Uptake and trafficking of different sized PLGA nanoparticles by dendritic cells in imiquimod-induced psoriasis-like mice model

Abstract

Psoriasis is an autoimmune inflammatory disease, where dendritic cells (DCs) play an important role in its pathogenesis. In our previous work, we have demonstrated that topical delivery of curcumin-loaded poly (lactic-co-glycolic acid) (PLGA) nanoparticles (NPs) could treat Imiquimod (IMQ)-induced psoriasis-like mice. The objective of this study is to further elucidate biofate of PLGA NPs after intradermal delivery including DCs uptake, and their further trafficking in psoriasis-like mice model by using fluorescence probes. Two-sized DiO/DiI-loaded PLGA NPs of 50 ± 4.9 nm (S-NPs) and 226 ± 7.8 nm (L-NPs) were fabricated, respectively. In vitro cellular uptake results showed that NPs could be internalized into DCs with intact form, and DCs preferred to uptake larger NPs. Consistently, in vivo study showed that L-NPs were more captured by DCs and NPs were firstly transported to skin-draining lymph nodes (SDLN), then to spleens after 8 h injection, whereas more S-NPs were transported into SDLN and spleens. Moreover, FRET imaging showed more structurally intact L-NPs distributed in skins and lymph nodes. In conclusion, particle size can affect the uptake and trafficking of NPs by DCs in skin and lymphoid system, which needs to be considered in NPs tailing to treat inflammatory skin disease like psoriasis.

Keywords: APCs, antigen-presenting cells; Biofate; CLSM, confocal laser scanning microscope; DCs, dendritic cells; DMF, dimethylformamide; Dendritic cells; DiI, 1,1′-dioctadecyl-3,3,3′,3′-tetramethylindocarbocyanine perchlorate; DiO, 3,3′-dioctadecyloxacarbocyanine perchlorate; Fluorescence; Fluorescence resonance energy transfer; Lymphoid organs; MLN, mesenteric lymph nodes; NPs, nanoparticles; PDI, polydispersity index; PFA, paraformaldehyde; PLGA nanoparticles; Psoriasis; SDLN, skin-draining lymph nodes; Uptake and trafficking.

Graphical abstract

Figure 1

Characterization of DiO/DiI-loaded PLGA nanoparticles. Size distribution of S-NPs (A) and L-NPs (B), and their zeta potential and PDI. Data are presented as mean ± SD (n = 3) (C). (D) TEM images of S-NPs and L-NPs. Scale bar = 100 nm.

Figure 2

Comparison of cellular uptake and integrity of two different-sized PLGA nanoparticles. (A) Counts of DCs that absorbed DiI-loaded PLGA nanoparticles (DiI: 5 μg/mL) after co-incubated with S-NPs (left) and L-NPs (right) for 1, 2 and 4 h. (B) Mean fluorescence intensity of DCs incubated with DiI-loaded PLGA nanoparticles (DiI: 5 μg/mL) for 1, 2 and 4 h, respectively. Data are presented as mean ± SD (n = 3), ∗∗∗∗P < 0.0001, ∗∗P < 0.01. (C) Confocal laser scanning imaging of DCs, which were respectively pre-incubated with DiO/DiI-loaded PLGA nanoparticles (DiO/DiI: 10 μg/mL) for 1 h as the starting point, and then medium was removed to observe the FRET up to 4 h. (Scale bar = 100 μm). (D) FRET ratio changes with time for S-NPs and L-NPs. Data are presented as mean ± SD (n = 3).

Figure 3

DCs uptake preference of intradermal injected DiO-loaded PLGA nanoparticles in IMQ-induced psoriasis-like mice. (A) Representative immunofluorescence staining of psoriasis-like mice skin section (a) that intradermally injected with DiO-loaded PLGA nanoparticles (after injection for 4 h), enlarged in (b). (B) Representative immunofluorescence staining of SDLN from psoriasis-like mice that intradermally injected with DiO-loaded PLGA nanoparticles (after injection for 1 h), enlarged in (b). Scale bar: 250 μm (enlargement: 50 μm). Blue: nucleus, green: DiO, red: anti-CD11c. (C) Flow cytometry gating strategy to identify and analyze DCs populations in different lymphoid organs. Gating is shown in one representative data of mice at 1 h exposure after treated with S-NPs. After the initial live gating in a forward scatter (FSC) and side scatter (SSC) plot, particle-positive cells were first gated. Among them, DCs from SDLN and spleens were gated in a CD45 versus CD11c plot. (D) PBS controls and different sizes of DiO-loaded PLGA nanoparticles were intradermally administrated in IMQ-induced psoriasis-like mice, and the single-cell suspensions obtained from lymphoid organs were analyzed at 1, 4, 8, 24, and 48 h after particle exposure. Among particle-positive subset in SDLN (D) and spleens (E), comparison between S-NPs and L-NPs detected in these tissues were made. Data are presented as mean ± SEM (n = 6), ∗∗P < 0.01, ∗P < 0.05.

Figure 4

In vivo integrity of DiO/DiI-loaded PLGA nanoparticles in skins and SDLN after IMQ-induced psoriasis-like mice were intradermally injected of different-sized DiO/DiI-loaded PLGA nanoparticles at 8 h. Confocal laser scanning imaging of IMQ-induced psoriasis-like mice dorsal skins (A), and SDLN (D). Scale bar = 250 nm. Lambda scan was shown as spectrum in one representative experiment of skin sections (B), and SDLN sections (E). FRET ratios were calculated by the emission intensities of donor (505 nm) and acceptor (565 nm) based on formula FRET ratio = I_{565 nm}/(I_{505 nm} + I_{565 nm}) and shown data pooled from separate experiments of skin sections (C), and SDLN sections (F). Data are presented as mean ± SD (n = 9), ∗∗∗∗P < 0.0001.