Katalin Dobra - Academia.edu (original) (raw)

Papers by Katalin Dobra

Experimental Cell Research, 2002

Syndecans are transmembrane proteoglycans implicated in the regulation of cell growth and differe... more Syndecans are transmembrane proteoglycans implicated in the regulation of cell growth and differentiation, by interacting with growth factors. Although syndecans play a major role in regulating cell morphology, little is known about their subcellular distribution and in vivo association with the cytoskeleton. To address this question, we investigated the subcellular distribution and dynamic rearrangement of syndecans-1,-2, and-4, using confocal laser microscopy. Furthermore, we monitored the spatial relation of syndecans to tubulin in both mitotic and interphase cells. Initially, the reactivity to syndecans was confined to the cytoplasm, staining of the cell membranes appearing later. Syndecan-1 also seems to translocate to the nucleus in a time-dependent manner. The mitotic spindle shows unexpectedly more syndecans than that found in interphase cells. After vinblastine treatment, both syndecan-1 and tubulin were recovered as paracrystalline occlusion bodies, and the nuclear reactivity to syndecan-1 disappeared, suggesting tubulin-mediated nuclear transport of this proteoglycan. Plasma membrane staining reappeared in the postmitotic cells. Nuclear translocation predominantly affected syndecan-1, whereas syndecan-2 and-4 remained in cytoplasm and cell membrane. This is the first report on regulated nuclear translocation and the presence of syndecan-1 in the mitotic spindle, where it may stabilize the mitotic machinery. The syndecan-1/ tubulin complex may also act as a vehicle for the transport of protein growth factors to the cell nucleus.

Differentiation, 2000

Malignant mesothelioma is a tumour originating from mesothelial cells, and it exhibits epithelial... more Malignant mesothelioma is a tumour originating from mesothelial cells, and it exhibits epithelial, fibrous, or biphasic differentiation. This tumour is highly resistant to therapy, and presence of a sarcomatous growth pattern has been associated with worse prognosis. A mesothelioma cell line with retained ability to differentiate into either epithelial or fibroblast-like phenotype was subjected to subtractive hybridisation in order to identify the genes coupled to tumour cell differentiation. Nine genes were found to be selectively overexpressed in the epithelial sub-line, compared to only two genes in the fibroblast-like phenotype. This may support the idea that the sarcomatous phenotype represents a less differentiated tumour. One of the genes that is differentially expressed by the epithelial cells was thioredoxin, a small redox-active protein associated with cell-growth and differentiation. This overexpression was accompanied by increased protein levels both intracellularly and in the medium. Thioredoxin is reduced by the selenoprotein thioredoxin reductase and NADPH. The activity of this enzyme was high in both cell sub-lines but induced 2-fold in the epithelially-differentiated cells. Overexpression of thioredoxin might be a factor behind the poor prognosis and reduced responsiveness to therapy of mesotheliomas. Epithelial differentiation in this cell line has previously been linked to increased synthesis of heparan sulphate proteoglycans. The possible formation of complexes including thioredoxin, thioredoxin re

Biomolecules

Extracellular vesicles (EVs), comprising exosomes, microvesicles, and apoptotic bodies, are relea... more Extracellular vesicles (EVs), comprising exosomes, microvesicles, and apoptotic bodies, are released by all cells into the extracellular matrix and body fluids, where they play important roles in intercellular communication and matrix remodeling in various pathological conditions. Malignant pleural mesothelioma (MPM) is a primary tumor of mesothelial origin, predominantly related to asbestos exposure. The detection of MPM at an early stage and distinguishing it from benign conditions and metastatic adenocarcinomas (AD) is sometimes challenging. Pleural effusion is often the first available biological material and an ideal source for characterizing diagnostic and prognostic factors. Specific proteins have previously been identified as diagnostic markers in effusion, but it is not currently known whether these are associated with vesicles or released in soluble form. Here, we study and characterize tumor heterogeneity and extracellular vesicle diversity in pleural effusion as diagnost...

Biomolecules

Malignant pleural mesothelioma (MPM) is a highly aggressive and therapy resistant pleural maligna... more Malignant pleural mesothelioma (MPM) is a highly aggressive and therapy resistant pleural malignancy that is caused by asbestos exposure. MPM is associated with poor prognosis and a short patient survival. The survival time is strongly influenced by the subtype of the tumor. Dyspnea and accumulation of pleural effusion in the pleural cavity are common symptoms of MPM. The diagnostic distinction from other malignancies and reactive conditions is done using histopathology or cytopathology, always supported by immunohistochemistry, and sometimes also by analyses of soluble biomarkers in effusion supernatant. We evaluated the soluble angiogenesis related molecules as possible prognostic and diagnostic biomarkers for MPM by Luminex multiplex assay. Pleural effusion from 42 patients with malignant pleural mesothelioma (MPM), 36 patients with adenocarcinoma (AD) and 40 benign (BE) effusions were analyzed for 10 different analytes that, in previous studies, were associated with angiogenesis...

