Theranostic Calcium Phosphate Nanoparticles With Potential for Multimodal Imaging and Drug Delivery

Abstract

Calcium phosphate (CaP) bioceramics closely resemble the natural human bone, which is a main reason for their popularity as bone substitutes. However, this compositional similarity makes it difficult to distinguish CaPs, especially in particulate form, from native bone by imaging modalities such as X-ray radiography, computed tomography (CT), and magnetic resonance imaging (MRI) to monitor the healing progress. External contrast agents can improve the imaging contrast of CaPs but can affect their physicochemical properties and can produce artifacts. In this work, we have attempted to improve the contrast of CaP nanoparticles via ion substitutions for multimodal imaging. Calcium-deficient hydroxyapatite (CDHA) nanoparticles with silver (Ag), gadolinium (Gd), and iron (Fe) substitution were prepared by a microwave-accelerated wet chemical process to improve the contrast in CT, T1 (spin-lattice), and T2 (spin-spin) MRI relaxation modes, respectively. Ag, Gd, and Fe were substituted at 0.25, 0.5, and 0.25 at.%, respectively. The ion-substituted CDHA (ICDHA) was found to be phase pure by X-ray diffraction (XRD) and Fourier transform infrared spectroscopy (FT-IR). Transmission electron microscopy (TEM) images showed that the ICDHA nanoparticles were platelet shaped and of 52 ± 2 nm length and 6 ± 1 nm width. The ICDHA showed high contrast in X-ray and CT compared to CDHA. The vibrating sample magnetometry (VSM) studies showed the ICDHA to exhibit paramagnetic behavior compared to diamagnetic CDHA, which was further confirmed by improved contrast in T1 and T2 MRI mode. In addition, the in vitro tetracycline drug loading and release was studied to investigate the capability of these nanoparticles for antibiotic drug delivery. It was found that a burst release profile was observed for 24 h with 47-52% tetracycline drug release. The ICDHA nanoparticles also showed in vitro antibacterial activity against Staphylococcus aureus and Escherichia coli due to Ag, which was further enhanced by antibiotic loading. In vitro biocompatibility studies showed that the triple-ion-substituted ICDHA nanoparticles were cytocompatible. Thus, the ion-substituted CDHA nanoparticles can have potential theranostic applications due to their multimodal image contrast, antibacterial activity, and drug delivery potential. Future work will be conducted with actual bone samples in vitro or in animal models.

Keywords: antibacterial; bone substitute; calcium phosphate nanoparticles; drug delivery; image contrast; ion substitution; multimodal imaging; theranostics.

Figure 1

XRD pattern of pure CDHA and triple-ion-substituted CDHA (ICDHA).

Figure 2

FT-IR spectra of pure CDHA and triple-ion-substituted CDHA (ICDHA).

Figure 3

(A) SEM image and (B) corresponding EDS spectra of ICDHA.

Figure 4

TEM image of (A) CDHA and (B) ICDHA.

Figure 5

VSM graph showing magnetization properties of CDHA and ICDHA.

Figure 6

(A) X-ray and (B) grayscale value graph of CDHA and ICDHA.

Figure 7

CT contrast of ICDHA samples compared with CDHA in HU (Hounsfield Units).

Figure 8

(A) T1-weighted MR image of ICDHA substituted samples dispersed at different concentrations in distilled water and (B) longitudinal relaxivity vs. concentration graph of ICDHA.

Figure 9

T2-weighted MR image of ICDHA substituted samples dispersed at different concentrations in distilled water.

Figure 10

Loading profile of tetracycline from ion-substituted CDHA (ICDHA) compared to pure CDHA (n = 3; data shown as mean ± SD; p < 0.05, one-way ANOVA).

Figure 11

Tetracycline release profile from ICDHA compared to that of CDHA (n = 3; data shown as mean ± SD; p < 0.05, one-way ANOVA).

Figure 12

Antibacterial activity of pure and tetracycline-loaded CDHA and ICDHA (n = 3; data shown as mean ± SD; p < 0.05, one-way ANOVA).

Figure 13

Biocompatibility of pure and tetracycline-loaded samples against Swiss 3T3 fibroblast cells by MTT assay for 24 and 48 h; TC, tetracycline (n = 3; data shown as mean ± SD; p < 0.05, two-way ANOVA).