Posterior corneal surface stability after femtosecond laser-assisted keratomileusis

The purpose of this study was to evaluate posterior corneal surface variation after femtosecond laser-assisted keratomileusis in patients with myopia and myopic astigmatism. Patients were evaluated by corneal tomography preoperatively and at 1, 6, and 12 months. We analyzed changes in the posterior corneal curvature, posterior corneal elevation, and anterior chamber depth. Moreover, we explored correlation between corneal ablation depth, residual corneal thickness, percentage of ablated corneal tissue, and preoperative corneal thickness. During follow-up, the posterior corneal surface did not have a significant forward corneal shift: no significant linear relationships emerged between the anterior displacement of the posterior corneal surface and corneal ablation depth, residual corneal thickness, or percentage of ablated corneal tissue

Investigating alcohol consumption during pregnancy for the prevention of Fetal Alcohol Spectrum Disorders (FASD)

The term FASD (Fetal Alcohol Spectrum Disorders) is used to describe the entire spectrum of pathologies and disorders caused by alcohol exposure in uterus. Alcohol assumed in pregnancy passes directly through the placental barrier causing a broad range of symptoms whose severity can greatly vary in degree. The alcohol teratogenic effect may result in physical damage and specific facial anomalies, growth delays, neurological defects along with intellectual disabilities and behavioral problems. Children affected show difficulties in verbal learning, memory, visual-spatial abilities, attention, logic and math abilities, information processing, executive functions as well as in many other domains and in general coping with daily life. Total abstention from alcohol during pregnancy is strongly recommended, as a safe threshold of consumption has not been established yet. Hence, the early identification of alcohol consumption in pregnancy is crucial. Specific methodologies to overcome difficulties related to the identification of alcohol behavior in pregnant women are needed and intervention protocols should be implemented to prevent damage in offsprings. This paper gives an overview on this pathology, from clinical delineation to epidemiology and risk factors with a special focus to promote alcohol-free pregnanc

A short story of 3AB-OS cancer stem cells, a possible model for studying cancer stemness

Cancer Stem Cells (CSCs) are thought to be the cause of cancer initiation, growth and development. Thus, a challenge in cancer research is their identification and eradication. In our laboratory, by chemical treatment of the human osteosarcoma (OS) MG63 cell line, we have isolated and characterized 3AB-OS cells, a human OS CSC line. 3AB-OS cells transdifferentiate in vitro into cells of the three derivatives germ layers and, when xenografted in athymic mice they are highly tumorigenic and recapitulate in vivo crucial features of human OS. They even express a reprogrammed energy metabolism, with a dependence on glycolytic metabolism more strong than parental MG63 cells. 3AB-OS cells have chromosomes showing a great number of abnormalities which are very similar to abnormalities found in both pediatric and adult osteosarcomas. In comparison with parental MG63 cells (where TP53 gene is hypermethylated, rearranged and in single copy), 3AB-OS cells have TP53 gene unmethylated, rearranged and in multiple copies. Moreover, the mutp53 (p53-R248W/P72R) is post-translationally stabilized, has nuclear localization and a gain of function. A great number of results obtained in our laboratories suggested that p53 mutation could be the “driver mutation” at the origin of the transformation of MG63 cells into 3AB-OS CSCs

Parthenolide induces caspase-independent and AIF-mediated cell death in human osteosarcoma and melanoma cells.

The mechanism of the cytotoxic effect exerted by parthenolide on tumor cells is not clearly defined today. This article shows that parthenolide stimulates in human osteosarcoma MG63 and melanoma SK-MEL-28 cells a mechanism of cell death, which is not prevented by z-VAD-fmk and other caspase inhibitors. In particular treatment with parthenolide rapidly stimulated (1-2 h) reactive oxygen species (ROS) generation by inducing activation of extracellular signal-regulated kinase 1/2 (ERK 1/2) and NADPH oxidase. This event caused depletion of thiol groups and glutathione, NF-κB inhibition, c-Jun N-terminal kinase (JNK) activation, cell detachment from the matrix, and cellular shrinkage. The increase of ROS generation together with the mitochondrial accumulation of Ca(2+) also favored dissipation of Δψm, which seemed primarily determined by permeability transition pore opening, since Δψm loss was partially prevented by the inhibitor cyclosporin A. Staining with Hoechst 33342 revealed in most cells, at 3-5 h of treatment, chromatin condensation, and fragmentation, while only few cells were propidium iodide (PI)-positive. In addition, at this stage apoptosis inducing factor (AIF) translocated to the nucleus and co-localized with areas of condensed chromatin. Prolonging the treatment (5-15 h) ATP content declined while PI-positive cells strongly augmented, denouncing the increase of necrotic effects. All these effects were prevented by N-acetylcysteine, while caspase inhibitors were ineffective. We suggest that AIF exerts a crucial role in parthenolide action. In accordance, down-regulation of AIF markedly inhibited parthenolide effect on the production of cells with apoptotic or necrotic signs. Taken together our results demonstrate that parthenolide causes in the two cell lines a caspase-independent cell death, which is mediated by AIF

Single low-dose cyclophosphamide combined with interleukin-12 gene therapy is superior to a metronomic schedule in inducing immunity against colorectal carcinoma in mice

