Puncturing of lyophilized tissue engineered vascular matrices enhances the efficiency of their recellularization

Data on in vitro engineered "off the shelf" matrices support the concept of endogenous cellular repopulation driving the graft's remodeling via immune-mediated response. This seems important to further accelerate the cell reconstitution and may play a crucial role when mononuclear cells are used. Nevertheless, studies on decellularized xenogeneic grafts showed only limited host cell repopulation post-implantation. This study aims at a systematic comparison of reseeding methods (dripping, injection, bathing in a cell suspension and combined puncturing-dripping method) to define the most efficient technique enhancing recellularization of tissue engineered vascular matrices (patches, vessels, small diameter and standard size valves) prior implantation. The constructs were analyzed histologically, biochemically and biomechanically. Various preconditioning treatments (wet, lyophilized and air-dried) combined with reseeding methods demonstrated the highest cell loading efficiency, despite applied crimping and flow stress, of lyophilization followed by puncturing-dripping technique. This novel seeding method allows for an efficient, time saving graft reseeding that can be used within a one-step cardiovascular clinical intervention. STATEMENT OF SIGNIFICANCE The concept of living tissue engineered, self-repairing, autologous cardiovascular replacements, was proposed alternatively to existing synthetic/xenogeneic prostheses. Recent studies in animal models demonstrate faster in vivo recellularization after grafts pre-seeding with cells prior implantation. Pre-seeded cells hold either, the ability to differentiate directionally or attract host cells, crucial for graft integration and remodeling. It is unclear, however, how efficient the pre-loading is and how well cells withstand the flow. The study presents a systematic overview on cell loading techniques of different cardiovascular constructs, tested under static and dynamic conditions. Comparison illustrates a significantly higher efficiency of cells loading in lyophilized tissues punctured before their standard seeding. This technique may beneficially accelerate remodeling of cardiovascular grafts in further in vivo studies

Human iPSCs and Genome Editing Technologies for Precision Cardiovascular Tissue Engineering

Induced pluripotent stem cells (iPSCs) originate from the reprogramming of adult somatic cells using four Yamanaka transcription factors. Since their discovery, the stem cell (SC) field achieved significant milestones and opened several gateways in the area of disease modeling, drug discovery, and regenerative medicine. In parallel, the emergence of clustered regularly interspaced short palindromic repeats (CRISPR)-associated protein 9 (CRISPR-Cas9) revolutionized the field of genome engineering, allowing the generation of genetically modified cell lines and achieving a precise genome recombination or random insertions/deletions, usefully translated for wider applications. Cardiovascular diseases represent a constantly increasing societal concern, with limited understanding of the underlying cellular and molecular mechanisms. The ability of iPSCs to differentiate into multiple cell types combined with CRISPR-Cas9 technology could enable the systematic investigation of pathophysiological mechanisms or drug screening for potential therapeutics. Furthermore, these technologies can provide a cellular platform for cardiovascular tissue engineering (TE) approaches by modulating the expression or inhibition of targeted proteins, thereby creating the possibility to engineer new cell lines and/or fine-tune biomimetic scaffolds. This review will focus on the application of iPSCs, CRISPR-Cas9, and a combination thereof to the field of cardiovascular TE. In particular, the clinical translatability of such technologies will be discussed ranging from disease modeling to drug screening and TE applications

Endothelial Progenitor Cell-Based in vitro Pre-Endothelialization of Human Cell-Derived Biomimetic Regenerative Matrices for Next-Generation Transcatheter Heart Valves Applications

Hemocompatibility of cardiovascular implants represents a major clinical challenge and, to date, optimal antithrombotic properties are lacking. Next-generation tissue-engineered heart valves (TEHVs) made from human-cell-derived tissue-engineered extracellular matrices (hTEMs) demonstrated their recellularization capacity in vivo and may represent promising candidates to avoid antithrombotic therapy. To further enhance their hemocompatibility, we tested hTEMs pre-endothelialization potential using human-blood-derived endothelial-colony-forming cells (ECFCs) and umbilical vein cells (control), cultured under static and dynamic orbital conditions, with either FBS or hPL. ECFCs performance was assessed via scratch assay, thereby recapitulating the surface damages occurring in transcatheter valves during crimping procedures. Our study demonstrated: feasibility to form a confluent and functional endothelium on hTEMs with expression of endothelium-specific markers; ECFCs migration and confluency restoration after crimping tests; hPL-induced formation of neo-microvessel-like structures; feasibility to pre-endothelialize hTEMs-based TEHVs and ECFCs retention on their surface after crimping. Our findings may stimulate new avenues towards next-generation pre-endothelialized implants with enhanced hemocompatibility, being beneficial for selected high-risk patients

