Femtosecond Electronic and Hydrogen Structural Dynamics in Ammonia Imaged with Ultrafast Electron Diffraction.

Directly imaging structural dynamics involving hydrogen atoms by ultrafast diffraction methods is complicated by their low scattering cross sections. Here we demonstrate that megaelectronvolt ultrafast electron diffraction is sufficiently sensitive to follow hydrogen dynamics in isolated molecules. In a study of the photodissociation of gas phase ammonia, we simultaneously observe signatures of the nuclear and corresponding electronic structure changes resulting from the dissociation dynamics in the time-dependent diffraction. Both assignments are confirmed by ab initio simulations of the photochemical dynamics and the resulting diffraction observable. While the temporal resolution of the experiment is insufficient to resolve the dissociation in time, our results represent an important step towards the observation of proton dynamics in real space and time.

Multichannel photodissociation dynamics in CS2 studied by ultrafast electron diffraction.

The structural dynamics of photoexcited gas-phase carbon disulfide (CS2) molecules are investigated using ultrafast electron diffraction. The dynamics were triggered by excitation of the optically bright 1B2(1Σu+) state by an ultraviolet femtosecond laser pulse centred at 200 nm. In accordance with previous studies, rapid vibrational motion facilitates a combination of internal conversion and intersystem crossing to lower-lying electronic states. Photodissociation via these electronic manifolds results in the production of CS fragments in the electronic ground state and dissociated singlet and triplet sulphur atoms. The structural dynamics are extracted from the experiment using a trajectory-fitting filtering approach, revealing the main characteristics of the singlet and triplet dissociation pathways. Finally, the effect of the time-resolution on the experimental signal is considered and an outlook to future experiments provided.

Transient vibration and product formation of photoexcited CS2 measured by time-resolved x-ray scattering.

We have observed details of the internal motion and dissociation channels in photoexcited carbon disulfide (CS2) using time-resolved x-ray scattering (TRXS). Photoexcitation of gas-phase CS2 with a 200 nm laser pulse launches oscillatory bending and stretching motion, leading to dissociation of atomic sulfur in under a picosecond. During the first 300 fs following excitation, we observe significant changes in the vibrational frequency as well as some dissociation of the C-S bond, leading to atomic sulfur in the both 1D and 3P states. Beyond 1400 fs, the dissociation is consistent with primarily 3P atomic sulfur dissociation. This channel-resolved measurement of the dissociation time is based on our analysis of the time-windowed dissociation radial velocity distribution, which is measured using the temporal Fourier transform of the TRXS data aided by a Hough transform that extracts the slopes of linear features in an image. The relative strength of the two dissociation channels reflects both their branching ratio and differences in the spread of their dissociation times. Measuring the time-resolved dissociation radial velocity distribution aids the resolution of discrepancies between models for dissociation proposed by prior photoelectron spectroscopy work.

Miniaturizing Transmon Qubits Using van der Waals Materials.

Quantum computers can potentially achieve an exponential speedup versus classical computers on certain computational tasks, recently demonstrated in superconducting qubit processors. However, the capacitor electrodes that comprise these qubits must be large in order to avoid lossy dielectrics. This tactic hinders scaling by increasing parasitic coupling among circuit components, degrading individual qubit addressability, and limiting the spatial density of qubits. Here, we take advantage of the unique properties of van der Waals (vdW) materials to reduce the qubit area by >1000 times while preserving the capacitance while maintaining quantum coherence. Our qubits combine conventional aluminum-based Josephson junctions with parallel-plate capacitors composed of crystalline layers of superconducting niobium diselenide and insulating hexagonal boron nitride. We measure a vdW transmon T1 relaxation time of 1.06 µs, demonstrating a path to achieve high-qubit-density quantum processors with long coherence times, and the broad utility of layered heterostructures in low-loss, high-coherence quantum devices.

Can We Accurately Identify Peritoneal Metastases Based on Their Appearance? An Assessment of the Current Practice of Intraoperative Gastrointestinal Cancer Staging.

