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Background and Aims: Dietary fat intake is associated with increased risk of colorectal cancer (CRC). We examined the role of high-fat diet (HFD) in driving CRC through modulating gut microbiota and metabolites., Methods: HFD or control diet was fed to mice littermates in CRC mouse models of an azoxymethane (AOM) model and Apc min/+ model, with or without antibiotics cocktail treatment. Germ-free mice for fecal microbiota transplantation were used for validation. Gut microbiota and metabolites were detected using metagenomic sequencing and high-performance liquid chromatography-mass spectrometry, respectively. Gut barrier function was determined using lipopolysaccharides level and transmission electron microscopy., Results: HFD promoted colorectal tumorigenesis in both AOM-treated mice and Apc min/+ mice compared with control diet-fed mice. Gut microbiota depletion using antibiotics attenuated colon tumor formation in HFD-fed mice. A significant shift of gut microbiota composition with increased pathogenic bacteria Alistipessp.Marseille-P5997 and Alistipessp.5CPEGH6, and depleted probiotic Parabacteroides distasonis, along with impaired gut barrier function was exhibited in HFD-fed mice. Moreover, HFD-modulated gut microbiota promotes colorectal tumorigenesis in AOM-treated germ-free mice, indicating gut microbiota was essential in HFD-associated colorectal tumorigenesis. Gut metabolites alteration, including elevated lysophosphatidic acid, which was confirmed to promote CRC cell proliferation and impair cell junction, was also observed in HFD-fed mice. Moreover, transfer of stools from HFD-fed mice to germ-free mice without interference increased colonic cell proliferation, impaired gut barrier function, and induced oncogenic genes expression., Conclusions: HFD drives colorectal tumorigenesis through inducing gut microbial dysbiosis, metabolomic dysregulation with elevated lysophosphatidic acid, and gut barrier dysfunction in mice., (Copyright © 2022 The Authors. Published by Elsevier Inc. All rights reserved.)

Biomechanical cues within tissue microenvironments are critical for maintaining homeostasis, and their disruption can contribute to malignant transformation and metastasis. Once transformed, metastatic cancer cells can migrate persistently by adapting (plasticity) to changes in the local fibrous extracellular matrix, and current strategies to recapitulate persistent migration rely exclusively on the use of aligned geometries. Here, the controlled interfiber spacing in suspended crosshatch networks of nanofibers induces cells to exhibit plasticity in migratory behavior (persistent and random) and the associated cytoskeletal arrangement. At dense spacing (3 and 6 µm), unexpectedly, elongated cells migrate persistently (in 1 dimension) at high speeds in 3-dimensional shapes with thick nuclei, and short focal adhesion cluster (FAC) lengths. With increased spacing (18 and 36 µm), cells attain 2-dimensional morphologies, have flattened nuclei and longer FACs, and migrate randomly by rapidly detaching their trailing edges that strain the nuclei by ∼35%. At 54-µm spacing, kite-shaped cells become near stationary. Poorly developed filamentous actin stress fibers are found only in cells on 3-µm networks. Gene-expression profiling shows a decrease in transcriptional potential and a differential up-regulation of metabolic pathways. The consistency in observed phenotypes across cell lines supports using this platform to dissect hallmarks of plasticity in migration in vitro .-Jana, A., Nookaew, I., Singh, J., Behkam, B., Franco, A. T., Nain, A. S. Crosshatch nanofiber networks of tunable interfiber spacing induce plasticity in cell migration and cytoskeletal response.

Nuclear alterations are a hallmark of many types of cancers, including prostate cancer (PCa). Recent evidence shows that subvisual changes, ones that may not be visually perceptible to a pathologist, to the nucleus and its ultrastructural components can precede visual histopathological recognition of cancer. Alterations to nuclear features, such as nuclear size and shape, texture, and spatial architecture, reflect the complex molecular-level changes that occur during oncogenesis. Quantitative nuclear morphometry, a field that uses computational approaches to identify and quantify malignancy-induced nuclear changes, can enable a detailed and objective analysis of the PCa cell nucleus. Recent advances in machine learning-based approaches can now automatically mine data related to these changes to aid in the diagnosis, decision making, and prediction of PCa prognoses. In this review, we use PCa as a case study to connect the molecular-level mechanisms that underlie these nuclear changes to the machine learning computational approaches, bridging the gap between the clinical and computational understanding of PCa. First, we will discuss recent developments to our understanding of the molecular events that drive nuclear alterations in the context of PCa: the role of the nuclear matrix and lamina in size and shape changes, the role of 3-dimensional chromatin organization and epigenetic modifications in textural changes, and the role of the tumor microenvironment in altering nuclear spatial topology. We will then discuss the advances in the applications of machine learning algorithms to automatically segment nuclei in prostate histopathological images, extract nuclear features to aid in diagnostic decision making, and predict potential outcomes, such as biochemical recurrence and survival. Finally, we will discuss the challenges and opportunities associated with translation of the quantitative nuclear morphometry methodology into the clinical space. Ultimately, accurate identification and quantification of nuclear alterations can contribute to the field of nucleomics and has applications for computationally driven precision oncologic patient care., (© 2018 Wiley Periodicals, Inc.)

Early detection of hepatocellular carcinoma (HCC) is crucial for successful therapy. The present work examined the value of ultrastructural morphometric image analysis of hepatocyte nuclei in patients with chronic hepatitis C virus (HCV) versus HCC cases with chronic HCV and the corresponding surgical tumor-free safe margins (TFMs), to highlight any early predictive signs of neoplastic cellular transformation. This work also performed an immunohistochemical assessment of cytokeratin 19 (CK19) and Ki-67-positive cells to visualize any associated proliferative activity in the examined groups. The results showed significant decrease in the hepatocyte nuclear surface areas in the HCC and TFMs versus those in the HCV cases. The hepatocyte nucleolar surface area was significantly increased in the HCC cases versus that in the HCV cases. This increase was associated with a significant increase in Ki-67-positive cells in the HCC cases compared to those in the other groups. Conversely, the mean number of CK 19-positive cells was significantly reduced in the HCC cases compared to the cell numbers in TFMs and HCV cases with severe hepatic fibrosis. Liver progenitor cells (LPCs) were discerned in the reactive ductules and canaliculo-ductular junctions that characterized TFMs. LPCs were sporadically distributed in the liver lobules and reactive bile ductules in the HCC samples. In conclusion, CK 19 represents an important marker for distinguishing between dysplastic and malignant liver nodules. Electron microscopic morphometric image analysis may be considered as adjunct factor for assessing hepatocyte malignant transformation. Wider scale studies are needed to authenticate these results.

Bioassays of planarian neoplasia highlight the potential of these organisms as useful standards to assess whether environmental toxins such as cadmium promote tumorigenesis. These studies complement other investigations into the exceptional healing and regeneration of planarians - processes that are driven by a population of active stem cells, or neoblasts, which are likely transformed during planarian tumor growth. Our goal was to determine if planarian tumorigenesis assays are amenable to mechanistic studies of cadmium carcinogenesis. To that end we demonstrate, by examining both counts of cell populations by size, and instances of mitosis, that the activity of the stem cell population can be monitored. We also provide evidence that specific biomodulators can affect the potential of planarian neoplastic growth, in that an inhibitor of metalloproteinases effectively blocked the development of the lesions. From these results, we infer that neoblast activity does respond to cadmium-induced tumor growth, and that metalloproteinases are required for the progression of cancer in the planarian., (Copyright © 2017 Elsevier Inc. All rights reserved.)

