research highlights

BIOLUMINESCENCE IMAGING Getting that inner glow in non-transgenic animals Y evtodiyenko, A. et al. Nat Commun 12, 2680 (2021)

Bioluminescent tools can give different Portable system (PBL) animals an inner glow that researchers can NNCO2H then take advantage of to image what’s going Cage on inside of their subjects. Bioluminescence X SS hν imaging (BLI) lacks some specificity that other imaging modalities offer, but the Caged luciferin probe plug detector imaging itself is relatively noninvasive and offers longitudinal monitoring of different The three components of the PBL. Credit: Yevtodiyenko, A. et al. (Springer Nature) 2021 biological processes. Getting the necessary components in to the animals can however be a bit more involved. The product of a biochemical reaction, bioluminescence probe – in this case, luciferin that has been clinical signs of toxicology observed in the comes from light generated when the rendered inert until the chemical bonds dogs’ blood serum, the team could then enzyme luciferase oxidizes its substrate, ‘caging’ it are broken by specific biological determine if the amount of bioluminescence luciferin. Luciferase and luciferin (as well processes or enzymes in the body. Next, from the luciferase corresponded to luciferin as many related variants) can be found in the animal is subcutaneously injected with concentrations. It did – all while requiring a number of – perhaps most a small plug that contains the luciferase 10x less luciferin than that needed in notably, the – but lab mice, and most enzyme. As the luciferin is uncaged, it will mice to see the same signal, says Goun. other animals for that matter, don’t come by flow through the blood stream and react “This means much lower doses of ‘caged’ the molecules naturally. with the luciferase as it passes by. A portable luciferin probes are needed, making Researchers that want to use light device developed by Goun and a team production of the compounds more bioluminescence imaging (BLI) must of international collaborators can then be practical,” she says. generate transgenic animals that express held above where the plug was injected, In a final proof of principle, the team luciferin in cells or tissues of interest, after where it will record any bioluminescence. injected a luciferase plug and three which they can be injected with luciferase The stronger the light signal, the greater the doses of luciferin into the arm of a human to prompt the light-generating chemical reaction in the animal. cadaver donated to the Swiss Federal reaction. The process works well and has Following validation of the components Institute of Technology Lausanne; the yielded valuable insights in many disease in vivo, the team tested the PBL’s ability to light detector picked up light for fifteen areas, but creating transgenic models detect an intracellular enzyme, cytochrome minutes, with a signal proportional to takes time, money, and extra animals. P450 (CYP450). CYP450 is a liver enzyme luciferin concentration. It’s also limited to those species that can involved in drug metabolism; testing for its The PBL system was years in the be genetically modified in the first place. activation is an important part of assessing making – Goun says she’d been dreaming The instruments involved can also be the toxicity of therapeutic candidates. of developing it since 2011, but it took time expensive and cumbersome – most BLI This is however a highly invasive process to find the optical engineers, including systems require a light-tight box and a that can involve many animals that must the National Institute of Standard and cooled charge-coupled device camera; these undergo multiple blood draws before being Technology’s Sergey Polyakov and can be constrained by subject size and are sacrificed for final analysis. The most George Eppeldauer, to collaborate with not always suitable for larger, non-rodent common isoform of CYP450, cytochrome on the necessary components. From this species. Such animals, including primates, P450 3A (Cyp3a) can be activated with first prototype device, she hopes to go canines, cats, and rabbits, are however injections of dexamethasone (DEX), so wireless with implantable sensors, similar important in research – particularly the team injected DEX into cohorts of to those used to record glucose levels. for toxicology and drug testing before luciferase-expressing and non-transgenic, The team is also exploring other kinds candidates can be moved on to clinical trials. wild-type FVB mice and then recorded for of caged luciferins. Elena Goun and her lab, now at the enzymatic activity under a conventional Meanwhile, Goun hopes the technology University of Missouri, Columbia, have BLI camera and the smaller PBL device. will prove useful for those labs that cannot been working to advance BLI technologies, They observed comparable readouts. They afford the expensive equipment currently with a motivation to help improve the then compared their imaging results with required for BLI. Avoiding the need for quality of animal experiments while conventional measures of Cyp3a activity, transgenic animals – including rodents – reducing the numbers needed. Writing in which involved sacrificing animals to should also help reduce the total number Nature Communications, she and colleagues analyze expression in their livers. PBL of animals needed for a BLI study, as will present a low cost (~500 to 600 USD), proved up to par. the ability to measure drug toxicity without portable BLI method that’s compatible with Moving on from mice, the team tried needing to sacrifice test subjects. non-transgenic animals. the PBL on a larger animal: dogs. First, The portable bioluminescent system they confirmed the system was indeed Ellen Nef (PBL) relies on injections, rather than safe for use in canines. Though luciferin is genetic modification. First, the subject extensively in mice without adverse effects, Published online: 14 June 2021 is intraperitoneally injected with a caged data wasn’t available for dogs. With no https://doi.org/10.1038/s41684-021-00798-9

168 Lab Animal | VOL 50 | July 2021 | 165–170 | www.nature.com/laban