[Show abstract][Hide abstract] ABSTRACT: Background:
The immunohistochemical detection of Enhancer of zeste homologue 2 (EZH2) proved to be a useful tool to recognize the malignant nature of tumors in a wide variety of neoplasms. The histological diagnostics of salivary gland tumors is a challenging task, and a reliable marker of malignancy would be extremely helpful.
EZH2 expression was investigated in 54 malignant and 40 benign salivary gland tumors of various histological types by standard immunohistochemistry.
The majority (n = 52) of the malignant tumors stained positively, while all the investigated benign tumors were negative for EZH2.
EZH2 expression in salivary gland tumors, similarly to the tumors of other organs is not characteristic for any tumor type, but is a solid marker of the malignant nature of the tumors.
[Show abstract][Hide abstract] ABSTRACT: Reproducible structural analysis was made on cirrhotic human liver samples in order to reveal potential connections between morphological and laboratory parameters.
Large histological samples were taken from segment VII of 56 cirrhotic livers removed in connection with liver transplantation. Picro Sirius red and immunohistochemically (smooth muscle actin [SMA], cytokeratin 7 [CK7], Ki-67) stained sections were digitalized and morphometric evaluation was performed.
The Picro Sirius-stained fibrotic area correlated with the average thickness of the three broadest septa, extent of SMA positivity, alkaline phosphatase (ALP) values and it was lower in the viral hepatitis related cirrhoses than in samples with non-viral etiology. The extent of SMA staining increased with the CK7-positive ductular reaction. The proliferative activity of the hepatocytes correlated positively with the Ki-67 labeling of the ductular cells and inversely with the septum thickness. These data support the potential functional connection among different structural components, for example, myofibroblasts, ductular reaction and fibrogenesis but challenges the widely proposed role of ductular cells in regeneration.
Unbiased morphological characterization of cirrhotic livers can provide valuable, clinically relevant information. Similar evaluation of routine core biopsies may increase the significance of this 'Gold Standard' examination.
[Show abstract][Hide abstract] ABSTRACT: We demonstrate that characteristic nodules emerge in cultures of several
malignant pleural mesothelioma (MPM) cell lines. Instead of excessive local
cell proliferation, the nodules arise by Myosin II-driven cell contractility.
The aggregation process can be prevented or reversed by suitable
pharmacological inhibitors of acto-myosin contractility. A cell-resolved
elasto-plastic model of the multicellular patterning process indicates that the
morphology and size of the nodules as well as the speed of their formation is
determined by the mechanical tension cells exert on their neighbors, and the
stability of cell-substrate adhesion complexes. A linear stability analysis of
a homogenous, self-tensioned Maxwell fluid indicates the unconditional presence
of a patterning instability.
[Show abstract][Hide abstract] ABSTRACT: Background: Comparison of tissue microarray results of 29 cervical cancer and 27 normal cervix tissue samples using immunohistochemistry revealed considerable reorganization of the fibrillar stroma of these tumors. Methods: In vitro models were used to throw light on the multifactorial process of tumor-stroma interaction, by means of studying the cooperation between tumor cells and fibroblasts. Fibroblasts from normal cervix and cervical cancers were grown either separately or in co-culture with CSCC7 cervical cancer cell line. Changes manifest in secreted glycoproteins, integrins and matrix metallo-proteases (MMPs) were explored. Results: While normal fibroblasts produced components of interstitial matrix and TGF-β1 that promoted cell proliferation, cancer-associated fibroblasts (CAFs) synthesized ample amounts of laminin-1. The following results support the significance of laminin-1 in the invasion of CSCC7 cells: 1.) Tumor-associated fibroblasts produced more laminin-1 and less components of fibrillar ECM than normal cells; 2.) The production of laminin chains was further increased when CSCC7 cells were grown in co-culture with fibroblasts; 3.) CSCC7 cells were capable of increasing their laminin production; 4.) Tumor cells predominantly expressed integrin α6β4 laminin receptors and migrated towards laminin. The integrin profile of both normal and tumor-associated fibroblasts was similar, expressing receptors for fibronectin, vitronectin and osteopontin. MMP-7 secreted by CSCC7 cells was upregulated by the presence of normal fibroblasts, whereas MMP-2 produced mainly by fibroblasts was activated in the presence of CSCC7 cells. Conclusions: Our results indicate that in addition to degradation of the basement membrane, invasion of cervical cancer is accomplished by the remodeling of the interstitial stroma, which process includes decrease and partial replacement of fibronectin and collagens by a laminin-rich matrix.
