Subcellular Partitioning and Analysis of Gd³⁺-Loaded Ultrashort Single-Walled Carbon Nanotubes

Magnetic resonance imaging (MRI) is of vast clinical utility, with tens of millions of scans performed annually. Chemical contrast agents (CAs) can greatly enhance the diagnostic potential of MRI, and ∼50% of MRI scans use CAs. However, CAs have significant limitations such as low contrast enhancement, lack of specificity, and potential toxicity. Recently developed, Gd³⁺-loaded ultrashort single-walled carbon nanotubes, also referred to as gadonanotubes or GNTs, exhibit ∼40 times the relaxivities of clinical CAs, representing a potential major advance in clinically relevant MRI CA materials. Although initial cytotoxicity and MRI studies have suggested great promise for GNTs, relatively little is known regarding their subcellular interactions, which are crucial for further, safe development of GNTs as CAs. In this work, we administered GNTs to a well-established human cell line (HeLa) and to murine macrophage-like cells (J774A.1). GNTs were not acutely cytotoxic and did not reduce proliferation, except for the highest exposure concentration of 27 μg/mL for J774A.1 macrophages, yet bulk uptake of GNTs occurred in minutes at picogram quantities, or millions of GNTs per cell. J774A.1 macrophages internalized substantially more GNTs than HeLa cells in a dose-dependent manner, and Raman imaging of the subcellular distribution of GNTs revealed perinuclear localization. Fluorescence intensity and lifetime imaging demonstrated that GNTs did not grossly alter subcellular compartments, including filamentous-actin structures. Together, these results provide subcellular evidence necessary to establish GNTs as a new MRI CA material