A light-driven enzymatic enantioselective radical acylation.

Enzymes are recognized as exceptional catalysts for achieving high stereoselectivities1-3, but their ability to control the reactivity and stereoinduction of free radicals lags behind that of chemical catalysts4. Thiamine diphosphate (ThDP)-dependent enzymes5 are well-characterized systems that inspired the development of N-heterocyclic carbenes (NHCs)6-8 but have not yet been proved viable in asymmetric radical transformations. There is a lack of a biocompatible and general radical-generation mechanism, as nature prefers to avoid radicals that may be harmful to biological systems9. Here we repurpose a ThDP-dependent lyase as a stereoselective radical acyl transferase (RAT) through protein engineering and combination with organophotoredox catalysis10. Enzyme-bound ThDP-derived ketyl radicals are selectively generated through single-electron oxidation by a photoexcited organic dye and then cross-coupled with prochiral alkyl radicals with high enantioselectivity. Diverse chiral ketones are prepared from aldehydes and redox-active esters (35 examples, up to 97% enantiomeric excess (e.e.)) by this method. Mechanistic studies reveal that this previously elusive dual-enzyme catalysis/photocatalysis directs radicals with the unique ThDP cofactor and evolvable active site. This work not only expands the repertoire of biocatalysis but also provides a unique strategy for controlling radicals with enzymes, complementing existing chemical tools.

Cryo-EM structure of HQNO-bound Alternative Complex III from the anoxygenic phototrophic bacterium Chloroflexus aurantiacus.

Alternative complex III (ACIII) couples quinol oxidation and electron acceptor reduction with potential transmembrane proton translocation. It is compositionally and structurally different from the cytochrome bc1/b6f complexes, but functionally replaces these enzymes in the photosynthetic and/or respiratory electron transport chains (ETCs) of many bacteria. However, the true compositions and architectures of ACIIIs remain unclear, as do their structural and functional relevance in mediating the ETCs. We here determined cryogenic electron microscopy structures of photosynthetic ACIII isolated from Chloroflexus aurantiacus (CaACIIIp), in apo-form and in complexed form bound to a menadiol analog 2-heptyl-4-hydroxyquinoline-N-oxide (HQNO). Besides six canonical subunits (ActABCDEF), the structures revealed conformations of two previously unresolved subunits, ActG and I, which contributed to the complex stability. We also elucidated the structural basis of menaquinol oxidation and subsequent electron transfer along the [3Fe-4S]-6 hemes wire to its periplasmic electron acceptors, using electron paramagnetic resonance (EPR), spectroelectrochemistry, enzymatic analyses and molecular dynamics (MD) simulations. A unique insertion loop in ActE was shown to function in determining the binding specificity of CaACIIIp for downstream electron acceptors. This study broadens our understanding of the structural diversity and molecular evolution of ACIIIs, enabling further investigation of the (mena)quinol oxidoreductases evolved coupling mechanism in bacterial energy conservation.

The calcium-dependent protein kinase CPK16 regulates hypoxia-induced ROS production by phosphorylating the NADPH oxidase RBOHD in Arabidopsis.

Reactive oxygen species (ROS) production is a key event in modulating plant responses to hypoxia and post-hypoxia reoxygenation. However, the molecular mechanism by which hypoxia-associated ROS homeostasis is controlled remains largely unknown. Here, we showed that the calcium-dependent protein kinase CPK16 regulates plant hypoxia tolerance by phosphorylating the plasma membrane-anchored NADPH oxidase respiratory burst oxidase homolog D (RBOHD) to regulate ROS production in Arabidopsis (Arabidopsis thaliana). In response to hypoxia or reoxygenation, CPK16 was activated through phosphorylation of its Ser274 residue. The cpk16 knockout mutant displayed enhanced hypoxia tolerance, whereas CPK16-overexpressing (CPK16-OE) lines showed increased sensitivity to hypoxic stress. In agreement with these observations, hypoxia and reoxygenation both induced ROS accumulation in the rosettes of CPK16-OEs more strongly than in the rosettes of the cpk16-1 mutant or the wild type. Moreover, CPK16 interacted with and phosphorylated the N-terminus of RBOHD at 4 serine residues (Ser133, Ser148, Ser163, and Ser347) that were necessary for hypoxia- and reoxygenation-induced ROS accumulation. Furthermore, the hypoxia-tolerant phenotype of cpk16-1 was fully abolished in the cpk16 rbohd double mutant. Thus, we have uncovered a regulatory mechanism by which the CPK16-RBOHD module shapes the ROS production during hypoxia and reoxygenation in Arabidopsis.