BMC Cancer 04/2015; 15(1):256. DOI:10.1186/s12885-015-1272-3 · 3.36 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The appearance of lung metastases is associated with poor outcome and the management of patients with secondary pulmonary tumours remains a clinical challenge. We examined the vascularisation process of lung metastasis in six different preclinical models and found that the tumours incorporated the pre-existing alveolar capillaries (i.e. vessel co-option). During the initial phase of vessel co-option, the incorporated capillaries were still sheathed by pneumocytes, but these incorporated vessels subsequently underwent different fates dependent on the model. In five of the models examined (B16, HT1080, HT25, C26 and MAT B-III), the tumour cells gradually stripped the pneumocytes from the vessels. These dissected pneumocytes underwent fragmentation, but the incorporated microvessels survived. In the sixth model (C38), the tumour cells failed to invade the alveolar walls. Instead, they induced the development of vascularised desmoplastic tissue columns. Finally, we examined the process of arterialisation in lung metastases and found that they became arterialized when their diameter grew to exceed 5 mm. In conclusion, our data show that lung metastases can vascularise by co-opting the pulmonary microvasculature. This is likely to have important clinical implications, especially with respect to anti-angiogenic therapies.
The Journal of Pathology 02/2015; 235(3). DOI:10.1002/path.4464 · 7.43 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Background and purpose:
The clinical effects of anti-angiogenic agents remain controversial. Therefore, elucidating the pharmacological properties of these compounds is a pivotal issue.
The effects of treatment with sunitinib on tumour and normal tissues of mice bearing C-26 adenocarcinoma cells were analysed by matrix-assisted laser desorption ionization MS imaging (MALDI-MSI). Expression of the key targets of sunitinib--angiogenic receptors--was studied by immunofluorescent labelling.
MALDI-MS assays showed that sunitinib and its fragment ions were present throughout tumour and normal tissues. Major metabolites were identified in blood and solid tissues, while minor drug metabolites were detectable only in blood. Tumour growth and intratumour VEGF receptor-2 expressions were significantly reduced in sunitinib-treated mice, while the expression of the other targeted receptors, PDGF receptor -α or -β and fibroblast growth factor receptor-1, remained unaffected. Within tumour tissue, the close proximity of sunitinib metabolites to the precursor ion suggested in situ metabolism of the administered drug. There were intratumour areas where the signal intensity of sunitinib correlated with expression of VEGF receptor-2.
Conclusions and implications:
This is the first study that demonstrates MALDI-MSI is a versatile platform to study the intratumour localization of an unlabelled anti-angiogenic drug. The combination of MALDI-MSI and immunofluorescence analysis can provide further insights into the molecular interaction of drug compounds and their targets within tumour tissue.
British Journal of Pharmacology 11/2014; 172(4). DOI:10.1111/bph.12990 · 4.84 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Whereas the role of the G-protein-coupled APJ receptor and its ligand, apelin, in angiogenesis has been well documented, the ability of the apelin/APJ system to induce lymphangiogenesis and lymphatic metastasis has been largely unexplored. To this end, we first show that APJ is expressed in lymphatic endothelial cells (LECs) and, moreover, that it responds to apelin by activating the apelinergic signaling cascade. We find that although apelin treatment does not influence the proliferation of LECs in vitro, it enhances their migration, protects them against UV irradiation-induced apoptosis, increases their spheroid numbers in 3D culture, stimulates their in vitro capillary-like tube formation and, furthermore, promotes the invasive growth of lymphatic microvessels in vivo in the matrigel plug assay. We also demonstrate that apelin overexpression in malignant cells is associated with accelerated in vivo tumor growth and with increased intratumoral lymphangiogenesis and lymph node metastasis. These results indicate that apelin induces lymphangiogenesis and, accordingly, plays an important role in lymphatic tumor progression. Our study does not only reveal apelin as a novel lymphangiogenic factor but might also open the door for the development of novel anticancer therapies targeting lymphangiogenesis.
[Show abstract][Hide abstract] ABSTRACT: A 2D model was previously presented that describes the gliding motility of human fibrosarcoma cells. The model was based on the observation that adhesions are present only on the outer rim of the leading lamella of the semicircular cell. The present model describes the organization of adhesions and the cytoskeleton of migrating HT1080 fibrosarcoma and LX2 hepatic stellate cells in three dimensions. The migration assays were performed in a modified Boyden chamber using fibronectin, Matrigel, or collagen I as chemoattractants. The distribution of the adhesions was analyzed by confocal laser scanning microscope, and following decoration with heavy meromyosin, the organization of actin filaments was analyzed by electron microscopy. Double labeling was performed to study the relationship of the actin and vimentin filament network in the moving cells. Vinculin containing adhesions were observed only at the front of the cell in the form of a ring while passing through a filter pore of the Boyden chamber. Actin filaments were present only below the plasma membrane, except the very tip of the leading lamella. Vimentin intermediate filaments were localized around the cell nucleus behind the actin filament-rich lamella.This paper describes a model of the organization of adhesions and the cytoskeleton of migrating cells in the Boyden chamber. The model is based on the observation that adhesions are present only at the leading edge of the cell. The results extend the earlier 2D model of cell locomotion into 3D.
