Exosomal miR-21-5p derived from endometrial stromal cells promotes angiogenesis by targeting TIMP3 in ovarian endometrial cysts.

Endometriosis is a multifactorial gynecological disease, with angiogenesis as a key hallmark. The role of exosomal microRNAs (miRNAs) in endometriosis is not well understood. This study investigates differentially expressed exosomal miRNAs linked to angiogenesis in endometriosis, clarifies their molecular mechanisms, and identifies potential targets. Primary endometrial stromal cells (ESCs) were cultured, and exosomes were extracted. In a co-culture system, ESC-derived exosomes were taken up by human umbilical vein endothelial cells (HUVECs). Endometriosis implant-ESC-derived exosomes (EI-EXOs) significantly promoted HUVEC proliferation, migration and tube formation compared to normal endometrium-exosomes (NE-EXOs), a finding consistent in vivo in mice. MiRNA sequencing and bioinformatics identified differentially expressed miR-21-5p from EI-EXOs, confirmed by RT-qPCR. The miR-21-5p inhibitor or GW4869 attenuated EI-EXO-induced HUVEC proliferation, migration, and tube formation. TIMP3 overexpression diminished the pro-angiogenic effect of EI-EXOs, which was reversed by adding EI-EXOs or upregulating miR-21-5p. These findings validate the crosstalk between ESCs and HUVECs mediated by exosomal miR-21-5p, and confirm the miR-21-5p-TIMP3 axis in promoting angiogenesis in endometriosis. KEY MESSAGES: ESC-derived exosomes were found to be taken up by recipient cells, i.e. HUVECs. Functionally, endometriosis implant-ESC-derived exosomes (EI-EXOs) could significantly promote the proliferation, migration and tube formation of HUVECs compared to normal endometrium-exosomes (NE-EXOs). Through miRNA sequencing and bioinformatics analysis, differentially expressed miR-21-5p released by EI-EXOs was chosen, as confirmed by qRT-PCR. miR-21-5p inhibitor or GW4869 was found to attenuate the proliferation, migration, and tube formation of HUVECs induced by EI-EXOs. In turn, TIMP3 overexpression diminished the pro-angiogenic effect of EI-EXOs, and this angiogenic phenotype was reversed once EI-EXOs were added or miR-21-5p was upregulated.

Network-based transcranial direct current stimulation enhances attention function in healthy young adults: a preliminary study.

Purpose: Attention, a complex cognitive process, is linked to the functional activities of the brain's dorsal attention network (DAN) and default network (DN). This study aimed to investigate the feasibility, safety, and blinding efficacy of a transcranial direct current stimulation (tDCS) paradigm designed to increase the excitability of the DAN while inhibiting the DN (DAN+/DN-tDCS) on attention function in healthy young adults. Methods: In this randomized controlled experiment, participants were assigned to either the DAN+/DN-tDCS group or the sham group. A single intervention session was conducted at a total intensity of 4 mA for 20 min. Participants completed the Attention Network Test (ANT) immediately before and after stimulation. Blinding efficacy and adverse effects were assessed post-stimulation. Results: Forty participants completed the study, with 20 in each group. Paired-sample t-test showed a significant post-stimulation improvement in executive effect performance (t = 2.245; p = 0.037) in the DAN+/DN-tDCS group. The sham group did not exhibit any significant differences in ANT performance. Participants identified the stimulation type with 52.50% accuracy, indicating no difference in blinding efficacy between groups (p = 0.241). Mild-to-moderate adverse effects, such as stinging, itching, and skin reddening, were reported in the DAN+/DN-tDCS group (p < 0.05). Conclusion: DAN+/DN-tDCS enhanced attention function in healthy young individuals, particularly in improving executive effect performance. This study presents novel strategies for enhancing attentional performance and encourages further investigation into the mechanisms and outcomes of these interventions across diverse populations.

Photocatalytic Hydrogen Peroxide Production by a Mixed Ligand-Functionalized Uranyl-Organic Framework.

