A MicroRNA Approach to Evaluating Elevated Prostate Cancer Risk in Cancer-Free Men.

Objective criteria are required for prostate cancer (PCa) risk assessment, treatment decisions, evaluation of therapy, and initial indications of recurrence. Circulating microRNAs were utilized as biomarkers to distinguish PCa patients from cancer-free subjects or those encountering benign prostate hyperplasia. A panel of 60 microRNAs was developed with established roles in PCa initiation, progression, metastasis, and recurrence. Utilizing the FirePlex® platform for microRNA analysis, we demonstrated the efficacy and reproducibility of a rapid, high-throughput, serum-based assay for PCa biomarkers that circumvents the requirement for extraction and fractionation of patient specimens supporting feasibility for expanded clinical research and diagnostic applications.

Approaches to neuropathic amputation-related pain: narrative review of surgical, interventional, and medical treatments.

BACKGROUND/IMPORTANCE: Neuropathic amputation-related pain can consist of phantom limb pain (PLP), residual limb pain (RLP), or a combination of both pathologies. Estimated of lifetime prevalence of pain and after amputation ranges between 8% and 72%. OBJECTIVE: This narrative review aims to summarize the surgical and non-surgical treatment options for amputation-related neuropathic pain to aid in developing optimized multidisciplinary and multimodal treatment plans that leverage multidisciplinary care. EVIDENCE REVIEW: A search of the English literature using the following keywords was performed: PLP, amputation pain, RLP. Abstract and full-text articles were evaluated for surgical treatments, medical management, regional anesthesia, peripheral block, neuromodulation, spinal cord stimulation, dorsal root ganglia, and peripheral nerve stimulation. FINDINGS: The evidence supporting most if not all interventions for PLP are inconclusive and lack high certainty. Targeted muscle reinnervation and regional peripheral nerve interface are the leading surgical treatment options for reducing neuroma formation and reducing PLP. Non-surgical options include pharmaceutical therapy, regional interventional techniques and behavioral therapies that can benefit certain patients. There is a growing evidence that neuromodulation at the spinal cord or the dorsal root ganglia and/or peripheral nerves can be an adjuvant therapy for PLP. CONCLUSIONS: Multimodal approaches combining pharmacotherapy, surgery and invasive neuromodulation procedures would appear to be the most promising strategy for preventive and treating PLP and RLP. Future efforts should focus on cross-disciplinary education to increase awareness of treatment options exploring best practices for preventing pain at the time of amputation and enhancing treatment of chronic postamputation pain.

Few-cycle laser pulse characterization on-target using high-harmonic generation from nano-scale solids.

We demonstrate high-harmonic generation for the time-domain observation of the electric field (HHG-TOE) and use it to measure the waveform of ultrashort mid-infrared (MIR) laser pulses interacting with ZnO thin-films or WS2 monolayers. The working principle relies on perturbing HHG in solids with a weak replica of the pump pulse. We measure the duration of few-cycle pulses at 3200 nm, in reasonable agreement with the results of established pulse characterization techniques. Our method provides a straightforward approach to accurately characterize femtosecond laser pulses used for HHG experiments right at the point of interaction.

AutoPhy: Automated phylogenetic identification of novel protein subfamilies.

Phylogenetic analysis of protein sequences provides a powerful means of identifying novel protein functions and subfamilies, and for identifying and resolving annotation errors. However, automation of functional clustering based on phylogenetic trees has been challenging and most of it is done manually. Clustering phylogenetic trees usually requires the delineation of tree-based thresholds (e.g., distances), leading to an ad hoc problem. We propose a new phylogenetic clustering approach that identifies clusters without using ad hoc distances or other pre-defined values. Our workflow combines uniform manifold approximation and projection (UMAP) with Gaussian mixture models as a k-means like procedure to automatically group sequences into clusters. We then apply a "second pass" clade identification algorithm to resolve non-monophyletic groups. We tested our approach with several well-curated protein families (outer membrane porins, acyltransferase, and nuclear receptors) and showed our automated methods recapitulated known subfamilies. We also applied our methods to a broad range of different protein families from multiple databases, including Pfam, PANTHER, and UniProt, and to alignments of RNA viral genomes. Our results showed that AutoPhy rapidly generated monophyletic clusters (subfamilies) within phylogenetic trees evolving at very different rates both within and among phylogenies. The phylogenetic clusters generated by AutoPhy resolved misannotations and identified new protein functional groups and novel viral strains.