[Show abstract][Hide abstract] ABSTRACT: The hepatic stem cells reside periportally forming the canals of Hering in normal liver. They can be identified by their unique immunophenotype in rat. The oval cells, the progenies of stem cells invade deep the liver parenchyma following activation and differentiate into focally arranged small - and eventually trabecularly ordered regular hepatocytes. We have observed that upon the completion of intense oval cell reactions narrow ductular structures are present in the parenchyma, we propose to call them parenchymal ductules. These parenchymal ductules have the same immunophenotype (CK7-/CK19+/AFP-/DLK-) as the resting stem cells of the canals of Hering, but different from them reside scattered in the parenchyma. In our present experiments we have investigated in an in vivo functional assay if the presence of these parenchymal ductules has any impact on a progenitor cell driven regeneration process. Parenchymal ductules were induced either by an established model of oval cell induction consisting of the administration of necrogenic dose of carbontetrachloride to 2-acetaminofluorene pretreated rats (AAF/CCl4) or a large necrogenic dose of diethylnitrosamine (DEN). The oval cells expanded faster and the foci evolved earlier after repeated injury in the livers with preexistent parenchymal ductules. When the animals were left to survive for one more year increased liver tumor formation was observed exclusively in the DEN treated rats. Thus repeated oval cell reactions are not necessarily carcinogenic. We conclude, that the expansion of hepatic stem cell compartment conceptually can be used to facilitate liver regeneration without an increased risk of tumorigenesis.
Stem cells and development 08/2013; 23(1). DOI:10.1089/scd.2013.0202 · 3.73 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: We have analyzed the architectural aspects of progenitor-cell-driven regenerative growth in rat liver by applying the 2-acetaminofluorene/partial hepatectomy experimental model. The regeneration is initiated by the proliferation of so-called oval cells. The oval cells at the proximal tips of the ductules have a more differentiated phenotype and higher proliferative rate. This preferential growth results in the formation of a seemingly random collection of small hepatocytes, called foci. These foci have no clonal origin, but possess a highly organized structure, which shows similarities to normal hepatic parenchyma. Therefore, they can easily remodel into the lobular structure. Eventually, the regenerated liver is constructed by enlarged hepatic lobules; no new lobules are formed during this process. The foci of the Solt-Farber experimental hepatocarcinogenesis model have identical morphological features; accordingly, they also represent only regenerative, not neoplastic, growth. Conclusion: Progenitor-cell-driven liver regeneration is a well-designed, highly organized tissue reaction, and better comprehension of the architectural events may help us to recognize this process and understand its role in physiological and pathological reactions. (HEPATOLOGY 2012).
[Show abstract][Hide abstract] ABSTRACT: The immunohistochemical demonstration of Enhancer of zeste homologue 2 (EZH2) proved to be a useful marker in several tumor types. It has been described to distinguish reliably hepatocellular carcinomas from liver adenomas and other benign hepatocellular lesions. However, no other types of malignant liver tumors were studied so far.
To evaluate the diagnostic value of this protein in hepatic tumors we have investigated the presence of EZH2 by immunohistochemistry in hepatocellular carcinomas and other common hepatic tumors.EZH2 expression was examined in 44 hepatocellular carcinomas, 23 cholangiocarcinomas, 31 hepatoblastomas, 16 other childhood tumor types (rhabdomyosarcoma, neuroblastoma, Wilms' tumor and rhabdoid tumor), 17 metastatic liver tumors 24 hepatocellular adenomas, 15 high grade dysplastic nodules, 3 biliary cystadenomas, 3 biliary hamartomas and 3 Caroli's diseases.
Most of the malignant liver tumors were positive for EZH2, but neither of the adenomas, cirrhotic/dysplastic nodules, reactive and hamartomatous biliary ductules stained positively.
