INDUCED PLURIPOTENT STEM CELLS: AN INNOVATIVE TOOL TO DISSECT OVARIAN CANCER PATHOGENESIS

Ovarian cancer (OC) has one of the highest death-to-incidence ratios among all tumor types, which points to the need for novel therapeutic and prognostic strategies. Indeed, the absence of relevant tumor cell lines that can recapitulate disease histopathology highlights an acute need for new model systems to study this pathology. In particular, it is still unclear whether the most common and aggressive form of this disease, high grade serous ovarian cancer (HGSOC), could arise from in the ovarian surface epithelium (OSE), as initially thought, or might be arising from the fimbrial epithelium. Here I addressed these issues in two complementary ways based on induced pluripotent stem cells: i) the modeling of Ovarian Cancer by somatic cell reprogramming to pluripotency of tumor cells; ii) the molecular characterization of HGSOC and its putative cells of origin. Somatic cell reprogramming, by erasing tumor-associated epigenetic marks while preserving the underlying genetic mutations, would allow for the first time the precise dissection of genetic and epigenetic contribution to this disease, through the differentiation of OC-iPSC into disease-relevant cell types. I demonstrated the feasibility of OC reprogramming through a non-integrative platform, showing that OC-derived iPSC are closely similar to human ESC, and proving their tumoral origin by whole exome sequencing. Moreover, I showed that independent iPSC clones derived from the same tumor upon trilineage differentiation in vivo show differential tumorigenic potential. For a more precise dissection of this phenotype, I set up a differentiation protocol that allows differentiation of pluripotent cells into mesodermal progenitors, that are precursors of both fimbria and OSE. To isolate a pure population of these cells, I resorted to CRISPR/Cas9 to integrate a selection cassette in the MIXL1 locus. By this approach, I was able to show correct gene targeting at the intended site, allowing also for selection of mesodermal progenitors upon differentiation of normal iPSC. The same approach translated to OC-derived iPSC would allow to study the effects of genetic mutations deprived of tumor-associated epigenetic marks during differentiation, both at the stage of mesodermal progenitors and in cells directed towards the female reproductive epithelium in vivo. The second approach relies on the identification of specific molecular features of fimbria and ovarian surface epithelium, the two putative cells of origin of HGSOC. On this side, I offer a first glimpse on molecular features of HGSOC cancer and normal gynecological tissues. I could show that specific DNA methylation signatures of fimbrial epithelial cells and ovarian surface epithelium cells are partially retained in tumor samples and stratify HGSOC samples according to the putative cell of origin of this tumor. Moreover, I show for the first time a description of histone modifications in primary HGSOC, concentrating on marks of activation/repression sitting on promoter regions (H3K4me3 and H3K27me3, respectively) and marks that characterize active/closed-poised enhancers (H3K4me1, H3K27ac and H3K27me3)

Food ingredients and supplements : is this the future?

The concept of adequate nutrition has been changing in the last years in light of the increased knowledge about the relationship between diet and health and the new evidence on the protective role of numerous bioactive compounds introduced with specific categories of foods. In particular, the concept of adequate nutrition has been substituted with that of optimal nutrition, in other terms, attention has been focused on nutritional needs to optimize physiological functions and promote health, minimizing the risk of development of degenerative diseases. Moreover, it is always more clear that nutritional needs can vary depending on the sub-groups of population considered and also on factors related to individual genetic characteristics, as supported by several nutrigenetic and nutrigenomic studies

Anthocyanins and metabolites resolve TNF-α-mediated production of E-selectin and adhesion of monocytes to endothelial cells

This study investigated the capacity of an anthocyanin-rich fraction (ACN-RF) from blueberry, single anthocyanins (cyanidin, delphinidin and malvidin-3-glucoside; Cy, Dp and Mv-3-glc) and related metabolites (protocatechuic, gallic and syringic acid; PrA, GA and SA) to resolve an inflammation-driven adhesion of monocytes (THP-1) on endothelial cell (HUVECs) and secretion of cell adhesion molecules E-selectin and vascular cell adhesion molecule 1 (VCAM-1). The adhesion of THP-1 to HUVECs was induced by tumour necrosis factor \u3b1 (TNF-\u3b1, 100\u202fng\u202fmL-1). Subsequently, ACN-RF, single ACNs and metabolites (from 0.01 to 10\u202f\u3bcg\u202fmL-1) were incubated for 24\u202fh. The adhesion was measured in a fluorescence spectrophotometer. E-selectin and VCAM-1 were quantified by ELISA. No toxicological effects were observed for the compounds and the doses tested. ACN-RF and Mv-3-glc reducedTHP-1 adhesion at all the concentrations with the maximum effect at 10 \u3bcg/ml (-60.2% for ACNs and-33.9% for Mv-3-glc). Cy-3-glc decreased the adhesion by about 41.8% at 10\u202f\u3bcg\u202fmL-1, while PrA and GA reduced the adhesion of THP-1 to HUVECs both at 1 and at 10\u202f\u3bcg\u202fmL-1 (-29.5% and -44.3% for PrA, respectively, and -18.0%and -59.3% for GA, respectively). At the same concentrations a significant reduction of E-selectin, but notVCAM-1 levels, was documented. No effect was observed following Dp-3-glc and SA supplementation. Overall, ACNs and metabolites seem to resolve, in a dose-dependent manner, the inflammation-driven adhesion of THP-1 to HUVECs by decreasing E-selectin concentrations. Interestingly, Mv-3-glc was active at physiologically relevant concentrations

