The cone of curves of Fano varieties of coindex four

We classify the cones of curves of Fano varieties of dimension greater or equal than five and (pseudo)index dim X -3, describing the number and type of their extremal rays.Comment: 27 pages; changed the numbering of Theorems, Definitions, Propositions, etc. in accordance with the published version to avoid incorrect reference

Microfluidic Platform for Adherent Single Cell High-Throughput Screening

This paper was presented at the 4th Micro and Nano Flows Conference (MNF2014), which was held at University College, London, UK. The conference was organised by Brunel University and supported by the Italian Union of Thermofluiddynamics, IPEM, the Process Intensification Network, the Institution of Mechanical Engineers, the Heat Transfer Society, HEXAG - the Heat Exchange Action Group, and the Energy Institute, ASME Press, LCN London Centre for Nanotechnology, UCL University College London, UCL Engineering, the International NanoScience Community, www.nanopaprika.eu.Traditionally, in vitro investigations on biology and physiology of cells rely on averaging the responses eliciting from heterogeneous cell populations, thus being unsuitable for assessing individual cell behaviors in response to external stimulations. In the last years, great interest has thus been focused on single cell analysis and screening, which represents a promising tool aiming at pursuing the direct and deterministic control over cause-effect relationships guiding cell behavior. In this regard, a high-throughput microfluidic platform for trapping and culturing adherent single cells was presented. A single cell trapping mechanism was implemented based on dynamic variation of fluidic resistances. A round-shaped culture chamber (Φ=250μm, h=25μm) was conceived presenting two connections with a main fluidic path: (i) an upper wide opening, and (ii) a bottom trapping junction which modulates the hydraulic resistance. Several layouts of the chamber were designed and computationally validated for the optimization of the single cell trapping efficacy. The optimized chamber layouts were integrated in a polydimethylsiloxane (PDMS) microfluidic platform presenting two main functionalities: (i) 288 chambers for trapping single cells, and (ii) a chaoticmixer based serial dilution generator for delivering both soluble factors and non-diffusive molecules under spatio-temporally controlled chemical patterns. The devices were experimentally validated and allowed for trapping individual U87-MG (human glioblastoma-astrocytoma epithelial-like) cells and culturing them up to 3 days

High-Throughput Microfluidic Platform for 3D Cultures of Mesenchymal Stem Cells, Towards Engineering Developmental Processes

The development of in vitro models to screen the effect of different concentrations, combinations and temporal sequences of morpho-regulatory factors on stem/progenitor cells is crucial to investigate and possibly recapitulate developmental processes with adult cells. Here, we designed and validated a microfluidic platform to (i) allow cellular condensation, (ii) culture 3D micromasses of human bone marrow-derived mesenchymal stromal cells (hBM-MSCs) under continuous flow perfusion, and (ii) deliver defined concentrations of morphogens to specific culture units. Condensation of hBM-MSCs was obtained within 3 hours, generating micromasses in uniform sizes (56.2 ± 3.9 μm). As compared to traditional macromass pellet cultures, exposure to morphogens involved in the first phases of embryonic limb development (i.e. Wnt and FGF pathways) yielded more uniform cell response throughout the 3D structures of perfused micromasses (PMMs), and a 34-fold higher percentage of proliferating cells at day 7. The use of a logarithmic serial dilution generator allowed to identify an unexpected concentration of TGFβ3 (0.1 ng/ml) permissive to hBM-MSCs proliferation and inductive to chondrogenesis. This proof-of-principle study supports the described microfluidic system as a tool to investigate processes involved in mesenchymal progenitor cells differentiation, towards a ‘developmental engineering’ approach for skeletal tissue regeneration

Rational curves and bounds on the Picard number of Fano manifolds

We prove that Generalized Mukai Conjecture holds for Fano manifolds $X$ of pseudoindex $i_X \ge (\dim X +3)/3$. We also give different proofs of the conjecture for Fano fourfolds and fivefolds.Comment: Modified the proof of the main theorem and eliminated a statement about the cone of curves. To appear in Geometriae Dedicat

High-throughput microfluidic platform for adherent single cells non-viral gene delivery

