Li, HH and Li, J and Wasserloos, KJ and Wallace, C and Sullivan, MG and Bauer, PM and Stolz, DB and Lee, JS and Watkins, SC and St Croix, CM and Pitt, BR and Zhang, LM
(2013)
Caveolae-dependent and -independent uptake of albumin in cultured rodent pulmonary endothelial cells.
PLoS ONE, 8 (11).
Abstract
Although a critical role for caveolae-mediated albumin transcytosis in pulmonary endothelium is well established, considerably less is known about caveolae-independent pathways. In this current study, we confirmed that cultured rat pulmonary microvascular (RPMEC) and pulmonary artery (RPAEC) endothelium endocytosed Alexa488-labeled albumin in a saturable, temperature-sensitive mode and internalization resulted in co-localization by fluorescence microscopy with cholera B toxin and caveolin-1. Although siRNA to caveolin-1 (cav-1) in RPAEC significantly inhibited albumin uptake, a remnant portion of albumin uptake was cav-1-independent, suggesting alternative pathways for albumin uptake. Thus, we isolated and cultured mouse lung endothelial cells (MLEC) from wild type and cav-1-/- mice and noted that ∼ 65% of albumin uptake, as determined by confocal imaging or live cell total internal reflectance fluorescence microscopy (TIRF), persisted in total absence of cav-1. Uptake of colloidal gold labeled albumin was evaluated by electron microscopy and demonstrated that albumin uptake in MLEC from cav-1-/- mice was through caveolae-independent pathway(s) including clathrin-coated pits that resulted in endosomal accumulation of albumin. Finally, we noted that albumin uptake in RPMEC was in part sensitive to pharmacological agents (amiloride [sodium transport inhibitor], Gö6976 [protein kinase C inhibitor], and cytochalasin D [inhibitor of actin polymerization]) consistent with a macropinocytosis-like process. The amiloride sensitivity accounting for macropinocytosis also exists in albumin uptake by both wild type and cav-1 -/- MLEC. We conclude from these studies that in addition to the well described caveolar-dependent pulmonary endothelial cell endocytosis of albumin, a portion of overall uptake in pulmonary endothelial cells is cav-1 insensitive and appears to involve clathrin-mediated endocytosis and macropinocytosis-like process. © 2013 Li et al.
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| Item Type: |
Article
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| Status: |
Published |
| Creators/Authors: |
| Creators | Email | Pitt Username | ORCID  |
|---|
| Li, HH | hul18@pitt.edu | HUL18 | | | Li, J | | | | | Wasserloos, KJ | | | | | Wallace, C | ctw14@pitt.edu | CTW14 | | | Sullivan, MG | | | | | Bauer, PM | | | | | Stolz, DB | donna.stolz@pitt.edu | DSTOLZ | | | Lee, JS | jsl26@pitt.edu | JSL26 | | | Watkins, SC | simon.watkins@pitt.edu | SWATKINS | | | St Croix, CM | claudette.stcroix@pitt.edu | CLS13 | 0000-0003-0794-4939 | | Pitt, BR | brucep@pitt.edu | BRUCEP | | | Zhang, LM | liz19@pitt.edu | LIZ19 | 0000-0003-3351-801X |
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| Contributors: |
| Contribution | Contributors Name | Email | Pitt Username | ORCID |
|---|
| Editor | Zhao, You-Yang | UNSPECIFIED | UNSPECIFIED | UNSPECIFIED |
|
| Centers: |
Other Centers, Institutes, Offices, or Units > Center for Biologic Imaging |
| Date: |
27 November 2013 |
| Date Type: |
Publication |
| Journal or Publication Title: |
PLoS ONE |
| Volume: |
8 |
| Number: |
11 |
| DOI or Unique Handle: |
10.1371/journal.pone.0081903 |
| Schools and Programs: |
School of Public Health > Environmental and Occupational Health
School of Medicine > Anesthesiology
School of Medicine > Cell Biology
School of Medicine > Critical Care Medicine
School of Medicine > Surgery |
| Refereed: |
Yes |
| Date Deposited: |
30 Jan 2014 18:10 |
| Last Modified: |
22 Jun 2021 13:56 |
| URI: |
http://d-scholarship.pitt.edu/id/eprint/20416 |
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