Excessive nucleic acid R-loops induce mitochondria-dependent epithelial cell necroptosis and drive spontaneous intestinal inflammation.

Oxidative stress, which can be activated by a variety of environmental risk factors, has been implicated as an important pathogenic factor for inflammatory bowel disease (IBD). However, how oxidative stress drives IBD onset remains elusive. Here, we found that oxidative stress was strongly activated in inflamed tissues from both ulcerative colitis patients and Crohn's disease patients, and it caused nuclear-to-cytosolic TDP-43 transport and a reduction in the TDP-43 protein level. To investigate the function of TDP-43 in IBD, we inducibly deleted exons 2 to 3 of Tardbp (encoding Tdp-43) in mouse intestinal epithelium, which disrupted its nuclear localization and RNA-processing function. The deletion gave rise to spontaneous intestinal inflammation by inducing epithelial cell necroptosis. Suppression of the necroptotic pathway with deletion of Mlkl or the RIP1 inhibitor Nec-1 rescued colitis phenotypes. Mechanistically, disruption of nuclear TDP-43 caused excessive R-loop accumulation, which triggered DNA damage and genome instability and thereby induced PARP1 hyperactivation, leading to subsequent NAD+ depletion and ATP loss, consequently activating mitochondrion-dependent necroptosis in intestinal epithelial cells. Importantly, restoration of cellular NAD+ levels with NAD+ or NMN supplementation, as well as suppression of ALKBH7, an α-ketoglutarate dioxygenase in mitochondria, rescued TDP-43 deficiency-induced cell death and intestinal inflammation. Furthermore, TDP-43 protein levels were significantly inversely correlated with γ-H2A.X and p-MLKL levels in clinical IBD samples, suggesting the clinical relevance of TDP-43 deficiency-induced mitochondrion-dependent necroptosis. Taken together, these findings identify a unique pathogenic mechanism that links oxidative stress to intestinal inflammation and provide a potent and valid strategy for IBD intervention.

Exploring the Spatial Landscape of the Estrogen Receptor Proximal Proteome With Antibody-Based Proximity Labeling.

Estrogen receptor α (ERα) drives the transcription of genes involved in breast cancer (BC) progression, relying on coregulatory protein recruitment for its transcriptional and biological activities. Mutation of ERα as well as aberrant recruitment of its regulatory proteins contribute to tumor adaptation and drug resistance. Therefore, understanding the dynamic changes in ERα protein interaction networks is crucial for elucidating drug resistance mechanisms in BC. Despite progress in studying ERα-associated proteins, capturing subcellular transient interactions remains challenging and, as a result, significant number of important interactions remain undiscovered. In this study, we employed biotinylation by antibody recognition (BAR), an innovative antibody-based proximity labeling (PL) approach, coupled with mass spectrometry to investigate the ERα proximal proteome and its changes associated with resistance to aromatase inhibition, a key therapy used in the treatment of ERα-positive BC. We show that BAR successfully detected most of the known ERα interactors and mainly identified nuclear proteins, using either an epitope tag or endogenous antibody to target ERα. We further describe the ERα proximal proteome rewiring associated with resistance applying BAR to a panel of isogenic cell lines modeling tumor adaptation in the clinic. Interestingly, we find that ERα associates with some of the canonical cofactors in resistant cells and several proximal proteome changes are due to increased expression of ERα. Resistant models also show decreased levels of estrogen-regulated genes. Sensitive and resistant cells harboring a mutation in the ERα (Y537C) revealed a similar proximal proteome. We provide an ERα proximal protein network covering several novel ERα-proximal partners. These include proteins involved in highly dynamic processes such as sumoylation and ubiquitination difficult to detect with traditional protein interaction approaches. Overall, we present BAR as an effective approach to investigate the ERα proximal proteome in a spatial context and demonstrate its application in different experimental conditions.

Structure of the human respiratory complex II.

