Beckman Laser Institute & Medical Clinic

Computed tomography (CT) scanners are essential for modern imaging but require around 600 projections from various angles. We present x-ray-induced acoustic computed tomography (XACT), a method that uses radiation-induced acoustic waves for three-dimensional (3D) x-ray imaging. These spherical acoustic waves travel through tissue at 1.5 × 10³ meters per second, much slower than x-rays, allowing ultrasound detectors to capture them and generate 3D images without mechanical scanning. We validate this theory by performing 3D numerical reconstructions of a human breast from a single x-ray projection and experimentally determining 3D structures of objects at different depths. Achieving resolutions of 0.4 millimeters in the XZ plane and 3.5 millimeters in the XY plane at a depth of 16 millimeters, XACT demonstrates the ability to produce 3D images from one x-ray projection, reducing radiation exposure and enabling gantry-free imaging. XACT shows great promise for biomedical and nondestructive testing applications, potentially replacing conventional CT.

BACKGROUND/OBJECTIVES: Although the use of radiation-sensitizing agents has been shown to enhance the effect of radiation on tumor cells, the blood-brain barrier (BBB) impedes these agents from reaching brain tumor sites when provided systemically. Localized methods of sensitizer delivery, utilizing hydrogels, have the potential to bypass the blood-brain barrier. This study examined the ability of photochemical internalization (PCI) of hydrogel-released bleomycin to enhance the growth-inhibiting effects of radiation on multi-cell glioma spheroids in vitro. METHODS: Loaded fibrin hydrogel layers were created by combining thrombin, fibrinogen, and bleomycin (BLM). Supernatants from these layers were collected, combined with photosensitizer, and added to F98 glioma spheroid cultures. Following light (PCI) and radiation treatment, at increasing dosages, spheroid growth was monitored for 14 days. RESULTS: PCI of released BLM significantly reduced the radiation dose required to achieve equivalent efficacy compared to radiation or BLM + RT alone. Both immediate and delayed RT delivery post-BLM-PCI resulted in similar degrees of growth inhibition. CONCLUSIONS: Non-degraded BLM was released from the fibrin hydrogel. PCI of BLM synergistically increased the growth-inhibiting effects of radiation treatment compared to radiation and BLM, as well as radiation acting as a single treatment.

To combat the rise of antibiotic-resistance in bacteria and the resulting effects on healthcare worldwide, new technologies are needed that can perform rapid antibiotic susceptibility testing (AST). Conventional clinical methods for AST rely on growth-based assays, which typically require long incubation times to obtain quantitative results, representing a major bottleneck in the determination of the optimal antibiotic regimen to treat patients. Here, we demonstrate a rapid AST method based on the metabolic activity measured by fluorescence lifetime imaging microscopy (FLIM). Using lab strains and clinical isolates of Escherichia coli with tetracycline-susceptible and resistant phenotypes as models, we demonstrate that changes in metabolic state associated with antibiotic susceptibility can be quantitatively tracked by FLIM. Our results show that the magnitude of metabolic perturbation resulting from antibiotic activity correlates with susceptibility evaluated by conventional metrics. Moreover, susceptible and resistant phenotypes can be differentiated in as short as 10 min after antibiotic exposure. This FLIM-AST (FAST) method can be applied to other antibiotics and provides insights into the nature of metabolic perturbations inside bacterial cells resulting from antibiotic exposure with single cell resolution.

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Severe brain injury can result in disorders of consciousness (DoC), including coma, vegetative state/unresponsive wakefulness syndrome, and minimally conscious state. Improved emergency and trauma medicine response, in addition to expanding efforts to prevent premature withdrawal of life-sustaining treatment, has led to an increased number of patients with prolonged DoC. High-quality bedside care of patients with DoC is key to improving long-term functional outcomes. However, there is a paucity of DoC-specific evidence guiding clinicians on efficacious bedside care that can promote medical stability and recovery of consciousness. This Viewpoint describes the state of current DoC bedside care and identifies knowledge and practice gaps related to patient care with DoC collated by the Care of the Patient in Coma scientific workgroup as part of the Neurocritical Care Society's Curing Coma Campaign. The gap analysis identified and organized domains of bedside care that could affect patient outcomes: clinical expertise, assessment and monitoring, timing of intervention, technology, family engagement, cultural considerations, systems of care, and transition to the post-acute continuum. Finally, this Viewpoint recommends future research and education initiatives to address and improve the care of patients with DoC.

The list of Curing Coma Campaign Collaborators included as a file in Supplementary Information has been updated. Flora Hammond, Raimund Helbok, and Naomi Niznick have been added to the list. Several spelling errors have been corrected. A revised file has been included.

The list of Curing Coma Campaign Collaborators included in the Acknowledgements section has been updated. Flora Hammond, Raimund Helbok, and Naomi Niznick have been added to the list. Several spelling errors have been corrected. A revised file has been included.

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We introduce a method utilizing single laser-generated cavitation bubbles to stimulate cellular mechanotransduction in dermal fibroblasts embedded within 3D hydrogels. We demonstrate that fibroblasts embedded in either amorphous or fibrillar hydrogels engage in Ca²⁺ signaling following exposure to an impulsive mechanical stimulus provided by a single 250 µm diameter laser-generated cavitation bubble. We find that the spatial extent of the cellular signaling is larger for cells embedded within a fibrous collagen hydrogel as compared to those embedded within an amorphous polyvinyl alcohol polymer (SLO-PVA) hydrogel. Additionally, for fibroblasts embedded in collagen, we find an increased range of cellular mechanosensitivity for cells that are polarized relative to the radial axis as compared to the circumferential axis. By contrast, fibroblasts embedded within SLO-PVA did not display orientation-dependent mechanosensitivity. Fibroblasts embedded in hydrogels and cultured in calcium-free media did not show cavitation-induced mechanotransduction; implicating calcium signaling based on transmembrane Ca²⁺ transport. This study demonstrates the utility of single laser-generated cavitation bubbles to provide local non-invasive impulsive mechanical stimuli within 3D hydrogel tissue models with concurrent imaging using optical microscopy. STATEMENT OF SIGNIFICANCE: Currently, there are limited methods for the non-invasive real-time assessment of cellular sensitivity to mechanical stimuli within 3D tissue scaffolds. We describe an original approach that utilizes a pulsed laser microbeam within a standard laser scanning microscope system to generate single cavitation bubbles to provide impulsive mechanostimulation to cells within 3D fibrillar and amorphous hydrogels. Using this technique, we measure the cellular mechanosensitivity of primary human dermal fibroblasts embedded in amorphous and fibrillar hydrogels, thereby providing a useful method to examine cellular mechanotransduction in 3D biomaterials. Moreover, the implementation of our method within a standard optical microscope makes it suitable for broad adoption by cellular mechanotransduction researchers and opens the possibility of high-throughput evaluation of biomaterials with respect to cellular mechanosignaling.