Aberrant changes in histone post-translational modifications are encountered frequently in diseases like cancer. Although histone H3 post-translational modifications have been extensively studied in context of diseases, the functionally important histone H2A PTM H2A119ub (H2Aub) has not gained much attention. In this study, we report that H2Aub markedly decreases in hepatocellular carcinoma. Usp21, a H2A deubiquitinase, is probably responsible for decrease in H2Aub. In addition, the H2Aub levels showed an inverse correlation with H3S10 phosphorylation (H3S10p) and the proliferative state of the cells. Downregulation of H2Aub is also associated with increased expression of growth factor gene lipocalin 2. Interestingly, we show that treatment of cells with histone deacetylase inhibitor trichostatin A results in increase of H2Aub and decrease in H3S10p. Our work for the first time suggests the in vivo association of H3S10p, H4ac, and H2A119ub with cellular transformation., (© 2016 by the Society for Experimental Biology and Medicine.)

Objective: Problems in pre-cancer diagnosis complicate cancer theragnosis as well as life expectancy. There is uncertainty regarding malignant transformation of oral submucous fibrosis (OSF), an oral pre-cancer with dysplastic (OSFWD) and non-dysplastic (OSFWT) subtypes. Understanding the structural, molecular and physical aspects of epithelial homeostasis may be useful., Materials and Methods: Histopathological grading of biopsy sections was performed using H&E staining. Alterations in epithelial surface architecture in different groups was evaluated using scanning electron microscopy (SEM). The expression of crucial epithelial genes (p63, CK-5/6, CK-10, E-cadherin and β-catenin) was studied by immunohistochemistry, Western blot and RT-PCR analysis., Results: SEM observations revealed that the surface epithelial ridge pattern became thick and dense, and pit pattern gradually decreased in OSFWD and oral squamous cell carcinoma (OSCC). p63, ΔNp63 and CK-5/6 were up-regulated in OSFWD and OSCC but down-regulated in OSFWT. CK-10 was down-regulated in OSFWD compared to OSFWT. Cytoplasmic expression of E-cadherin and β-catenin was elevated in dysplastic and cancerous conditions. Moreover, statistical correlation between SEM features (ridges and pits) and molecular attributes demonstrated a significant positive relationship between the ridge-to-pit ratio and p63 population density (r=0.85) and the ridge-to-pit ratio and CK-5/6 intensity (r=0.63)., Conclusions: Molecular changes related to epithelial progressive maturation and cellular proliferation are correlated with concomitant alteration of epithelial surface architecture which helps to predict the malignant potentiality of OSF., (Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://group.bmj.com/group/rights-licensing/permissions.)

Background: Nuclear alterations are a well-known manifestation of cancer. However, little is known about the early, microscopically-undetectable stages of malignant transformation. Based on the phenomenon of field cancerization, the tissue in the field of a tumor can be used to identify and study the initiating events of carcinogenesis. Morphological changes in nuclear organization have been implicated in the field of colorectal cancer (CRC), and we hypothesize that characterization of chromatin alterations in the early stages of CRC will provide insight into cancer progression, as well as serve as a biomarker for early detection, risk stratification and prevention., Methods: For this study we used transmission electron microscopy (TEM) images of nuclei harboring pre-neoplastic CRC alterations in two models: a carcinogen-treated animal model of early CRC, and microscopically normal-appearing tissue in the field of human CRC. We quantify the chromatin arrangement using approaches with two levels of complexity: 1) binary, where chromatin is separated into areas of dense heterochromatin and loose euchromatin, and 2) grey-scale, where the statistics of continuous mass-density distribution within the nucleus is quantified by its spatial correlation function., Results: We established an increase in heterochromatin content and clump size, as well as a loss of its characteristic peripheral positioning in microscopically normal pre-neoplastic cell nuclei. Additionally, the analysis of chromatin density showed that its spatial distribution is altered from a fractal to a stretched exponential., Conclusions: We characterize quantitatively and qualitatively the nanoscale structural alterations preceding cancer development, which may allow for the establishment of promising new biomarkers for cancer risk stratification and diagnosis. The findings of this study confirm that ultrastructural changes of chromatin in field carcinogenesis represent early neoplastic events leading to the development of well-documented, microscopically detectable hallmarks of cancer.

Hepatic oval cells are considered as facultative progenitor/stem cells of liver and able to differentiate into either hepatocytes or biliary epithelial cells. The transformed oval cells by carcinogen possess potential to develop carcinomas in animal models. In order to better understand the molecular mechanism in carcinogenetic process, we used a proteomic approach to assess the early changes in protein expression of oval cells (OC3W3-15) initiated by methylnitronitrosoguanidine (MNNG). Meanwhile, we compared cell biologic characteristics of the MNNG treated OC3W3-15 cells and control oval cells by electron microscopy, flow cytometry, karyotype and soft agar assay. The mRNA levels of GGT and GSTP1 determined by real-time PCR were also detected in both cell lines. Our results showed that MNNG-treated OC3W3-15 cells exhibited characteristics of malignant transformation, including growth rate, chromosomal aberrations, abnormal DNA content, and the ability to form colonies. The cells expressed higher levels of the tumor marker AFP, GGT and GSTP1 mRNA than that of control cells. Significant changes of several proteins involved in the malignant transformation process, including cell cycle related proteins, proteins involved in organism development and cell differentiation, are found in OC3W3-15 cells. The proteins may provide early affection in malignant transformation of hepatic oval cells, and yield further insight into mechanism of carcinogenesis of hepatocellular carcinoma., (Copyright © 2013 Elsevier GmbH. All rights reserved.)

The classical sacrococcygeal chordoma tumor presents with a typical morphology of lobulated myxoid tumor tissue with cords, strands and nests of tumor cells. The population of cells consists of small non-vacuolated cells, intermediate cells with a wide range of vacuolization and large heavily vacuolated (physaliferous) cells. To date analysis was only performed on bulk tumor mass because of its rare incidence, lack of suited model systems and technical limitations thereby neglecting its heterogeneous composition. We intended to clarify whether the observed cell types are derived from genetically distinct clones or represent different phenotypes. Furthermore, we aimed at elucidating the differences between small non-vacuolated and large physaliferous cells on the genomic and transcriptomic level. Phenotype-specific analyses of small non-vacuolated and large physaliferous cells in two independent chordoma cell lines yielded four candidate genes involved in chordoma cell development. UCHL3, coding for an ubiquitin hydrolase, was found to be over-expressed in the large physaliferous cell phenotype of MUG-Chor1 (18.7-fold) and U-CH1 (3.7-fold) cells. The mannosyltransferase ALG11 (695-fold) and the phosphatase subunit PPP2CB (18.6-fold) were found to be up-regulated in large physaliferous MUG-Chor1 cells showing a similar trend in U-CH1 cells. TMEM144, an orphan 10-transmembrane family receptor, yielded contradictory data as cDNA microarray analysis showed up- but RT-qPCR data down-regulation in large physaliferous MUG-Chor1 cells. Isolation of few but morphologically identical cells allowed us to overcome the limitations of bulk analysis in chordoma research. We identified the different chordoma cell phenotypes to be part of a developmental process and discovered new genes linked to chordoma cell development representing potential targets for further research in chordoma tumor biology.

