The effect of blood protein adsorption on cellular uptake of anatase TiO₂ nanoparticles

Figures & data

Figure 1 Thermal analysis of TiO₂ NP samples A.5, A.10, and A.40.

Notes: (A) TGA; and (B) TPD–MS.

Abbreviations: NP, nanoparticle; TGA, thermogravimetric analysis; TPD–MS, temperature programmed desorption–mass spectrometry; T, temperature.

Figure 2 XPS of TiO₂ NP samples A.5, A.10, and A.40.

Notes: (A) Survey scan; (B) atomic percent of surface elements; (C) high resolution scan of O 1s.

Abbreviations: NP, nanoparticle; XPS, X-ray photoelectron spectroscopy; At%, atomic percent; pos, position.

Table 1 Agglomerate sizes expressed as hydrodynamic diameter of TiO₂ NPs (100 mg/L) in RPMI 1640 cell culture medium with (100 mg/L) and without proteins, after 24 h and 3 h (in brackets) rotation at 37°C

Sample	Hydrodynamic diameter ± SD (nm)
	Without proteins	HSA	Glbs	Fib
A.5	2,779±130	573±6	*	*
	(2,874±62)	(547±6)	(*)	(660±24)
A.10	3,242±244	572±14	*	*
	(3,211±39)	(580±10)	(*)	(573±17)
A.40	1,439±41	1,275±30	*	*
	(1,800±70)	(1,500±64)	(*)	(962±121)

Note:

* It was impossible to measure the hydrodynamic diameter because the samples were too polydispersed (polydispersity index >0.6).

Abbreviations: NPs, nanoparticles; RPMI, Roswell Park Memorial Institute; h, hours; SD, standard deviation; HSA, human serum albumin; Glbs, γ-globulins; Fib, fibrinogen.

Table 2 Zeta potentials of TiO₂ NPs in RPMI medium with and without proteins, after 24 h rotation at 37°C

Sample	Zeta potential ± SD (mV)
	Without proteins	HSA	Glbs	Fib
A.5	−18.2±1.6	−12.0±1.0	−5.4±0.5	−9.0±0.5
A.10	−18.0±1.3	−13.4±0.6	−5.9±0.4	−7.6±0.5
A.40	−20.6±1.7	−17.6±1.1	−16.4±1.1	−18.4±1.4

Abbreviations: NPs, nanoparticles; RPMI, Roswell Park Memorial Institute; h, hours; SD, standard deviation; HSA, human serum albumin; Glbs, γ-globulins; Fib, fibrinogen.

Figure 3 Quantity adsorbed of proteins onto TiO₂ NPs after 2-h and 24-h rotations at 37°C. The results for samples A.5, A.10, and A.40 are expressed as medians with interquartile range.

Abbreviations: HSA, human serum albumin; Glbs, γ-globulins; Fib, fibrinogen; h, hours; quant, quantity; ads, adsorbed; NPs, nanoparticles.

Figure 4 Degree of cellular uptake, showing the concentration (in μg/L) of titanium element associated with L929 fibroblasts, after 24 h exposure to TiO₂ NPs suspended in different media. The results are expressed as medians with interquartile range.

Note: *Statistical significance (P<0.05, Dunn’s multiple comparison test).

Abbreviations: HSA, human serum albumin; Glbs, γ-globulins; Fib, fibrinogen; h, hours; NPs, nanoparticles.

Figure 5 URI images of L929 fibroblasts at 1000× magnification (scale bar: 10 μm), after 24 h exposure to 0.5 mg/L of TiO₂ NPs (sample A.5) in different media.

Notes: (A) Control cells not exposed to NPs. (B) Protein-free. (C) HSA. (D) Glb. (E) Fib. Bright spots are NPs/agglomerates either internalized or attached to the cell membrane.

Abbreviations: URI, ultra-high resolution imaging; h, hours; NPs, nanoparticles; HSA, human serum albumin; Glbs, γ-globulins; Fib, fibrinogen.