Thermoprotection by a cell membrane-localized metacaspase in a green alga.

Caspases are restricted to animals, while other organisms, including plants, possess metacaspases (MCAs), a more ancient and broader class of structurally related yet biochemically distinct proteases. Our current understanding of plant MCAs is derived from studies in streptophytes, and mostly in Arabidopsis (Arabidopsis thaliana) with 9 MCAs with partially redundant activities. In contrast to streptophytes, most chlorophytes contain only 1 or 2 uncharacterized MCAs, providing an excellent platform for MCA research. Here we investigated CrMCA-II, the single type-II MCA from the model chlorophyte Chlamydomonas (Chlamydomonas reinhardtii). Surprisingly, unlike other studied MCAs and similar to caspases, CrMCA-II dimerizes both in vitro and in vivo. Furthermore, activation of CrMCA-II in vivo correlated with its dimerization. Most of CrMCA-II in the cell was present as a proenzyme (zymogen) attached to the plasma membrane (PM). Deletion of CrMCA-II by genome editing compromised thermotolerance, leading to increased cell death under heat stress. Adding back either wild-type or catalytically dead CrMCA-II restored thermoprotection, suggesting that its proteolytic activity is dispensable for this effect. Finally, we connected the non-proteolytic role of CrMCA-II in thermotolerance to the ability to modulate PM fluidity. Our study reveals an ancient, MCA-dependent thermotolerance mechanism retained by Chlamydomonas and probably lost during the evolution of multicellularity.

SPIRO - the automated Petri plate imaging platform designed by biologists, for biologists.

Phenotyping of model organisms grown on Petri plates is often carried out manually, despite the procedures being time-consuming and laborious. The main reason for this is the limited availability of automated phenotyping facilities, whereas constructing a custom automated solution can be a daunting task for biologists. Here, we describe SPIRO, the Smart Plate Imaging Robot, an automated platform that acquires time-lapse photographs of up to four vertically oriented Petri plates in a single experiment, corresponding to 192 seedlings for a typical root growth assay and up to 2500 seeds for a germination assay. SPIRO is catered specifically to biologists' needs, requiring no engineering or programming expertise for assembly and operation. Its small footprint is optimized for standard incubators, the inbuilt green LED enables imaging under dark conditions, and remote control provides access to the data without interfering with sample growth. SPIRO's excellent image quality is suitable for automated image processing, which we demonstrate on the example of seed germination and root growth assays. Furthermore, the robot can be easily customized for specific uses, as all information about SPIRO is released under open-source licenses. Importantly, uninterrupted imaging allows considerably more precise assessment of seed germination parameters and root growth rates compared with manual assays. Moreover, SPIRO enables previously technically challenging assays such as phenotyping in the dark. We illustrate the benefits of SPIRO in proof-of-concept experiments which yielded a novel insight on the interplay between autophagy, nitrogen sensing, and photoblastic response.

Dynamical Franz-Keldysh Effect in Diamond in the Deep Ultraviolet Probed by Transient Absorption and Dispersion Spectroscopy Using a Miniature Beamline.

Here, we introduce a miniature beamline for transient absorption and dispersion spectroscopy, using a tailored deep ultraviolet field immediately after the noncollinear generation without subsequent optical elements. We explore the near-band-gap region in diamond in the presence of a few-femtosecond pump pulse where the delayed dynamical Franz-Keldysh effect and the almost instantaneous optical Kerr effect coexist.

Reduction of Escherichia coli O157:H7 in Finishing Cattle Fed Corn Genetically Modified to Produce Increased Concentrations of Alpha Amylase in the Corn Kernel.

