Atomic force microscopy for revealing micro/nanoscale mechanics in tumor metastasis: from single cells to microenvironmental cues.

Mechanics are intrinsic properties which appears throughout the formation, development, and aging processes of biological systems. Mechanics have been shown to play important roles in regulating the development and metastasis of tumors, and understanding tumor mechanics has emerged as a promising way to reveal the underlying mechanisms guiding tumor behaviors. In particular, tumors are highly complex diseases associated with multifaceted factors, including alterations in cancerous cells, tissues, and organs as well as microenvironmental cues, indicating that investigating tumor mechanics on multiple levels is significantly helpful for comprehensively understanding the effects of mechanics on tumor progression. Recently, diverse techniques have been developed for probing the mechanics of tumors, among which atomic force microscopy (AFM) has appeared as an excellent platform enabling simultaneously characterizing the structures and mechanical properties of living biological systems ranging from individual molecules and cells to tissue samples with unprecedented spatiotemporal resolution, offering novel possibilities for understanding tumor physics and contributing much to the studies of cancer. In this review, we survey the recent progress that has been achieved with the use of AFM for revealing micro/nanoscale mechanics in tumor development and metastasis. Challenges and future progress are also discussed.

El organofosforado clorpirifos como disruptor estrogénico y factor de riesgo para el cáncer de mama / The organophosphorus chlorpyrifos as endocrine disruptor and breast cancer factor risk

El clorpirifos (CPF) es un insecticida de amplio espectro que se utiliza en Argentina y en otros países de Latinoamérica. Se emplea para el control de plagas en la producción de frutas, hortalizas, cereales y plantas ornamentales. El principal mecanismo de acción descripto para este insecticida es la inhibición de la acetilcolinesterasa. Sin embargo, reportes más recientes sugieren múltiples efectos del plaguicida independientes de la inhibición de esa enzima. El objetivo de este trabajo es transmitir a la comunidad los resultados de nuestras investigaciones obtenidos utilizando diferentes dosis de CPF en distintos modelos experimentales, tanto in vitro como in vivo. En relación a esto, hemos evidenciado una acción del CPF sobre el sistema redox celular que conduce al incremento de especies reactivas del oxígeno y consecuentemente a la activación de diferentes vías de señalización. Además, hemos determinado que el insecticida CPF puede comportarse como un disruptor endócrino modulando la acción de los estrógenos y alterando la normal estructura del tejido mamario. Nuestros resultados alertan sobre el impacto que este compuesto podría tener sobre la salud, sugiriendo la necesidad de revisar su uso dado que manifiesta acciones a dosis encontradas en el ambiente.

Chlorpyrifos (CPF) is a broad spectrum insecticide used in Argentina and other Latin American countries. It is commonly used for pest control in the production of fruits, vegetables, cereals and ornamental plants. The main mechanism of action described for this insecticide is the inhibition of acetylcholinesterase activity. However, more recent reports suggest multiple effects for this pesticide in an independent way from the inhibition of this enzyme. The objective of this work is to convey to the community the results of our investigations obtained using different doses of CPF in various experimental models, both in vitro and in vivo. In this connection, we have shown a CPF action on the cellular redox system which leads to increased reactive oxygen species and the consequent activation of different signaling pathways. In addition, we have determined that the insecticide CPF acts as an endocrine disruptor modulating the action of estrogen and altering the normal structure of breast tissue. Our findings warn about the impact that this compound might have on health, suggesting the need to review its use since adverse actions were found at environmentally relevant doses.

Modeling and integral X-ray, optical, and MRI visualization of multiorgan metastases of orthotopic 4T1 breast carcinoma in BALB/c mice.

