Longitudinal dynamics of farmer and livestock nasal and faecal microbiomes and resistomes.

Globally, half a billion people are employed in animal agriculture and are directly exposed to the associated microorganisms. However, the extent to which such exposures affect resident human microbiomes is unclear. Here we conducted a longitudinal profiling of the nasal and faecal microbiomes of 66 dairy farmers and 166 dairy cows over a year-long period. We compare farmer microbiomes to those of 60 age-, sex- and ZIP code-matched people with no occupational exposures to farm animals (non-farmers). We show that farming is associated with microbiomes containing livestock-associated microbes; this is most apparent in the nasal bacterial community, with farmers harbouring a richer and more diverse nasal community than non-farmers. Similarly, in the gut microbial communities, we identify more shared microbial lineages between cows and farmers from the same farms. Additionally, we find that shared microbes are associated with antibiotic resistance genes. Overall, our study demonstrates the interconnectedness of human and animal microbiomes.

Swimming kinematics of rainbow trout behind cylinder arrays: the effect of vortex street periodicity and turbulence kinetic energy.

Fish in the wild often contend with complex flows that are produced by natural and artificial structures. Research into fish interactions with turbulence often investigates metrics such as turbulence kinetic energy (TKE) or fish positional location, with less attention paid to the specific interactions between vortex organization and body swimming kinematics. Here we compare the swimming kinematics of rainbow trout ( Oncorhynchus mykiss ) holding station in flows produced by two different 3 x 5 cylinder arrays. We systematically utilized computational fluid dynamics to generate one array that produced a Kármán vortex street with high vortex periodicity and TKE (KVS array), and another that produced low periodicity and TKE, similar to a parallel vortex street (PVS array). The only difference in swimming kinematics between cylinder arrays was an increased tail beat amplitude in the KVS array. In both cylinder arrays, the tail beat frequency decreased and snout amplitude increased compared with the freestream. The center of mass amplitude was greater in the PVS array than in only the freestream, however, suggesting some buffeting of the body by the fluid. Notably, we did not observe Kármán gaiting in the KVS array as in previous studies. We hypothesize that this is because (1) vorticity was dissipated in the region where fish held station in this study and (2) cylinder arrays produced vortices that were in-line rather than staggered. These results are the first to quantify the kinematics and behavior of fishes swimming in the wake of multiple cylinder arrays, which has important implications for biomechanics, fluid dynamics, and fisheries management. SUMMARY STATEMENT: The swimming kinematics of rainbow trout are largely preserved across two, 3 x 5 cylinder array treatments that differed in vortex periodicity and turbulence kinetic energy.

Fantastic beasts and how to study them: rethinking experimental animal behavior.

Humans have been trying to understand animal behavior at least since recorded history. Recent rapid development of new technologies has allowed us to make significant progress in understanding the physiological and molecular mechanisms underlying behavior, a key goal of neuroethology. However, there is a tradeoff when studying animal behavior and its underlying biological mechanisms: common behavior protocols in the laboratory are designed to be replicable and controlled, but they often fail to encompass the variability and breadth of natural behavior. This Commentary proposes a framework of 10 key questions that aim to guide researchers in incorporating a rich natural context into their experimental design or in choosing a new animal study system. The 10 questions cover overarching experimental considerations that can provide a template for interspecies comparisons, enable us to develop studies in new model organisms and unlock new experiments in our quest to understand behavior.

Kinematics and muscle activity of pectoral fins in rainbow trout (Oncorhynchus mykiss) station holding in turbulent flow.

Pectoral fins play a crucial role in fish locomotion. Despite fishes living in complex fluid environments that exist in rivers and tidal flows, the role of the pectoral fins in navigating turbulent flows is not well understood. This study investigated the kinematics and muscle activity of pectoral fins in rainbow trout as they held station in the unsteady flows behind a D-section cylinder. We observed two distinct pectoral fin behaviors, one during braking and the other during Kármán gaiting. These behaviors were correlated to whole-body movements in response to the hydrodynamic conditions of specific regions in the cylinder wake. Sustained fin extensions during braking, where the fin was held out to maintain its position away from the body and against the flow, were associated with the cessation of forward body velocity, where the fish avoided the suction region directly downstream of the cylinder. Transient fin extensions and retractions during Kármán gaiting controlled body movements in the cross-stream direction. These two fin behaviors had different patterns of muscle activity. All braking events required recruitment from both the abductor and adductor musculature to actively extend a pectoral fin. In contrast, over 50% of fin extension movements during Kármán gaiting proceed in the absence of muscle activity. We reveal that in unsteady fluid environments, pectoral fin movements are the result of a complex combination of passive and active mechanisms that deviate substantially from canonical labriform locomotion, the implications of which await further work on the integration of sensory and motor systems.

