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Histotripsy is an emerging noninvasive, non-thermal, and non-ionizing focused ultrasound (US) therapy that can be used to destroy targeted tissue. Histotripsy has evolved from early laboratory prototypes to clinical systems which have been comprehensively evaluated in the preclinical environment to ensure safe translation to human use. This review summarizes the observations and results from preclinical histotripsy studies in the liver, kidney, and pancreas. Key findings from these studies include the ability to make a clinically relevant treatment zone in each organ with maintained collagenous architecture, potentially allowing treatments in areas not currently amenable to thermal ablation. Treatments across organ capsules have proven safe, including in anticoagulated models which may expand patients eligible for treatment or eliminate the risk associated with taking patients off anti-coagulation. Treatment zones are well-defined with imaging and rapidly resorb, which may allow improved evaluation of treatment zones for residual or recurrent tumor. Understanding the effects of histotripsy in animal models will help inform physicians adopting histotripsy for human clinical use.

Originally described by Davis et al in 2013, 4D-Digital Subtraction Angiography (4D-DSA) has developed into a commercially available application of DSA in the angiography suite. 4D-DSA provides the user with 3D time-resolved images, allowing observation of a contrast bolus at any desired viewing angle through the vasculature and at any time point during the acquisition (any view at any time). 4D-DSA mitigates some limitations that are intrinsic to both 2D- and 3D-DSA images. The clinical applications for 4D-DSA include evaluations of AVMs and AVFs, intracranial aneurysms, and atherosclerotic occlusive disease. Recent advances in blood flow quantification using 4D-DSA indicate that these data provide both the velocity and geometric information necessary for the quantification of blood flow. In this review, we will discuss the development, acquisition, reconstruction, and current neurovascular applications of 4D-DSA volumes., (© 2021 by American Journal of Neuroradiology.)

Background and Purpose: Quantification of blood flow using a 4D-DSA would be useful in the diagnosis and treatment of cerebrovascular diseases. A protocol optimizing identification of density variations in the time-density curves of a 4D-DSA has not been defined. Our purpose was to determine the contrast injection protocol most likely to result in the optimal pulsatility signal strength., Materials and Methods: Two 3D-printed patient-specific models were used and connected to a pulsatile pump and flow system, which delivered 250-260 mL/min to the model. Contrast medium (Isovue, 370 mg I/mL, 75% dilution) was injected through a 6F catheter positioned upstream from the inlet of the model. 4D-DSA acquisitions were performed for the following injection rates: 1.5, 2.0, 2.5, 3.0 and 3.5 mL/s for 8 seconds. To determine pulsatility, we analyzed the time-density curve at the inlets using the oscillation amplitude and a previously described numeric metric, the sideband ratio. Vascular geometry from 4D-DSA reconstructions was compared with ground truth and micro-CT measurements of the model. Dimensionless numbers that characterize hemodynamics, Reynolds and Craya-Curtet, were calculated for each injection rate., Results: The strongest pulsatility signal occurred with the 2.5 mL/s injections. The largest oscillation amplitudes were found with 2.0- and 2.5-mL/s injections. Geometric accuracy was best preserved with injection rates of >1.5 mL/s., Conclusions: An injection rate of 2.5 mL/s provided the strongest pulsatility signal in the 4D-DSA time-density curve. Geometric accuracy was best preserved with injection rates above 1.5 mL/s. These results may be useful in future in vivo studies of blood flow quantification., (© 2019 by American Journal of Neuroradiology.)

Purpose: Particle image velocimetry (PIV), an in vitro experimentation technique that optically measures velocity components to analyze fluid velocity fields, has become increasingly popular to study flow dynamics in various vascular territories. However, it can be difficult and expensive to create patient-specific clear models for PIV due to the importance of refractive index matching of the model and the fluid. We aim to implement and test the use of poly-vinyl alcohol (PVA) in a lost-core casting technique to create low-cost, patient-specific models for PIV., Methods: Anonymized patient vascular anatomies were segmented and processed in Mimics/3Matic to create patient-specific cores from 3D digital subtraction angiographies. The cores were 3D-printed with PVA and post-processed with a 80:20 water:glue mixture to smooth the surface. Two silicones, Sylgard 184 and Solaris, were used to encapsulate the model and the PVA core was dissolved using warm water. Computed tomography scans were used to evaluate geometric accuracy using circumferences and surface differences in the model., Results: Mean geometric differences in circumference along the inlet centerline and the mean surface difference in the aneurysm between the final Silicone Model and the desired STL Print geometry were statistically insignificant (0.6 mm, 95% CI [- 1.4, 2.8] and 0.3 mm 95% CI [- 0.1, 0.7], respectively). Particle illumination within each model was successful. The cost of one 10 cm × 10 cm × 5 cm model was $69., Conclusion: This technique was successful to implement and test the use of PVA in a lost-core casting technique to create low-cost, patient-specific in vitro models for PIV experimentation.

