Spliced X-box binding protein 1 couples the unfolded protein response to hexosamine biosynthetic pathway.

The hexosamine biosynthetic pathway (HBP) generates uridine diphosphate N-acetylglucosamine (UDP-GlcNAc) for glycan synthesis and O-linked GlcNAc (O-GlcNAc) protein modifications. Despite the established role of the HBP in metabolism and multiple diseases, regulation of the HBP remains largely undefined. Here, we show that spliced X-box binding protein 1 (Xbp1s), the most conserved signal transducer of the unfolded protein response (UPR), is a direct transcriptional activator of the HBP. We demonstrate that the UPR triggers HBP activation via Xbp1s-dependent transcription of genes coding for key, rate-limiting enzymes. We further establish that this previously unrecognized UPR-HBP axis is triggered in a variety of stress conditions. Finally, we demonstrate a physiologic role for the UPR-HBP axis by showing that acute stimulation of Xbp1s in heart by ischemia/reperfusion confers robust cardioprotection in part through induction of the HBP. Collectively, these studies reveal that Xbp1s couples the UPR to the HBP to protect cells under stress.

Posttranscriptional regulation of de novo lipogenesis by glucose-induced O-GlcNAcylation.

O-linked ß-N-acetyl glucosamine (O-GlcNAc) is attached to proteins under glucose-replete conditions; this posttranslational modification results in molecular and physiological changes that affect cell fate. Here we show that posttranslational modification of serine/arginine-rich protein kinase 2 (SRPK2) by O-GlcNAc regulates de novo lipogenesis by regulating pre-mRNA splicing. We found that O-GlcNAc transferase O-GlcNAcylated SRPK2 at a nuclear localization signal (NLS), which triggers binding of SRPK2 to importin α. Consequently, O-GlcNAcylated SRPK2 was imported into the nucleus, where it phosphorylated serine/arginine-rich proteins and promoted splicing of lipogenic pre-mRNAs. We determined that protein nuclear import by O-GlcNAcylation-dependent binding of cargo protein to importin α might be a general mechanism in cells. This work reveals a role of O-GlcNAc in posttranscriptional regulation of de novo lipogenesis, and our findings indicate that importin α is a "reader" of an O-GlcNAcylated NLS.

Direct stimulation of de novo nucleotide synthesis by O-GlcNAcylation.

O-linked ß-N-acetyl glucosamine (O-GlcNAc) is at the crossroads of cellular metabolism, including glucose and glutamine; its dysregulation leads to molecular and pathological alterations that cause diseases. Here we report that O-GlcNAc directly regulates de novo nucleotide synthesis and nicotinamide adenine dinucleotide (NAD) production upon abnormal metabolic states. Phosphoribosyl pyrophosphate synthetase 1 (PRPS1), the key enzyme of the de novo nucleotide synthesis pathway, is O-GlcNAcylated by O-GlcNAc transferase (OGT), which triggers PRPS1 hexamer formation and relieves nucleotide product-mediated feedback inhibition, thereby boosting PRPS1 activity. PRPS1 O-GlcNAcylation blocked AMPK binding and inhibited AMPK-mediated PRPS1 phosphorylation. OGT still regulates PRPS1 activity in AMPK-deficient cells. Elevated PRPS1 O-GlcNAcylation promotes tumorigenesis and confers resistance to chemoradiotherapy in lung cancer. Furthermore, Arts-syndrome-associated PRPS1 R196W mutant exhibits decreased PRPS1 O-GlcNAcylation and activity. Together, our findings establish a direct connection among O-GlcNAc signals, de novo nucleotide synthesis and human diseases, including cancer and Arts syndrome.

The anti-immune dengue subgenomic flaviviral RNA is present in vesicles in mosquito saliva and is associated with increased infectivity.