ROCK1/2 signaling contributes to corticosteroid-refractory acute graft-versus-host disease.

Patients with corticosteroid-refractory acute graft-versus-host disease (aGVHD) have a low one-year survival rate. Identification and validation of novel targetable kinases in patients who experience corticosteroid-refractory-aGVHD may help improve outcomes. Kinase-specific proteomics of leukocytes from patients with corticosteroid-refractory-GVHD identified rho kinase type 1 (ROCK1) as the most significantly upregulated kinase. ROCK1/2 inhibition improved survival and histological GVHD severity in mice and was synergistic with JAK1/2 inhibition, without compromising graft-versus-leukemia-effects. ROCK1/2-inhibition in macrophages or dendritic cells prior to transfer reduced GVHD severity. Mechanistically, ROCK1/2 inhibition or ROCK1 knockdown interfered with CD80, CD86, MHC-II expression and IL-6, IL-1ß, iNOS and TNF production in myeloid cells. This was accompanied by impaired T cell activation by dendritic cells and inhibition of cytoskeletal rearrangements, thereby reducing macrophage and DC migration. NF-κB signaling was reduced in myeloid cells following ROCK1/2 inhibition. In conclusion, ROCK1/2 inhibition interferes with immune activation at multiple levels and reduces acute GVHD while maintaining GVL-effects, including in corticosteroid-refractory settings.

Role of Rho-Associated Kinase in the Pathophysiology of Cerebral Cavernous Malformations.

Cerebral cavernous malformations (CCMs) are vascular lesions characterized by a porous endothelium. The lack of a sufficient endothelial barrier can result in microbleeds and frank intracerebral hemorrhage. A primary mechanism for lesion development is a sequence variant in at least 1 of the 3 CCM genes (CCM1, CCM2, and CCM3), which influence various signaling pathways that lead to the CCM phenotype. A common downstream process associated with CCM gene loss of function involves overactivation of RhoA and its effector Rho-associated kinase (ROCK). In this study, we review RhoA/ROCK-related mechanisms involved in CCM pathophysiology as potential therapeutic targets. Literature searches were conducted in PubMed using combinations of search terms related to RhoA/ROCK and CCMs. In endothelial cells, CCM1, CCM2, and CCM3 proteins normally associate to form the CCM protein complex, which regulates the functions of a wide variety of protein targets (e.g., MAP3K3, SMURF1, SOK-1, and ICAP-1) that directly or indirectly increase RhoA/ROCK activity. Loss of CCM complex function and increased RhoA/ROCK activity can lead to the formation of stress fibers that contribute to endothelial junction instability. Other RhoA/ROCK-mediated pathophysiologic outcomes include a shift to a senescence-associated secretory phenotype (primarily mediated by ROCK2), which is characterized by endothelial cell migration, cell cycle arrest, extracellular matrix degradation, leukocyte chemotaxis, and inflammation. ROCK represents a potential therapeutic target, and direct (fasudil, NRL-1049) and indirect (statins) ROCK inhibitors have demonstrated various levels of efficacy in reducing lesion burden in preclinical models of CCM. Current (atorvastatin) and planned (NRL-1049) clinical studies will determine the efficacy of ROCK inhibitors for CCM in humans, for which no US Food and Drug Administration-approved or EU-approved pharmacologic treatment exists.

Effects of n-butanol production on metabolism and the photosystem in Synecococcus elongatus PCC 7942 based on metabolic flux and target proteome analyses.

