Advanced deep-tissue imaging and manipulation enabled by biliverdin reductase knockout.

We developed near-infrared (NIR) photoacoustic and fluorescence probes, as well as optogenetic tools from bacteriophytochromes, and enhanced their performance using biliverdin reductase-A knock-out model (Blvra-/-). Blvra-/- elevates endogenous heme-derived biliverdin chromophore for bacteriophytochrome-derived NIR constructs. Consequently, light-controlled transcription with IsPadC-based optogenetic tool improved up to 25-fold compared to wild-type cells, with 100-fold activation in Blvra-/- neurons. In vivo , light-induced insulin production in Blvra-/- reduced blood glucose in diabetes by ∼60%, indicating high potential for optogenetic therapy. Using 3D photoacoustic, ultrasound, and two-photon fluorescence imaging, we overcame depth limitations of recording NIR probes. We achieved simultaneous photoacoustic imaging of DrBphP in neurons and super-resolution ultrasound localization microscopy of blood vessels ∼7 mm deep in the brain, with intact scalp and skull. Two-photon microscopy provided cell-level resolution of miRFP720-expressing neurons ∼2.2 mm deep. Blvra-/- significantly enhances efficacy of biliverdin-dependent NIR systems, making it promising platform for interrogation and manipulation of biological processes.

Subventricular zone stem cell niche injury is associated with intestinal perforation in preterm infants and predicts future motor impairment.

Brain injury is highly associated with preterm birth. Complications of prematurity, including spontaneous or necrotizing enterocolitis (NEC)-associated intestinal perforations, are linked to lifelong neurologic impairment, yet the mechanisms are poorly understood. Early diagnosis of preterm brain injuries remains a significant challenge. Here, we identified subventricular zone echogenicity (SVE) on cranial ultrasound in preterm infants following intestinal perforations. The development of SVE was significantly associated with motor impairment at 2 years. SVE was replicated in a neonatal mouse model of intestinal perforation. Examination of the murine echogenic subventricular zone (SVZ) revealed NLRP3-inflammasome assembly in multiciliated FoxJ1+ ependymal cells and a loss of the ependymal border in this postnatal stem cell niche. These data suggest a mechanism of preterm brain injury localized to the SVZ that has not been adequately considered. Ultrasound detection of SVE may serve as an early biomarker for neurodevelopmental impairment after inflammatory disease in preterm infants.

The sound of blood: photoacoustic imaging in blood analysis.

Blood analysis is a ubiquitous and critical aspect of modern medicine. Analyzing blood samples requires invasive techniques, various testing systems, and samples are limited to relatively small volumes. Photoacoustic imaging (PAI) is a novel imaging modality that utilizes non-ionizing energy that shows promise as an alternative to current methods. This paper seeks to review current applications of PAI in blood analysis for clinical use. Furthermore, we discuss obstacles to implementation and future directions to overcome these challenges. Firstly, we discuss three applications to cellular analysis of blood: sickle cell, bacteria, and circulating tumor cell detection. We then discuss applications to the analysis of blood plasma, including glucose detection and anticoagulation quantification. As such, we hope this article will serve as inspiration for PAI's potential application in blood analysis and prompt further studies to ultimately implement PAI into clinical practice.

Three-dimensional non-invasive brain imaging of ischemic stroke by integrated photoacoustic, ultrasound and angiographic tomography (PAUSAT).

We present an ischemic stroke study using our newly-developed PAUSAT system that integrates photoacoustic computed tomography (PACT), high-frequency ultrasound imaging, and acoustic angiographic tomography. PAUSAT is capable of three-dimensional (3D) imaging of the brain morphology, blood perfusion, and blood oxygenation. Using PAUSAT, we studied the hemodynamic changes in the whole mouse brain induced by two common ischemic stroke models: the permanent middle cerebral artery occlusion (pMCAO) model and the photothrombotic (PT) model. We imaged the same mouse brains before and after stroke, and quantitatively compared the two stroke models. We observed clear hemodynamic changes after ischemic stroke, including reduced blood perfusion and oxygenation. Such changes were spatially heterogenous. We also quantified the tissue infarct volume in both stroke models. The PAUSAT measurements were validated by laser speckle imaging and histology. Our results have collectively demonstrated that PAUSAT can be a valuable tool for non-invasive longitudinal studies of neurological diseases at the whole-brain scale.

Integrated Photoacoustic, Ultrasound, and Angiographic Tomography (PAUSAT) for NonInvasive Whole-Brain Imaging of Ischemic Stroke.