BACKGROUND: Peritoneal lesions are common findings during operative abdominal cancer staging. The decision to perform biopsy is made subjectively by the surgeon, a practice the authors hypothesized to be imprecise. This study aimed to describe optical characteristics differentiating benign peritoneal lesions from peritoneal metastases. METHODS: The study evaluated laparoscopic images of 87 consecutive peritoneal lesions biopsied during staging laparoscopies for gastrointestinal malignancies from 2014 to 2017. A blinded survey assessing these lesions was completed by 10 oncologic surgeons. Three senior investigators categorized optical features of the lesions. Computer-aided digital image processing and machine learning was used to classify the lesions. RESULTS: Of the 87 lesions, 28 (32%) were metastases. On expert survey, surgeons on the average misidentified 36 ± 19% of metastases. Multivariate analysis identified degree of nodularity, border transition, and degree of transparency as independent predictors of metastases (each p < 0.03), with an area under the receiver operating characteristics curve (AUC) of 0.82 (95% confidence interval [CI], 0.72-0.91). Image processing demonstrated no difference using image color segmentation, but showed a difference in gradient magnitude between benign and metastatic lesions (AUC, 0.66; 95% CI 0.54-0.78; p = 0.02). Machine learning using a neural network with a tenfold cross-validation obtained an AUC of only 0.47. CONCLUSIONS: To date, neither experienced oncologic surgeons nor computerized image analysis can differentiate peritoneal metastases from benign peritoneal lesions with an accuracy that is clinically acceptable. Although certain features correlate with the presence of metastases, a substantial overlap in optical appearance exists between benign and metastatic peritoneal lesions. Therefore, this study suggested the need to perform biopsy for all peritoneal lesions during operative staging, or at least to lower the threshold significantly.

On the limits of observing motion in time-resolved X-ray scattering.

Limits on the ability of time-resolved X-ray scattering (TRXS) to observe harmonic motion of amplitude, A and frequency, ω0, about an equilibrium position, R0, are considered. Experimental results from a TRXS experiment at the LINAC Coherent Light Source are compared to classical and quantum theories that demonstrate a fundamental limitation on the ability to observe the amplitude of motion. These comparisons demonstrate dual limits on the spatial resolution through Qmax and the temporal resolution through ωmax for observing the amplitude of motion. In the limit where ωmax ≈ ω0, the smallest observable amplitude of motion is A = 2 π/ Qmax. In the limit where ωmax≥2 ω0, A≤2 π/ Qmax is observable provided there are sufficient statistics. This article is part of the theme issue 'Measurement of ultrafast electronic and structural dynamics with X-rays'.

Improved, Shorter-Latency Carcinogen-Induced Hepatocellular Carcinoma Model in Pigs.

Large animal models are important tools for hepatocellular carcinoma (HCC) research, especially in studies of hepatic vasculature, interventional techniques, and radiofrequency or microwave hyperthermia. Currently, diethylnitrosamine (DENA)-induced HCC in pigs is the only large animal model for in situ HCC with a tumor latency of 10-26 months. While phenobarbital (PB) is often used to accelerate DENA-induced HCC in rodents, it has not been previously studied in the porcine model. Therefore, we hypothesize that the addition of PB in the DENA-induced HCC porcine model will accelerate tumor latency compared to DENA alone. HCC and benign lesions were seen on serial MRI and confirmed on histopathology. Liver and tumors were further characterized by CT angiography, vascular corrosion casting, and permittivity measurements.

Observation of Quantum Interferences via Light-Induced Conical Intersections in Diatomic Molecules.

We observe energy-dependent angle-resolved diffraction patterns in protons from strong-field dissociation of the molecular hydrogen ion H_{2}^{+}. The interference is a characteristic of dissociation around a laser-induced conical intersection (LICI), which is a point of contact between two surfaces in the dressed two-dimensional Born-Oppenheimer potential energy landscape of a diatomic molecule in a strong laser field. The interference magnitude and angular period depend strongly on the energy difference between the initial state and the LICI, consistent with coherent diffraction around a cone-shaped potential barrier whose width and thickness depend on the relative energy of the initial state and the cone apex. These findings are supported by numerical solutions of the time-dependent Schrödinger equation for similar experimental conditions.

Multiscale benchmarking of drug delivery vectors.

