Cellular uptake and intracellular distribution of the nanoparticles studied by CLSM.

<p>bEnd3 cells were cultured on collagen IV-coated glass slides and were treated with a) PEGylated HI 6 dichloride monohydrate-loaded nanoparticles or b) ApoE-modified HI 6 dichloride monohydrate-loaded nanoparticles for 4 h at 37°C. The green autofluorescence of the nanoparticles was used for detection. Red: cytosol stained with CellTracker™ Red CMTPX, blue: nucleus stained with DAPI. Pictures were taken within inner sections of the cells.</p

Long time measurement of transendothelial electrical resistance (TER) after nanoparticle addition.

<p>pBCEC were seeded on collagen IV-coated Transwell inserts and incubated with the free drug or 0.26 mg nanoparticles per cm² growth area of the ApoE-modified (NP-ApoE) as well as the PEGylated (NP-PEG) nanoparticulate formulations, which were loaded with 1000 µM of HI 6 dichloride monohydrate (HI 6-DCL) at 37°C by adding the nanoparticles into the upper/apical compartment of the Transwell system. The TER was measured automatically every hour by impedance measurement. A magnification of the area of interest is highlighted in the red quadrangle. As control the measurement of the TER of a Transwell insert without cells is shown.</p

Transport study of adsorptively HI 6 dichloride monohydrate-loaded nanoparticles on an <i>in vitro</i> BBB model.

<p>Transport study of adsorptively HI 6 dichloride monohydrate-loaded nanoparticles on an <i>in vitro</i> BBB model.</p

Transport study of adsorptively HI 6 dimethanesulfonate-loaded nanoparticles on an <i>in vitro</i> BBB model.

<p>*Mann-Whitney U Test: significant difference between NP-ApoE and free HI 6-DMS (P<0.05).</p

Measurement of transendothelial electrical resistance (TER) and the capacitance (C_cl).

<p>pBCEC were seeded on collagen IV-coated Transwell inserts, which were placed in the cellZscope. The transendothelial electrical resistance (TER) and the capacitance (C_cl) of the pBCEC were measured automatically every hour by impedance measurement. As control the measurement of the transendothelial electrical resistance (TER) of a Transwell insert without cells is shown.</p

Transport study of adsorptively obidoxime-loaded nanoparticles on an <i>in vitro</i> BBB model.

<p>Transport study of adsorptively obidoxime-loaded nanoparticles on an <i>in vitro</i> BBB model.</p

Co-incubation of bEnd3 cells with the different nanoparticulate formulations and LDL.

<p>*: cells without NP incubation, with LDL.</p><p>One representative experiment out n>3 is shown.</p

Specific cellular binding of the ApoE-modified nanoparticles studied by flow cytometry.

<p>bEnd3 cells were incubated with ApoE-modified nanoparticles (NP-ApoE) or control nanoparticles without ApoE modification (NP-PEG) for 4 h at 37°C and 4°C, respectively. Flow cytometry analysis was performed to quantify their cellular binding. The data are shown as histograms of the FL1-H-channel (autofluorescence of the nanoparticles) as well as in the table with the analysis of the Y mean fluorescence and the percentage of positive cells. Green: NP-ApoE, red: NP-PEG, blue: untreated control.</p

Co-incubation of bEnd3 cells with the different nanoparticulate formulations and LDL+RAP.

<p>*: cells without NP incubation, with LDL+RAP;</p><p>One representative experiment out n>3 is shown.</p

Determination of the receptor state of the bEnd3 cells.

<p>C: control only with the secondary antibody.</p