Presented here is an experimental ischemic stroke study using our newly developed noninvasive imaging system that integrates three acoustic-based imaging technologies: photoacoustic, ultrasound, and angiographic tomography (PAUSAT). Combining these three modalities helps acquire multi-spectral photoacoustic tomography (PAT) of the brain blood oxygenation, high-frequency ultrasound imaging of the brain tissue, and acoustic angiography of the cerebral blood perfusion. The multi-modal imaging platform allows the study of cerebral perfusion and oxygenation changes in the whole mouse brain after stroke. Two commonly used ischemic stroke models were evaluated: the permanent middle cerebral artery occlusion (pMCAO) model and the photothrombotic (PT) model. PAUSAT was used to image the same mouse brains before and after a stroke and quantitatively analyze both stroke models. This imaging system was able to clearly show the brain vascular changes after ischemic stroke, including significantly reduced blood perfusion and oxygenation in the stroke infarct region (ipsilateral) compared to the uninjured tissue (contralateral). The results were confirmed by both laser speckle contrast imaging and triphenyltetrazolium chloride (TTC) staining. Furthermore, stroke infarct volume in both stroke models was measured and validated by TTC staining as the ground truth. Through this study, we have demonstrated that PAUSAT can be a powerful tool in noninvasive and longitudinal preclinical studies of ischemic stroke.

Multiscale photoacoustic tomography using reversibly switchable thermochromics.

Significance: Based on acoustic detection of optical absorption, photoacoustic tomography (PAT) allows functional and molecular imaging beyond the optical diffusion limit with high spatial resolution. However, multispectral functional and molecular PAT is often limited by decreased spectroscopic accuracy and reduced detection sensitivity in deep tissues, mainly due to wavelength-dependent optical attenuation and inaccurate acoustic inversion. Aim: Previous work has demonstrated that reversible color-shifting can drastically improve the detection sensitivity of PAT by suppressing nonswitching background signals. We aim to develop a new color switching-based PAT method using reversibly switchable thermochromics (ReST). Approach: We developed a family of ReST with excellent water dispersion, biostability, and temperature-controlled color changes by surface modification of commercial thermochromic microcapsules with the hydrophilic polysaccharide alginate. Results: The optical absorbance of the ReST was switched on and off repeatedly by modulating the surrounding temperature, allowing differential photoacoustic detection that effectively suppressed the nonswitching background signal and substantially improved image contrast and detection sensitivity. We demonstrate reversible thermal-switching imaging of ReST in vitro and in vivo using three PAT modes at different length scales. Conclusions: ReST-enabled PAT is a promising technology for high-sensitivity deep tissue imaging of molecular activity in temperature-related biomedical applications, such as cancer thermotherapy.

Plasmonic nanorod probes' journey inside plant cells for in vivo SERS sensing and multimodal imaging.

Nanoparticle-based platforms are gaining strong interest in plant biology and bioenergy research to monitor and control biological processes in whole plants. However, in vivo monitoring of biomolecules using nanoparticles inside plant cells remains challenging due to the impenetrability of the plant cell wall to nanoparticles beyond the exclusion limits (5-20 nm). To overcome this physical barrier, we have designed unique bimetallic silver-coated gold nanorods (AuNR@Ag) capable of entering plant cells, while conserving key plasmonic properties in the near-infrared (NIR). To demonstrate cellular internalization and tracking of the nanorods inside plant tissue, we used a comprehensive multimodal imaging approach that included transmission electron microscopy (TEM), confocal fluorescence microscopy, two-photon luminescence (TPL), X-ray fluorescence microscopy (XRF), and photoacoustics imaging (PAI). We successfully acquired SERS signals of nanorods in vivo inside plant cells of tobacco leaves. On the same leaf samples, we applied orthogonal imaging methods, TPL and PAI techniques for in vivo imaging of the nanorods. This study first demonstrates the intracellular internalization of AuNR@Ag inside whole plant systems for in vivo SERS analysis in tobacco cells. This work demonstrates the potential of this nanoplatform as a new nanotool for intracellular in vivo biosensing for plant biology.

Sound out the impaired perfusion: Photoacoustic imaging in preclinical ischemic stroke.

Acoustically detecting the optical absorption contrast, photoacoustic imaging (PAI) is a highly versatile imaging modality that can provide anatomical, functional, molecular, and metabolic information of biological tissues. PAI is highly scalable and can probe the same biological process at various length scales ranging from single cells (microscopic) to the whole organ (macroscopic). Using hemoglobin as the endogenous contrast, PAI is capable of label-free imaging of blood vessels in the brain and mapping hemodynamic functions such as blood oxygenation and blood flow. These imaging merits make PAI a great tool for studying ischemic stroke, particularly for probing into hemodynamic changes and impaired cerebral blood perfusion as a consequence of stroke. In this narrative review, we aim to summarize the scientific progresses in the past decade by using PAI to monitor cerebral blood vessel impairment and restoration after ischemic stroke, mostly in the preclinical setting. We also outline and discuss the major technological barriers and challenges that need to be overcome so that PAI can play a more significant role in preclinical stroke research, and more importantly, accelerate its translation to be a useful clinical diagnosis and management tool for human strokes.