The central role of iron in human nutrition : from folk to contemporary medicine

Iron is a fundamental element in human history, from the dawn of civilization to contemporary days. The ancients used the metal to shape tools, to forge weapons, and even as a dietary supplement. This last indication has been handed down until today, when martial therapy is considered fundamental to correct deficiency states of anemia. The improvement of the martial status is mainly targeted with dietary supplements that often couple diverse co-factors, but other methods are available, such as parenteral preparations, dietary interventions, or real-world approaches. The oral absorption of this metal occurs in the duodenum and is highly dependent upon its oxidation state, with many absorption influencers possibly interfering with the intestinal uptake. Bone marrow and spleen represent the initial and ultimate step of iron metabolism, respectively, and the most part of body iron circulates bound to specific proteins and mainly serves to synthesize hemoglobin for new red blood cells. Whatever the martial status is, today\u2019s knowledge about iron biochemistry allows us to embrace exceedingly personalized interventions, which however owe their success to the mythical and historical events that always accompanied this metal

Effect of fiber and protein-enriched pasta formulations on satiety-related sensations and afternoon snacking in Italian healthy female subjects

The objective of the study was to investigate the effect of consuming different fiber and protein-enriched pasta formulations on satiety response and on mid-afternoon energy intake. Twenty Italian young healthy female subjects participated to a randomized repeated measure study design developed to evaluate the effect on satiety and energy intake of five different pasta formulations, i.e. high fiber, high fiber and high protein, high protein from soy, high protein from egg white, and standard commercial pasta consumed at lunch. The formulations together with a portion of fruit were consumed on five different occasions followed by an ad libitum snack meal proposed 2 h later. Before, immediately after the lunch consumption, and every 30 min until snack time, satiety sensations were assessed by visual analogue scales. In addition, mid-afternoon energy and macronutrient intake consumed with the snacks were calculated. Compared to the control pasta, all the formulations significantly affected satiety-related sensations. Palatability-related attributes of pasta were positively correlated to snack energy intake, whereas fullness sensation was negatively correlated. Among the formulations tested only the fiber and protein-enriched pasta significantly reduced energy intake following the ad libitum snack consumption (p < 0.05). Overall, these findings suggest that pasta enriched with a combination of fiber and protein, might be effective in the modulation of appetite sensations, thus suggesting a new concept-pasta formulation for the modulation of eating behavior. These results are interesting considering that pasta is a staple food in different target groups of the population

Tracking of Normal and Malignant Progenitor Cell Cycle Transit in a Defined Niche.

While implicated in therapeutic resistance, malignant progenitor cell cycle kinetics have been difficult to quantify in real-time. We developed an efficient lentiviral bicistronic fluorescent, ubiquitination-based cell cycle indicator reporter (Fucci2BL) to image live single progenitors on a defined niche coupled with cell cycle gene expression analysis. We have identified key differences in cell cycle regulatory gene expression and transit times between normal and chronic myeloid leukemia progenitors that may inform cancer stem cell eradication strategies

Intra and inter-day repeatability of peripheral arterial function: suitability and potential limitations

The present study aimed to investigate the inter- and intraday repeatability of reactive hyperemia index (RHI) measured by Endo-PAT in healthy volunteers. Interday RHI repeatability was tested in two consecutive days in a group of thirty-one male subjects. Intraday repeatability was investigated at baseline and after 2 and 4 h in a group of sixteen volunteers. Data were evaluated by analysis of variance. Bland-Altman plot, coefficient of variation (CV), coefficient of repeatability (CR) and intraclass correlation coefficient (ICC) were measured. While interday RHI repeatability was found to be reliable (CV: 6.0%; CR: 0.51; ICC: 0.77), multiple evaluations within the same day significantly (p<0.001) affected RHI (repeatability of the measurement - CV: 18.8%; CR: 1.26; ICC: 0.48). In particular, a significant increase in RHI occurred at 4 h compared to 2 h (+16.8%; p<0.05) and to baseline (+30.1%; p<0.05). In conclusion, RHI showed good interday but poor intraday repeatability. Multiple evaluations increased RHI especially in subjects with endothelial dysfunction who improved or reversed their impairment. These results show the potential limitations of multiple Endo-PAT measurements within the same day and the importance of standardizing the protocols before RHI evaluations