The widespread use of gene therapy as a therapeutic tool relies on the development of DNA-carrying vehicles devoid of any safety concerns. In contrast to viral vectors, non-viral gene carriers show promise in this perspective, although their low transfection efficiency leads to the necessity to carry out further optimizations. In order to overcome the limitations of traditional macroscale approaches, which mainly consist of time-consuming and simplified models, a microfluidic strategy has been developed to carry out transfection studies on single cells in a high-throughput and deterministic fashion. A single cell trapping mechanism has been implemented, based on the dynamic variation of fluidic resistances. For this purpose, we designed a round-shaped culture chamber integrated with a bottom trapping junction, which modulates the hydraulic resistance. Several layouts of the chamber were designed and computationally validated for optimization of the single cell trapping efficacy. The optimized chamber layout was integrated in a polydimethylsiloxane (PDMS) microfluidic platform presenting two main functionalities: (i) 288 chambers for trapping single cells, and (ii) a serial dilution generator with chaotic mixing properties, able to deliver to the chambers both soluble factors and non-diffusive particles (i.e., polymer/DNA complexes, polyplexes) under spatio-temporally controlled chemical patterns. The devices were experimentally validated and allowed the trapping of individual human glioblastoma–astrocytoma epithelial-like cells (U87-MG) with a trapping efficacy of about 40%. The cells were cultured within the device and underwent preliminary transfection experiments using 25 kDa linear polyethylenimine (lPEI)-based polyplexes, confirming the potentiality of the proposed platform for the future high-throughput screening of gene delivery vectors and for the optimization of transfection protocols

Lab-on-Chip for Testing Myelotoxic Effect of Drugs and Chemicals

This paper was presented at the 4th Micro and Nano Flows Conference (MNF2014), which was held at University College, London, UK. The conference was organised by Brunel University and supported by the Italian Union of Thermofluiddynamics, IPEM, the Process Intensification Network, the Institution of Mechanical Engineers, the Heat Transfer Society, HEXAG - the Heat Exchange Action Group, and the Energy Institute, ASME Press, LCN London Centre for Nanotechnology, UCL University College London, UCL Engineering, the International NanoScience Community, www.nanopaprika.eu.In the last twenty years, one of the main goals in the drug discovery field has been the development of reliable in vitro models. In particular, in 2006 the European Centre for the Validation of Alternative Methods (ECVAM) has approved the Colony forming Unit-Granulocytes-Macrophages (CFU-GM) test, which is the first and currently unique test applied to evaluate the myelotoxicity of xenobiotics in vitro. The present work aimed at miniaturizing this in vitro assay by developing and validating a Lab-on-Chip (LoC) platform consisting of a high number of bioreactor chambers with screening capabilities in a high-throughput regime

LivHeart: A Multi Organ-on-Chip Platform to Study Off-Target Cardiotoxicity of Drugs Upon Liver Metabolism

The drug discovery and development process is still long, costly, and highly risky. The principal attrition factor is undetected toxicity, with hepatic and cardiac toxicities playing a critical role and being the main responsible of safety-related drug withdrawals from the market. Multi Organs-on-Chip (MOoC) represent a disruptive solution to study drug-related effects on several organs simultaneously and to efficiently predict drug toxicity in preclinical trials. Specifically focusing on drug safety, different technological features are applied here to develop versatile MOoC platforms encompassing two culture chambers for generating and controlling the type of communication between a metabolically competent liver model and a functional 3D heart model. The administration of the drug Terfenadine, a cardiotoxic compound liver-metabolized into the noncardiotoxic Fexofenadine, proved that liver metabolism and a fine control over drug diffusion are fundamental to elicit a physio-pathological cardiac response. From these results, an optimized LivHeart platform is developed to house a liver model and a cardiac construct that can be mechanically trained to achieve a beating microtissue, whose electrophysiology can be directly recorded in vitro. The platform is proved able to predict off-target cardiotoxicity of Terfenadine after liver metabolism both in terms of cell viability and functionality

Fabrication of 3D cell-laden hydrogel microstructures through photo-mold patterning

Native tissues are characterized by spatially organized three-dimensional (3D) microscaled units which functionally define cells–cells and cells–extracellular matrix interactions. The ability to engineer biomimetic constructs mimicking these 3D microarchitectures is subject to the control over cell distribution and organization. In the present study we introduce a novel protocol to generate 3D cell laden hydrogel micropatterns with defined size and shape. The method, named photo-mold patterning (PMP), combines hydrogel micromolding within polydimethylsiloxane (PDMS) stamps and photopolymerization through a recently introduced biocompatible ultraviolet (UVA) activated photoinitiator (VA-086). Exploiting PDMS micromolds as geometrical constraints for two methacrylated prepolymers (polyethylene glycol diacrylate and gelatin methacrylate), micrometrically resolved structures were obtained within a 3 min exposure to a low cost and commercially available UVA LED. The PMP was validated both on a continuous cell line (human umbilical vein endothelial cells expressing green fluorescent protein, HUVEC GFP) and on primary human bone marrow stromal cells (BMSCs). HUVEC GFP and BMSCs were exposed to 1.5% w/v VA-086 and UVA light (1 W, 385 nm, distance from sample = 5 cm). Photocrosslinking conditions applied during the PMP did not negatively affect cells viability or specific metabolic activity. Quantitative analyses demonstrated the potentiality of PMP to uniformly embed viable cells within 3D microgels, creating biocompatible and favorable environments for cell proliferation and spreading during a seven days' culture. PMP can thus be considered as a promising and cost effective tool for designing spatially accurate in vitro models and, in perspective, functional constructs