Cross-system comparisons of drug delivery vectors are essential to ensure optimal design. An in-vitro experimental protocol is presented that separates the role of the delivery vector from that of its cargo in determining the cell response, thus allowing quantitative comparison of different systems. The technique is validated through benchmarking of the dose-response of human fibroblast cells exposed to the cationic molecule, polyethylene imine (PEI); delivered as a free molecule and as a cargo on the surface of CdSe nanoparticles and Silica microparticles. The exposure metrics are converted to a delivered dose with the transport properties of the different scale systems characterized by a delivery time, τ. The benchmarking highlights an agglomeration of the free PEI molecules into micron sized clusters and identifies the metric determining cell death as the total number of PEI molecules presented to cells, determined by the delivery vector dose and the surface density of the cargo.

Photon counting compressive depth mapping.

We demonstrate a compressed sensing, photon counting lidar system based on the single-pixel camera. Our technique recovers both depth and intensity maps from a single under-sampled set of incoherent, linear projections of a scene of interest at ultra-low light levels around 0.5 picowatts. Only two-dimensional reconstructions are required to image a three-dimensional scene. We demonstrate intensity imaging and depth mapping at 256 × 256 pixel transverse resolution with acquisition times as short as 3 seconds. We also show novelty filtering, reconstructing only the difference between two instances of a scene. Finally, we acquire 32 × 32 pixel real-time video for three-dimensional object tracking at 14 frames-per-second.

RND Pump-Mediated Efflux of Amotosalen, a Compound Used in Pathogen Inactivation Technology to Enhance Safety of Blood Transfusion Products, May Compromise Its Gram-Negative Anti-Bacterial Activity.

Pathogen inactivation is a strategy to improve the safety of transfusion products. The only pathogen reduction technology for blood products currently approved in the US utilizes a psoralen compound, called amotosalen, in combination with UVA light to inactivate bacteria, viruses, and protozoa. Psoralens have structural similarity to bacterial multidrug efflux pump substrates. As these efflux pumps are often overexpressed in multidrug-resistant pathogens, we tested whether contemporary drug-resistant pathogens might show resistance to amotosalen and other psoralens based on multidrug efflux mechanisms through genetic, biophysical, and molecular modeling analysis. The main efflux systems in Enterobacterales, Acinetobacter baumannii, and Pseudomonas aeruginosa are tripartite resistance-nodulation-cell division (RND) systems, which span the inner and outer membranes of Gram-negative pathogens, and expel antibiotics from the bacterial cytoplasm into the extracellular space. We provide evidence that amotosalen is an efflux substrate for the E. coli AcrAB, Acinetobacter baumannii AdeABC, and P. aeruginosa MexXY RND efflux pumps. Furthermore, we show that the MICs for contemporary Gram-negative bacterial isolates for these species and others in vitro approached and exceeded the concentration of amotosalen used in the approved platelet and plasma inactivation procedures. These findings suggest that otherwise safe and effective inactivation methods should be further studied to identify possible gaps in their ability to inactivate contemporary, multidrug-resistant bacterial pathogens. IMPORTANCE Pathogen inactivation is a strategy to enhance the safety of transfused blood products. We identify the compound, amotosalen, widely used for pathogen inactivation, as a bacterial multidrug efflux substrate. Specifically, experiments suggest that amotosalen is pumped out of bacteria by major efflux pumps in E. coli, Acinetobacter baumannii, and Pseudomonas aeruginosa. Such efflux pumps are often overexpressed in multidrug-resistant pathogens. Importantly, the MICs for contemporary multidrug-resistant Enterobacterales, Acinetobacter baumannii, Pseudomonas aeruginosa, Burkholderia spp., and Stenotrophomonas maltophilia isolates approached or exceeded the amotosalen concentration used in approved platelet and plasma inactivation procedures, potentially as a result of efflux pump activity. Although there are important differences in methodology between our experiments and blood product pathogen inactivation, these findings suggest that otherwise safe and effective inactivation methods should be further studied to identify possible gaps in their ability to inactivate contemporary, multidrug-resistant bacterial pathogens.

Validating the use of stereo-video cameras to conduct remote measurements of sea turtles.