Background and Study Aims: Esophageal adenocarcinoma (EAC) has a dismal prognosis unless treated early or prevented at the precursor stage of Barrett's esophagus-associated dysplasia. However, some patients with cancer or dysplastic Barrett's esophagus (DBE) may not be captured by current screening and surveillance programs. Additional screening techniques are needed to determine who would benefit from endoscopic screening or surveillance. Partial wave spectroscopy (PWS) microscopy (also known as nanocytology) measures the disorder strength (Ld ), a statistic that characterizes the spatial distribution of the intracellular mass at the nanoscale level and thus provides insights into the cell nanoscale architecture beyond that which is revealed by conventional microscopy. The aim of the present study was to compare the disorder strength measured by PWS in normal squamous epithelium in the proximal esophagus to determine whether nanoscale architectural differences are detectable in the field area of EAC and Barrett's esophagus., Methods: During endoscopy, proximal esophageal squamous cells were obtained by brushings and were fixed in alcohol and stained with standard hematoxylin and Cyto-Stain. The disorder strength of these sampled squamous cells was determined by PWS., Results: A total of 75 patient samples were analyzed, 15 of which were pathologically confirmed as EAC, 13 were DBE, and 15 were non-dysplastic Barrett's esophagus; 32 of the patients, most of whom had reflux symptoms, acted as controls. The mean disorder strength per patient in cytologically normal squamous cells in the proximal esophagus of patients with EAC was 1.79-times higher than that of controls (P<0.01). Patients with DBE also had a disorder strength 1.63-times higher than controls (P<0.01)., Conclusion: Intracellular nanoarchitectural changes were found in the proximal squamous epithelium in patients harboring distal EAC and DBE using PWS. Advances in this technology and the biological phenomenon of the field effect of carcinogenesis revealed in this study may lead to a useful tool in non-invasive screening practices in DBE and EAC., (© Georg Thieme Verlag KG Stuttgart · New York.)

Purpose: To report the outcomes of choroidal nevi monitoring and to compare the detection of ultrasonographic hollowness, a risk factor of malignant transformation, from a B-scan with results from an A-scan examination., Design: Retrospective cohort study., Methods: Standardized A- and B-scan echography and ophthalmoscopy in 358 consecutive patients with median age 69 years and baseline choroidal nevus higher or lower than 1.5 mm in 51 (14%) and 307 (86%) eyes, respectively., Results: No growth or change in echographic or ophthalmoscopic features were found in 307 nevi with a median elevation of 0.9 mm (range, 0.7 to 1.5 mm) and a median follow-up of 6 years (range, 4 to 9 years). After 2 to 6 years, decreased internal reflectivity on standardized A-scan and ultrasonographic hollowness on B-scan were detected in 7 (18%) of 38 initially highly reflective choroidal nevi (thickness, 1.98 ± 0.37 mm); 2 (5%) nevi grew into melanoma 15 years after the first observation. Of 13 choroidal nevi (thickness, 2.75 ± 0.66 mm), with initial atypical medium to high reflectivity on standardized A-scan (100%) and hollowness on B-scan (85%), 6 (46%) were plaqued 6 to 15 months later because of the presence of multiple risk factors., Conclusions: No changes in thickness or echographic features of choroidal nevi elevated up to 1.5 mm were recorded during the follow-up period. In initially typical choroidal nevi higher than 1.5 mm, strong agreement was detected between decreased reflectivity on standardized A-scan and ultrasonographic hollowness on B-scan found after up to 15 years of stability. The vast majority of choroidal nevi with initial atypical standardized A-scan features showed ultrasonographic hollowness., (Copyright © 2013 Elsevier Inc. All rights reserved.)

Background: The detection of oral cancer at an early stage is an optimal strategy and is the most effective approach for preventing further progression. The rationale of the study was to evaluate the epithelial maturation pattern in oral mucosa exposed to tobacco/alcohol and on dysplastic oral mucosa using the scanning electron microscope., Methods: Fifteen subjects were selected based on clinical examination and divided into three groups: group 1-patients with apparently normal oral mucosa; group 2-patients with oral mucosa exposed to tobacco/alcohol; group 3-patients with clinical diagnosis of leukoplakia. An incisional biopsy was performed from the buccal mucosa. One part of the specimen was prepared for light microscopy and the other part was prepared for scanning electron microscopy., Results: Light microscopy revealed nonkeratinized stratified squamous epithelium in group 1, while group 2 demonstrated hyperparakeratinized stratified squamous epithelium with mild cytological atypia, and group 3 showed architectural and cytological changes. Scanning electron microscopy demonstrated flat-surfaced cells with equidistant parallel microridges in group 1, while group 2 showed irregular and widened microridges with numerous pits and absence of honeycomb pattern. Group 3 showed irregularly arranged broad and swollen cells with numerous pits and irregular microvilli projecting over the surface., Conclusion: The present study establishes the relationship of the surface abnormalities to the tendency of the cells to become malignant and thus serves as a tool in early detection of squamous cell carcinoma. It also emphasizes the need of routine follow-up in these high-risk patients for progression of carcinoma.

Objective: Tumours with neuroendocrine differentiation frequently express chromogranin A, synaptophysin and somatostatin receptors. The role of neuroendocrine differentiation in head and neck squamous cell carcinoma is not yet clear., Method: The presence of chromogranin A, synaptophysin and somatostatin receptors was studied immunohistochemically in 78 head and neck squamous cell carcinoma specimens., Results: Sparse chromogranin A expression was found in 41 per cent, associated with high chromogranin A messenger RNA expression and the presence of dense core granules. Low synaptophysin expression was found in 18 per cent. The highest staining scores were found for somatostatin receptor 5 (82 per cent), followed by somatostatin receptor 1 (69 per cent) and somatostatin receptor 2 (54 per cent), whereas somatostatin receptors 3 and 4 expression was low. Expression was not correlated with tumour stage or survival., Conclusion: Cells with neuroendocrine differentiation are sparsely scattered in some head and neck squamous cell carcinomas. Their pathophysiological role is elusive. In contrast, somatostatin receptor and particularly somatostatin receptor 5 expression is frequent in head and neck squamous cell carcinoma. Somatostatin receptor expression is not considered to indicate neuroendocrine differentiation in head and neck squamous cell carcinoma.

Background and Aim: Accumulating evidence suggests that the extracellular matrix play important roles in intercellular communications and contribute to the development of a number of diseases, including diseases of the gastrointestinal tract. The present study examined the structural characteristics and alterations of the extracellular matrix of the mucosa stroma in the Barrett's esophagus metaplasia-dysplasia-adenocarcinoma sequence., Methods: A total of 41 esophageal tissue specimens (15 esophageal adenocarcinoma, 10 Barrett's esophagus intestinal metaplasia, seven dysplasia and nine normal esophagus) were studied. The present study used transmission electron microscopy and computerized quantitative electron-microscopic analysis in order to investigate the characteristics of the extracellular matrix of the mucosa., Results: The study revealed that marked structural alterations of the mucosa stroma, relating to changes in the distribution and appearance of collagen fibers as well as to changes in numbers of matrix microvesicles, occur in Barrett's esophagus and esophageal adenocarcinoma. It was found that there were 3.1 times more microvesicles in the stroma in Barrett's esophagus than in the stroma of the normal esophagus (P<0.0001) and that there were 5.8 times more microvesicles in esophageal adenocarcinoma than in the normal esophagus (P<0.0001). There were 1.9 times more microvesicles in esophageal adenocarcinoma than in Barrett's esophagus (P=0.0043)., Conclusions: The study demonstrates distinctive alterations of the mucosa stroma extracellular matrix in the metaplasia-dysplasia-adenocarcinoma sequence. The findings suggest that the redistribution of collagen fibers and increases in numbers of matrix microvesicles may play roles in the formation of specialized intestinal metaplasia and the development of adenocarcinoma., (© 2012 Journal of Gastroenterology and Hepatology Foundation and Blackwell Publishing Asia Pty Ltd.)

