Isolation of single circulating trophoblasts from maternal circulation for noninvasive fetal copy number variant profiling

ObjectiveTo develop a multi-step workflow for the isolation of circulating extravillous trophoblasts (cEVTs) by describing the key steps enabling a semi-automated process, including a proprietary algorithm for fetal cell origin genetic confirmation and copy number variant (CNV) detection. MethodsDetermination of the limit of detection (LoD) for submicroscopic CNV was performed by serial experiments with genomic DNA and single cells from Coriell cell line biobank with known imbalances of different sizes. A pregnancy population of 372 women was prospectively enrolled and blindly analyzed to evaluate the current workflow. ResultsAn LoD of 800 Kb was demonstrated with Coriell cell lines. This level of resolution was confirmed in the clinical cohort with the identification of a pathogenic CNV of 800 Kb, also detected by chromosomal microarray. The mean number of recovered cEVTs was 3.5 cells per sample with a significant reverse linear trend between gestational age and cEVT recovery rate and number of recovered cEVTs. In twin pregnanices, evaluation of zygosity, fetal sex and copy number profiling was performed in each individual cell. ConclusionOur semi-automated methodology for the isolation and single-cell analysis of cEVTS supports the feasibility of a cell-based noninvasive prenatal test for fetal genomic profiling. © 2022 A. Menarini Biomarkers Singapore Pte Ltd. Prenatal Diagnosis published by John Wiley & Sons Ltd

Stemming Cancer: Functional Genomics of Cancer Stem Cells in Solid Tumors

Cancer stem cells (CSCs) were discovered about 15 years ago in hematopoietic cancers. Subsequently, cancer stem cells were discovered in various solid tumors. Based on parallels with normal stem cells, a developmental process of cancer stem cells follows paths of organized, hierarchical structure of cells with different degrees of maturity. While some investigators have reported particular markers as identification of cancer stem cells, these markers require further research. In this review, we focus on the functional genomics of cancer stem cells. Functional genomics provides useful information on the signaling pathways which are consecutively activated or inactivated amongst those cells. This information is of particular importance for cancer research and clinical treatment in many respects. (1) Understanding of self-renewal mechanisms crucial to tumor growth. (2) Allow the identification of new, more specific marker for CSCs, and (3) pathways that are suitable as future targets for anti-cancer drugs. This is of particular importance, because today’s chemotherapy targets the proliferating cancer cells sparing the relatively slow dividing cancer stem cells. The first step on this long road therefore is to analyze genome-wide expression-profiles within the same type of cancer and then between different types of cancer, encircling those target genes and pathways, which are specific to these cells

Zinc and ageing: third Zincage conference

The importance of Zn for optimal functioning of the immune system and antioxidant stress response is well documented. Zn homeostasis influences development and function of immune cells, activity of stress-related and antioxidant proteins [metallothioneins (MT), chaperones, ApoJ, Poly(ADP-Ribose) polymerase-1 (PARP-1) and Methionione Sulfoxide Reductase (Msr), Superoxide Dismutase (SOD)], and helps to maintain genomic integrity and stability. During ageing, the intake of Zn decreases due to inadequate diet and/or intestinal malabsorption, contributing to frailty, general disability and increased incidence of age-related degenerative diseases (cancer, infections and atherosclerosis). Although many factors contributing to Zn deficiency have been identified, the biochemical markers of Zn deficiency as well as the possibility to achieve relevant health benefits through Zn supplementation in the elderly are still a matter for evaluation. Taking into account that Zn homeostasis is regulated by proteins and enzymes for which polymorphisms have been previously found to be associated with successful/unsuccessful ageing, genetic screening might be of added value in evaluating the individual response to Zn supplementation. Biochemical, immunological, dietary and genetic studies aimed at understanding the impact of Zn in healthy ageing, the effect of Zn supplementation in the elderly and finally formulating a rationale for the promotion of correct Zn supplementation were discussed at the international Zincage conference held in Ancona in January 2007

Cupricyclins, Novel Redox-Active Metallopeptides Based on Conotoxins Scaffold

Highly stable natural scaffolds which tolerate multiple amino acid substitutions represent the ideal starting point for the application of rational redesign strategies to develop new catalysts of potential biomedical and biotechnological interest. The knottins family of disulphide-constrained peptides display the desired characteristics, being highly stable and characterized by hypervariability of the inter-cysteine loops. The potential of knottins as scaffolds for the design of novel copper-based biocatalysts has been tested by engineering a metal binding site on two different variants of an ω-conotoxin, a neurotoxic peptide belonging to the knottins family. The binding site has been designed by computational modelling and the redesigned peptides have been synthesized and characterized by optical, fluorescence, electron spin resonance and nuclear magnetic resonance spectroscopy. The novel peptides, named Cupricyclin-1 and -2, bind one Cu2+ ion per molecule with nanomolar affinity. Cupricyclins display redox activity and catalyze the dismutation of superoxide anions with an activity comparable to that of non-peptidic superoxide dismutase mimics. We thus propose knottins as a novel scaffold for the design of catalytically-active mini metalloproteins

How do cardiologists select patients for dual antiplatelet therapy continuation beyond 1 year after a myocardial infarction? Insights from the EYESHOT Post-MI Study

Background: Current guidelines suggest to consider dual antiplatelet therapy (DAPT) continuation for longer than 12 months in selected patients with myocardial infarction (MI). Hypothesis: We sought to assess the criteria used by cardiologists in daily practice to select patients with a history of MI eligible for DAPT continuation beyond 1 year. Methods: We analyzed data from the EYESHOT Post-MI, a prospective, observational, nationwide study aimed to evaluate the management of patients presenting to cardiologists 1 to 3 years from the last MI event. Results: Out of the 1633 post-MI patients enrolled in the study between March and December 2017, 557 (34.1%) were on DAPT at the time of enrolment, and 450 (27.6%) were prescribed DAPT after cardiologist assessment. At multivariate analyses, a percutaneous coronary intervention (PCI) with multiple stents and the presence of peripheral artery disease (PAD) resulted as independent predictors of DAPT continuation, while atrial fibrillation was the only independent predictor of DAPT interruption for patients both at the second and the third year from MI at enrolment and the time of discharge/end of the visit. Conclusions: Risk scores recommended by current guidelines for guiding decisions on DAPT duration are underused and misused in clinical practice. A PCI with multiple stents and a history of PAD resulted as the clinical variables more frequently associated with DAPT continuation beyond 1 year from the index MI

Regulation of the fibroblast growth factor receptor in early Xenopus embryos.

Recent evidence suggests that fibroblast growth factor (FGF) is a primary mesoderm inducer in Xenopus development. We have isolated a full-length cDNA clone for the Xenopus FGF receptor. Like other FGF receptors, the Xenopus homolog is a membrane-spanning protein with a split intracellular tyrosine kinase domain. The Xenopus FGF receptor mRNA is present as a maternal message whose levels are constant through early development. There is no specific regional localization of the transcript by analysis of FGF receptor mRNA levels in microdissected embryonic tissue. In isolated animal-pole blastomeres, FGF receptor mRNA declines over 16 hr in culture and this loss can be prevented by incubation with FGF or activin. Despite the presence of the FGF receptor mRNA in the oocyte, oocytes in culture do not respond to added FGF. However, injection of exogenous Xenopus FGF receptor transcripts into oocytes does generate a functional response to FGF. Our data suggest that posttranscriptional response to FGF. Our data suggest that posttranscriptional mechanisms regulate the FGF receptor in the oocyte and early embryo and further suggest that mesoderm-inducing factors influence receptor mRNA levels during the time of early tissue formation