Journal of Clinical Pathology

AimsAccurate and reliable diagnosis is essential for lung cancer treatment. The study aim was to ... more AimsAccurate and reliable diagnosis is essential for lung cancer treatment. The study aim was to investigate interpathologist diagnostic concordance for pulmonary tumours according to WHO diagnostic criteria.MethodsFifty-two unselected lung and bronchial biopsies were diagnosed by a thoracic pathologist based on a broad spectrum of immunohistochemical (IHC) stainings, molecular data and clinical/radiological information. Slides stained with H&E, thyroid transcription factor-1 (TTF-1) clone SPT24 and p40 were scanned and provided digitally to 20 pathologists unaware of reference diagnoses. The pathologists independently diagnosed the cases and stated if further diagnostic markers were deemed necessary.ResultsIn 31 (60%) of the cases, ≥80% of the pathologists agreed with each other and with the reference diagnosis. Lower agreement was seen in non-small cell neuroendocrine tumours and in squamous cell carcinoma with diffuse TTF-1 positivity. Agreement with the reference diagnosis range...

Cancers

Malignant mesothelioma (MM) is a rare but highly aggressive cancer that primarily originates from... more Malignant mesothelioma (MM) is a rare but highly aggressive cancer that primarily originates from the pleura, peritoneum or pericardium. There is a well-established link between asbestos exposure and progression of MM. Direct invasion of the surrounding tissues is the main feature of MM, which is dependent on dysregulated communication between the mesothelium and the microenvironment. This communication is dependent on the dynamic organization of the cytoskeleton. We have analyzed the organization and function of key cytoskeletal components in MM cell lines of increasing malignancies measured as migratory and invasive properties, and we show that highly malignant and invasive MM cells have an organization of the actin filament and vimentin systems that is distinct from the less malignant MM cell lines. In addition, the Hippo tumor suppressor pathway was inactivated in the invasive MM cells, which was seen as increased YAP nuclear localization.

Cancers

Malignant mesothelioma (MM) is an aggressive tumor of the serosal cavities. Angiogenesis is impor... more Malignant mesothelioma (MM) is an aggressive tumor of the serosal cavities. Angiogenesis is important for mesothelioma progression, but so far, anti-angiogenic agents have not improved patient survival. Our hypothesis is that better understanding of the regulation of angiogenesis in this tumor would largely improve the success of such a therapy. Syndecan-1 (SDC-1) is a transmembrane heparan sulfate proteoglycan that acts as a co-receptor in various cellular processes including angiogenesis. In MM, the expression of SDC-1 is generally low but when present, SDC-1 associates to epithelioid differentiation, inhibition of tumor cell migration and favorable prognosis, meanwhile SDC-1 decrease deteriorates the prognosis. In the present study, we studied the effect of SDC-1 overexpression and silencing on MM cells ability to secrete angiogenic factors and monitored the downstream effect of SDC-1 modulation on endothelial cells proliferation, wound healing, and tube formation. This was done ...

Translational Lung Cancer Research

The global incidence of malignant mesothelioma (MM) causes considerable disease burden, suffering... more The global incidence of malignant mesothelioma (MM) causes considerable disease burden, suffering and health care costs. Beside preventive measures and ban the use of asbestos, early diagnosis would largely improve the chance of curative treatment. Current histologic criteria, however, requiring presence of invasion in the surrounding fatty tissue fail to identify MM in sufficiently early stage. Unilateral accumulation of pleural effusion is one of the earliest clinical manifestations of MM that occurs in approximately 90% of the patients. Therapeutic thoracocenthesis is necessary to remove the fluid and to relieve patients' symptoms. This effusion is easily accessible and offers early and minimally invasive diagnosis by combining cytology with immunologic, molecular-and biomarker analyses. Typically, the fluid is rich in malignant cells and cell groups, but incipient stages of the disease may be difficult to recognize as the malignant cells can be masked by presence of inflammatory or reactive mesothelial cells. Recurrent, hemorrhagic and cell rich effusion should always be suspicious for MM and adequately prepared and analyzed to provide necessary information for subsequent therapy. Importantly, early detection of MM by integrating cytology and molecular approaches has high sensitivity and positive predictive value and has a major impact on patient survival. Thus, a conclusive positive MM cytology should lead to treatment without delay. This review summarizes molecular and diagnostic criteria of MM diagnosis.

Cytopathology

Serous effusion fluid is one of the most commonly encountered specimens in routine cytopathology ... more Serous effusion fluid is one of the most commonly encountered specimens in routine cytopathology practice. It provides invaluable information about the patient and the clinical status; but to get the most of it, specimen handling and processing must be carried out properly. Cytomorphology is the basis of a successful analysis which should complemented by ancillary tests when needed. A wide spectrum of ancillary techniques - ranging from immunocytochemistry and flow cytometry to different assays of molecular pathology - can be applied to serous effusions. This article describes the acquisition and management of serous effusion fluids, methods for preservation and transportation, different techniques of cytopreparation, application of immunocytochemistry, flow cytometry, and fluorescence in-situ hybridization (FISH), as well as DNA extraction for polymerase chain reaction (PCR) and next generation sequencing (NGS). Principles of bio-banking of effusion samples are also discussed which is getting more important in correlation with the developments in personalized medicine. This article is protected by copyright. All rights reserved.