Microvesicles (or MVs) are plasma membrane-derived vesicles released from most eukaryotic cells constitutively during early apoptosis or at higher levels after chemical or physical stress conditions. This review looks at some of the functions of MVs in terms of intercellular communication and ensuant signal transduction, including the transport of proteins (unconventional protein export) as well as of mRNA and microRNA. MVs also have roles in membrane repair, the removal of misfolded proteins, and in the control of apoptosis. We also discuss the role MVs have been shown to have in invasive growth and metastasis as well as in hypoxia in tumours and cerebral ischaemia. The association of MVs in infectious and autoimmune disease is also summarised together with their possible use as therapeutic agents.

The genetic code has received a great amount of attention from investigators, and the media since its discovery, and then again with the sequencing of the human genome in 2000. A decade later, investigators are beginning to look beyond the raw sequence to other mechanisms that affect gene expression. The main function of the nucleus is to maintain the genome and regulate gene expression. Changes in the expression of genes can drastically change the properties of the cell therefore giving the nucleus a role as the cell's "command post." In the past few years, one of the most notable discoveries in the study of the nucleus is that this organelle is not homogeneous. It is also not randomly organized; everything within the nucleus has a specific location with a specific function. Chromosome location within the nucleus relative to its center is directly related to transcription level. Additionally, there are specific regions of the nucleus where content and function differ. The various structures of the nucleus such as the membranes and matrix that supply support to the well protected chromatin offer ever increasing layers of complexity to the nucleus. Here, we focus on the nuclear matrix and its possible effects on signaling and cellular transformation leading to cancer.

The capacity of HDAC inhibitor sodium butyrate to induce senescence in cells derived from rat embryonic fibroblasts transformed by E1A+E1B19 kDa oncogenes has been studied. These transformants are resistant to apoptosis in response to gamma-irradiation and growth factor deprivation. The process of cell senescence was investigated by the analysis of cell growth curves, G1/S and G2/M cell cycle arrest, and senescent associated beta-galactosidase expression. The irreversibility of sodium butyrate antiproliferative activity was analyzed by clonogenic assay. We show that sodium butyrate suppresses proliferation and induces senescence in the E1A+E1B19 kDa transformed cells. Interestingly, NaB induces growth arrest due to accumulation of cells in G2/M phase, these cells are not tetraploid but mainly binuclear. Thus, in case of NaB induced senescence in E1A+E1B19 kDa transformed fibroblasts, the observed suppression of cell proliferation may be the result of cytokinesis failure leading to formation of binuclear and multinuclear cells incapable to proliferate.

Zebrafish are an ideal model organism to research cancer. Zebrafish embryos and larvae are optically translucent, which has made imaging multiple processes in development and disease possible. When coupled with fluorescent imaging techniques, zebrafish are fast becoming a model of choice for following tumor formation. This is highlighted by recent studies using fluorescent proteins to image xenograft transplantation, neovascularization, growth responses to drug treatments, and self-renewal. Fluorescent labeled tumors can be generated in zebrafish by multiple methods including chemical mutagenesis, oncogene expression by mosaic or stable transgenesis, or genetic mutations that are predisposing to cancer. In this chapter, we highlight the studies that have employed fluorescence to follow critical aspects of tumorigenesis, with particular focus on providing methods for labeling, isolating, transplanting, and imaging fluorescently labeled tumors in zebrafish., (Copyright © 2011 Elsevier Inc. All rights reserved.)

Perianal fistula is a very common problem in general population. Ninety percent of perianal fistulae arise from infected anal glands, and they often give rise to perianal abscesses. Very rarely perianal fistulae and abscesses undergo malignant transformation and give rise to carcinomas, mainly adenocarcinomas. We are reporting a case of squamous cell carcinoma arising from long-standing perianal fistula and how we managed it surgically., (© 2010 The Authors. Journal Compilation © 2010 Blackwell Publishing Ltd and Medicalhelplines.com Inc.)

Epithelioid sarcoma is an uncommon but well-described malignancy, which is found predominantly in the soft tissues of the young and middle-aged, and which pursues an indolent to aggressive course. It shows a degree of both mesenchymal and epithelial differentiation. Myofibroblastic differentiation has been recorded in epithelioid sarcoma for some time, the evidence being based mainly on the presence of smooth-muscle-type myofilaments and, more recently, on alpha-smooth-muscle actin and muscle-specific actin immunostaining. Myofibroblastic differentiation based on the stricter criterion of the fibronectin fibril/fibronexus junction has not so far been demonstrated except for a single atypical case with spindle-cell morphology and a cytokeratin-negative immunophenotype. The authors describe a conventional epithelioid sarcoma showing myofibroblastic differentiation based on the presence of fibronectin fibrils and fibronexus junctions. The patient was a 35-year-old Chinese male with a tumor that initially developed on his left forefinger: it recurred, then metastasised first to the left arm and eventually to the scalp. Histologically, the tumors had the typical features of epithelioid sarcoma: by immunohistochemistry, cytokeratin, epithelial membrane antigen, and vimentin were positive. By electron microscopy, rough endoplasmic reticulum, intermediate filaments, peripheral myofilaments, and fibronexus junctions were observed. This is the first epithelioid sarcoma of conventional histological type to show myofibroblastic differentiation on the basis of the more stringent criterion of the fibronexus. The findings are discussed in relation to the unusually varied differentiation reported for this tumor.

Conclusions: Numerical and structural centrosome abnormalities play a critical role in the tumor progression of in head and neck squamous cell carcinoma (HNSCC) and may provide useful information as a prognostic factor for these patients., Objectives: Centrosome alterations are often linked with aneuploidy, cell transformation, and tumor progress. We investigated centrosome abnormalities in HNSCC and correlated these variables to clinicopathological parameters and clinical follow up data of the patients., Methods: Retrospective analysis of numerical and structural alterations of centrosomes in tumor tissues and corresponding normal epithelium (n=50 and 31, respectively). Immunohistochemistry was performed using an anti-gamma-tubulin antibody. Image acquisition was done by an Orthoplan microscope, centrosomes were segmented interactively, and area as well as mean optical density was measured. Aneuploidy was evaluated by fluorescence in situ hybridization in a subset of cases (n=29)., Results: Numerical and structural centrosome abnormalities differed significantly between normal squamous epithelium and tumor cells (both P<0.0001). Especially numerical centrosome abnormalities were significantly associated with T category and tumor stage (both P<0.0001) and the occurrence of distant metastasis (P=0.002 and P=0.019, respectively). Numerical centrosome abnormalities correlated also with disease free survival of the patients (P=0.032) as well as shorter overall survival (P=0.003).