Hydrogen peroxide (H2O2) production driven by solar energy has received enormous attention due to its high efficiency, low cost, and environmental friendliness characteristics. Searching for new photocatalytic materials for H2O2 production is one of the most important targets. In this work, a new three-dimensional (3D) uranyl-organic framework material was constructed with mixed ligands via a solvothermal reaction and used for photocatalytic H2O2 production. The mixed ligand strategy not only benefits the construction of a 3D uranyl-organic framework but also introduces strong photon absorption groups into the framework. The thiophene and pyridine rings in the framework enhance photon absorption and carrier transfer. In addition, with the assistance of the hydrogen abstraction reaction of uranyl centers, the H2O2 production rate reaches 345 µmol h-1 g-1. This study provides a new blueprint for exploring the artificial photosynthesis of H2O2 through uranium-based metal-organic frameworks.

Protocol for preparing mammalian skin samples encompassing hair follicles for spatial transcriptomics.

Spatial transcriptomics enables a single-cell resolution view of gene expression patterns in tissues, providing insight into their biological functions. However, applying this approach to the skin presents inherent challenges. Here, we present a protocol for preparing mammalian skin samples encompassing hair follicles for spatial transcriptomics. We describe steps for sample preparation, embedding, acquisition of frozen slices, RNA quality control, tissue mounting, fixation, staining, and imaging. We then detail procedures for permeabilization, reverse transcription, and cDNA collection. For complete details on the use and execution of this protocol, please refer to Chen et al.1.

Urinary microbiome dysbiosis is associated with an inflammatory environment and perturbed fatty acids metabolism in the pathogenesis of bladder cancer.

BACKGROUND: Bladder cancer is a common malignancy with high recurrence rate. Early diagnosis and recurrence surveillance are pivotal to patients' outcomes, which require novel minimal-invasive diagnostic tools. The urinary microbiome is associated with bladder cancer and can be used as biomarkers, but the underlying mechanism is to be fully illustrated and diagnostic performance to be improved. METHODS: A total of 23 treatment-naïve bladder cancer patients and 9 non-cancerous subjects were enrolled into the Before group and Control group. After surgery, 10 patients from the Before group were further assigned into After group. Void mid-stream urine samples were collected and sent for 16S rDNA sequencing, targeted metabolomic profiling, and flow cytometry. Next, correlations were analyzed between microbiota, metabolites, and cytokines. Finally, receiver operating characteristic (ROC) curves of the urinary biomarkers were plotted and compared. RESULTS: Comparing to the Control group, levels of IL-6 (p < 0.01), IL-8 (p < 0.05), and IL-10 (p < 0.05) were remarkably elevated in the Before group. The α diversity of urine microbiome was also significantly higher, with the feature microbiota positively correlated to the level of IL-6 (r = 0.58, p < 0.01). Significant differences in metabolic composition were also observed between the Before and Control groups, with fatty acids and fatty acylcarnitines enriched in the Before group. After tumor resection, cytokine levels and the overall microbiome structure in the After group remained similar to that of the Before group, but fatty acylcarnitines were significantly reduced (p < 0.05). Pathway enrichment analysis revealed beta-oxidation of fatty acids was significantly involved (p < 0.001). ROC curves showed that the biomarker panel of Actinomycetaceae + arachidonic acid + IL-6 had superior diagnostic performance, with sensitivity of 0.94 and specificity of 1.00. CONCLUSIONS: Microbiome dysbiosis, proinflammatory environment and altered fatty acids metabolism are involved in the pathogenesis of bladder cancer, which may throw light on novel noninvasive diagnostic tool development.

Si-doped carbonized polymer dot as robust hydrophilic coating using for high efficiency antifogging.

Hydrophilic coating can prevent surface from fogging but its application is limited by low mechanical performance. In this study, a hydrophilic coating was prepared by crosslinking the Si-doped carbonized polymer dot (Si-CPD) with 3-glycidyloxypropyltrimethoxysilane (GPTMS) and ethylene oxide (EO). The hydrophilic coating can be used as robust hydrophilic anti-fogging coating. The Si-CPD derived from ethylene diamine tetraacetic acid (EDTA) and aminopropyl oligosiloxanes (APOS) was successfully prepared via one-step hydrothermal method. Then, a resin solution was prepared by mixing Si-CPD, GPTMS and EO. Epoxy group of GPTMS and EO can react with amino group of Si-CPD. Finally, a composite coating with antifogging function can be obtained by simple heating curing. Due to the introduction of hydroxyl which derived from EO, the coating shows excellent antifogging performance. Meanwhile, the presence of inorganic component endows the coating with outstanding mechanical performance. The coating has great potential in related applications, such as optical lenses, mirrors and other transparency substrates.

