Surfactants influence polymer nanoparticle fate within the brain

Abstract

Drug delivery to the brain is limited by poor penetration of pharmaceutical agents across the blood-brain barrier (BBB), within the brain parenchyma, and into specific cells of interest. Nanotechnology can overcome these barriers, but its ability to do so is dependent on nanoparticle physicochemical properties including surface chemistry. Surface chemistry can be determined by a number of factors, including by the presence of stabilizing surfactant molecules introduced during the formulation process. Nanoparticles coated with poloxamer 188 (F68), poloxamer 407 (F127), and polysorbate 80 (P80) have demonstrated uptake in BBB endothelial cells and enhanced accumulation within the brain. However, the impact of surfactants on nanoparticle fate, and specifically on brain extracellular diffusion or intracellular targeting, must be better understood to design nanotherapeutics to efficiently overcome drug delivery barriers in the brain. Here, we evaluated the effect of the biocompatible and commonly used surfactants cholic acid (CHA), F68, F127, P80, and poly (vinyl alcohol) (PVA) on poly (lactic-co-glycolic acid)-poly (ethylene glycol) (PLGA-PEG) nanoparticle transport to and within the brain. The inclusion of these surfactant molecules decreases diffusive ability through brain tissue, reflecting the surfactant's role in encouraging cellular interaction at short length and time scales. After in vivo administration, PLGA-PEG/P80 nanoparticles demonstrated enhanced penetration across the BBB and subsequent internalization within neurons and microglia. Surfactants incorporated into the formulation of PLGA-PEG nanoparticles therefore represent an important design parameter for controlling nanoparticle fate within the brain.

Keywords: Blood-brain barrier; Brain drug delivery; Cellular uptake; Diffusion; Polymeric nanoparticles; Surfactant.

Figure 1.

Characterization of nanoparticle stability and diffusion in brain tissue. (A) After incubation in rat serum, the PLGA/F127 formulation increased in average size, while all other formulations remained close to their original size. (B) Nanoparticle diffusion trajectories through brain tissue were analyzed to calculate ensemble-averaged mean squared displacement at time lags up to 6.5 s. (C) Log of D_b at 0.8 s were extracted for each trajectory (1 dot = 1 trajectory). (D) The aspect ratio of the MSD curve for each trajectory was extracted and classified as subdiffusive (<0) or superdiffusive (>0). (E) The anomalous exponent α was extracted for each trajectory and classified as superdiffusive (>1), normal (1), or subdiffusive (<1). The (F) trappedness and (G) efficiency of each trajectory was calculated and plotted as violin plots.

Figure 2.

Assessment of surfactant effects on nanoparticle transport in organotypic brain slices. (A) Flow cytometry analysis indicate that all formulations with surfactant demonstrated similar levels of microglial uptake within 4 h. PLGA-PEG/DI achieved elevated levels of uptake. (B) Propidium iodide (PI)-positive cell counts, as a proportion of total cells, demonstrate no significant differences in cytotoxicity across all treatment conditions.

Figure 3.

Distribution of biodegradable, PEGylated nanoparticles (red) in the brain and major organs at t=4h. (A) PLGA-PEG/P80 nanoparticles, unlike all other formulations, exhibit significantly higher accumulation (p=0.0280) in the brain parenchyma (left bars, solid fill) compared to brain capillaries (right bars, hashed). (B) PLGA-PEG/P80 nanoparticles can internalize within some microglia (green, top) and neurons (green, bottom) in the brain parenchyma. (C) PLGA-PEG/DI, PLGA-PEG/CHA, PLGA-PEG/F68, PLGA-PEG/F127, and PLGA-PEG/PVA nanoparticles do not exhibit patterns of microglial (top row) or neuronal (bottom row) uptake, and instead appear associated within the vasculature. (D) Nanoparticles demonstrate accumulation in the serum, liver, and spleen, with minimal signal from the kidney, heart, and lungs. (A, D): Each dot represents one pup for a total of n=4 (brain) or n=5 (major organs). (B-C): All cell nuclei (blue) are stained with DAPI and all scale bars represent 20 μm.

Figure 4.

PLGA-PEG/P80 surface analysis and serum protein adsorption. Positive ion (A) and negative ion (B) peak ratio from PDG-PEG/DI and PLG-PED/P80 ToF-SIMS data. (C) Compared to the PLGA-PEG/DI control, PLGA-PEG/P80 nanoparticles exhibited increased protein adsorption (p=0.0382). (D) PLGA-PEG/P80 demonstrated a negative shift in ζ-potential after plasma incubation, which was not observed with the PLGA-PEG/DI control.