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ICAM-1-Targeted Nanocarriers Attenuate Endothelial Release of Soluble ICAM-1, an Inflammatory Regulator.

ABSTRACT: Targeting of drug nanocarriers (NCs) to intercellular adhesion molecule-1 (ICAM-1), an endothelial-surface protein overexpressed in many pathologies, has shown promise for therapeutic delivery into and across this lining. Yet, due to the role of ICAM-1 in inflammation, the effects of targeting this receptor need investigation. Since ICAM-1 binding by natural ligands (leukocyte integrins) results in release of the "soluble ICAM-1" ectodomain (sICAM-1), an inflammatory regulator, we investigated the influence of targeting ICAM-1 with NCs on this process. For this, sICAM-1 was measured by ELISA from cell-medium supernatants, after incubation of endothelial cell (EC) monolayers in the absence versus presence of anti-ICAM NCs. In the absence of NCs, ECs released sICAM-1 when treated with a pro-inflammatory cytokine (TNFα). This was reduced by inhibiting matrix metalloproteinases MMP-9 or MMP-2, yet inhibiting both did not render additive effects. Release of sICAM-1 mainly occurred at the basolateral versus apical side, and both MMP-9 and MMP-2 influenced apical release, while basolateral release depended on MMP-9. Interestingly, anti-ICAM NCs reduced sICAM-1 to a greater extent than MMP inhibition, both at the apical and basolateral sides. This effect was enhanced with time, although NCs had been removed after binding to cells, ruling out a "trapping" effect of NCs. Instead, inhibiting anti-ICAM NC endocytosis counteracted their inhibition on sICAM-1 release. Hence, anti-ICAM NCs inhibited sICAM-1 release by mobilizing ICAM-1 from the cell-surface into intracellular vesicles. Since elevated levels of sICAM-1 associate with numerous diseases, this effect represents a secondary benefit of using ICAM-1-targeted NCs for drug delivery.

SUBMITTER: Manthe RL

PROVIDER: S-EPMC5510616 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Delivery of drugs into the endothelium by nanocarriers targeted to endothelial determinants may improve treatment of vascular maladies. This is the case for intercellular adhesion molecule 1 (ICAM-1), a glycoprotein overexpressed on endothelial cells (ECs) in many pathologies. ICAM-1-targeted nanocarriers bind to and are internalized by ECs via a non-classical pathway, CAM-mediated endocytosis. In this work we studied the effects of endothelial adaptation to physiological flow on the endocytosis of model polymer nanocarriers targeted to ICAM-1 (anti-ICAM/NCs, ~180 nm diameter). Culturing established endothelial-like cells (EAhy926 cells) and primary human umbilical vein ECs (HUVECs) under 4 dyn/cm(2) laminar shear stress for 24 h resulted in flow adaptation: cell elongation and formation of actin stress fibers aligned to the flow direction. Fluorescence microscopy showed that flow-adapted cells internalized anti-ICAM/NCs under flow, although at slower rate versus non flow-adapted cells under static incubation (~35% reduction). Uptake was inhibited by amiloride, whereas marginally affected by filipin and cadaverine, implicating that CAM-endocytosis accounts for anti-ICAM/NC uptake under flow. Internalization under flow was more modestly affected by inhibiting protein kinase C, which regulates actin remodeling during CAM-endocytosis. Actin recruitment to stress fibers that maintain the cell shape under flow may delay uptake of anti-ICAM/NCs under this condition by interfering with actin reorganization needed for CAM-endocytosis. Electron microscopy revealed somewhat slow, yet effective endocytosis of anti-ICAM/NCs by pulmonary endothelium after i.v. injection in mice, similar to that of flow-adapted cell cultures: ~40% (30 min) and 80% (3 h) internalization. Similar to cell culture data, uptake was slightly faster in capillaries with lower shear stress. Further, LPS treatment accelerated internalization of anti-ICAM/NCs in mice. Therefore, regulation of endocytosis of ICAM-1-targeted nanocarriers by flow and endothelial status may modulate drug delivery into ECs exposed to different physiological (capillaries vs. arterioles/venules) or pathological (ischemia, inflammation) levels and patterns of blood flow.

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ICAM-1-Targeted Nanocarriers Attenuate Endothelial Release of Soluble ICAM-1, an Inflammatory Regulator.

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