Human complex II is a key protein complex that links two essential energy-producing processes: the tricarboxylic acid cycle and oxidative phosphorylation. Deficiencies due to mutagenesis have been shown to cause mitochondrial disease and some types of cancers. However, the structure of this complex is yet to be resolved, hindering a comprehensive understanding of the functional aspects of this molecular machine. Here, we have determined the structure of human complex II in the presence of ubiquinone at 2.86 Å resolution by cryoelectron microscopy, showing it comprises two water-soluble subunits, SDHA and SDHB, and two membrane-spanning subunits, SDHC and SDHD. This structure allows us to propose a route for electron transfer. In addition, clinically relevant mutations are mapped onto the structure. This mapping provides a molecular understanding to explain why these variants have the potential to produce disease.

Rice transcription factor OsWRKY37 positively regulates flowering time and grain fertility under copper deficiency.

Efficient uptake, translocation, and distribution of Cu to rice (Oryza sativa) spikelets is crucial for flowering and yield production. However, the regulatory factors involved in this process remain unidentified. In this study, we isolated a WRKY transcription factor gene induced by Cu deficiency, OsWRKY37, and characterized its regulatory role in Cu uptake and transport in rice. OsWRKY37 was highly expressed in rice roots, nodes, leaf vascular bundles, and anthers. Overexpression of OsWRKY37 promoted the uptake and root-to-shoot translocation of Cu in rice under -Cu condition but not under +Cu condition. While mutation of OsWRKY37 significantly decreased Cu concentrations in the stamen, the root-to-shoot translocation and distribution ratio in brown rice affected pollen development, delayed flowering time, decreased fertility, and reduced grain yield under -Cu condition. yeast one-hybrid, transient co-expression and EMSAs, together with in situ RT-PCR and RT-qPCR analysis, showed that OsWRKY37 could directly bind to the upstream promoter region of OsCOPT6 (copper transporter) and OsYSL16 (yellow stripe-like protein) and positively activate their expression levels. Analyses of oscopt6 mutants further validated its important role in Cu uptake in rice. Our study demonstrated that OsWRKY37 acts as a positive regulator involved in the uptake, root-to-shoot translocation, and distribution of Cu through activating the expression of OsCOPT6 and OsYSL16, which is important for pollen development, flowering, fertility, and grain yield in rice under Cu deficient conditions. Our results provide a genetic strategy for improving rice yield under Cu deficient condition.

mTORC2-AKT signaling to PFKFB2 activates glycolysis that enhances stemness and tumorigenicity of intestinal epithelial cells.

Although elevated glycolysis has been widely recognized as a hallmark for highly proliferating cells like stem cells and cancer, its regulatory mechanisms are still being updated. Here, we found a previously unappreciated mechanism of mammalian target of rapamycin complex 2 (mTORC2) in regulating glycolysis in intestinal stem cell maintenance and cancer progression. mTORC2 key subunits expression levels and its kinase activity were specifically upregulated in intestinal stem cells, mouse intestinal tumors, and human colorectal cancer (CRC) tissues. Genetic ablation of its key scaffolding protein Rictor in both mouse models and cell lines revealed that mTORC2 played an important role in promoting intestinal stem cell proliferation and self-renewal. Moreover, utilizing mouse models and organoid culture, mTORC2 loss of function was shown to impair growth of gut adenoma and tumor organoids. Based on these findings, we performed RNA-seq and noticed significant metabolic reprogramming in Rictor conditional knockout mice. Among all the pathways, carbohydrate metabolism was most profoundly altered, and further studies demonstrated that mTORC2 promoted glycolysis in intestinal epithelial cells. Most importantly, we showed that a rate-limiting enzyme in regulating glycolysis, 6-phosphofructo-2-kinase (PFKFB2), was a direct target for the mTORC2-AKT signaling. PFKFB2 was phosphorylated upon mTORC2 activation, but not mTORC1, and this process was AKT-dependent. Together, this study has identified a novel mechanism underlying mTORC2 activated glycolysis, offering potential therapeutic targets for treating CRC.

A chemical timer approach to delayed chemiluminescence.

Although the onset time of chemical reactions can be manipulated by mechanical, electrical, and optical methods, its chemical control remains highly challenging. Herein, we report a chemical timer approach for manipulating the emission onset time of chemiluminescence (CL) reactions. A mixture of Mn2+, NaHCO3, and a luminol analog with H2O2 produced reactive oxygen species (ROS) radicals and other superoxo species (superoxide containing complex) with high efficiency, accompanied by strong and immediate CL emission. Surprisingly, the addition of thiourea postponed CL emission in a concentration-dependent manner. The delay was attributed to a slow-generation-scavenging mechanism, which was found to be generally applicable not only to various types of CL reagents and ROS radical scavengers but also to popular chromogenic reactions. The precise regulation of CL kinetics was further utilized in dynamic chemical coding with improved coding density and security. This approach provides a powerful platform for engineering chemical reaction kinetics using chemical timers, which is of application potential in bioassays, biosensors, CL microscopic imaging, microchips, array chips, and informatics.

Secretome of Cancer-Associated Fibroblasts (CAFs) Influences Drug Sensitivity in Cancer Cells.

Resistance is a major problem with effective cancer treatment and the stroma forms a significant portion of the tumor mass but traditional drug screens involve cancer cells alone. Cancer-associated fibroblasts (CAFs) are a major tumor stroma component and its secreted proteins may influence the function of cancer cells. The majority of secretome studies compare different cancer or CAF cell lines exclusively. Here, we present the direct characterization of the secreted protein profiles between CAFs and KRAS mutant-cancer cell lines from colorectal, lung, and pancreatic tissues using multiplexed mass spectrometry. 2573 secreted proteins were annotated, and differential analysis highlighted understudied CAF-enriched secreted proteins, including Wnt family member 5B (WNT5B), in addition to established CAF markers, such as collagens. The functional role of CAF secreted proteins was explored by assessing its effect on the response to 97 anticancer drugs since stromal cells may cause a differing cancer drug response, which may be missed on routine drug screening using cancer cells alone. CAF secreted proteins caused specific effects on each of the cancer cell lines, which highlights the complexity and challenges in cancer treatment and so the importance to consider stromal elements.

Targeting Neuronal GPR65 With Delayed BTB09089 Treatment Improves Neurorehabilitation Following Ischemic Stroke.

BACKGROUND: GPR65 (G protein-coupled receptor 65) can sense extracellular acidic environment to regulate pathophysiological processes. Pretreatment with the GPR65 agonist BTB09089 has been proven to produce neuroprotection in acute ischemic stroke. However, whether delayed BTB09089 treatment and neuronal GPR65 activation promote neurorestoration remains unknown. METHODS: Ischemic stroke was induced in wild-type (WT) or GPR65 knockout (GPR65-/-) mice by photothrombotic ischemia. Male mice were injected intraperitoneally with BTB09089 every other day at days 3, 7, or 14 poststroke. AAV-Syn-GPR65 (adenoassociated virus-synapsin-GPR65) was utilized to overexpress GPR65 in the peri-infarct cortical neurons of GPR65-/- and WT mice. Motor function was monitored by grid-walk and cylinder tests. The neurorestorative effects of BTB09089 were observed by immunohistochemistry, Golgi-Cox staining, and Western blotting. RESULTS: BTB09089 significantly promoted motor outcomes in WT but not in GPR65-/- mice, even when BTB09089 was delayed for 3 to 7 days. BTB09089 inhibited the activation of microglia and glial scar progression in WT but not in GPR65-/- mice. Meanwhile, BTB09089 reduced the decrease in neuronal density in WT mice, but this benefit was abolished in GPR65-/- mice and reemerged by overexpressing GPR65 in peri-infarct cortical neurons. Furthermore, BTB09089 increased the GAP43 (growth-associated protein-43) and synaptophysin puncta density, dendritic spine density, dendritic branch length, and dendritic complexity by overexpressing GPR65 in the peri-infarct cortical neurons of GPR65-/- mice, which was accompanied by increased levels of p-CREB (phosphorylated cAMP-responsive element-binding protein). In addition, the therapeutic window of BTB09089 was extended to day 14 by overexpressing GPR65 in the peri-infarct cortical neurons of WT mice. CONCLUSIONS: Our findings indicated that delayed BTB09089 treatment improved neurological functional recovery and brain tissue repair poststroke through activating neuronal GRP65. GPR65 overexpression may be a potential strategy to expand the therapeutic time window of GPR65 agonists for neurorehabilitation after ischemic stroke.

Redox-Regulated and Guest-Driven Transformations of Aromatic Oligoamide Foldamers in Advanced Structures.

In this study, we demonstrate that an aromatic oligoamide sequence assembles into a trimeric helix-turn-helix architecture with a disulfide linkage, and upon cleavage of this linkage, it reconstructs into an antiparallel double helix. The antiparallel double helix is accessible to encapsulate a diacid guest within its cavity, forming a 2:1 host-guest complex. In contrast, hydrogen-bonding interactions between the trimeric-assembled structure and guests induce a conformational shift in the trimeric helix, resulting in a cross-shaped double-helix complex at a 2:2 host-guest ratio. Interconversions between the trimeric helix and the antiparallel double helix, along with their respective host-guest complexes, can be initiated through thiol/disulfide redox-mediated regulation.

Chiral Nanostructured Ag Films for Multicarbon Products from CO2 Electroreduction.

The formation of multicarbon products from CO2 electroreduction is challenging on materials other than Cu-based catalysts. Ag has been known to be a typical metal catalyst, producing CO in CO2 electroreduction. The formation of C2+ products by Ag has never been reported because the carbon-carbon (C-C) coupling is an unfavorable process due to the high reaction barrier energy of *OCCO. Here, we propose that the chirality-induced spin polarization of chiral nanostructured Ag films (CNAFs) can promote the formation of triplet OCCO by regulating its parallel electron spin alignment, and the helical lattice distortion of nanostructures can decrease the reaction energy of *OCCO, which triggers C-C coupling and promotes subsequent *OCCO hydrogenation to facilitate the generation of C2+ products. The CNAFs with helically lattice-distorted nanoflakes were fabricated via electrodeposition using phenylalanine as the symmetry-breaking agent. C2+ products (C2H4, C2H6, C3H8, C2H5OH, and CH3COOH) with a Faradaic efficiency of ∼4.7% and a current density of ∼22 mA/cm2 were generated in KHCO3 electrolytes under 12.5 atm of CO2 (g). Our findings propose that the chiral nanostructured materials can regulate the multifunctionality of catalytic performance in the catalytic reactions with triplet intermediates and products.

Enzyme Core Spherical Nucleic Acid That Enables Enhanced Cuproptosis and Antitumor Immune Response through Alleviating Tumor Hypoxia.

Cuproptosis, a copper-dependent cell death process, has been confirmed to further activate the immune response and mediate the immune resistance. However, hypoxic tumor microenvironment hampers cuproptosis sensitivity and suppresses the body's antitumor immune response. Herein, we have successfully immobilized and functionalized catalase (CAT) with long single-stranded DNA containing polyvalent CpG sequences through rolling circle amplification (RCA) techniques, obtaining an enzyme-cored spherical nucleic acid nanoplatform (CAT-ecSNA-Cu) to deliver copper ions for cuproptosis. The presence of long-stranded DNA-protected CAT enhances mitochondrial respiration by catalyzing the conversion of H2O2 to O2, thereby sensitizing cuproptosis. Meanwhile, increased tumor oxygenation suppresses the expression of the hypoxia-inducible factor-1 (HIF-1) protein, resulting in the alleviation of the immunosuppressive tumor microenvironment. Of note, cuproptosis induces immunogenic cell death (ICD), which facilitates dendritic cell (DC) maturation and enhances antigen presentation through polyCpG-supported Toll-like receptor 9 (TLR9) activation. Furthermore, cuproptosis-induced PD-L1 upregulation in tumor cells complements checkpoint blockers (αPD-L1), enhancing antitumor immunity. The strategy of enhancing cuproptosis-mediated antitumor immune responses by alleviating hypoxia effectively promotes the activation and proliferation of effector T cells, ultimately leading to long-term immunity against cancer.

Galectin-3 contributes to pathogenesis of IgA nephropathy.

IgA nephropathy (IgAN) is the most common type of glomerulonephritis that frequently progresses to kidney failure. However, the molecular pathogenesis underlying IgAN remains largely unknown. Here, we investigated the role of galectin-3 (Gal-3), a galactoside-binding protein in IgAN pathogenesis, and showed that Gal-3 expression by the kidney was significantly enhanced in patients with IgAN. In both TEPC-15 hybridoma-derived IgA-induced, passive, and spontaneous "grouped" ddY IgAN models, Gal-3 expression was clearly increased with disease severity in the glomeruli, peri-glomerular regions, and some kidney tubules. Gal-3 knockout (KO) in the passive IgAN model had significantly improved proteinuria, kidney function and reduced severity of kidney pathology, including neutrophil infiltration and decreased differentiation of Th17 cells from kidney-draining lymph nodes, despite increased percentages of regulatory T cells. Gal-3 KO also inhibited the NLRP3 inflammasome, yet it enhanced autophagy and improved kidney inflammation and fibrosis. Moreover, administration of 6-de-O-sulfated, N-acetylated low-molecular-weight heparin, a competitive Gal-3 binding inhibitor, restored kidney function and improved kidney lesions in passive IgAN mice. Thus, our results suggest that Gal-3 is critically involved in IgAN pathogenesis by activating the NLRP3 inflammasome and promoting Th17 cell differentiation. Hence, targeting Gal-3 action may represent a new therapeutic strategy for treatment of this kidney disease.

Impact of UGT1A4 and UGT2B7 polymorphisms on lamotrigine plasma concentration in patients with bipolar disorder.

PURPOSE: The purpose of this study was to evaluate the effect of UGT1A4 and UGT2B7 polymorphisms on the plasma concentration of lamotrigine in Chinese patients with bipolar disorder. METHODS: A total of 104 patients were included in this study. Steady-state plasma lamotrigine concentrations were determined in each patient after at least 21 days of continuous treatment with a set dose of the drug. Lamotrigine plasma concentrations were ascertained using ultra-performance liquid chromatography. Simultaneously, plasma samples were used for patient genotyping. RESULTS: The age, sex, BMI, daily lamotrigine dose, plasma lamotrigine concentration, and lamotrigine concentration/dose ratio of patients exhibited significant differences, and these were associated with differences in the genotype [ UGT1A4 -142T>G and UGT2B7 -161C>T ( P  < 0.05)]. Patients with the GG and GT genotypes in UGT1A4 -142T>G had significantly higher lamotrigine concentration/dose values (1.6 ±â€…1.1 and 1.7 ±â€…0.5 µg/ml per mg/kg) than those with the TT genotype (1.4 ±â€…1.1 µg/ml per mg/kg). Likewise, patients with the UGT2B7 -161C>T TT genotype had significantly higher lamotrigine concentration/dose values (1.6 ±â€…1.1 µg/ml per mg/kg) than those with the CC genotype (1.3 ±â€…1.3 µg/ml per mg/kg). Multiple linear regression analysis showed that sex, lamotrigine dose, UGT1A4 -142T>G, and UGT2B7 -161C>T were the most important factors influencing lamotrigine pharmacokinetics ( P  < 0.001). CONCLUSION: The study results suggest that the UGT1A4 -142T>G and UGT2B7 -161C>T polymorphisms affect lamotrigine plasma concentrations in patients with bipolar disorder.

Effects of CYP3A5*3 genetic polymorphisms on the pharmacokinetics of perampanel in Chinese pediatric patients with epilepsy.

PURPOSE: This study was the first to evaluate the effect of CYP3A5*3 gene polymorphisms on plasma concentration of perampanel (PER) in Chinese pediatric patients with epilepsy. METHODS: We enrolled 98 patients for this investigation. Plasma PER concentrations were measured using liquid chromatography-tandem mass spectrometry. Leftover samples from standard therapeutic drug monitoring were allocated for genotyping analysis. The primary measure of efficacy was the rate of seizure reduction with PER treatment at the final checkup. RESULTS: The plasma concentration showed a linear correlation with the daily dose taken ( r  = 0.17; P  < 0.05). The ineffective group showed a significantly lower plasma concentration of PER (490.5 ±â€…297.1 vs. 633.8 ±â€…305.5 µg/ml; P  = 0.019). For the mean concentration-to-dose (C/D) ratio, the ineffective group showed a significantly lower C/D ratio of PER (3.2 ±â€…1.7 vs. 3.8 ±â€…2.0; P  = 0.040). The CYP3A5*3 CC genotype exhibited the highest average plasma concentration of PER at 562.8 ±â€…293.9 ng/ml, in contrast to the CT and TT genotypes at 421.1 ±â€…165.6 ng/ml and 260.0 ±â€…36.1 ng/ml. The mean plasma PER concentration was significantly higher in the adverse events group (540.8 ±â€…285.6 vs. 433.0 ±â€…227.2 ng/ml; P  = 0.042). CONCLUSION: The CYP3A5*3 gene's genetic polymorphisms influence plasma concentrations of PER in Chinese pediatric patients with epilepsy. Given that both efficacy and potential toxicity are closely tied to plasma PER levels, the CYP3A5*3 genetic genotype should be factored in when prescribing PER to patients with epilepsy.

