Search

Background: We previously reported that modulation of cytokeratin18 induces pleomorphism of liver cells, higher cell motility, and higher drug sensitivity to sorafenib treatment of hepatoma cells. These relationships were established by in vitro experiments. The aim of this study was to determine the in vivo association between cytokeratin expression and tumor behavior, as well as cancer stem cells of hepatocellular carcinoma and intra-hepatic cholangiocarcinoma in Taiwan. Methods: Cytokeratins and sal-like protein 4 expression was determined in 83 hepatocellular carcinoma and 30 intra-hepatic cholangiocarcinoma specimens by immunohistochemistry. The relationship between cytokeratins and sal-like protein 4 expression with hepatitis virus infection, clinicopathologic factors, and survival was analyzed. Further, the correlation among cytokeratins and sal-like protein 4 expression was studied. Results: In addition to cytokeratin8/18, the expression of cytokeratin7/19 and sal-like protein 4 was noted in hepatocellular carcinoma; however, only cytokeratin19 expression had a significant correlation with poor overall survival and poor disease-free survival. The expression of cytokeratins and sal-like protein 4 was not correlated with hepatitis virus infection. The expression of cytokeratin19, but not 7, 8, and 18, was correlated with sal-like protein 4 expression in hepatocellular carcinoma. Cytokeratin7 expression was decreased and the sal-like protein 4 expression was absent in all 30 intra-hepatic cholangiocarcinoma cases. The expression of cytokeratins had not statistically significant correlation with overall and disease-free survival in patients with intra-hepatic cholangiocarcinoma. Conclusions: The expression of cytokeratin19 was associated with sal-like protein 4 expression, as well as poor overall and disease-free survival in hepatocellular carcinoma patients in Taiwan.

Unstable cytokeratins are associated with tumor transformation in the development of human hepatocellular carcinoma. We previously demonstrated that the cytokeratin 18 was modulated and that a histone H3-specific modification occured, among members of the histone family, during the development of human hepatocellular carcinoma. Evidence suggested that the modification of histone H3 was highly correlated with the modulation of cytokeratin 18 and probably plays an important role in tumorigenesis of hepatocytes. Aberrant expression of histone deacetylase leading to imbalance between acetylation and deacetylation of histones may exhibit regulatory roles in tumor transformation. Recently we found that overexpression of histone deacetylase-1 and hypoacetylation of histone H3 were associated with hepatocellular carcinoma. The underlying roles of histone H3 modulation are discussed in this mini-review article.

Synemin is a large intermediate filament protein that has been identified in all types of muscle cells. It plays a role in human muscle diseases; however, the role of synemin in tumor cell transformation has rarely been investigated. Because hepatocellular carcinoma cells are morphologically different from normal human hepatocytes, we hypothesized that altered synemin expression and cytoskeletal disorganization might underlie this pleomorphic transformation. To test this hypothesis, we studied synemin expression in hepatocellular carcinoma and liver tissues by immunohistochemistry and immunoblotting. In addition, we analyzed the expression level and organization of all cytoskeletal elements after synemin knock-down in human Chang liver cells. Previously we found that plectin knock-down in human Chang liver cells causes a reduction in cytokeratin 18 expression with effects on intermediate filament disorganization and altered cellular morphology. In this study we also compared the effects of synemin knock-down and plectin knock-down on the cytoskeleton expression and organization. The results revealed that synemin expression was down-regulated in human hepatocellular carcinoma compared with normal liver, which is similar to the plectin expression. Surprisingly, the expression of cytoskeletal elements (cytokeratin 18, actin and tubulin) was not influenced by synemin knock-down in human Chang liver cells. The organization of cytoskeletal networks was also unaltered after synemin knock-down. In conclusion, both plectin and synemin are down-regulated in human hepatocellular carcinoma in vivo and transformed human liver cell in vitro. However, the mechanism of cell transformation caused by synemin knock-down is different from that of plectin knock-down. Plectin, but not synemin, knock-down provoked liver cell transformation via suppressing cytokeratin 18 expression and disrupting intermediate filament networks. Synemin knock-down did not influence the cytoskeleton expression and organization of human Chang liver cells.

Background Plectin is one of the cytolinker proteins that play a crucial role in maintaining the integrity of cellular architecture. It is a component of desmosome complexes connecting cytoskeletal proteins and trans-membrane molecules. In epithelial cells, plectin connects cytokeratins and integrin α6β4 in hemidesmosomes anchoring to the extracellular matrix. In addition to the function of molecular adherent, plectin has been reported to exhibit functions affecting cellular signals and responsive activities mediated by stress, cellular migration, polarization as well as the dynamic movement of actin filaments. Plectin deficiency in hepatocellular carcinoma results in abnormal expression of cytokeratin 18 and disassembled hemidesmosome. Therefore, it is hypothesized that the plectin deficiency-mediated collapse of cytoskeleton may modulate cellular motility that is associated with consequent metastatic behaviors of cancer cells. Methods and results The cellular motility of plectin-deficient Chang liver cells generated by transient knockdown were analyzed by trans-well migration assay and the results revealed a higher migration rate. The confocal microscopy also demonstrated less organized and more polarized morphology as well as more focal adhesion kinase activity in comparison with that of the mock Chang liver cells. Furthermore, plectin-knockdown in Chang liver cells was associated with a higher activity of Rac1-GTPase in accordance with the results of the Rac1 pull-down assay. The immunohistochemical assay on human hepatocellular carcinoma showed that the expression of focal adhesion kinase was increased in the invasive front of tumor. Conclusion Plectin-deficient human hepatic cells exhibit higher cell motility associated with increase in focal adhesion kinase activity that are comparable to the properties of invasive hepatocellular carcinoma.