Cattle are recognized as the principal reservoir for Escherichia coli O157:H7 and preharvest food safety efforts often focus on decreasing shedding of this pathogen in cattle feces. Enogen® corn (EC; Syngenta Seeds, LLC) is genetically modified to produce enhanced concentrations of α-amylase in the corn kernel endosperm. Research has demonstrated improvements in feed efficiency for cattle fed EC and research has not yet explored whether improved digestion impacts foodborne pathogen populations in cattle. Therefore, this study explored effects of finishing diets containing EC on Escherichia coli O157:H7 prevalence in cattle. A 2 × 2 factorial experiment was conducted with steers (n = 960) fed diets consisting of 2 types of silage (EC or Control) and grain (EC or Control), fed daily ad libitum. Steers were grouped into 12 blocks by incoming body weight, blocks were randomly assigned to one of four pens, and pens were randomly assigned to one diet. Cattle were sampled using rectoanal mucosal swabs in cohorts of 298-337 cattle per day, for a total of 3 sampling days (15-16 days apart). Escherichia coli O157:H7 prevalence rates ranged from not detected (0/75) to 10.0% (8/80) depending on sampling day. Tests for the silage × corn interaction, and the main effects of silage and corn, were not significant (p > 0.05); however, EC reduced the odds of Escherichia coli O157:H7 prevalence by 43% compared to the control corn diet (p = 0.07). Diets containing EC tended to decrease Escherichia coli O157:H7 prevalence in feedlot cattle; however, this reduction was not significant. Before a conclusion can be drawn about impact of EC on Escherichia coli O157:H7 in cattle, further research is necessary to (1) determine if this tendency is due to increased alpha amylase activity and (2) elucidate impact on Escherichia coli O157:H7 prevalence and concentration, as well as a possible mechanism of action.

Impact of the Probiotic Organism Megasphaera elsdenii on Escherichia coli O157:H7 Prevalence in Finishing Cattle.

Feedlot cattle commonly shed the foodborne pathogen Escherichia coli O157:H7 in their feces. Megasphaera elsdenii (ME), a lactic acid-utilizing bacterium, is commonly administered to cattle to avoid lactate accumulation in the rumen and to control ruminal acidosis. The impact of administering ME on foodborne pathogen prevalence, specifically E. coli O157:H7, has not been explored. The purpose of this study was to quantify E. coli O157:H7 prevalence in finishing cattle administered ME. Cattle (n = 448) were assigned to treatments in a randomized complete block design with repeated measurements over two sampling periods. Treatments were arranged as a 2 × 2 factorial containing: ruminally protected lysine (RPL; included for a complementary study) fed at 0% or 0.45% of diet dry matter; with or without ME. Freeze-dried ME was administered as an oral drench (1 × 1010 CFU/steer on day one) and then top dressed onto basal diets (1 × 107 CFU/steer) daily thereafter. Rectoanal mucosal swabs (RAMS) were obtained from animals before harvest to determine the E. coli O157:H7 prevalence. The inclusion of RPL (P = 0.2136) and ME (P = 0.5012) did not impact E. coli O157:H7 prevalence, and RPL was not included in any significant interactions (P > 0.05). A significant interaction was observed between ME and sampling period (P = 0.0323), indicating that the effect of ME on E. coli O157:H7 prevalence varied over the sampling period. A diet containing ME reduced the odds of E. coli O157:H7 prevalence by 50% during sampling period 1 (8.0% and 14.7% for cattle with and without ME, respectively) and increased the odds by 23% during sampling period 2 (10.8% and 8.9% for cattle with and without ME, respectively). Administering ME in cattle diets did not impact E. coli O157:H7 in feedlot cattle. This is the first study to investigate the use of ME as a preharvest food safety intervention in cattle, and additional research is necessary to determine the efficacy.

Single-FiO2 lung modelling with machine learning: a computer simulation incorporating volumetric capnography.

We investigated whether machine learning (ML) analysis of ICU monitoring data incorporating volumetric capnography measurements of mean alveolar PCO2 can partition venous admixture (VenAd) into its shunt and low V/Q components without manipulating the inspired oxygen fraction (FiO2). From a 21-compartment ventilation / perfusion (V/Q) model of pulmonary blood flow we generated blood gas and mean alveolar PCO2 data in simulated scenarios with shunt values from 7.3% to 36.5% and a range of FiO2 settings, indirect calorimetry and cardiac output measurements and acid- base and hemoglobin oxygen affinity conditions. A 'deep learning' ML application, trained and validated solely on single FiO2 bedside monitoring data from 14,736 scenarios, then recovered shunt values in 500 test scenarios with true shunt values 'held back'. ML shunt estimates versus true values (n = 500) produced a linear regression model with slope = 0.987, intercept = -0.001 and R2 = 0.999. Kernel density estimate and error plots confirmed close agreement. With corresponding VenAd values calculated from the same bedside data, low V/Q flow can be reported as VenAd-shunt. ML analysis of blood gas, indirect calorimetry, volumetric capnography and cardiac output measurements can quantify pulmonary oxygenation deficits as percentage shunt flow (V/Q = 0) versus percentage low V/Q flow (V/Q > 0). High fidelity reports are possible from analysis of data collected solely at the operating FiO2.

