11 research outputs found
Protocol to generate an in vitro model to study vascular calcification using human endothelial and smooth muscle cells
Vascular calcification is a systemic disease characterized by calcium salt deposition
within vascular walls. Here, we present a protocol for establishing an
advanced dynamic in vitro co-culture system using endothelial and smooth muscle
cells to replicate vascular tissue complexity. We describe steps for cell culture
and seeding in a double-flow bioreactor that recreates the action of blood in humans.
We then detail the induction of calcification, setting up of the bioreactor,
followed by cell viability assessment and calcium quantification
Type-specific inflammatory responses of vascular cells activated by interaction with virgin and aged microplastics
Microplastics (MPs) are recognized as a major environmental problem due to their ubiquitous presence in ecosystems and bioaccumulation in food chains. Not only humans are continuously exposed to these pollutants through ingestion and inhalation, but recent findings suggest they may trigger vascular inflammation and potentially worsen the clinical conditions of cardiovascular patients. Here we combine headspace analysis by needle trap microextraction-gas chromatography-mass spectrometry (HS-NTME-GC-MS) and biological assays to evaluate the effects of polystyrene, high- and low-density polyethylene MPs on phenotype, metabolic activity, and pro-inflammatory status of Vascular Smooth Muscle Cells (VSMCs) the most prominent cells in vascular walls. Virgin and artificially aged MPs (4 weeks at 40 °C and 750 W/m2 simulated solar irradiation) were comparatively tested at 1 mg/mL to simulate a realistic exposure scenario. Our results clearly show the activation of oxidative stress and inflammatory processes when VSMCs were cultured with aged polymers, with significant overexpression of IL-6 and TNF-α. In addition, volatile organic compounds (VOCs), including pentane, acrolein, propanal, and hexanal as the main components, were released by VSMCs into the headspace. Type-specific VOC response profiles were induced on vascular cells from different MPs
Vascular Calcification: In Vitro Models under the Magnifying Glass.
Vascular calcification is a systemic disease contributing to cardiovascular morbidity and mortality. The pathophysiology of vascular calcification involves calcium salt deposition by vascular smooth muscle cells that exhibit an osteoblast-like phenotype. Multiple conditions drive the phenotypic switch and calcium deposition in the vascular wall; however, the exact molecular mechanisms and the connection between vascular smooth muscle cells and other cell types are not fully elucidated. In this hazy landscape, effective treatment options are lacking. Due to the pathophysiological complexity, several research models are available to evaluate different aspects of the calcification process. This review gives an overview of the in vitro cell models used so far to study the molecular processes underlying vascular calcification. In addition, relevant natural and synthetic compounds that exerted anticalcifying properties in in vitro systems are discussed
Cardiac tissue engineering: Multiple approaches and potential applications
The overall increase in cardiovascular diseases and, specifically, the ever-rising exposure to cardiotoxic compounds has greatly increased in vivo animal testing; however, mainly due to ethical concerns related to experimental animal models, there is a strong interest in new in vitro models focused on the human heart. In recent years, human pluripotent stem cells-derived cardiomyocytes (hiPSC-CMs) emerged as reference cell systems for cardiac studies due to their biological similarity to primary CMs, the flexibility in cell culture protocols, and the capability to be amplified several times. Furthermore, the ability to be genetically reprogrammed makes patient-derived hiPSCs, a source for studies on personalized medicine. In this mini-review, the different models used for in vitro cardiac studies will be described, and their pros and cons analyzed to help researchers choose the best fitting model for their studies. Particular attention will be paid to hiPSC-CMs and three-dimensional (3D) systems since they can mimic the cytoarchitecture of the human heart, reproducing its morphological, biochemical, and mechanical features. The advantages of 3D in vitro heart models compared to traditional 2D cell cultures will be discussed, and the differences between scaffold-free and scaffold-based systems will also be spotlighte
Novel in vitro evidence on the beneficial effect of quercetin treatment in vascular calcification.
