Figure 6.

Antares2 expression in vivo following injection of Antares2 mRNA-loaded exosomes.A, combined light and bioluminescence images showing DTZ-triggered, Antares2-catalyzed light emission from the eye of mice that had been injected intravitreally with exosome/mRNA formulation and then exposed to topical DTZ solution 36 h later. Light emission, measured in radiance (photons/s/area (cm²)/steradian), is reflected in the color, with red denoting the areas of highest light emission and blue denoting the areas of lower but still significant light emission. Similar results were observed in a second animal that was subjected to the same treatment. Bar, 1 cm. B, combined light and bioluminescence images showing DTZ-triggered, Antares2-catalyzed light emission from the muscle of mice that had been injected i.m. in the left thigh with exosome/mRNA formulation and reinjected in both legs i.m. with DTZ solution 48 h later. This experiment was performed in a single animal. Bar, 1 cm. C, immunoblot of cell lysates prepared from HEK293 cells (HEK) and from cells that had been transfected 2 days earlier with plasmids designed to express Antares2-2a and Antares2-myc, using anti-mKate antibodies to detect the products of Antares2 expression and anti-HSP90 antibodies to confirm the presence of HEK293 proteins in each sample. D, immunoblot of tissue protein extracts collected 2 days after mice had been injected i.m. with either mRNA-loaded exosomes (animals 1 and 2) or PBS (control). Tissue protein samples were probed with (upper blots) antibodies specific for mKate to detect Antares2 and with (lower blots) antibodies specific for HSP90. Arrowheads denote position of Antares2 proteins detected by the anti-mKate antibodies in skeletal muscle of treated animals.