemical agents and acetic acids (15 patients, group A) and TACE (18 patients, group B)., Results: Tumor diameter and serum AFP level reduced to 86.6% and 83.3% in group A, and 55.5% and 40% in group B, respectively. There was significant difference between the two groups (P<0.01). The 1, 2, 3, 4-year survival rates of group A were 96.7%, 86.6%, 51.3%, 33.3%, respectively and in group B were 66.7%, 44.4%, 16.7%, 0%, respectively (P<0.01)., Conclusion: TACE combined with percutaneous injection of chemical agents and acetic acids is efficacious to increase the survival rate of patients with PLC.

Aim: To develop atumor vaccine by fusion of H22 hepatocarcinoma cells and DC, and to study its protective and therapeutical effect against H22 cell., Methods: H22-DC vaccine was produced by PEG fusion of H22 and DC induced by cytokine released from splenic mononuclear cells, sorted by CD11c magnetic microbead marker. It was injected through the tail vein of the mice and the H(22)-DC oncogenesis was detected in the liver, spleen and lung. In order to study the therapeutical and protective effect of H(22)-DC against tumor H(22), two groups were divided: immune group and therapeutic group. Immune group was further divided into P, D, HD and H subgroups, immunized by PBS, DC, H(22)-DC and inactivated H(22), respectively, and attacked by H(22) cell. The tumor size, tumor weight, mice survival time and tumor latent period were recorded and statistically analyzed; Therapeutical group was divided into three subgroups of P, D and HD, and attacked by H(22), then treated with PBS, DC, and H(22)-DC, respectively. Pathology and flow cytometry were also applied to study the mechanism how the H(22)-DC vaccine attacked on the H(22) cell., Results: 1. No oncogenesis was found in spleen, lung and liver after H22-DC injection. 2. Hybrid vaccine immunized mice had strongest CTL activity. 3. In the immune group, latent period was longer in HD subgroup than that in P, H and D subgroup; and tumor size and weight were smaller in HD subgroup than that in P, H and D subgroup. 4. In therapeutic group, tumor size was smaller in HD subgroup than that in P, D subgroup., Conclusion: 1. H22-DC tumor vaccine is safe without oncogenesis in vivo. 2. Hybrid vaccine can stimulate potent specific CTL activity against H22. 3. H22-DC vaccine has distinctive prophylatic effect on tumor H22 and can inhibit the tumor growth.

Aim: To observe the biological specialization of human peripheral blood dendritic cells (DC) and cord blood derived DC and its effects on effector cells killing human hepatocarcinoma cell line BEL-7402. in vitro., Methods: The DC biological characteristics were detected with immunohistochemical and MTT assay. Two antitumor experiment groups are divided: peripheral blood DC and cord blood DC groups. Peripheral blood DC groups used LAK cells as the effector cells and BEL-7402 as target cells, while cord blood DC groups used CTL induced by tumor antigen twice pulsed DC as effector cells and BEL-7402 as target cells, additional peripheral blood DC and cord blood DC are added to observe its stimulating activities to effector cells. The effector's cytotoxicity to tumor cells were detected with neutral red colorimetric assay at two effector/target ratios of 5:1 and 10:1., Results: Peripheral blood DC and cord blood DC highly expressed HLA-ABC, HLA-DR, HLA-DQ, CD54 and S-100 protein. The stimulating activities to lymphocyte proliferation were compared between experimental groups (DC added) and control group (no DC added), in six experiment subgroups,the DC/lymphocyte ratio was sequentially 0.25:100, 0.5:100, 1:100, 2:100, 4:100 and 8:100.A values were sequentially 0.75396+/-0.009, 0.84916+/-0.010, 0.90894+/-0.012, 0.98371+/-0.007, 1.01299+/-0.006 and 1.20384+/-0.006 in peripheral blood DC groups and 0.77650+/-0.005, 0.83008+/-0.007, 0.92725+/-0.007, 1.05990+/-0.010, 1.15583+/-0.011, 1.22983+/-0.011 in cord blood DC groups. A value was 0.59517+/-0.005 in control group. The stimulating activities were higher in experimental groups than in control group (P<0.01), which were increased when the DC concentration was enlarged (P<0.01). Two differently derived DCs had the same phenotypes and similar stimulating activities (P<0.05). In peripheral blood DC groups, the cytotoxicity of the LD groups (experimental groups) and L groups (control group) was 58.16%+/-2.03% (5:1), 46.18%+/-2.25% (10:1) and 38.13%+/-1.29% (5:1) and 65.40%+/-1.56% (10:1) respectively; in cord blood DC groups, TD groups (experimental groups) and T groups (control groups) were 69.71%+/-2.33 % (5:1), 77.64%+/-1.94% (10:1) and 56.89%+/-1.82% (5:1) and 60.99%+/-1.42% (10:1) respectively.The cytotoxicity activities were enhanced with increased effector/target ratio (P<0.01). At the same effector/target ratio, the cytotoxicity of experimental groups were bigger than that of control groups (P<0.01). The cytotoxicity activities of cord blood DC groups were higher than that of peripheral blood DC groups (P<0.01)., Conclusion: Peripheral blood DC and cord blood DC are mature DC in morphology and function, both can enhance the effector cell killing activities to hepatocarcinoma cells. DC pulsed with tumor antigen can induce higher specific CTL activity than unpulsed DC.