The gut microbiota promotes distal tissue regeneration via RORγ+ regulatory T cell emissaries.

Specific microbial signals induce the differentiation of a distinct pool of RORγ+ regulatory T (Treg) cells crucial for intestinal homeostasis. We discovered highly analogous populations of microbiota-dependent Treg cells that promoted tissue regeneration at extra-gut sites, notably acutely injured skeletal muscle and fatty liver. Inflammatory meditators elicited by tissue damage combined with MHC-class-II-dependent T cell activation to drive the accumulation of gut-derived RORγ+ Treg cells in injured muscle, wherein they regulated the dynamics and tenor of early inflammation and helped balance the proliferation vs. differentiation of local stem cells. Reining in IL-17A-producing T cells was a major mechanism underlying the rheostatic functions of RORγ+ Treg cells in compromised tissues. Our findings highlight the importance of gut-trained Treg cell emissaries in controlling the response to sterile injury of non-mucosal tissues.

Fossil evidence for a pharyngeal origin of the vertebrate pectoral girdle.

The origin of vertebrate paired appendages is one of the most investigated and debated examples of evolutionary novelty1-7. Paired appendages are widely considered as key innovations that enabled new opportunities for controlled swimming and gill ventilation and were prerequisites for the eventual transition from water to land. The past 150 years of debate8-10 has been shaped by two contentious theories4,5: the ventrolateral fin-fold hypothesis9,10 and the archipterygium hypothesis8. The latter proposes that fins and girdles evolved from an ancestral gill arch. Although studies in animal development have revived interest in this idea11-13, it is apparently unsupported by fossil evidence. Here we present palaeontological support for a pharyngeal basis for the vertebrate shoulder girdle. We use computed tomography scanning to reveal details of the braincase of Kolymaspis sibirica14, an Early Devonian placoderm fish from Siberia, that suggests a pharyngeal component of the shoulder. We combine these findings with refreshed comparative anatomy of placoderms and jawless outgroups to place the origin of the shoulder girdle on the sixth branchial arch. These findings provide a novel framework for understanding the origin of the pectoral girdle. Our evidence clarifies the location of the presumptive head-trunk interface in jawless fishes and explains the constraint on branchial arch number in gnathostomes15. The results revive a key aspect of the archipterygium hypothesis and help reconcile it with the ventrolateral fin-fold model.

MicroRNAs Cause Accelerated Decay of Short-Tailed Target mRNAs.

MicroRNAs (miRNAs) specify the recruitment of deadenylases to mRNA targets. Despite this recruitment, we find that miRNAs have almost no effect on steady-state poly(A)-tail lengths of their targets in mouse fibroblasts, which motivates the acquisition of pre-steady-state measurements of the effects of miRNAs on tail lengths, mRNA levels, and translational efficiencies. Effects on translational efficiency are minimal compared to effects on mRNA levels, even for newly transcribed target mRNAs. Effects on target mRNA levels accumulate as the mRNA population approaches steady state, whereas effects on tail lengths peak for recently transcribed target mRNAs and then subside. Computational modeling of this phenomenon reveals that miRNAs cause not only accelerated deadenylation of their targets but also accelerated decay of short-tailed target molecules. This unanticipated effect of miRNAs largely prevents short-tailed target mRNAs from accumulating despite accelerated target deadenylation. The net result is a nearly imperceptible change to the steady-state tail-length distribution of targeted mRNAs.

The Dynamics of Cytoplasmic mRNA Metabolism.

For all but a few mRNAs, the dynamics of metabolism are unknown. Here, we developed an experimental and analytical framework for examining these dynamics for mRNAs from thousands of genes. mRNAs of mouse fibroblasts exit the nucleus with diverse intragenic and intergenic poly(A)-tail lengths. Once in the cytoplasm, they have a broad (1000-fold) range of deadenylation rate constants, which correspond to cytoplasmic lifetimes. Indeed, with few exceptions, degradation appears to occur primarily through deadenylation-linked mechanisms, with little contribution from either endonucleolytic cleavage or deadenylation-independent decapping. Most mRNA molecules degrade only after their tail lengths fall below 25 nt. Decay rate constants of short-tailed mRNAs vary broadly (1000-fold) and are larger for short-tailed mRNAs that have previously undergone more rapid deadenylation. This coupling helps clear rapidly deadenylated mRNAs, enabling the large range in deadenylation rate constants to impart a similarly large range in stabilities.