Transcriptional landscape and dynamics involved in sugar and acid accumulation during apple fruit development.

In fleshy fruit, sugars and acids are central components of fruit flavor and quality. To date, the mechanisms underlying transcriptional regulation of sugar and acid during fruit development remain largely unknown. Here, we combined ATAC-seq with RNA-seq to investigate the genome-wide chromatin accessibility and to identify putative transcription factors related to sugar and acid accumulation during apple (Malus domestica) fruit development. By integrating the differentially accessible regions and differentially expressed genes, we generated a global data set of promoter-accessibility and expression-increased genes. Using this strategy, we constructed a transcriptional regulatory network enabling screening for key transcription factors and target genes involved in sugar and acid accumulation. Among these transcription factors, 5 fruit-specific DNA binding with one finger genes were selected to confirm their regulatory effects, and our results showed that they could affect sugar or acid concentration by regulating the expression of sugar or acid metabolism-related genes in apple fruits. Our transcriptional regulatory network provides a suitable platform to identify candidate genes that control sugar and acid accumulation. Meanwhile, our data set will aid in analyzing other characteristics of apple fruit that have not been illuminated previously. Overall, these findings support a better understanding of the regulatory dynamics during apple fruit development and lay a foundation for quality improvement of apple.

Dihydroxanthene-based monoamine oxidase A-activated photosensitizers for photodynamic/photothermal therapy of tumors.

Small molecule photosensitizers for combined in vivo tailored cancer diagnostics and photodynamic/photothermal therapy are desperately needed. Monoamine oxidase A (MAO-A)-activated therapeutic and diagnostic compounds provide great selectivity because MAO-A can be employed as a biomarker for associated Tumors. In order to screen photosensitizers with photodynamic therapeutic potential, we have created a range of near-infrared fluorescent molecules in this work by combining dihydroxanthene parent with various heterocyclic fluorescent dyes. The NIR fluorescent diagnostic probe, DHMQ, was created by combining the screened fluorescent dye matrices with the propylamino group, which is the recognition moiety of MAO-A, based on the oxidative deamination mechanism of the enzyme. This probe has a low toxicity level and can identify MAO-A precisely. It has the ability to use fluorescence imaging on mice and cells to track MAO-A activity in real-time. It has strong phototoxicity and can produce singlet oxygen when exposed to laser light. The temperature used in photothermal imaging can get up to 50 °C, which can harm tumor cells permanently and have a positive phototherapeutic impact on tumors grown from SH-SY5Y xenograft mice. The concept of using MAO-A effectively in diseases is expanded by the MAO-A-activated diagnostic-integrated photosensitizers, which offer a new platform for in vivo cancer diagnostics and targeted anticancer treatment.

Molecular weight control of poly-γ-glutamic acid reveals novel insights into extracellular polymeric substance synthesis in Bacillus licheniformis.

BACKGROUND: The structural diversity of extracellular polymeric substances produced by microorganisms is attracting particular attention. Poly-gamma-glutamic acid (γ-PGA) is a widely studied extracellular polymeric substance from Bacillus species. The function of γ-PGA varies with its molecular weight (Mw). RESULTS: Herein, different endogenous promoters in Bacillus licheniformis were selected to regulate the expression levels of pgdS, resulting in the formation of γ-PGA with Mw values ranging from 1.61 × 103 to 2.03 × 104 kDa. The yields of γ-PGA and exopolysaccharides (EPS) both increased in the pgdS engineered strain with the lowest Mw and viscosity, in which the EPS content was almost tenfold higher than that of the wild-type strain. Subsequently, the compositions of EPS from the pgdS engineered strain also changed. Metabolomics and RT-qPCR further revealed that improving the transportation efficiency of EPS and the regulation of carbon flow of monosaccharide synthesis could affect the EPS yield. CONCLUSIONS: Here, we present a novel insight that increased pgdS expression led to the degradation of γ-PGA Mw and changes in EPS composition, thereby stimulating EPS and γ-PGA production. The results indicated a close relationship between γ-PGA and EPS in B. licheniformis and provided an effective strategy for the controlled synthesis of extracellular polymeric substances.

Transcriptional factor MdESE3 controls fruit acidity by activating genes regulating malic acid content in apple.