Background and Purpose: 4D-DSA provides time-resolved 3D-DSA volumes with high temporal and spatial resolutions. The purpose of this study is to investigate a shifted least squares method to estimate the blood velocity from the 4D DSA images. Quantitative validation was performed using a flow phantom with an ultrasonic flow probe as ground truth. Quantification of blood velocity in human internal carotid arteries was compared with measurements generated from 3D phase-contrast MR imaging., Materials and Methods: The centerlines of selected vascular segments and the time concentration curves of each voxel along the centerlines were determined from the 4D-DSA dataset. The temporal shift required to achieve a minimum difference between any point and other points along the centerline of a segment was calculated. The temporal shift as a function of centerline point position was fit to a straight line to generate the velocity. The proposed shifted least-squares method was first validated using a flow phantom study. Blood velocities were also estimated in the 14 ICAs of human subjects who had both 4D-DSA and phase-contrast MR imaging studies. Linear regression and correlation analysis were performed on both the phantom study and clinical study, respectively., Results: Mean velocities of the flow phantom calculated from 4D-DSA matched very well with ultrasonic flow probe measurements with 11% relative root mean square error. Mean blood velocities of ICAs calculated from 4D-DSA correlated well with phase-contrast MR imaging measurements with Pearson correlation coefficient r = 0.835., Conclusions: The availability of 4D-DSA provides the opportunity to use the shifted least-squares method to estimate velocity in vessels within a 3D volume., (© 2018 by American Journal of Neuroradiology.)

Background and Purpose: Safe and effective use of newly developed devices for aneurysm treatment requires the ability to make accurate measurements in the angiographic suite. Our purpose was to determine the parameters that optimize the geometric accuracy of three-dimensional (3D) vascular reconstructions., Methods: An in vitro flow model consisting of a peristaltic pump, plastic tubing, and 3D printed patient-specific aneurysm models was used to simulate blood flow in an intracranial aneurysm. Flow rates were adjusted to match values reported in the literature for the internal carotid artery. 3D digital subtraction angiography acquisitions were obtained using a commercially available biplane angiographic system. Reconstructions were done using Edge Enhancement (EE) or Hounsfield Unit (HU) kernels and a Normal or Smooth image characteristic. Reconstructed images were analyzed using the vendor's aneurysm analysis tool. Ground truth measurements were derived from metrological scans of the models with a microCT. Aneurysm volume, surface area, dome height, minimum and maximum ostium diameter were determined for the five models., Results: In all cases, measurements made with the EE kernel most closely matched ground truth values. Differences in values derived from reconstructions displayed with Smooth or Normal image characteristics were small and had only little impact on the geometric parameters considered., Conclusions: Reconstruction parameters impact the accuracy of measurements made using the aneurysm analysis tool of a commercially available angiographic system. Absolute differences between measurements made using reconstruction parameters determined as optimal in this study were, overall, very small. The significance of these differences, if any, will depend on the details of each individual case., Competing Interests: Competing interests: None declared., (Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://www.bmj.com/company/products-services/rights-and-licensing/.)

Although slugs and snails play important roles in terrestrial ecosystems and cause considerable damage on a variety of crop plants, knowledge about the mechanisms of plant immunity to molluscs is limited. We found slugs to be natural herbivores of Arabidopsis thaliana and therefore investigated possible resistance mechanisms of this species against several molluscan herbivores. Treating wounded leaves with the mucus residue ('slime trail') of the Spanish slug Arion lusitanicus increased wound-induced jasmonate levels, suggesting the presence of defence elicitors in the mucus. Plants deficient in jasmonate biosynthesis and signalling suffered more damage by molluscan herbivores in the laboratory and in the field, demonstrating that JA-mediated defences protect A. thaliana against slugs and snails. Furthermore, experiments using A. thaliana mutants with altered levels of specific glucosinolate classes revealed the importance of aliphatic glucosinolates in defending leaves and reproductive structures against molluscs. The presence in mollusc faeces of known and novel metabolites arising from glutathione conjugation with glucosinolate hydrolysis products suggests that molluscan herbivores actively detoxify glucosinolates. Higher levels of aliphatic glucosinolates were found in plants during the night compared to the day, which correlated well with the nocturnal activity rhythms of slugs and snails. Our data highlight the function of well-known antiherbivore defence pathways in resistance against slugs and snails and suggest an important role for the diurnal regulation of defence metabolites against nocturnal molluscan herbivores., (© 2013 John Wiley & Sons Ltd.)