Point 1: Stereo-video camera systems (SVCSs) are a promising tool to remotely measure body size of wild animals without the need for animal handling. Here, we assessed the accuracy of SVCSs for measuring straight carapace length (SCL) of sea turtles. Point 2: To achieve this, we hand captured and measured 63 juvenile, subadult, and adult sea turtles across three species: greens, Chelonia mydas (n = 52); loggerheads, Caretta caretta (n = 8); and Kemp's ridley, Lepidochelys kempii (n = 3) in the waters off Eleuthera, The Bahamas and Crystal River, Florida, USA, between May and November 2019. Upon release, we filmed these individuals with the SVCS. We performed photogrammetric analysis to extract stereo SCL measurements (eSCL), which were then compared to the (manual) capture measurements (mSCL). Point 3: mSCL ranged from 25.9 to 89.2 cm, while eSCL ranged from 24.7 to 91.4 cm. Mean percent bias of eSCL ranged from -0.61% (±0.11 SE) to -4.46% (±0.31 SE) across all species and locations. We statistically analyzed potential drivers of measurement error, including distance of the turtle to the SVCS, turtle angle, image quality, turtle size, capture location, and species. Point 4: Using a linear mixed effects model, we found that the distance between the turtle and the SVCS was the primary factor influencing measurement error. Our research suggests that stereo-video technology enables high-quality measurements of sea turtle body size collected in situ without the need for hand-capturing individuals. This study contributes to the growing knowledge base that SVCS are accurate for body size measurements independent of taxonomic clade.

Survivorship and growth in staghorn coral (Acropora cervicornis) outplanting projects in the Florida Keys National Marine Sanctuary.

Significant population declines in Acropora cervicornis and A. palmata began in the 1970s and now exceed over 90%. The losses were caused by a combination of coral disease and bleaching, with possible contributions from other stressors, including pollution and predation. Reproduction in the wild by fragment regeneration and sexual recruitment is inadequate to offset population declines. Starting in 2007, the Coral Restoration Foundation™ evaluated the feasibility of outplanting A. cervicornis colonies to reefs in the Florida Keys to restore populations at sites where the species was previously abundant. Reported here are the results of 20 coral outplanting projects with each project defined as a cohort of colonies outplanted at the same time and location. Photogrammetric analysis and in situ monitoring (2007 to 2015) measured survivorship, growth, and condition of 2419 colonies. Survivorship was initially high but generally decreased after two years. Survivorship among projects based on colony counts ranged from 4% to 89% for seven cohorts monitored at least five years. Weibull survival models were used to estimate survivorship beyond the duration of the projects and ranged from approximately 0% to over 35% after five years and 0% to 10% after seven years. Growth rate averaged 10 cm/year during the first two years then plateaued in subsequent years. After four years, approximately one-third of surviving colonies were ≥ 50 cm in maximum diameter. Projects used three to sixteen different genotypes and significant differences did not occur in survivorship, condition, or growth. Restoration times for three reefs were calculated based on NOAA Recovery Plan (NRP) metrics (colony abundance and size) and the findings from projects reported here. Results support NRP conclusions that reducing stressors is required before significant population growth and recovery will occur. Until then, outplanting protects against local extinction and helps to maintain genetic diversity in the wild.

Sensing Temperature in Vitro and in Cells Using a BODIPY Molecular Probe.

Boron dipyrromethene (BODIPY) molecular rotors have shown sensitivity toward viscosity, polarity, and temperature. Here, we report a 1,3,5,7-tetramethyl-8-phenyl-BODIPY modified with a polyethylene glycol (PEG) chain, for temperature sensing and live cell imaging. This new PEG-BODIPY dye presents an increase in nonradiative decay as temperature increases, which directly influences its lifetime. This change in lifetime is dependent on changes in both temperature and viscosity at low viscosity values, but is only dependent on temperature at high viscosity values. The dependence of fluorescence lifetime with temperature allows for temperature monitoring in vitro and in cells, with sub degree resolution. When in contact with cells, the PEG-BODIPY spontaneously penetrates and stains the cell but not the nucleus. Furthermore, no significant cell toxicity was found even at 100 µM concentration. Using fluorescence lifetime imaging microscopy (FLIM), we were able to observe the changes in the lifetime of PEG-BODIPY within the cell at different temperatures. The use of FLIM and molecular probes such as PEG-BODIPY can provide important information about cellular temperature and heat dissipation upon medically relevant stimuli, such as radiofrequency ablation and photodynamic therapy.