Bizarre tail weaponry in a transitional ankylosaur from subantarctic Chile.

Armoured dinosaurs are well known for their evolution of specialized tail weapons-paired tail spikes in stegosaurs and heavy tail clubs in advanced ankylosaurs1. Armoured dinosaurs from southern Gondwana are rare and enigmatic, but probably include the earliest branches of Ankylosauria2-4. Here we describe a mostly complete, semi-articulated skeleton of a small (approximately 2 m) armoured dinosaur from the late Cretaceous period of Magallanes in southernmost Chile, a region that is biogeographically related to West Antarctica5. Stegouros elengassen gen. et sp. nov. evolved a large tail weapon unlike any dinosaur: a flat, frond-like structure formed by seven pairs of laterally projecting osteoderms encasing the distal half of the tail. Stegouros shows ankylosaurian cranial characters, but a largely ancestral postcranial skeleton, with some stegosaur-like characters. Phylogenetic analyses placed Stegouros in Ankylosauria; specifically, it is related to Kunbarrasaurus from Australia6 and Antarctopelta from Antarctica7, forming a clade of Gondwanan ankylosaurs that split earliest from all other ankylosaurs. The large osteoderms and specialized tail vertebrae in Antarctopelta suggest that it had a tail weapon similar to Stegouros. We propose a new clade, the Parankylosauria, to include the first ancestor of Stegouros-but not Ankylosaurus-and all descendants of that ancestor.

Endothelial-derived angiocrine signals induce and sustain regenerative lung alveolarization.

To identify pathways involved in adult lung regeneration, we employ a unilateral pneumonectomy (PNX) model that promotes regenerative alveolarization in the remaining intact lung. We show that PNX stimulates pulmonary capillary endothelial cells (PCECs) to produce angiocrine growth factors that induce proliferation of epithelial progenitor cells supporting alveologenesis. Endothelial cells trigger expansion of cocultured epithelial cells, forming three-dimensional angiospheres reminiscent of alveolar-capillary sacs. After PNX, endothelial-specific inducible genetic ablation of Vegfr2 and Fgfr1 in mice inhibits production of MMP14, impairing alveolarization. MMP14 promotes expansion of epithelial progenitor cells by unmasking cryptic EGF-like ectodomains that activate the EGF receptor (EGFR). Consistent with this, neutralization of MMP14 impairs EGFR-mediated alveolar regeneration, whereas administration of EGF or intravascular transplantation of MMP14(+) PCECs into pneumonectomized Vegfr2/Fgfr1-deficient mice restores alveologenesis and lung inspiratory volume and compliance function. VEGFR2 and FGFR1 activation in PCECs therefore increases MMP14-dependent bioavailability of EGFR ligands to initiate and sustain alveologenesis.

A giant soft-shelled egg from the Late Cretaceous of Antarctica.

Egg size and structure reflect important constraints on the reproductive and life-history characteristics of vertebrates1. More than two-thirds of all extant amniotes lay eggs2. During the Mesozoic era (around 250 million to 65 million years ago), body sizes reached extremes; nevertheless, the largest known egg belongs to the only recently extinct elephant bird3, which was roughly 66 million years younger than the last nonavian dinosaurs and giant marine reptiles. Here we report a new type of egg discovered in nearshore marine deposits from the Late Cretaceous period (roughly 68 million years ago) of Antarctica. It exceeds all nonavian dinosaur eggs in volume and differs from them in structure. Although the elephant bird egg is slightly larger, its eggshell is roughly five times thicker and shows a substantial prismatic layer and complex pore structure4. By contrast, the new fossil, visibly collapsed and folded, presents a thin eggshell with a layered structure that lacks a prismatic layer and distinct pores, and is similar to that of most extant lizards and snakes (Lepidosauria)5. The identity of the animal that laid the egg is unknown, but these preserved morphologies are consistent with the skeletal remains of mosasaurs (large marine lepidosaurs) found nearby. They are not consistent with described morphologies of dinosaur eggs of a similar size class. Phylogenetic analyses of traits for 259 lepidosaur species plus outgroups suggest that the egg belonged to an individual that was at least 7 metres long, hypothesized to be a giant marine reptile, all clades of which have previously been proposed to show live birth6. Such a large egg with a relatively thin eggshell may reflect derived constraints associated with body shape, reproductive investment linked with gigantism, and lepidosaurian viviparity, in which a 'vestigial' egg is laid and hatches immediately7.

