Numerous light-based diagnostic and therapeutic devices are routinely used in the clinic. These devices have a familiar look as items plugged in the wall or placed at patients' bedsides, but recently, many new ideas have been proposed for the realization of implantable or wearable functional devices. Many advances are being fuelled by the development of multifunctional materials for photonic healthcare devices. However, the finite depth of light penetration in the body is still a serious constraint for their clinical applications. In this Review, we discuss the basic concepts and some examples of state-of-the-art implantable and wearable photonic healthcare devices for diagnostic and therapeutic applications. First, we describe emerging multifunctional materials critical to the advent of next-generation implantable and wearable photonic healthcare devices and discuss the path for their clinical translation. Then, we examine implantable photonic healthcare devices in terms of their properties and diagnostic and therapeutic functions. We next describe exemplary cases of noninvasive, wearable photonic healthcare devices across different anatomical applications. Finally, we discuss the future research directions for the field, in particular regarding mobile healthcare and personalized medicine.

Multifunctional materials for implantable and wearable photonic healthcare devices.

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.

http://science.sciencemag.org/content/sci/359/6378/935.full.pdf

Single-cell bioluminescence imaging of deep tissue in freely moving animals

Naturally occurring protein switches have been repurposed for the development of biosensors and reporters for cellular and clinical applications1. However, the number of such switches is limited, and reengineering them is challenging. Here we show that a general class of protein-based biosensors can be created by inverting the flow of information through de novo designed protein switches in which the binding of a peptide key triggers biological outputs of interest2. The designed sensors are modular molecular devices with a closed dark state and an open luminescent state; analyte binding drives the switch from the closed to the open state. Because the sensor is based on the thermodynamic coupling of analyte binding to sensor activation, only one target binding domain is required, which simplifies sensor design and allows direct readout in solution. We create biosensors that can sensitively detect the anti-apoptosis protein BCL-2, the IgG1 Fc domain, the HER2 receptor, and Botulinum neurotoxin B, as well as biosensors for cardiac troponin I and an anti-hepatitis B virus antibody with the high sensitivity required to detect these molecules clinically. Given the need for diagnostic tools to track the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)3, we used the approach to design sensors for the SARS-CoV-2 spike protein and antibodies against the membrane and nucleocapsid proteins. The former, which incorporates a de novo designed spike receptor binding domain (RBD) binder4, has a limit of detection of 15 pM and a luminescence signal 50-fold higher than the background level. The modularity and sensitivity of the platform should enable the rapid construction of sensors for a wide range of analytes, and highlights the power of de novo protein design to create multi-state protein systems with new and useful functions. A modular de novo designed biosensor platform consisting of a cage and key molecule is developed, and used to create sensors for seven distinct proteins including the spike protein from SARS-CoV-2 and anti-SARS antibodies.

/pdf/de-novo-design-of-modular-and-tunable-protein-biosensors-2ef8wakjwu.pdf

De novo design of modular and tunable protein biosensors.

Although fluorescent reporters and biosensors have become indispensable tools in biological and biomedical fields, fluorescence measurements require external excitation light, thereby limiting their use in thick tissues and live animals. Bioluminescent reporters and biosensors may potentially overcome this hurdle because they use enzyme-catalyzed exothermic biochemical reactions to generate excited-state emitters. This review first introduces the development of bioluminescent reporters, and next, their applications in sensing biological changes in vitro and in vivo as biosensors. Lastly, we discuss chemiluminescent sensors that produce photons in the absence of luciferases. This review aims to explore fundamentals and experimental insights and to emphasize the yet-to-be-reached potential of next-generation luminescent reporters and biosensors.