The role of Atg16 in autophagy, anthocyanin biosynthesis, and programmed cell death in leaves of the lace plant (Aponogeton madagascariensis).

Aponogeton madagascariensis, commonly known as the lace plant, produces leaves that form perforations by programmed cell death (PCD). Leaf development is divided into several stages beginning with "pre-perforation" furled leaves enriched with red pigmentation from anthocyanins. The leaf blade is characterized by a series of grids known as areoles bounded by veins. As leaves develop into the "window stage", anthocyanins recede from the center of the areole towards the vasculature creating a gradient of pigmentation and cell death. Cells in the middle of the areole that lack anthocyanins undergo PCD (PCD cells), while cells that retain anthocyanins (non-PCD cells) maintain homeostasis and persist in the mature leaf. Autophagy has reported roles in survival or PCD promotion across different plant cell types. However, the direct involvement of autophagy in PCD and anthocyanin levels during lace plant leaf development has not been determined. Previous RNA sequencing analysis revealed the upregulation of autophagy-related gene Atg16 transcripts in pre-perforation and window stage leaves, but how Atg16 affects PCD in lace plant leaf development is unknown. In this study, we investigated the levels of Atg16 in lace plant PCD by treating whole plants with either an autophagy promoter rapamycin or inhibitors concanamycin A (ConA) or wortmannin. Following treatments, window and mature stage leaves were harvested and analyzed using microscopy, spectrophotometry, and western blotting. Western blotting showed significantly higher Atg16 levels in rapamycin-treated window leaves, coupled with lower anthocyanin levels. Wortmannin-treated leaves had significantly lower Atg16 protein and higher anthocyanin levels compared to the control. Mature leaves from rapamycin-treated plants generated significantly fewer perforations compared to control, while wortmannin had the opposite effect. However, ConA treatment did not significantly change Atg16 levels, nor the number of perforations compared to the control, but anthocyanin levels did increase significantly in window leaves. We propose autophagy plays a dual role in promoting cell survival in NPCD cells by maintaining optimal anthocyanin levels and mediating a timely cell death in PCD cells in developing lace plant leaves. How autophagy specifically affects anthocyanin levels remained unexplained.

Pulmonary gas exchange evaluated by machine learning: a computer simulation.

Using computer simulation we investigated whether machine learning (ML) analysis of selected ICU monitoring data can quantify pulmonary gas exchange in multi-compartment format. A 21 compartment ventilation/perfusion (V/Q) model of pulmonary blood flow processed 34,551 combinations of cardiac output, hemoglobin concentration, standard P50, base excess, VO2 and VCO2 plus three model-defining parameters: shunt, log SD and mean V/Q. From these inputs the model produced paired arterial blood gases, first with the inspired O2 fraction (FiO2) adjusted to arterial saturation (SaO2) = 0.90, and second with FiO2 increased by 0.1. 'Stacked regressor' ML ensembles were trained/validated on 90% of this dataset. The remainder with shunt, log SD, and mean 'held back' formed the test-set. 'Two-Point' ML estimates of shunt, log SD and mean utilized data from both FiO2 settings. 'Single-Point' estimates used only data from SaO2 = 0.90. From 3454 test gas exchange scenarios, two-point shunt, log SD and mean estimates produced linear regression models versus true values with slopes ~ 1.00, intercepts ~ 0.00 and R2 ~ 1.00. Kernel density and Bland-Altman plots confirmed close agreement. Single-point estimates were less accurate: R2 = 0.77-0.89, slope = 0.991-0.993, intercept = 0.009-0.334. ML applications using blood gas, indirect calorimetry, and cardiac output data can quantify pulmonary gas exchange in terms describing a 20 compartment V/Q model of pulmonary blood flow. High fidelity reports require data from two FiO2 settings.

