(725d) In Vivo Endothelial Uptake of Nanoparticles: Impact of Disturbed Flow and Degraded Glycocalyx
Methods: Mouse models of endothelial dysfunction apply partial ligation of the left carotid artery (LCA) to model acute disturbed flow in the mice, which has been reported to induce endothelial dysfunction.6,7 Seven C57BL/6 mice at 4 weeks of age underwent this partial LCA ligation surgery. Polymer- and biotin-coated 10 nm gold nanospheres (GNS) were administered to the mice at day 26 after LCA ligation. In the case of targeted delivery to the endothelial cell surface, an anti-ICAM peptide was conjugated to the GNS surface before it was administered to the mice. The animals were euthanized 2 days after receiving the GNS without or conjugated with anti-ICAM. The LCA and right carotid artery (RCA) were separately cryopreserved and sectioned onto slides. GCX was imaged after incubating LCA and RCA sections with antibody to block GNS, followed by biotinylated wheat germ agglutinin and horseradish peroxidase conjugated to streptavidin. GNS were imaged after incubation with horseradish peroxidase conjugated to streptavidin. We then applied a fluorescent reagent that reacts with horseradish peroxidase. The blood vessels were imaged by fluorescence microscopy and analyzed using ImageJ for GCX coverage and GNS uptake.
Results: The partial LCA ligation model achieved the goal of creating a vessel with disturbed flow. The model also provided a convenient comparison with a healthy RCA experiencing streamlined flow. The vessel walls of the LCA exhibited a more discontinuous GCX layer on the intima as compared to the RCA, decreasing from 76.3 ± 10.2 % in the RCA to 21.2 ± 5.9 % in the LCA. This observable dysfunction correlated to increased nanoparticle uptake, as the LCA took in approximately 2.5-fold more GNS than the RCA did, based on the fluorescence signal detected in the histology images. The studies to improve endothelial cell surface-specific localization of these nanoparticles by targeting ICAM are still ongoing.
Conclusions: A partial LCA ligation was performed to acutely disturb blood flow in a mouse vessel and observe resultant endothelial GCX dysfunction as well as passive targeting of GNS to affected areas. The affected LCA exhibited lack of continuous GCX layer, as well as increased localization of plolymer-coated GNS that are designed to deliver drug therapies. These results indicate that vessel and GCX dysfunction, both precursors of atherosclerosis and cardiovascular disease, can be induced in a mouse model to study targeted drug delivery. Passive nanoparticle uptake differences between the healthy RCA and disturbed LCA indicate a role of GCX infiltration of nanoparticles to the endothelial cells. The ongoing investigation will determine the effectiveness of active targeting with ICAM to localize these nanoparticles to the endothelial cell surface. This strategy of targeting dysfunctional vessels based on the GCX offers a new approach in cardiovascular disease therapy and prevention.
Acknowledgements: We appreciate funding from National Institutes of Health (K01 HL125499) and the Northeastern University Provostâs Tier 1 Grant.
- Weinbaum, S.; et. al, Annu Rev Biomed Eng 2007, 9,121-167.
- Becker, B. et. al, Br J Clin Pharmacol 2015, 80, 389-402.
- Cheng, et. al, Int. J. Nanomed 2016, 21, 3305-3315.
- Cheng, et. al, Int. J. Nanomed 2019, 14, 319-333.
- Nakashima et. al, Arteriosclerosis, Thrombosis, and Vascular Biology 1998, 18, 842-851.
- Kumar, S.; et. al, J. Lab. Invest. 2017, 97, 935-945
- Mitra, et. al, Trans. Med. Comm. 2018, 3-10.