Development and Applications of Bioluminescent and Chemiluminescent Reporters and Biosensors

Spontaneous transient depolarizations in mitochondrial membrane potential (Δψ m ), mitochondrial flickers, have been observed in isolated mitochondria and intact cells using the fluorescent probe, tetramethylrhodamine ethyl ester (TMRE). In theory, the ratio of [TMRE] in cytosol and mitochondrion allows Δψ m to be calculated with the Nernst equation, but this has proven difficult in practice due to fluorescence quenching and binding of dye to mitochondrial membranes. We developed a new method to determine the amplitude of flickers in terms of millivolts of depolarization. TMRE fluorescence was monitored using high-speed, high-sensitivity three-dimensional imaging to track individual mitochondria in freshly dissociated smooth muscle cells. Resting mitochondrial fluorescence, an exponential function of resting Δψ m , varied among mitochondria and was approximately normally distributed. Spontaneous changes in mitochondrial fluorescence, indicating depolarizations and repolarizations in Δψ m , were observed. The depolarizations were reversible and did not result in permanent depolarization of the mitochondria. The magnitude of the flickers ranged from 100 mV with a mean of 17.6 ′ 1.0 mV (n = 360) and a distribution skewed to smaller values. Nearly all mitochondria flickered, and they did so independently of one another, indicating that mitochondria function as independent units in the myocytes employed here.

Quantitative analysis of Spontaneous mitochondrial depolarizations

Red-shifted bioluminescence reporters are desirable for biological imaging. We describe the development of red-shifted luciferins based on synthetic coelenterazine analogs and corresponding mutants of NanoLuc that enable bright bioluminescence. One pair in particular showed superior in vitro and in vivo sensitivity over commonly used bioluminescence reporters. We adapted this pair to develop a bioluminescence resonance-energy-based Antares reporter called Antares2, which offers improved signal from deep tissues.

https://escholarship.org/content/qt3jd2f6kn/qt3jd2f6kn.pdf?t=pbah0c

Red-shifted luciferase–luciferin pairs for enhanced bioluminescence imaging

Mitochondrial membrane potential (MMP) is a frequently used indicator for mitochondrial function Herein, we report a photostable near-infrared (NIR) fluorescent dye for monitoring MMP This new probe, named NIMAP, is non-fluorescent in aqueous solution and can be activated by cell membranes, providing high fluorescence contrast and low background fluorescence NIMAP has been validated for monitoring MMP in living mammalian cells and in mice Due to the large fluorescence response, low fluorescence background, high photostability, and excellent tissue penetration resulting from red-shifted excitation and emission in the "optical window" above 600 nm, broad applications of this new probe are expected

/pdf/a-membrane-activatable-near-infrared-fluorescent-probe-with-3bmgwlart9.pdf

A membrane-activatable near-infrared fluorescent probe with ultra-photostability for mitochondrial membrane potentials

Exposure to cigarette smoke represents the largest source of preventable death and disease in the United States. This may be in part due to the nature of the delayed harmful effects as well as the lack of awareness of the scope of harm presented by these products. The presence of “light” versions further clouds the harmful effects of tobacco products. While active smoking in expectant mothers may be reduced by educational and outreach campaigns, exposure to secondhand smoke is often involuntary yet may harm the developing embryo. In this study, we show that the main component of secondhand smoke, sidestream cigarette smoke, from several brands, including harm-reduction versions, triggered unsuccessful hatching at 3 dpf and reduced overall survival at 6 dpf in developing zebrafish. At non-lethal concentrations, craniofacial defects with different severity based on the cigarette smoke extract were noted by 6 dpf. All tested products, including harm-reduction products, significantly impacted cartilage formation and/or bone mineralization in zebrafish embryos, independent of whether the bones/cartilage formed from the mesoderm or neural crest. Together, these results in a model system often used to detect embryonic malformations imply that exposure of a woman to secondhand smoke while pregnant may lead to mineralization issues in the skeleton of her newborn, ultimately adding a direct in utero association to the increased fracture risk observed in children of mothers exposed to cigarette smoke.

/pdf/embryonic-exposure-to-cigarette-smoke-extract-impedes-1iju75o6.pdf

Omran Karmach

Papers

Red-shifted luciferase–luciferin pairs for enhanced bioluminescence imaging

A membrane-activatable near-infrared fluorescent probe with ultra-photostability for mitochondrial membrane potentials

Embryonic Exposure to Cigarette Smoke Extract Impedes Skeletal Development and Evokes Craniofacial Defects in Zebrafish

Hyperglycemia Induced Oxidative Stress Alters the FOXO/AMPK Pathway Which Leads to Loss of Pluripotency

Chapter 10 Prostate Cancer and the Therapeutic Potential of Pomegranate