Cancer Cytopathology

BACKGROUND: Malignant mesothelioma (MM) is an aggressive, fatal tumor. Current therapeutic option... more BACKGROUND: Malignant mesothelioma (MM) is an aggressive, fatal tumor. Current therapeutic options only marginally improve survival. Programmed cell death ligand 1 (PD-L1) is a dominant mediator of immunosuppression, binding to programmed cell death 1 (PD-1). PD-L1 is up-regulated in cancer cells, and the PD-1/PD-L1 pathway plays a critical role in tumor immune evasion, thus providing a target for antitumor therapy. Further, a correlation between PD-L1 expression and prognosis has been reported. Studies performed on histological material have revealed expression of PD-L1 in MM, but no study has been performed on MM effusions thus far. METHODS: PD-L1 expression was determined by a commercially available antibody (clone 28-8) in 74 formalin-fixed, paraffin-embedded cell blocks from body effusions obtained at diagnosis from patients with MM. The presence of MM cells was confirmed with CK5/6, calretinin, and EMA and the admixture of macrophages was assessed with CD68. Only cases containing more than 100 tumor cells were assessed. Membranous staining in tumor cells was considered positive. Survival time was calculated from the appearance of the first malignant effusion until death. RESULTS: Reactivity was observed in 23 of 61 (38%) of cases and was classified as 1%-5% (n 5 9 cases), >5%-10% (n 5 4 cases), >10%-50% (n 5 4 cases), and >50% (n 5 6 cases) positive cells. Survival times did not differ significantly between patients with PD-L1-positive and PD-L1-negative tumors. CONCLUSION: MM effusions are suitable for immune-cytochemical assessment of PD-L1 expression in malignant cells and the results are similar to those reported for histological specimens.

Matrix Biology, 2016

Synthesis, deposition, and interactions of hyaluronan (HA) with its cellular receptor CD44 are cr... more Synthesis, deposition, and interactions of hyaluronan (HA) with its cellular receptor CD44 are crucial events that regulate the onset and progression of tumors. The intracellular signaling pathways initiated by HA interactions with CD44 leading to tumorigenic responses are complex. HA molecules may perform dual functions depending on their concentration and size. Overexpression of variant isoforms of CD44 (CD44v) is most commonly linked to cancer progression, whereas their loss is associated with inhibition of tumor growth. In this review, we highlight that regulation of HA synthases (HASes) by post-translational modifications such as O-GlcNAcylation and ubiquitination, or by environmental factors or the action of microRNAs is important for HA synthesis and secretion in the tumor microenvironment. Moreover, we focus on the roles and interactions of CD44 with various proteins that reside extra-and intracellularly, as well as on cellular membranes with particular reference to the CD44-HA axis in cancer stem cell functions, and the importance of CD44/CD44v6 targeting to inhibit tumorigenesis in colon cancer cell models.

Journal of translational medicine, Oct 19, 2016

The goal of biomarker research is to identify clinically valid markers. Despite decades of resear... more The goal of biomarker research is to identify clinically valid markers. Despite decades of research there has been disappointingly few molecules or techniques that are in use today. The "1st International NTNU Symposium on Current and Future Clinical Biomarkers of Cancer: Innovation and Implementation", was held June 16th and 17th 2016, at the Knowledge Center of the St. Olavs Hospital in Trondheim, Norway, under the auspices of the Norwegian University of Science and Technology (NTNU) and the HUNT biobank and research center. The Symposium attracted approximately 100 attendees and invited speakers from 12 countries and 4 continents. In this Symposium original research and overviews on diagnostic, predictive and prognostic cancer biomarkers in serum, plasma, urine, pleural fluid and tumor, circulating tumor cells and bioinformatics as well as how to implement biomarkers in clinical trials were presented. Senior researchers and young investigators presented, reviewed and vi...