Aire Inhibits the Generation of a Perinatal Population of Interleukin-17A-Producing γδ T Cells to Promote Immunologic Tolerance.

Aire's primary mechanism of action is to regulate transcription of a battery of genes in medullary thymic epithelial cells (mTECs) and, consequently, negative selection of effector T cells and positive selection of regulatory T cells. We found that Aire-deficient mice had expanded thymic and peripheral populations of perinatally generated IL-17A+Vγ6+Vδ1+ T cells, considered to be "early responders" to tissue stress and drivers of inflammatory reactions. Aire-dependent control of Il7 expression in mTECs regulated the size of thymic IL-17A+Vγ6+Vδ1+ compartments. In mice lacking Aire and γδ T cells, certain tissues typically targeted in the "Aire-less" disease, notably the retina, were only minimally infiltrated. IL-17A+Vγ6+Vδ1+ cells were present in the retina of wild-type mice and expanded very early in Aire-deficient mice. A putatively parallel population of IL-17A+Vγ9+Vδ2+ T cells was increased in humans lacking Aire. Thus, Aire exerts multi-faceted autoimmune control that extends to a population of innate-like T cells.

Lateral Flow Assay Biotesting by Utilizing Plasmonic Nanoparticles Made of Inexpensive Metals─Replacing Colloidal Gold.

Nanoparticles (NPs) can be conjugated with diverse biomolecules and employed in biosensing to detect target analytes in biological samples. This proven concept was primarily used during the COVID-19 pandemic with gold-NP-based lateral flow assays (LFAs). Considering the gold price and its worldwide depletion, here we show that novel plasmonic NPs based on inexpensive metals, titanium nitride (TiN) and copper covered with a gold shell (Cu@Au), perform comparable to or even better than gold nanoparticles. After conjugation, these novel nanoparticles provided high figures of merit for LFA testing, such as high signals and specificity and robust naked-eye signal recognition. Since the main cost of Au NPs in commercial testing kits is the colloidal synthesis, our development with the Cu@Au and the laser-ablation-fabricated TiN NPs is exciting, offering potentially inexpensive plasmonic nanomaterials for various bioapplications. Moreover, our machine learning study showed that biodetection with TiN is more accurate than that with Au.

FcγRIIA-specific DARPins as novel tools in blood cell analysis and platelet aggregation.

Fc receptors are involved in a variety of physiologically and disease-relevant responses. Among them, FcγRIIA (CD32a) is known for its activating functions in pathogen recognition and platelet biology, and, as potential marker of T lymphocytes latently infected with HIV-1. The latter has not been without controversy due to technical challenges complicated by T-B cell conjugates and trogocytosis as well as a lack of antibodies distinguishing between the closely related isoforms of FcγRII. To generate high-affinity binders specific for FcγRIIA, libraries of designed ankyrin repeat proteins (DARPins) were screened for binding to its extracellular domains by ribosomal display. Counterselection against FcγRIIB eliminated binders cross-reacting with both isoforms. The identified DARPins bound FcγRIIA with no detectable binding for FcγRIIB. Their affinities for FcγRIIA were in the low nanomolar range and could be enhanced by cleavage of the His-tag and dimerization. Interestingly, complex formation between DARPin and FcγRIIA followed a two-state reaction model, and discrimination from FcγRIIB was based on a single amino acid residue. In flow cytometry, DARPin F11 detected FcγRIIA+ cells even when they made up less than 1% of the cell population. Image stream analysis of primary human blood cells confirmed that F11 caused dim but reliable cell surface staining of a small subpopulation of T lymphocytes. When incubated with platelets, F11 inhibited their aggregation equally efficient as antibodies unable to discriminate between both FcγRII isoforms. The selected DARPins are unique novel tools for platelet aggregation studies as well as the role of FcγRIIA for the latent HIV-1 reservoir.

