Introduction.

The chemical and physical properties of O₂ facilitate a wide variety of methods for monitoring partial pressure of O₂ (pO₂), and include magnetic resonance (MR) based techniques. Changes in tissue O₂ concentrations produce changes in the relaxation rate R₁ (= 1/T₁) of water thus T₁ demonstrates a sensitivity to dissolved O₂ (which acts as a T₁-shortening paramagnetic probe). An O₂-induced increase in R₁ has the potential to provide measurements in fluctuations in the O₂ level of tissue. Jordan et al ¹ describes an MR method allowing for the rapid mapping of changes in tissue oxygenation based on the higher solubility of O₂ in lipids than in water (MOBILE). Lipid nanocapsules (LNCs) are core-shell based nanoparticles wherein an oil-filled core, capable of encapsulating hydrophobic drugs, is surrounded by a polyethylene glycol (PEG) layer, and used in therapeutic cancer treatments ². In this study, we used the MOBILE MR sequence to determine tissue hypoxia using lipids in the form of LNCs.

Methods.

LNCs were synthesized via the phase-inversion temperature method ³ and osmotically adjusted for in vivo use. Size and polydispersity (PDI) were determined by dynamic light scattering analysis. MR imaging was performed using a 7T scanner (Biospec 70/20 Avance III, Bruker Wissembourg, France) equipped with BGA12S gradient system (675mT/m). Prior to in vivo experimentation, the ability of the LNCs to respond to change in pO₂ was assessed in vitro using distinct samples with differing O₂ levels, namely 0 % (100 % argon), 21 %, and 100 % (corresponding to 0, 0.2, and 1 atm, respectively), and differing temperatures, from which lipid T₁ was measured using the MOBILE MR sequence ¹. For in vivo validation, LNCs were injected into the right hind femoral muscle (n = 5) of C3H mice and T₁ relaxation rates were measured whilst the animal was breathing air or undergoing a carbogen gas (95 % O₂, 5 % CO₂) challenge to induce a change in muscle pO₂. T₁ relaxation was determined by drawing a ROI around the lipid hypersignal and analyzing the T₁ relaxation rates' pixel-by-pixel using a home-made program written in MatLab (The MathWorks, Inc., Natick, MA) to discern a T₁ mean value and map of the muscle pO₂.

Results.

LNCs were synthesized with an average hydrodynamic size of 53.9 nm ±0.7 (PDI 0.04 ±0.01). For in vitro validation T₁ values of 306 ms, 420 ms and 450 ms were measured after bubbling in 100% O₂, air or argon, respectively at 37 °C. The pO₂ calibration curve was calculated from the values using the equation: R₁ = 3.0952 + 0.0115 * PO₂ - 0.0258 * T (°C) (Fig. 1). For in vivo validation, the osmotically adjusted LNC preparation was injected into femoral mouse muscle. In all explored animals a decrease in T₁ was observed following the carbogen challenge (mean T_{1 Air} = 390 ±4 ms and mean T_{1 Carbogen} = 372 ±7 ms, p = 0.04) (Fig. 2), and T₁ maps demonstrating the T₁ values of air and carbogen breathing were composed (Fig. 3).

Conclusion.

Preliminary results indicate that tissue pO₂ can be mapped using the MOBILE MR sequence after a single introduction of LNCs. The lipid T₁ map was sensitive to variations in oxygenation induced by a carbogen challenge.

Acknowledgements

The authors would like to thank the NanoFar Erasmus Mundus program and the ‘Comité Inter-Regional Grand Ouest de La Ligue Contre le Cancer’ (CIRGO) for providing funding for this project.