Acidity is a key factor controlling fruit flavor and quality. In a previous study, combined transcriptome and methylation analyses identified a P3A-type ATPase from apple (Malus domestica), MdMa11, which regulates vacuolar pH when expressed in Nicotiana benthamiana leaves. In this study, the role of MdMa11 in controlling fruit acidity was verified in apple calli, fruits, and plantlets. In addition, we isolated an APETALA2 domain-containing transcription factor, designated MdESE3, based on yeast one-hybrid (Y1H) screening using the MdMa11 promoter as bait. A subcellular localization assay indicated that MdESE3 localized to the nucleus. Analyses of transgenic apple calli, fruits, and plantlets, as well as tomatoes, demonstrated that MdESE3 enhances fruit acidity and organic acid accumulation. Meanwhile, chromatin immunoprecipitation quantitative PCR, luciferase (LUC) transactivation assays, and GUS reporter assays indicated that MdESE3 could bind to the ethylene-responsive element (ERE; 5'-TTTAAAAT-3') upstream of the MdMa11 transcription start site, thereby activating its expression. Furthermore, MdtDT, MdDTC2, and MdMDH12 expression increased in apple fruits and plantlets overexpressing MdESE3 and decreased in apple fruits and plantlets where MdESE3 was silenced. The ERE was found in MdtDT and MdMDH12 promoters, but not in the MdDTC2 promoter. The Y1H, LUC transactivation assays, and GUS reporter assays indicated that MdESE3 could bind to the MdtDT and MdMDH12 promoters and activate their expression. Our findings provide valuable functional validation of MdESE3 and its role in the transcriptional regulation of MdMa11, MdtDT, and MdMDH12 and malic acid accumulation in apple.

An insertion in the promoter of a malate dehydrogenase gene regulates malic acid content in apple fruit.

Malic acid is an important flavor determinant in apple (Malus × domestica Borkh.) fruit. One known variation controlling malic acid is the A/G single nucleotide polymorphism in an aluminum-activated malate transporter gene (MdMa1). Nevertheless, there are still differences in malic acid content in apple varieties with the same Ma1 genotype (Ma1/Ma1 homozygous), such as 'Honeycrisp' (high malic acid content) and 'Qinguan' (low malic acid content), indicating that other loci may influence malic acid and fruit acidity. Here, the F1 (Filial 1) hybrid generation of 'Honeycrisp' × 'Qinguan' was used to analyze quantitative trait loci for malic acid content. A major locus (Ma7) was identified on chromosome 13. Within this locus, a malate dehydrogenase gene, MDH1 (MdMa7), was the best candidate for further study. Subcellular localization suggested that MdMa7 encodes a cytosolic protein. Overexpression and RNA interference of MdMa7 in apple fruit increased and decreased malic acid content, respectively. An insertion/deletion (indel) in the MdMa7 promoter was found to affect MdMa7 expression and malic acid content in both hybrids and other cultivated varieties. The insertion and deletion genotypes were designated as MA7 and ma7, respectively. The transcription factor MdbHLH74 was found to stimulate MdMa7 expression in the MA7 genotype but not in the ma7 genotype. Transient transformation of fruit showed that MdbHLH74 affected MdMa7 expression and malic acid content in 'Gala' (MA7/MA7) but not in 'Fuji' (ma7/ma7). Our results indicated that genetic variation in the MdMa7 (MDH1) promoter alters the binding ability of the transcription factor MdbHLH74, which alters MdMa7 (MDH1) transcription and the malic acid content in apple fruit, especially in Ma1/Ma1 homozygous accessions.

Inter- and trans-generational impacts of real-world PM2.5 exposure on male-specific primary hypogonadism.

Exposure to PM2.5, a harmful type of air pollution, has been associated with compromised male reproductive health; however, it remains unclear whether such exposure can elicit transgenerational effects on male fertility. Here, we aim to examine the effect of paternal exposure to real-world PM2.5 on the reproductive health of male offspring. We have observed that paternal exposure to real-world PM2.5 can lead to transgenerational primary hypogonadism in a sex-selective manner, and we have also confirmed this phenotype by using an external model. Mechanically, we have identified small RNAs (sRNAs) that play a critical role in mediating these transgenerational effects. Specifically, miR6240 and piR016061, which are present in F0 PM sperm, regulate intergenerational transmission by targeting Lhcgr and Nsd1, respectively. We have also uncovered that piR033435 and piR006695 indirectly regulate F1 PM sperm methylation by binding to the 3'-untranslated region of Tet1 mRNA. The reduced expression of Tet1 resulted in hypermethylation of several testosterone synthesis genes, including Lhcgr and Gnas, impaired Leydig cell function and ultimately led to transgenerational primary hypogonadism. Our findings provide insights into the mechanisms underlying the transgenerational effects of paternal PM2.5 exposure on reproductive health, highlighting the crucial role played by sRNAs in mediating these effects. The findings underscore the significance of paternal pre-conception interventions in alleviating the adverse effects of environmental pollutants on reproductive health.

A spatiotemporal atlas of mouse liver homeostasis and regeneration.

The mechanism by which mammalian liver cell responses are coordinated during tissue homeostasis and perturbation is poorly understood, representing a major obstacle in our understanding of many diseases. This knowledge gap is caused by the difficulty involved with studying multiple cell types in different states and locations, particularly when these are transient. We have combined Stereo-seq (spatiotemporal enhanced resolution omics-sequencing) with single-cell transcriptomic profiling of 473,290 cells to generate a high-definition spatiotemporal atlas of mouse liver homeostasis and regeneration at the whole-lobe scale. Our integrative study dissects in detail the molecular gradients controlling liver cell function, systematically defining how gene networks are dynamically modulated through intercellular communication to promote regeneration. Among other important regulators, we identified the transcriptional cofactor TBL1XR1 as a rheostat linking inflammation to Wnt/ß-catenin signaling for facilitating hepatocyte proliferation. Our data and analytical pipelines lay the foundation for future high-definition tissue-scale atlases of organ physiology and malfunction.

The association between different physical activity (PA) patterns and cardiometabolic index (CMI) in US adult population from NHANES (2007-2016).

Background: Physical activity (PA) is widely recommended for preventing and combating obesity, but the most effective PA pattern for treating obesity remains unclear. Cardiometabolic index (CMI), derived from waist height ratio and triglycerides to high-density lipoprotein-cholesterol ratio, is a novel indicator for evaluating obesity. However, the relationship between different PA patterns and CMI remains unelucidated. Objective: This study aimed to explore the association between different PA patterns and CMI in U.S. adults. Methods: Participants with complete information in CMI, PA patterns, and other covariates in the National Health and Nutrition Examination Survey database (2007-2016) were included in this study. Multivariate linear regression models were utilized to explore the relationship between PA patterns and CMI. Moreover, stratified analyses, interaction tests and restricted cubic spline (RCS) regression analysis were used to investigate the stability and nonlinearity of the association, respectively. Results: A total of 16,442 adults were included in this study. After adjusting for all potential covariates, only the regularly active group was significantly associated with CMI reduction (ß = -0.13, 95% CI: 0.19 to -0.07, P < 0.0001), while the weekend warriors group did not achieve equivalent CMI reduction (ß = -0.09, 95% CI: 0.32 to 0.14, P = 0.4204). Subgroup analyses and interaction tests revealed that the CMI-PA association was more pronounced in the subgroup with age≤45 or >60, with higher education level, and who are current drinkers. Furthermore, RCS analysis indicated that total PA in a week was significantly, nonlinearly associated with CMI in non-inactive adults, and that a total of PA more than 330 min can reap favorable CMI reduction. Conclusion: Being regularly active is associated with significant CMI reduction, while being weekend warriors and insufficiently active do not achieve equivalent benefits. For non-inactive individuals, engaging in PA for more than 330 min weekly helps to reduce CMI effectively.

Suppression of A-to-I RNA-editing enzyme ADAR1 sensitizes hepatocellular carcinoma cells to oxidative stress through regulating Keap1/Nrf2 pathway.