Ultrasound Doppler as an Imaging Modality for Selection of Murine 4T1 Breast Tumors for Combination Radiofrequency Hyperthermia and Chemotherapy.

Noninvasive radiofrequency-induced (RF) hyperthermia has been shown to increase the perfusion of chemotherapeutics and nanomaterials through cancer tissue in ectopic and orthotopic murine tumor models. Additionally, mild hyperthermia (37°C-45°C) has previously shown a synergistic anticancer effect when used with standard-of-care chemotherapeutics such as gemcitabine and Abraxane. However, RF hyperthermia treatment schedules remain unoptimized, and the mechanisms of action of hyperthermia and how they change when treating various tumor phenotypes are poorly understood. Therefore, pretreatment screening of tumor phenotypes to identify key tumors that are predicted to respond more favorably to hyperthermia will provide useful mechanistic data and may improve therapeutic outcomes. Herein, we identify key biophysical tumor characteristics in order to predict the outcome of combinational RF and chemotherapy treatment. We demonstrate that ultrasound imaging using Doppler mode can be utilized to predict the response of combinational RF and chemotherapeutic therapy in a murine 4T1 breast cancer model.

Development of photoactive Sweet-C60 for pancreatic cancer stellate cell therapy.

AIM: Glycoconjugated C60 derivatives are of particular interest as potential cancer targeting agents due to an upregulated metabolic glucose demand, especially in the case of pancreatic adenocarcinoma and its dense stroma, which is known to be driven by a subset of pancreatic stellate cells. MATERIALS & METHODS: Herein, we describe the synthesis and biological characterization of a hexakis-glucosamine C60 derivative (termed 'Sweet-C60'). RESULTS: Synthesized fullerene derivative predominantly accumulates in the nucleus of pancreatic stellate cells; is inherently nontoxic up to concentrations of 1 mg/ml; and is photoactive when illuminated with blue and green light, allowing its use as a photodynamic therapy agent. CONCLUSION: Obtained glycoconjugated nanoplatform is a promising nanotherapeutic for pancreatic cancer.

Chemotherapy and Radiofrequency-Induced Mild Hyperthermia Combined Treatment of Orthotopic Pancreatic Ductal Adenocarcinoma Xenografts.

Patients with pancreatic ductal adenocarcinomas (PDAC) have one of the poorest survival rates of all cancers. The main reason for this is related to the unique tumor stroma and poor vascularization of PDAC. As a consequence, chemotherapeutic drugs, such as nab-paclitaxel and gemcitabine, cannot efficiently penetrate into the tumor tissue. Non-invasive radiofrequency (RF) mild hyperthermia treatment was proposed as a synergistic therapy to enhance drug uptake into the tumor by increasing tumor vascular inflow and perfusion, thus, increasing the effect of chemotherapy. RF-induced hyperthermia is a safer and non-invasive technique of tumor heating compared to conventional contact heating procedures. In this study, we investigated the short- and long-term effects (~20 days and 65 days, respectively) of combination chemotherapy and RF hyperthermia in an orthotopic PDAC model in mice. The benefit of nab-paclitaxel and gemcitabine treatment was confirmed in mice; however, the effect of treatment was statistically insignificant in comparison to saline treated mice during long-term observation. The benefit of RF was minimal in the short-term and completely insignificant during long-term observation.

Non-Invasive Radiofrequency Field Treatment of 4T1 Breast Tumors Induces T-cell Dependent Inflammatory Response.

Previous work using non-invasive radiofrequency field treatment (RFT) in cancer has demonstrated its therapeutic potential as it can increase intratumoral blood perfusion, localization of intravenously delivered drugs, and promote a hyperthermic intratumoral state. Despite the well-known immunologic benefits that febrile hyperthermia can induce, an investigation of how RFT could modulate the intra-tumoral immune microenvironment had not been studied. Thus, using an established 4T1 breast cancer model in immune competent mice, we demonstrate that RFT induces a transient, localized, and T-cell dependent intratumoral inflammatory response. More specifically we show that multi- and singlet-dose RFT promote an increase in tumor volume in immune competent Balb/c mice, which does not occur in athymic nude models. Further leukocyte subset analysis at 24, 48, and 120 hours after a single RFT show a rapid increase in tumoral trafficking of CD4+ and CD8+ T-cells 24 hours post-treatment. Additional serum cytokine analysis reveals an increase in numerous pro-inflammatory cytokines and chemokines associated with enhanced T-cell trafficking. Overall, these data demonstrate that non-invasive RFT could be an effective immunomodulatory strategy in solid tumors, especially for enhancing the tumoral trafficking of lymphocytes, which is currently a major hindrance of numerous cancer immunotherapeutic strategies.