Primary and metastatic carcinomas have a reactive stroma characterized by many myofibroblasts. These cells have also been documented in nonepithelial malignancies, such as sarcomas, malignant melanoma, and lymphoid tumors but in generally far fewer numbers. In non-Hodgkin's lymphoma, Hodgkin's disease, and leukemia, myofibroblasts are rather rarely documented. In particular, there appear to be no reports of myofibroblasts in either primary bone-marrow/peripheral blood leukemia or secondary deposits of leukemia. In this paper, a case of a relapsed chronic myeloid leukemia appearing in an inguinal lymph node is described, containing many myofibroblasts. The case is detailed and presented with a discussion on the role of myofibroblasts in the progression of nonepithelial cancers.

Understanding cell functioning at the nanoscale has been hampered in part by the diffraction limited resolution of optical microscopy. We developed partial wave spectroscopic (PWS) microscopy that is capable of quantifying statistical properties of cell structure at the nanoscale. Our animal and human studies demonstrated that alterations in the nanoscale cell architecture is one of the earliest events in carcinogenesis and precedes any other known morphological changes at larger length scales (i.e. microarchitecture).

Carcinomas are widely thought to derive from epithelial cells with malignant progression often associated with an epithelial-mesenchymal transition (EMT). We have characterized tumors generated by spontaneously transformed human mesenchymal cells (TMC) previously obtained in our laboratory. Immunohistopathological analyses identified these tumors as poorly differentiated carcinomas, suggesting that a mesenchymal-epithelial transition (MET) was involved in the generation of TMC. This was corroborated by microarray and protein expression analysis that showed that almost all mesenchymal-related genes were severely repressed in these TMC. Interestingly, TMC also expressed embryonic antigens and were able to integrate into developing blastocysts with no signs of tumor formation, suggesting a dedifferentiation process was associated with the mesenchymal stem cell (MSC) transformation. These findings support the hypothesis that some carcinomas are derived from mesenchymal rather than from epithelial precursors.

Significant improvement in the understanding of mesenchymal stem cell (MSC) biology has opened the way to their clinical use. However, concerns regarding the possibility that MSCs undergo malignant transformation have been raised. We investigated the susceptibility to transformation of human bone marrow (BM)-derived MSCs at different in vitro culture time points. MSCs were isolated from BM of 10 healthy donors and propagated in vitro until reaching either senescence or passage (P) 25. MSCs in the senescence phase were closely monitored for 8 to 12 weeks before interrupting the cultures. The genetic characterization of MSCs was investigated through array-comparative genomic hybridization (array-CGH), conventional karyotyping, and subtelomeric fluorescent in situ hybridization analysis both before and after prolonged culture. MSCs were tested for the expression of telomerase activity, human telomerase reverse transcriptase (hTERT) transcripts, and alternative lengthening of telomere (ALT) mechanism at different passages. A huge variability in terms of proliferative capacity and MSCs life span was noted between donors. In eight of 10 donors, MSCs displayed a progressive decrease in proliferative capacity until reaching senescence. In the remaining two MSC samples, the cultures were interrupted at P25 to pursue data analysis. Array-CGH and cytogenetic analyses showed that MSCs expanded in vitro did not show chromosomal abnormalities. Telomerase activity and hTERT transcripts were not expressed in any of the examined cultures and telomeres shortened during the culture period. ALT was not evidenced in the MSCs tested. BM-derived MSCs can be safely expanded in vitro and are not susceptible to malignant transformation, thus rendering these cells suitable for cell therapy approaches.

Aims: To assess neuronal differentiation in oligodendrogliomas (ODGs)., Methods and Results: An electron microscopic and immunohistochemical study of 41 consecutive cases was performed. In all cases, tumour cells with neuritic structures were identified ultrastructurally, including synapses and neurosecretory granules. For the immunohistochemical identification of synaptophysin, monoclonal antibody clones 27G12, Snp88 and SY38 and a polyclonal antibody were compared in optimized protocols on slides from a spectrum of tissues and 16 ODGs. 27G12 gave the best signal-to-noise ratio, while SY38 gave the poorest. When 27G12 was applied on all 41 ODGs, widespread immunoreactivity was obtained in 100%. Among three antibodies to chromogranin compared similarly, clone LK2H10 and a polyclonal antibody gave identical patterns of immunoreactivity, whereas clone DAK-A3 gave weaker reactions. When LK2H10 was applied on all tumours, staining was found in 12 (29%). All tumours but one stained strongly for glial fibrillary acidic protein and all for synapsin I. Fluorescence in situ hybridization analysis showed a concomitant 1p/19q deletion in 12/16 ODGs., Conclusions: Our study provides evidence for widespread neuronal differentiation in ODGs, suggesting that these tumours may be derived from progenitor cells with limited commitment. Antibody selection and protocol optimization are mandatory for reliable immunohistochemistry results.

A brief review of the cytoskeleton dynamic structure in cells of two types: fibroblasts and epitheliocytes. Differences have been described between the functions of y-actin filaments and beta-actomyosin bundles. Tubulogenesis and angiogenesis have been considered as consequences of partial epitheliomesenchymal transformation and neoplastic transformation as a consequence of gamma-actomyosin bundles disturbance.

Antioxidants are one of the key players in tumorigenesis, several natural and synthetic antioxidants were shown to have anticancer effects. In the present investigation the efficacy of silymarin on the antioxidant status of N-nitrosodiethylamine (NDEA) induced hepatocarcinogenesis in Wistar albino male rats were assessed. The animals were divided into five groups. The animals in the groups 1 and 3 were normal control and silymarin control, respectively. Groups 2, 4 and 5 were administered with 0.01% NDEA in drinking water for 15 weeks to induce hepatocellular carcinoma (HCC). Starting 1 week prior to NDEA administration group 4 animals were treated with silymarin in diet for 16 weeks, 10 weeks after NDEA administration group 5 animals were treated with silymarin and continued till the end of the experiment period (16 weeks). After the experimental period the body weight, relative liver weight, number of nodules, size of nodules, the levels of lipid peroxidation, glutathione (GSH), and the activities of antioxidant enzymes were assessed in both haemolysate and liver tissue. In group 2 hepatocellular carcinoma induced animals there was an increase in the number of nodules, relative liver weight. The levels of lipid peroxides were elevated with subsequent decrease in the body weight, (glutathione) GSH, superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx), glutathione reductase (GR), glucose-6-phosphate dehydrogenase (G6PD). In contrast, silymarin + NDEA treated groups 4 and 5 animals showed a significant decrease in the number of nodules with concomitant decrease in the lipid peroxidation status. The levels of GSH and the activities of antioxidant enzymes in both haemolysate and liver were improved when compared with hepatocellular carcinoma induced group 2 animals. The electron microscopy studies were also carried out which supports the chemopreventive action of the silymarin against NDEA administration during liver cancer progression. These findings suggest that silymarin suppresses NDEA induced hepatocarcinogenesis by modulating the antioxidant defense status of the animals.