Male fin whales, Balaenoptera physalus, produce a song consisting of 20 Hz notes at regularly spaced time intervals. Previous studies identified regional differences in fin whale internote intervals (INI), but seasonal changes within populations have not been closely examined. To understand the patterns of fin whale song in the western North Atlantic, the seasonal abundance and acoustic features of fin whale song are measured from two years of archival passive acoustic recordings at two representative locations: Massachusetts Bay and New York Bight. Fin whale 20 Hz notes are detected on 99% of recorded days. In both regions, INI varies significantly throughout the year as two distinct periods: a "short-INI" season in September-January (9.6 s) and a "long-INI" season in March-May (15.1 s). February and June-August are transitional-INI months, with higher variability. Note abundance decreases with increasing INI, where note abundance is significantly lower in April-August than in September-January. Short-INI and high note abundance correspond to the fin whale reproductive season. The temporal variability of INI may be a mechanism by which fin whale individuals encode and communicate a variety of behaviorally relevant information.

Glucosinolates are sulfur-rich plant metabolites of the order Brassicales that function in the defense of plants against pests and pathogens. They are also important in human society as flavor components, cancer-prevention agents, and crop biofumigants. Since glucosinolates may represent up to 30 % of the total sulfur content of plant organs, their accumulation should depend intimately on the sulfur status of the entire plant. Here we review the literature on how sulfur supply affects glucosinolate content. In field and greenhouse experiments involving soil, hydroponic and tissue culture media, sulfur fertilisation usually led to an increase in glucosinolate content ranging from 25 % to more than 50-fold, depending on the plant species, amount of sulfur applied, and type of treatment. The effect was greater on glucosinolates derived from the sulfur amino acid, methionine, than on glucosinolates derived from tryptophan. These changes are regulated not by simple mass action effects, but by extensive changes in gene transcription. In sulfur-deficient plants, there is a general down-regulation of glucosinolate biosynthetic genes which accompanies an up-regulation of genes controlling sulfur uptake and assimilation. Glucosinolates may be considered a potential source of sulfur for other metabolic processes under low-sulfur conditions, since increased breakdown of glucosinolates has been reported under sulfur deficiency. However, the pathway for sulfur mobilisation from glucosinolates has not been determined. The breakdown of indolic glucosinolates to form auxin in roots under sulfur-deficient conditions may help stimulate root formation for sulfur uptake.

Desert locusts (Schistocerca gregaria) occasionally feed on Schouwia purpurea, a plant that contains tenfold higher levels of glucosinolates than most other Brassicaceae. Whereas this unusually high level of glucosinolates is expected to be toxic and/or deterrent to most insects, locusts feed on the plant with no apparent ill effects. In this paper, we demonstrate that the desert locust, like larvae of the diamondback moth (Plutella xylostella), possesses a glucosinolate sulfatase in the gut that hydrolyzes glucosinolates to their corresponding desulfonated forms. These are no longer susceptible to cleavage by myrosinase, thus eliminating the formation of toxic glucosinolate hydrolysis products. Sulfatase is found throughout the desert locust gut and can catalyze the hydrolysis of all of the glucosinolates present in S. purpurea. The enzyme was detected in all larval stages of locusts as well as in both male and female adults feeding on this plant species. Glucosinolate sulfatase activity is induced tenfold when locusts are fed S. purpurea after being maintained on a glucosinolate-free diet, and activity declines when glucosinolates are removed from the locust diet. A detoxification system that is sensitive to the dietary levels of a plant toxin may minimize the physiological costs of toxin processing, especially for a generalist insect herbivore that encounters large variations in plant defense metabolites while feeding on different species.