A model of highly metastasizing orthotopic allogeneic breast carcinoma was reproduced and standardized in experiments on BALB/c mice. 4T1 cells characterized by high metastatic activity were transfected with red fluorescent protein (RFP) gene or firefly luciferase (Luc2) gene. Unmodified 4T1 cells and modified 4T1-RFP and 4T1-Luc2 cells were subcutaneously injected to mature female mice into the second mammary fat pads. Quantitative evaluation of the primary node and visceral metastases was performed using magnetic-resonance imaging, X-ray and optical tomography. Modification of 4T1 cells with RFP gene considerably reduced their invasive and metastatic potential and led to spontaneous regression of the primary tumor in 20% cases. Modification of 4T1 cells with Luc2 gene had practically no effect on proliferative, invasive, and metastatic characteristics of the tumor and provided the possibility of quantitative analysis of the primary tumor dynamics by the luminescence intensity. The survival median in mice receiving unmodified 4T1 cells and transfected 4T1-RFP and 4Т1-Luc2 cells was 32, 42, and 38 days, respectively. Neither primary node nor tumor metastases accumulated gadolinium-containing contrast agent and Alasens fluorescent tracer. After implantation of 4T1 and 4Т1-Luc2 cells, multiple metastases were more often detected in the lungs, liver, spleen, spine, and regional lymph nodes and less frequently in the brain, which corresponded to metastasizing profile of human breast cancer. The developed model of orthotopic breast carcinoma 4T1 in BALB/c mice with complex detection of multiple organ metastases using X-ray microCT, optical, and MRI can be recommended for preclinical studies of new antitumor preparations.

Atomic force microscopy-based microrheology reveals significant differences in the viscoelastic response between malign and benign cell lines.

Mechanical phenotyping of cells by atomic force microscopy (AFM) was proposed as a novel tool in cancer cell research as cancer cells undergo massive structural changes, comprising remodelling of the cytoskeleton and changes of their adhesive properties. In this work, we focused on the mechanical properties of human breast cell lines with different metastatic potential by AFM-based microrheology experiments. Using this technique, we are not only able to quantify the mechanical properties of living cells in the context of malignancy, but we also obtain a descriptor, namely the loss tangent, which provides model-independent information about the metastatic potential of the cell line. Including also other cell lines from different organs shows that the loss tangent (G″/G') increases generally with the metastatic potential from MCF-10A representing benign cells to highly malignant MDA-MB-231 cells.

El sistema lewis hoy. nuevos enfoques / The lewis system today. new approaches

La expresión de antígenos (Ags) Lewis depende de alelos heredados en loci independientes, el gen Secretor (SE) que codifica la fucosiltransferasa 2 (FUT2) y el gen Lewis (LE) que codifica la fucosiltransferasa 3 (FUT3). El gen Se codifica una glicosiltransferasa que adiciona una fucosa en la cadena precursora de tipo 1 formando el Ag H en secreciones y fluidos. Como los azúcares inmunodominantes del Ag A y B pueden ser agregados a la cadena H de tipo 1, la FUT2 también controla la expresión de Ag A y B en las secreciones. El gen se es un alelo no funcional. El gen Le codifica una transferasa diferente que adiciona una fucosa en el 2do carbono en el precursor de tipo 1. El alelo le no es funcional. Las FUT2 y FUT3 interactúan para la formación de Ags Lewis en secreciones y fluídos. Los Ags Lewis en los eritrocitos no son en realidad parte integral de la membrana, están adsorbidos sobre la superficie en forma pasiva a partir del plasma. Están ampliamente distribuidos en tejidos humanos, eritrocitos, endotelio, riñón, tracto genitourinario, epitelio gastrointestinal y son receptores para algunos patógenos. Los anticuerpos (Acs) anti-Lewis en general no son clínicamente significativos, aunque se han publicado algunos casos de reacciones transfusionales hemolíticas, enfermedad hemolítica fetoneonatal y rechazo de transplante renal. Este trabajo es una revisión sobre los Ags del Sistema Lewis enfocada hacia sus diferentes funciones biológicas y su importancia en campos variados fuera del Banco de Sangre y la Inmunohematología tradicional.

The expression of Lewis blood group antigens depends on the alleles inherited at independent loci, FUT2 Secretor gene (SE) and FUT3 Lewis gene (LE). The Se and Le alleles encode separate fucosyltransferases that interact to form Lewis antigens in secretions and fluids. The Lewis antigens on red blood cells are not integral to the membrane but are passively adsorbed from the plasma. The allele Se encodes a transferase that adds fucose to type 1 precursor chains in secretions and fluids to form type 1 H antigen. Because A and B terminal sugars may be added to type 1 H chains, FUT2 also controls A and B expression in secretions. The FUT2 allele se gen is a nonfunctional allele. The FUT3 allele Le encodes a transferase that adds a fucose in other position in type 1 H precursor. The FUT3 allele le gen is a nonfunctional allele. The Le antigens are widely distributed in human tissues and fluids and are receptors for some pathogenic organisms. Lewis antibodies are rare and clinically no significant, although there are some reports of hemolytic transfusion reactions, hemolytic disease of the newborn and renal transplant rejection. This review focuses on different biological functions of Lewis antigens and their importance in some fields other than Blood Banks and traditional.