Malignant mesothelioma is a highly aggressive tumor with median survival ranging from 4 to 12 mon... more Malignant mesothelioma is a highly aggressive tumor with median survival ranging from 4 to 12 months and, despite intense therapeutical efforts, it is invariably fatal. Mesothelioma cells are unique in the sense that they possess a biphasic growth potential and can be stimulated by serum growth factors to differentiate into stable epithelial or fibrous phenotypes. The prognosis of this tumor varies greatly depending on the differences in growth pattern, the most important predictor of poor prognosis being a fibrous phenotype. To study the molecular basis of mesothelial differentiation, we used benign and malignant mesothelial cells in various stages of phenotypic differentiation. In order to evaluate the impact of proteoglycans (PG) on this process, a series of PGs were analyzed by semiquantitative reversed transcriptase polymerase chain reaction. The cells with epithelial phenotype showed increased expression of syndecan-2, syndecan-4 and hyaluronan synthase, and fibroblast-like cells expressed more matrix PGs: versican, decorin and biglycan. The PG profile may serve as a "fingerprint", and reflect the maturation of mesothelial cells. The functional importance of syndecans in mesothelial differentiation was further shown by antisense targeting; down-regulation of each particular syndecan caused a loss of epithelial morphology. Syndecans-1 and-4 are also needed for cell adhesion. The differentiation of mesothelioma cells was influenced by treatment with various growth factors (TGF-b2, EGF, FGF-2, IGF-I and PDGF-BB). These factors affected the proliferation and morphology of mesothelioma cells to various extents, and the PG profile changed, in parallel, with an induced epithelial-mesenchymal transition. Exposure to EGF and IGF-I caused a fibroblast-like morphology simultaneously with a reduction in the syndecan expression levels. At the same time, the levels of shed syndecan-1 increased in the culture medium. The involvement of other regulatory molecules in mesothelioma differentiation was assessed by subtractive hybridization, which has revealed a limited number of genes being differentially expressed between cells of epithelial or fibrous phenotypes. Most of these genes were recovered from the epithelial cells, which may indicate a more mature phenotype. The expression level of thioredoxin reductase, a small redox-active protein involved in drug resistance, was extremely high in both cell sub-lines, and may reflect the generic insensitivity of mesotheliomas to chemotherapy. Although syndecans play a major role in regulating cell morphology, little is known about their subcellular distribution. Using confocal laser microscopy we found a substantial proportion of syndecans at intracellular locations, and syndecan-1 accumulated in the nucleus in a time-dependent manner. There was a close spatial relation of syndecans to tubulin in both interphase and mitotic cells. Vinblastine treatment interfered with the nuclear transport, and syndecan-1 and tubulin copolymerize in paracrystalline occlusion bodies, in parallel with impaired nuclear transport. These findings suggest a tubulin-mediated transport mechanism. TGF-b2 reduced the proliferation rate of mesothelioma cells, concomitantly with a delay in nuclear transport of syndecan-1. These data show that all syndecans are involved in maintaining the epithelial morphology, and that various amounts and translocation of syndecans may participate in molecular switches that regulate cell differentiation and proliferation. The above mechanisms may represent crucial steps, and possible future targets for therapy, that can be used to improve the management of patients with malignant mesothelioma.

In Vivo, 2009

Fibrosarcoma is an uncommon soft tissue tumor with a complex cell microenvironment, particularly ... more Fibrosarcoma is an uncommon soft tissue tumor with a complex cell microenvironment, particularly rich in glycosaminoglycans/proteoglycans (GAGs/PGs). Chondroitin sulfate proteoglycans (CSPGs) participate in the modulation of various cellular functions, including adhesion and migration. The role of chondroitin sulfate (CS) chains on adhesion, chemotaxis and migration of poorly differentiated fibrosarcoma B6FS cell was studied, utilizing exogenous CS treatment and chondroitinase digestions as well as specific modulators of CS synthesis. Cleavage of cell-associated CS chains and specific inhibition of endogenous CS production severely impaired these fibrosarcoma cell functions. These results show that the reduction of endogenous CSPG expression as well as cleavage of the CS chain inhibited fibrosarcoma cell motility, migration and adhesion. Treatment with free CS chains enhanced cell chemotaxis and migration, whereas adhesion was inhibited. CS chains were found to upregulate cell motility through the MAPK pathway, specifically through JNK, whereas CS-induced migration was found to require tyrosine kinase dependent pathways. This study suggests a new role of CS on tumor cell adhesion, chemotaxis and migration.

Anticancer Research, 2004

Background: Hyaluronan is one of the main components of the extracellular matrix. It is synthesiz... more Background: Hyaluronan is one of the main components of the extracellular matrix. It is synthesized at the cell plasma membrane by specific hyaluronan synthases (HAS). Although a large number of studies have described hyaluronan in pleural effusion from malignant mesothelioma, the source of hyaluronan in malignant mesothelioma has been subject to controversy. Materials and Methods: The mRNA expression of all three HAS in malignant mesothelioma cells was studied using RT-PCR. The hyaluronan production in culture medium of malignant mesothelioma cells was also examined using highperformance liquid chromatography (HPLC). Results: We found that 9/10 malignant mesothelioma cell lines and one primary culture of malignant mesothelioma cells expressed HAS-1, while 10/10 malignant mesothelioma cell lines and one primary culture of malignant mesothelioma cells expressed HAS-2 and HAS-3. In addition, we demonstrated hyaluronan in the culture medium of 6 out of 10 malignant mesothelioma cell lines and one primary culture of malignant mesothelioma cells. Conclusion: Our results show that malignant mesothelioma cells express all three HAS and synthesize hyaluronan. The expression of HAS isoforms and hyaluronan in malignant mesothelioma cells in cultures and previous observations by other investigators indicate that these cells are, at least in part, responsible for hyaluronan synthesis in vivo. Materials and Methods Reagents. RPMI 1640, FCS, penicillin, streptomycin and Lglutamine were obtained from Life Technologies (Paisley, UK). Cells and culture conditions. Primary culture of malignant mesothelioma cells was obtained from pleural fluid of a patient with malignant mesothelioma. Ten human malignant mesothelioma cell lines were used:

Experimental Cell Research, 2002

Syndecans are transmembrane proteoglycans implicated in the regulation of cell growth and differe... more Syndecans are transmembrane proteoglycans implicated in the regulation of cell growth and differentiation, by interacting with growth factors. Although syndecans play a major role in regulating cell morphology, little is known about their subcellular distribution and in vivo association with the cytoskeleton. To address this question, we investigated the subcellular distribution and dynamic rearrangement of syndecans-1,-2, and-4, using confocal laser microscopy. Furthermore, we monitored the spatial relation of syndecans to tubulin in both mitotic and interphase cells. Initially, the reactivity to syndecans was confined to the cytoplasm, staining of the cell membranes appearing later. Syndecan-1 also seems to translocate to the nucleus in a time-dependent manner. The mitotic spindle shows unexpectedly more syndecans than that found in interphase cells. After vinblastine treatment, both syndecan-1 and tubulin were recovered as paracrystalline occlusion bodies, and the nuclear reactivity to syndecan-1 disappeared, suggesting tubulin-mediated nuclear transport of this proteoglycan. Plasma membrane staining reappeared in the postmitotic cells. Nuclear translocation predominantly affected syndecan-1, whereas syndecan-2 and-4 remained in cytoplasm and cell membrane. This is the first report on regulated nuclear translocation and the presence of syndecan-1 in the mitotic spindle, where it may stabilize the mitotic machinery. The syndecan-1/ tubulin complex may also act as a vehicle for the transport of protein growth factors to the cell nucleus.

Differentiation, 2000

Malignant mesothelioma is a tumour originating from mesothelial cells, and it exhibits epithelial... more Malignant mesothelioma is a tumour originating from mesothelial cells, and it exhibits epithelial, fibrous, or biphasic differentiation. This tumour is highly resistant to therapy, and presence of a sarcomatous growth pattern has been associated with worse prognosis. A mesothelioma cell line with retained ability to differentiate into either epithelial or fibroblast-like phenotype was subjected to subtractive hybridisation in order to identify the genes coupled to tumour cell differentiation. Nine genes were found to be selectively overexpressed in the epithelial sub-line, compared to only two genes in the fibroblast-like phenotype. This may support the idea that the sarcomatous phenotype represents a less differentiated tumour. One of the genes that is differentially expressed by the epithelial cells was thioredoxin, a small redox-active protein associated with cell-growth and differentiation. This overexpression was accompanied by increased protein levels both intracellularly and in the medium. Thioredoxin is reduced by the selenoprotein thioredoxin reductase and NADPH. The activity of this enzyme was high in both cell sub-lines but induced 2-fold in the epithelially-differentiated cells. Overexpression of thioredoxin might be a factor behind the poor prognosis and reduced responsiveness to therapy of mesotheliomas. Epithelial differentiation in this cell line has previously been linked to increased synthesis of heparan sulphate proteoglycans. The possible formation of complexes including thioredoxin, thioredoxin re

Biomolecules

Extracellular vesicles (EVs), comprising exosomes, microvesicles, and apoptotic bodies, are relea... more Extracellular vesicles (EVs), comprising exosomes, microvesicles, and apoptotic bodies, are released by all cells into the extracellular matrix and body fluids, where they play important roles in intercellular communication and matrix remodeling in various pathological conditions. Malignant pleural mesothelioma (MPM) is a primary tumor of mesothelial origin, predominantly related to asbestos exposure. The detection of MPM at an early stage and distinguishing it from benign conditions and metastatic adenocarcinomas (AD) is sometimes challenging. Pleural effusion is often the first available biological material and an ideal source for characterizing diagnostic and prognostic factors. Specific proteins have previously been identified as diagnostic markers in effusion, but it is not currently known whether these are associated with vesicles or released in soluble form. Here, we study and characterize tumor heterogeneity and extracellular vesicle diversity in pleural effusion as diagnost...

Biomolecules

Malignant pleural mesothelioma (MPM) is a highly aggressive and therapy resistant pleural maligna... more Malignant pleural mesothelioma (MPM) is a highly aggressive and therapy resistant pleural malignancy that is caused by asbestos exposure. MPM is associated with poor prognosis and a short patient survival. The survival time is strongly influenced by the subtype of the tumor. Dyspnea and accumulation of pleural effusion in the pleural cavity are common symptoms of MPM. The diagnostic distinction from other malignancies and reactive conditions is done using histopathology or cytopathology, always supported by immunohistochemistry, and sometimes also by analyses of soluble biomarkers in effusion supernatant. We evaluated the soluble angiogenesis related molecules as possible prognostic and diagnostic biomarkers for MPM by Luminex multiplex assay. Pleural effusion from 42 patients with malignant pleural mesothelioma (MPM), 36 patients with adenocarcinoma (AD) and 40 benign (BE) effusions were analyzed for 10 different analytes that, in previous studies, were associated with angiogenesis...

