Neuronal activity-induced, equilibrative nucleoside transporter-dependent, somatodendritic adenosine release revealed by a GRAB sensor.

The purinergic signaling molecule adenosine (Ado) modulates many physiological and pathological functions in the brain. However, the exact source of extracellular Ado remains controversial. Here, utilizing a newly optimized genetically encoded GPCR-Activation-Based Ado fluorescent sensor (GRABAdo), we discovered that the neuronal activity-induced extracellular Ado elevation is due to direct Ado release from somatodendritic compartments of neurons, rather than from the axonal terminals, in the hippocampus. Pharmacological and genetic manipulations reveal that the Ado release depends on equilibrative nucleoside transporters but not the conventional vesicular release mechanisms. Compared with the fast-vesicular glutamate release, the Ado release is slow (~40 s) and requires calcium influx through L-type calcium channels. Thus, this study reveals an activity-dependent second-to-minute local Ado release from the somatodendritic compartments of neurons, potentially serving modulatory functions as a retrograde signal.

MR Uniformity Ratio Estimates to Evaluate Ventricular Mechanical Dyssynchrony and Prognosis After ST-Segment Elevation Myocardial Infarction.

BACKGROUND: The impact of left ventricular mechanical dyssynchrony (LVMD) on the long-term prognosis of ST-segment elevation myocardial infarction (STEMI) is unclear. HYPOTHESIS: MR uniformity ratio estimates (URE) can detect LVMD and assess STEMI prognosis. STUDY TYPE: Retrospective analysis of a prospective multicenter registry (EARLY-MYO trial, NCT03768453). POPULATION: Overall, 450 patients (50 females) with first-time STEMI were analyzed, as well as 40 participants without cardiovascular disease as controls. FIELD STRENGTH/SEQUENCE: 3.0-T, balanced steady-state free precession cine and late gadolinium enhancement imaging. ASSESSMENT: MRI data were acquired within 1 week of symptom onset. Major adverse cardiovascular events (MACEs), including cardiovascular death, nonfatal re-infarction, hospitalization for heart failure, and stroke, were the primary clinical outcomes. LVMD was represented by circumferential URE (CURE) and radial URE (RURE) calculated using strain measurements. The patients were grouped according to clinical outcomes or URE values. Patients' clinical characteristics and MR indicators were compared. STATISTICAL TESTS: The Student's t-test, Mann-Whitney U test, chi-square test, Fisher's exact test, receiver operating characteristic curve analysis with area under the curve, Kaplan-Meier analysis, Cox regression, logistic regression, intraclass correlation coefficient, c-index, and integrated discrimination improvement were used. P < 0.05 was considered statistically significant. RESULTS: CURE and RURE were significantly lower in patients with STEMI than in controls. The median follow-up was 60.5 months. Patients with both lower CURE and RURE values experienced a significantly higher incidence of MACEs by 3.525-fold. Both CURE and RURE were independent risk factors for MACEs. The addition of UREs improved diagnostic efficacy and risk stratification based on infarct size and left ventricular ejection fraction (LVEF). The indicators associated with LVMD included male sex, serum biomarkers (peak creatine phosphokinase and cardiac troponin I), infarct size, and LVEF. DATA CONCLUSION: CURE and RURE may be useful to evaluate long-term prognosis after STEMI. EVIDENCE LEVEL: 4 TECHNICAL EFFICACY: Stage 2.

Role of the hydrocarbon molecular structure in CNT growth on Fe-Al catalysts.

Upgrading plastic wastes into high-value products via the thermochemical process is one of the most attractive topics. Although carbon nanotubes (CNTs) have been successfully synthesized from plastic pyrolysis gas over Fe-, Co-, or Ni-based catalysts, a deep discussion about the reaction mechanism was seldom mentioned in the literature. Herein, this work was intended to study the growth mechanism of CNTs from hydrocarbons on Fe-Al2O3 catalysts. C5-C7 hydrocarbons were used to synthesize CNTs in a high-temperature fixed-bed reactor, and the carbon products and cracked gas were analyzed in detail. The CNT yield was in the order of cyclohexane, cyclohexene > n-hexane > n-heptane > n-pentane, 1-hexene. It was proposed that CNT growth on Fe-Al2O3 catalysts was mainly determined by the yield and structure of six-membered cyclic species, which was tailored by the carbon chain length, C-C/CC bonds, and linear/cyclic structures of C5-C7 hydrocarbons. Compared with n-hexane, the six-membered rings of cyclohexane and cyclohexene promoted six-membered cyclic species formation, increasing CNT and benzene yields; the seven-membered carbon chain of n-heptane promoted methyl-six-membered cyclic species formation, decreasing CNT and benzene yields while increasing the toluene yield; the five-membered carbon chain of n-pentane and the CC bond of 1-hexene inhibited six-membered cyclic species formation, decreasing CNT and benzene yields. This work revealed the structure-activity relationship between C5-C7 hydrocarbons and CNT growth, which may direct the process design and optimization of CNT synthesis from plastic pyrolysis gas.

A Complete Endocannabinoid Signaling System Modulates Synaptic Transmission between Human Induced Pluripotent Stem Cell-Derived Neurons.

The endocannabinoid system (ECS) modulates synaptic function to regulate many aspects of neurophysiology. It adapts to environmental changes and is affected by disease. Thus, the ECS presents an important target for therapeutic development. Despite recent interest in cannabinoid-based treatments, few preclinical studies are conducted in human systems. Human induced pluripotent stem cells (hiPSCs) provide one possible solution to this issue. However, it is not known if these cells have a fully functional ECS. Here, we show that hiPSC-derived neuron/astrocyte cultures exhibit a complete ECS. Using Ca2+ imaging and a genetically encoded endocannabinoid sensor, we demonstrate that they not only respond to exogenously applied cannabinoids but also produce and metabolize endocannabinoids. Synaptically driven [Ca2+]i spiking activity was inhibited (EC50 = 48 ± 13 nM) by the efficacious agonist [R(+)-[2,3-dihydro-5-methyl-3-[(morpholinyl)methyl]pyrolol [1,2,3-de]-1,4-benzoxazin-yl]-(1-naphthalenyl)methanone mesylate] (Win 55,212-2) and by the endogenous ligand 2-arachidonoyl glycerol (2-AG; EC50 = 2.0 ± 0.6 µm). The effects of Win 55212-2 were blocked by a CB1 receptor-selective antagonist. Δ9-Tetrahydrocannabinol acted as a partial agonist, maximally inhibiting synaptic activity by 47 ± 14% (EC50 = 1.4 ± 1.9 µm). Carbachol stimulated 2-AG production in a manner that was independent of Ca2+ and blocked by selective inhibition of diacylglycerol lipase. 2-AG returned to basal levels via a process mediated by monoacylglycerol lipase as indicated by slowed recovery in cultures treated with 4-[Bis(1,3-benzodioxol-5-yl)hydroxymethyl]-1-piperidinecarboxylic acid 4-nitrophenyl ester (JZL 184). Win 55,212-2 markedly desensitized CB1 receptor function following a 1-day exposure, whereas desensitization was incomplete following 7-day treatment with JZL 184. This human cell culture model is well suited for functional analysis of the ECS and as a platform for drug development. SIGNIFICANCE STATEMENT: Despite known differences between the human response to cannabinoids and that of other species, an in vitro human model demonstrating a fully functional endocannabinoid system has not been described. Human induced pluripotent stem cells (hiPSCs) can be obtained from skin samples and then reprogrammed into neurons for use in basic research and drug screening. Here, we show that hiPSC-derived neuronal cultures exhibit a complete endocannabinoid system suitable for mechanistic studies and drug discovery.

Bioinformatic analysis of structures and encoding genes of Escherichia coli surface polysaccharides sheds light on the heterologous biosynthesis of glycans.

BACKGROUND: Surface polysaccharides (SPs), such as lipopolysaccharide (O antigen) and capsular polysaccharide (K antigen), play a key role in the pathogenicity of Escherichia coli (E. coli). Gene cluster for polysaccharide antigen biosynthesis encodes various glycosyltransferases (GTs), which drive the process of SP synthesis and determine the serotype. RESULTS: In this study, a total of 7,741 E. coli genomic sequences were chosen for systemic data mining. The monosaccharides in both O and K antigens were dominated by D-hexopyranose, and the SPs in 70-80% of the strains consisted of only the five most common hexoses (or some of them). The linkages between the two monosaccharides were mostly α-1,3 (23.15%) and ß-1,3 (20.49%) bonds. Uridine diphosphate activated more than 50% of monosaccharides for glycosyltransferase reactions. These results suggest that the most common pathways could be integrated into chassis cells to promote glycan biosynthesis. We constructed a database (EcoSP, http://ecosp.dmicrobe.cn/ ) for browse this information, such as monosaccharide synthesis pathways. It can also be used for serotype analysis and GT annotation of known or novel E. coli sequences, thus facilitating the diagnosis and typing. CONCLUSIONS: Summarizing and analyzing the properties of these polysaccharide antigens and GTs are of great significance for designing glycan-based vaccines and the synthetic glycobiology.

Development and evaluation of serological screening based on one dried plasma spot for HIV, syphilis, and HCV.

BACKGROUND: In the effort to prevent and control HIV/AIDS, China has established a national sentinel surveillance system. However, some sentinel sites face limitations in environmental resources and accessibility, prompting the exploration of alternative sample strategies. Dried plasma spots (DPS) samples are viewed as promising alternatives to traditional plasma samples due to their advantages, including sample stability, easy storage, and convenient transport. This study aims to develop a method for screening HIV, Treponema pallidum (TP), and Hepatitis C Virus (HCV) using DPS samples and assess their performance. METHODS: Based on existing commercial assay kits, a detection method was established through the optimization of experimental parameters, including the amount of plasma on filter paper, the volume of elution solution applied to dried plasma spots, the size of dried plasma spots, elution solution volume, elution solution components, elution temperature, and elution time. A series of laboratory evaluation panels were constructed for laboratory assessments, including the laboratory basic panel, laboratory interference panel, and laboratory precision panel. Additionally, clinical samples were used for evaluation. RESULTS: Optimal conditions for DPS sample extraction were: plasma volume, 100 µL; DPS size, whole spot; eluent volume, 500 µL; eluent, PBS with 1‰ Tween20; elution time, 2 h; elution temperature, room temperature. A total of 619 paired plasma/DPS samples were tested by both methods. The DPS-based ELISA method exhibited 100% sensitivity/specificity for HIV, 98.6%/100% for TP, and 99.6%/100% for HCV. Kappa values between the plasma samples and DPS samples were 100% for HIV, 99% for TP, and 100% for HCV. The DPS-based ELISA method failed to detect 1 HCV mono-infected sample and TP in 1 HIV/HCV/TP co-infected sample. For the HIV/HCV/TP co-infected sample, the S/CO in the plasma sample was 2.143 and in the DPS sample was 0.5. For HCV, the S/CO (sample OD/cut-off) was 3.049 in the plasma sample and 0.878 in the DPS sample. CONCLUSIONS: A single DPS, following one-time standardized processing, can be used to detect HIV, HCV, and TP. Researching and establishing laboratory testing methods better suited for China's sentinel surveillance have significant practical applications in improving HIV testing in resource-constrained environments.

Superrepellency of underwater hierarchical structures on Salvinia leaf.

Biomimetic superhydrophobic surfaces display many excellent underwater functionalities, which attribute to the slippery air mattress trapped in the structures on the surface. However, the air mattress is easy to collapse due to various disturbances, leading to the fully wetted Wenzel state, while the water filling the microstructures is difficult to be repelled to completely recover the air mattress even on superhydrophobic surfaces like lotus leaves. Beyond superhydrophobicity, here we find that the floating fern, Salvinia molesta, has the superrepellent capability to efficiently replace the water in the microstructures with air and robustly recover the continuous air mattress. The hierarchical structures on the leaf surface are demonstrated to be crucial to the recovery. The interconnected wedge-shaped grooves between epidermal cells are key to the spontaneous spreading of air over the entire leaf governed by a gas wicking effect to form a thin air film, which provides a base for the later growth of the air mattress in thickness synchronously along the hairy structures. Inspired by nature, biomimetic artificial Salvinia surfaces are fabricated using 3D printing technology, which successfully achieves a complete recovery of a continuous air mattress to exactly imitate the superrepellent capability of Salvinia leaves. This finding will benefit the design principles of water-repellent materials and expand their underwater applications, especially in extreme environments.

Peak early diastolic strain rate improves prediction of adverse cardiovascular outcomes in patients with ST-elevation myocardial infarction.

BACKGROUND: The prognostic role of diastolic dysfunction measured by the circumferential peak early diastolic strain rate (PEDSR) on ST-elevation myocardial infarction (STEMI) is not completely established. OBJECTIVES: We aimed to investigate the prognostic value of diastolic function by measuring PEDSR within 1 week after STEMI. METHODS: The cardiac magnetic resonance (CMR) pictures of 420 subjects from a clinical registry study (NCT03768453) were analyzed and the composite major adverse cardiac events (MACEs) were followed up. RESULTS: The PEDSR of patients was significantly lower compared with that of control subjects (P < 0.001). Within the median follow-up period of 52 months, PEDSR of patients who experienced MACEs deceased more significantly than that of patients without MACEs (P < 0.001). After adjusting with clinical or CMR indexes, per 0.1/s reduction of PEDSR increased the risks of MACEs to 1.402 or 1.376 fold and the risk of left ventricular (LV) remodeling to 1.503 or 1.369 fold. When PEDSR divided by best cutoff point, significantly higher risk of MACEs (P < 0.001) and more remarkable LV remodeling (P < 0.001) occurred in patients with PEDSR ≤ 0.485/s. Moreover, when adding the PEDSR to the conventional prognostic factors such as LV ejection fraction and infarction size, better prognostic risk classification models were created. Finally, aging, tobacco use, remarkable LV remodeling, and a low LV ejection fraction were factors related with the reduction of PEDSR. CONCLUSIONS: Diastolic dysfunction has an important prognostic effect on patients with STEMI. Measurement of the PEDSR in the acute phase could serve as an effective index to predict the long-term risk of MACEs and cardiac remodeling.

Progress of newborn screening in China. / 中国新生儿筛查进展.

Newborn screening (NBS) plays a significant role in reducing the risk of birth defects. NBS in China began in the early 1980s. Under the protection of laws and regulations and the leadership of the national health administration, approved screening centers in public hospitals took the responsibility for publicity, screening, diagnosis, treatment, follow-up and management of birth defects. As of 2022, 31 provinces (autonomous regions and municipalities directly under the central government) have carried out NBS for phenylketonuria, congenital hypothyroidism, and hearing loss, 23 provinces have carried out screening for glucose-6-phosphate dehydrogenase (with a screening rate of 89.24%), and 24 provinces have carried out screening for congenital adrenal cortical hyperplasia (91.45% screening rate). Over the past four decades, screening techniques have evolved from bacterial inhibition, fluorescence analysis, and tandem mass spectrometry for the detection of biochemical markers to genetic testing, which has greatly contributed to the expansion of the types of diseases screened for. The combined use of metabolomics and genomics is currently being explored. Effective management and rigorous quality control of NBS are prerequisites for improving the quality and ensuring the accuracy of screening. The Quality Management System for Newborn Screening System Network (QMS-NBS), established by the National Center for Clinical Laboratories, covers all screening centers and related blood collection agencies. The operation of the QMS-NBS allows the quality and performance of screening to be transparent and measurable, ensuring the quality and efficiency of screening. This article provides an overview of the history of NBS, especially the evolution of policies for the NBS in China, the construction of screening institutions, the number of newborns screened, the incidence rates of screened diseases, the changes in screening technology, the expansion of new diseases screened for, and the quality control of NBS. Overall, the progress in NBS in China has not only benefited from the development and standardization at the technological level, but also benefited from the construction of policies, regulations and ethics.

CRISPR-Based Genome Editing and Its Applications in Woody Plants.

CRISPR/Cas-based genome editing technology provides straightforward, proficient, and multifunctional ways for the site-directed modification of organism genomes and genes. The application of CRISPR-based technology in plants has a vast potential value in gene function research, germplasm innovation, and genetic improvement. The complexity of woody plants genome may pose significant challenges in the application and expansion of various new editing techniques, such as Cas9, 12, 13, and 14 effectors, base editing, particularly for timberland species with a long life span, huge genome, and ploidy. Therefore, many novel optimisms have been drawn to molecular breeding research based on woody plants. This review summarizes the recent development of CRISPR/Cas applications for essential traits, including wood properties, flowering, biological stress, abiotic stress, growth, and development in woody plants. We outlined the current problems and future development trends of this technology in germplasm and the improvement of products in woody plants.

Ultra-brilliant GeV betatronlike radiation from energetic electrons oscillating in frequency-downshifted laser pulses.

Electrons can be accelerated to GeV energies with high collimation via laser wakefield acceleration in the bubble regime and emit bright betatron radiation in a table-top size. However, the radiation brightness is usually limited to the third-generation synchrotron radiation facilities operating at similar photon energies. Using a two-stage plasma configuration, we propose a novel scheme for generating betatronlike radiation with an extremely high brilliance. In this scheme, the relativistic electrons inside the bubble injected from the first stage can catch up with the frequency-downshifted laser pulse formed in the second stage. The laser red shift originates from the phase modulation, together with the group velocity dispersion, which enables more energy to be transfered from the laser pulse to γ-photons, giving rise to ultra-brilliant betatronlike radiation. Multi-dimensional particle-in-cell simulations indicate that the radiated γ-photons have the cut-off energy of GeV and a peak brilliance of 1026 photons s-1 mm-2 mrad-2 per 0.1%BW at 1 MeV, which may have diverse applications in various fields.

Using Reflectometric Interference Spectroscopy to Real-Time Monitor Amphiphile-Induced Orientational Responses of Liquid-Crystal-Loaded Silica Colloidal Crystal Films.

An approach to optical transduction and amplification of amphiphile-triggered orientational responses of liquid crystals (LCs) based on the interference effect was developed. The sensitive substrate was obtained by lading 4'-pentyl-4-cyanobiphenyl (5CB) into three-dimensionally ordered silica colloidal crystal (SCC) films. Changes in the optical thickness (ΔOT) of the substrates, which are inverted by their Fabry-Perot fringes, depend on the changes of the refractive index caused by the differences in the orientations of LCs. The orientation changes of LCs loading into SCC films have the effect of amplifying signals. These are based on the interactions between surfactants (alkyl trimethylammonium halides (CnTABs, n = 8, 10, 12, 14, and 16) and sodium lauryl sulfonate (SLS)) and LCs, which induce a particular orientation of the LCs molecules. In this flowing system, the reversibility of the signal response for the adsorption of amphiphile was related to the length of the surfactant chain and its critical micelle concentration (CMC). A new method capable of real-time sensing adsorbate-triggered anchoring transitions based on LC-infiltrated SCC films was accomplished. These results provide basics and principles for online, label-free, and real-time analysis of molecules and their interactions in a flowing environment based on the interference effect.

Interference Effect of Silica Colloidal Crystal Films and Their Applications to Biosensing.

With the aim to develop better and more reliable interference effective substrates, silica colloidal crystal films with different sphere diameters and film thicknesses were successfully made by an improved vertical deposition method and a systematic investigation of their reflectometric interference spectroscopy (RIfS) properties are presented in this work. The influence of silica sphere diameter and film thickness on the RIfS signals was studied. The results showed that the film thickness is the key factor of RIfS signals. An RIfS system was set up by using a silica colloidal crystal film as an interference effective substrate. The influence of film thickness on the response to refractive index changes of the proposed system was also investigated. When the influence of film thickness on RIfS signals and refractive index response we considered together, silica colloidal crystal films with a thickness between 4 and 6 µm were chosen for sensor construction. Monitoring the digestive process of gelatin with trypsin was also demonstrated by combining gelatin-modified silica colloidal crystal films with RIfS. The system showed excellent sensitivity with a wide linear range and could achieve real-time measurement of each process. It has been proved that this is a promising method to construct biosensors using silica colloidal crystal films as interference-sensitive substrates.

Ordered Silica Nanosphere Templates: Large-Area Assembly and Characterization.

Ordered silica nanosphere templates, which are usually known as colloidal crystals, are most widely used to prepare ordered porous nanostructure materials among the templates to fabricate nanostructure materials. We present here a method for the simultaneous assembly of multiple ordered silica nanosphere templates with same quality, in which a glass trough together with a stand was used as the experimental cell. Two different diameters of silica colloidal particles were selected for our experiments, namely ∼190 nm and ∼280 nm. The growth process, thickness and optical properties of films of silica nanospheres on substrates were studied. The particle sedimentation and solvent evaporation both play a role in determining particle volume fractions. In addition, the standard deviation of the diameter of the particles affects the optical properties of the films along the growth direction. There was almost no difference observed in our measurements of the film thickness and optical properties for both the same regions of different films and different regions of the same film along the direction perpendicular to the growth direction. The elucidation of the growth process and characterization of the film properties achieved in this study could help us to obtain better quality templates. This is the first systematic study of the evolution of the thickness and optical properties of ordered silica nanosphere films formed by a simultaneous assembly process.

[Clinical and laboratory characteristics of juvenile myelomonocytic leukemia].

OBJECTIVE: To study the clinical and laboratory characteristics of juvenile myelomonocytic leukemia (JMML). METHODS: The clinical characteristics and laboratory results were retrospectively analyzed in 10 children with newly diagnosed JMML. They were compared with those of 28 children with myelodysplastic syndrome (MDS) and 44 children with chronic myeloid leukemia (CML). RESULTS: Compared with the children with CML or MDS, the children with JMML had significantly higher rates of skin rashes, ecchymosis, and lymphadenectasis, a significantly lower serum cholinesterase (ChE) level, and a significantly higher fetal hemoglobin level (P<0.05). The white blood cell count of children with JMML was significantly higher than that of children with MDS, but significantly lower than that of children with CML (P<0.05). In addition, the myeloid/erythroid ratio and rate of dyshaematopoiesis were significantly lower in children with JMML than those in children with CML or MDS. The children with JMML had a significantly higher expression of mature monocyte marker CD14 than those with CML or MDS (P<0.05). The levels of myeloid markers CD33, CD11b, CD13, and CD15 in children with JMML were significantly higher than those in children with MDS, but significantly lower than those in children with CML (P<0.05). The levels of CD2 and CD7 in children with JMML were higher than those in children with CML, but lower than those in children with MDS (P<0.05). CONCLUSIONS: Skin rashes, ecchymosis, lymphadenectasis, and ChE reduction are more common in children with JMML than in those with CML or MDS, while dyshaematopoiesis is less common. In addition, CD14 level increases significantly in children with JMML.

Disseminated Nonleukemic Myeloid Sarcoma of the Spleen With Involvement of the Liver in an Infant.

Nonleukemic myeloid sarcoma (MS) is a rare tumor that can occur in several locations without myeloid leukemia. We reported a first case of nonleukemic MS of the spleen involving the liver in a 5-month-old boy presenting with hematochezia, petechial hemorrhage, fever, and hepatosplenomegaly. Bone marrow trephine biopsy and immunophenotypic flow cytometry revealed no evidence of myeloid leukemia. The patient underwent liver biopsy and splenectomy. Clinicopathology and immunohistochemistry suggested a disseminated nonleukemic MS. The patient died of respiratory failure on the seventh postoperative day. Early diagnosis of a disseminated nonleukemic MS may be quite important for patient survival and it should be considered one of the differential diagnoses of hepatosplenomegaly with atypical clinical features.

Preparation methods, structural characteristics, and biological activity of polysaccharides from Platycodon grandiflorus.

Platycodon grandiflorus (P. grandiflorus), a traditional Chinese medicinal herb used for both medicine and food, has a long history of treating respiratory infections, bronchitis, pneumonia, and other lung-related diseases. The therapeutic effects of P. grandiflorus are attributed to its chemical components, including polysaccharides. Among these components, Platycodon grandiflorus polysaccharides (PGP) are recognized as one of the most important and abundant active ingredients, exhibiting various biological activities such as prebiotic, antioxidant, antiviral, anticancer, antiangiogenic, and immune regulatory properties. Incorporating the principles of traditional Chinese medicine, carrier concepts, and modern targeted drug delivery technologies, PGP can influence the target sites and therapeutic effects of other drugs while also serving as a drug carrier for targeted and precise treatments. Therefore, it is essential to provide a comprehensive review of the extraction, separation, purification, physicochemical properties, and biological activities of PGP. In the future, by integrating new concepts, technologies, and processes, further references and guidance can be provided for the comprehensive development of PGP. This will contribute to the advancement of P. grandiflorus in various fields such as pharmaceuticals, health products, and food.

Shared genetic architecture highlights the bidirectional association between major depressive disorder and fracture risk.

Background: There is limited evidence suggesting that osteoporosis might exacerbate depressive symptoms, while more studies demonstrate that depression negatively affects bone density and increases fracture risk. Aims: To explore the relationship between major depressive disorder (MDD) and fracture risk. Methods: We conducted a nested case-control analysis (32 670 patients with fracture and 397 017 individuals without fracture) and a matched cohort analysis (16 496 patients with MDD and 435 492 individuals without MDD) in the same prospective UK Biobank data set. Further, we investigated the shared genetic architecture between MDD and fracture with linkage disequilibrium score regression and the MiXeR statistical tools. We used the conditional/conjunctional false discovery rate approach to identify the specific shared loci. We calculated the weighted genetic risk score for individuals in the UK Biobank and logistic regression was used to confirm the association observed in the prospective study. Results: We found that MDD was associated with a 14% increase in fracture risk (hazard ratio (HR) 1.14, 95% CI 1.14 to 1.15, p<0.001) in the nested case-control analysis, while fracture was associated with a 72% increase in MDD risk (HR 1.72, 95% CI 1.64 to 1.79, p<0.001) in the matched cohort analysis, suggesting a longitudinal and bidirectional relationship. Further, genetic summary data suggested a genetic overlap between MDD and fracture. Specifically, we identified four shared genomic loci, with the top signal (rs7554101) near SGIP1. The protein encoded by SGIP1 is involved in cannabinoid receptor type 1 signalling. We found that genetically predicted MDD was associated with a higher risk of fracture and vice versa. In addition, we found that the higher expression level of SGIP1 in the spinal cord and muscle was associated with an increased risk of fracture and MDD. Conclusions: The genetic pleiotropy between MDD and fracture highlights the bidirectional association observed in the epidemiological analysis. The shared genetic components (such as SGIP1) between the diseases suggest that modulating the endocannabinoid system could be a potential therapeutic strategy for both MDD and bone loss.

P2X7 receptor-dependent increase in endocannabinoid 2-arachidonoyl glycerol production by neuronal cells in culture: Dynamics and mechanism.

BACKGROUND AND PURPOSE: Neurotransmission and neuroinflammation are controlled by local increases in both extracellular ATP and the endocannabinoid 2-arachidonoyl glycerol (2-AG). While it is known that extracellular ATP stimulates 2-AG production in cells in culture, the dynamics and molecular mechanisms that underlie this response remain poorly understood. Detection of real-time changes in eCB levels with the genetically encoded sensor, GRABeCB2.0, can address this shortfall. EXPERIMENTAL APPROACH: 2-AG and arachidonoylethanolamide (AEA) levels in Neuro2a (N2a) cells were measured by LC-MS, and GRABeCB2.0 fluorescence changes were detected using live-cell confocal microscopy and a 96-well fluorescence plate reader. KEY RESULTS: 2-AG and AEA increased GRABeCB2.0 fluorescence in N2a cells with EC50 values of 81 and 58 nM, respectively; both responses were reduced by the cannabinoid receptor type 1 (CB1R) antagonist SR141617 and absent in cells expressing the mutant-GRABeCB2.0. ATP increased only 2-AG levels in N2a cells, as measured by LC-MS, and induced a transient increase in the GRABeCB2.0 signal within minutes primarily via activation of P2X7 receptors (P2X7R). This response was dependent on diacylglycerol lipase ß activity, partially dependent on extracellular calcium and phospholipase C activity, but not controlled by the 2-AG hydrolysing enzyme, α/ß-hydrolase domain containing 6 (ABHD6). CONCLUSIONS AND IMPLICATIONS: Considering that P2X7R activation increases 2-AG levels within minutes, our results show how these molecular components are mechanistically linked. The specific molecular components in these signalling systems represent potential therapeutic targets for the treatment of neurological diseases, such as chronic pain, that involve dysregulated neurotransmission and neuroinflammation.

Transcription directionality is licensed by Integrator at active human promoters.

A universal characteristic of eukaryotic transcription is that the promoter recruits RNA polymerase II (RNAPII) to produce both precursor mRNAs (pre-mRNAs) and short unstable promoter upstream transcripts (PROMPTs) toward the opposite direction. However, how the transcription machinery selects the correct direction to produce pre-mRNAs is largely unknown. Here, through multiple acute auxin-inducible degradation systems, we show that rapid depletion of an RNAPII-binding protein complex, Integrator, results in robust PROMPT accumulation throughout the genome. Interestingly, the accumulation of PROMPTs is compensated by the reduction of pre-mRNA transcripts in actively transcribed genes. Consistently, Integrator depletion alters the distribution of polymerase between the sense and antisense directions, which is marked by increased RNAPII-carboxy-terminal domain Tyr1 phosphorylation at PROMPT regions and a reduced Ser2 phosphorylation level at transcription start sites. Mechanistically, the endonuclease activity of Integrator is critical to suppress PROMPT production. Furthermore, our data indicate that the presence of U1 binding sites on nascent transcripts could counteract the cleavage activity of Integrator. In this process, the absence of robust U1 signal at most PROMPTs allows Integrator to suppress the antisense transcription and shift the transcriptional balance in favor of the sense direction.