During breast cancer development, the luminal space of the mammary acinar unit fills with proliferating epithelial cells that exhibit growth factor-independence, cell attachment defects, and a more invasive fibroblastic phenotype. Here, we used primary cultures of mammary epithelial cells derived from genetically engineered mice to identify caveolin-1 (Cav-1) as a critical factor for maintaining the normal architecture of the mammary acinar unit. Isolated cultures of normal mammary epithelial cells retained the capacity to generate mammary acini within extracellular matrix. However, those from Cav-1 (-/-) mice exhibited defects in three-dimensional acinar architecture, including disrupted lumen formation and epidermal growth factor-independent growth due to hyperactivation of the p42/44 mitogen-activated protein kinase cascade. In addition, Cav-1-null mammary epithelial cells deprived of exogenous extracellular matrix underwent a spontaneous epithelial-mesenchymal transition, with reorganization of the actin cytoskeleton, and E-cadherin redistribution. Mechanistically, these phenotypic changes appear to be caused by increases in matrix metalloproteinase-2/9 secretion and transforming growth factor-beta/Smad-2 hyperactivation. Finally, loss of Cav-1 potentiated the ability of growth factors (hepatocyte growth factor and basic fibroblast growth factor) to induce mammary acini branching, indicative of a more invasive fibroblastic phenotype. Thus, a Cav-1 deficiency profoundly affects mammary epithelia by modulating the activation state of important signaling cascades. Primary cultures of Cav-1-deficient mammary epithelia will provide a valuable new model to study the spatial/temporal progression of mammary cell transformation.

The keratin filament network is an important part of the cytoskeleton. It is involved in the regulation of shape and viscoelasticity of epithelial cells. The morphology of keratin networks depends on post-translational modifications of keratin monomers. In-vitro studies indicated that network characteristics, such as filament crosslink density, determines the biophysical properties of the filament network. This report presents a quantitative method for the morphological analysis of keratin filament networks. Visualization of filaments was based on prefixation extraction of epithelial cells and scanning electron microscopy (SEM). SEM images were processed by a skeletonization algorithm to obtain a graph structure that represents individual filaments as well as their connections. This method was applied to investigate the effects of transforming growth factor alpha (TGFalpha) on the morphology of keratin networks in pancreatic cancer cells. TGFalpha contributes to pancreatic cancer progression and activates signalling pathways phosphorylating keratin monomers. Using this new method, a significant alteration to the keratin network morphology could be detected in response to TGFalpha.

Experiments conducted by the authors, as well as clinical studies, show that in addition to hormonal mis-balance, development of mammary gland cancer is significantly influenced by the action of low frequency electromagnetic fields on epiphysis. By reducing the production of melatonin it increases the risk of development of mammary gland tumors. The review of scientific literature indicates that pinealocytes are the main morphological substrate responsible for functional activity of pineal cells. When estimating large specific weight of lipids, many researchers point to the dependence of their presence in epiphysis on the level of melatonin. It is thought that hormones of pineal cells are deposited in the form of lipid drops. The most characteristic feature in the structure of pinealocytes is the presence of large number of various size complexes in their bodies and appendices. According to many authors this could be the form of depositing the secretory products of pinealocytes, such as melatonin, biological amines, etc. Ultrastructure characteristic of pinealocytes described in the review provides deeper understanding of the fine structure morpho-physiology of epiphysis, enable to point out the peculiarities of its functionality under the influence of electromagnetic fields and discover the organizational structure of pineal body at a time of the mammary gland tumor development.

Most cancers arise from the stepwise accumulation of genetic changes. There is also evidence for defects in the machinery and checkpoints for maintenance of normal diploid chromosome complements, resulting in genetic instability that helps fuel the accumulation of mutations that contribute to the development of cancer. The proto-oncogene protein kinase B (PKB/Akt), and its regulators including phosphatidylinositol 3' kinase and PTEN, has been shown to play critical roles in the regulation of multiple cellular functions such as transcription, cell survival, cell cycle progression, angiogenesis and cell motility--all of which are important to the malignant process. Here, we report the use of a membrane targeted PKBbeta, the activation of which is under the control of a 4-hydroxy-Tamoxifen-responsive estrogen-receptor (ER) ligand binding domain. Induction of PKBbeta-ER activity in human kidney epithelial cells (HEK293) resulted in changes in cellular growth, size, and in the appearance of aneuploid cells. Over time, in a PKBbeta-dependent manner, cells also underwent extensive multinucleation caused due to a combination of both endomitosis and cell fusion. These findings suggest that chronic activation of PKBbeta may contribute to genetic instability and autophagy, properties commonly found in tumor cells.

We recently reported that uPARAP/Endo180 can mediate the cellular uptake and lysosomal degradation of collagen by cultured fibroblasts. Here, we show that uPARAP/Endo180 has a key role in the degradation of collagen during mammary carcinoma progression. In the normal murine mammary gland, uPARAP/Endo180 is widely expressed in periductal fibroblast-like mesenchymal cells that line mammary epithelial cells. This pattern of uPARAP/Endo180 expression is preserved during polyomavirus middle T-induced mammary carcinogenesis, with strong uPARAP/Endo180 expression by mesenchymal cells embedded within the collagenous stroma surrounding nests of uPARAP/Endo180-negative tumor cells. Genetic ablation of uPARAP/Endo180 impaired collagen turnover that is critical to tumor expansion, as evidenced by the abrogation of cellular collagen uptake, tumor fibrosis, and blunted tumor growth. These studies identify uPARAP/Endo180 as a key mediator of collagen turnover in a pathophysiological context.

The role of enterochromaffin-like (ECL) cells in gastric carcinogenesis is not fully understood. Spontaneous tumours developing in hypergastrinemic female cotton rats have an adenocarcinoma phenotype, but numerous cells in the dysplastic mucosa as well as in the carcinomas are positive for neuroendocrine markers. In the present study of female cotton rats with 2 and 8 months' hypergastrinemia, the oxyntic mucosa of the stomach was examined histologically and immunolabelled for histidine decarboxylase (HDC) and pancreastatin, and hyperplastic and neoplastic ECL cells were evaluated by electron microscopy. These animals developed hyperplasia of the oxyntic mucosa in general and of the ECL cells in particular after 2 months and dysplasia and carcinomas after 8 months. The immunoreactivity of the ECL cells in the oxyntic mucosa was increased at 2 months and declined at 8 months. These histological changes were associated with progressive loss of secretory vesicles and granules in ECL cells. We suggest that ECL cells in hypergastrinemic cotton rats dedifferentiate with time and that the gastric carcinomas may develop from ECL cells.

The rearrangement of the actin cytoskeleton has been shown to play a critical role in the development of transformation and malignant phenotype of cancer cells. Rho family GTPases regulate the arrangement of the actin cytoskeleton. By wound-healing assay, we have found that NIH 3T3 fibroblast cells move towards the wound- gaps by extending filopodial and lamellipdial structures at the leading edge of the moving cells. We have inactivated the function of Rho GTPases of v-Ras transformed NIH 3T3 cells by overexpressing Rho GTPase-activating (RhoGAP) domain of RhoGAP of p190. We have observed that inactivation of Rho, Rac and Cdc42 GTPases by overexpressing RHG causes inhibition of: (i) polymerization of actin to form filaments, (ii) formation of lamellipodia, filopodia and stress fibres, (iii) cell motility, (iv) cell spreading and (v) cell-to-cell adhesions. These results further strengthen the current knowledge on the role of Rho, Rac and Cdc42 GTPases in the regulation of the rearrangement of actin cytoskeleton. Our results, for the first time, demonstrate that RhoGAP domain of RhoGAP could be used to study the molecular mechanism of Ras-mediated signalling in growth, differentiation and carcinogenesis.