BACKGROUND: A-to-I RNA editing is an abundant post-transcriptional modification event in hepatocellular carcinoma (HCC). Evidence suggests that adenosine deaminases acting on RNA 1 (ADAR1) correlates to oxidative stress that is a crucial factor of HCC pathogenesis. The present study investigated the effect of ADAR1 on survival and oxidative stress of HCC, and underlying mechanisms. METHODS: ADAR1 expression was measured in fifty HCC and normal tissues via real-time quantitative PCR, and immunohistochemistry. For stable knockdown or overexpression of ADAR1, adeno-associated virus vectors carrying sh-ADAR1 or ADAR1 overexpression were transfected into HepG2 and SMMC-7721 cells. Transfected cells were exposed to oxidative stress agonist tBHP or sorafenib Bay 43-9006. Cell proliferation, apoptosis, and oxidative stress were measured, and tumor xenograft experiment was implemented. RESULTS: ADAR1 was up-regulated in HCC and correlated to unfavorable clinical outcomes. ADAR1 deficiency attenuated proliferation of HCC cells and tumor growth and enhanced apoptosis. Moreover, its loss facilitated intracellular ROS accumulation, and elevated Keap1 and lowered Nrf2 expression. Intracellular GSH content and SOD activity were decreased and MDA content was increased in the absence of ADAR1. The opposite results were observed when ADAR1 was overexpressed. The effects of tBHP and Bay 43-9006 on survival, apoptosis, intracellular ROS accumulation, and Keap1/Nrf2 pathway were further exacerbated by simultaneous inhibition of ADAR1. CONCLUSIONS: The current study unveils that ADAR1 is required for survival and oxidative stress of HCC cells, and targeting ADAR1 may sensitize HCC cells to oxidative stress via modulating Keap1/Nrf2 pathway.

Predictive value of bilirubin and serum γ-glutamyltranspeptidase levels in type-2 diabetes mellitus patients with acute coronary syndrome.

BACKGROUND: Cardiovascular disease is a major complication of diabetes mellitus (DM). Type-2 DM (T2DM) is associated with an increased risk of cardiovascular events and mortality, while serum biomarkers may facilitate the prediction of these outcomes. Early differential diagnosis of T2DM complicated with acute coronary syndrome (ACS) plays an important role in controlling disease progression and improving safety. AIM: To investigate the correlation of serum bilirubin and γ-glutamyltranspeptidase (γ-GGT) with major adverse cardiovascular events (MACEs) in T2DM patients with ACS. METHODS: The clinical data of inpatients from January 2022 to December 2022 were analyzed retrospectively. According to different conditions, they were divided into the T2DM complicated with ACS group (T2DM + ACS, n = 96), simple T2DM group (T2DM, n = 85), and simple ACS group (ACS, n = 90). The clinical data and laboratory indices were compared among the three groups, and the correlations of serum total bilirubin (TBIL) levels and serum γ-GGT levels with other indices were discussed. T2DM + ACS patients received a 90-day follow-up after discharge and were divided into event (n = 15) and nonevent (n = 81) groups according to the occurrence of MACEs; Univariate and multivariate analyses were further used to screen the independent influencing factors of MACEs in patients. RESULTS: The T2DM + ACS group showed higher γ-GGT, total cholesterol, low-density lipoprotein cholesterol (LDL-C) and glycosylated hemoglobin (HbA1c) and lower TBIL and high-density lipoprotein cholesterol levels than the T2DM and ACS groups (P < 0.05). Based on univariate analysis, the event and nonevent groups were significantly different in age (t = 3.3612, P = 0.0011), TBIL level (t = 3.0742, P = 0.0028), γ-GGT level (t = 2.6887, P = 0.0085), LDL-C level (t = 2.0816, P = 0.0401), HbA1c level (t = 2.7862, P = 0.0065) and left ventricular ejection fraction (LEVF) levels (t=3.2047, P = 0.0018). Multivariate logistic regression analysis further identified that TBIL level and LEVF level were protective factor for MACEs, and age and γ-GGT level were risk factors (P < 0.05). CONCLUSION: Serum TBIL levels are decreased and γ-GGT levels are increased in T2DM + ACS patients, and the two indices are significantly negatively correlated. TBIL and γ-GGT are independent influencing factors for MACEs in such patients.

Protocol for optimized dissociation of human scalp tissue for hair follicle transcriptomics by scRNA-seq.

Single-cell RNA sequencing (scRNA-seq) is a powerful tool for studying transcriptomics. Here, we present an optimized protocol for dissociating human scalp tissue and acquiring high-quality single-cell suspension for scRNA-seq to study transcriptomics of human hair follicles. We describe steps for human scalp tissue cleaning, subcutaneous fat removal, mechanical mincing, and enzymatic digestion. We then detail procedures for cleaning, resuspending, a cell viability assay, and library construction.