Although n-butanol (BuOH) is an ideal fuel because of its superior physical properties, it has toxicity to microbes. Previously, a Synechococcus elongatus PCC 7942 derivative strain that produces BuOH from CO2 was developed by introducing six heterologous genes (BUOH-SE strain). To identify the bottleneck in BuOH production, the effects of BuOH production and its toxicity on central metabolism and the photosystem were investigated. Parental (WT) and BUOH-SE strains were cultured under autotrophic conditions. Consistent with the results of a previous study, BuOH production was observed only in the BUOH-SE strain. Isotopically non-stationary 13C-metabolic flux analysis revealed that the CO2 fixation rate was much larger than the BuOH production rate in the BUOH-SE strain (1.70 vs 0.03 mmol gDCW-1 h-1), implying that the carbon flow for BuOH biosynthesis was less affected by the entire flux distribution. No large difference was observed in the flux of metabolism between the WT and BUOH-SE strains. Contrastingly, in the photosystem, the chlorophyll content and maximum O2 evolution rate per dry cell weight of the BUOH-SE strain were decreased to 81% and 43% of the WT strain, respectively. Target proteome analysis revealed that the amounts of some proteins related to antennae (ApcA, ApcD, ApcE, and CpcC), photosystem II (PsbB, PsbU, and Psb28-2), and cytochrome b6f complex (PetB and PetC) in photosystems decreased in the BUOH-SE strain. The activation of photosynthesis would be a novel approach for further enhancing BuOH production in S. elongatus PCC 7942.

Anion efflux mediates transduction in the hair cells of the zebrafish lateral line.

Hair cells are the principal sensory receptors of the vertebrate auditory system, where they transduce sounds through mechanically gated ion channels that permit cations to flow from the surrounding endolymph into the cells. The lateral line of zebrafish has served as a key model system for understanding hair cell physiology and development, often with the belief that these hair cells employ a similar transduction mechanism. In this study, we demonstrate that these hair cells are exposed to an unregulated external environment with cation concentrations that are too low to support transduction. Our results indicate that hair cell excitation is instead mediated by a substantially different mechanism involving the outward flow of anions. Further investigation of hair cell transduction in a diversity of sensory systems and species will likely yield deep insights into the physiology of these unique cells.

Establishing a framework for quality of inpatient care for Parkinson's disease: A study on inpatient medication administration.

BACKGROUND: The complexity of antiparkinsonian medications makes patients vulnerable to medication deviations. This study examines the frequency and outcomes of deviations between outpatient and inpatient medication administrations in patients with Parkinson's disease (PD). METHODS: We included hospital admissions of patients with PD during a 12-month period at the Cleveland Clinic Main and Fairview campuses. Outpatient regimens were compared with hospital medication administration records to establish rates of deviations in terms of levodopa equivalent daily dose (LEDD) difference, timing deviations/omissions of time-critical medications, substitution of levodopa compounds, and administration of antidopaminergic medications. Logistic regression analyses were used to investigate associations with length of stay (LOS), readmission rates, and mortality. RESULTS: The study included 492 patients with 725 admissions. Of those on time-critical medications, 43% had a LEDD deviation and 19% had levodopa formulation substitutions. Of the admission days with known outpatient timing regimens, 47% had an average deviation of more than 30 min and 22% had at least one missed levodopa dose. LOS was longer with each additional day of over-dose (4%), under-dose (14%), missed dose (21%), timing deviation (15%) and substitution (19%), (all p < 0.0001). Administration of antidopaminergic medications (9.9% of admissions) was associated with increased 30-day readmission/death (OR 1.85, p = 0.041), 90-day mortality (OR 2.2, p = 0.018), and LOS (7.6 vs. 3.8 days, p < 0.0001). LEDD underdose was associated with 30-day readmission/death (OR 1.78, p = 0.025) and 90-day mortality (OR 1.14, CI 1.05-1.24, p = 0.002). CONCLUSIONS: Deviations between outpatient and hospital regimens, and administration of antidopaminergic medications, were associated with poor outcomes.

Chylomicrons Regulate Lacteal Permeability and Intestinal Lipid Absorption.