Endosomal WASH and exocyst complexes control exocytosis of MT1-MMP at invadopodia.

Remodeling of the extracellular matrix by carcinoma cells during metastatic dissemination requires formation of actin-based protrusions of the plasma membrane called invadopodia, where the trans-membrane type 1 matrix metalloproteinase (MT1-MMP) accumulates. Here, we describe an interaction between the exocyst complex and the endosomal Arp2/3 activator Wiskott-Aldrich syndrome protein and Scar homolog (WASH) on MT1-MMP­containing late endosomes in invasive breast carcinoma cells. We found that WASH and exocyst are required for matrix degradation by an exocytic mechanism that involves tubular connections between MT1-MMP­positive late endosomes and the plasma membrane in contact with the matrix. This ensures focal delivery of MT1-MMP and supports pericellular matrix degradation and tumor cell invasion into different pathologically relevant matrix environments. Our data suggest a general mechanism used by tumor cells to breach the basement membrane and for invasive migration through fibrous collagen-enriched tissues surrounding the tumor.

18F-FDG PET/TC con imagen hepática en dos tiempos en la sospecha de recidiva del cáncer colorrectal / Dual-time point images of the liver with 18F-FDG PET/CT in suspected recurrence from colorectal cancer

Objetivo. Analizar el potencial de la PET/TC usando imagen tardía del hígado en pacientes con sospecha de recidiva de cáncer colorrectal. Material y métodos. Se han incluido prospectivamente 71 pacientes (22 mujeres, 49 hombres) con edad de 65±11 años y sospecha clínica, analítica o radiológica de recurrencia. Se realizó PET/TC después de la inyección de 4,07MBq/kg de 18F-FDG con imagen de cuerpo entero a los 60min (imagen estándar) y hepática a las 2h (imagen tardía). Se efectuó análisis visual y cuantitativo mediante SUV de los hallazgos de la PET/TC. Se obtuvo confirmación de las lesiones por estudio histopatológico y/o seguimiento mínimo de 6 meses. Resultados. Se diagnosticaron metástasis hepáticas en 37/71 pacientes (79 metástasis). Un total de 38/71 pacientes mostraban enfermedad extra-hepática en forma de recidiva local (10), adenopatías abdominopélvicas (3) o mediastínicas (3), metástasis óseas (1) o pulmonares (16) y carcinomatosis (10). Se calculó la sensibilidad y especificidad para el diagnóstico de metástasis hepáticas en base a cada paciente para la imagen estándar (81 y 91%) y la imagen tardía (95 y 97%). El número de metástasis hepáticas diagnosticadas fue mayor con la imagen tardía (66/79) que con la imagen estándar (57/79). La sensibilidad y especificidad de la PET/TC en lesiones extra-hepáticas fue de 84 y 70%, contribuyendo al diagnóstico no sospechado de 5 tumores sincrónicos. Conclusiones. La PET/TC es recomendable para descartar enfermedad extra-hepática en sospecha de recidiva de cáncer colorrectal. La realización de imagen tardía mejora la sensibilidad de la PET/TC en el diagnóstico de metástasis hepáticas(AU)

