Two peptides targeting endothelial receptors are internalized into murine brain endothelial cells.
The blood-brain barrier (BBB) is one of the main obstacles for therapies targeting brain diseases. Most macromolecules fail to pass the tight BBB, formed by brain endothelial cells interlinked by tight junctions. A wide range of small, lipid-soluble molecules can enter the brain parenchyma via diffu...
| Main Authors: | , , , , , , |
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| Format: | Article |
| Language: | English |
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Public Library of Science (PLoS)
2021-01-01
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| Series: | PLoS ONE |
| Online Access: | https://doi.org/10.1371/journal.pone.0249686 |
| _version_ | 1819037972237910016 |
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| author | Diána Hudecz Sara Björk Sigurdardóttir Sarah Christine Christensen Casper Hempel Andrew J Urquhart Thomas Lars Andresen Morten S Nielsen |
| author_facet | Diána Hudecz Sara Björk Sigurdardóttir Sarah Christine Christensen Casper Hempel Andrew J Urquhart Thomas Lars Andresen Morten S Nielsen |
| author_sort | Diána Hudecz |
| collection | DOAJ |
| description | The blood-brain barrier (BBB) is one of the main obstacles for therapies targeting brain diseases. Most macromolecules fail to pass the tight BBB, formed by brain endothelial cells interlinked by tight junctions. A wide range of small, lipid-soluble molecules can enter the brain parenchyma via diffusion, whereas macromolecules have to transcytose via vesicular transport. Vesicular transport can thus be utilized as a strategy to deliver brain therapies. By conjugating BBB targeting antibodies and peptides to therapeutic molecules or nanoparticles, it is possible to increase uptake into the brain. Previously, the synthetic peptide GYR and a peptide derived from melanotransferrin (MTfp) have been suggested as candidates for mediating transcytosis in brain endothelial cells (BECs). Here we study uptake, intracellular trafficking, and translocation of these two peptides in BECs. The peptides were synthesized, and binding studies to purified endocytic receptors were performed using surface plasmon resonance. Furthermore, the peptides were conjugated to a fluorophore allowing for live-cell imaging studies of their uptake into murine brain endothelial cells. Both peptides bound to low-density lipoprotein receptor-related protein 1 (LRP-1) and the human transferrin receptor, while lower affinity was observed against the murine transferrin receptor. The MTfp showed a higher binding affinity to all receptors when compared to the GYR peptide. The peptides were internalized by the bEnd.3 mouse endothelial cells within 30 min of incubation and frequently co-localized with endo-lysosomal vesicles. Moreover, our in vitro Transwell translocation experiments confirmed that GYR was able to cross the murine barrier and indicated the successful translocation of MTfp. Thus, despite binding to endocytic receptors with different affinities, both peptides are able to transcytose across the murine BECs. |
| first_indexed | 2024-12-21T08:29:54Z |
| format | Article |
| id | doaj.art-922698fa4c0a47eb9b69cb886bed49d9 |
| institution | Directory Open Access Journal |
| issn | 1932-6203 |
| language | English |
| last_indexed | 2024-12-21T08:29:54Z |
| publishDate | 2021-01-01 |
| publisher | Public Library of Science (PLoS) |
| record_format | Article |
| series | PLoS ONE |
| spelling | doaj.art-922698fa4c0a47eb9b69cb886bed49d92022-12-21T19:10:14ZengPublic Library of Science (PLoS)PLoS ONE1932-62032021-01-01164e024968610.1371/journal.pone.0249686Two peptides targeting endothelial receptors are internalized into murine brain endothelial cells.Diána HudeczSara Björk SigurdardóttirSarah Christine ChristensenCasper HempelAndrew J UrquhartThomas Lars AndresenMorten S NielsenThe blood-brain barrier (BBB) is one of the main obstacles for therapies targeting brain diseases. Most macromolecules fail to pass the tight BBB, formed by brain endothelial cells interlinked by tight junctions. A wide range of small, lipid-soluble molecules can enter the brain parenchyma via diffusion, whereas macromolecules have to transcytose via vesicular transport. Vesicular transport can thus be utilized as a strategy to deliver brain therapies. By conjugating BBB targeting antibodies and peptides to therapeutic molecules or nanoparticles, it is possible to increase uptake into the brain. Previously, the synthetic peptide GYR and a peptide derived from melanotransferrin (MTfp) have been suggested as candidates for mediating transcytosis in brain endothelial cells (BECs). Here we study uptake, intracellular trafficking, and translocation of these two peptides in BECs. The peptides were synthesized, and binding studies to purified endocytic receptors were performed using surface plasmon resonance. Furthermore, the peptides were conjugated to a fluorophore allowing for live-cell imaging studies of their uptake into murine brain endothelial cells. Both peptides bound to low-density lipoprotein receptor-related protein 1 (LRP-1) and the human transferrin receptor, while lower affinity was observed against the murine transferrin receptor. The MTfp showed a higher binding affinity to all receptors when compared to the GYR peptide. The peptides were internalized by the bEnd.3 mouse endothelial cells within 30 min of incubation and frequently co-localized with endo-lysosomal vesicles. Moreover, our in vitro Transwell translocation experiments confirmed that GYR was able to cross the murine barrier and indicated the successful translocation of MTfp. Thus, despite binding to endocytic receptors with different affinities, both peptides are able to transcytose across the murine BECs.https://doi.org/10.1371/journal.pone.0249686 |
| spellingShingle | Diána Hudecz Sara Björk Sigurdardóttir Sarah Christine Christensen Casper Hempel Andrew J Urquhart Thomas Lars Andresen Morten S Nielsen Two peptides targeting endothelial receptors are internalized into murine brain endothelial cells. PLoS ONE |
| title | Two peptides targeting endothelial receptors are internalized into murine brain endothelial cells. |
| title_full | Two peptides targeting endothelial receptors are internalized into murine brain endothelial cells. |
| title_fullStr | Two peptides targeting endothelial receptors are internalized into murine brain endothelial cells. |
| title_full_unstemmed | Two peptides targeting endothelial receptors are internalized into murine brain endothelial cells. |
| title_short | Two peptides targeting endothelial receptors are internalized into murine brain endothelial cells. |
| title_sort | two peptides targeting endothelial receptors are internalized into murine brain endothelial cells |
| url | https://doi.org/10.1371/journal.pone.0249686 |
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