Our immunostainings confirm that EZH2 is a sensitive marker of hepatocellular carcinoma, but its specificity is very low, since almost all the investigated malignant liver tumors were positive regardless of their histogenesis. Based on these results EZH2 is a sensitive marker of malignancy in hepatic tumors. In routine surgical pathology EZH2 could be most helpful to diagnose cholangiocarcinomas, because as far as we know this is the first marker to distinguish transformed and reactive biliary structures. Although hepatoblastomas also express EZH2, the diagnostic significance of this observation seems to be quite limited whereas, the structurally similar, other blastic childhood tumors are also positive. VIRTUAL SLIDES: The virtual slide(s) for this article can be found here: http://www.diagnosticpathology.diagnomx.eu/vs/1173195902735693.
[Show abstract][Hide abstract] ABSTRACT: Syndecans are transmembrane heparan sulphate proteoglycans. Their role in the development of the malignant phenotype is ambiguous and depends upon the particular type of cancer. Nevertheless, syndecans are promising targets in cancer therapy, and it is important to elucidate the mechanisms controlling their various cellular effects. According to earlier studies, both syndecan-1 and syndecan-2 promote malignancy of HT-1080 human fibrosarcoma cells, by increasing the proliferation rate and the metastatic potential and migratory ability, respectively. To better understand their tumour promoter role in this cell line, syndecan expression levels were modulated in HT-1080 cells and the growth rate, chemotaxis and invasion capacity were studied. For in vivo testing, syndecan-1 overexpressing cells were also inoculated into mice. Overexpression of full length or truncated syndecan-1 lacking the entire ectodomain but containing the four juxtamembrane amino acids promoted proliferation and chemotaxis. These effects were accompanied by a marked increase in syndecan-2 protein expression. The pro-migratory and pro-proliferative effects of truncated syndecan-1 were not observable when syndecan-2 was silenced. Antisense silencing of syndecan-2, but not that of syndecan-1, inhibited cell migration. In vivo, both full length and truncated syndecan-1 increased tumour growth and metastatic rate. Based on our in vitro results, we conclude that the tumour promoter role of syndecan-1 observed in HT-1080 cells is independent of its ectodomain; however, in vivo the presence of the ectodomain further increases tumour proliferation. The enhanced migratory ability induced by syndecan-1 overexpression is mediated by syndecan-2. Overexpression of syndecan-1 also leads to activation of IGF1R and increased expression of Ets-1. These changes were not evident when syndecan-2 was overexpressed. These findings suggest the involvement of IGF1R and Ets-1 in the induction of syndecan-2 synthesis and stimulation of proliferation by syndecan-1. This is the first report demonstrating that syndecan-1 enhances malignancy of a mesenchymal tumour cell line, via induction of syndecan-2 expression.
PLoS ONE 06/2012; 7(6):e39474. DOI:10.1371/journal.pone.0039474 · 3.23 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The proliferative response of hepatocytes in vivo can be induced by two mechanisms: severe damage to hepatic tissue results in regenerative growth and so-called primary hepatocyte mitogens can initiate liver cell proliferation without preceding loss of parenchyma. The regulation of the two responses is quite different. The decreased regenerative response of cirrhotic/fibrotic liver is well known, and is a severe obstacle to surgery of the diseased liver. In the present experiments we investigated the efficiency of a primary hepatocyte mitogen 1,4-Bis[2-(3,5-dichloropyridyloxy)]benzene (TCPOBOB) on two different liver cirrhosis/fibrosis models in mice induced by chronic administration of CCl(4) and thioacetamide respectively. BrdU incorporation and cyclin A expression established clearly that there is a reduced but still powerful mitogenic response of the fibrotic livers. Therefore, primary hepatocyte mitogens appear to be suitable to be used to rescue the regenerative response of cirrhotic livers.
International Journal of Experimental Pathology 01/2012; 93(2):125-9. DOI:10.1111/j.1365-2613.2011.00803.x · 2.17 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Angiogenesis is believed to be essential for the growth of metastatic tumors in the brain. We analyzed the vascularization of tumors formed by 4 epithelial cell lines (C38, ZR75, HT25, and H1650) and a fibrosarcoma (HT1080) cell line injected into the brains of mice. No peritumoral angiogenesis was observed. Tumors apparently acquired their vasculature by incorporation of native vessels. Vessel density was lower, but vessel diameter and vascular cell proliferation were higher within all tumors versus those in the peritumoral tissue. There was an inverse correlation between the number of incorporated vessels and vascular cell proliferation. Epithelial tumors with pushing growth patterns had lower vessel density and elevated vascular cell proliferation compared with invasive tumors. The incorporated vessels retained their normal structure, with the exception of astrocyte foot processes that were replaced by tumor cells. Attachment to the vascular basement membrane led to the differentiation of the ZR75 breast cancer cells. In the HT1080 metastases, there was intussusceptive angiogenesis, that is, the fibrosarcoma cells that were attached to the vessel caused lumen splitting and filled the developing pillars. Branching angiogenesis was not observed either in the tumors or in control cerebral wounds. These data suggest that sprouting angiogenesis is not needed for the incipient growth of cerebral metastases and that tumor growth in this model is a result of incorporation of host vessels.
