Prussian blue nanosphere-embedded in situ hydrogel for photothermal therapy by peritumoral administration

Abstract

To establish an injectable hydrogel containing Prussian blue (PB) nanospheres for photothermal therapy against cancer, PB nanospheres were prepared by one-pot synthesis and the thermosensitive Pluronic F127 was used as the hydrogel matrix. The PB nanospheres and the hydrogel were characterized by shape, particle size, serum stability, photothermal performance upon repeated 808 nm laser irradiation, as well as the rheological features. The effect of the PB nanospheres and the hydrogel were evaluated qualitatively and quantitatively in 4T1 mouse breast cancer cells. The retention, photothermal efficacy, therapeutic effects and systemic toxicity of the hydrogel were assessed in a tumor-bearing mouse model. The PB nanospheres had a diameter of about 150 nm and exhibited satisfactory serum stability, photo-heat convert ability and repeated laser exposure stability. The hydrogel encapsulation did not negatively influence the above features of the photothermal agent. The nanosphere-containing hydrogel showed a phase transition at body temperature and, as a result, a long retention time in vivo. The photothermal agent-embedded hydrogel displayed promising photothermal therapeutic effects in the tumor-bearing mouse model with little-to-no systemic toxicity after peritumoral administration.

Keywords: Hydrogel; In situ; Injectable; Nanospheres; Photothermal; Prussian blue; Thermosensitive.

Graphical abstract

Figure 1

Schematic illustration.

Figure 2

In vitro characterization of the PB nanospheres. (A) TEM image of the PB nanospheres (the scale bar represents 100 nm); (B) particle sizes of the PB nanospheres before (red) and after 3-day incubation in serum at 37 °C (green); (C) and (D) the thermographs and corresponding heating curves of the PB nanospheres at varied concentrations upon 808 nm laser exposure (1 W/cm²); (E) the heating curves of the PB nanospheres upon repeated 808 nm irradiation (1 W/cm²); (F) the visible absorption profiles of the PB nanospheres before and after 5-cycle 808 nm irradiation (1 W/cm²); (G) particle sizes of the PB nanospheres before (red) and after (green) 5-cycle 808 nm irradiation (1 W/cm²).

Figure 3

In vitro characterization of the PB hydrogel. (A) SEM images of the blank Pluronic F127 hydrogel and the PB nanosphere-loaded hydrogel (the scale bars in the upper and lower graphs represent 100 and 2 µm, respectively); (B) and (C) the thermographs and corresponding heating curves of the PB hydrogel and blank hydrogel upon 808 nm irradiation (1 W/cm²); (D) pictures of the blank hydrogel and PB hydrogel at 4 and 37 °C; (E) the rheology curves of the blank hydrogel and PB hydrogel; (F) particle size of the PB nanospheres embedded in the hydrogel after different storage times at 37 °C; (G) the heating profiles of the PB hydrogel upon 808 nm laser exposure (1 W/cm²) after different storage times at 37 °C.

Figure 4

The anti-cancer effects of the PB nanospheres and the hydrogel against 4T1 mouse breast cancer cells. (A) The fluorescence images of the 4T1 cells after being treated by the PB nanospheres differently, the red color represents the dead cells stained by propidium iodide and the blue color represents the nucleus of all the cells; the white line is the boundary of the 808 nm laser; (B) the fluorescence images of the lower 4T1 cells after being treated by the blank hydrogel and PB hydrogel in the upper transwell, the red color represents the dead cells stained by propidium iodide and the blue color represents the nucleus of all the cells; (C) the survival rates of the 4T1 cells and the L02 cells measured by the MTT test after being treated with the PB nanospheres, ^**P < 0.01. Scale bar: 200 μm.

Figure 5

In vivo performances of the PB hydrogel in a tumor-bearing mouse model. (A) The retention of the DiR-labeled hydrogel and free DiR solution after peritumoral administration (Ex/Em: 740/820 nm); (B) and (C) the thermographs and the corresponding heating curves of the tumor sites after peritumoral administration of the blank hydrogel or PB hydrogel or the PB nanodispersions plus 808 nm irradiation (1 W/cm²), ^*P < 0.05; D) pictures of the tumor tissues after photothermal therapy (1 W/cm², 5 min) over eight sequential days, the red circle represents visibly undetectable tumor sample; (E) statistic graph of the tumor weights, ^*P < 0.05; F) hematoxylin & eosin staining of the tumor tissues.

Figure 6

(A) Body weight curves of the tumor-bearing mice throughout the experimental period; (B) hematoxylin & eosin staining of the main organs of the PB hydrogel-treated tumor-bearing mouse; (C) particle sizing distribution of the PB nanospheres (red) and the urine from the PB nanodispersions i.v. treated mouse (blue).