Prognostic and predictive impact of metastatic organ involvement on maintenance therapy in advanced metastatic colorectal cancer: Subgroup analysis of patients treated within the PanaMa trial (AIO KRK 0212).

Despite molecular selection, patients (pts) with RAS wildtype mCRC represent a heterogeneous population including diversity in metastatic spread. We investigated metastatic patterns for their prognostic and predictive impact on maintenance therapy with 5-fluorouracil/folinic acid ± panitumumab. The study population was stratified according to (1) number of involved metastatic sites (single vs multiple organ metastasis), liver-limited disease vs (2) liver metastasis plus one additional site, and (3) vs liver metastasis plus ≥two additional sites. Kaplan-Meier method and Cox regressions were used to correlate efficacy endpoints. Single organ metastasis was observed in 133 pts (53.6%) with 102 pts (41.1%) presenting with liver-limited disease, while multiple organ metastases were reported in 114 pts (46.0). Multiple compared to single organ metastases were associated with less favorable PFS (HR 1.48, 95% CI 1.13-1.93; P = .004) and OS (HR 1.37, 95% CI 0.98-1.93; P = .068) of maintenance therapy. While metastatic spread involving one additional extrahepatic site was not associated with clearly impaired survival compared to liver-limited disease, pts with liver metastasis plus ≥two additional sites demonstrated less favorable PFS (HR 1.92, 95% CI 1.30-2.83; P < .001), and OS (HR 2.38, 95% CI 1.51-3.76; P < .001) of maintenance therapy. Pmab-containing maintenance therapy appeared active in both pts with multiple (HR 0.58; 95% CI, 0.39-0.86; P = .006) as well as to a lesser numerical extent in pts with single organ metastasis (HR 0.83; 95% CI, 0.57-1.21; P = .332; Interaction P = .183). These data may support clinical decisions when EGFR-based maintenance therapy is considered.

The Autophagic and Apoptotic Death of Forebrain Neurons of Rats with Global Brain Ischemia Is Diminished by the Intranasal Administration of Insulin: Possible Mechanism of Its Action.

Insulin is a promising neuroprotector. To better understand the mechanism of insulin action, it was important to show its ability to diminish autophagic neuronal death in animals with brain ischemic and reperfusion injury. In forebrain ischemia and reperfusion, the number of live neurons in the hippocampal CA1 region and frontal cortex of rats decreased to a large extent. Intracerebroventricular administration of the autophagy and apoptosis inhibitors to ischemic rats significantly increased the number of live neurons and showed that the main part of neurons died from autophagy and apoptosis. Intranasal administration of 0.5 IU of insulin per rat (before ischemia and daily during reperfusion) increased the number of live neurons in the hippocampal CA1 region and frontal brain cortex. In addition, insulin significantly diminished the level of autophagic marker LC3B-II in these forebrain regions, which markedly increased during ischemia and reperfusion. Our studies demonstrated for the first time the ability of insulin to decrease autophagic neuronal death, caused by brain ischemia and reperfusion. Insulin administered intranasally activated the Akt-kinase (activating the mTORC1 complex, which inhibits autophagy) and inhibited the AMP-activated protein kinase (which activates autophagy) in the hippocampus and frontal cortex of rats with brain ischemia and reperfusion.

The last ceratosaur of Asia: a new noasaurid from the Early Cretaceous Great Siberian Refugium.

The noasaurid ceratosaur Kiyacursor longipes gen. et sp. nov. is described based on a fragmentary skeleton including cervical vertebra, pectoral girdle, humerus and hind limbs from the Lower Cretaceous (Aptian) Ilek Formation at Shestakovo 1 locality in Western Siberia, Russia. This is the first ceratosaur from the Early Cretaceous of Asia, extending the stratigraphic range of Ceratosauria by 40 Myr on that continent. Kiyacursor shares unique hind limb proportions with Elaphrosaurus and Limusaurus, suggesting improved cursorial ability. These taxa show an ostrich-like specialization of the pes, with a large third metatarsal and greatly reduced second metatarsal. By contrast, all other fast running non-avian theropod dinosaurs have an arctometatarsalian pes, with the third metatarsal strongly reduced proximally. The new taxon lived in the Early Cretaceous ecosystem containing a number of other Jurassic relics, such as stem salamanders, protosuchian and shartegosuchid crocodyliforms, tritylodontid synapsids and docodontan mammaliaforms.