Journal of Clinical Pathology

AimsAccurate and reliable diagnosis is essential for lung cancer treatment. The study aim was to ... more AimsAccurate and reliable diagnosis is essential for lung cancer treatment. The study aim was to investigate interpathologist diagnostic concordance for pulmonary tumours according to WHO diagnostic criteria.MethodsFifty-two unselected lung and bronchial biopsies were diagnosed by a thoracic pathologist based on a broad spectrum of immunohistochemical (IHC) stainings, molecular data and clinical/radiological information. Slides stained with H&E, thyroid transcription factor-1 (TTF-1) clone SPT24 and p40 were scanned and provided digitally to 20 pathologists unaware of reference diagnoses. The pathologists independently diagnosed the cases and stated if further diagnostic markers were deemed necessary.ResultsIn 31 (60%) of the cases, ≥80% of the pathologists agreed with each other and with the reference diagnosis. Lower agreement was seen in non-small cell neuroendocrine tumours and in squamous cell carcinoma with diffuse TTF-1 positivity. Agreement with the reference diagnosis range...

Cancers

Malignant mesothelioma (MM) is a rare but highly aggressive cancer that primarily originates from... more Malignant mesothelioma (MM) is a rare but highly aggressive cancer that primarily originates from the pleura, peritoneum or pericardium. There is a well-established link between asbestos exposure and progression of MM. Direct invasion of the surrounding tissues is the main feature of MM, which is dependent on dysregulated communication between the mesothelium and the microenvironment. This communication is dependent on the dynamic organization of the cytoskeleton. We have analyzed the organization and function of key cytoskeletal components in MM cell lines of increasing malignancies measured as migratory and invasive properties, and we show that highly malignant and invasive MM cells have an organization of the actin filament and vimentin systems that is distinct from the less malignant MM cell lines. In addition, the Hippo tumor suppressor pathway was inactivated in the invasive MM cells, which was seen as increased YAP nuclear localization.

Cancers

Malignant mesothelioma (MM) is an aggressive tumor of the serosal cavities. Angiogenesis is impor... more Malignant mesothelioma (MM) is an aggressive tumor of the serosal cavities. Angiogenesis is important for mesothelioma progression, but so far, anti-angiogenic agents have not improved patient survival. Our hypothesis is that better understanding of the regulation of angiogenesis in this tumor would largely improve the success of such a therapy. Syndecan-1 (SDC-1) is a transmembrane heparan sulfate proteoglycan that acts as a co-receptor in various cellular processes including angiogenesis. In MM, the expression of SDC-1 is generally low but when present, SDC-1 associates to epithelioid differentiation, inhibition of tumor cell migration and favorable prognosis, meanwhile SDC-1 decrease deteriorates the prognosis. In the present study, we studied the effect of SDC-1 overexpression and silencing on MM cells ability to secrete angiogenic factors and monitored the downstream effect of SDC-1 modulation on endothelial cells proliferation, wound healing, and tube formation. This was done ...

Translational Lung Cancer Research

The global incidence of malignant mesothelioma (MM) causes considerable disease burden, suffering... more The global incidence of malignant mesothelioma (MM) causes considerable disease burden, suffering and health care costs. Beside preventive measures and ban the use of asbestos, early diagnosis would largely improve the chance of curative treatment. Current histologic criteria, however, requiring presence of invasion in the surrounding fatty tissue fail to identify MM in sufficiently early stage. Unilateral accumulation of pleural effusion is one of the earliest clinical manifestations of MM that occurs in approximately 90% of the patients. Therapeutic thoracocenthesis is necessary to remove the fluid and to relieve patients' symptoms. This effusion is easily accessible and offers early and minimally invasive diagnosis by combining cytology with immunologic, molecular-and biomarker analyses. Typically, the fluid is rich in malignant cells and cell groups, but incipient stages of the disease may be difficult to recognize as the malignant cells can be masked by presence of inflammatory or reactive mesothelial cells. Recurrent, hemorrhagic and cell rich effusion should always be suspicious for MM and adequately prepared and analyzed to provide necessary information for subsequent therapy. Importantly, early detection of MM by integrating cytology and molecular approaches has high sensitivity and positive predictive value and has a major impact on patient survival. Thus, a conclusive positive MM cytology should lead to treatment without delay. This review summarizes molecular and diagnostic criteria of MM diagnosis.

Cytopathology

Serous effusion fluid is one of the most commonly encountered specimens in routine cytopathology ... more Serous effusion fluid is one of the most commonly encountered specimens in routine cytopathology practice. It provides invaluable information about the patient and the clinical status; but to get the most of it, specimen handling and processing must be carried out properly. Cytomorphology is the basis of a successful analysis which should complemented by ancillary tests when needed. A wide spectrum of ancillary techniques - ranging from immunocytochemistry and flow cytometry to different assays of molecular pathology - can be applied to serous effusions. This article describes the acquisition and management of serous effusion fluids, methods for preservation and transportation, different techniques of cytopreparation, application of immunocytochemistry, flow cytometry, and fluorescence in-situ hybridization (FISH), as well as DNA extraction for polymerase chain reaction (PCR) and next generation sequencing (NGS). Principles of bio-banking of effusion samples are also discussed which is getting more important in correlation with the developments in personalized medicine. This article is protected by copyright. All rights reserved.