Loop-mediated isothermal amplification linked a nanoparticles-based biosensor for detecting Epstein-Barr virus.

Epstein-Barr virus (EBV) is a ubiquitous gamma herpesvirus that maintains a lifelong latent association with B lymphocytes. Here, a rapid and reliable diagnosis platform for detecting EBV infection, employing loop-mediated isothermal amplification (LAMP) combined with a gold nanoparticles-based lateral flow biosensors (AuNPs-LFB) (termed LAMP Amplification Mediated AuNPs-LFB Detection, LAMAD), was developed in the current study. A set of specific LAMP primers targeting the Epstein-Barr nuclear antigen (EBNA) leader protein (EBNA-LP) gene was designed and synthesized. Subsequently, these templates extracted from various pathogens and whole blood samples were used to optimize and evaluate the EBV-LAMAD assay. As a result, the limit of detection (LoD) of the EBV-LAMAD assay was 45 copies/reaction. The EBV-LAMAD assay can detect all representative EBV pathogens used in the study, and of note, no cross-reactions were observed with other non-EBV organisms. Moreover, the whole workflow of the EBV-LAMAD assay can be completed within 70 min, including rapid EBV template preparation, EBV-LAMP amplification, and AuNPs-LFB-mediated detection. Taken together, the EBV-LAMAD assay targeting the EBNA-LP gene is a rapid, simplified, sensitive, reliable, and easy-to-use detection protocol that can be used as a competitive potential diagnostic/screening tool for EBV infection in clinical settings, especially in basic laboratories in resource-limited regions. KEY POINTS: • A novel, simplified, and easy-to-use AuNPs-LFB biosensor was designed and prepared. • LAMP combined with an AuNPs-LFB targeting the novel EBNA-LP gene was established. • EBV-LAMAD is a rapid, sensitive, and reliable detection protocol for EBV infection.

Protocol for acquiring high-quality fresh mouse lung spatial transcriptomics data.

Spatial transcriptomics analysis allows the examination of the biological characteristics and spatial distribution of individual lung cells at a single-cell resolution. However, due to the presence of cavities in the alveoli of the lungs, it is challenging to section them for spatial transcriptomics experiments. Here, we present a protocol for acquiring high-quality fresh mouse lung spatial transcriptomics data. We describe steps for lung perfusion, acquiring frozen slices, collecting cDNA from lung sections, and data analysis. For complete details on the use and execution of this protocol, please refer to Jiang et al.1.

The Influence of Dexmedetomidine as an Adjuvant in Intrathecal Labor Analgesia: A Multicenter Study on Efficacy and Maternal Satisfaction.

In this study, we examined the impact of dexmedetomidine (DEX) on the effectiveness of epidural analgesia and labor outcomes. We administered different doses of DEX combined with 0.1% ropivacaine for epidural analgesia to evaluate the clinical effects and safety. To assess the effects of different concentrations of DEX in parturient women receiving epidural analgesia, we conducted a randomized double-blind trial. We selected 400 parturient women and randomly assigned them to 4 groups, with 100 parturient women in each group: S0.1 (0.1 µg/mL DEX), S0.2 (0.2 µg/mL DEX), S0.3 (0.3 µg/mL DEX), and a control group (0.3 µg/mL sufentanil). Post-analgesia, we recorded the Bromage score, duration of labor, method of delivery, bleeding, neonatal Apgar score, adverse reactions, and maternal satisfaction. The number of patients with a Bromage score of ≥2 and the incidence of bradycardia were higher in the S0.3 group compared with the other 3 groups (P < .05), whereas the high satisfaction rate was lower in the S0.3 group (P < .05). Moreover, we found that the number of times that additional patient-controlled analgesia was administered was higher in the S0.1 group compared with the remaining 3 groups (P < .05). The control group exhibited a higher incidence of pruritus than the other 3 groups (P < .05). In conclusion, when administering spinal anesthesia for the relief of labor pain, epidural analgesia with 0.1% ropivacaine combined with 0.2 µg/mL DEX provides relatively ideal analgesic effects, higher maternal satisfaction, and reduces the incidence of pruritus, compared with the combination of 0.1% ropivacaine and 0.3 µg/mL sufentanil.