Polarized Th2 cells attenuate high-fat-diet induced obesity through the suppression of lipogenesis.

Immune cells, such as macrophages, B cells, neutrophils and T cell subsets, have been implicated in the context of obesity. However, the specific role of Th2 cells in adipose tissue function has remained elusive. Eight-week-old male CD3ε─/─ mice were randomly divided into two groups (≥ 5 mice per group): one received intravenous injection of Th2 cells isolated from LATY136F mice, while the other receiving PBS as a control. Both of groups were subjected to a high-fat diet (HFD). The adoptive transfer of polarized Th2 cells led to a significant reduction in obesity following a HFD. This reduction was accompanied by improvements in hepatic steatosis, glucose intolerance, and insulin resistance. Mechanistically, Th2 cell treatment promoted oxidative phosphorylation of adipocytes, thereby contributing to a reduction of lipid droplet accumulation. These findings suggest that Th2 cell therapy represents a novel approach for treating diet-induced obesity and other diseases involving lipid droplet accumulation disorders.

Identification of Omitted Pollutants in Environmental Water via In Situ Thin-Film Microextraction.

Sample preparation is an inevitable step in the screening workflow for the identification of unknown pollutants in the aquatic environment. However, the possible loss of pollutants during sample preparation has aroused serious concern but remains not effectively resolved. This study shows that high-risk pollutants omitted in solid-phase extraction (SPE) can be identified via in situ thin-film microextraction (TFME) coupled with high-resolution mass spectrometry. It was observed that a total of 541 features showed higher mass spectrometry signal intensity by using in situ TFME in comparison with SPE. Subsequently, 28 compounds were identified from the features with higher intensity by comparing the recorded tandem mass spectra with the online database and validating with standards. Notably, six out of these compounds were completely omitted using SPE, including a dye, drug, and industrial product. It was confirmed that the lower extraction efficiencies of SPE were attributed to the limited sample volumes, the losses of compounds during sample transportation and storage, and the entrapment of compounds in SPE columns. This study demonstrates that in situ TFME should be used as a supplementary technique to SPE for extending the coverage of pollutants in the screening workflows.

On-Site Ratiometric Analysis of UO22+ with High Selectivity.

Uranyl ions (UO22+) are recognized as important indicators for monitoring sudden nuclear accidents. However, the interferences coexisting in the complicated environmental matrices impart serious constraints on the reliability of current on-site monitoring methods. Herein, a novel ratiometric method for the highly sensitive and selective detection of UO22+ is reported based on a [Eu(diaminoterephthalic acid)] (Eu-DATP) metal-organic framework. Benefiting from the unique chemical structure of Eu-DATP, energy transfer from DATP to UO22+ was enabled, resulting in the up-regulated fluorescence of UO22+ and the simultaneous down-regulated fluorescence of Eu3+. The limit of detection reached as low as 2.7 nM, which was almost 2 orders of magnitude below the restricted limit in drinking water set by the United States Environmental Protection Agency (130 nM). The Eu-DATP probe showed excellent specificity to UO22+ over numerous interfering species, as the intrinsic emissions of UO22+ were triggered. This unprecedentedly high selectivity is especially beneficial for monitoring UO22+ in complicated environmental matrices with no need for tedious sample pretreatment, such as filtration and digestion. Then, by facilely equipping a Eu-DATP-based sampler on a drone, remotely controlled sampling and on-site analysis in real water samples were realized. The concentrations of UO22+ were determined to be from 16.5 to 23.5 nM in the river water of the Guangzhou downtown area, which was consistent with the results determined by the gold-standard inductively coupled plasma mass spectrometry. This study presents a reliable and convenient method for the on-site analysis of UO22+.