Mosquito transmission of dengue viruses to humans starts with infection of skin resident cells at the biting site. There is great interest in identifying transmission-enhancing factors in mosquito saliva in order to counteract them. Here we report the discovery of high levels of the anti-immune subgenomic flaviviral RNA (sfRNA) in dengue virus 2-infected mosquito saliva. We established that sfRNA is present in saliva using three different methods: northern blot, RT-qPCR and RNA sequencing. We next show that salivary sfRNA is protected in detergent-sensitive compartments, likely extracellular vesicles. In support of this hypothesis, we visualized viral RNAs in vesicles in mosquito saliva and noted a marked enrichment of signal from 3'UTR sequences, which is consistent with the presence of sfRNA. Furthermore, we show that incubation with mosquito saliva containing higher sfRNA levels results in higher virus infectivity in a human hepatoma cell line and human primary dermal fibroblasts. Transfection of 3'UTR RNA prior to DENV2 infection inhibited type I and III interferon induction and signaling, and enhanced viral replication. Therefore, we posit that sfRNA present in salivary extracellular vesicles is delivered to cells at the biting site to inhibit innate immunity and enhance dengue virus transmission.

Deciphering the roles of subcellular distribution and interactions involving the MEF2 binding region, the ankyrin repeat binding motif and the catalytic site of HDAC4 in Drosophila neuronal morphogenesis.

BACKGROUND: Dysregulation of nucleocytoplasmic shuttling of histone deacetylase 4 (HDAC4) is associated with several neurodevelopmental and neurodegenerative disorders. Consequently, understanding the roles of nuclear and cytoplasmic HDAC4 along with the mechanisms that regulate nuclear entry and exit is an area of concerted effort. Efficient nuclear entry is dependent on binding of the transcription factor MEF2, as mutations in the MEF2 binding region result in cytoplasmic accumulation of HDAC4. It is well established that nuclear exit and cytoplasmic retention are dependent on 14-3-3-binding, and mutations that affect binding are widely used to induce nuclear accumulation of HDAC4. While regulation of HDAC4 shuttling is clearly important, there is a gap in understanding of how the nuclear and cytoplasmic distribution of HDAC4 impacts its function. Furthermore, it is unclear whether other features of the protein including the catalytic site, the MEF2-binding region and/or the ankyrin repeat binding motif influence the distribution and/or activity of HDAC4 in neurons. Since HDAC4 functions are conserved in Drosophila, and increased nuclear accumulation of HDAC4 also results in impaired neurodevelopment, we used Drosophila as a genetic model for investigation of HDAC4 function. RESULTS: Here we have generated a series of mutants for functional dissection of HDAC4 via in-depth examination of the resulting subcellular distribution and nuclear aggregation, and correlate these with developmental phenotypes resulting from their expression in well-established models of neuronal morphogenesis of the Drosophila mushroom body and eye. We found that in the mushroom body, forced sequestration of HDAC4 in the nucleus or the cytoplasm resulted in defects in axon morphogenesis. The actions of HDAC4 that resulted in impaired development were dependent on the MEF2 binding region, modulated by the ankyrin repeat binding motif, and largely independent of an intact catalytic site. In contrast, disruption to eye development was largely independent of MEF2 binding but mutation of the catalytic site significantly reduced the phenotype, indicating that HDAC4 acts in a neuronal-subtype-specific manner. CONCLUSIONS: We found that the impairments to mushroom body and eye development resulting from nuclear accumulation of HDAC4 were exacerbated by mutation of the ankyrin repeat binding motif, whereas there was a differing requirement for the MEF2 binding site and an intact catalytic site. It will be of importance to determine the binding partners of HDAC4 in nuclear aggregates and in the cytoplasm of these tissues to further understand its mechanisms of action.

Zymograph profiling reveals a divergent evolution of sirtuin that may originate from class III enzymes.