Journal of Neuropathology and Experimental Neurology 11/2011; 70(11):979-91. DOI:10.1097/NEN.0b013e318233afd7 · 3.80 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: One of the hallmarks of intussusceptive angiogenesis is the development of intraluminal connective tissue pillars. The exact mechanism of pillar formation has not yet been elucidated. By using electron and confocal microscopy, we observed intraluminal nascent pillars that contain a collagen bundle covered by endothelial cells (ECs) in the vasculature of experimental tumors. We proposed a new mechanism for the development of these pillars. First, intraluminal endothelial bridges are formed. Second, localized dissolution of the basement membrane occurs and a bridging EC attaches to a collagen bundle in the underlying connective tissue. A pulling force is then exerted by the actin cytoskeleton of the ECs via specific attachment points, which contain vinculin, to the collagen bundle, resulting in suction and subsequent transport of the collagen bundle into and through the vessel lumen. Third, the pillar matures through the immigration of connective tissue cells and the deposition of new collagenous connective tissue. The proposed simple mechanism generates a connection between the processes of endothelial bridging and intussusceptive angiogenesis and identifies the source of the force behind pillar formation. Moreover, it ensures the rapid formation of pillars from pre-existing building blocks and the maintenance of EC polarity. To describe it, we coined the term inverse sprouting.
American Journal Of Pathology 09/2011; 179(3):1573-85. DOI:10.1016/j.ajpath.2011.05.033 · 4.59 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Male infertility is an increasing problem in all domestic species including man. Localization and identification of genes involved in defects causing male infertility provide valuable information of specific events in sperm development. Correct condensation of the sperm head and development of the acrosome are required for fertile sperm. In the Finnish Yorkshire pig population a knobbed acrosome defect (KAD) has been reported which appears to be of genetic origin. In previous studies we have shown that a large number of affected spermatozoa have a cystic swelling anterior to the apical part of the acrosome.
Characterization of the knobbed acrosome affected sperm revealed that both the acrosomal granules and chromatin are affected. This type of KAD appears to be a previously unknown and serious form of the defect. A genome wide scan with PorcineSNP60 Genotyping BeadChip defined the KAD associated region within 0.7 Mbp on porcine chromosome 15. Two genes, STK17b and HECW2, located within this region were sequenced. The expression of these genes appeared comparable in KA-affected and control boars. The known function of HECW2 in acrosome development highlighted this gene as a good candidate responsible for the KAD. One nonsynonymous SNP was identified within the HECW2 gene. However, as this mutation was found in homozygous state in individuals with normal sperm, this is not likely to be the causal mutation.
In this study we identified two candidate genes for a severe defect affecting both the sperm acrosome and chromatin that causes infertility. One of these genes, HECW2, plays an important role in ubiquitination, a prerequisite for chromatin remodelling and acrosome formation, highlighting the involvement of this gene in the knobbed acrosome defect and male infertility.
[Show abstract][Hide abstract] ABSTRACT: Transforming growth factor (TGF)-β-1 is a very efficient inhibitor of hepatocyte proliferation in various in vivo and in vitro experimental systems. However, there are no data on whether it can influence the mitogenic response induced by primary hepatocyte mitogens.
In this study, we compared the proliferative response in the liver between wild-type and transgenic mice, overexpressing active TGF-β-1 in their liver following the treatment by a primary hepatocyte mitogen TCPOBOP (1,4-bis[2-(3,5-dichloropyridyloxy)]benzene).
The proliferative response was characterized by the immunohistochemical examination of pulse and cumulative bromodeoxyuridine labelling and by quantitative real-time polymerase chain reaction analysis of cell cycle-related genes.
Neither of the applied techniques revealed significant differences between the two groups of mice; furthermore, we observed the upregulation of TGF-β-1 expression following the mitogenic treatment.
TGF-β-1 does not inhibit the primary mitogen-induced proliferative response of the hepatocytes. This observation may provide an explanation for the divergent consequences of hepatic proliferations induced by partial hepatectomy or primary mitogenic treatment.
Liver international: official journal of the International Association for the Study of the Liver 11/2010; 30(10):1505-10. DOI:10.1111/j.1478-3231.2010.02324.x · 4.85 Impact Factor