Onset of Bloch oscillations in the almost-strong-field regime.

In the field of high-order harmonic generation from solids, the electron motion typically exceeds the edge of the first Brillouin zone. In conventional nonlinear optics, on the other hand, the excursion of band electrons is negligible. Here, we investigate the transition from conventional nonlinear optics to the regime where the crystal electrons begin to explore the first Brillouin zone. It is found that the nonlinear optical response changes abruptly already before intraband currents due to ionization become dominant. This is observed by an interference structure in the third-order harmonic generation of few-cycle pulses in a non-collinear geometry. Although approaching Keldysh parameter γ = 1, this is not a strong-field effect in the original sense, because the iterative series still converges and reproduces the interference structure. The change of the nonlinear interband response is attributed to Bloch motion of the reversible (or transient or virtual) population, similar to the Bloch motion of the irreversible (or real) population which affects the intraband currents that have been observed in high-order harmonic generation.

Resonant Perfect Absorption Yielded by Zero-Area Pulses.

We propose and study the manipulation of the macroscopic transient absorption of an ensemble of open two-level systems via temporal engineering. The key idea is to impose an ultrashort temporal gate on the polarization decay of the system by transient absorption spectroscopy, thus confining its free evolution and the natural reshaping of the excitation pulse. The numerical and analytical results demonstrate that even at moderate optical depths, the resonant absorption of light can be reduced or significantly enhanced by more than 5 orders of magnitude relative to that without laser manipulation. The achievement of the quasicomplete extinction of light at the resonant frequency, here referred to as resonant perfect absorption, arises from the full destructive interference between the excitation pulse and its subpulses developed and tailored during propagation, and is revealed to be connected with the formation of zero-area pulses in the time domain.

Anaerobic Fungal Mevalonate Pathway Genomic Biases Lead to Heterologous Toxicity Underpredicted by Codon Adaptation Indices.

Anaerobic fungi are emerging biotechnology platforms with genomes rich in biosynthetic potential. Yet, the heterologous expression of their biosynthetic pathways has had limited success in model hosts like E. coli. We find one reason for this is that the genome composition of anaerobic fungi like P. indianae are extremely AT-biased with a particular preference for rare and semi-rare AT-rich tRNAs in E coli, which are not explicitly predicted by standard codon adaptation indices (CAI). Native P. indianae genes with these extreme biases create drastic growth defects in E. coli (up to 69% reduction in growth), which is not seen in genes from other organisms with similar CAIs. However, codon optimization rescues growth, allowing for gene evaluation. In this manner, we demonstrate that anaerobic fungal homologs such as PI.atoB are more active than S. cerevisiae homologs in a hybrid pathway, increasing the production of mevalonate up to 2.5 g/L (more than two-fold) and reducing waste carbon to acetate by ~90% under the conditions tested. This work demonstrates the bioproduction potential of anaerobic fungal enzyme homologs and how the analysis of codon utilization enables the study of otherwise difficult to express genes that have applications in biocatalysis and natural product discovery.

Apoptosis is not conserved in plants as revealed by critical examination of a model for plant apoptosis-like cell death.