Cancer Cytopathology

BACKGROUND: Malignant mesothelioma (MM) is an aggressive, fatal tumor. Current therapeutic option... more BACKGROUND: Malignant mesothelioma (MM) is an aggressive, fatal tumor. Current therapeutic options only marginally improve survival. Programmed cell death ligand 1 (PD-L1) is a dominant mediator of immunosuppression, binding to programmed cell death 1 (PD-1). PD-L1 is up-regulated in cancer cells, and the PD-1/PD-L1 pathway plays a critical role in tumor immune evasion, thus providing a target for antitumor therapy. Further, a correlation between PD-L1 expression and prognosis has been reported. Studies performed on histological material have revealed expression of PD-L1 in MM, but no study has been performed on MM effusions thus far. METHODS: PD-L1 expression was determined by a commercially available antibody (clone 28-8) in 74 formalin-fixed, paraffin-embedded cell blocks from body effusions obtained at diagnosis from patients with MM. The presence of MM cells was confirmed with CK5/6, calretinin, and EMA and the admixture of macrophages was assessed with CD68. Only cases containing more than 100 tumor cells were assessed. Membranous staining in tumor cells was considered positive. Survival time was calculated from the appearance of the first malignant effusion until death. RESULTS: Reactivity was observed in 23 of 61 (38%) of cases and was classified as 1%-5% (n 5 9 cases), >5%-10% (n 5 4 cases), >10%-50% (n 5 4 cases), and >50% (n 5 6 cases) positive cells. Survival times did not differ significantly between patients with PD-L1-positive and PD-L1-negative tumors. CONCLUSION: MM effusions are suitable for immune-cytochemical assessment of PD-L1 expression in malignant cells and the results are similar to those reported for histological specimens.

Matrix Biology, 2016

Synthesis, deposition, and interactions of hyaluronan (HA) with its cellular receptor CD44 are cr... more Synthesis, deposition, and interactions of hyaluronan (HA) with its cellular receptor CD44 are crucial events that regulate the onset and progression of tumors. The intracellular signaling pathways initiated by HA interactions with CD44 leading to tumorigenic responses are complex. HA molecules may perform dual functions depending on their concentration and size. Overexpression of variant isoforms of CD44 (CD44v) is most commonly linked to cancer progression, whereas their loss is associated with inhibition of tumor growth. In this review, we highlight that regulation of HA synthases (HASes) by post-translational modifications such as O-GlcNAcylation and ubiquitination, or by environmental factors or the action of microRNAs is important for HA synthesis and secretion in the tumor microenvironment. Moreover, we focus on the roles and interactions of CD44 with various proteins that reside extra-and intracellularly, as well as on cellular membranes with particular reference to the CD44-HA axis in cancer stem cell functions, and the importance of CD44/CD44v6 targeting to inhibit tumorigenesis in colon cancer cell models.

Journal of translational medicine, Oct 19, 2016

The goal of biomarker research is to identify clinically valid markers. Despite decades of resear... more The goal of biomarker research is to identify clinically valid markers. Despite decades of research there has been disappointingly few molecules or techniques that are in use today. The "1st International NTNU Symposium on Current and Future Clinical Biomarkers of Cancer: Innovation and Implementation", was held June 16th and 17th 2016, at the Knowledge Center of the St. Olavs Hospital in Trondheim, Norway, under the auspices of the Norwegian University of Science and Technology (NTNU) and the HUNT biobank and research center. The Symposium attracted approximately 100 attendees and invited speakers from 12 countries and 4 continents. In this Symposium original research and overviews on diagnostic, predictive and prognostic cancer biomarkers in serum, plasma, urine, pleural fluid and tumor, circulating tumor cells and bioinformatics as well as how to implement biomarkers in clinical trials were presented. Senior researchers and young investigators presented, reviewed and vi...