Heterologous pulcherrimin production in Saccharomyces cerevisiae confers inhibitory activity on Botrytis conidiation.

Pulcherrimin is an iron (III) chelate of pulcherriminic acid that plays a role in antagonistic microbial interactions, iron metabolism, and stress responses. Some bacteria and yeasts produce pulcherriminic acid, but so far, pulcherrimin could not be produced in Saccharomyces cerevisiae. Here, multiple integrations of the Metschnikowia pulcherrima PUL1 and PUL2 genes in the S. cerevisiae genome resulted in red colonies, which indicated pulcherrimin formation. The coloration correlated positively and significantly with the number of PUL1 and PUL2 genes. The presence of pulcherriminic acid was confirmed by mass spectrometry. In vitro competition assays with the plant pathogenic fungus Botrytis caroliana revealed inhibitory activity on conidiation by an engineered, strong pulcherrimin-producing S. cerevisiae strain. We demonstrate that the PUL1 and PUL2 genes from M. pulcherrima, in multiple copies, are sufficient to transfer pulcherrimin production to S. cerevisiae and represent the starting point for engineering and optimizing this biosynthetic pathway in the future.

Quantitative intra- and intergeneric taxonomic relationships among Micrococcaceae strains reveal contradictions in the historical assignments of the strains and indicate the need for species reclassification.

This article reports the results of quantitative intra- and intergeneric taxonomic relationships among Micrococcaceae strains and a novel endophytic bacterium (SG) isolated from a suspension culture of Arabidopsis thaliana (L.) Heynh in our laboratory. The known strain Rothia sp. ND6WE1A was used as a reference one for SG. Whole-genome sequencing and phylogenetic analysis were based on the 16S rRNA test. Quantitative analysis for the nucleotide identity (ANI) and calculation of evolutionary distances were based on the identified amino acids (AAI) test indicating the generic assignment of the reference strain within and between the identified monophyletic groups of Micrococcaceae. The amino acid data structure of Rothia sp. ND6WE1A was compared against the UniProt database (250 million records) of close lineage of Micrococcaceae, including other Rothia spp. These data presented unique and evolutionary amino acid alignments, eventually expected in the new SG isolate as well. The metagenomic entries of the respective genome and proteome, characterized at the genus and species levels, could be considered for evolutionary taxonomic reclassification of the isolated and the reference strain (SG + Rothia sp. ND6WE1A). Therefore, our results warrant further investigations on the isolated SG strain.

Observing the Role of Electron Delocalization in Electronic Transport by Incorporating Actinides into Ligated Metal-Chalcogenide Superatoms.

Since delocalization of electronic states is a prerequisite for exerting unique electron transport properties, early actinides (An) with highly delocalized 5f/6d orbitals are natural candidates. However, given the experimental difficulties of such radioactive compounds and the complex relativistic effects in theoretical studies, understanding the electronic structure and bonding of actinides is underdeveloped on the periodic table. A further challenge is the very complicated electronic structures encountered in the confinement of actinides, as vividly illustrated by the weakly radioactive Th(Thorium)-encapsulated metal chalcogenide clusters, Th@Co6Te8L6 (L = PH3, PMe3, PEt3). Here we report the electronic structure and the electron transport properties of the Th@Co6Te8L6 clusters and compare them with those of the hollow Co6Te8L6 clusters using the nonequilibrium Green's function combined with relativistic density functional theory (NEGF-DFT). We found that the equilibrium conductance in Th@Co6Te8(PH3)6 (0.76 G0) has been greatly improved over that in Co6Te8(PH3)6 (0.03 G0), which has also been verified under an applied different bias voltage. The covalent bonding character between 6d (Th) and 3d (Co) atomic orbitals resulting from steric confinement is the source of the performance enhancement and a most important factor governing the accessibility of such 5f/6d orbitals. The results are of significance to the rapidly developing field of molecular nanoelectronics.

