Cellular Uptake and Delivery-Dependent Effects of Tb³⁺-Doped Hydroxyapatite Nanorods

With the increasing interest in hydroxyapatite (HA) nanostructures for use in biomedicine, the systematic evaluation of their potential effects on biological systems is becoming critically important. In this work, we report the in vitro cellular uptake, in vivo tissue distributions and toxicity of Tb³⁺-doped HA (HA-Tb) after short-, intermediate-, and long-term exposure. Transmission electron microscopy analysis indicated that HA-Tb was taken up by cells via vesicle endocytosis. Cell proliferation and cytotoxicity assay, combined with confocal laser scanning microscopy, indicated excellent cell viability with no changes in cell morphology at the examined doses. Three HA-Tb delivery methods (intraperitoneal, intragastric, and intravenous) resulted in similar time-dependent tissue distributions, while intraperitoneal injection produced the highest bioavailability. HA-Tb initially accumulated in livers and intestines of rats (4 h to one day after administration), then became increasingly distributed in the kidney and bladder (seven days), and finally decreased in all tissues after 30 to 90 days. No histopathological abnormalities or lesions related to treatment with HA-Tb were observed. These results suggest that HA-Tb has minimal in vitro and in vivo toxicity, regardless of the delivery mode, time, and dose. The findings provide a foundation for the design and development of HA for biological applications.