The major class of glucosinolates in Arabidopsis thaliana (L.) Heynh. are biosynthesized from methionine involving a three-step chain-elongation cycle. Each passage through the cycle results in the net addition of a single methylene group, with up to six cycles of elongation occurring in A. thaliana. The first reaction of the cycle is catalyzed by a methylthioalkylmalate synthase (MAMS), which condenses a omega-methylthio-2-oxoalkanoic acid with acetyl-CoA. Here we have demonstrated that MAM1, one of two similar genes in the A. thaliana ecotype Columbia, encodes a MAMS catalyzing the condensing reactions of the first two elongation cycles but not those of further cycles. The Columbia ecotype is dominated by compounds that have undergone only two elongation cycles. The A. thaliana MAM1 protein exhibits basic sequence similarity to other previously described enzymes catalyzing the condensation of 2-oxo acids and acetyl-CoA, such as isopropylmalate synthase (EC 2.3.3.13), an enzyme of leucine biosynthesis, and homocitrate synthase (EC 2.3.3.14). It also shares similar properties with them, including the catalytic requirements for a divalent metal ion and an adenine nucleotide. However, the MAM1 protein does not show activity with the substrates of any of these other enzymes, and was chromatographically separable from isopropylmalate synthase in extracts of A. thaliana. Thus, MAM1 is exclusively an enzyme of secondary metabolism, distinct from primary metabolic enzymes catalyzing similar reactions.

Glucosinolates are a group of sulfur-rich thioglucoside natural products common in the Brassicaceae and related plant families. The first phase in the formation of many glucosinolates involves the chain extension of the amino acid methionine. Additional methylene groups are inserted into the side chain of methionine by a three-step elongation cycle involving 2-oxo acid intermediates. This investigation demonstrated the first step of this chain elongation cycle in a partially-purified preparation from arugula (Eruca sativa). The 2-oxo acid derived from methionine, 4-methylthio-2-oxobutanoic acid, was shown to condense with acetyl-CoA to form 2-(2'-methylthioethyl)malate. The catalyst, designated as a 2-(omega-methylthioalkyl)malate synthase, belongs to a family of enzymes that mediate the condensation of acyl-CoAs with 2-oxo acids, including citrate synthase of the citric acid cycle, and 2-isopropylmalate synthase of leucine biosynthesis. The 2-(omega-methylthioalkyl)malate synthase studied here shares properties with other enzymes of this class, but appears chromatographically distinct and is found only in extracts of plant species producing glucosinolates from chain-elongated methionine derivatives. Although the principal glucosinolates of arugula are formed from methionine that has undergone two rounds of chain elongation to form dihomomethionine, studies with substrates and substrate analogs of different chain lengths showed that the isolated enzyme is responsible only for the condensation step of the first round of elongation.

Arabidopsis and other Brassicaceae produce an enormous diversity of aliphatic glucosinolates, a group of methionine (Met)-derived plant secondary compounds containing a beta-thio-glucose moiety, a sulfonated oxime, and a variable side chain. We fine-scale mapped GSL-ELONG, a locus controlling variation in the side-chain length of aliphatic glucosinolates. Within this locus, a polymorphic gene was identified that determines whether Met is extended predominantly by either one or by two methylene groups to produce aliphatic glucosinolates with either three- or four-carbon side chains. Two allelic mutants deficient in four-carbon side-chain glucosinolates were shown to contain independent missense mutations within this gene. In cell-free enzyme assays, a heterologously expressed cDNA from this locus was capable of condensing 2-oxo-4-methylthiobutanoic acid with acetyl-coenzyme A, the initial reaction in Met chain elongation. The gene methylthioalkylmalate synthase1 (MAM1) is a member of a gene family sharing approximately 60% amino acid sequence similarity with 2-isopropylmalate synthase, an enzyme of leucine biosynthesis that condenses 2-oxo-3-methylbutanoate with acetyl-coenzyme A.

Biosynthesis of sucrose from triacylglycerol requires the bypass of the CO2-evolving reactions of the tricarboxylic acid (TCA) cycle. The regulation of the TCA cycle bypass during lipid mobilization was examined. Lipid mobilization in Brassica napus was initiated shortly after imbibition of the seed and proceeded until 2 d postimbibition, as measured by in vivo [1-14C]acetate feeding to whole seedlings. The activity of NAD+-isocitrate dehydrogenase (a decarboxylative enzyme) was not detected until 2 d postimbibition. RNA-blot analysis of B. napus seedlings demonstrated that the mRNA for NAD+-isocitrate dehydrogenase was present in dry seeds and that its level increased through the 4 d of the experiment. This suggested that NAD+-isocitrate dehydrogenase activity was regulated by posttranscriptional mechanisms during early seedling development but was controlled by mRNA level after the 2nd or 3rd d. The activity of fumarase (a component of the nonbypassed section of the TCA cycle) was low but detectable in B. napus seedlings at 12 h postimbibition, coincident with germination, and increased for the next 4 d. RNA-blot analysis suggested that fumarase activity was regulated primarily by the level of its mRNA during germination and early seedling development. It is concluded that posttranscriptional regulation of NAD+-isocitrate dehydrogenase activity is one mechanism of restricting carbon flux through the decarboxylative section of the TCA cycle during lipid mobilization in germinating oilseeds.

