Background The incidence of diabetes and diabetic peripheral neuropathy continues to rise, and studies have shown that macrophages play an important role in their pathogenesis. Here we present a novel two-color fluorescently labeled perfluorocarbon nanoemulsion to monitor phagocytic macrophages in vitro and in vivo using non-invasive near infrared fluorescence imaging and fluorescence microscopy in diabetic neuropathy. We next applied this nanoemulsion to the Leprdb/db model of Type 2 Diabetes Mellitus to track macrophages along the length of the peripheral sensory pathway of the hindlimb, where diabetic peripheral neuropathy tends to originate.
Methods In vitro assessment of the nanoemulsion was performed by measuring fluorescent signal stability, colloidal stability, and macrophage uptake and subsequent viability. In vivo tracking of the nanoemulsion within Leprdb/db and wild-type mice was performed using both near infra-red fluorescent imaging and confocal microscopy to assess its biodistribution within phagocytic macrophages along the peripheral sensory apparatus of the hindlimb.
Results In vitro experiments show two-color nanoemulsion had high levels of fluorescent and colloidal stability, and that it was readily incorporated into RAW 264.7 macrophages. In vivo tracking revealed distribution of the two-color nanoemulsion to macrophages within most tissues of Leprdb/db and wild-type mice which persisted for several weeks, however it did not cross the blood brain barrier. Reduced fluorescence was seen in sciatic nerves of both Leprdb/db and wildtype mice, implying that the nanoemulsion may also have difficulty crossing an intact blood nerve barrier. Additionally, distribution of the nanoemulsion in Lepr db/db mice was reduced in several tissues as compared to wild-type mice. This reduction in biodistribution appears to be caused by the increased number of adipose tissue macrophages in Leprdb/db mice.
Conclusions The nanoemulsion in this study has the ability to identify phagocytic macrophages in the Leprdb/db model using both near infra-red fluorescent imaging and fluorescence microscopy. Based on the drug loading capacity of this new nanoemulsion and the role of inflammatory macrophages in diabetes, we foresee this agent being a useful tool in the assessment and treatment of diabetic peripheral neuropathy.