Arsenic and cadmium (Cd(+2)) are human carcinogens, and epidemiological studies have implicated both pollutants in the development of urinary bladder cancer. Despite this epidemiological base, it is unknown if either Cd(+2) or arsenite (As(+3)) can directly cause the malignant transformation of human urothelial cells. The goal of this study was to determine if Cd(+2) and/or As(+3) are able to cause the malignant transformation of human urothelial cells. The strategy employed was to expose the nontumorigenic urothelial cell line UROtsa to long-term in vitro exposure to Cd(+2) and As(+3), with the endpoint being the ability of the cells to form colonies in soft agar and tumors when heterotransplanted into nude mice. It was demonstrated that a long-term exposure to either 1 M Cd(+2) or 1 M As(+3) resulted in the selection of cells that were able to form colonies in soft agar and tumors when heterotransplanted into nude mice. The histology of the tumor heterotransplants produced by UROtsa cells malignantly transformed by Cd(+2) had epithelial features consistent with those of a classic transitional-cell carcinoma of the bladder. The histology of the tumor heterotransplants produced by cells malignantly transformed by As(+3) was unique in that the cells displayed a prominent squamoid differentiation.

Objectives: Endometriosis is an illness accompanied by invasion features, but malignant changes appear extremely seldom. Metallothionein (MT) is a protein and takes part in the detoxicating processes of the organism. MT is located, among others, in benign and malignant neoplasms in animals as well as humans, mainly in the S phase of cellular cycle, and that is why MT is considered to be both an index of cell proliferation and tumor progress., Materials and Methods: 34 specimens from 21 women with ovary endometriosis (III degree according to AFS) have been examined. The specimens were obtained during surgery and they were histopathologically verified. The material was coloured by H + E and by van Gieson method, and MT was determined immunohistochemically. The measurement of the cells number containing MT was performed with measurement system Multi-Scan Base V8.08, with the microscope Axiophot, Zeiss Jena in so-called measurements areas, with the surface 18802 microns 2., Results: High MT capacity was found in the epithelial cells in the endometriosis focus. This high MT capacity may imply that there exists the proliferation process in the focuses of external endometriosis. Simultaneously, the lowest MT capacity was discovered in glandular ducts., Conclusions: Proliferating epithelial cells contain the highest capacity of MT, which indicates increase of number of dividing cells particularly in the S phase of cellular cycle and that is why MT can be considered one of the markers of ovary endometriosis.

Eighty-two mesenchymal tumors of the gastrointestinal tract were examined by electron microscopy for the purposes of subtyping for diagnostic precision and of understanding cellular differentiation. Tumors were subclassified into leiomyoma/leiomyosarcoma, tumors of the interstitial cell of Cajal (equivalent to traditionally defined GISTs [Miettinen et al. Hum Pathol. 1999; 30:1213-1220; Mod Pathol. 2000; 13:1134-1142]), gastrointestinal autonomic nerve tumors (GANTs), and fibroblastic and myofibroblastic tumors, using criteria from the literature. Leiomyoma/leiomyosarcoma were diagnosed by myofilaments, attachment plaques, plasmalemmal caveolae, and lamina; GIST by processes or cell bodies full of intermediate filaments, solitary focal densities amid intermediate filaments, attachment plaques with incomplete lamina, scarce myofilaments, and smooth endoplasmic reticulum; GANTs by neuroendocrine granules, cell bodies/processes full of intermediate filaments (more rarely microtubules), and smooth endoplasmic reticulum; fibroblastic/myofibroblastic tumors by abundant rough endoplasmic reticulum, myofilaments, and fibronexuses. Seventy-three tumors (89%) were successfully subclassified, as 5 leiomyoma/leiomyosarcoma (6%), 36 GISTs (44%), 22 GANTs (27%), 10 fibroblastic and myofibroblastic tumors (12%). Results indicated overlap between poorly differentiated leiomyosarcoma and GIST, and between GIST and GANT. GANT is emphasized as a neuronal tumor identifiable by electron microscopy, and thereby distinguishable from GIST.

For a long time now we have been putting forwards the hypothesis that, when two cells belonging to different tissues of the same organism fuse, the nucleous/nuclei of the resulting "fused" cell could lay into a cytoplasm possibly containing factors influencing the activation or deactivation of genes able to interfere with the differentiation "iter" proper of the parent cells. As a consequence, the fused cells could undergo neoplastic transformation. In order to give an experimental support to such an hypothesis, mononuclear cells and granulocytes were isolated from the white blood cells of the same individual, mixed (at 1/1 ratio) and fused, using polyethylen glycol (PEG). The fused cells were then cultured into a suitable medium, supplemented with phytohemoagglutinin (PHE), a lectin which is able to stimulate at least one of the nuclei. Samples from these cultures were withdrawn at progressive time-intervals and stained with May Grümwald-Giemsa. Microscopic examination showed the appearance of cells morphologically resembling Hodgkin's and Reed Sternberg cells, or leukemic cells. A crown of mononucleated cells was often surrounding such anomalous cells, as happens in the early stage of killer cells action against target cells. In two out of fifty cultures, cells grew without need for serum supply, suggesting the onset of an autocrine system.

Transmission electron microscopy (EM) has been used to identify the ultrastructural details of normal and cancerous human oral mucosa. However, inconsistent reports of structural descriptions have rendered transmission EM valueless in the diagnosis of oral squamous cell carcinoma (SCC) or as a prognostic indicator. To identify features of dysplasia for diagnostic purposes, normal mucosa, severe dysplasia, oral SCC and normal margin adjacent to oral SCC were used to compare the ultrastructural features of normal and premalignant oral mucosa and oral SCC. The preparatory stages of dehydration, embedding, cutting and positive staining for transmission EM were modified and tested to improve ultrastructural definition. Thin and discontinuous basal laminas were found in mucosa with severe dysplasia and normal margin adjacent to oral SCC. No basal lamina was identified in oral SCC. This study has shown that there are some ultrastructural changes during malignant transformation of oral mucosa. Together with other laboratory investigative techniques, transmission EM may be helpful in detecting malignant changes in oral mucosa., (Copyright 2002 The British Association of Oral and Maxillofacial Surgeons.)

The term radial segmentation (RS) is applied to a characteristic nuclear deformation observed in peripheral blood in vitro in some neoplastic and normal cells. It concerns al mononuclear cells. RS positive lymphocytes were found as CD 4 cells. Different conditions and substances, i.e. anticoagulant or cyclosporin can induce the formation of RS nuclei in vitro. Elevated ratio of RS nuclei in peripheral blood has been observed in patients with some autoimmunological diseases i.e. rheumatoid arthritis, diabetes mellitus insulin-dependent, sarcoidosis. Reduced ratio of RS nuclei was observed in neoplastic diseases. The aim of the study was to analyze the incidence of RS nuclei of lymphocytes in vitro in peripheral blood (induced by cyclosporin) in patients with non-small cell lung cancer (NSCLC). Heparinized peripheral blood was obtained from controls and patients with primary NSCLC. The blood was incubated with cyclosporin, and then lymphocytes were isolated by Lymphoprep. RS positive lymphocytes were counted in smears stained with MGG stain. In peripheral blood from healthy donors the average incidence of lymphocytes RS was 3.314% and in patients with NSCLC 4.481% respectively. The difference between controls and patients with NSCLC was not significant. No correlation was found between incidence of RS and stadium of lung cancer.

