Effect of surface charge and agglomerate degree of magnetic iron oxide nanoparticles on KB cellular uptake in vitro

doi:10.1016/j.colsurfb.2009.05.031

Colloids and Surfaces B: Biointerfaces

Volume 73, Issue 2, 15 October 2009, Pages 294-301

https://doi.org/10.1016/j.colsurfb.2009.05.031 Get rights and content

Abstract

We synthesized three types of magnetic iron oxide nanoparticles (MNPs), which were meso-2,3-dimercaptosuccinic acid (DMSA) coated MNPs (DMSA@MNPs, 17.3 ± 4.8 nm, negative charge), chitosan (CS) coated MNPs (CS@MNPs, 16.5 ± 6.1 nm, positive charge) and magnetic nanoparticles agglomerates, formed by electronic aggregation between DMSA@MNPs and CS (CS-DMSA@MNPs, 85.7 ± 72.9 nm, positive charge) respectively. The interactions of these MNPs with Oral Squamous Carcinoma Cell KB were investigated. The results showed that cellular uptakes of MNPs were on the dependence of incubation time, nanoparticles concentration and nanoparticles properties such as surface charge, size, etc. The cellular uptake was enhanced with the increase of incubation time and nanoparticles concentration. Although all MNPs could enter to cells, we observed apparent differences in the magnitude of nanoparticles uptaken. The cellular uptake of CS-DMSA@MNPs by KB cells was the highest and that of DMSA@MNPs was the lowest among the three types of MNPs. The same conclusions were drawn via the reduction of water proton relaxation times $T_{2}^{*}$ , resulting from the different iron load of labeled cells using a 1.5 T clinical MR imager. The finding of this study will have implications in the chemical design of nanomaterials for biomedical applications.

Introduction

The diagnostic and therapeutic applications of magnetic iron oxide nanoparticles (MNPs) such as drug delivery, magnetic resonance imaging (MRI), hyperthermia techniques, cell separation and tissue repair expanded enormously in the past decades [1], [2], [3], [4], [5], [6]. Such applications of MNPs owed to their low toxicity to human beings and the possibility to exploit their outstanding magnetic properties [7], [8], For those applications, it is desired that MNPs have potential to interact with cells strongly and enter into them. So understanding the interactions of nanoparticles with cells is a key gap in nanotechnology that needs to be filled. Nanoparticle size [9], [10], surface chemistry [11], and charge [12] have a profound effect on internalization capability. Considerable researches have been devoted to improving the cell internalizing efficiency by changing the size of nanoparticles [13], by the attachment on the nanoparticle surface with protein transduction domains [14], [15], or by utilizing transfection agents [16], [17].

Noted that, due to the magnetic dipole–dipole attractions between nanoparticles, these particles are prone to agglomerate and form large clusters, resulting in increased particle size. So far only limited data are available regarding MNPs agglomerations within the biological systems. A recent study on the in vitro uptake of oxide nanoparticles in human lung cells has underlined the effect of nanopaticles agglomeration and size [18]. Wick et al. explored the cytotoxic effects about the degree and kind of agglomerations of carbon nanotube [19]. But to the best of our knowledge, not much is known about their impact on biological systems including humans.

In present study, we prepared three types of MNPs, which were DMSA coated MNPs (17.3 ± 4.8 nm, negative charge), CS coated MNPs (16.5 ± 6.1 nm, positive charge) and CS magnetic agglomerates (85.7 ± 72.9 nm, positive charge) through electrostatic interaction respectively. Subsequently, we evaluated the effect of these MNPs by utilizing Oral Squamous Carcinoma Cell KB as an in vitro model. CS magnetic agglomerates were more uptakes by KB cells than CS coated MNPs, while DMSA coated MNPs were less uptakes than CS coated MNPs. These in vitro findings can be applied to in vivo studies for further investigations on quantitative assessment of health effects.

Section snippets

Synthesis of chitosan coated MNPs

Magnetic nanoparticles were synthesized by co-precipitation of ferrous and ferric chlorides in aqueous solution [20]. Solutions of FeCl₃·6H₂O and FeCl₂·4H₂O (molar ratio 2:1) were mixed and precipitated with concentrated ammonia while being stirred vigorously under N₂ protecting. The black precipitate was formed and washed several times with deionized water. The final magnetite nanoparticles were dispersed in deionized water with pH 3.0 and oxidized into more stable MNPs (γ-Fe₂O₃) by air at the

Characterizations of magnetic nanoparticles

Fig. 1 showed FTIR spectra of CS, CS@MNPs, DMSA@MNPs and CS-DMSA@MNPs. The characteristic absorption bands of CS (Fig. 1(a)) appeared at 1646 cm⁻¹(amide I), 1618 cm⁻¹ (amide II) and 1400 cm⁻¹(amide III). In the spectrum of CS@MNPs (Fig. 1(b)) and CS-DMSA@MNPs (Fig. 1(d)), compared with the specrum of CS, the 1646 cm⁻¹ peak of –NH₂ bending vibration shift to 1621 cm⁻¹, and a new peak 1406 cm⁻¹ appeared. It could be attributed to the linkage between carboxyl group and ammonium ion [23]. The results

Conclusions

In conclusion, low-toxic DMSA@MNPs, CS@MNPs and CS-DMSA@MNPs have been prepared and characterized in vitro by various physicochemical means. Cellular uptakes of MNPs by KB cells were dependent on incubation time, nanoparticle concentration and properties. The higher cellular uptake of CS-DMSA@MNPs compared with CS@MNPs results from the high surface charge and large agglomerate size. CS@MNPs with positive zeta potential also showed higher cellular uptake compared to DMSA@MNPs with negative zeta

Acknowledgements

This work has been carried out under financial support of the National Natural Science Foundation of China (nos. 60571031, 60501009 and 60725101) and National Basic Research Program of China (nos. 2006CB933206 and 2006CB705602).

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Effect of surface charge and agglomerate degree of magnetic iron oxide nanoparticles on KB cellular uptake in vitro

Abstract

Introduction

Section snippets

Synthesis of chitosan coated MNPs

Characterizations of magnetic nanoparticles

Conclusions

Acknowledgements

Nanotoday

Adv. Drug Deliv. Rev.

Cancer Lett.

Biomaterials

Biomaterials

Int. J. Pharm.

Toxicol. Lett.

J. Mol. Liquids

Biomaterials

Biomaterials

Eng. Aspects

J. Magn. Magn. Mater.

Biomaterials

Science

Science

J. Am. Chem. Soc.

Adv. Mater.