BACKGROUND: Lymphatic vessels are responsible for tissue drainage, and their malfunction is associated with chronic diseases. Lymph uptake occurs via specialized open cell-cell junctions between capillary lymphatic endothelial cells (LECs), whereas closed junctions in collecting LECs prevent lymph leakage. LEC junctions are known to dynamically remodel in development and disease, but how lymphatic permeability is regulated remains poorly understood. METHODS: We used various genetically engineered mouse models in combination with cellular, biochemical, and molecular biology approaches to elucidate the signaling pathways regulating junction morphology and function in lymphatic capillaries. RESULTS: By studying the permeability of intestinal lacteal capillaries to lipoprotein particles known as chylomicrons, we show that ROCK (Rho-associated kinase)-dependent cytoskeletal contractility is a fundamental mechanism of LEC permeability regulation. We show that chylomicron-derived lipids trigger neonatal lacteal junction opening via ROCK-dependent contraction of junction-anchored stress fibers. LEC-specific ROCK deletion abolished junction opening and plasma lipid uptake. Chylomicrons additionally inhibited VEGF (vascular endothelial growth factor)-A signaling. We show that VEGF-A antagonizes LEC junction opening via VEGFR (VEGF receptor) 2 and VEGFR3-dependent PI3K (phosphatidylinositol 3-kinase)/AKT (protein kinase B) activation of the small GTPase RAC1 (Rac family small GTPase 1), thereby restricting RhoA (Ras homolog family member A)/ROCK-mediated cytoskeleton contraction. CONCLUSIONS: Our results reveal that antagonistic inputs into ROCK-dependent cytoskeleton contractions regulate the interconversion of lymphatic junctions in the intestine and in other tissues, providing a tunable mechanism to control the lymphatic barrier.

An ATP-sensitive phosphoketolase regulates carbon fixation in cyanobacteria.

Regulation of CO2 fixation in cyanobacteria is important both for the organism and global carbon balance. Here we show that phosphoketolase in Synechococcus elongatus PCC7942 (SeXPK) possesses a distinct ATP-sensing mechanism, where a drop in ATP level allows SeXPK to divert precursors of the RuBisCO substrate away from the Calvin-Benson-Bassham cycle. Deleting the SeXPK gene increased CO2 fixation particularly during light-dark transitions. In high-density cultures, the Δxpk strain showed a 60% increase in carbon fixation and unexpectedly resulted in sucrose secretion without any pathway engineering. Using cryo-EM analysis, we discovered that these functions were enabled by a unique allosteric regulatory site involving two subunits jointly binding two ATP, which constantly suppresses the activity of SeXPK until the ATP level drops. This magnesium-independent ATP allosteric site is present in many species across all three domains of life, where it may also play important regulatory functions.

Understanding the molecular mechanisms of statin pleiotropic effects.

Statins represent the cornerstone of pharmacotherapy for the prevention of atherosclerotic cardiovascular disease. These medications not only reduce low-density lipoprotein cholesterol (LDL-C) via inhibition of 3-hydroxy-3-methylglutarate attached to CoA reductase, the key rate-limiting step in the cholesterol biosynthetic pathway, but also upregulate expression of the low-density lipoprotein receptor, improving serum clearance. Given LDL-C is a causal risk factor for the development of atherosclerosis, these complementary mechanisms largely explain why statin therapy leads to reductions in major adverse cardiovascular events. However, decades of basic and clinical research have suggested that statins may exert other effects independent of LDL-C lowering, termed pleiotropic effects, which have become a topic of debate among the scientific community. While some literature suggests statins may improve plaque stability, reduce inflammation and thrombosis, decrease oxidative stress, and improve endothelial function and vascular tone, other studies have suggested potential harmful pleiotropic effects related to increased risk of muscle-related side effects, diabetes, hemorrhagic stroke, and cognitive decline. Furthermore, the introduction of newer, non-statin LDL-C lowering therapies, including ezetimibe, proprotein convertase subtilisin/Kexin Type 9, and bempedoic acid, have challenged the statin pleiotropy theory. This review aims to provide a historical background on the development of statins, explore the mechanistic underpinnings of statin pleiotropy, review the available literature, and provide up to date examples that suggest statins may exert effects outside of LDL-C lowering and the cardiovascular system.

Recording central nervous system responses of freely-swimming marine and freshwater fishes with a customizable, implantable AC differential amplifier.

BACKGROUND: Fish have adapted to a diversity of environments but the neural mechanisms underlying natural aquatic behaviors are not well known. NEW METHOD: We have developed a small, customizable AC differential amplifier and surgical procedures for recording multi-unit extracellular signals in the CNS of marine and freshwater fishes. RESULTS: Our minimally invasive amplifier allowed fish to orient to flow and respond to hydrodynamic and visual stimuli. We recorded activity in the cerebellum and optic tectum during these behaviors. COMPARISON WITH EXISTING METHODS: Our system is very low-cost, hydrodynamically streamlined, and capable of high-gain in order to allow for recordings from freely behaving, fast fishes in complex fluid environments. CONCLUSIONS: Our tethered approach allows access to record neural activity in a diversity of adult fishes in the lab, but can also be modified for data logging in the field.