Macrophage Polarization Contributes to the Anti-Tumoral Efficacy of Mesoporous Nanovectors Loaded with Albumin-Bound Paclitaxel.

Therapies targeted to the immune system, such as immunotherapy, are currently shaping a new, rapidly developing branch of promising cancer treatments, offering the potential to change the prognosis of previously non-responding patients. Macrophages comprise the most abundant population of immune cells in the tumor microenvironment (TME) and can undergo differentiation into functional phenotypes depending on the local tissue environment. Based on these functional phenotypes, tumor-associated macrophages (TAMs) can either aid tumor progression (M2 phenotype) or inhibit it (M1 phenotype). Presence of M2 macrophages and a high ratio of M2/M1 macrophages in the TME are clinically associated with poor prognosis in many types of cancers. Herein, we evaluate the effect of macrophage phenotype on the transport and anti-cancer efficacy of albumin-bound paclitaxel (nAb-PTX) loaded into porous silicon multistage nanovectors (MSV). Studies in a coculture of breast cancer cells (3D-spheroid) with macrophages and in vivo models were conducted to evaluate the therapeutic efficacy of MSV-nAb-PTX as a function of macrophage phenotype. Association with MSV increased drug accumulation within the macrophages and the tumor spheroids, shifting the inflammation state of the TME toward the pro-inflammatory, anti-tumorigenic milieu. Additionally, the treatment increased macrophage motility toward cancer cells, promoting the active transport of therapeutic nanovectors into the tumor lesion. Consequently, apoptosis of cancer cells was increased and proliferation decreased in the MSV-nAb-PTX-treated group as compared to controls. The results also confirmed that the tested system shifts the macrophage differentiation toward an M1 phenotype, possessing an anti-proliferative effect toward the breast cancer cells. These factors were further incorporated into a mathematical model to help analyze the synergistic effect of the macrophage polarization state on the efficacy of MSV-nAb-PTX in alleviating hypovascularized tumor lesions. In conclusion, the ability of MSV-nAb-PTX to polarize TAM to the M1 phenotype, causing (1) enhanced penetration of the drug-carrying macrophages to the center of the tumor lesion and (2) increased toxicity to tumor cells may explain the increased anti-cancer efficacy of the system in comparison to nAb-PTX and other controls.

Pancreatic adenocarcinoma response to chemotherapy enhanced with non-invasive radio frequency evaluated via an integrated experimental/computational approach.

Although chemotherapy combined with radiofrequency exposure has shown promise in cancer treatment by coupling drug cytotoxicity with thermal ablation or thermally-induced cytotoxicity, limited access of the drug to tumor loci in hypo-vascularized lesions has hampered clinical application. We recently showed that high-intensity short-wave capacitively coupled radiofrequency (RF) electric-fields may reach inaccessible targets in vivo. This non-invasive RF combined with gemcitabine (Gem) chemotherapy enhanced drug uptake and effect in pancreatic adenocarcinoma (PDAC), notorious for having poor response and limited therapeutic options, but without inducing thermal injury. We hypothesize that the enhanced cytotoxicity derives from RF-facilitated drug transport in the tumor microenvironment. We propose an integrated experimental/computational approach to evaluate chemotherapeutic response combined with RF-induced phenotypic changes in tissue with impaired transport. Results show that RF facilitates diffusive transport in 3D cell cultures representing hypo-vascularized lesions, enhancing drug uptake and effect. Computational modeling evaluates drug vascular extravasation and diffusive transport as key RF-modulated parameters, with transport being dominant. Assessment of hypothetical schedules following current clinical protocol for Stage-IV PDAC suggests that unresponsive lesions may be growth-restrained when exposed to Gem plus RF. Comparison of these projections to experiments in vivo indicates that synergy may result from RF-induced cell phenotypic changes enhancing drug transport and cytotoxicity, thus providing a potential baseline for clinically-focused evaluation.