Sirtuins are a group of NAD+-dependent deacylases that conserved in three domains of life and comprehensively involved in the regulation of gene transcription, chromosome segregation, RNA splicing, apoptosis, and aging. Previous studies in mammalian cells have revealed that sirtuins not only exist as multiple copies, but also show distinct deacylase activities in addition to deacetylation. However, the understanding of sirtuin zymographs in other organisms with respect to molecular evolution remains at an early stage. Here, we systematically analyze the sirtuin activities in representative species from archaea, bacteria, and eukaryotes, using both the HPLC assay and a 7-amino-4-methylcoumarin-based fluorogenic method. Global profiling suggests that the deacylase activities of sirtuins could be divided into three categories and reveals undifferentiated zymographs of class III sirtuins, especially for those from bacteria and archaea. Nevertheless, initial differentiation of enzymatic activity was also observed for the class III sirtuins at both paralog and ortholog levels. Further phylogenetic analyses support a divergent evolution of sirtuin that may originate from class III sirtuins. Together, this work demonstrates a comprehensive panorama of sirtuin zymographs and provides new insights into the cellular specific regulation and molecular evolution of sirtuins.

METTL16 promotes osteosarcoma progression by downregulating VPS33B in an m6 A-dependent manner.

N6-methyladenosine (m6 A) is one of the main epitranscriptomic modifications that accelerates the progression of malignant tumors by modifying RNA. Methyltransferase-like 16 (METTL16) is a newly identified methyltransferase that has been found to play an important oncogenic role in a few malignancies; however, its function in osteosarcoma (OS) remains unclear. In this study, METTL16 was found to be upregulated in OS tissues, and associated with poor prognosis in OS patients. Functionally, METTL16 substantially promoted OS cell proliferation, migration, and invasion in vitro and OS growth in vivo. Mechanistically, vacuolar protein sorting protein 33b (VPS33B) was identified as the downstream target of METTL16, which induced m6 A modification of VPS33B and impaired the stability of the VPS33B transcript, thereby degrading VPS33B. In addition, VPS33B was found to be downregulated in OS tissues, VPS33B knockdown markedly attenuated shMETTL16-mediated inhibition on OS progression. Finally, METTL16/VPS33B might facilitate OS progression through PI3K/AKT pathway. In summary, this study revealed an important role for the METTL16-mediated m6 A modification in OS progression, implying it as a promising target for OS treatment.

Elimination of CaMKIIδ Autophosphorylation by CRISPR-Cas9 Base Editing Improves Survival and Cardiac Function in Heart Failure in Mice.

BACKGROUND: Cardiovascular diseases are the main cause of worldwide morbidity and mortality, highlighting the need for new therapeutic strategies. Autophosphorylation and subsequent overactivation of the cardiac stress-responsive enzyme CaMKIIδ (Ca2+/calmodulin-dependent protein kinase IIδ) serves as a central driver of multiple cardiac disorders. METHODS: To develop a comprehensive therapy for heart failure, we used CRISPR-Cas9 adenine base editing to ablate the autophosphorylation site of CaMKIIδ. We generated mice harboring a phospho-resistant CaMKIIδ mutation in the germline and subjected these mice to severe transverse aortic constriction, a model for heart failure. Cardiac function, transcriptional changes, apoptosis, and fibrosis were assessed by echocardiography, RNA sequencing, terminal deoxynucleotidyl transferase dUTP nick end labeling staining, and standard histology, respectively. Specificity toward CaMKIIδ gene editing was assessed using deep amplicon sequencing. Cellular Ca2+ homeostasis was analyzed using epifluorescence microscopy in Fura-2-loaded cardiomyocytes. RESULTS: Within 2 weeks after severe transverse aortic constriction surgery, 65% of all wild-type mice died, and the surviving mice showed dramatically impaired cardiac function. In contrast to wild-type mice, CaMKIIδ phospho-resistant gene-edited mice showed a mortality rate of only 11% and exhibited substantially improved cardiac function after severe transverse aortic constriction. Moreover, CaMKIIδ phospho-resistant mice were protected from heart failure-related aberrant changes in cardiac gene expression, myocardial apoptosis, and subsequent fibrosis, which were observed in wild-type mice after severe transverse aortic constriction. On the basis of identical mouse and human genome sequences encoding the autophosphorylation site of CaMKIIδ, we deployed the same editing strategy to modify this pathogenic site in human induced pluripotent stem cells. It is notable that we detected a >2000-fold increased specificity for editing of CaMKIIδ compared with other CaMKII isoforms, which is an important safety feature. While wild-type cardiomyocytes showed impaired Ca2+ transients and an increased frequency of arrhythmias after chronic ß-adrenergic stress, CaMKIIδ-edited cardiomyocytes were protected from these adverse responses. CONCLUSIONS: Ablation of CaMKIIδ autophosphorylation by adenine base editing may offer a potential broad-based therapeutic concept for human cardiac disease.

Vortex-Enhanced Microfluidic Chip for Efficient Mixing and Particle Capturing Combining Acoustics with Inertia.

Vortex-based microfluidics has received significant attention for its unique characteristics of high efficiency, flexible control, and label-free properties for the past decades. Herein, we present a vortex-based acousto-inertial chip that allows both fluid and particle manipulation within a significantly wider flow range and lower excitation voltage. Composed of contraction-expansion array structures and vibrating microstructures combined with bubbles and sharp edges, such a configuration results in more vigorous vortical fluid motions. The overall improvement in device performance comes from the synergistic effect of acoustics and inertia, as well as the positive feedback loop formed by vibrating bubbles and sharp edges. We characterize flow patterns in the microchannels by fluorescence particle tracer experiments and uncover single- and double-vortex modes over a range of sample flow rates and excitation voltages. On this basis, the ability of rapid and efficient sample homogenization up to a flow rate of 200 µL/min under an excitation voltage of 15 Vpp is verified by a two-fluid fluorescence mixing experiment. Moreover, the recirculation motion of particles in microvortices is investigated by using a high-speed imaging system. We also quantitatively measure the particle velocity variation on the trajectory and illustrate the capturing mechanism, which results from the interaction of the microvortices, particle dynamics, and composite microstructure perturbations. Further utilizing the shear forces derived by microvortices, our acousto-inertial chip is demonstrated to lysis red blood cells (RBCs) in a continuous, reagent-free manner. The high controllability and multifunction of this technology allow for the development of multistep miniaturized "lab-on-chip" analytical systems, which could significantly broaden the application of microvortex technology in biological, chemical, and clinical applications.

Mining the interpretable prognostic features from pathological image of intrahepatic cholangiocarcinoma using multi-modal deep learning.

BACKGROUND: The advances in deep learning-based pathological image analysis have invoked tremendous insights into cancer prognostication. Still, lack of interpretability remains a significant barrier to clinical application. METHODS: We established an integrative prognostic neural network for intrahepatic cholangiocarcinoma (iCCA), towards a comprehensive evaluation of both architectural and fine-grained information from whole-slide images. Then, leveraging on multi-modal data, we conducted extensive interrogative approaches to the models, to extract and visualize the morphological features that most correlated with clinical outcome and underlying molecular alterations. RESULTS: The models were developed and optimized on 373 iCCA patients from our center and demonstrated consistent accuracy and robustness on both internal (n = 213) and external (n = 168) cohorts. The occlusion sensitivity map revealed that the distribution of tertiary lymphoid structures, the geometric traits of the invasive margin, the relative composition of tumor parenchyma and stroma, the extent of necrosis, the presence of the disseminated foci, and the tumor-adjacent micro-vessels were the determining architectural features that impacted on prognosis. Quantifiable morphological vector extracted by CellProfiler demonstrated that tumor nuclei from high-risk patients exhibited significant larger size, more distorted shape, with less prominent nuclear envelope and textural contrast. The multi-omics data (n = 187) further revealed key molecular alterations left morphological imprints that could be attended by the network, including glycolysis, hypoxia, apical junction, mTORC1 signaling, and immune infiltration. CONCLUSIONS: We proposed an interpretable deep-learning framework to gain insights into the biological behavior of iCCA. Most of the significant morphological prognosticators perceived by the network are comprehensible to human minds.

Roxadustat reduces left ventricular mass index compared to rHuEPO in haemodialysis patients in a randomized controlled trial.