Plectin is a versatile cytoplasmic cross-linking protein that connects intermediate filaments to microfilaments, microtubules, and membrane adhesion sites. The cross-linking functions of plectin help organize the cytoskeleton into a stable meshwork important for maintaining uniformity in cell size and shape. As cells of hepatocellular carcinoma are morphologically different from normal human hepatocytes, we hypothesized that altered plectin expression and cytoskeletal organization underlies this pleomorphic transformation. To test this hypothesis, we analyzed expression levels and organization of all cytoskeletal elements, including intermediate filaments, microfilaments, and microtubules, after plectin knockdown in human Chang liver cells. We found that expression of cytokeratin 18, but not actin or tubulin, was downregulated by suppression of plectin protein. Furthermore, cytokeratin networks were partially collapsed and actin-rich stress fibers were increased. The organization of microtubule networks, by contrast, was unaltered. These findings support our hypothesis that, via effects on cytoskeletal organization, plectin deficiency might play an important role in the transformation of human liver cells.

Plectin is a cross-linking protein that organizes the cytoskeleton into a stable meshwork that helps maintain the uniform size and shape of cells. As cells of hepatocellular carcinoma are morphologically different from healthy human hepatocytes, we hypothesized that plectin deficiency and cytoskeletal disorganization underlies this pleomorphic transformation. To test this hypothesis we induced apoptosis as the most accessible pathway for creating plectin deficiency status in vivo. We analyzed expression levels and organization of plectin and other cytoskeletal elements, including intermediate filaments, microfilaments, and microtubules, after staurosporine-induced apoptosis in human Chang liver cells. The results revealed the expression of plectin and cytokeratin 18 were downregulated in hepatocellular carcinoma tissues in vivo. The expression of actin and tubulin, however, were not altered. In vitro analysis indicated that plectin and cytokeratin 18 were cleaved following staurosporine-treatment of human Chang liver cells. Time course experiments revealed that plectin was cleaved 2h earlier than cytokeratin 18. The organization of plectin and cytokeratin 18 networks collapsed after staurosporine-treatment. Conclusively, degradation of plectin induced by staurosporine-treatment in liver cells resulted in cytoskeleton disruption and induced morphological changes in these cells by affecting the expression and organization of cytokeratin 18.

Intermediate filaments are important in building the cellular architecture. Previously we found cytokeratin18 was modulated in human hepatocellular carcinoma. Plectin is a cross-linking protein that organizes the cytoskeleton into a stable meshwork, which can maintain the uniform size and shape of hepatocytes. Because the cells of hepatocellular carcinoma were morphologically different from the hepatocytes, we speculated that expression of plectin and organization of intermediate filament might play roles in the pleomorphism of hepatocellular carcinoma cells. In this paper, we studied the plectin expression of hepatocellular carcinoma and liver tissues by immunohistochemistry and immunoblot. The results revealed that plectin was deficient and cytokeratin18 was modulated in hepatocellular carcinoma. Furthermore, we knockdown the plectin mRNA in Chang cells, the result revealed the plectin was deficient and the organization of cytokeratin18 was altered. Conclusively, this study offers a hypothesis that plectin deficient might play an important role in the tumorigenesis of hepatocellular carcinoma.

Low-power laser therapy has become popular in clinical applications including promoting wound healing and pain relief. However, effects of this photoradiation on human hepatoma cells are rarely studied. Previously, we found 808 nm gallium aluminum arsenide (GaAlAs) continuous wave laser had an inhibitory effect on the proliferation of human hepatoma cell lines HepG2 and J-5 at the energy density of 5.85 and 11.7 J/cm(2), respectively. The aim of the present study was to evaluate the possible mechanism of action of this photoradiation on HepG2 and J-5 cells. HepG2 and J-5 cells were cultured in 24-well plates for 24 h. After photoradiation by 130 mW 808 nm GaAlAs continuous wave laser for different time intervals (0, 30, 60, 90, 120, 150, and 180 s), Western blot and immunofluorescent staining were used to examine the expression and distribution of histone and cytoskeletal proteins. The cell counts as well as histone and synemin expression of HepG2 and J-5 cells were reduced by photoradiation at the energy density of 5.85 and 11.7 J/cm(2), respectively. Furthermore, the architecture of cytoskeletons and the distribution of intermediate filament-associated proteins (plectin and synemin) were disorganized by photoradiation. Photoradiation by 808 nm GaAlAs continuous wave laser at the energy density of 5.85 and 7.8 J/cm(2) inhibited the survival of human hepatoma cell lines. The mechanism might reduce synthesis of histone and synemin. Reduced histone synthesis might further reduce the proliferation rate of these cells. Reduced synemin synthesis might result in the destruction of the cytoskeleton. Therefore, the net effects by this photoradiation were reduced cell survival.