BACKGROUND: Animals and plants diverged over one billion years ago and evolved unique mechanisms for many cellular processes, including cell death. One of the most well-studied cell death programmes in animals, apoptosis, involves gradual cell dismantling and engulfment of cellular fragments, apoptotic bodies, through phagocytosis. However, rigid cell walls prevent plant cell fragmentation and thus apoptosis is not applicable for executing cell death in plants. Furthermore, plants are devoid of the key components of apoptotic machinery, including phagocytosis as well as caspases and Bcl-2 family proteins. Nevertheless, the concept of plant "apoptosis-like programmed cell death" (AL-PCD) is widespread. This is largely due to superficial morphological resemblances between plant cell death and apoptosis, and in particular between protoplast shrinkage in plant cells killed by various stimuli and animal cell volume decrease preceding fragmentation into apoptotic bodies. RESULTS: Here, we provide a comprehensive spatio-temporal analysis of cytological and biochemical events occurring in plant cells subjected to heat shock at 40-55 °C and 85 °C, the experimental conditions typically used to trigger AL-PCD and necrotic cell death, respectively. We show that cell death under both conditions was not accompanied by membrane blebbing or formation of apoptotic bodies, as would be expected during apoptosis. Instead, we observed instant and irreversible permeabilization of the plasma membrane and ATP depletion. These processes did not depend on mitochondrial functionality or the presence of Ca2+ and could not be prevented by an inhibitor of ferroptosis. We further reveal that the lack of protoplast shrinkage at 85 °C, the only striking morphological difference between cell deaths induced by 40-55 °C or 85 °C heat shock, is a consequence of the fixative effect of the high temperature on intracellular contents. CONCLUSIONS: We conclude that heat shock-induced cell death is an energy-independent process best matching definition of necrosis. Although the initial steps of this necrotic cell death could be genetically regulated, classifying it as apoptosis or AL-PCD is a terminological misnomer. Our work supports the viewpoint that apoptosis is not conserved across animal and plant kingdoms and demonstrates the importance of focusing on plant-specific aspects of cell death pathways.

The relationship of the Phantom® Lapidus Intramedullary Nail System to neurologic and tendinous structures in the foot: An anatomic study.

BACKGROUND: The purpose of our cadaveric study was to determine the proximity of nail insertion and interlocking mechanisms in the Phantom® Lapidus Intramedullary Nail System to neurologic and tendinous structures in the foot. METHODS: We used 10 fresh-frozen human lower-extremity specimen cadavers. For each specimen, the Nail System was inserted as described in the published technique guide. We then performed dissection on the tibialis anterior tendon, extensor hallucis longus tendon, and medial dorsal cutaneous branch of the superficial peroneal nerve and we measured and averaged the distances from each of these structures from the nail. RESULTS: The tibialis anterior tendon was in closest proximity to the insertion of the proximal medial interlock K-wire with an average distance of 0.4mm from the tendon. The extensor hallucis longus tendon was in closest proximity to nail insertion with an average distance of 1.2mm. The medial dorsal cutaneous branch of the superficial peroneal nerve was in closest proximity to the distal interlock K-wire with an average distance of 7.5mm. CONCLUSIONS: The tibialis anterior tendon, extensor hallucis longus tendon, and the medial dorsal cutaneous branch of the superficial peroneal nerve are at risk with the insertion of the nail system. Blunt dissection should be performed using this system with a path to bone before instrumentation to reduce the risk of nerve and tendon injury in the foot.

Systematic Review and Meta-Analysis of Racial and Ethnic Differences in Dementia Caregivers' Well-Being.

BACKGROUND AND OBJECTIVES: Studies comparing racial/ethnic differences on measures of psychological and physical well-being for dementia caregivers have reported differences between minority and white caregivers. Recruitment methods often differ for minority and white participants due to enrollment targets and may lead to biased comparisons, especially in convenience samples. We aimed to examine racial/ethnic differences in dementia caregiver outcomes and to determine whether differences vary between studies with population-based or convenience samples. RESEARCH DESIGN AND METHODS: We systematically reviewed articles with primary data from PubMed, Google Scholar, and PsycINFO. We included studies comparing African American or Hispanic/Latino to white dementia caregivers on measures of psychological well-being or physical well-being. Reviewers screened titles and abstracts, reviewed full texts and conducted risk-of-bias assessments. Meta-analyses were conducted to assess effects by race/ethnicity and study bias. RESULTS: A total of 159 effects were extracted from 38 studies, 2 of which were population based. Random-effects models revealed small but statistically significant effects with better psychological well-being in African American caregivers compared with white caregivers in both population-based (d = -0.22) and convenience sample studies (d = -0.21). Hispanics/Latino caregivers reported lower levels of physical well-being than white caregivers (d = 0.12), though these effects varied by level of rated study bias. DISCUSSION AND IMPLICATIONS: Consistency across study methods raises confidence in the validity of previous reports of better psychological well-being in African American caregivers. Future studies should use population-based samples with subgroups of Hispanic/Latino, Asian American, and American Indian caregivers that are culturally distinct on factors such as country of origin and tribe.