The use of conventional cytotoxic agents at metronomic schedules, alone or in combination with targeted agents or immunotherapy, is being explored as a promising anticancer strategy. We previously reported a potent antitumor effect of a single low-dose cyclophosphamide and interleukin-12 (IL-12) gene therapy against advanced gastrointestinal carcinoma, in mice. Here, we assessed whether the delivery of IL-12 by gene therapy together with metronomic cyclophosphamide exerts antitumor effects in a murine model of colorectal carcinoma. This combination therapy was able, at least in part, to reverse immunosuppression, by decreasing the number of regulatory T cells (Tregs) as well as of splenic myeloid-derived suppressor cells (MDSC s). However, metronomic cyclophosphamide plus IL-12 gene therapy failed to increase the number of tumor-infiltrating T lymphocytes and, more importantly, to induce a specific antitumor immune response. With respect to this, cyclophosphamide at a single low dose displayed a superior anticancer profile than the same drug given at a metronomic schedule. Our results may have important implications in the design of new therapeutic strategies against colorectal carcinoma using cyclophosphamide in combination with immunotherapy.Fil: Malvicini, Mariana. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad Austral; ArgentinaFil: Alaniz, Laura Daniela. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad Austral; ArgentinaFil: Bayo Fina, Juan Miguel. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad Austral; ArgentinaFil: García, Mariana Gabriela. Universidad Austral; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Piccioni, Flavia Valeria. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad Austral; ArgentinaFil: Fiore, Esteban Juan. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad Austral; ArgentinaFil: Atorrasagasti, María Catalina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad Austral; ArgentinaFil: Aquino, Jorge Benjamin. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad Austral; ArgentinaFil: Matar, Pablo. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad Nacional de Rosario. Facultad de Ciencias Medicas. Instituto de Genetica Experimental; ArgentinaFil: Mazzolini Rizzo, Guillermo Daniel. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad Austral; Argentin

Parthenolide induces superoxide anion production by stimulating EGF receptor in MDA-MB-231 breast cancer cells.

The sesquiterpene lactone parthenolide (PN) has recently attracted considerable attention because of its anti-microbial, anti-inflammatory and anticancer effects. However, the mechanism of its cytotoxic action on tumor cells remains scarcely defined. We recently provided evidence that the effect exerted by PN in MDA-MB-231 breast cancer cells was mediated by the production of reactive oxygen species (ROS). The present study shows that PN promoted the phosphorylation of EGF receptor (phospho-EGFR) at Tyr1173, an event which was observed already at 1  h of incubation with 25  µM PN and reached a peak at 8-16  h. This effect seemed to be a consequence of ROS production, because N-acetylcysteine (NAC), a powerful ROS scavenger, prevented the increment of phospho-EGFR levels. In addition fluorescence analyses performed using dihydroethidium demonstrated that PN stimulated the production of superoxide anion already at 2-3  h of incubation and the effect further increased prolonging the time of treatment, reaching a peak at 8-16  h. Superoxide anion production was markedly hampered by apocynin, a well known NADPH oxidase (NOX) inhibitor, suggesting that the effect was dependent on NOX activity. The finding that AG1478, an EGFR kinase inhibitor, substantially blocked both EGFR phosphorylation and superoxide anion production strongly suggested that phosphorylation of EGFR can be responsible for the activation of NOX with the consequent production of superoxide anion. Therefore, EGFR phosphorylation can exert a key role in the production of superoxide anion and ROS induced by PN in MDA-MB-231 cells

Nanostructures for SERS in living cell

Surface-enhanced Raman spectroscopy (SERS) has received renewed interest in recent years in fields such as trace analysis, biorelated diagnosis, and living cell study. However, the interference of impurities left on the surface from the preparation process of substrates limits to some extent the application of SERS. In the present paper, we propose a method to prepare clean SERS substrates by a combined method of hydrothermal green synthesis and thermal treatment to obtain a clean and impurity-free surface for SERS measurements, suitable for cells growth. The goal of such activity was the study of the membrane proteome, with special attention to prion protein (PrPC), in its physiological ambient. SERS has been used to evidence the PrPC-Cu(II) interaction in a rat neuroblastoma cell line (B104), known to overexpress the cellular prion protein PrPC

The oxygen radicals involved in the toxicity induced by parthenolide in MDA-MB-231 cells

It has been shown that the sesquiterpene lactone parthenolide lowers the viability of MDA-MB-231 breast cancer cells, in correlation with oxidative stress. The present report examined the different radical species produced during parthenolide treatment and their possible role in the toxicity caused by the drug. Time course experiments showed that in the first phase of treatment (0-8 h), and in particular in the first 3 h, parthenolide induced dichlorofluorescein (DCF) signal in a large percentage of cells, while dihydroethidium (DHE) signal was not stimulated. Since the effect on DCF signal was suppressed by apocynin and diphenyleneiodonium (DPI), two inhibitors of NADPH oxidase (NOX), we suggest that parthenolide rapidly stimulated NOX activity with production of superoxide anion (O2•-), which was converted by superoxide dismutase 1 (SOD1) into hydrogen peroxide (H2O2). In the second phase of treatment (8-16 h), parthenolide increased the number of positive cells to DHE signal. Since this event was not prevented by apocynin and DPI and was associated with positivity of cells to MitoSox Red, a fluorochrome used to detect mitochondrial production of O2•-, we suggest that parthenolide induced production of O2•- at the mitochondrial level independently by NOX activity in the second phase of treatment. Finally, in this phase, most cells became positive to hydroxyphenyl fluorescein (HPF) signal, a fluorescent probe to detect highly reactive oxygen species (hROS), such as hydroxyl radical and peroxynitrite. Therefore, parthenolide between 8-16 h of treatment induced generation of O2•- and hROS, in close correlation with a marked reduction in cell viability