Mr Rolando Matos


Davide Maselli, Rolando S. Matos, Robert D. Johnson, Davide Martella, Davide Martella, Valeria Caprettini, Valeria Caprettini, Ciro Chiappini, Ciro Chiappini, Patrizia Camelliti, Paola Campagnolo (2022)Porcine Organotypic Epicardial Slice Protocol: A Tool for the Study of Epicardium in Cardiovascular Research, In: Frontiers in cardiovascular medicine9920013 Frontiers Media S.A

The epicardium has recently gained interest in the cardiovascular field due to its capacity to support heart regeneration after ischemic injury. Models to study the epicardium of large animals in vitro are limited and mainly based on epicardial cell isolation/differentiation from stem cells, followed by 2D cells culture. In this method paper, we describe the procedure to obtain and culture 3D organotypic heart slices presenting an intact epicardium, as a novel model to study the epicardial physiology and activation. Epicardial slices are obtained from porcine hearts using a high-precision vibratome and retain a healthy epicardial layer embedded in its native extracellular environment and connected with other cardiac cells (cardiomyocytes, fibroblasts, vascular cells etc.). Epicardial slices can be cultured for 72 h, providing an ideal model for studying the epicardium physiology or perform pharmacological interventions/gene therapy approaches. We also report on methods to assesses the viability and composition of the epicardial slices, and evaluate their architecture in 3D through tissue decoloration. Finally, we present a potential application for a nanomaterial-based gene transfer method for tracking of epicardial cells within the slice. Crucially, given the similarity in morphology and physiology of porcine heart with its human counterpart, our system provides a platform for translational research while providing a clinically relevant and ethical alternative to the use of small animals in this type of research.

GAVRIELLE REBECCA UNTRACHT, ROLANDO SZILVESZTER MATOS, NIKOLAOS DIKAIOS, MARIAM BAPIR, ABDULLAH KHAN DURRANI, Teemapron Butsabong, PAOLA CAMPAGNOLO, DAVID SAMPSON, CHRISTIAN HEISS, DANUTA SAMPSON (2021)OCTAVA: An open-source toolbox for quantitative analysis of optical coherence tomography angiography images, In: Plos One16(12)pp. 1-22 Public Library of Science

Optical coherence tomography angiography (OCTA) performs non-invasive visualization and characterization of microvasculature in research and clinical applications mainly in ophthalmology and dermatology. A wide variety of instruments, imaging protocols, processing methods and metrics have been used to describe the microvasculature, such that comparing different study outcomes is currently not feasible. With the goal of contributing to standardization of OCTA data analysis, we report a user-friendly, open-source toolbox, OCTAVA (OCTA Vascular Analyzer), to automate the pre-processing, segmentation, and quantitative analysis of en face OCTA maximum intensity projection images in a standardized workflow. We present each analysis step, including optimization of filtering and choice of segmentation algorithm, and definition of metrics. We perform quantitative analysis of OCTA images from different commercial and non-commercial instruments and samples and show OCTAVA can accurately and reproducibly determine metrics for characterization of microvasculature. Wide adoption could enable studies and aggregation of data on a scale sufficient to develop reliable microvascular biomarkers for early detection, and to guide treatment, of microvascular disease.

Rolando S. Matos, Davide Maselli, John H. McVey, Christian Heiss, Paola Campagnolo (2022)3D printed bioreactor enabling the pulsatile culture of native and angioplastied large arteries, In: Frontiers in Cardiovascular Medicine9864580 Frontiers Media

Routine interventions such as balloon angioplasty, result in vascular activation and remodeling, often requiring re-intervention. 2D in vitro models and small animal experiments have enabled the discovery of important mechanisms involved in this process, however the clinical translation is often underwhelming. There is a critical need for an ex vivo model representative of the human vascular physiology and encompassing the complexity of the vascular wall and the physical forces regulating its function. Vascular bioreactors for ex vivo culture of large vessels are viable alternatives, but their custom-made design and insufficient characterization often hinders the reproducibility of the experiments. The objective of the study was to design and validate a novel 3D printed cost-efficient and versatile perfusion system, capable of sustaining the viability and functionality of large porcine arteries for 7 days and enabling early post-injury evaluations. MultiJet Fusion 3D printing was used to engineer the EasyFlow insert, converting a conventional 50 ml centrifuge tube into a mini bioreactor. Porcine carotid arteries either left untreated or injured with an angioplasty balloon, were cultured under pulsatile flow for up to 7 days. Pressure, heart rate, medium viscosity and shear conditions were adjusted to resemble arterial in vivo hemodynamics. Tissue viability, cell activation and matrix remodeling were analyzed by immunohistochemistry, and vascular function was monitored by duplex ultrasound. Culture conditions in the EasyFlow bioreactor preserved endothelial coverage and smooth muscle organization and extracellular matrix structure in the vessel wall, as compared to static culture. Injured arteries presented hallmarks of early remodeling, such as intimal denudation, smooth muscle cell disarray and media/adventitia activation in flow culture. Duplex ultrasound confirmed continuous pulsatile blood flow conditions, dose-dependent vasodilator response to nitroglycerin in untreated vessels and impaired dilator response in angioplastied vessels. The scope of this work is to validate a low-cost, robust and reproducible system to explore the culture of native and injured large arteries under pulsatile flow. While the study of vascular pathology is beyond the scope of the present paper, our system enables future investigations and provides a platform to test novel therapies and devices ex vivo, in a patient relevant system.

DAVIDE MASELLI, ROLANDO SZILVESZTER MATOS, ROBERT DAVID JOHNSON, C Chiappini, PATRIZIA CAMELLITI, PAOLA CAMPAGNOLO (2021)Epicardial slices: a 3D organotypic model for the study of epicardial activation and differentiation, In: NPJ Regenerative medicine Springer

The epicardium constitutes an untapped reservoir for cardiac regeneration. Upon heart injury, the adult epicardium re-activates, leading to epithelial-to-mesenchymal transition (EMT), migration and differentiation. While interesting mechanistic and therapeutic findings arose from lower vertebrates and rodent models, the introduction of an experimental system representative of large mammals would undoubtedly facilitate translational advancements. Here, we apply innovative protocols to obtain living 3D organotypic epicardial slices from porcine hearts, encompassing the epicardial/myocardial interface. In culture, our slices preserve the in vivo architecture and functionality, presenting a continuous epicardium overlaying a healthy and connected myocardium. Upon thymosin β4 treatment of the slices, the epicardial cells become activated upregulating epicardial and EMT genes, resulting in epicardial cell mobilization and differentiation into epicardial-derived mesenchymal cells. Our 3D organotypic model enables to investigate the reparative potential of the adult epicardium, offering an advanced tool to explore ex vivo the complex 3D interactions occurring within the native heart environment

VALERIA MASTRULLO, DANIEL RUTGER VAN DER VEEN, Priyanka Gupta, ROLANDO SZILVESZTER MATOS, JONATHAN DAVID JOHNSTON, JOHN HENDERSON MCVEY, Paolo Madeddu, EIRINI VELLIOU, PAOLA CAMPAGNOLO (2022)Pericytes' circadian clock affect endothelial cells' synchronization and angiogenesis in a 3D tissue engineered scaffold, In: Frontiers in pharmacology. Frontiers