Vascular calcification is a pathological chronic condition characterized by calcium
crystal deposition in the vessel wall and is a recurring event in atherosclerosis,
chronic kidney disease, and diabetes. The lack of effective therapeutic treatments
opened the research to natural products, which have shown promising potential in
inhibiting the pathological process in different experimental models. This study
investigated the anti-calcifying effects of Quercetin and Berberine extracts on
vascular smooth muscle cells (VSMCs) treated with an inorganic phosphate
solution for 7 days. Quercetin has shown the highest anti-calcifying activity, as
revealed by the intracellular quantitative assay and morphological analysis.
Confocal microscopy revealed downregulation of RUNX2, a key marker for
calcified phenotype, which was otherwise upregulated in calcified VSMCs. To
investigate the anti-inflammatory activity of Quercetin, culture media were
subjected to immunometric assays to quantify the levels of IL-6 and TNF-α, and
the caspase-1 activity. As expected, calcified VSMCs released a large quantity of
inflammatory mediators, significantly decreasing in the presence of Quercetin. In
summary, our findings suggest that Quercetin counteracted calcification by
attenuating the VSMC pathological phenotypic switch and reducing the
inflammatory response. In our opinion, these preliminary in vitro findings could
be the starting point for further investigations into the beneficial effects of
Quercetin dietary supplementation against vascular calcification
Dynamic Cellular Model as an Emerging Platform to Reproduce the Complexity of Human Vascular Calcification In Vitro
Vascular calcification (VC) is a cardiovascular disease characterized by calcium salt deposition in vascular smooth muscle cells (VSMCs). Standard in vitro models used in VC investigations are based on VSMC monocultures under static conditions. Although these platforms are easy to use, the absence of interactions between different cell types and dynamic conditions makes these models insufficient to study key aspects of vascular pathophysiology. The present study aimed to develop a dynamic endothelial cell-VSMC co-culture that better mimics the in vivo vascular microenvironment. A double-flow bioreactor supported cellular interactions and reproduced the blood flow dynamic. VSMC calcification was stimulated with a DMEM high glucose calcification medium supplemented with 1.9 mM NaH2PO4/Na2HPO4 (1:1) for 7 days. Calcification, cell viability, inflammatory mediators, and molecular markers (SIRT-1, TGFβ1) related to VSMC differentiation were evaluated. Our dynamic model was able to reproduce VSMC calcification and inflammation and evidenced differences in the modulation of effectors involved in the VSMC calcified phenotype compared with standard monocultures, highlighting the importance of the microenvironment in controlling cell behavior. Hence, our platform represents an advanced system to investigate the pathophysiologic mechanisms underlying VC, providing information not available with the standard cell monoculture
Sharing real-world experiences to optimize the management of olaparib toxicities: a practical guidance from an Italian expert panel
Olaparib is the first poly(ADP-ribose) polymerase inhibitor approved as maintenance therapy of recurrent ovarian cancer (OC) patients with a BRCA mutation. To achieve the maximum clinical benefit, adherence to olaparib must be persistent. However, in clinical practice, this is challenged by the frequent suboptimal management of toxicities. In view of the expanding use of olaparib also in Italy, physicians must learn how to adequately and promptly manage drug toxicities not to unnecessarily interrupt or reduce the dose. The experts agreed that nausea,vomiting, anemia, and fatigue are the most frequent events experienced by OC patients on olaparib, and that these toxicities usually develop early during treatment, are mainly of grade 1–2 and transient and can be managed with simple non-pharmacological interventions. By sharing their real-world experiences, the panel prepared, for each toxicity, an algorithm organized by grade and besides the procedures indicated in the local label, included supportive care interventions based also on nutritional and lifestyle modifications and psycho-oncology consultation. Moreover, in view of the tablet entry into the Italian market, the full and reduced dosages of capsules and tablets were compared. This practical guidance is intended to be a tool to support especially less-experienced physicians in the management of these complex patients, with the aim to help preventing the worsening of patients’ conditions and the unnecessary interruption/reduction of olaparib dosage, which may jeopardize treatment efficacy