Simvastatin interferes with cancer 'stem-cell' plasticity reducing metastasis in ovarian cancer.

Cell plasticity of 'stem-like' cancer-initiating cells (CICs) is a hallmark of cancer, allowing metastasis and cancer progression. Here, we studied whether simvastatin, a lipophilic statin, could impair the metastatic potential of CICs in high-grade serous ovarian cancer (HGS-ovC), the most lethal among the gynecologic malignancies. qPCR, immunoblotting and immunohistochemistry were used to assess simvastatin effects on proteins involved in stemness and epithelial-mesenchymal cell plasticity (EMT). Its effects on tumor growth and metastasis were evaluated using different models (e.g., spheroid formation and migration assays, matrigel invasion assays, 3D-mesomimetic models and cancer xenografts). We explored also the clinical benefit of statins by comparing survival outcomes among statin users vs non-users. Herein, we demonstrated that simvastatin modifies the stemness and EMT marker expression patterns (both in mRNA and protein levels) and severely impairs the spheroid assembly of CICs. Consequently, CICs become less metastatic in 3D-mesomimetic models and show fewer ascites/tumor burden in HGS-ovC xenografts. The principal mechanism behind statin-mediated effects involves the inactivation of the Hippo/YAP/RhoA pathway in a mevalonate synthesis-dependent manner. From a clinical perspective, statin users seem to experience better survival and quality of life when compared with non-users. Considering the high cost and the low response rates obtained with many of the current therapies, the use of orally or intraperitoneally administered simvastatin offers a cost/effective and safe alternative to treat and potentially prevent recurrent HGS-ovCs.

A cytosolic source of calcium unveiled by hydrogen peroxide with relevance for epithelial cell death.

Oxidative stress releases intracellular calcium, which plays a pathogenic role in mammalian cell death. Here we report a search for the source of oxidative calcium in HeLa cells based on confocal epifluorescence microscopy. H(2)O(2) caused a rapid increase in cytosolic calcium, which was followed by mitochondrial Ca(2+) loading. Combined mitochondrial uncoupling with full depletion of thapsigargin-sensitive stores abrogated inositol 1,4,5-trisphosphate-mediated calcium release but failed to inhibit H(2)O(2)-induced calcium release, observation that was confirmed in MDCK cells. Prevention of peroxide-induced acidification with a pH clamp was also ineffective, discarding a role for endosomal/lysosomal Ca(2+)/H(+) exchange. Lysosomal integrity was not affected by H(2)O(2). Mature human erythrocytes also reacted to peroxide by releasing intracellular calcium, thus directly demonstrating the cytosolic source. Glutathione depletion markedly sensitized cells to H(2)O(2), an effect opposite to that achieved by DTT. Iron chelation was ineffective. In summary, our results show the existence of a previously unrecognized sulfhydryl-sensitive source of pathogenic calcium in the cytosol of mammalian cells.

Resistencia microtraccional de capa de adhesivo contaminada con sangre / Microtensile bond strength of adhesive layer contaminated with blood on human sound enamel

Objetivo: Evaluar la influencia de la contaminación con sangre en una capa de adhesivo formada sobre esmalte humano y su posterior descontaminación con NaOCl (2.5 por ciento) y etanol (70°) en la resistencia microtraccional. Métodos: La superficie vestibular de 80 premolares humanos fue fresada para obtener superficies planas sobre las que se aplicó un adhesivo de grabado y lavado siguiendo las indicaciones del fabricante (Adper Single Bond 2, 3M ESPE). Los premolares fueron distribuidos aleatoriamente en cuatro grupos: Grupo 1 (control), Grupo 2 (contaminación con sangre), Grupo 3 (descontaminación con NaOCl 2.5 por ciento) y Grupo 4 (descontaminación con etanol 70°). Luego, sobre cada premolar se confeccionó una corona de resina compuesta (Filtek Z350, 3M ESPE) y fueron cortados para obtener cuerpos de prueba de 1mm2 de sección transversal, los cuales fueron termociclados (5500 ciclos, 5-55°C) y traccionados hasta su límite de ruptura (Micro Tensile Tester, Bisco). Los resultados fueron analizados estadísticamente (ANOVA, Scheffe, p<0.05). Resultados: La resistencia microtraccional del Grupo 1 (24.8 MPa) fue significativamente superior al resto de los grupos (p<0.05). Las diferencias entre los grupos 2, 3 y 4 no fueron estadísticamente significativas (p>0.05). Conclusión: La contaminación con sangre de la capa de adhesivo interfiere significativamente en la resistencia microtraccional. La descontaminación con NaOCl o etanol no logró una recuperación de la resistencia microtraccional.