Malignant mesothelioma is a highly aggressive tumor with median survival ranging from 4 to 12 mon... more Malignant mesothelioma is a highly aggressive tumor with median survival ranging from 4 to 12 months and, despite intense therapeutical efforts, it is invariably fatal. Mesothelioma cells are unique in the sense that they possess a biphasic growth potential and can be stimulated by serum growth factors to differentiate into stable epithelial or fibrous phenotypes. The prognosis of this tumor varies greatly depending on the differences in growth pattern, the most important predictor of poor prognosis being a fibrous phenotype. To study the molecular basis of mesothelial differentiation, we used benign and malignant mesothelial cells in various stages of phenotypic differentiation. In order to evaluate the impact of proteoglycans (PG) on this process, a series of PGs were analyzed by semiquantitative reversed transcriptase polymerase chain reaction. The cells with epithelial phenotype showed increased expression of syndecan-2, syndecan-4 and hyaluronan synthase, and fibroblast-like cells expressed more matrix PGs: versican, decorin and biglycan. The PG profile may serve as a "fingerprint", and reflect the maturation of mesothelial cells. The functional importance of syndecans in mesothelial differentiation was further shown by antisense targeting; down-regulation of each particular syndecan caused a loss of epithelial morphology. Syndecans-1 and-4 are also needed for cell adhesion. The differentiation of mesothelioma cells was influenced by treatment with various growth factors (TGF-b2, EGF, FGF-2, IGF-I and PDGF-BB). These factors affected the proliferation and morphology of mesothelioma cells to various extents, and the PG profile changed, in parallel, with an induced epithelial-mesenchymal transition. Exposure to EGF and IGF-I caused a fibroblast-like morphology simultaneously with a reduction in the syndecan expression levels. At the same time, the levels of shed syndecan-1 increased in the culture medium. The involvement of other regulatory molecules in mesothelioma differentiation was assessed by subtractive hybridization, which has revealed a limited number of genes being differentially expressed between cells of epithelial or fibrous phenotypes. Most of these genes were recovered from the epithelial cells, which may indicate a more mature phenotype. The expression level of thioredoxin reductase, a small redox-active protein involved in drug resistance, was extremely high in both cell sub-lines, and may reflect the generic insensitivity of mesotheliomas to chemotherapy. Although syndecans play a major role in regulating cell morphology, little is known about their subcellular distribution. Using confocal laser microscopy we found a substantial proportion of syndecans at intracellular locations, and syndecan-1 accumulated in the nucleus in a time-dependent manner. There was a close spatial relation of syndecans to tubulin in both interphase and mitotic cells. Vinblastine treatment interfered with the nuclear transport, and syndecan-1 and tubulin copolymerize in paracrystalline occlusion bodies, in parallel with impaired nuclear transport. These findings suggest a tubulin-mediated transport mechanism. TGF-b2 reduced the proliferation rate of mesothelioma cells, concomitantly with a delay in nuclear transport of syndecan-1. These data show that all syndecans are involved in maintaining the epithelial morphology, and that various amounts and translocation of syndecans may participate in molecular switches that regulate cell differentiation and proliferation. The above mechanisms may represent crucial steps, and possible future targets for therapy, that can be used to improve the management of patients with malignant mesothelioma.

In Vivo, 2009

Fibrosarcoma is an uncommon soft tissue tumor with a complex cell microenvironment, particularly ... more Fibrosarcoma is an uncommon soft tissue tumor with a complex cell microenvironment, particularly rich in glycosaminoglycans/proteoglycans (GAGs/PGs). Chondroitin sulfate proteoglycans (CSPGs) participate in the modulation of various cellular functions, including adhesion and migration. The role of chondroitin sulfate (CS) chains on adhesion, chemotaxis and migration of poorly differentiated fibrosarcoma B6FS cell was studied, utilizing exogenous CS treatment and chondroitinase digestions as well as specific modulators of CS synthesis. Cleavage of cell-associated CS chains and specific inhibition of endogenous CS production severely impaired these fibrosarcoma cell functions. These results show that the reduction of endogenous CSPG expression as well as cleavage of the CS chain inhibited fibrosarcoma cell motility, migration and adhesion. Treatment with free CS chains enhanced cell chemotaxis and migration, whereas adhesion was inhibited. CS chains were found to upregulate cell motility through the MAPK pathway, specifically through JNK, whereas CS-induced migration was found to require tyrosine kinase dependent pathways. This study suggests a new role of CS on tumor cell adhesion, chemotaxis and migration.

Anticancer Research, 2004

Background: Hyaluronan is one of the main components of the extracellular matrix. It is synthesiz... more Background: Hyaluronan is one of the main components of the extracellular matrix. It is synthesized at the cell plasma membrane by specific hyaluronan synthases (HAS). Although a large number of studies have described hyaluronan in pleural effusion from malignant mesothelioma, the source of hyaluronan in malignant mesothelioma has been subject to controversy. Materials and Methods: The mRNA expression of all three HAS in malignant mesothelioma cells was studied using RT-PCR. The hyaluronan production in culture medium of malignant mesothelioma cells was also examined using highperformance liquid chromatography (HPLC). Results: We found that 9/10 malignant mesothelioma cell lines and one primary culture of malignant mesothelioma cells expressed HAS-1, while 10/10 malignant mesothelioma cell lines and one primary culture of malignant mesothelioma cells expressed HAS-2 and HAS-3. In addition, we demonstrated hyaluronan in the culture medium of 6 out of 10 malignant mesothelioma cell lines and one primary culture of malignant mesothelioma cells. Conclusion: Our results show that malignant mesothelioma cells express all three HAS and synthesize hyaluronan. The expression of HAS isoforms and hyaluronan in malignant mesothelioma cells in cultures and previous observations by other investigators indicate that these cells are, at least in part, responsible for hyaluronan synthesis in vivo. Materials and Methods Reagents. RPMI 1640, FCS, penicillin, streptomycin and Lglutamine were obtained from Life Technologies (Paisley, UK). Cells and culture conditions. Primary culture of malignant mesothelioma cells was obtained from pleural fluid of a patient with malignant mesothelioma. Ten human malignant mesothelioma cell lines were used: