The FLRT3-UNC5B checkpoint pathway inhibits T cell-based cancer immunotherapies.

Cancers exploit coinhibitory receptors on T cells to escape tumor immunity, and targeting such mechanisms has shown remarkable clinical benefit, but in a limited subset of patients. We hypothesized that cancer cells mimic noncanonical mechanisms of early development such as axon guidance pathways to evade T cell immunity. Using gain-of-function genetic screens, we profiled axon guidance proteins on human T cells and their cognate ligands and identified fibronectin leucine-rich transmembrane protein 3 (FLRT3) as a ligand that inhibits T cell activity. We demonstrated that FLRT3 inhibits T cells through UNC5B, an axon guidance receptor that is up-regulated on activated human T cells. FLRT3 expressed in human cancers favored tumor growth and inhibited CAR-T and BiTE + T cell killing and infiltration in humanized cancer models. An FLRT3 monoclonal antibody that blocked FLRT3-UNC5B interactions reversed these effects in an immune-dependent manner. This study supports the concept that axon guidance proteins mimic T cell checkpoints and can be targeted for cancer immunotherapy.

Advances in the differentiation of pluripotent stem cells into vascular cells.

Blood vessels constitute a closed pipe system distributed throughout the body, transporting blood from the heart to other organs and delivering metabolic waste products back to the lungs and kidneys. Changes in blood vessels are related to many disorders like stroke, myocardial infarction, aneurysm, and diabetes, which are important causes of death worldwide. Translational research for new approaches to disease modeling and effective treatment is needed due to the huge socio-economic burden on healthcare systems. Although mice or rats have been widely used, applying data from animal studies to human-specific vascular physiology and pathology is difficult. The rise of induced pluripotent stem cells (iPSCs) provides a reliable in vitro resource for disease modeling, regenerative medicine, and drug discovery because they carry all human genetic information and have the ability to directionally differentiate into any type of human cells. This review summarizes the latest progress from the establishment of iPSCs, the strategies for differentiating iPSCs into vascular cells, and the in vivo transplantation of these vascular derivatives. It also introduces the application of these technologies in disease modeling, drug screening, and regenerative medicine. Additionally, the application of high-tech tools, such as omics analysis and high-throughput sequencing, in this field is reviewed.

Neutralizing antibodies to block viral entry and for identification of entry inhibitors.

Neutralizing antibodies (NAbs) are naturally produced by our immune system to combat viral infections. Clinically, neutralizing antibodies with potent efficacy and high specificity have been extensively used to prevent and treat a wide variety of viral infections, including Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2), Human Immunodeficiency Virus (HIV), Dengue Virus (DENV) and Hepatitis B Virus (HBV). An overwhelmingly large subset of clinically effective NAbs operates by targeting viral envelope proteins to inhibit viral entry into the host cell. Binding of viral envelope protein to the host receptor is a critical rate limiting step triggering a cascade of downstream events, including endocytosis, membrane fusion and pore formation to allow viral entry. In recent years, improved structural knowledge on these processes have allowed researchers to also leverage NAbs as an indispensable tool in guiding discovery of novel antiviral entry inhibitors, providing drug candidates with high efficacy and pan-genus specificity. This review will summarize the latest progresses on the applications of NAbs as effective entry inhibitors and as important tools to develop antiviral therapeutics by high-throughput drug screenings, rational design of peptidic entry inhibitor mimicking NAbs and in silico computational modeling approaches.

Geography, phylogeny and host switch drive the coevolution of parasitic Gyrodactylus flatworms and their hosts.

BACKGROUND: Gyrodactylus is a lineage of monogenean flatworm ectoparasites exhibiting many features that make them a suitable model to study the host-parasite coevolutionary dynamics. Previous coevolutionary studies of this lineage mainly relied on low-power datasets (a small number of samples and a single molecular marker) and (now) outdated algorithms. METHODS: To investigate the coevolutionary relationship of gyrodactylids and their fish hosts in high resolution, we used complete mitogenomes (including two newly sequenced Gyrodactylus species), a large number of species in the single-gene dataset, and four different coevolutionary algorithms. RESULTS: The overall coevolutionary fit between the parasites and hosts was consistently significant. Multiple indicators confirmed that gyrodactylids are generally highly host-specific parasites, but several species could parasitize either multiple (more than 5) or phylogenetically distant fish hosts. The molecular dating results indicated that gyrodactylids tend to evolve towards high host specificity. Speciation by host switch was identified as a more important speciation mode than co-speciation. Assuming that the ancestral host belonged to Cypriniformes, we inferred four major host switch events to non-Cypriniformes hosts (mostly Salmoniformes), all of which occurred deep in the evolutionary history. Despite their relative rarity, these events had strong macroevolutionary consequences for gyrodactylid diversity. For example, in our dataset, 57.28% of all studied gyrodactylids parasitized only non-Cypriniformes hosts, which implies that the evolutionary history of more than half of all included lineages could be traced back to these major host switch events. The geographical co-occurrence of fishes and gyrodactylids determined the host use by these gyrodactylids, and geography accounted for most of the phylogenetic signal in host use. CONCLUSIONS: Our findings suggest that the coevolution of Gyrodactylus flatworms and their hosts is largely driven by geography, phylogeny, and host switches.

Independent activation of CREB3L2 by glucose fills a regulatory gap in mouse ß-cells by co-ordinating insulin biosynthesis with secretory granule formation.

OBJECTIVE: Although individual steps have been characterized, there is little understanding of the overall process whereby glucose co-ordinates the biosynthesis of insulin with its export out of the endoplasmic reticulum (ER) and incorporation into insulin secretory granules (ISGs). Here we investigate a role for the transcription factor CREB3L2 in this context. METHODS: MIN6 cells and mouse islets were analysed by immunoblotting after treatment with glucose, fatty acids, thapsigargin and various inhibitors. Knockdown of CREB3L2 was achieved using si or sh constructs by transfection, or viral delivery. In vivo metabolic phenotyping was conducted after deletion of CREB3L2 in ß-cells of adult mice using Ins1-CreER+. Islets were isolated for RNAseq and assays of glucose-stimulated insulin secretion (GSIS). Trafficking was monitored in islet monolayers using a GFP-tagged proinsulin construct that allows for synchronised release from the ER. RESULTS: With a Km ≈3.5 mM, glucose rapidly (T1/2 0.9 h) increased full length (FL) CREB3L2 followed by a slower rise (T1/2 2.5 h) in its transcriptionally-active cleavage product, P60 CREB3L2. Glucose stimulation repressed the ER stress marker, CHOP, and this was partially reverted by knockdown of CREB3L2. Activation of CREB3L2 by glucose was not due to ER stress, however, but a combination of O-GlcNAcylation, which impaired proteasomal degradation of FL-CREB3L2, and mTORC1 stimulation, which enhanced its conversion to P60. cAMP generation also activated CREB3L2, but independently of glucose. Deletion of CREB3L2 inhibited GSIS ex vivo and, following a high-fat diet (HFD), impaired glucose tolerance and insulin secretion in vivo. RNAseq revealed that CREB3L2 regulated genes controlling trafficking to-and-from the Golgi, as well as a broader cohort associated with ß-cell compensation during a HFD. Although post-Golgi trafficking appeared intact, knockdown of CREB3L2 impaired the generation of both nascent ISGs and proinsulin condensates in the Golgi, implying a defect in ER export of proinsulin and/or its processing in the Golgi. CONCLUSION: The stimulation of CREB3L2 by glucose defines a novel, rapid and direct mechanism for co-ordinating the synthesis, packaging and storage of insulin, thereby minimizing ER overload and optimizing ß-cell function under conditions of high secretory demand. Upregulation of CREB3L2 also potentially contributes to the benefits of GLP1 agonism and might in itself constitute a novel means of treating ß-cell failure.

Natural compounds as obesity pharmacotherapies.

Obesity has become a serious global public health problem, affecting over 988 million people worldwide. Nevertheless, current pharmacotherapies have proven inadequate. Natural compounds have garnered significant attention due to their potential antiobesity effects. Over the past three decades, ca. 50 natural compounds have been evaluated for the preventive and/or therapeutic effects on obesity in animals and humans. However, variations in the antiobesity efficacies among these natural compounds have been substantial, owing to differences in experimental designs, including variations in animal models, dosages, treatment durations, and administration methods. The feasibility of employing these natural compounds as pharmacotherapies for obesity remained uncertain. In this review, we systematically summarized the antiobesity efficacy and mechanisms of action of each natural compound in animal models. This comprehensive review furnishes valuable insights for the development of antiobesity medications based on natural compounds.

Clinical efficacy and safety of tigecycline based on therapeutic drug monitoring for carbapenem-resistant Gram-negative bacterium pneumonia in intensive care units.

BACKGROUND: We investigated the associations between the different doses of tigecycline, its efficacy and safety, and the role of tigecycline therapeutic drug monitoring for patients in the intensive care unit. METHODS: This study was a single-center cohort including patients infected with multidrug-resistant Acinetobacter baumannii (MDR-AB) and multidrug-resistant Klebsiella pneumoniae (MDR-KP) causing pulmonary infections. The steady-state plasma concentration after tigecycline administration was determined by High-Performance Liquid Chromatography (HPLC) in patients admitted to the ICU between October 2020 and December 2021. Multivariate analyses of tigecycline's clinical efficacy and safety were performed to control confounding factors. RESULTS: For this study, we included 45 patients and 45 blood samples to determine steady-state trough concentrations of tigecycline. All patients were divided into the High Dose (HD) and Standard Dose (SD) groups. The median trough concentration of tigecycline was 0.56 µg/mL in the HD group, which was higher than in the SD group (0,21 µg/mL), p = 0.000. There was no significant difference between the two groups of patients in terms of bacterial eradication rate, mortality rate, and clinical efficacy. Multiple regression analysis showed that the ICU days were correlated with mortality OR 1.030(1.005-1.056), p = 0.017. APACHE II was significantly associated with clinical efficacy OR 0.870(0.755-1.002), p = 0.045. The level of fibrinogen decline in the HD group was significantly higher than in the SD group (-3.05 ± 1.67 vs -1.75 ± 1.90), p = 0.038. We identified that age and tigecycline treatment duration influenced fibrinogen decline. CONCLUSIONS: Tigecycline plasma concentrations are significantly increased when using a high dose. However, the plasma concentration of tigecycline is not correlated with clinical efficacy and adverse reactions. Fibrinogen decline appears to be related to the patient's age and days of tigecycline. Large sample data are still needed to confirm the clinical guidance significance of tigecycline TDM.

LI-RADS category 3, 4, and M observations: a multiple parameters diagnostic model for hepatocellular carcinoma.

BACKGROUND: Hepatic lesions categorized as LR-3, LR-4, and LR-M are challenging to accurately assess and diagnose. PURPOSE: To combine potential clinical and/or magnetic resonance imaging (MRI) features for a more comprehensive hepatocellular carcinoma (HCC) versus non-HCC diagnosis for patients with LR-3, LR-4, and LR-M graded lesions. METHODS: Data were consecutively retrieved from 82 at-risk patients with LR-3 (n = 43), LR-4 (n = 20), and LR-M (n = 23) lesions. Significant findings for the differentiation of HCC and non-HCC, including MRI features and clinical factors, were identified with univariable and multivariable analyses. The variables for a prediction model were selected through stepwise use of Akaike's Information Criterion (AIC) to build multivariable logistic regression model. RESULTS: Serum alpha-fetoprotein (AFP) >16.2 ng/mL (odds ratio [OR] = 22.4; P = 0.006), septum (OR = 52.1; P = 0.011), and hepatobiliary phase (HBP) hypointensity (OR = 40.2; P = 0.001) were confirmed as independent predictors of HCC. When combining the three predictors and mild-moderate T2 hyperintensity, the model (AIC = 50.91) showed good accuracy with a C-index of 0.948. CONCLUSION: In at-risk patients with LR-3, LR-4, or LR-M lesions, integrating AFP, septum, HBP hypointensity, and mild-moderate T2 hyperintensity achieved high diagnostic performance for the diagnosis of HCC.

Delta-radiomics models based on multi-phase contrast-enhanced magnetic resonance imaging can preoperatively predict glypican-3-positive hepatocellular carcinoma.

Objectives: The aim of this study is to investigate the value of multi-phase contrast-enhanced magnetic resonance imaging (CE-MRI) based on the delta radiomics model for identifying glypican-3 (GPC3)-positive hepatocellular carcinoma (HCC). Methods: One hundred and twenty-six patients with pathologically confirmed HCC (training cohort: n = 88 and validation cohort: n = 38) were retrospectively recruited. Basic information was obtained from medical records. Preoperative multi-phase CE-MRI images were reviewed, and the 3D volumes of interest (VOIs) of the whole tumor were delineated on non-contrast T1-weighted imaging (T1), arterial phase (AP), portal venous phase (PVP), delayed phase (DP), and hepatobiliary phase (HBP). One hundred and seven original radiomics features were extracted from each phase, and delta-radiomics features were calculated. After a two-step feature selection strategy, radiomics models were built using two classification algorithms. A nomogram was constructed by combining the best radiomics model and clinical risk factors. Results: Serum alpha-fetoprotein (AFP) (p = 0.013) was significantly related to GPC3-positive HCC. The optimal radiomics model is composed of eight delta-radiomics features with the AUC of 0.805 and 0.857 in the training and validation cohorts, respectively. The nomogram integrated the radiomics score, and AFP performed excellently (training cohort: AUC = 0.844 and validation cohort: AUC = 0.862). The calibration curve showed good agreement between the nomogram-predicted probabilities and GPC3 actual expression in both training and validation cohorts. Decision curve analysis further demonstrates the clinical practicality of the nomogram. Conclusion: Multi-phase CE-MRI based on the delta-radiomics model can non-invasively predict GPC3-positive HCC and can be a useful method for individualized diagnosis and treatment.

The evaluation of cognitive-behavioral therapy-based intervention on type 2 diabetes patients with comorbid metabolic syndrome: a randomized controlled trial.

BACKGROUND: Cognitive behavior therapy (CBT) has been applied in intervention research in diabetes patients with satisfying results. However, there was no research on type 2 diabetes (T2DM) patients with comorbidities. This study aimed to investigate the effectiveness of CBT on psychological variables, behavior variables, quality of life, sleep quality, and physical variables among adult T2DM patients with comorbid metabolic syndrome (MS). METHODS: 281 patients aged 18-75 years were recruited from Ningbo First Hospital in China from October 2021 to March 2022. Patients were randomized to the intervention group (IG, N = 148) or control group (CG, N = 133). Patients in the IG received 12 CBT-based sessions during a six-month intervention time. Patients in the CG received the usual care only. Univariate General Linear Model was used to analyze the effect of CBT-based interventions. The analysis was conducted by SPSS Version 28. RESULTS: Results indicated that CBT-based intervention was superior in the following aspects: relieving depression symptoms: IG (4.11 ± 4.35 vs. 1.99 ± 2.12), CG (3.40 ± 3.26 vs. 2.32 ± 1.88), interaction effect (F = 4.074, P = 0.044); enhancing diabetes self-care behaviors: IG (26.79 ± 12.18 vs. 37.49 ± 10.83), CG (25.82 ± 13.71 vs. 31.96 ± 11.72), interaction effect (F = 5.242, P = 0.022); promoting the efficacy of CBT: IG (47.45 ± 6.83 vs. 50.76 ± 4.98), CG (46.74 ± 6.94 vs. 47.87 ± 5.11), interaction effect (F = 5.198, P = 0.023); improving subjective sleep quality: IG (0.93 ± 0.68 vs. 0.69 ± 0.63), CG (1.03 ± 0.72 vs. 1.01 ± 0.68), interaction effect (F = 3.927, P = 0.048). CONCLUSIONS: The CBT-based intervention was beneficial in improving depression symptoms, diabetes self-care behaviors, the efficacy of CBT, and sleep quality in T2DM patients with comorbid MS. The downtrend of body mass index, systolic blood pressure, diastolic pressure, and glycated hemoglobin was larger in the intervention group but not to a significant level. TRIAL REGISTRATION: This study has been prospectively registered at Australia New Zealand Clinical Trials Registry (Registration ID: ACTRN12621001348842 website: https://www.anzctr.org.au/trial/MyTrial.aspx ).

Potential for NPY receptor-related therapies for polycystic ovary syndrome: an updated review.

Polycystic ovary syndrome (PCOS) is a complex endocrine disease that can cause female infertility and bring economic burden to families and to society. The clinical and/or biochemical manifestations include hyperandrogenism, persistent anovulation, and polycystic ovarian changes, often accompanied by insulin resistance and obesity. Although its pathogenesis is unclear, PCOS involves the abnormal regulation of the hypothalamic-pituitary-ovarian axis and the abnormal activation of GnRH neurons. Neuropeptide Y (NPY) is widely distributed in the arcuate nucleus of the hypothalamus and functions as the physiological integrator of two neuroendocrine systems, one governing feeding and the other controlling reproduction. In recent years, an increasing number of studies have focused on the improvement of the reproductive and metabolic status of PCOS through the therapeutic application of NPY and its receptors. In this review, we summarize the central and peripheral regulation of NPY and its receptors in the development of PCOS and discuss the potential for NPY receptor-related therapies for PCOS.

Critical role of lateral habenula circuits in the control of stress-induced palatable food consumption.

Chronic stress fuels the consumption of palatable food and can enhance obesity development. While stress- and feeding-controlling pathways have been identified, how stress-induced feeding is orchestrated remains unknown. Here, we identify lateral habenula (LHb) Npy1r-expressing neurons as the critical node for promoting hedonic feeding under stress, since lack of Npy1r in these neurons alleviates the obesifying effects caused by combined stress and high fat feeding (HFDS) in mice. Mechanistically, this is due to a circuit originating from central amygdala NPY neurons, with the upregulation of NPY induced by HFDS initiating a dual inhibitory effect via Npy1r signaling onto LHb and lateral hypothalamus neurons, thereby reducing the homeostatic satiety effect through action on the downstream ventral tegmental area. Together, these results identify LHb-Npy1r neurons as a critical node to adapt the response to chronic stress by driving palatable food intake in an attempt to overcome the negative valence of stress.

Monitoring AKT activity and targeting in live tissue and disease contexts using a real-time Akt-FRET biosensor mouse.

Aberrant AKT activation occurs in a number of cancers, metabolic syndrome, and immune disorders, making it an important target for the treatment of many diseases. To monitor spatial and temporal AKT activity in a live setting, we generated an Akt-FRET biosensor mouse that allows longitudinal assessment of AKT activity using intravital imaging in conjunction with image stabilization and optical window technology. We demonstrate the sensitivity of the Akt-FRET biosensor mouse using various cancer models and verify its suitability to monitor response to drug targeting in spheroid and organotypic models. We also show that the dynamics of AKT activation can be monitored in real time in diverse tissues, including in individual islets of the pancreas, in the brown and white adipose tissue, and in the skeletal muscle. Thus, the Akt-FRET biosensor mouse provides an important tool to study AKT dynamics in live tissue contexts and has broad preclinical applications.

Deficiency of neuropeptide Y attenuates neointima formation after vascular injury in mice.

BACKGROUND: Restenosis after percutaneous coronary intervention (PCI) limits therapeutic revascularization. Neuropeptide Y (NPY), co-stored and co-released with the sympathetic nervous system, is involved in this process, but its exact role and underlying mechanisms remain to be fully understood. This study aimed to investigate the role of NPY in neointima formation after vascular injury. METHODS: Using the left carotid arteries of wild-type (WT, NPY-intact) and NPY-deficient (NPY-/-) mice, ferric chloride-mediated carotid artery injury induced neointima formation. Three weeks after injury, the left injured carotid artery and contralateral uninjured carotid artery were collected for histological analysis and immunohistochemical staining. RT-qPCR was used to detect the mRNA expression of several key inflammatory markers and cell adhesion molecules in vascular samples. Raw264.7 cells were treated with NPY, lipopolysaccharide (LPS), and lipopolysaccharide-free, respectively, and RT-qPCR was used to detect the expression of these inflammatory mediators. RESULTS: Compared with WT mice, NPY-/- mice had significantly reduced neointimal formation three weeks after injury. Mechanistically, immunohistochemical analysis showed there were fewer macrophages and more vascular smooth muscle cells in the neointima of NPY-/- mice. Moreover, the mRNA expression of key inflammatory markers such as interleukin-6 (IL-6), transforming growth factor-ß1 (TGF-ß1), and intercellular adhesion molecule-1 (ICAM-1) was significantly lower in the injured carotid arteries of NPY-/- mice, compared to that in the injured carotid arteries of WT mice. In RAW264.7 macrophages, NPY significantly promoted TGF-ß1 mRNA expression under unactivated but not LPS-stimulated condition. CONCLUSIONS: Deletion of NPY attenuated neointima formation after artery injury, at least partly, through reducing the local inflammatory response, suggesting that NPY pathway may provide new insights into the mechanism of restenosis.

Identification of indocyanine green as a STT3B inhibitor against mushroom α-amanitin cytotoxicity.

The "death cap", Amanita phalloides, is the world's most poisonous mushroom, responsible for 90% of mushroom-related fatalities. The most fatal component of the death cap is α-amanitin. Despite its lethal effect, the exact mechanisms of how α-amanitin poisons humans remain unclear, leading to no specific antidote available for treatment. Here we show that STT3B is required for α-amanitin toxicity and its inhibitor, indocyanine green (ICG), can be used as a specific antidote. By combining a genome-wide CRISPR screen with an in silico drug screening and in vivo functional validation, we discover that N-glycan biosynthesis pathway and its key component, STT3B, play a crucial role in α-amanitin toxicity and that ICG is a STT3B inhibitor. Furthermore, we demonstrate that ICG is effective in blocking the toxic effect of α-amanitin in cells, liver organoids, and male mice, resulting in an overall increase in animal survival. Together, by combining a genome-wide CRISPR screen for α-amanitin toxicity with an in silico drug screen and functional validation in vivo, our study highlights ICG as a STT3B inhibitor against the mushroom toxin.