Aim. To analyze the potential improvement of 18F-fluorodeoxyglucose (FDG) PET/CT using additional delayed images of the liver in operated colorectal cancer. Material and Methods. The study prospectively included 71 patients (22 women, 49 men) with mean age of 65±11 years with clinical, analytic or radiological suspicion of current disease. A whole body PET/CT scan was performed at 60min. (standard images) and after 2 hr (delayed images) post-injection of 4.07 MBq/Kg of 18F-FDG. Visual and quantitative SUV analysis of PET/CT findings was done. All findings were confirmed by histopathology and/or at least 6 months follow-up. Results. Thirty-seven out of 71 patients were diagnosed of liver metastases (79 metastases). In 38/71 cases there was extra-hepatic disease in the form of local recurrence (10), abdominopelvic (3) or mediastinal (3) lymph nodes, bone (1) or lung metastases (16) and carcinomatosis (10). Sensitivity and specificity in the diagnosis of liver metastases in a patient-by-patient basis in standard (81% and 91%) and in delayed images (95% y 97%) was calculated. The number of lesions detected in delayed images was significantly higher (66/79) than in standard images (57/79). Sensitivity and specificity for PET/CT in the diagnosis of extra-hepatic disease was 84% and 70%, contributing to the detection of synchronous tumors in 5 patients. Conclusions. PET/CT may be useful in the diagnosis of extra-hepatic disease in suspected recurrence of colorectal cancer. Delayed images on PET/CT may increase the sensitivity to identify liver metastases(AU)

Live cell imaging in live animals with fluorescent proteins.

The discovery, cloning, and characterization of GFP and related proteins of many colors have enabled live cell imaging to an unprecedented extent and resolution. Essentially, any cellular process can be imaged with a fluorescent protein. These proteins serve as genetic reporters and therefore can be used to follow cellular processes over indefinite periods in vivo as well as in vitro. The brightness and specific spectra of fluorescent proteins allow them to be imaged in vivo, using specific filters, without interference from autofluorescence. This chapter describes the development of live imaging in live animals with subcellular resolution, emphasizing the study of in vivo cell biology of cancer growth, spread, and metastasis.

Real-time imaging reveals the single steps of brain metastasis formation.

Brain metastasis frequently occurs in individuals with cancer and is often fatal. We used multiphoton laser scanning microscopy to image the single steps of metastasis formation in real time. Thus, it was possible to track the fate of individual metastasizing cancer cells in vivo in relation to blood vessels deep in the mouse brain over minutes to months. The essential steps in this model were arrest at vascular branch points, early extravasation, persistent close contacts to microvessels and perivascular growth by vessel cooption (melanoma) or early angiogenesis (lung cancer). Inefficient steps differed between the tumor types. Long-term dormancy was only observed for single perivascular cancer cells, some of which moved continuously. Vascular endothelial growth factor-A (VEGF-A) inhibition induced long-term dormancy of lung cancer micrometastases by preventing angiogenic growth to macrometastases. The ability to image the establishment of brain metastases in vivo provides new insights into their evolution and response to therapies.

Drug testing on 3D in vitro tissues trapped on a microcavity chip.

Close to realistic responses to anti-cancer drugs are not adequately provided in monolayer or single cells assays. 3-dimensional multicellular cultures (spheroids) mimicking in vivo-like conditions are established as cell biological models for microtumors/metastases. For a non-invasive real-time monitoring of the electrical parameters of such spheroid cultures we designed, fabricated and tested a 3D multifunctional electrode-based microcavity array. In a non-adherent assay acute tests with tumor spheroids were done maintaining their spherical shape and cellular arrangement. The sensor chip with 15 individual square microcavities containing four gold electrodes each was used for impedance spectroscopy to analyze the tissue models in terms of morphological and structural changes. Cell type specific differences in the spectra and varying responses to several anti-tumor drugs were found. Further development of the prototype will provide a promising tool for the use in pharmacological high-throughput studies.

Interactions between colon cancer cells and hepatocytes in rats in relation to metastasis.

Adhesion of cancer cells to endothelium is considered an essential step in metastasis. However, we have shown in a previous study that when rat colon cancer cells are administered to the vena portae, they get stuck mechanically in liver sinusoids. Then, endothelial cells retract rapidly and cancer cells bind to hepatocytes. We investigated the molecular nature of these interactions between colon cancer cells and hepatocytes. Cancer cells in coculture with hepatocytes became rapidly activated with distinct morphological changes. Cancer cells formed long cytoplasmic protrusions towards hepatocytes in their close vicinity and these protrusions attached to microvilli of hepatocytes. Then, adhering membrane areas were formed by both cell types. Integrin subunits alphav, alpha6 and beta1 but not alphaL, beta2, beta3 and CD44 and CD44v6 were expressed on the cancer cells. In conclusion, colon cancer cells show an active behaviour to bind to hepatocytes, likely involving the integrin subunits alphav, alpha6 and beta1, indicating that early events in colon cancer metastasis in liver are distinctly different than assumed thus far.