The purpose of this study was to assess the effectiveness and safety of Gadolite Oral Suspension as a gastrointestinal (GI) contrast agent for MRI in a phase II and two phase III multicenter clinical trials. Gadolite was administered to 306 patients with known or suspected abdominal and/or pelvic disease. MRI with T1- and T2-weighted sequences was performed before and after ingestion. Efficacy was evaluated by having two masked readers rate the certainty of their MR diagnosis (0 = uncertain, 1 = probable, 2 = definite) on randomly presented pre- and post-Gadolite Oral Suspension enhanced images. Principal investigators also evaluated the images and established the final diagnosis. Vital signs, clinical chemistries, and adverse events were documented. Blood and urine samples were analyzed for gadolinium content to determine whether Gadolite Oral Suspension was absorbed systemically. Certainty in MR diagnosis increased significantly (P < .001) for both blinded readers between pre- and post-Gadolite images (.49-1.18 for reader 1: .46-1.53 for reader 2). Sensitivity, specificity, and accuracy also increased for both masked readers. No gadolinium was detected in blood or urine samples. There were no serious adverse events and no apparent drug-related trends in mean vital signs or laboratory values. Gadolite is a highly effective, safe, and well tolerated contrast agent for clinical use with MRI.

A study of the biosynthesis of coenzyme A (CoA), a critical cofactor in the metabolism of lipids and other molecules in higher plants, was initiated. Pantothenate kinase was partially purified from spinach leaves. This enzyme was predominantly localized in the chloroplast with very little activity observed in the mitochondria or cytosol. DEAE-agarose chromatography resolved two pantothenate kinase activity peaks which differed in their requirement for reductant, stability upon boiling, and reactivity in the presence of spinach holo-acyl carrier protein (ACP) I. One active peak of this enzyme was further purified on Cibacron blue 3GA to yield a preparation containing pantothenate kinase enriched to 20% of the total protein within the fraction. Pantothenate kinase was inhibited by malonyl-CoA, but not by CoASH or acetyl-CoA, and the activity was stabilized by the phosphatase inhibitors sodium molybdate, sodium tungstate, and the phosphatase substrate glycerol 2-phosphate, but was inhibited by sodium fluoride. Further experiments demonstrated a linear increase in pantothenate kinase activity during spinach seed germination, consistent with a role for this enzyme in the developmental utilization of seed triacylglycerol.

Forty patients with karyotypically proven Turner syndrome were prospectively studied using magnetic resonance imaging (MRI) and echocardiography in order to determine the frequency of cardiovascular anomalies and to assess the utility of both imaging modalities as methods for cardiovascular evaluation in Turner syndrome. Cardiovascular anomalies were found in 45% of patients. A high absolute prevalence of bicuspid aortic valve (17.5%) and aortic coarctation (12.5%) were observed relative to comparable series. Of clinically significant abnormalities, three of five aortic coarctations and four of five ascending aortic dilatations were solely MRI detected and not evident at echocardiographic examination. MRI is thus seen as a valuable adjunct to echocardiography in the cardiovascular evaluation of Turner syndrome patients. The usefulness of MRI primarily relates to its ability to provide excellent visualisation of the entire thoracic aorta where a large proportion of clinically significant anomalies occur in Turner syndrome.

Leaves of common sage (Salvia officinalis) accumulate monoterpenes in glandular trichomes at levels exceeding 15 milligrams per gram fresh weight at maturity, whereas sage cells in suspension culture did not accumulate detectable levels of monoterpenes (<0.3 nanograms per gram fresh weight) at any stage of the growth cycle, even in the presence of a polystyrene resin trap. Monoterpene biosynthesis from [U-(14)C]sucrose was also virtually undetectable in this cell culture system. In vitro assay of each of the enzymes required for the sequential conversion of the ubiquitous isoprenoid precursor geranyl pyrophosphate to (+)-camphor (a major monoterpene product of sage) in soluble extracts of the cells revealed the presence of activity sufficient to produce (+)-camphor at a readily detectable level (>0.3 micrograms per gram fresh weight) at the late log phase of growth. Other monoterpene synthetic enzymes were present as well. In vivo measurement of the ability to catabolize (+)-camphor in these cells indicated that degradative capability exceeded biosynthetic capacity by at least 1000-fold. Therefore, the lack of monoterpene accumulation in undifferentiated sage cultures could be attributed to a low level of biosynthetic activity (relative to the intact plant) coupled to a pronounced capacity for monoterpene catabolism.