Usefulness of the maggic score in predicting the competing risk of non-sudden death in heart failure patients receiving an implantable cardioverter-defibrillator: A sub-analysis of the observo-icd registry

The role of prognostic risk scores in predicting the competing risk of non-sudden death in heart failure patients with reduced ejection fraction (HFrEF) receiving an implantable cardioverterdefibrillator (ICD) is unclear. To this goal, we evaluated the accuracy and usefulness of the MetaAnalysis Global Group in Chronic Heart Failure (MAGGIC) score. The present analysis included 1089 HFrEF ICD recipients enrolled in the OBSERVO-ICD registry (NCT02735811). During a median follow-up of 36 months (1st\u20133rd IQR 25\u201348 months), 193 patients (17.7%) experienced at least one appropriate ICD therapy, and 133 patients died (12.2%) without experiencing any ICD therapy. The frequency of patients receiving ICD therapies was stable around 17\u201319% across increasing tertiles of 3-year MAGGIC probability of death, whereas non-sudden mortality increased (6.4% to 9.8% to 20.8%, p < 0.0001). Accuracy of MAGGIC score was 0.60 (95% CI, 0.56\u20130.64) for the overall outcome, 0.53 (95% CI, 0.49\u20130.57) for ICD therapies and 0.65 (95% CI, 0.60\u20130.70) for non-sudden death. In patients with higher 3-year MAGGIC probability of death, the increase in the competing risk of non-sudden death during follow-up was greater than that of receiving an appropriate ICD therapy. Results were unaffected when analysis was limited to ICD shocks only. The MAGGIC risk score proved accurate and useful in predicting the competing risk of non-sudden death in HFrEF ICD recipients. Estimation of mortality risk should be taken into greater consideration at the time of ICD implantation

A method to generate perfusable physiologic-like vascular channels within a liver-on-chip model

Data availability: The data that support the findings of this study are available from the corresponding author upon reasonable request.Supplementary Material: Further data on collagen–fibrin gel characterization, immunofluorescent staining of peculiar endothelial markers, rocker platform setup, and an upgraded version of the current ECM-mediated-contact platform are presented in the supplementary material available online at: https://pubs.aip.org/bmf/article-supplement/2925768/zip/064103_1_5.0170606.suppl_material/ (zip file).Copyright © 2023 Author(s). The human vasculature is essential in organs and tissues for the transport of nutrients, metabolic waste products, and the maintenance of homeostasis. The integration of vessels in in vitro organs-on-chip may, therefore, improve the similarity to the native organ microenvironment, ensuring proper physiological functions and reducing the gap between experimental research and clinical outcomes. This gap is particularly evident in drug testing and the use of vascularized models may provide more realistic insights into human responses to drugs in the pre-clinical phases of the drug development pipeline. In this context, different vascularized liver models have been developed to recapitulate the architecture of the hepatic sinusoid, exploiting either porous membranes or bioprinting techniques. In this work, we developed a method to generate perfusable vascular channels with a circular cross section within organs-on-chip without any interposing material between the parenchyma and the surrounding environment. Through this technique, vascularized liver sinusoid-on-chip systems with and without the inclusion of the space of Disse were designed and developed. The recapitulation of the Disse layer, therefore, a gap between hepatocytes and endothelial cells physiologically present in the native liver milieu, seems to enhance hepatic functionality (e.g., albumin production) compared to when hepatocytes are in close contact with endothelial cells. These findings pave the way to numerous further uses of microfluidic technologies coupled with vascularized tissue models (e.g., immune system perfusion) as well as the integration within multiorgan-on-chip settings.This work was partially financed by the European Union's Horizon 2020 research and innovation programme under the Marie Skłodowska-Curie Grant Agreement No 800924 (International Cancer Research Fellowships-2 [iCARE-2])