Dendritic polyglycerol nanoparticles show charge dependent bio-distribution in early human placental explants and reduce hCG secretion

Abstract

A thorough understanding of nanoparticle bio-distribution at the feto-maternal interface will be a prerequisite for their diagnostic or therapeutic application in women of childbearing age and for teratologic risk assessment. Therefore, the tissue interaction of biocompatible dendritic polyglycerol nanoparticles (dPG-NPs) with first- trimester human placental explants were analyzed and compared to less sophisticated trophoblast-cell based models. First-trimester human placental explants, BeWo cells and primary trophoblast cells from human term placenta were exposed to fluorescence labeled, ∼5 nm dPG-NPs, with differently charged surfaces, at concentrations of 1 µM and 10 nM, for 6 and 24 h. Accumulation of dPGs was visualized by fluorescence microscopy. To assess the impact of dPG-NP on trophoblast integrity and endocrine function, LDH, and hCG releases were measured. A dose- and charge-dependent accumulation of dPG-NPs was observed at the early placental barrier and in cell lines, with positive dPG-NP-surface causing deposits even in the mesenchymal core of the placental villi. No signs of plasma membrane damage could be detected. After 24 h we observed a significant reduction of hCG secretion in placental explants, without significant changes in trophoblast apoptosis, at low concentrations of charged dPG-NPs. In conclusion, dPG-NP's surface charge substantially influences their bio-distribution at the feto-maternal interface, with positive charge facilitating trans-trophoblast passage, and in contrast to more artificial models, the first-trimester placental explant culture model reveals potentially hazardous influences of charged dPG-NPs on early placental physiology.

Keywords: BeWo; Dendritic polyglycerol nanoparticles; early human placenta; hCG; nanotoxicology; primary trophoblasts.

Figure 1.

Schematic illustration of rhodamine B labeled dendritic polyglycerol (dPG-RB), dPG sulfate (dPGS-RB), and dPG amine (dPGNH₂-RB). Counter ions are not shown for clarity reasons.

Figure 2.

Localization of differently charged (neutral (Neu; A,B), negative (Neg; C–F), positive (Pos; G–L)), labeled dPG-NPs (pink) in exposed primary cytotrophoblasts (A–D,G,H), and BeWo cells (B,E,F,I–L). Exposure concentrations: 1 µM (A–J) and 10 nM (K,L)), exposure time: 6 h (C,E,G,I,K), and 24 h (A,B,D,F,H,J,L). Nuclear staining: DAPI (blue). Confocal images, 400x. Patchy perinuclear and nuclear localization was seen after 6 h of exposure to 1 µM neg. dPG-NPs (C,E) and pos. dPG-NPs (G,I), increasing after 24 h. Notably, in BeWo cells pos. dPG-NP deposits were also detected after exposing them to a 10 nM concentration for 6 h (K) and 24 h (L). No neutral dPG-NP deposits were detected in both cell types (A,B).

Figure 3.

Localization of differently charged (neutral (Neu; C), negative (Neg; A,D), positive (Pos; B, E–F)), labeled dPG-NPs (orange), in exposed placental explants. Exposure concentration: 1 µM, exposure-time: 6 h (A,B) and 24 h (C–F). Nuclear staining: DAPI (blue). Fluorescence microscopy images, 200× (A,B), 1000× (C–F). Local staining of cytoplasm and nuclei in syncytiotrophoblast (ST, arrows) was detected after 6 h of exposure to neg. and pos. dPG-NPs (A,B), increasing after 24 h. Notably, pos. dPG-NP deposits, were localized at the basal membrane (thin arrows, F) close to the cytotrophoblast cell layer (CT) and in some cells of mesenchymal core (short arrowheads) (E,F). No signal was observed for neu. dPG-NPs (C).

Figure 4.

LDH- and hCG-levels in culture supernatants of placental explants, BeWo cells, and primary cytotrophoblast cells, exposed to neutral (Neu), positively (Pos), and negatively charged (Neg) dPG-NPs for 24 h, at concentrations of 1 µM and 10 nM and to 1% Triton^®X100. LDH data are shown in relative units as ratio to control values, while hCG data are presented in mU mL⁻¹. *(p < 0.05), **(p < 0.01), and ***(p < 0.001) indicate statistically significant differences.