Analysis of ovarian cancer cell secretome during epithelial to mesenchymal transition reveals a protein signature associated with advanced stages of ovarian tumors.

Ovarian cancer (OvCA) is the most lethal neoplasia among gynecologic malignancies and faces high rates of new cases particularly in South America. In special, the High Grade Serous Ovarian Carcinoma (HGSC) presents very poor prognosis with deaths caused mainly by metastasis. Among several mechanisms involved in metastasis, the Epithelial to Mesenchymal Transition (EMT) molecular reprogramming represents a model for latest stages of cancer progression. EMT promotes important cellular changes in cellular adhesion and cell-cell communication, which particularly depends on the paracrine signaling from neighbor cells. Considering the importance of cellular communication during EMT and metastasis, here we analyzed the changes in the secretome of the ovarian cancer cell line Caov-3 induced to EMT by Epidermal Growth Factor (EGF). Using a combination of GEL-LC-MS/MS and stable isotopic metabolic labelling (SILAC), we identified up-regulated candidates during EMT as a starting point to identify relevant proteins for HGSC. Based on public databases, our candidate proteins were validated and prioritized for further analysis. Importantly, several of the protein candidates were associated with cellular vesicles, which are important to the cell-cell communication and metastasis. Furthermore, the association of candidate proteins with gene expression data uncovered a subset of proteins correlated with the mesenchymal subtype of ovarian cancer. Based on this relevant molecular signature for aggressive ovarian cancer, supported by protein and gene expression data, we developed a targeted proteomic method to evaluate individual OvCA clinical samples. The quantitative information obtained for 33 peptides, representative of 18 proteins, was able to segregate HGSC from other tumor types. Our study highlighted the richness of the secretome and EMT to reveal relevant proteins for HGSC, which could be used in further studies and larger patient cohorts as a potential stratification signature for ovarian cancer tumor that could guide clinical conduct for patient treatment.

Comparative study of quercetin and its two glycoside derivatives quercitrin and rutin against methylmercury (MeHg)-induced ROS production in rat brain slices.

The hypothesis that methylmercury (MeHg) potently induces formation of reactive oxygen species (ROS) in the brain is supported by observations on the neuroprotective effects of various classes of antioxidants. Flavonoids have been reported to possess divalent metal chelating properties, antioxidant activities and to readily permeate the blood-brain barrier. They can also provide neuroprotection in a wide array of cellular and animal models of neurological diseases. Paradoxically, in vivo administration of quercetin displays unexpected synergistic neurotoxic effect with MeHg. Considering this controversy and the limited data on the interaction of MeHg with other flavonoids, the potential protective effect of quercetin and two of its glycoside analogs (i.e., rutin and quercitrin) against MeHg toxicity were evaluated in rat cortical brain slices. MeHg (100 microM) caused lipid peroxidation and ROS generation. Quercitrin (10 microg/mL) and quercetin (10 microg/mL) protected mitochondria from MeHg (5 microM)-induced changes. In contrast, rutin did not afford a significant protective effect against MeHg (100 microM)-induced lipid peroxidation and ROS production in cortical brain slices. MeHg-generated ROS in cortical slices was dependent upon an increase in intracellular Ca(2+) levels, because the over-production of MeHg-induced H(2)O(2) in mitochondria occurred with a concomitant increase in Ca(2+) transient. Here, we have extended the characterization of mechanisms associated with the neuroprotective effects of quercetin against MeHg-induced toxicity in isolated mitochondria, by performing an array of parallel studies in brain slices. We provide novel data establishing that (1) Ca(2+) plays a central role in MeHg toxicity and (2) in brain slices MeHg induces mitochondrial oxidative stress both via direct interaction with mitochondria (as previously reported in in vitro studies) as well as via mitochondria-independent (or indirect) mechanisms.