Aim: To evaluate the influence of blood contamination of the adhesive layer and posterior decontamination with NaOCl (2.5 percent and ethanol (70º) on microtensile bond strength on human enamel. Methods: Vestibular surfaces of eighty human premolars were grounded to obtain flat surfaces. An etch-and-rinse adhesive was applied according to the manufacturer’s instructions (Adper Single bond 2, 3M ESPE). Teeth were randomly assigned into 4 groups: Group 1 (control), Group 2 (blood contamination), Group 3 (decontamination with NaOCl 2.5 percent) and Group 4 (decontamination with ethanol 70º). Then, a 4mm composite crown (Filtek Z350) was made and the teeth were vertically sectioned to obtain 1mm cross-section stick shape specimens. Specimens were thermocycled (5500 cycles, 5-55ºC) and pulled under tension until failure (Micro Tensile Tester, Bisco). Results were statistically analyzed (ANOVA, Scheffe’s test, p<0.05). Results: Microtensile bond strength in group 1 (24.8 MPa) was significantly higher than groups 2, 3 and 4 (p<0.05). The differences between groups 2, 3 and 4 were not significant (p>0.05). Conclusion: Blood contamination significantly interferes in microtensile bond strength. Decontaminating the blood residues with NaOCl or ethanol did not show a recovery of bond strength.

Regulation of adenosine transport by D-glucose in human fetal endothelial cells: involvement of nitric oxide, protein kinase C and mitogen-activated protein kinase.

The effects of elevated D-glucose on adenosine transport were investigated in human cultured umbilical vein endothelial cells isolated from normal pregnancies. Elevated D-glucose resulted in a time- (8-12 h) and concentration-dependent (half-maximal at 10+/-2 mM) inhibition of adenosine transport, which was associated with a reduction in the Vmax for nitrobenzylthioinosine (NBMPR)-sensitive (es) saturable nucleoside with no significant change in Km. d-Fructose (25 mM), 2-deoxy-D-glucose (25 mM) or D-mannitol (20 mM) had no effect on adenosine transport. Adenosine transport was inhibited following incubation of cells with the protein kinase C (PKC) activator phorbol 12-myristate 13-acetate (PMA; 100 nM, 30 min to 24 h). D-Glucose-induced inhibition of transport was abolished by calphostin C (100 nM, an inhibitor of PKC), and was not further reduced by PMA. Increased PKC activity in the membrane (particulate) fraction of endothelial cells exposed to D-glucose or PMA was blocked by calphostin C but was unaffected by NG-nitro-L-arginine methyl ester (L-NAME; 100 microM, an inhibitor of nitric oxide synthase (NOS)) or PD-98059 (10 microM, an inhibitor of mitogen-activated protein kinase kinase 1). D-Glucose and PMA increased endothelial NOS (eNOS) activity, which was prevented by calphostin C or omission of extracellular Ca2+ and unaffected by PD-98059. Adenosine transport was inhibited by S-nitroso-N-acetyl-l, d-penicillamine (SNAP; 100 microM, an NO donor) but was increased in cells incubated with L-NAME. The effect of SNAP on adenosine transport was abolished by PD-98059. Phosphorylation of mitogen-activated protein kinases p44mapk (ERK1) and p42mapk (ERK2) was increased in endothelial cells exposed to elevated D-glucose (25 mM for 30 min to 24 h) and the NO donor SNAP (100 microM, 30 min). The effect of D-glucose was blocked by PD-98059 or L-NAME, which also prevented the inhibition of adenosine transport mediated by elevated D-glucose. Our findings provide evidence that D-glucose inhibits adenosine transport in human fetal endothelial cells by a mechanism that involves activation of PKC, leading to increased NO levels and p42-p44mapk phosphorylation. Thus, the biological actions of adenosine appear to be altered under conditions of sustained hyperglycaemia.

Elevated expression of glucose transporter-1 in hypothalamic ependymal cells not involved in the formation of the brain-cerebrospinal fluid barrier.