Tailoring crystallinity for hemocompatible and durable PEEK cardiovascular implants

Polymers have the potential to replace metallic or bioprosthetic heart valve components due to superior durability and inertness while allowing for native tissue-like flexibility. Despite these appealing properties, certain polymers such as polyetheretherketone (PEEK) have issues with hemocompatibility, which have previously been addressed through assorted complex processes. In this paper, we explore the enhancement of PEEK hemocompatibility with polymer crystallinity. Amorphous, semi-crystalline and crystalline PEEK are investigated in addition to a highly crystalline carbon fiber (CF)/PEEK composite material (CFPEEK). The functional group density of the PEEK samples is determined, showing that higher crystallinity results in increased amount of surface carbonyl functional groups. The increase of crystallinity (and negatively charged groups) appears to cause significant reductions in platelet adhesion (33 vs. 1.5 % surface coverage), hemolysis (1.55 vs. 0.75 %∙cm$^{-2}$), and thrombin generation rate (4840 vs. 1585 mU/mL/min/cm$^{2}$). In combination with the hemocompatibility study, mechanical characterization demonstrates that tailoring crystallinity is a simple and effective method to control both hemocompatibility and mechanical performance of PEEK. Furthermore, the results display that CFPEEK composite performed very well in all categories due to its enhanced crystallinity and complete carbon encapsulation, allowing the unique properties of CFPEEK to empower new concepts in cardiovascular device design

Combining Cell Technologies With Biomimetic Tissue Engineering Applications: A New Paradigm for Translational Cardiovascular Therapies

Cardiovascular disease is a major cause of morbidity and mortality worldwide and, to date, the clinically available prostheses still present several limitations. The design of next-generation regenerative replacements either based on cellular or extracellular matrix technologies can address these shortcomings. Therefore, tissue engineered constructs could potentially become a promising alterative to the current therapeutic options for patients with cardiovascular diseases. In this review, we selectively present an overview of the current tissue engineering tools such as induced pluripotent stem cells, biomimetic materials, computational modeling, and additive manufacturing technologies, with a focus on their application to translational cardiovascular therapies. We discuss how these advanced technologies can help the development of biomimetic tissue engineered constructs and we finally summarize the latest clinical evidence for their use, and their potential therapeutic outcome

Next-generation tissue-engineered heart valves with repair, remodelling and regeneration capacity

Valvular heart disease is a major cause of morbidity and mortality worldwide. Surgical valve repair or replacement has been the standard of care for patients with valvular heart disease for many decades, but transcatheter heart valve therapy has revolutionized the field in the past 15 years. However, despite the tremendous technical evolution of transcatheter heart valves, to date, the clinically available heart valve prostheses for surgical and transcatheter replacement have considerable limitations. The design of next-generation tissue-engineered heart valves (TEHVs) with repair, remodelling and regenerative capacity can address these limitations, and TEHVs could become a promising therapeutic alternative for patients with valvular disease. In this Review, we present a comprehensive overview of current clinically adopted heart valve replacement options, with a focus on transcatheter prostheses. We discuss the various concepts of heart valve tissue engineering underlying the design of next-generation TEHVs, focusing on off-the-shelf technologies. We also summarize the latest preclinical and clinical evidence for the use of these TEHVs and describe the current scientific, regulatory and clinical challenges associated with the safe and broad clinical translation of this technology.</p

Optimal plant density and nitrogen rates for improving off-season corn yields in Brazil

Integrating plant density and nitrogen (N) management is a strategy for improving corn yields, especially for off-season corn production in the tropics. This study tested the hypothesis that increasing plant densities and N rates promotes yield gains for off-season corn production in high-yielding environments. The aim of the study was to investigate the yield performances of two hybrid versions (DKB PRO and DKB PRO3) submitted to three plant densities (55,000; 65,000 and 75,000 plants ha−1) and four N rates (control, 60, 120 and 180 kg ha−1 N). Field trials were undertaken at Uberlândia-MG (site1 and 2) and Pedro Afonso-TO (site 3), Brazil from which data on corn yield parameters were collected and analyzed. Multivariate analysis separated the three trial areas into two groups, presenting high (sites 1 and 2) and low yields (site 3), which were related to weather conditions. There was no influence of a hybrid version or plant densities on crop yields at site 1 or 2. In contrast, there was a positive response to increasing plant densities and the use of DKB PRO3 at site 3. A significant response to N was observed at sites 2 and 3, following a plateau model. Our results suggest that N application rates and plant densities do have the potential to increase off-season corn yields in low yielding environments

The Long-Baseline Neutrino Experiment: Exploring Fundamental Symmetries of the Universe

The preponderance of matter over antimatter in the early Universe, the dynamics of the supernova bursts that produced the heavy elements necessary for life and whether protons eventually decay --- these mysteries at the forefront of particle physics and astrophysics are key to understanding the early evolution of our Universe, its current state and its eventual fate. The Long-Baseline Neutrino Experiment (LBNE) represents an extensively developed plan for a world-class experiment dedicated to addressing these questions. LBNE is conceived around three central components: (1) a new, high-intensity neutrino source generated from a megawatt-class proton accelerator at Fermi National Accelerator Laboratory, (2) a near neutrino detector just downstream of the source, and (3) a massive liquid argon time-projection chamber deployed as a far detector deep underground at the Sanford Underground Research Facility. This facility, located at the site of the former Homestake Mine in Lead, South Dakota, is approximately 1,300 km from the neutrino source at Fermilab -- a distance (baseline) that delivers optimal sensitivity to neutrino charge-parity symmetry violation and mass ordering effects. This ambitious yet cost-effective design incorporates scalability and flexibility and can accommodate a variety of upgrades and contributions. With its exceptional combination of experimental configuration, technical capabilities, and potential for transformative discoveries, LBNE promises to be a vital facility for the field of particle physics worldwide, providing physicists from around the globe with opportunities to collaborate in a twenty to thirty year program of exciting science. In this document we provide a comprehensive overview of LBNE's scientific objectives, its place in the landscape of neutrino physics worldwide, the technologies it will incorporate and the capabilities it will possess.Comment: Major update of previous version. This is the reference document for LBNE science program and current status. Chapters 1, 3, and 9 provide a comprehensive overview of LBNE's scientific objectives, its place in the landscape of neutrino physics worldwide, the technologies it will incorporate and the capabilities it will possess. 288 pages, 116 figure

Implicating genes, pleiotropy, and sexual dimorphism at blood lipid loci through multi-ancestry meta-analysis

Publisher Copyright: © 2022, The Author(s).Background: Genetic variants within nearly 1000 loci are known to contribute to modulation of blood lipid levels. However, the biological pathways underlying these associations are frequently unknown, limiting understanding of these findings and hindering downstream translational efforts such as drug target discovery. Results: To expand our understanding of the underlying biological pathways and mechanisms controlling blood lipid levels, we leverage a large multi-ancestry meta-analysis (N = 1,654,960) of blood lipids to prioritize putative causal genes for 2286 lipid associations using six gene prediction approaches. Using phenome-wide association (PheWAS) scans, we identify relationships of genetically predicted lipid levels to other diseases and conditions. We confirm known pleiotropic associations with cardiovascular phenotypes and determine novel associations, notably with cholelithiasis risk. We perform sex-stratified GWAS meta-analysis of lipid levels and show that 3–5% of autosomal lipid-associated loci demonstrate sex-biased effects. Finally, we report 21 novel lipid loci identified on the X chromosome. Many of the sex-biased autosomal and X chromosome lipid loci show pleiotropic associations with sex hormones, emphasizing the role of hormone regulation in lipid metabolism. Conclusions: Taken together, our findings provide insights into the biological mechanisms through which associated variants lead to altered lipid levels and potentially cardiovascular disease risk.Peer reviewe