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Preparation and In Vitro Characterization of Magnetized miR-modified Endothelial Cells
Published on: May 2, 2017
Core-Shell Polymer-Based Nanoparticles Deliver miR-155-5p to Endothelial Cells
Joana C Antunes1, Louise Benarroch1, Fernanda C Moraes1
1Université de Paris, LVTS, INSERM U1148, Université Paris 13, 75018 Paris, France.
Abstract:
Heart failure occurs in over 30% of the worldwide population and most commonly originates from cardiovascular diseases such as myocardial infarction. microRNAs (miRNAs) target and silence specific mRNAs, thereby regulating gene expression. Because the endogenous miR-155-5p has been ascribed to vasculoprotection, loading it onto positively charged, core-shell poly(isobutylcyanoacrylate) (PIBCA)-polysaccharide nanoparticles (NPs) was attempted. NPs showed a decrease (p < 0.0001) in surface electrical charge (ζ potential), with negligible changes in size or shape when loaded with the anionic miR-155-5p. Presence of miR-155-5p in loaded NPs was further quantified. Cytocompatibility up to 100 μg/mL of NPs for 2 days with human coronary artery endothelial cells (hCAECs) was documented. NPs were able to enter hCAECs and were localized in the endoplasmic reticulum (ER). Expression of miR-155-5p was increased within the cells by 75-fold after 4 hours of incubation (p < 0.05) and was still noticeable at day 2. Differences between loaded NP-cultured cells and free miRNA, at days 1 (p < 0.05) and 2 (p < 0.001) suggest the ability of prolonged load release in physiological conditions. Expression of miR-155-5p downstream target BACH1 was decreased in the cells by 4-fold after 1 day of incubation (p < 0.05). This study is a first proof of concept that miR-155-5p can be loaded onto NPs and remain intact and biologically active in endothelial cells (ECs). These nanosystems could potentially increase an endogenous cytoprotective response and decrease damage within infarcted hearts.
Insights
This study successfully loaded miR-155-5p onto nanoparticles (NPs) for potential heart failure treatment. These NPs deliver the microRNA into cells, showing prolonged activity and reduced damage markers.
Area of Science:
- Biomedical Engineering
- Nanotechnology
- Molecular Biology
Background:
- Heart failure affects over 30% globally, often stemming from cardiovascular diseases like myocardial infarction.
- MicroRNAs (miRNAs) regulate gene expression by targeting specific mRNAs.
- Endogenous miR-155-5p is recognized for its vasculoprotective properties.
Purpose of the Study:
- To develop a nanoparticle (NP) delivery system for miR-155-5p.
- To assess the stability and biological activity of miR-155-5p loaded onto poly(isobutylcyanoacrylate)-polysaccharide NPs.
- To evaluate the potential of these NPs in enhancing endothelial cell protection.
Main Methods:
- miR-155-5p was loaded onto positively charged poly(isobutylcyanoacrylate)-polysaccharide NPs.
- Nanoparticle characteristics (size, charge, shape) and miR-155-5p loading were quantified.
- Cytocompatibility and cellular uptake of NPs by human coronary artery endothelial cells (hCAECs) were assessed.
- Intracellular miR-155-5p expression, NP localization, and downstream target gene (BACH1) expression were measured over time.
Main Results:
- NP loading with miR-155-5p resulted in a significant decrease in ζ potential with minimal changes in size or shape.
- NPs were cytocompatible with hCAECs up to 100 μg/mL for 2 days and were internalized by the cells, localizing in the endoplasmic reticulum.
- Intracellular miR-155-5p levels increased significantly (75-fold) within 4 hours and remained elevated for 2 days, indicating prolonged release.
- A significant decrease in the miR-155-5p target gene, BACH1, was observed, suggesting functional activity.
- NP-delivered miRNA showed prolonged activity compared to free miRNA.
Conclusions:
- This study demonstrates a successful proof of concept for loading miR-155-5p onto NPs.
- The NPs maintain miR-155-5p integrity and biological activity within endothelial cells.
- These miR-155-5p-loaded NPs hold promise for enhancing endogenous cytoprotective responses and mitigating damage in infarcted hearts.
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