Adenopatía cervical como presentación de adenocarcinoma de próstata / Cervical adenopathy presentation of adenocarcinoma of prostate

Los reportes de metástasis de cáncer de próstata demuestran la diseminación linfática regional, siendo la metástasis a linfáticos cervicales infrecuentes y poco reportados en la literatura. En el presente trabajo aportamos una particular observación de esta entidad. Presentando un caso clínico donde se evidencia masa latero cervical izquierda, dolor abdominal y diarrea. Al realizar la biopsia de adenopatía cervical y de la lesión anal que reportaron adenocarcinoma prostático poco diferenciado. Se realizó colonoscopia en vista de lesión anal al examen físico, con biopsia cuyos resultados fueron carcinoma poco diferenciado de próstata en foco necrótico infiltrante a recto. Los estudios complementarios demostraron adenopatías para aórticas abdominales bilaterales. Las adenopatías cervicales en cuello en hombres adultos debe contemplar dentro de sus diagnósticos diferenciales el carcinoma de próstata, debido a que el adecuando tratamiento puede prolongar la sobrevida en estos pacientes

The metastases of prostate cancer shows the regional lymphatic dissemination, being the cervical lymphatic metastases to infrequent and little reported in Literature. In the present work we contributed a new observation of this organization. We displayed a clinical case with lateral mass cervical left, abdominal pain and diarrheic. When making biopsy of cervical adenopathy reported adenocarcinoma prostate metastases. Colonoscopy with biopsy was made whose results were carcinoma little differentiated infiltrated necrotic center to rectum. The complementary studies demonstrated bilateral aortic adenopathys without injuries in thorax. Within the diagnosis differential of the cervical adenopathys in neck in adult men it must consider the prostate carcinoma, because in an early diagnosis and adapting treatment it can prolong the survive

Cannibalism: a way to feed on metastatic tumors.

Cannibalism of tumors is an old story for pathologists, but it remained a mystery for at least one century. Recent data highlighted tumor cannibalism as a key advantage in tumor malignancy, possibly involved in resistance of tumors to the specific immune reaction. However, new data suggests also that metastatic tumor cells may use this peculiar function to feed in conditions of low nutrient supply. This makes malignant cancer cells more similar to microorganisms, rather than to normal cells undergoing malignant transformation. In cytological or histological samples of human tumors it is common to detect cells with one or many vacuoles, possibly containing cells under degradation, that push the nucleus to the periphery giving it the shape of a crescent moon. The cannibal cells may feed on sibling tumor cells, but also of the lymphocytes that should kill them. Cannibal cells eat everything without distinguishing between the feeding materials, with a mechanism that mostly differ from typical phagocytosis. Despite such phenomenon is considered mainly non-selective, a molecular framework of factors that contribute to cannibalism has been described. This machinery includes the presence of an acidic environment that allows a continuous activation of specific lytic enzymes, such as cathepsin B. Cannibalism occurs in apparently well defined structures whose main actors are big caveolar-like vacuoles and a connection between caveolin-1 and the actin cytoskeleton through the actin-linker molecule ezrin. Each of the components of the cannibal framework may represent specific tumor targets for future new strategies against cancer.

Angiotropic melanoma and extravascular migratory metastasis: a review.

How metastases develop is poorly understood. The concept of intravascular dissemination of cancer cells has been widely accepted as a central paradigm. In addition to this explanation, however, other mechanisms may be operable. Ultrastructural studies have identified in malignant melanoma an angio-tumoral complex, in which tumor cells are linked to endothelium by a matrix containing laminin without evidence of intravasation. This observation has suggested that melanoma cells may migrate along the external surface of vessels and other anatomic structures, a mechanism termed "extravascular migratory metastasis" (EVMM). Angiotropism (melanoma cells cuffing the external surface of vessels) is the histopathologic counterpart of the angio-tumoral complex. The authors have recently drawn attention to the importance of angiotropism as a biologic phenomenom and prognostic factor in melanoma and as a likely correlate of EVMM. In addition, recent experimental studies strongly suggest a correlation of angiotropism of melanoma cells with EVMM. These studies, including cocultures of melanoma cells with capillarylike structures in vitro and the growth of green fluorescent protein-labeled melanoma cells in the shell-less chick chorioallantoic membrane model, have demonstrated the migration of angiotropic melanoma cells along the vascular channels, supporting the concept of EVMM. The new field of EVMM reviewed in this paper may prove useful in elucidating the molecular interactions involved in melanoma metastasis.