Glucose transporters play an essential role in the acquisition of glucose by the brain. Elevated expression of glucose transporter-1 has been detected in endothelial cells of the blood-brain barrier and in choroid plexus cells of the blood-cerebrospinal fluid barrier. On the other hand, there is a paucity of information on the expression of glucose transporters in the ependymal cells that line the walls of the cerebral ventricles. The tanycytes are specialized ependymal cells localized in circumventricular organs such as the median eminence that can be segregated into at least three types, alpha, beta1 and beta2. The beta2 tanycytes form tight junctions and participate in the formation of the cerebrospinal fluid-median eminence barrier. Using immunocytochemistry and in situ hybridization, we analyzed the expression of hexose transporters in rat and mouse hypothalamic tanycytes. In both species, immunocytochemical analysis revealed elevated expression of glucose transporter-1 in alpha and beta1 tanycytes. Intense anti-glucose transporter-1 staining was observed in cell processes located throughout the arcuate nucleus, in the end-feet reaching the lateral sulcus of the infundibular region, and in cell processes contacting the hypothalamic capillaries. On the other hand, there was very low expression of glucose transporter-1 in beta2 tanycytes involved in barrier function. In contrast with the results of the cytochemical analysis, in situ hybridization revealed that tanycytes alpha, beta1, and beta2 express similar levels of glucose transporter-1 mRNA. Further analysis using anti-glial fibrillary acidic protein antibodies to identify areas rich in astrocytes revealed that astrocytes were absent from areas containing alpha and beta1 tanycytes, but were abundant in regions containing the barrier-forming beta2 tanycytes. Overall, our data reveal a lack of correlation between participation in barrier function and expression of glucose transporter-1 in hypothalamic tanycytes. Given the virtual absence of astrocytes in areas rich in alpha and beta1 tanycytes, we speculate whether the tanycytes might have astrocyte-like functions and participate in the metabolic coupling between glia and neurons in the hypothalamic area.

Virus de la hepatitis C en un grupo de pacientes hematológicos y oncohematológicos / Hepatitis C virus in a group of hematological and oncohematological patients

Background: Little information is available in Chile about hepatitis C virus (HCV) in hematological and oncohematological patients. Aim: To evaluate the prevalence of hepatitis C virus markers in a group of hematological and oncohematological pediatric patients seen at Valdivia Regional Hospital. Patients and methods: Antibodies against virus C, determined by ELISA and viral RNA, determined using RT-polymerase chain reaction, were measured in 54 blood samples from children with hematological diseases (34 with Acute Lymphoblastic Leukaemia, 4 with Hodgkin Diseases, 4 with Haemolytic Anemia, 5 with Sarcomas, 2 with Non-Hodgkin Lymphoma, 2 with Thrombocytopenic Purpura, 1 with an Ependimoma, one with a Wilms Tumor and 1 with a Von Willebrand Disease). Results: All samples were negative for antibodies against hepatitis C virus. Viral RNA was found in four children, all with a diagnosis of acute lymphoblastic leukemia and who received chemotherapy and multiple transfusions. Conclusions: The prevalence of Viral RNA for hepatitis C virus in oncohematological patients in our study is high and associated with the use of chemotherapy and multiple transfusions

Estudio descriptivo de brote de hepatitis viral aguda A, en comuna de Empedrado, VII región, enero-junio 1997 / Descriptive study of acute viral hepatitis A, at Empedrado, VII region, january-june 1997

Con el objetivo de conocer la realidad ocurrida en el brote de Hepatitis presentado en Empedrado, se llevó a cabo un estudio descriptivo con un universo de los casos. Se encontraron 64 casos mediante el diagnóstico clínico y de laboratorio en el consultorio de Empedrado; Se analizaron las variables edad, sexo, ruralidad, higiene ambiental (eliminación de excretas, presencia de agua potable e higiene personal), número de casos que presentan Hepatitis Aguda Fulminante, recaída de la enfermedad y los que desarrollan enfermedad a pesar de haber sido inmunizados. Se encontró un predominio de casos femeninos, entre 0 a 9 años, de procedencia urbana, que contaban con agua potable, con un deficiente higiene personal y eliminación de excretas en pozo negro. De los Contactos vacunados, la enfermedad se desarrolló e un bajo porcentaje. Del total de los casos hubo uno que evolucinó a Hepatitis Aguda Fulminante y otro que presentó recaída de la enfermedad