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Background: The limited radiosensitivity of osteosarcoma poses a challenge in applying radiotherapy, necessitating the search for effective radiosensitizing targets., Methods: The lentiviral vectors were employed to establish CDKN2C-overexpressing (CDKN2C-OE) and CDKN2C-negative control (CDKN2C-NC) HOS and U2OS osteosarcoma cells. Cells were treated with or without irradiation (IR) to assess radiosensitization via viability, proliferation, apoptosis, and cell cycle analysis. A mouse model with subcutaneous tumors from CDKN2C-OE and CDKN2C-NC HOS cells evaluated tumor growth post-IR. Immunohistochemical staining and Western blot analysis were conducted to confirm model establishment and explore mechanisms., Results: CDKN2C-OE combined with IR inhibited cell viability and proliferation, promoting apoptosis in vitro and inhibiting tumor growth in vivo. CDKN2C-OE inhibited G1 phase progression post-IR by suppressing Cyclin-dependent kinase 4 (CDK4) expression and Thr172 phosphorylation, reducing retinoblastoma protein (RB) phosphorylation at Ser807/811. CDKN2C-OE did not primarily impact the cell cycle by regulating the expression of CDK6 and Cyclin D1. Furthermore, when CDKN2C-OE was combined with IR, the expression of BAX, Caspase-3, and its active cleavage product, cleaved Caspase-3, was upregulated., Conclusions: Our research results indicate that overexpression of CDKN2C enhances radiosensitivity in osteosarcoma through the induction of G1 phase arrest and subsequent apoptosis. G1 phase arrest is mediated by the suppression of CDK4 expression and Thr172 phosphorylation, which consequently affects the expression of phosphorylated RB at the Ser807/811 sites., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024. Published by Elsevier Inc.)

Objectives: To evaluate the relative contribution of basal hyperglycemia (BHG) and postprandial hyperglycemia (PHG) to the time in range (TIR) categories and adverse pregnancy outcomes in pregnant women with type 1 diabetes mellitus (T1DM)., Materials and Methods: This observational study included 112 pregnancies with T1DM from the CARNATION study who wore continuous glucose monitoring (CGM) devices during pregnancy. The data from CGM were analyzed for TIR (range, 3.5-7.8 mmol/L), areas under the curve of PHG, area under the curve of BHG, and BHG and PHG contribution rates. The contribution rates of BHG and PHG to the different levels of TIR (<60%, 60-78%, ≥78%) and adverse pregnancy outcomes were analyzed., Results: The participants' average age was 28.8 ± 3.9 years with a diabetes duration of 8.4 ± 6.2 years. All women experienced a mean TIR of 75.6 ± 19.0% and a mean glycated hemoglobin of 6.2 ± 1.1% during pregnancy. The BHG contribution accounted for 74.9% (36.8, 100), 69.2% (13.4, 100), and 66.5% (10.0, 100) (P < .001) and PHG accounted for 25.1% (0, 63.2), 30.8% (0, 86.6), and 33.5% (0, 90.0) (P < .001) when participants experienced the TIR<60%, 60%-78%, and ≥78%, respectively. Participants with higher BHG contribution rates tended to have more adverse pregnancy outcomes., Conclusion: Basal hyperglycemia was the major contributor to TIR during pregnancy. Along with controlling PHG, pregnant women with T1DM who did not reach the target of TIR may benefit more from the optimization of insulin regimens focusing on reducing basal glucose., (© The Author(s) 2024. Published by Oxford University Press on behalf of the Endocrine Society. All rights reserved. For commercial re-use, please contact reprints@oup.com for reprints and translation rights for reprints. All other permissions can be obtained through our RightsLink service via the Permissions link on the article page on our site—for further information please contact journals.permissions@oup.com.)

RIG-I-like receptors (RLRs) are cytosolic RNA sensors critical for antiviral immunity. RLR activation is regulated by polyubiquitination and oligomerization following RNA binding. Yet, little is known about how RLRs exploit subcellular organelles to facilitate their posttranslational modifications and activation. Endosomal adaptor TAPE regulates the endosomal TLR and cytosolic RLR pathways. The potential interplay between RIG-I signaling and endosomes has been explored. Here, we report that endosomes act as platforms for facilitating RIG-I polyubiquitination and complex formation. RIG-I was translocated onto endosomes to form signaling complexes upon activation. Ablation of endosomes impaired RIG-I signaling to type I IFN activation. TAPE mediates the interaction and polyubiquitination of RIG-I and TRIM25. TAPE-deficient myeloid cells were defective in type I IFN activation upon RNA ligand and virus challenges. Myeloid TAPE deficiency increased the susceptibility to RNA virus infection in vivo. Our work reveals endosomes as signaling platforms for RIG-I activation and antiviral immunity.

Background: Open-source automated insulin delivery systems are increasingly adopted yet predominantly discussed outside of Asia. This study aimed to describe efficacy and safety of android artificial pancreas (AAPS) in people with type 1 diabetes mellitus (T1DM) from mainland China., Methods: This real-world study recruited people who initiated AAPS for ≥ 3 months between 2019 and 2024. Key outcomes included glycated hemoglobin A1c (HbA1c) and metrics from continuous glucose monitoring, rates of diabetic ketoacidosis (DKA) and severe hypoglycemia., Findings: 292 (male, 46·9 %) participants aged 25·7 (14·7, 35·0) years were included, with 183 (62·7 %) and 68 (23·3 %) using AAPS for 6 and 12 months. Prior-AAPS HbA1c was 7·6 ± 1·7 % with 44·5 % achieving < 7·0 %. After 3 months, mean HbA1c improved by -1·5 ± 2·0 % to 6·3 ± 0·8 % (P < 0.01), with 82·9 % achieving < 7.0 %. Time in range 3·9-10·0 mmol/L (TIR) improved to 78·8 ± 12·9 %, with 80·5 % achieving > 70 %, followed by time below 3·9 mmol/L of 3·9 (2·1, 6·1) %. After 12 months, HbA1c and TIR remained similar at 6·4 ± 1·0 % and 77·9 ± 12·2 %. No DKA and severe hypoglycemia was observed., Interpretation: Real-world data from mainland China highlights current uptake of open-source AAPS with potential glycemic benefits. No safety signals are seen. More support to enhance access and utilization of all AID systems in this region is warranted., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Objective: As more states legalize cannabis in the US, marketing from the cannabis industry and news coverage of cannabis have increased. Sources of information on cannabis can influence beliefs about risks and benefits. Yet, little is known about how the use and influence of specific sources of information have changed over time., Method: We conducted a longitudinal study of 5053 US adults between 2017 and 2021. Participants were asked about sources of information on cannabis risks and benefits and which sources were most influential using a web-based survey at three different time points (2017, 2020, 2021). We evaluated changes in the use/influence of multiple sources of information from 2017 to 2021 and examined interactions with age, cannabis use and state cannabis legal status., Results: The largest increases in sources of information on cannabis benefits and risks were in use of health professionals (+5.5% for benefits and +17.4% for risks). The largest declines were in use of traditional media (TV, radio, newspapers) (-12.3% and -11.4%). Use of cannabis advertisements and dispensaries/other industry sources also significantly increased. Health professionals were the most influential source of information in all three waves regardless of age, cannabis use or state legal status., Conclusions: Participants sought information from different sources, and increasingly relied on health professionals as a primary source, highlighting the need to train healthcare providers about cannabis risks and benefits. While fewer people used traditional media, use of industry sources increased, underscoring the need for accurate cannabis information sources.

The trithorax group (TrxG) complex is an important protein in the regulation of plant histone methylation. The ABSENT, SMALL, OR HOMEOTIC DISCS 1 (ASH1) gene family, as important family members of the TrxG complex, has been shown to regulate tolerance to abiotic stress and growth and development in many plants. In this study, we identified nine GhASH1s in upland cotton. Bioinformatics analysis revealed that GhASH1s contain a variety of cis-acting elements related to stress resistance and growth and development. The transcriptome expression profiles revealed that GhASHH1.A and GhASHH2.A genes expression were upregulated in flower organs and in response to external temperature stress. The results of virus-induced gene silencing (VIGS) indicated that GhASHH1.A and GhASHH2.A genes silencing reduced the ability of cotton to adapt to temperature stress and delayed the development of the flowering phenotype. We also showed that the silencing of these two target genes did not induce early flowering at high temperature (32 °C), suggesting that GhASHH1.A and GhASHH2.A might regulate cotton flowering in response to temperature. These findings provide genetic resources for future breeding of early-maturing and temperature-stress-tolerant cotton varieties.

Background: The triglyceride-glucose (TyG) index is identified as an alternative indicator of insulin resistance (IR) and is associated with macro- and micro-vascular diseases among patients with type 2 diabetes mellitus. The relationship between the TyG index and IR and its impact on diabetic kidney disease (DKD) remains unclear among adults with type 1 diabetes mellitus(T1DM)., Methods: This study comprised a cross-sectional analysis using data from the Guangdong T1DM Translational Medicine Study (GTT) and a longitudinal analysis using data from the type 1 diabetes (T1D) Exchange registry study. Correlation analysis was used to investigate the association between the TyG index and IR. Logistic regression and Cox proportional hazards regression were performed to explore the impact of the TyG index on DKD risk., Results: The GTT Study included 836 adults (216 with DKD and 620 without DKD). A significant correlation existed between the TyG index and the estimated glucose disposal rate (r=-0.64, p < 0.01). The TyG index was a risk factor for DKD after confounder adjustment (OR = 1.34, 95% CI:1.03-1.74). The T1D Exchange registry study included 8,771 adults (2,050 with DKD and 6,721 without DKD). After adjusting confounding factors, the TyG index was identified as an independent risk factor for DKD at enrollment, with the highest risk of DKD incidence observed in the highest TyG tertile group (OR = 1.92, 95%CI:1.67-2.20). During a median follow-up of 44.58(21.84, 67.09) months, the risk of developing DKD was increased by 32% at every 1 SD increase of the TyG index over time among participants without DKD at enrollment., Conclusions: The TyG index could be used to assess IR and was identified as an independent risk factor of DKD among adults with T1DM., (© 2024. The Author(s).)

Irritability, a state of excessive reactivity to negative emotional stimuli, is common in individuals with autism spectrum disorder (ASD). Although it has a significant negative impact of patients' disease severity and quality of life, the neural mechanisms underlying irritability in ASD remain largely unclear. We have previously demonstrated that male mice lacking the Coiled-coil and C2 domain containing 1a (Cc2d1a) in forebrain excitatory neurons recapitulate numerous ASD-like behavioral phenotypes, including impaired social behaviors and pronounced repetitive behaviors. Here, using the bottle-brush test (BBT) to trigger and evaluate aggressive and defensive responses, we show that Cc2d1a deletion increases irritability-like behavior in male but not female mice, which is correlated with reduced number of oxytocin (OXT)-expressing neurons in the paraventricular nucleus (PVN) of the hypothalamus. Intranasal OXT administration or chemogenetic activation of OXT neurons in the PVN rescues irritability-like behavior in Cc2d1a conditional knockout (cKO) mice. Administration of a selective melanocortin receptor 4 agonist, RO27-3225, which potentiates endogenous OXT release, also alleviates irritability-like behavior in Cc2d1a cKO mice, an effect blocked by a specific OXT receptor antagonist, L-368,899. We additionally identify a projection connecting the posterior ventral segment of the medial amygdala (MeApv) and ventromedial nucleus of the ventromedial hypothalamus (VMHvl) for governing irritability-like behavior during the BBT. Chemogenetic suppression of the MeApv-VMHvl pathway alleviates irritability-like behavior in Cc2d1a cKO mice. Together, our study uncovers dysregulation of OXT system in irritability-like behavior in Cc2d1a cKO mice during the BBT and provide translatable insights into the development of OXT-based therapeutics for clinical interventions., (© 2024. The Author(s), under exclusive licence to American College of Neuropsychopharmacology.)

Oncidium hybridum Lodd. is an important ornamental flower that is used as both a cut flower and a potted plant around the world; additionally, its pseudobulbs serve as essential carriers for floral organs and flower development. The NAM gene family is crucial for managing responses to various stresses as well as regulating growth in plants. However, the mechanisms by which NAM genes regulate the development of pseudobulbs remain unclear. In this study, a total of 144 NAM genes harboring complete structural domains were identified in O. hybridum . The 144 NAM genes were systematically classified into 14 distinct subfamilies via phylogenetic analysis. Delving deeper into the conserved motifs revealed that motifs 1-6 exhibited remarkable conservation, while motifs 7-10 presented in a few NAM genes only. Notably, NAM genes sharing identical specific motifs were classified into the same subfamily, indicating functional relatedness. Furthermore, the examination of occurrences of gene duplication indicated that the NAM genes display 16 pairs of tandem duplications along with five pairs of segmental duplications, suggesting their role in genetic diversity and potential adaptive evolution. By conducting a correlation analysis integrating transcriptomics and metabolomics at four stages of pseudobulb development, we found that OhNAM023 , OhNAM030 , OhNAM007 , OhNAM019 , OhNAM083 , OhNAM047 , OhNAM089 , and OhNAM025 exhibited significant relationships with the endogenous plant hormones jasmonates (JAs), hinting at their potential involvement in hormonal signaling. Additionally, OhNAM089 , OhNAM025 , OhNAM119 , OhNAM055 , and OhNAM136 showed strong links with abscisic acid (ABA) and abscisic acid glucose ester (ABA-GE), suggesting the possible regulatory function of these NAM genes in plant growth and stress responses. The 144 NAM genes identified in this study provide a basis for subsequent research and contribute to elucidating the intricate molecular mechanisms of NAM genes in Oncidium and potentially in other species.

Plants generate a range of physiological and molecular responses to sustain their growth and development when suffering heat stress. Avocado is a type of tropical fruit tree with high economic value. Most avocado cultivars delete, wither, or even die when exposed to heat stress for a long time, which seriously restricts the introduction and cultivation of avocados. In this study, samples of a heat-intolerant variety (' Hass ') were treated under heat stress, and the transcriptomics and metabolomics were analyzed, with the expectation of providing information on the variety improvement and domestication of avocados. The differentially expressed genes identified using transcriptome analysis mainly involved metabolic pathways such as plant hormone signal transduction, plant-pathogen interaction, and protein processing in the endoplasmic reticulum. Combined transcriptome and metabolome analysis indicated that the down-regulation of Hass.g03.10206 and Hass.g03.10205 in heat shock-like proteins may result in the reduced Trehalose and Sinapoyl aldehyde content. Metabolomics analysis results indicated that the decrease in Trehalose and Sinapoyl aldehyde content may be an important factor for heat intolerance. These results provide important clues for understanding the physiological mechanisms of adaptation to heat stress in avocados.

In this work, we developed a dual-targeting probe consisted of well-defined hyaluronan (HA) oligosaccharide and glucose (Glc) labeled with Rhodamine B (HGR). The probe was designed to enhance tumor targeting both in vitro and in vivo, by simultaneously targeting CD44 and Glc transporter 1 (GLUT1). The HA oligosaccharide component was crucial for accurately assessing the impact of sugar chain structure on targeting efficacy, while its unoccupied carboxyl groups could minimize interference with HA's binding affinity to CD44. In vitro studies demonstrated that HGR possessed remarkable cytocompatibility and superior targeting abilities compared to single-targeting probes. It displayed a marked preference for CD44 high /GLUT1 high cells rather than CD44 low /GLUT1 low cells. In vivo studies using murine models further confirmed the significantly enhanced targeting efficacy and excellent biocompatibility of HGR. Therefore, this designed dual-targeting probe holds potential for clinical tumor detection., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Introduction: Coal represents a significant natural resource in our world, and its quality and commercial value is primarily determined by its heating capacity. Numerous scientists worldwide have attempted to explore the impact of various environmental factors on coal rank, yet their conclusions are often inconsistent., Methods: In this study, the Illumina MiSeq sequencing approach was used to analyze the bacterial community from a low-rank coal mine as well as a high-rank mine. Moreover, we investigated the relationship between the physical and chemical properties of the coal and the bacterial composition., Results: Overall, we found that the high-rank coal exhibited higher heating value but higher total sulfur and lead levels. Considering the community of bacteria, the abundances of Phascolarctobacterium and Anaerostipes were highly elevated in the high-rank coal group. Most interestingly, the Anaerostipes abundance was correlated with coal quality positively. Additionally, the co-occurrence network of the bacterial community in the high-rank coal group showed much higher complexity. The bacterial functional potential predictions indicated elevated levels of phosphoenolpyruvate carboxykinase ATP, succinate dehydrogenase fumarate reductase flavoprotein subunit, and methylenetetrahydrofolate dehydrogenase NADP methenyltetrahydrofolate cyclohydrolase pathways., Conclusion: This study revealed that high-rank coal had more complicated co-occurrence network and elevated Anaerostipes abundance, which may suggest a potential biological pathway that can be explored to enhance coal quality., Competing Interests: HZ and JX were employed by the Shaanxi Coalbed Methane Development Co., Shaanxi Coal and Chemical Industry Group Co., Ltd. GL, JY, and HC were employed by the Shaanxi Coal Industry Company Limited. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Zou, Tian, Xu, Li, Chen, Yang, Ling, Shen and Guo.)

Cancer presents a significant health threat, necessitating the development of more precise, efficient, and less damaging treatment approaches. To address this challenge, we employed the 1-ethyl-(3-dimethyl aminopropyl) carbodiimide/ N -hydroxy succinimide (EDC/NHS) catalytic system and utilized quaternized chitosan oligosaccharide (HTCOSC) as a drug carrier to construct a nanoparticle delivery system termed HTCOSC-cRGD-ES2-MTX (CREM). This system specifically targets integrin αvβ3 on tumor cell surfaces and enables simultaneous loading of the antiangiogenic agent ES2 (IVRRADRAAVP) and the chemotherapy drug methotrexate (MTX). Due to its amphiphilic properties, CREM self-assembles into nanoparticles in aqueous solution, exhibiting an average diameter of 179.47 nm. Comparative studies demonstrated that CREM, in contrast to free ES2 and MTX-free nanoparticles (CRE), significantly suppressed the proliferation of EAhy926 endothelial cells and B16 melanoma cells in vitro , resulting in inhibition rates of 71.18 and 82.25%, respectively. Furthermore, CREM exhibited a hemolysis rate below 2%, indicating excellent in vitro antiangiogenic and antitumor activity as well as favorable blood compatibility. Additionally, both CRE and CREM demonstrated favorable tumor targeting capabilities through the specific binding action of cyclic RGD (cRGD) to integrin αvβ3. Further in vivo investigations revealed that CREM induced apoptosis in tumor cells via the mitochondrial apoptotic pathway and reduced the expression of angiogenic factors such as vascular endothelial growth factor (VEGF), thereby inhibiting tumor angiogenesis. This potent antitumor effect was evident through a tumor suppression rate of 80.19%. Importantly, histopathological staining (HE staining) demonstrated the absence of significant toxic side effects of CREM on various organs compared to MTX. In conclusion, the CREM nano drug delivery system synergistically enhances the therapeutic efficacy of antiangiogenic drugs and chemotherapeutic agents, thus offering a novel targeted approach for cancer treatment.

Key Message: Two genomic regions associated with FFBN and HFFBN and a potential regulatory gene (GhE6) of HFFBN were identified through the integration of RTM-GWAS and meta‑QTL analyses. Abstract The first fruit branch node (FFBN) and the height of the first fruit branch node (HFFBN) are two important traits that are related to plant architecture and early maturation in upland cotton. Several studies have been conducted to elucidate the genetic basis of these traits in cotton using biparental and natural populations. In this study, by using 9,244 SNP linkage disequilibrium block (SNPLDB) loci from 315 upland cotton accessions, we carried out restricted two-stage multilocus and multiallele genome-wide association studies (RTM-GWASs) and identified promising haplotypes/alleles of the four stable and true major SNPLDB loci that were significantly associated with FFBN and HFFBN. Additionally, a meta-quantitative trait locus (MQTL) analysis was conducted on 274 original QTLs that were reported in 27 studies, and 40 MQTLs associated with FFBN and HFFBN were identified. Through the integration of the RTM-GWAS and meta‑QTL analyses, two stable and true major SNPLDBs (LDB&#95;5&#95;15144433 and LDB&#95;16&#95;37952328) that were distributed in the two MQTLs were identified. Ultimately, 142 genes in the two genomic regions were annotated, and three candidate genes associated with FFBN and HFFBN were identified in the genomic region (A05:14.64-15.64 Mb) via RNA-Seq and qRT‒PCR. The results of virus-induced gene silencing (VIGS) experiments indicated that GhE6 was a key gene related to HFFBN and that GhDRM1 and GhGES were important genes associated with early flowering in upland cotton. These findings will aid in the future identification of molecular markers and genetic resources for developing elite early-maturing cultivars with ideal plant characteristics., (© 2024. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Cell-cell interactions play an important role in many biological processes, and various methods have been developed for controlling the cell-cell interactions. However, the effective and rapid control of intercellular interactions remains challenging. Herein, we report a novel, rapid, and effective electrochemical strategy without destroying the basic life processes for the dynamic control of intercellular interactions via liposome fusion. In the proposed system, bioorthogonal chemical groups and hydroquinone (HQ)- and aminooxy (AO)-tethered ligands were modified on the surface of living cells on the basis of the liposome fusion, enabling dynamical intercellular assemblies. Upon application of the corresponding oxidative potential, the "off-state" HQ could be oxidized to the "on-state" quinone (Q), which subsequently reacts with AO-tethered ligands to form stable oxime linkages under physiological conditions. This reaction effectively shortens the distance between cells, promoting the formation of cell clusters. When the corresponding reverse reductive potential is applied, the oxime linkage is cleaved, resulting in the release of the cells. Furthermore, we employed HQ- and AO-tethered ligands to modify mitochondria, inducing mitochondrial aggregation. This noninvasive and label-free strategy allows for the dynamic reversible regulation of intercellular interactions, enhancing our understanding of intercellular communication networks, and has the potential for improving the antitumor therapy efficacy.

Disturbing cellular redox homeostasis within malignant cells, particularly improving reactive oxygen species (ROS), is one of the effective strategies for cancer therapy. The ROS generation based on nanozymes presents a promising strategy for cancer treatment. However, the therapeutic efficacy is limited due to the insufficient catalytic activity of nanozymes or their high dependence on hydrogen peroxide (H 2 O 2 ) or oxygen. Herein, we reported a nanozyme (CSA) based on well-defined CuSe hollow nanocubes (CS) uniformly covered with Ag nanoparticles (AgNPs) to disturb cellular redox homeostasis and catalyze a cascade of intracellular biochemical reactions to produce ROS for the synergistic therapy of breast cancer. In this system, CSA could interact with the thioredoxin reductase (TrxR) and deplete the tumor microenvironment-activated glutathione (GSH), disrupting the cellular antioxidant defense system and augmenting ROS generation. Besides, CSA possessed high peroxidase-mimicking activity toward H 2 O 2 , leading to the generation of various ROS including hydroxyl radical ( • OH), superoxide radicals ( • O 2 - ), and singlet oxygen ( 1 O 2 ), facilitated by the Cu(II)/Cu(I) redox and H 2 O 2 cycling, and plentiful catalytically active metal sites. Additionally, due to the absorption and charge separation performance of AgNPs, the CSA exhibited excellent photothermal performance in the second near-infrared (NIR-II, 1064 nm) region and enhanced the photocatalytic ROS level in cancer cells. Owing to the inhibition of TrxR activity, GSH depletion, high peroxidase-mimicking activity of CSA, and abundant ROS generation, CSA displays remarkable and specific inhibition of tumor growth.

Background: The SET domain group (SDG) genes encode histone lysine methyltransferases, which regulate gene transcription by altering chromatin structure and play pivotal roles in plant flowering determination. However, few studies have investigated their role in the regulation of flowering in upland cotton., Results: A total of 86 SDG genes were identified through genome-wide analysis in upland cotton (Gossypium hirsutum). These genes were unevenly distributed across 25 chromosomes. Cluster analysis revealed that the 86 GhSDGs were divided into seven main branches. RNA-seq data and qRT‒PCR analysis revealed that lysine methyltransferase 3 (KMT3) genes were expressed at high levels in stamens, pistils and other floral organs. Using virus-induced gene silencing (VIGS), functional characterization of GhKMT3;1a and GhKMT3;2a revealed that, compared with those of the controls, the GhKMT3;1a- and GhKMT3;2a-silenced plants exhibited later budding and flowering and lower plant heightwere shorter. In addition, the expression of flowering-related genes (GhAP1, GhSOC1 and GhFT) significantly decreased and the expression level of GhSVP significantly increased in the GhKMT3;1a- and GhKMT3;2a-silenced plants compared with the control plants., Conclusion: A total of 86 SDG genes were identified in upland cotton, among which GhKMT3;1a and GhKMT3;2a might regulate flowering by affecting the expression of GhAP1, GhSOC1, GhFT and GhSVP. These findings will provide genetic resources for advanced molecular breeding in the future., (© 2024. The Author(s).)

Background: Inguinal hernia is a relatively common condition. Most patients with inguinal hernia require surgery. At present, mesh repair is one of the most effective methods to treat inguinal hernia, but insertion of the mesh can cause inflammation. Dexamethasone (DEX) can treat inflammation, but the mechanism by which DEX alleviates inflammation caused by inguinal hernia mesh placement remains unclear., Method: We randomly divided rats into groups: negative control (NC), inguinal hernia (IH), polypropylene mesh (PM), DEX treatment, and miR-155 treatment groups. RT-qPCR was performed to determine the expression of miR-155. ELISA was implemented to determine the secretion of IL-1β, IL-6, and IL-18. Western blotting was used to detect caspase-1, JAK1, p-JAK1, STAT3, and p-STAT3 expression. A dual-luciferase reporter gene array identified a connection between miR-155 and JAK1., Results: The results revealed that the expression of miR-155, IL-1β, IL-6, and IL-18 was upregulated in the PM group. After DEX treatment, the secretion of miR-155, caspase-1, IL-1β, IL-6, and IL-18 decreased. Dual luciferase results confirmed that miR-155 induced the targeted downregulation of JAK1, while a miR-155 mimic reversed the therapeutic effect of DEX, and the expression levels of p-JAK1 and p-STAT3 increased., Conclusion: DEX regulates the JAK1/STAT3 signaling pathway through miR-155 to relieve inflammation caused by inguinal hernia meshes., (© 2024. The Author(s), under exclusive licence to Springer-Verlag France SAS, part of Springer Nature.)

Efficient remediation of soil contaminated by polycyclic aromatic hydrocarbons (PAHs) is challenging. To determine whether soil ecoenzyme stoichiometry influences PAH degradation under biostimulation and bioaugmentation, this study initially characterized soil ecoenzyme stoichiometry via a PAH degradation experiment and subsequently designed a validation experiment to answer this question. The results showed that inoculation of PAH degradation consortia ZY-PHE plus vanillate efficiently degraded phenanthrene with a K value of 0.471 (depending on first-order kinetics), followed by treatment with ZY-PHE and control. Ecoenzyme stoichiometry data revealed that the EEA C:N , vector length and angle increased before day five and decreased during the degradation process. In contrast, EEA N:P decreased and then increased. These results indicated that the rapid PAH degradation period induced more C limitation and organic P mineralization. Correlation analysis indicated that the degradation rate K was negatively correlated with vector length, EEA C:P , and EEA N:P , suggesting that C limitation and relatively less efficient P mineralization could inhibit biodegradation. Therefore, incorporating liable carbon and acid phosphatase or soluble P promoted PAH degradation in soils with ZY-PHE. This study provides novel insights into the relationship between soil ecoenzyme stoichiometry and PAH degradation. It is suggested that soil ecoenzyme stoichiometry be evaluated before designing bioremeiation stragtegies for PAH contanminated soils., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Background and Purpose: Knowledge concerning the use AI models for the classification of glenohumeral osteoarthritis (GHOA) and avascular necrosis (AVN) of the humeral head is lacking. We aimed to analyze how a deep learning (DL) model trained to identify and grade GHOA on plain radiographs performs. Our secondary aim was to train a DL model to identify and grade AVN on plain radiographs., Patients and Methods: A modified ResNet-type network was trained on a dataset of radiographic shoulder examinations from a large tertiary hospital. A total of 7,139 radiographs were included. The dataset included various projections of the shoulder, and the network was trained using stochastic gradient descent. Performance evaluation metrics, area under the receiver operating characteristic curve (AUC), sensitivity, and specificity were used to assess the network's performance for each outcome., Results: The network demonstrated AUC values ranging from 0.73 to 0.93 for GHOA classification and > 0.90 for all AVN classification classes. The network exhibited lower AUC for mild cases compared with definitive cases of GHOA. When none and mild grades were combined, the AUC increased, suggesting difficulties in distinguishing between these 2 grades., Conclusion: We found that a DL model can be trained to identify and grade GHOA on plain radiographs. Furthermore, we show that a DL model can identify and grade AVN on plain radiographs. The network performed well, particularly for definitive cases of GHOA and any level of AVN. However, challenges remain in distinguishing between none and mild GHOA grades.

Key Message: The silencing of GhGASA14 and the identification of superior allelic variation in its coding region indicate that GhGASA14 may positively regulate flowering and the response to GA 3 . Gibberellic acid-stimulated Arabidopsis (GASA), a member of the gibberellin-regulated short amino acid family, has been extensively investigated in several plant species and found to be critical for plant growth and development. However, research on this topic in cotton has been limited. In this study, we identified 38 GhGASAs that were dispersed across 18 chromosomes in upland cotton, and all of these genes had a GASA core domain. Transcriptome expression patterns and qRT-PCR results revealed that GhGASA9 and GhGASA14 exhibited upregulated expression not only in the floral organs but also in the leaves of early-maturing cultivars. The two genes were functionally characterized by virus-induced gene silencing (VIGS), and the budding and flowering times after silencing the target genes were later than those of the control (TRV:00). Compared with that in the water-treated group (MOCK), the flowering period of the different fruiting branches in the GA 3 -treated group was more concentrated. Interestingly, allelic variation was detected in the coding sequence of GhGASA14 between early-maturing and late-maturing accessions, and the frequency of this favorable allele was greater in high-latitude cotton cultivars than in low-latitude ones. Additionally, a significant linear relationship was observed between the expression level of GhGASA14 and flowering time among the 12 upland cotton accessions. Taken together, these results indicated that GhGASA14 may positively regulate flowering time and respond to GA 3 . These findings could lead to the use of valuable genetic resources for breeding early-maturing cotton cultivars in the future., (© 2024. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

It still remains challenging to design multifunctional therapeutic reagents for effective cancer therapy under a unique tumor microenvironment including insufficient endogenous H 2 O 2 and O2, low pH, and a high concentration of glutathione (GSH). In this work, a CO-based phototherapeutic system triggered by photogenerated holes, which consisted of ionic liquid (IL), the CO prodrug Mn 2 (CO) 10 , and iridium(III) porphyrin (IrPor) modified carbonized ZIF-8-doped graphitic carbon nitride nanocomposite (IL/ZCN@Ir(CO)), was designed for cascade hypoxic tumors. Upon light irradiation, the photogenerated holes on IL/ZCN@Ir(CO) oxidize water into H 2 O 2 , which subsequently induces Mn 2 (CO) 10 to release CO. Meanwhile, IrPor can convert H 2 O 2 to hydroxyl radical (•OH) and subsequent singlet oxygen ( 1 O 2 ), which further triggers CO release. Moreover, the degraded MnO 2 shows activity for glutathione (GSH) depletion and mimics peroxidase, leading to GSH reduction and •OH production in tumors. Thus, this strategy can in situ release high concentrations of CO and reactive oxygen species (ROS) and deplete GSH to efficiently induce cell apoptosis under hypoxic conditions, which has a high inhibiting effect on the growth of tumors, offering an attractive strategy to amplify CO and ROS generation to meet therapeutic requirements in cancer treatment.

Human milk oligosaccharides (HMOs) are intricate glycans that promote healthy growth of infants and have been incorporated into infant formula as food additives. Despite their importance, the limited availability of asymmetrically branched HMOs hinders the exploration of their structure and function relationships. Herein, we report an enzymatic modular strategy for the efficient synthesis of these HMOs. The key branching enzyme for the assembly of branched HMOs, human β1,6-N-acetylglucosaminyltransferase 2 (GCNT2), was successfully expressed in Pichia pastoris for the first time. Then, it was integrated with six other bacterial glycosyltransferases to establish seven glycosylation modules. Each module comprises a one-pot multi-enzyme (OPME) system for in-situ generation of costly sugar nucleotide donors, combined with a glycosyltransferase for specific glycosylation. This approach enabled the synthesis of 31 branched HMOs and 13 linear HMOs in a stepwise manner with well-programmed synthetic routes. The binding details of these HMOs with related glycan-binding proteins were subsequently elucidated using glycan microarray assays to provide insights into their biological functions. This comprehensive collection of synthetic HMOs not only serves as standards for HMOs structure identification in complex biological samples but also significantly enhances the fields of HMOs glycomics, opening new avenues for biomedical applications., Competing Interests: Declaration of competing interest There is no conflict of interest to report., (Copyright © 2024. Published by Elsevier Ltd.)

The visualization of drugs in living systems has become key techniques in modern therapeutics. Recent advancements in optical imaging technologies and molecular design strategies have revolutionized drug visualization. At the subcellular level, super-resolution microscopy has allowed exploration of the molecular landscape within individual cells and the cellular response to drugs. Moving beyond subcellular imaging, researchers have integrated multiple modes, like optical near-infrared II imaging, to study the complex spatiotemporal interactions between drugs and their surroundings. By combining these visualization approaches, researchers gain supplementary information on physiological parameters, metabolic activity, and tissue composition, leading to a comprehensive understanding of drug behavior. This review focuses on cutting-edge technologies in drug visualization, particularly fluorescence imaging, and the main types of fluorescent molecules used. Additionally, we discuss current challenges and prospects in targeted drug research, emphasizing the importance of multidisciplinary cooperation in advancing drug visualization. With the integration of advanced imaging technology and molecular design, drug visualization has the potential to redefine our understanding of pharmacology, enabling the analysis of drug micro-dynamics in subcellular environments from new perspectives and deepening pharmacological research to the levels of the cell and organelles., Competing Interests: We declare that we have no financial or personal relationships with other people or organizations that can inappropriately influence our work., (© 2024 The Authors.)

Blocking the tumor nutrient supply through angiogenic inhibitors is an effective treatment approach for malignant tumors. However, using angiogenic inhibitors alone may not be enough to achieve a significant tumor response. Therefore, we recently designed a universal drug delivery system combining chemotherapy and anti-angiogenic therapy to target tumor cells while minimizing drug-related side effects. This system (termed as PCCE) is composed of biomaterial chondroitin sulfate (CS), the anti-angiogenic peptide ES2, and paclitaxel (PTX), which collectively enhance antitumor properties. Interestingly, the PCCE system is conferred exceptional cell membrane permeability due to inherent characteristics of CS, including CD44 receptor-mediated endocytosis. The PCCE could respond to the acidic and high glutathione conditions, thereby releasing PTX and ES2. PCCE could effectively inhibit the proliferation, migration, and invasion of tumor cells and cause apoptosis, while PCCE can affect the endothelial cells tube formation and exert anti-angiogenic function. Consistently, more potent in vivo antitumor efficacy and non-toxic sides were demonstrated in B16F10 xenograft mouse models. PCCE can achieve excellent antitumor activity via modulating angiogenic and apoptosis-related factors. In summary, we have successfully developed an intelligent and responsive CS-based nanocarrier known as PCCE for delivering various antitumor drugs, offering a promising strategy for treating malignant tumors., Competing Interests: Declaration of competing interest The authors declare no conflict of interests in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Unlike chemosynthetic drugs designed for specific molecular and disease targets, active small-molecule natural products typically have a wide range of bioactivities and multiple targets, necessitating extensive screening and development. To address this issue, we propose a strategy for the direct in situ microdynamic examination of potential drug candidates to rapidly identify their effects and mechanisms of action. As a proof-of-concept, we investigated the behavior of mussel oligosaccharide (MOS-1) by tracking the subcellular dynamics of fluorescently labeled MOS-1 in cultured cells. We recorded the entire dynamic process of the localization of fluorescein isothiocyanate (FITC)-MOS-1 to the lysosomes and visualized the distribution of the drug within the cell. Remarkably, lysosomes containing FITC-MOS-1 actively recruited lipid droplets, leading to fusion events and increased cellular lipid consumption. These drug behaviors confirmed MOS-1 is a candidate for the treatment of lipid-related diseases. Furthermore, in a high-fat HepG2 cell model and in high-fat diet-fed apolipoprotein E ( ApoE ) -/- mice, MOS-1 significantly promoted triglyceride degradation, reduced lipid droplet accumulation, lowered serum triglyceride levels, and mitigated liver damage and steatosis. Overall, our work supports the prioritization of in situ visual monitoring of drug location and distribution in subcellular compartments during the drug development phase, as this methodology contributes to the rapid identification of drug indications. Collectively, this methodology is significant for the screening and development of selective small-molecule drugs, and is expected to expedite the identification of candidate molecules with medicinal effects., Competing Interests: The authors declare that there are no conflicts of interest., (© 2024 The Authors.)

Minimally invasive thermal therapy is a successful alternative treatment to surgery in solid tumors with high complete ablation rates, however, tumor recurrence remains a concern. Central memory CD8 + T cells (T CM ) play important roles in protection from chronic infection and cancer. Here we find, by single-cell RNA analysis of human breast cancer samples, that although the memory phenotype of peripheral CD8 + T cells increases slightly after microwave ablation (MWA), the metabolism of peripheral CD8 + T cells remains unfavorable for memory phenotype. In mouse models, glycolysis inhibition by 2-deoxy-D-glucose (2DG) in combination with MWA results in long-term anti-tumor effect via enhancing differentiation of tumor-specific CD44 hi CD62L + CD8 + T CM cells. Enhancement of CD8 + T CM cell differentiation determined by Stat-1, is dependent on the tumor-draining lymph nodes (TDLN) but takes place in peripheral blood, with metabolic remodeling of CD8 + T cells lasting the entire course of the the combination therapy. Importantly, in-vitro glycolysis inhibition in peripheral CD8 + T cells of patients with breast or liver tumors having been treated with MWA thrice leads to their differentiation into CD8 + T CM cells. Our work thus offers a potential strategy to avoid tumor recurrence following MWA therapy and lays down the proof-of-principle for future clinical trials., (© 2024. The Author(s).)

Frequently, subcellular-targeted drugs tend to accumulate in lysosomes after cellular absorption, a process termed the lysosomal trap. This accumulation often interferes with the drug's ability to bind to its target, resulting in decreased efficiency. Existing methods for addressing lysosome-induced drug resistance mainly involve improving the structures of small molecules or enveloping drugs in nanomaterials. Nonetheless, these approaches can lead to changes in the drug structure or potentially trigger unexpected reactions within organisms. To address these issues, we introduced a strategy that involves inactivating the lysosome with the use of Ag nanoparticles (Cy3.5@Ag NPs). In this method, the Cy3.5@Ag NPs gradually accumulate inside lysosomes, leading to permeation of the lysosomal membrane and subsequent lysosomal inactivation. In addition, Cy3.5@Ag NPs also significantly affected the motility of lysosomes and induced the occurrence of lysosome passivation. Importantly, coincubating Cy3.5@Ag NPs with various subcellular-targeted drugs was found to significantly increase the efficiency of these treatments. Our strategy illustrates the potential of using lysosomal inactivation to enhance drug efficacy, providing a promising therapeutic strategy for cancer.

Objectives: This study aims to assess exposure to e-cigarette advertising across multiple marketing channels among U.S. youth and to examine whether racial/ethnic disparities exist in exposure to e-cigarette advertisements., Study Design: This is a cross-sectional study., Methods: Cross-sectional data were drawn from a longitudinal survey of participants recruited from two nationally representative panels (NORC's AmeriSpeak® and GfK's KnowledgePanel). A total of 2043 youth aged 13-17 completed the initial 2018 survey, and 2013 youth completed the follow-up survey in 2019 (including a replenishment sample of 690 youth). Outcome variables were self-reported e-cigarette advertisement exposure in the past three months through various sources, such as television, point of sale, and online/social media. Generalized estimating equation models were used to estimate the adjusted odds ratios (AOR) of the association between racial/ethnic identity and e-cigarette advertisement exposure., Results: The prevalence of reported exposure to e-cigarette advertisements through any channel was 79.8% (95% CI: 77.1-82.2) in 2018 and 74.9% (95% CI: 72.5-77.1) in 2019, respectively. Point of sale was the most common source of e-cigarette advertisement exposure in both years. Non-Hispanic Black and non-Hispanic Asian youth were more likely to report exposure to e-cigarette advertisements through television (AOR = 2.07, 95% CI: 1.44-2.99 and AOR = 2.11, 95% CI: 1.17-3.82, respectively) and online/social media (AOR = 1.61; 95% CI: 1.11-2.33 and AOR = 1.99, 95% CI: 1.10-3.59, respectively) channels compared with non-Hispanic White youth., Conclusions: A substantial proportion of U.S. youth reported exposure to e-cigarette advertising through a variety of marketing channels. Significant racial/ethnic disparities existed, with non-Hispanic Black and Asian youth reporting more marketing exposure than their non-Hispanic White counterparts., (Copyright © 2024 The Royal Society for Public Health. Published by Elsevier Ltd. All rights reserved.)

Mycoplasma pneumoniae causes respiratory tract infections, affecting both children and adults, with varying degrees of severity ranging from mild to life-threatening. In recent years, a new class of regulatory RNAs called long non-coding RNAs (lncRNAs) has been discovered to play crucial roles in regulating gene expression in the host. Research on lncRNAs has greatly expanded our understanding of cellular functions involving RNAs, and it has significantly increased the range of functions of lncRNAs. In lung cancer, transcripts associated with lncRNAs have been identified as regulators of airway and lung inflammation in a process involving protein complexes. An excessive immune response and antibacterial immunity are closely linked to the pathogenesis of M. pneumoniae . The relationship between lncRNAs and M. pneumoniae infection largely involves lncRNAs that participate in antibacterial immunity. This comprehensive review aimed to examine the dysregulation of lncRNAs during M. pneumoniae infection, highlighting the latest advancements in our understanding of the biological functions and molecular mechanisms of lncRNAs in the context of M. pneumoniae infection and indicating avenues for investigating lncRNAs-related therapeutic targets., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Yang, Zhou, Su, Zhang, Chen, Liu and Ling.)

Introduction: Depression is a complex psychiatric disorder with substantial societal impact. While current antidepressants offer moderate efficacy, their adverse effects and limited understanding of depression's pathophysiology hinder the development of more effective treatments. Amidst this complexity, the role of neuroinflammation, a recognized but poorly understood associate of depression, has gained increasing attention. This study investigates hydroxytyrosol (HT), an olive-derived phenolic antioxidant, for its antidepressant and anti-neuroinflammatory properties based on mitochondrial protection. Methods: In vitro studies on neuronal injury models, the protective effect of HT on mitochondrial ultrastructure from inflammatory damage was investigated in combination with high-resolution imaging of mitochondrial substructures. In animal models, depressive-like behaviors of chronic restraint stress (CRS) mice and chronic unpredictable mild stress (CUMS) rats were examined to investigate the alleviating effects of HT. Targeted metabolomics and RNA-Seq in CUMS rats were used to analyze the potential antidepressant pathways of HT. Results: HT protected mitochondrial ultrastructure from inflammatory damage, thus exerting neuroprotective effects in neuronal injury models. Moreover, HT reduced depressive-like behaviors in mice and rats exposed to CRS and CUMS, respectively. HT's influence in the CRS model included alleviating hippocampal neuronal damage and modulating cytokine production, mitochondrial dysfunction, and brain-derived neurotrophic factor (BDNF) signaling. Targeted metabolomics in CUMS rats revealed HT's effect on neurotransmitter levels and tryptophan-kynurenine metabolism. RNA-Seq data underscored HT's antidepressant mechanism through the BDNF/TrkB signaling pathways, key in nerve fiber functions, myelin formation, microglial differentiation, and neural regeneration. Discussion: The findings underscore HT's potential as an anti-neuroinflammatory treatment for depression, shedding light on its antidepressant effects and its relevance in nutritional psychiatry. Further investigations are warranted to comprehensively delineate its mechanisms and optimize its clinical application in depression treatment., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest. The author(s) declared that they were an editorial board member of Frontiers, at the time of submission. This had no impact on the peer review process and the final decision., (Copyright © 2024 Li, Shao, Sun, Guo, Zhang, Zeng, Fang, Liu, Wang, Liu and Ling.)

Background: Acute superior mesenteric venous thrombosis (ASMVT) decreases junction-associated protein expression and intestinal epithelial cell numbers, leading to intestinal epithelial barrier disruption. Pyroptosis has also recently been found to be one of the important causes of mucosal barrier defects. However, the role and mechanism of pyroptosis in ASMVT are not fully understood., Methods: Differentially expressed microRNAs (miRNAs) in the intestinal tissues of ASMVT mice were detected by transcriptome sequencing (RNA-Seq). Gene expression levels were determined by RNA extraction and reverse transcription-quantitative PCR (RT-qPCR). Western blot and immunofluorescence staining analysis were used to analyze protein expression. H&E staining was used to observe the intestinal tissue structure. Cell Counting Kit-8 (CCK-8) and fluorescein isothiocyanate/propidine iodide (FITC/PI) were used to detect cell viability and apoptosis, respectively. Dual-luciferase reporter assays prove that miR-138-5p targets NLRP3., Results: miR-138-5p expression was downregulated in ASMVT-induced intestinal tissues. Inhibition of miR-138-5p promoted NLRP3-related pyroptosis and destroyed tight junctions between IEC-6 cells, ameliorating ASMVT injury. miR-138-5p targeted to downregulate NLRP3. Knockdown of NLRP3 reversed the inhibition of proliferation, apoptosis, and pyroptosis and the decrease in tight junction proteins caused by suppression of miR-138-5p; however, this effect was later inhibited by overexpressing HMGB1. miR-138-5p inhibited pyroptosis, promoted intestinal epithelial tight junctions and alleviated ASMVT injury-induced intestinal barrier disruption via the NLRP3/HMGB1 axis., Competing Interests: The authors declare there are no competing interests., (©2024 Li et al.)

Introduction: Abiotic stress during growth readily reduces cotton crop yield. The different survival tactics of plants include the activation of numerous stress response genes, such as BREVIS RADIX ( BRX )., Methods: In this study, the BRX gene family of upland cotton was identified and analyzed by bioinformatics method, three salt-tolerant and cold-resistant GhBRX genes were screened. The expression of GhBRX.1 , GhBRX.2 and GhBRXL4.3 in upland cotton was silenced by virus-induced gene silencing (VIGS) technique. The physiological and biochemical indexes of plants and the expression of related stress-response genes were detected before and after gene silencing. The effects of GhBRX.1 , GhBRX.2 and GhBRXL4.3 on salt and cold resistance of upland cotton were further verified., Results and Discussion: We discovered 12, 6, and 6 BRX genes in Gossypium hirsutum , Gossypium raimondii and Gossypium arboreum , respectively. Chromosomal localization indicated that the retention and loss of GhBRX genes on homologous chromosomes did not have a clear preference for the subgenomes. Collinearity analysis suggested that segmental duplications were the main force for BRX gene amplification. The upland cotton genes GhBRX.1 , GhBRX.2 and GhBRXL4.3 are highly expressed in roots, and GhBRXL4.3 is also strongly expressed in the pistil. Transcriptome data and qRT‒PCR validation showed that abiotic stress strongly induced GhBRX.1 , GhBRX.2 and GhBRXL4.3 . Under salt stress and low-temperature stress conditions, the activities of superoxide dismutase (SOD), peroxidase (POD) and catalase (CAT) and the content of soluble sugar and chlorophyll decreased in GhBRX.1- , GhBRX.2- and GhBRXL4.3 -silenced cotton plants compared with those in the control (TRV: 00). Moreover, GhBRX.1 -, GhBRX.2 - and GhBRXL4.3 -silenced cotton plants exhibited greater malondialdehyde (MDA) levels than did the control plants. Moreover, the expression of stress marker genes ( GhSOS1 , GhSOS2 , GhNHX1 , GhCIPK6 , GhBIN2 , GhSnRK2.6 , GhHDT4D , GhCBF1 and GhPP2C ) decreased significantly in the three target genes of silenced plants following exposure to stress. These results imply that the GhBRX.1 , GhBRX.2 and GhBRXL4.3 genes may be regulators of salt stress and low-temperature stress responses in upland cotton., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as potential conflicts of interest., (Copyright © 2024 Wei, Ju, Zhang, Ling, Luo, Li, Xu, Su, Zhang and Wang.)

Dental fluorosis, resulting from long-term environmental exposure to fluoride, is prevalent among diverse populations worldwide. Severe fluorosis not only compromises the aesthetic appeal of teeth but also impairs their functionality. This study aims to investigate the oral microbiome in dental fluorosis and the health individuals of adolescents living in the endemic fluorosis area of Guizhou, China through full-length 16S rDNA sequencing. Fourty-six individuals meet the sampling criteria, and we divided these samples into the following groups: a healthy group ( H = 23) and a dental fluorosis group ( F = 23), and two subgroups of Miao ethnicity: a healthy Miao group (Hm = 13) and a dental fluorosis Miao group (Fm = 15). A total of 660,389 high-quality sequences were obtained, and 12,007 Amplicon Sequence Variants (ASVs) were identified, revealing significant variations in oral microbiome between Fm and Hm groups. The composition of oral microbiota was similar between the H and F groups. At the genus level, Pseudopropionibacterium and at the species level, Streptococcus oralis&#95;subsp.dentisani&#95;clade&#95;058 were less abundant in group F than in group H ( P < 0.05). Further analysis revealed that the abundance of Capnocytophaga gingivalis and Kingella denitrificans was significantly lower in Fm fluorosis patients than in the Hm group ( P < 0.05). Based on the LEfSe analysis, the potential core biomarkers in the oral of Fm fluorosis patients were identified at different taxonomic levels, ranging from phylum to species. These include Gammaproteobacteria , Prevotella sp&#95;HMT&#95;304 , Gemella sanguinis , and Gracilibacteria&#95; ( GN02 ). Network analysis revealed that the microbiota in the fluorosis group exhibited more complex interactions with each other than the healthy group. Notably, within the Hm group, the potential biomarkers Capnocytophaga gingivalis and Kingella denitrificans exhibited a positive correlation. Finally, we employed PICRUSt2 analysis to explore the abundance clustering of the top 30 functional units in each sample, and we found that the metabolic pathway compositions of the four groups were similar. In summary, our findings suggest that the microbial composition of plaque in Hm patients with dental fluorosis is significantly altered, and we identified the potential marker microorganisms that contribute to these changes., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Luo, Shao, Hong, Zhang, Zhou, Zhou, Rao, Zhao, Dong, Zhu, Ling, Cui, Guan, Luo, He, Qi, Liao and Hong.)

Annexins (ANNs) are a structurally conserved protein family present in almost all plants. In the present study, 27 GhANNs were identified in cotton and were unevenly distributed across 14 chromosomes. Transcriptome data and RT-qPCR results revealed that multiple GhANNs respond to at least two abiotic stresses. Similarly, the expression levels of GhANN4 and GhANN11 were significantly upregulated under heat, cold, and drought stress. Using virus-induced gene silencing (VIGS), functional characterization of GhANN4 and GhANN11 revealed that, compared with those of the controls, the leaf wilting of GhANN4 -silenced plants was more obvious, and the activities of catalase (CAT), peroxidase (POD), and superoxide dismutase (SOD) were lower under NaCl and PEG stress. Moreover, the expression of stress marker genes ( GhCBL3 , GhDREB2A , GhDREB2C , GhPP2C , GhRD20-2 , GhCIPK6 , GhNHX1 , GhRD20-1 , GhSOS1 , GhSOS2 and GhSnRK2.6 ) was significantly downregulated in GhANN4 -silenced plants after stress. Under cold stress, the growth of the GHANN11 -silenced plants was significantly weaker than that of the control plants, and the activities of POD, SOD, and CAT were also lower. However, compared with those of the control, the elasticity and orthostatic activity of the GhANN11 -silenced plants were greater; the POD, SOD, and CAT activities were higher; and the GhDREB2C , GhHSP , and GhSOS2 expression levels were greater under heat stress. These results suggest that different GhANN family members respond differently to different types of abiotic stress.

Aim: To assess the efficacy of artificial pancreas systems (APS) use among pregnant women with type 1 diabetes mellitus (T1DM) by conducting a meta-analysis., Methods: We searched five databases, including EMBASE, Web of Science, PubMed, Cochrane Library and SCOPUS, for literature on APS use among pregnant women with T1DM before October 9, 2023. The primary endpoint was 24-hour time in range (TIR; 3.5-7.8 mmol/L). Secondary endpoints included glycaemic metrics for 24-hour (mean blood glucose [MBG], time above range [TAR], time below range [TBR]), and overnight TIR and TBR., Results: We identified four randomized controlled trials involving 164 participants; one study with 16 participants focused on overnight APS use, and the other three focused on 24-hour APS use. Compared with standard care, APS exhibited a favourable effect on 24-hour TIR (standard mean difference [SMD] = 0.53, 95% confidence interval [CI] 0.25, 0.80, P < 0.001), overnight TIR (SMD = 0.67, 95% CI 0.39, 0.95, P < 0.001), and overnight TBR (<3.5 mmol/L; SMD = -0.49, 95% CI -0.77, -0.21 P < 0.001), while there was no significant difference in 24-hour TAR, 24-hour TBR, or MBG between the two groups. We further conducted subgroup analyses after removing the trial focused on overnight APS use and showed that 24-hour APS use reduced not only the 24-hour TIR (SMD = 0.41, 95% CI 0.12, 0.71; P = 0.007) but also the 24-hour TBR (<2.8 mmol/L; SMD = -0.77, 95% CI -1.32, -0.23, P = 0.006)., Conclusion: Our findings suggest that APS might improve 24-hour TIR and overnight glycaemic control, and 24-hour APS use also significantly reduced 24-hour TBR (2.8 mmol/L) among pregnant women with T1DM., (© 2023 John Wiley & Sons Ltd.)

Phosphatidylinositol 4-phosphate 5-kinases (PIP5Ks), essential enzymes in the phosphatidylinositol signaling pathway, are crucial for the abiotic stress responses and the overall growth and development of plants. However, the GhPIP5Ks had not been systematically studied, and their function in upland cotton was unknown. This study identified a total of 28 GhPIP5Ks , and determined their chromosomal locations, gene structures, protein motifs and cis-acting elements via bioinformatics analysis. A quantitative real-time PCR (qRT‒PCR) analysis showed that most GhPIP5Ks were upregulated under different stresses. A virus-induced gene silencing (VIGS) assay indicated that the superoxide dismutase (SOD), peroxidase (POD), and catalase (CAT) activities were significantly decreased, while malondialdehyde (MDA) content were significantly increased in GhPIP5K2 - and GhPIP5K22 -silenced upland cotton plants under abiotic stress. Furthermore, the expression of the stress marker genes GhHSFB2A , GhHSFB2B , GhDREB2A , GhDREB2C , GhRD20-1 , GhRD29A , GhBIN2 , GhCBL3 , GhNHX1 , GhPP2C , GhCBF1 , GhSnRK2.6 and GhCIPK6 was significantly decreased in the silenced plants after exposure to stress. These results revealed that the silencing of GhPIP5K2 and GhPIP5K22 weakened the tolerance to abiotic stresses. These discoveries provide a foundation for further inquiry into the actions of the GhPIP5K gene family in regulating the response and resistance mechanisms of cotton to abiotic stresses.

We report a draft genome assembly of Trichoderma longibrachiatum isolate GEV 3550, obtained from Florida, United States of America., Competing Interests: The authors declare no conflict of interest.

Natural compounds, such as carotenoids, flavonoids, anthocyanins, or terpenoids, are physiologically active components found in plants (pigments), often known as phytochemicals or phytonutrients. The in vitro cytotoxic and anticolon cancer effects of biologically bavachin, bavachinin, artepillin C, and aromadendrin compounds against SW48, SNU-C1, COLO 205, RKO, LS411N, and SW1417 cancer cell lines were assessed. Results of enzymes and antibacterial, antifungal were in level of micromolar that is good impacts. These natural compounds may be antidiabetic, anticancer, and antibacterial candidates for drug design. IC 50 results were obtained between 14-19 and 5-119 µM for α-amylase and α-glucosidase, respectively. Good inhibitor Bavachinin was detected for both enzymes (IC 50 for α-amylase: 14.37 µM and IC 50 for α-glucosidase: 5.27 µM). The chemical activities of aromadendrin, artepillin C, bavachin, and bavachinin against pancreatic α-amylase and α-glucosidase were assessed by conducting the molecular docking study. The chemical activities of aromadendrin, artepillin C, bavachin, and bavachinin against some of the expressed surface receptor proteins (CD44, CD47, CXCR4, EGFR, folate receptor, HER2, and endothelin receptor) in the mentioned cell lines were investigated using the molecular docking calculations. The results illustrated the atomic-level properties and potential interactions. These chemicals have high binding affinities to the enzymes and proteins, according to the docking scores. In addition, the compounds formed strong contacts with the enzymes and receptors. Thus, these compounds could be potential inhibitors for enzymes and cancer cells., (© 2023 Wiley Periodicals LLC.)

Background and Objective: Connexin 43 (Cx43) is the main gap junction (GJ) protein and hemichannel protein in bone tissue. It is involved in the formation of hemichannels and GJs and establishes channels that can communicate directly to exchange substances and signals, affecting the structure and function of osteocytes. CX43 is very important for the normal development of bone tissue and the establishment and balance of bone reconstruction. However, the molecular mechanisms by which CX43 regulates osteoblast function and homeostasis have been less well studied, and this article provides a review of research in this area., Methods: We searched the PubMed, EMBASE, Cochrane Library, and Web of Science databases for studies published up to June 2023 using the keywords Connexin 43/Cx43 and Osteocytes. Screening of literatures according to inclusion and exclusion guidelines and summarized the results., Key Content and Findings: Osteocytes, osteoblasts, and osteoclasts all express Cx43 and form an overall network through the interaction between GJs. Cx43 is not only involved in the mechanical response of bone tissue but also in the regulation of signal transduction, which could provide new molecular markers and novel targets for the treatment of certain bone diseases., Conclusions: Cx43 is expressed in osteoblasts, osteoclasts, and osteoclasts and plays an important role in regulating the function, signal transduction, and mechanotransduction of osteocytes. This review offers a new contribution to the literature by summarizing the relationship between Cx43, a key protein of bone tissue, and osteoblasts., Competing Interests: Conflicts of Interest: All authors have completed the ICMJE uniform disclosure form (available at https://aoj.amegroups.com/article/view/10.21037/aoj-23-65/coif). The authors have no conflicts of interest to declare., (2024 Annals of Joint. All rights reserved.)

Recently, various nanomaterials have drawn increasing attention for enhanced tumor therapy. However, a lack of tumor uptake and insufficient generation of cytotoxic agents have largely limited the antitumor efficacy in vivo. Herein, a multifunctional nanoplatform (IL@CPPor(CO)) was constructed with pH-responsive copper peroxide nanoparticles (CPNP) that are capable of self-supplying H 2 O 2 , a radical-sensitive carbonic oxide (CO) donor (Fe 3 (CO) 12 ), photosensitizer Iridium(III) meso-tetra ( N -methyl-4-pyridyl)porphyrin pentachloride (IrPor), and ionic liquid (IL) for enhanced oncotherapy. Under acidic conditions, the CPNP could decompose to release H 2 O 2 and Cu 2+ . The concomitant generation of H 2 O 2 could efficiently trigger Fe 3 (CO) 12 to release the CO in situ. On the other hand, Cu 2+ possesses both glutathione depletion and Fenton-like properties. In addition, IrPor has both peroxidase-like activity and photosensitizer properties to produce reactive oxygen species (ROS) in tumors. The released ROS could trigger the rapid intracellular release of CO. More importantly, released CO and ROS could promote cell apoptosis and improve the therapeutic efficacy. Moreover, due to the pH-dependent ROS generation property, the IL@CPPor(CO) exhibited high tumor accumulation, low toxicity, and good biocompatibility, which enabled effective tumor growth inhibition with minimal side effects in vivo. This work provides a novel multifunctional nanoplatform that combined photodynamic therapy with CDT and CO to improve therapeutic efficacy.

Background: Phalaenopsis is an important ornamental plant that has great economic value in the world flower market as one of the most popular flower resources., Objective: To investigate the flower colour formation of Phalaenopsis at the transcription level, the genes involved in flower color formation were identified from RNA-seq in this study., Methods: In this study, white and purple petals of Phalaenopsis were collected and analyzed to obtained (1) differential expression genes (DEGs) between white and purple flower color and (2) the association between single nucleotide polymorphisms (SNP) mutations and DEGs at the transcriptome level., Results: The results indicated that a total of 1,175 DEGs were identified, and 718 and 457 of them were up- and down-regulated genes, respectively. Gene Ontology and pathway enrichment showed that the biosynthesis of the secondary metabolites pathway was key to color formation, and the expression of 12 crucial genes (C4H, CCoAOMT, F3'H, UA3'5'GT, PAL, 4CL, CCR, CAD, CALDH, bglx, SGTase, and E1.11.17) that are involved in the regulation of flower color in Phalaenopsis., Conclusion: This study reported the association between the SNP mutations and DEGs for color formation at RNA level, and provides a new insight to further investigate the gene expression and its relationship with genetic variants from RNA-seq data in other species., (© 2023. The Author(s) under exclusive licence to The Genetics Society of Korea.)

Low acrosin activity (LAA) is associated with sperm function anomaly and poor outcomes of in vitro fertilization. In this study, we confirm that 993 semen samples with LAA had a reduced sperm motility and low in vitro fertilization rate in comparison with 1332 normal controls (NC). Proteomic comparison between 11 LAA and 11 NC sperm samples identified 35 upregulated and 99 downregulated proteins in the LAA group. Indeed, proteomic data showed that acrosome enzymes Spam1 and Acrosin were among the downregulated proteins in the LAA group, which was validated by quantitative PCR and immunefluorescent staining of sperm cells. The KEEG pathway analysis revealed a deficiency of GSH and Gln biosynthesis in LAA sperm cells. Immunofluorescent staining of sperms and quantitative PCR verified downregulation of GLUL and GCLC, the key enzymes for GSH and Gln biosynthesis. Moreover, the results of ELISA assay confirmed low levels of GSH and Gln in LAA sperm cells. Mechanistic studies showed that addition of 10 mM H 2 O 2 to semen samples led to a significant reduction of acrosin activity and sperm motility, most possibly by triggering premature acrosome release. In contrast, the presence of 20 mM GSH blocked the oxidative effects of H 2 O 2 . Since GSH counteracts the oxidative stress and Gln participates in TCA cycling, their deficiency may affect the redox balance as well as energy production of sperm cells. These findings shed new light on the pathological mechanisms of infertility associated with LAA. Male infertility patients could benefit from GSH supplement by improvement of acrosin activity and other sperm functions., (© 2023. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

The design and development of electrospun nanofibrous yarns (ENYs) have attracted intensive attentions in the fields of biomedical textiles and tissue engineering, but the inferior fiber arrangement structure, low yarn eveness, and poor tensile properties of currently-obtained ENYs has been troubled for a long time. In this study, a series of innovative strategies which combined a modified electrospinning method with some traditional textile processes like hot stretching, twisting, and plying, were designed and implemented to generate poly (L-lactic-acid) (PLLA) ENYs with adjustable morphology, structure, and tensile properties. PLLA ENYs made from bead-free and uniform PLLA nanofibers were fabricated by our modified electrospinning method, but the as-spun PLLA ENYs exhibited relatively lower fiber alignment degree and tensile properties. A hot stretching technique was explored to process the primary PLLA ENYs to improve the fiber alignment and crystallinity, resulting in a 779.7% increasement for ultimate stress and a 470.4% enhancement for Young's modulus, respectively. Then, the twisting post-treatment was applied to process as-stretched PLLA ENYs, and the tensile performances of as-twisted ENYs was found to present a trend of first increasing and then decreasing with the increasing of twisting degree. Finally, the PLLA threads made from different numbers of as-stretched PLLA ENYs were also manufactured with a traditional plying process, demonstrating the feasibility of further improving the yarn diameter and tensile properties. In all, this study reported a simple and cost-effective technique roadmap which could generate high performance PLLA nanofiber-constructed yarns or threads with controllable structures like highly aligned fiber orientation, twisted structure, and plied structure., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier Ltd. All rights reserved.)

Introduction: Tobacco harm reduction (THR) discourse has been divisive for the tobacco control community, partially because it sometimes aligns public health and tobacco industry interests. Industry funding is contentious as it influences study outcomes, and is not always disclosed in scientific publications. This study examines the role of disclosed and undisclosed industry support on THR publications via social network analysis., Methods: We reviewed 826 English-language manuscripts (1992-2016) to determine disclosed and undisclosed industry (pharmaceutical, tobacco, and e-cigarette) and non-industry (including government) support received by 1405 authors. We used social network analysis to identify the most influential authors in THR discourse by assessing the number of their collaborators on publications, the frequency of connecting other authors in the network, and tendency to form groups based on the presence of sponsorship disclosures, sources of funding, and THR stance., Results: About 284 (20%) out of 1405 authors were supported by industry. Industry-sponsored authors were more central and influential in the network: with twice as many publications (Median = 4), 1.25 as many collaborators on publications (Median = 5), and higher likelihood of connecting other authors and thus having more influence in the network, compared to non-industry-sponsored authors. E-cigarette industry-sponsored authors had the strongest association with undisclosed industry support., Conclusions: Authors with industry support exerted a stronger influence on the THR scientific discourse than non-industry-supported authors. Journals should continue adhering to strict policies requiring conflicts of interest disclosures. An increase in public health spending on tobacco control research may be necessary to achieve funding parity., (© The Author(s) 2022. Published by Oxford University Press on behalf of the Society for Research on Nicotine and Tobacco. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Background: Reducing tobacco depictions in media has significant health benefits because tobacco placements in films normalise tobacco use and are linked to youth initiation. Cannabis depictions may have similar effects. Placing restrictions on film incentives has been suggested; however, it remains an unexplored strategy for reducing tobacco depictions. We investigated whether states and localities that offer film incentives have established funding restrictions to deter tobacco or cannabis depictions., Methods: We conducted a descriptive cross-sectional survey of official state and local government websites in the USA that listed film incentives. We coded policy level (ie, state, local), incentive type (ie, tax credit, rebate), incentive amounts, programme qualifiers and presence and characteristics of tobacco and cannabis restrictions., Results: Quantifiable tax incentives at the state level offered over $1.6 billion to producers. Among 50 US states and Washington, DC, 39 provided film incentives: 4 restricted tobacco and 0 restricted cannabis. Among the 238 local film offices we identified, 24 offered incentives; 5 restricted tobacco and 1 restricted cannabis. All the incentive restrictions excluded tobacco or cannabis purchases from reimbursement; there were no penalties for tobacco or cannabis depictions., Conclusions: Film incentives are large financial commitments that may undermine public health since they allow depictions of tobacco and cannabis that contribute to initiation and use. Few states or localities limit film incentives related to tobacco or cannabis, and the existing restrictions are unlikely to deter depictions or product placement. Restrictions on incentives are an underused tool for deterring tobacco and cannabis depictions in film., Competing Interests: Competing interests: None declared., (© Author(s) (or their employer(s)) 2023. No commercial re-use. See rights and permissions. Published by BMJ.)

Neovascularization is crucial to the occurrence and progression of tumors, and the development of antiangiogenic drugs has essential theoretical value and clinical significance. However, antiangiogenesis therapy alone cannot meet the needs of tumor therapy. Meanwhile, polysaccharides are ideal drug carriers with promising applications in drug modification and delivery. In this research, we developed a novel redox and acid sensitive nanodrug (CDDP-CS-Cys-EA, CCEA) composed of chondroitin sulfate (CS), antiangiogenic peptide (endostatin2-alft1, EA) and chemotherapeutic drug (cisplatin, CDDP). CCEA exhibited redox and acid responsiveness, better blood hemocompatibility (hemolysis rate < 5 %), the ability to target tumors (CD44-mediated endocytosis), and strong antiangiogenesis and antitumor characteristics in vitro. Moreover, CCEA showed excellent antitumor activity and low toxicity in B16 xenograft mice. It also has been confirmed that CCEA induced tumor cell apoptosis through promoting the expression of Bax, suppressing the expression of Bcl-2, decreasing mitochondrial membrane potential, releasing cytochrome C (Cyto C), and enhancing the activities of Caspase 9 and Caspase 3. The results of this paper provided a theoretical basis and insight for the development of antitumor drugs., Competing Interests: Declaration of competing interest The authors have no conflicts of interest to declare., (Copyright © 2023. Published by Elsevier Ltd.)

The characteristics of acoustic emission signals generated in the process of rock deformation and fission contain rich information on internal rock damage. The use of acoustic emissions monitoring technology can analyze and identify the precursor information of rock failure. At present, in the field of acoustic emissions monitoring and the early warning of rock fracture disasters, there is no real-time identification method for a disaster precursor characteristic signal. It is easy to lose information by analyzing the characteristic parameters of traditional acoustic emissions to find signals that serve as precursors to disasters, and analysis has mostly been based on post-analysis, which leads to poor real-time recognition of disaster precursor characteristics and low application levels in the engineering field. Based on this, this paper regards the acoustic emissions signal of rock fracture as a kind of speech signal generated by rock fracture uses this idea of speech recognition for reference alongside spectral analysis (STFT) and Mel frequency analysis to realize the feature extraction of acoustic emissions from rock fracture. In deep learning, based on the VGG16 convolutional neural network and AlexNet convolutional neural network, six intelligent real-time recognition models of rock fracture and key acoustic emission signals were constructed, and the network structure and loss function of traditional VGG16 were optimized. The experimental results show that these six deep-learning models can achieve the real-time intelligent recognition of key signals, and Mel, combined with the improved VGG16, achieved the best performance with 87.68% accuracy and 81.05% recall. Then, by comparing multiple groups of signal recognition models, Mel+VGG-FL proposed in this paper was verified as having a high recognition accuracy and certain recognition efficiency, performing the intelligent real-time recognition of key acoustic emission signals in the process of rock fracture more accurately, which can provide new ideas and methods for related research and the real-time intelligent recognition of rock fracture precursor characteristics.

Objective: Glucose management indicator (GMI) is a core metric derived from continuous glucose monitoring (CGM) and is widely used to evaluate glucose control in patients with diabetes. No study has explored the pregnancy-specific GMI. This study aimed to derive a best-fitting model to calculate GMI from mean blood glucose (MBG) obtained from CGM among pregnant women with type 1 diabetes mellitus (T1DM)., Methods: A total of 272 CGM data and corresponding laboratory HbA1c from 98 pregnant women with T1DM in the CARNATION study were analysed in this study. Continuous glucose monitoring data were collected to calculate MBG, time-in-range (TIR), and glycaemic variability parameters. The relationships between the MBG and HbA1c during pregnancy and postpartum were explored. Mix-effect regression analysis with polynomial terms and cross-validation method was conducted to investigate the best-fitting model to calculate GMI from MBG obtained by CGM., Results: The pregnant women had a mean age of 28.9 ± 3.8 years, with a diabetes duration of 8.8 ± 6.2 years and a mean body mass index (BMI) of 21.1 ± 2.5 kg/m 2 . The HbA1c levels were 6.1 ± 1.0% and 6.4 ± 1.0% during pregnancy and at postpartum (p = 0.024). The MBG levels were lower during pregnancy than those at postpartum (6.5 ± 1.1 mmol/L vs. 7.1 ± 1.5 mmol/L, p = 0.008). After adjusting the confounders of haemoglobin (Hb), BMI, trimesters, disease duration, mean amplitude of glycaemic excursions and CV%, we developed a pregnancy-specific GMI-MBG equation: GMI for pregnancy (%) = 0.84-0.28* [Trimester] + 0.08 * [ BMI in kg/m 2 ] + 0.01 * [Hb in g/mL] + 0.50 * [MBG in mmol/L]., Conclusions: We derived a pregnancy-specific GMI equation, which should be recommended for antenatal clinical care., Clinical Trial Registry Number: ChiCTR1900025955., (© 2023 John Wiley & Sons Ltd.)