Bioluminescence is a natural light source based on luciferase catalysis of its substrate luciferin. We performed directed evolution on firefly luciferase using a red-shifted and highly deliverable luciferin analog to establish AkaBLI, an all-engineered bioluminescence in vivo imaging system. AkaBLI produced emissions in vivo that were brighter by a factor of 100 to 1000 than conventional systems, allowing noninvasive visualization of single cells deep inside freely moving animals. Single tumorigenic cells trapped in the mouse lung vasculature could be visualized. In the mouse brain, genetic labeling with neural activity sensors allowed tracking of small clusters of hippocampal neurons activated by novel environments. In a marmoset, we recorded video-rate bioluminescence from neurons in the striatum, a deep brain area, for more than 1 year. AkaBLI is therefore a bioengineered light source to spur unprecedented scientific, medical, and industrial applications.

In preclinical cancer research, bioluminescence imaging with firefly luciferase and D-luciferin has become a standard to monitor biological processes both in vitro and in vivo. However, the emission maximum (λmax) of bioluminescence produced by D-luciferin is 562 nm where light is not highly penetrable in biological tissues. This emphasizes a need for developing a red-shifted bioluminescence imaging system to improve detection sensitivity of targets in deep tissue. Here we characterize the bioluminescent properties of the newly synthesized luciferin analogue, AkaLumine-HCl. The bioluminescence produced by AkaLumine-HCl in reactions with native firefly luciferase is in the near-infrared wavelength ranges (λmax=677 nm), and yields significantly increased target-detection sensitivity from deep tissues with maximal signals attained at very low concentrations, as compared with D-luciferin and emerging synthetic luciferin CycLuc1. These characteristics offer a more sensitive and accurate method for non-invasive bioluminescence imaging with native firefly luciferase in various animal models.

Protein cages have been utilized as templates in the development of biomaterials. Here we report protein engineering of the ferritin (Fr) cage for encapsulating carbon monoxide releasing molecules (CORMs) and release of CO gas which serves as a cell signaling molecule. The protein cages enable us to increase the half-life for CO release, providing a release rate that is 18-fold slower than the rate of a typical CORM, Ru(CO)3Cl(glycinate) (CORM-3). Moreover, the uptake ratio of the composite is about 4-fold greater than that of CORM-3. We found that these effects enhance the activation of nuclear factor κB 10-fold higher than CORM-3. The protein cage of Fr thus provides the basis for new CORMs that can be used for in vitro cell research.

The western pattern diet is rich not only in fat and calorie but also in phosphate. Negative impacts of excessive fat and calorie intake on health are widely accepted, whereas potential harms of excessive phosphate intake are poorly recognized. Here we show the mechanism by which dietary phosphate damages the kidney. When phosphate intake was excessive relative to the functioning nephron number, circulating fibroblast growth factor-23 (FGF23), a hormone that increases phosphate excretion per nephron, was increased to maintain phosphate homeostasis.FGF23 suppressed phosphate reabsorption in renal tubules and thus raised the phosphate concentration in the tubular fluid. Once it exceeded a threshold, microscopic particles containing calcium phosphate crystals appeared in the tubular lumen, which damaged tubular cells through binding to Toll-like receptor-4 expressed on them. Persistent tubular damage induced interstitial fibrosis, reduced the nephron number, and further boosted FGF23 to trigger a deterioration spiral leading to progressive nephron loss. In humans, progression of chronic kidney disease (CKD) ensued when the serum FGF23 level exceeded 53 pg/mL. The present study identified the calcium phosphate particles in the renal tubular fluid as an effective therapeutic target to decelerate nephron loss during the course of aging and CKD progression.

Although the current murine model of bone metastasis using intracardiac (IC) injection successfully recapitulates the process of bone metastasis, further progress in the study of bone metastasis requires a new model to circumvent some limitations of this model. Here, we present a new murine model of bone metastasis achieved by injecting cancer cells through the intra-caudal arterial (CA). This model does not require high technical proficiency, predominantly delivers cancer cells to bone marrow of hind limbs with much higher efficiency than IC injection, and greatly shortens the period of overt bone metastasis development. Moreover, CA injection barely causes acute death of mice, enabling us to inject a larger number of cancer cells to further accelerate the development of bone metastasis with a wide variety of cell lines. Our model may open a new avenue for understanding the bone metastatic processes and development of drugs preventing bone metastasis and recurrence.