Fig 2: Cytotoxicity activity of Pt metal clusters. (a) Cellular uptake of Pt MCs in HeLa cells. HR-TEM amorphous aggregations of Pt clusters circled in blue and inset: AC-HAADF-STEM images of a HeLa sample treated with Pt MCs. (b) Cytotoxicity of Pt MCs against tumoral and non-tumoral cell lines by MTT assay. Data are the mean ± SD of three dose-response independent experiments. Cisplatin (0.1–50 µM) was used as the standard compound. (c) Cytotoxicity profiles of Pt MCs (left) and cisplatin (right) including ovarian cancer cells (A2780) and cisplatin-resistant cells (A2780cis). (d) Top: Cell morphology and fluorescence microscopy images of annexin-V/propidium iodide double-staining of HeLa cells after treatment with Pt MCs. Cells were labelled with annexin-V-FITC (?exc/?em 485/535 nm; green fluorescence) and propidium iodide (?exc/?em 535/617 nm; red fluorescence) and visualized with a Leica fluorescence microscope PAULA. Bottom: Caspase-3 activation assay and LDH release by HeLa cells upon treatment with platinum metal clusters. Data are the mean ± SD of three independent dose-response experiments. Asterisks indicate the confidence degree of the measurements between groups by the student's t-test (***p < 0.001 ns: non-significant). (e) DNA damage induced by Pt MCs against HeLa cells by comet assay. Left: Fluorescence microscopy images (green fluorescence) of comet experiments. Cells were treated with PtMCs and incubated for 24 h. Images are representative of three independent experiments. Right: Quantitation of DNA damage by means of a 6-class visual classification score. Data represent the mean ± SD of three independent experiments. Asterisks indicate the confidence degree of the measurements between groups by the DNA damage (***p < 0.001).

Fig 3: Analysis of infection of Vero E6 cells with VSV-based pseudovirus SARS-CoV-2 (MOI 0.1) pre-treated with the different UA-based formulations. In (A), the images reported were an example of each stimulation for GFP fluorescence expression obtained at 40× magnification by Paula (Leica DM1000); in (B) the analysis of GFP expression in cells obtained by the image analysis using Image J Software and the results were expressed as mean ± SD (%) compared to infected control cells (0 line) of four independent experiments. In (C), cell viability measured by MTT test after infection and expressed as mean ± SD (%) compared to uninfected control cells (0 line) obtained from four independent experiments. In (D) UA spectrophotometry quantification at 281 nm by Tecan (Infinite 200 Pro Mplex, Tecan, Milan, Italy) and data are expressed as ng/mL ± SD of four independent experiments. Solution 1 = 150 mcg/mL ß-CD +HPMC; Solution 2 = 150 mcg/mL UA+ 150 mcg/mL ß-CD + 1% HPMC; Solution 3 = 75 mcg/mL UA+ 150 mcg/mL ß-CD + 1% HPMC; HEXEDRA+ = 75 mcg/mL UA+ 150 mcg/mL ß-CD + 0.1% Tpmix + 1% HPMC. * p < 0.05 vs. Control; ** p < 0.05 vs. Solution 1; y p < 0.05 vs. Solution 2; yy p < 0.05 vs. Solution 3; f p < 0.05 vs. Infected control.

Fig 4: Analysis of infection of Vero E6 and HNEpC cells with VSV-based pseudovirus SARS-CoV-2 (MOI 0.1) pre-treated with the different UA-based formulations. The images reported were an example of each stimulation for GFP fluorescence expression obtained at 40× magnification by Paula (Leica DM1000). The positive cells were counted analyzing GFP expression in cells by the Image J software and the results were expressed as mean ± SD (%) compared to infected control cells (0 line) of four independent experiments. The abbreviations are the same reported in Figure 2. * p < 0.05 vs. infected control; ** p < 0.05 vs. Solution 1; y p < 0.05 vs. Solution 2.

Fig 5: Analysis of infection of Vero E6 cells with VSV-based pseudovirus SARS-CoV-2 (MOI 0.1) pre-treated with the different UA-based formulations. The images reported were an example of each stimulation for GFP fluorescence expression obtained at 40× magnification by Paula (Leica DM1000). The positive cells were counted analyzing GFP expression in cells by the Image J software and the results were expressed as mean ± SD (%) compared to infected control cells (0 line) of four independent experiments. The abbreviations are the same reported in Figure 2. * p < 0.05 vs. infected control; ** p < 0.05 vs. Solution 1; y p < 0.05 vs. Solution 2; yy p < 0.05 vs. Solution 3.