Interrupted Dance of Five Heteroatoms: Reinventing Ozonolysis to Make Geminal Alkoxyhydroperoxides from C═N Bonds.

Four heteroatoms dance in the cascade of four pericyclic reactions initiated by ozonolysis of C═N bonds. Switching from imines to semicarbazones introduces the fifth heteroatom that slows this dance, delays reaching the thermodynamically favorable escape path, and allows efficient interception of carbonyl oxides (Criegee intermediates, CIs) by an external nucleophile. The new three-component reaction of alcohols, ozone, and oximes/semicarbazones greatly facilitates synthetic access to monoperoxyacetals (alkoxyhydroperoxides).

Synthesis of ω-functionalized ketones from strained cyclic alcohols by ring-opening and cross-recombination between alkyl and N-oxyl radicals.

Radical ring-opening oxyimidation of cyclobutanols and cyclopropanols with the formation of ω-functionalized ketones was discovered. The oxidative C-O coupling proceeds via the interception of a primary alkyl radical generated from a cyclic alcohol with a reactive radical generated in situ, which is an electron-deficient N-oxyl radical. The developed conditions allow for the balanced generation rates of carbon- and N-oxyl radicals, which are necessary for their selective cross-recombination. Thus, typical competitive dimerization processes of carbon-centered radicals, their intermolecular cyclization, and N-oxyl radical self-decay are suppressed. The method is applicable to a wide range of cyclobutanols and results in oxyimidated ketones in yields of up to 82%.

Reshaping and induction of optical activity in gold@silver nanocuboids by chiral glutathione molecules.

Core-shell gold-silver cuboidal nanoparticles were produced, with either concave or straight facets. Their incubation with a low concentration of chiral l-glutathione (GSH) biomolecules was found to produce near UV plasmonic extinction and induced circular dichroism (CD) peaks. The effect is sensitive to the silver shell thickness. The GSH molecules were found to cause redistribution of silver in the shell, removing silver atoms from edges/corners and re-depositing them at the nanocuboid facets, probably through some redox and complexation processes between the silver and thiol group of the GSH. Other thiolated chiral biomolecules (and drug molecules) did not show this effect. The emerging near UV surface plasmon resonance is a silver slab resonance, which might also possess some multipolar resonance nature. The concave-shaped nanocuboids exhibited stronger induced plasmonic CD relative to the nanocuboids with straight facets.

Lifetime healthcare expenditures across socioeconomic groups.

BACKGROUND: A socioeconomic gradient affects healthcare expenditures and longevity in opposite directions as less affluent individuals have higher current healthcare expenditures but simultaneously enjoy shorter lives. Yet, it is unclear whether this cross-sectional healthcare expenditure gradient persists from a lifetime perspective. This paper analyzes lifetime healthcare expenditures across socioeconomic groups using detailed individual-level healthcare expenditure data for the entire Danish population. METHOD: Using full population healthcare expenditures from Danish registries, we estimate lifetime healthcare expenditures as age-specific mean healthcare expenditures times the probability of being alive at each age. Our data enables the estimation of lifetime healthcare expenditures by sex, socioeconomic status, and by various types of healthcare expenditure. RESULTS: Once we account for mortality differences and all types of healthcare expenditures, all socioeconomic groups spend an almost equal amount on healthcare throughout a lifetime. Lower socioeconomic groups incur the lowest lifetime hospital expenditures, whereas higher socioeconomic groups experience the highest lifetime expenditures on long-term care services. Our findings remain robust across various socioeconomic measures and alternative estimation methodologies. CONCLUSION: Improving the health status of lower socioeconomic groups to align with that of higher socioeconomic groups is costly but may ultimately reduce current healthcare expenditures. Enhanced health outcomes likely increase lifespan, leading to extended periods of healthcare consumption. However, since all socioeconomic groups tend to have similar lifetime healthcare expenditures, this prolonged consumption has limited impact on overall lifetime healthcare costs. Additionally, a significant benefit is the deferment of healthcare expenditures into the future. Overall, our results diminish concerns about socially inequitable utilization of healthcare resources while socioeconomic differences in health and longevity persist, even in a universal healthcare system.