Effects of hyperthermia in human neuroblastoma cells.

Neuroblastoma is a childhood malignancy derived from the developing sympathetic nervous system (SNS) and often diagnosed during early infancy. To investigate its metastatic properties, also in response to anti-cancer treatment, we have studied hyperthermia (HT) effects on the ultrastructure of SK-N-MC human neuroblastoma tumor cells. Cells undergoing HT showed a significant increase in apoptotic and necrotic events, but also a rearrangement of the cellular shape, appearing as cell detachment and rounding. The most striking effects of HT can be so correlated to primary tumor mass decrease and to a certain impairment of cell adhesion properties and consequently metastatic diffusion potential.

Differentiation of classic medulloblastoma into metastatic large cell medulloblastoma with focal rhabdoid differentiation in the absence of posterior fossa recurrence.

A case of classic medulloblastoma that metastasized, despite the absence of local recurrence, to extraneural sites 7 years after treatment is reported. The metastases were, in contrast to the primary tumor, of large cell type and displayed abortive myogenic and, in one site, also rhabdoid differentiation. The primary tumor expressed microtubule-associated protein 1B and neuron-specific nuclear protein (NeuN), and was desmin negative. The metastases were also positive for microtubule-associated protein 1B and NeuN, although the expression of the latter marker was weak and/or focal in two of four metastases and absent in the rhabdoid metastasis. They were, in contrast with the primary tumor, all strongly positive for desmin. The hSNF5/INI1 was expressed in the nuclei of all cells of the primary tumor and the metastases, including the one with rhabdoid differentiation. Two metastases were studied by cytogenetics. The composite karyotype of a large cell metastasis was 45~46,XY,add(1)(p36.1),t(2;8)(p21;q24.1),add(3)(q25),t(9;15)(q22;q13),add(12)(p11.2), +1approximately2mar,inc[cp12]/46,XY[12], while the rhabdoid metastasis contained additional changes including monosomy 22. These findings indicate that some rhabdoid (atypical teratoid/rhabdoid) tumors of the cerebellum and medulloblastoma may be histogenetically related.

The use of electron microscopy to refine diagnoses in the daily practice of cytopathology.

Conceived as a screening tool, cytology is a field that since the 1980s has become more diagnostic in its scope. The advent of the fine-needle aspiration biopsy (FNAB) is responsible for cytology's new place in pathology. In the everyday practice of cytopathology, about 85-90% of the nongynecologic cases can be diagnosed with the use of routine stains (i.e., Papanicolaou and Diff Quik). The other 10-15% of the cases require the use of ancillary diagnostic techniques for a precise diagnosis. Immunohistochemistry helps solve approximately 50% of these cases, and the other half of these challenging cases are best approached and diagnosed by using electron microscopy (EM). In their practice, the authors obtain cytologic samples for EM routinely in difficult cases. Unfortunately, a percentage of these cases collected for ultrastructural evaluation do not have enough cells after processing, and others only have a few diagnostic cells available. In the cases in which at least a handful of cells are available, EM is almost invariably helpful in one way or another, either making a definitive diagnosis or refining the diagnosis. A sampling of FNAB cases from the authors' everyday practice is prevented to illustrate the use of EM in the practice of cytopathology. The cases have been selected from among the most common diagnostic challenges to highlight the important role that ultrastructural evaluation plays in a busy cytology practice. In our practice ultrastructural evaluation is a piece of the puzzle, which, along with the clinical history, clinical impression, light microscopic/cytologic features, and other ancillary techniques (IHC, flow cytometry, and molecular pathology), help compile an accurate diagnosis. Many times EM is the most important component of the diagnostic algorithm.

Differential thymosin beta 10 expression levels and actin filament organization in tumor cell lines with different metastatic potential.

BACKGROUND: To investigate the differential expression levels of thymosin beta 10 (T beta 10) and the corresponding changes of actin filament organization in human tumor cell lines with different metastatic potential. METHODS: Four groups of nine human tumor cell lines with different metastatic potential were analyzed for the amount of T beta 10 mRNAs by Northern blot and for their peptide expression levels by immunohistochemistry. The filamentous actin (F-actin) was observed by staining of TRITC-phalloidin to detect changes in actin organization. RESULTS: In comparison with non-/weakly metastatic counterparts, T beta 10 was upregulated in highly metastatic human lung cancer, malignant melanoma and breast cancer cell lines. Staining of TRITC-phalloidin revealed less actin bundles, a fuzzy network of shorter filaments and some F-actin aggregates in the highly metastatic tumor cells. Meanwhile, the actin filaments were robust and orderly arranged in the non-/weakly metastatic cancer cell lines. CONCLUSION: T beta 10 levels correlate positively with the metastatic capacity in human tumors currently examined. The increasing metastatic potential of tumor cells is accompanied by a loss of F-actin, poorly arranged actin skeleton organizations and presence of F-actin aggregates. There is a consistent correlation between the elevated T beta 10 expression and the disrupted actin skeleton.

A novel concept of glomeruloid body formation in experimental cerebral metastases.

Glomeruloid bodies (GBs), tumor-associated vascular structures with a superficial resemblance to renal glomeruli, are important histopathological features of glioblastoma multiforme, but have also been described in other types of tumors and in cerebral metastases. The purpose of this study was to elucidate the pathogenesis of these lesions in an appropriate murine model of experimental brain metastases. To do so, we injected cells from 5 different tumor lines into the internal carotid artery of mice and investigated the development, composition, and fate of GBs growing within tumor nodules. Immunohistochemical analyses and 3-dimensional reconstruction of the cerebral vasculature showed clearly that the proliferating and migrating tumor cells pull the capillaries (and the adjacent capillary branching points) into the tumor cell nest. Initially, this process lead to the appearance of simple coiled vascular structures, which later developed into chaotic and tortuous vascular aggregates with multiple narrowed afferent and efferent microvessels. Despite the absence of sprouting angiogenesis, the very low level of endothelial cell proliferation index and the ruptures of the stretched and narrowed capillary segments observed frequently between the metastatic tumor nodules, necrosis was scarce in these lesions, implying that the blood supply from the multiple afferent microvessels and from the preexistent vascular bed sufficed to provide the tumor cells with oxygen and nutrients.

Metastatic balloon cell melanoma in a dog.

BACKGROUND: Balloon cell melanoma is a rare variant of amelanotic melanoma that is difficult to differentiate from sebaceous cell carcinoma, liposarcoma, and other clear cell neoplasms without immunohistochemistry or ultrastructural evidence of melanin or melanosomes. OBJECTIVE: The purpose of this report was to describe the clinical, cytologic, histologic, immunohistochemical, and ultrastructural findings in a dog with metastatic balloon cell melanoma. METHODS: A 6-year-old female Golden Retriever was evaluated for a white, flocculent infiltrate in the anterior chamber of the left eye and an enlarged left prescapular lymph node. Cytologic evaluation of the eye and lymph node were performed following aqueocentesis and fine-needle aspiration, respectively. The affected lymph node was examined histologically and stained for cytokeratin, vimentin, S-100, and Melan A. Following euthanasia a necropsy was performed and samples of the affected lymph node were examined by electron microscopy. RESULTS: Cytologic examination of the lymph node and aqueocentesis sample revealed round neoplastic cells that had abundant clear vacuolated cytoplasm. A tentative diagnosis of metastatic sebaceous cell carcinoma or clear cell neoplasm was made. Histologically, the affected lymph node had similar polygonal clear cells arranged in sheets and packets divided by delicate fibrovascular stroma. Immunohistochemical staining of the cells was negative for cytokeratin but positive for vimentin, weakly positive for S-100, and strongly positive for Melan A. At necropsy, metastatic lesions were identified in the diaphragm, heart, lung, kidneys, left eye, prescapular and sublumbar lymph nodes, and multiple skin sites. Ultrastructural examination of neoplastic lymph nodes revealed many membrane-bound vacuoles, myelin-like figures, and rare melanosomes. CONCLUSION: Immunohistochemical staining and ultrastructural features of the neoplastic cells supported a diagnosis of metastatic balloon cell melanoma.