A Novel HIF-2α/ARNT Signaling Pathway Protects Against Microvascular Dysfunction and heart failure After Myocardial Infarction.

RATIONALE: Cardiac microvascular leakage and inflammation are triggered during myocardial infarction (MI) and contribute to heart failure. Hypoxia-inducible factor 2α (Hif2α) is highly expressed in endothelial cells (ECs) and rapidly activated by myocardial ischemia, but whether it has a role in endothelial barrier function during MI is unclear. OBJECTIVE: To test our hypothesis that the expression of Hif2α and its binding partner aryl hydrocarbon nuclear translocator (ARNT) in ECs regulate cardiac microvascular permeability in infarcted hearts. METHODS AND RESULTS: Experiments were conducted with mice carrying an inducible EC-specific Hif2α-knockout (ecHif2α-/-) mutation, with mouse cardiac microvascular endothelial cells (CMVECs) isolated from the hearts of ecHif2α-/- mice after the mutation was induced, and with human CMVECs and umbilical-vein endothelial cells transfected with ecHif2α siRNA. After MI induction, echocardiographic assessments of cardiac function were significantly lower, while measures of cardiac microvascular leakage (Evans blue assay), plasma IL6 levels, and cardiac neutrophil accumulation and fibrosis (histology) were significantly greater, in ecHif2α-/- mice than in control mice, and RNA-sequencing analysis of heart tissues from both groups indicated that the expression of genes involved in vascular permeability and collagen synthesis was enriched in ecHif2α-/- hearts. In cultured ECs, ecHif2α deficiency was associated with declines in endothelial barrier function (electrical cell impedance assay) and the reduced abundance of tight-junction proteins, as well as an increase in the expression of inflammatory markers, all of which were largely reversed by the overexpression of ARNT. We also found that ARNT, but not Hif2α, binds directly to the IL6 promoter and suppresses IL6 expression. CONCLUSIONS: EC-specific deficiencies in Hif2α expression significantly increase cardiac microvascular permeability, promote inflammation, and reduce cardiac function in infarcted mouse hearts, and ARNT overexpression can reverse the upregulation of inflammatory genes and restore endothelial-barrier function in Hif2α-deficient ECs.

Tie2-Cre-Induced Inactivation of Non-Nuclear Estrogen Receptor-α Signaling Abrogates Estrogen Protection Against Vascular Injury.

Using the Cre-loxP system, we generated the first mouse model in which estrogen receptor-α non-nuclear signaling was inactivated in endothelial cells. Estrogen protection against mechanical vascular injury was impaired in this model. This result indicates the pivotal role of endothelial estrogen receptor-α non-nuclear signaling in the vasculoprotective effects of estrogen.

SARS-CoV-2 N protein mediates intercellular nucleic acid dispersion, a feature reduced in Omicron.

The coronavirus nucleocapsid (N) protein is known to bind to nucleic acids and facilitate viral genome encapsulation. Here we report that the N protein can mediate RNA or DNA entering neighboring cells through ACE2-independent, receptor (STEAP2)-mediated endocytosis, and achieve gene expression. The effect is more pronounced for the N protein of wild-type SARS-CoV-2 than that of the Omicron variant and other human coronaviruses. This effect is enhanced by RANTES (CCL5), a chemokine induced by N protein, and lactate, a metabolite produced in hypoxia, to cause more damage. These findings might explain the clinical observations in SARS-CoV-2-infected cases. Moreover, the N protein-mediated function can be inhibited by N protein-specific monoclonal antibodies or p38 mitogen-activated protein kinase inhibitors. Since the N-protein-mediated nucleic acid endocytosis involves a receptor commonly expressed in many types of cells, our findings suggest that N protein may have an additional role in SARS-CoV-2 pathogenesis.

Climate change challenges, plant science solutions.

Climate change is a defining challenge of the 21st century, and this decade is a critical time for action to mitigate the worst effects on human populations and ecosystems. Plant science can play an important role in developing crops with enhanced resilience to harsh conditions (e.g. heat, drought, salt stress, flooding, disease outbreaks) and engineering efficient carbon-capturing and carbon-sequestering plants. Here, we present examples of research being conducted in these areas and discuss challenges and open questions as a call to action for the plant science community.