The purpose of this study is to investigate the mechanism of tumor spread in the pagetoid spread of germ cell tumors in the rete testis (PSRT). Twenty consecutive cases of germ cell tumor of the testis (9 seminomas, 3 embryonal carcinomas, and 8 teratocarcinomas) were retrieved to identify the cases with PSRT. The areas of pagetoid spread were examined by the serial sectioning of the entire thickness of the tissue block. Available fresh tissue was submitted for electron microscopic study. Ten cases were associated with PSRT and had focal or extensive areas of intratubular germ cell neoplasia (IGCN) in the proximity of the tumor and the rete testis (RT). In the remaining 10 cases, 6 were associated with IGCN distant from the RT and the last 4 were not associated with IGCN. Seminiferous tubules with IGCN were seen connecting with the RT with pagetoid spread. Isolated single intraepithelial tumor cells also were identified at the periphery of the areas with PSRT. Electron microscopic study of the RT of 4 cases with PSRT (2 seminomas, 1 embryonal carcinoma, and 1 teratocarcinoma) revealed desmosome-type junctions between tumor cells with RT epithelial cells. Direct tumor expansion and cell motility as mechanisms of tumor spread in PSRT does not explain the presence of isolated cells and desmosome-type junctions of the tumor cells as demonstrated in this study. The authors believe that the field effect plays an important part in the pathogenesis of this pagetoid spread in the RT. It is likely that this field effect is induced by the germ cell tumor and is operated through the immature germ cells or undifferentiated epithelial cells in the RT adjacent to the tumor cells.

Metastasis is responsible for most cancer deaths. A better understanding of the process provides opportunities to develop new treatments to prevent metastasis. This article summarizes findings from experimental in vivo videomicroscopy and quantitative studies on metastatic inefficiency, which indicate that early steps in hematogenous metastasis may be quite efficient, but that regulation of cancer cell growth in secondary sites determines metastatic outcome. The authors have identified three key stages of this growth regulation: survival of a subset of single cells, proliferation of a subset of these cells to form preangiogenic micrometastases, and persistence of growth of a subset of these to form vascularized metastases. Formation of clinically relevant metastases is determined by the proportion of cells that proceeds successfully through each stage, and surviving single cells and preangiogenic micrometastases both represent possible sources of tumor dormancy.

Skin cancers, both non-melanoma and melanoma, usually progress through sequential steps towards malignant transformation, leading to mutant clones and precancerous lesions. Prevention of skin cancers relies on reduction of exposure to solar radiation and may be evaluated by measuring induction of intermediate-effect biomarkers such as sunburn cells or p53 mutations in the epidermis, actinic (solar) keratoses, UV-induced immunosuppression or naevi. Sunburn cells (apoptotic keratinocytes) and p53 mutations are indicators of UV-induced DNA lesions as early steps of malignant transformation of epidermal keratinocytes. Actinic keratoses are premalignant sun-induced skin lesions, characterized as keratinized patches with aberrant cell differentiation and proliferation; they represent risk factors for basal-cell carcinoma and melanoma and are precursors of squamous-cell carcinoma. Studies in humans have investigated UV-induced immunosuppression and its modulation by topical sunscreen application, focusing on contact hypersensitivity as measured by immunization or response to haptens, or on modulation of stimulation of allogeneic lymphocytes by epidermal cells, or local release of immunomodulatory molecules such as cis-urocanic acid or interleukin-10. Naevi are focal collections of melanocytes, usually found at the junction of the epidermis and dermis or at various depths in the dermis. Common acquired naevi arise after birth both spontaneously and in response to sun exposure. Most acquired naevi are clonal, while most melanocytes in non-naeval areas are not. Although it is not yet certain whether naevi represent premalignant lesions or risk factors, many melanomas arise in acquired naevi, and the number of naevi constitutes the best predictor of individual risk of melanoma. The presence of large (i.e., >5 mm) or atypical naevi (i.e., large naevi with non-uniform colour and irregular borders) is associated with elevated melanoma risk, independently of the number of smaller naevi. Children seem particularly vulnerable to sun-induced biological events involved in the genesis of melanoma, and the greatest increase in naevus numbers per unit of skin surface occurs before adolescence. Therefore, the distribution of naevi and their development in children are relevant to understanding melanoma occurrence in adults.

Breast cancer is thought to originate through progressively aberrant precursor lesions, paralleled by increasing morphological changes. The aim of this study was to quantify nuclear features by image cytometry in invasive breast cancer and its early (hyperplasia) and late (ductal carcinoma in situ) precursor lesions, in order to objectively describe nuclear changes in the spectrum of proliferative intraductal and invasive breast lesions. Image cytometry was performed on tissue sections of 20 samples of normal breast tissue, 71 of usual ductal hyperplasia (UDH), nine of atypical ductal hyperplasia (ADH), and 11 of well-differentiated and 13 of poorly differentiated ductal carcinoma in situ (DCIS) lesions. The invasive breast carcinomas consisted of 19 well-differentiated and 24 poorly differentiated lesions. Through the spectrum from normal breast tissue to invasive carcinoma, progressive changes in many nuclear features were measured. Significant differences were found between nuclei of florid ductal hyperplasia compared with mild and moderate ductal hyperplastic lesions, suggesting that florid ductal hyperplasia may be a more advanced lesion than assumed and may contain cancer precursor cells. No differences were found between ADH and well-differentiated DCIS, suggesting that these lesions are closely related. Feature values of well-differentiated DCIS were comparable to values found in well-differentiated invasive carcinoma and the same applied to poorly differentiated DCIS and invasive lesions. These results support the hypothesis that breast cancer develops through different routes of progression, one leading to well-differentiated invasive cancer through well-differentiated DCIS, and one leading to poorly differentiated invasive cancer through poorly differentiated DCIS. In conclusion, image cytometry reveals progressive changes in nuclear morphological and subvisual chromatin distribution features in the spectrum from intraductal proliferations to invasive breast cancer. This provides evidence for a progression from usual to atypical ductal hyperplasia and then to invasive cancer, through different routes for well-differentiated and poorly differentiated lesions.

The neoplastic conversion of a normal cell to a malignant one is a multistage process that occurs after a series of molecular alterations. Several chemical and physical agents can alter the morphology of different types of cells. Scanning and transmission electron microscopy have been valuable in evaluating changes that occur in the progression of transformation. MCF-10F, a spontaneously immortalized human breast epithelial cell line (Soule et al., 1990), was treated with benz(alpha)pyrene (BP) (Calaf and Russo, 1993) and then transfected with the c-Ha-ras oncogene (Calaf et al., 1995). The phenotypic changes of breast cancer progression were studied through the use of scanning and transmission electron microscopy. Activated oncogenes have been detected in a variety of malignant tumors and the altered expression of certain genes seems to play a role in the cancer process. Carcinogen-treated and transfected cells showed a progression of changes in the morphology, anchorage independent growth, invasiveness and capability of tumor formation in the SCID mice. This in vitro cancer model can parallel the progression of breast cancer seen through molecular changes that occur and have been observed during the natural development of this disease.