BACKGROUND: Left ventricular hypertrophy (LVH) is highly prevalent in haemodialysis (HD) patients and is associated with an increased risk of death. Roxadustat and recombinant human erythropoietin (rHuEPO, abbreviated as EPO) are the main treatment strategies for renal anaemia in HD patients, but it has not been clear whether there is a difference in their effect on LVH. METHODS: In this multi-centre, prospective, randomized trial of 12-month duration, study participants were randomized in a 1:1 ratio to the roxadustat group or the EPO group. The doses of both treatment regimens were adjusted so that the patients had a haemoglobin level of 10.0-12.0 g per dL. The primary study endpoint was the change from baseline to 12 months in the left ventricular mass index (LVMI, g/m2) measured by echocardiography. RESULTS: In total, 114 patients were enrolled. The mean age was 50 years, and the median dialysis duration was 33 months. Sixty-one patients were men, and 24 were diabetic. LVMI decreased from 116.18 ± 27.84 to 110.70 ± 25.74 g/m2 in the roxadustat group. However, it increased from 109.35 ± 23.41 to 114.99 ± 28.46 g/m2 in the EPO group, with a significant difference in the change in LVMI between the two groups [-5.48 (-11.60 to 0.65) vs. 5.65 (0.74 to 10.55), p < 0.05]. Changes in left ventricular mass, end-diastolic volume and 6-min walk test seemed superior in the roxadustat group. There were no significant differences in other cardiac geometry, biochemical parameters and major adverse cardiovascular events between the two groups. CONCLUSIONS: Compared to EPO, roxadustat is more helpful in the regression of LVH in HD patients.

Research Progress on the Assembly of Large DNA Fragments.

Synthetic biology, a newly and rapidly developing interdisciplinary field, has demonstrated increasing potential for extensive applications in the wide areas of biomedicine, biofuels, and novel materials. DNA assembly is a key enabling technology of synthetic biology and a central point for realizing fully synthetic artificial life. While the assembly of small DNA fragments has been successfully commercialized, the assembly of large DNA fragments remains a challenge due to their high molecular weight and susceptibility to breakage. This article provides an overview of the development and current state of DNA assembly technology, with a focus on recent advancements in the assembly of large DNA fragments in Escherichia coli, Bacillus subtilis, and Saccharomyces cerevisiae. In particular, the methods and challenges associated with the assembly of large DNA fragment in different hosts are highlighted. The advancements in DNA assembly have the potential to facilitate the construction of customized genomes, giving us the ability to modify cellular functions and even create artificial life. It is also contributing to our ability to understand, predict, and manipulate living organisms.

WNT-inhibitory factor 1-mediated glycolysis protects photoreceptor cells in diabetic retinopathy.

BACKGROUND: In diabetic retinopathy (DR), hypoxia-inducible factor (HIF-1α) induces oxidative stress by upregulating glycolysis. This process leads to neurodegeneration, particularly photoreceptor cell damage, which further contributes to retinal microvascular deterioration. Further, the regulation of Wnt-inhibitory factor 1 (WIF1), a secreted Wnt signaling antagonist, has not been fully characterized in neurodegenerative eye diseases. We aimed to explore the impact of WIF1 on photoreceptor function within the context of DR. METHOD: Twelve-week-old C57BL/KsJ-db/db mice were intravitreally injected with WIF1 overexpression lentivirus. After 4 weeks, optical coherence tomography (OCT), transmission electron microscopy (TEM), H&E staining, and electroretinography (ERG) were used to assess the retinal tissue and function. The potential mechanism of action of WIF1 in photoreceptor cells was explored using single-cell RNA sequencing. Under high-glucose conditions, 661 W cells were used as an in vitro DR model. WIF1-mediated signaling pathway components were assessed using quantitative real-time PCR, immunostaining, and western blotting. RESULT: Typical diabetic manifestations were observed in db/db mice. Notably, the expression of WIF1 was decreased at the mRNA and protein levels. These pathological manifestations and visual function improved after WIF1 overexpression in db/db mice. TEM demonstrated that WIF1 restored damaged mitochondria, the Golgi apparatus, and photoreceptor outer segments. Moreover, ERG indicated the recovery of a-wave potential amplitude. Single-cell RNA sequencing and in vitro experiments suggested that WIF1 overexpression prevented the expression of glycolytic enzymes and lactate production by inhibiting the canonical Wnt signaling pathway, HIF-1α, and Glut1, thereby reducing retinal and cellular reactive oxygen species levels and maintaining 661 W cell viability. CONCLUSIONS: WIF1 exerts an inhibitory effect on the Wnt/ß-catenin-HIF-1α-Glut1 glycolytic pathway, thereby alleviating oxidative stress levels and mitigating pathological structural characteristics in retinal photoreceptor cells. This mechanism helps preserve the function of photoreceptor cells in DR and indicates that WIF1 holds promise as a potential therapeutic candidate for DR and other neurodegenerative ocular disorders.

Characterization of dengue virus 3'UTR RNA binding proteins in mosquitoes reveals that AeStaufen reduces subgenomic flaviviral RNA in saliva.

Dengue viruses (DENV) are expanding global pathogens that are transmitted through the bite of mosquitoes, mostly Aedes aegypti. As RNA viruses, DENV rely on RNA-binding proteins (RBPs) to complete their life cycle. Alternatively, RBPs can act as restriction factors that prevent DENV multiplication. While the importance of RBPs is well-supported in humans, there is a dearth of information about their influence on DENV transmission by mosquitoes. Such knowledge could be harnessed to design novel, effective interventions against DENV. Here, we successfully adapted RNA-affinity chromatography coupled with mass spectrometry-a technique initially developed in mammalian cells-to identify RBPs in Ae. aegypti cells. We identified fourteen RBPs interacting with DENV serotype 2 3'UTR, which is involved in the viral multiplication and produces subgenomic flaviviral RNA (sfRNA). We validated the RNA affinity results for two RBPs by confirming that AePur binds the 3'UTR, whereas AeStaufen interacts with both 3'UTR and sfRNA. Using in vivo functional evaluation, we determined that RBPs like AeRan, AeExoRNase, and AeRNase have pro-viral functions, whereas AeGTPase, AeAtu, and AePur have anti-viral functions in mosquitoes. Furthermore, we showed that human and mosquito Pur homologs have a shared affinity to DENV2 RNA, although the anti-viral effect is specific to the mosquito protein. Importantly, we revealed that AeStaufen mediates a reduction of gRNA and sfRNA copies in several mosquito tissues, including the salivary glands and that AeStaufen-mediated sfRNA reduction diminishes the concentration of transmission-enhancing sfRNA in saliva, thereby revealing AeStaufen's role in DENV transmission. By characterizing the first RBPs that associate with DENV2 3'UTR in mosquitoes, our study unravels new pro- and anti-viral targets for the design of novel therapeutic interventions as well as provides foundation for studying the role of RBPs in virus-vector interactions.

The mechanism by which cannabidiol (CBD) suppresses TNF-α secretion involves inappropriate localization of TNF-α converting enzyme (TACE).

Cannabidiol (CBD) is a phytocannabinoid derived from Cannabis sativa that exerts anti-inflammatory mechanisms. CBD is being examined for its putative effects on the neuroinflammatory disease, multiple sclerosis (MS). One of the major immune mediators that propagates MS and its mouse model experimental autoimmune encephalomyelitis (EAE) are macrophages. Macrophages can polarize into an inflammatory phenotype (M1) or an anti-inflammatory phenotype (M2a). Therefore, elucidating the impact on macrophage polarization with CBD pre-treatment is necessary to understand its anti-inflammatory mechanisms. To study this effect, murine macrophages (RAW 264.7) were pre-treated with CBD (10 µM) or vehicle (ethanol 0.1 %) and were either left untreated (naive; cell media only), or stimulated under M1 (IFN-γ + lipopolysaccharide, LPS) or M2a (IL-4) conditions for 24 hr. Cells were analyzed for macrophage polarization markers, and supernatants were analyzed for cytokines and chemokines. Immunofluorescence staining was performed on M1-polarized cells for the metalloprotease, tumor necrosis factor-α-converting enzyme (TACE), as this enzyme is responsible for the secretion of TNF-α. Overall results showed that CBD decreased several markers associated with the M1 phenotype while exhibiting less effects on the M2a phenotype. Significantly, under M1 conditions, CBD increased the percentage of intracellular and surface TNF-α but decreased secreted TNF-α. This phenomenon might be mediated by TACE as staining showed that CBD sequestered TACE intracellularly. CBD also prevented RelA nuclear translocation. These results suggest that CBD may exert its anti-inflammatory effects by reducing M1 polarization and decreasing TNF-α secretion via inappropriate localization of TACE and RelA.

Association Between Home Renovation and Sleeping Problems Among Children Aged 6-18 Years: A Nationwide Survey in China.

BACKGROUND: Although the indoor environment has been proposed to be associated with childhood sleep health, to our knowledge no study has investigated the association between home renovation and childhood sleep problems. METHODS: The study included 186,470 children aged 6-18 years from the National Chinese Children Health Study (2012-2018). We measured childhood sleeping problems via the Chinese version of the Sleep Disturbance Scale for Children (C-SDSC). Information on home renovation exposure within the recent 2 years was collected via parent report. We estimated associations between home renovation and various sleeping problems, defined using both continuous and categorized (binary) C-SDSC t-scores, using generalized mixed models. We fitted models with city as a random effect variable, and other covariates as fixed effects. RESULTS: Out of the overall participants, 89,732 (48%) were exposed to recent home renovations. Compared to the unexposed group, children exposed to home renovations had higher odds of total sleep disorder (odd ratios [OR] = 1.3; 95% confidence interval [CI] = 1.2, 1.4). Associations varied when we considered different types of home renovation materials. Children exposed to multiple types of home renovation had higher odds of sleeping problems. We observed similar findings when considering continuous C-SDSC t-scores. Additionally, sex and age of children modified the associations of home renovation exposure with some of the sleeping problem subtypes. CONCLUSIONS: We found that home renovation was associated with higher odds of having sleeping problems and that they varied when considering the type of renovation, cumulative exposure, sex, and age differences.

PmAGAMOUS recruits polycomb protein PmLHP1 to regulate single-pistil morphogenesis in Japanese apricot.

Japanese apricot (Prunus mume Sieb. et Zucc.) is a traditional fruit tree with a long history. Multiple pistils (MP) lead to the formation of multiple fruits, decreasing fruit quality and yield. In this study, the morphology of flowers was observed at 4 stages of pistil development: undifferentiated stage (S1), predifferentiation stage (S2), differentiation stage (S3), and late differentiation stage (S4). In S2 and S3, the expression of PmWUSCHEL (PmWUS) in the MP cultivar was significantly higher than that in the single-pistil (SP) cultivar, and the gene expression of its inhibitor, PmAGAMOUS (PmAG), also showed the same trend, indicating that other regulators participate in the regulation of PmWUS during this period. Chromatin immunoprecipitation-qPCR (ChIP-qPCR) showed that PmAG could bind to the promoter and the locus of PmWUS, and H3K27me3 repressive marks were also detected at these sites. The SP cultivar exhibited an elevated level of DNA methylation in the promoter region of PmWUS, which partially overlapped with the region of histone methylation. This suggests that the regulation of PmWUS involves both transcription factors and epigenetic modifications. Also, the gene expression of Japanese apricot LIKE HETEROCHROMATIN PROTEIN (PmLHP1), an epigenetic regulator, in MP was significantly lower than that in SP in S2 to 3, contrary to the trend in expression of PmWUS. Our results showed that PmAG recruited sufficient PmLHP1 to maintain the level of H3K27me3 on PmWUS during the S2 of pistil development. This recruitment of PmLHP1 by PmAG inhibits the expression of PmWUS at the precise time, leading to the formation of 1 normal pistil primordium.

Allelic variation of PmCBF03 contributes to the altitude and temperature adaptability in Japanese apricot (Prunus mume Sieb. et Zucc.).

Japanese apricot is an important subtropical deciduous fruit tree in China, widely distributed in different altitude areas. How does it adapt to the different temperature environments in these areas? In this study, we identified a low-temperature transcription factor PmCBF03 on chromosome 7 through adaptive analysis of populations at different altitudes, which has an early termination single nucleotide polymorphism mutation. There were two different types of variation, PmCBF03A type in high-altitude areas and PmCBF03T type in low-altitude areas. PmCBF03A gene increased the survival rate, Fv/Fm values, antioxidant enzyme activity, and expression levels of antioxidant enzyme genes, and reducing electrolyte leakage and accumulation of reactive oxygen species in transgenic Arabidopsis under low temperature and freezing stress. Simultaneously, PmCBF03A gene promoted the dormancy of transgenic Arabidopsis seeds than wild-type. Biochemical analysis demonstrated that PmCBF03A directly bound to the DRE/CRT element in the promoters of the PmCOR413, PmDAM6 and PmABI5 genes, promoting their transcription and enhanced the cold resistance and dormancy of the overexpressing PmCBF03A lines. While PmCBF03T gene is unable to bind to the promoters of PmDAM6 and PmABI5 genes, leading to early release of dormancy to adapt to the problem of insufficient chilling requirement in low-altitude areas.

Experiment study on focusing pattern prediction of particles in asymmetric contraction-expansion array channel.

Contraction-expansion array (CEA) microchannel is a typical structure applied on particle/cell manipulation. The prediction of the particle focusing pattern in CEA microchannel is worthwhile to be investigate deeply. Here, we demonstrated a virtual boundary method by flow field analysis and theoretical derivation. The calculating method of the virtual boundary location, related to the Reynolds number (Re) and the structure parameter RW, was proposed. Combining the approximate Poiseuille flow pattern based on the virtual boundary method with the simulation results of Dean flow, the main line pattern and the main/lateral lines pattern were predicted and validated in experiments. The transformation from the main line pattern to the main/lateral lines pattern can be facilitated by increasing Re, decreasing RW , and decreasing α. An empirical formula was derived to characterize the critical condition of the transformation. The virtual boundary method can provide a guidance for asymmetric CEA channel design and contribute to the widespread application of microfluidic particle focusing.

Differential inhibition of sildenafil and macitentan on saxagliptin metabolism.

The development of diabetes mellitus (DM) is generally accompanied by erectile dysfunction (ED) and pulmonary arterial hypertension (PAH), which increases the use of combination drug therapy and the risk of drug-drug interactions. Saxagliptin for the treatment of DM, sildenafil for the treatment of ED and PAH, and macitentan for the treatment of PAH are all substrates of CYP3A4, which indicates their potential involvement in drug-drug interactions. Therefore, we investigated potential pharmacokinetic interactions between saxagliptin and sildenafil/macitentan. We investigated this speculation both in vitro and in vivo, and explored the underlying mechanism using in vitro hepatic metabolic models and molecular docking assays. The results showed that sildenafil substantially inhibited the metabolism of saxagliptin by occupying the catalytic site of CYP3A4 in a competitive manner, leading to the alterations in the pharmacokinetic properties of saxagliptin in terms of increased maximum plasma concentration (Cmax), area under the plasma concentration-time curve from time 0 to 24 h (AUC(0-t)), area under the plasma concentration-time curve from time 0 extrapolated to infinite time (AUC(0-∞)), decreased clearance rate (CLz/F), and prolonged terminal half-life (t1/2). In contrast, a slight inhibition was observed in saxagliptin metabolism when concomitantly used with macitentan, as no pharmacokinetic parameters were altered, except for CLz/F. Thus, dosage adjustment of saxagliptin may be required in combination with sildenafil to achieve safe therapeutic plasma concentrations and reduce the risk of potential toxicity, but it is not necessary for co-administration with macitentan.