Development and Application of a Comprehensive Nontargeted Screening Strategy for Aristolochic Acid Analogues.

Aristolochic acid analogs (AAAs) are naturally occurring carcinogenic and toxic compounds that pose a safety threat to pharmaceuticals and the environment. It is challenging to screen AAAs due to their lack of characteristic mass spectral fragmentation and their presence of structural diversity. A comprehensive nontargeted screening strategy was proposed by taking into account diverse factors and incorporating various self-developed techniques, and a Python3-based toolkit called AAAs_finder was developed for its implementation. The main procedures consist of virtual structure and ultraviolet and visible (UV) spectra database creation, exact mass and UV spectra-based suspect data extraction, tandem mass spectra (MS/MS) anthropomorphic interpretation, and multicondition retention time (RT) prediction-based candidate structures ranking. To initially assess screening feasibility, eight hypothetical unknown samples were subjected to nontargeted screening using the AAAs_finder toolkit and two other advanced tools. The results showed that the former successfully identified all, while the latter two only managed to identify two and three, respectively, indicating that our strategy was more feasible. After that, the strategy was carefully evaluated for false positives and false negatives, instrument dependence, reproducibility, and sensitivity. After the evaluation, the strategy was successfully applied to the screening of AAAs in real samples, such as herbal medicine, spiked soil, and water. Overall, this study proposed a nontargeted screening strategy and toolkit independent of characteristic mass spectral fragmentation and able to overcome challenges posed by structural diversity for the AAAs screening, which is also valuable for other classes of compounds.

Pulsed Direct Current Arc-Induced Nanoelectrospray Ionization Mass Spectrometry.

This study explores the innovative field of pulsed direct current arc-induced nanoelectrospray ionization mass spectrometry (DCAI-nano-ESI-MS), which utilizes a low-temperature direct current (DC) arc to induce ESI during MS analyses. By employing a 15 kV output voltage, the DCAI-nano-ESI source effectively identifies various biological molecules, including angiotensin II, bradykinin, cytochrome C, and soybean lecithin, showcasing impressive analyte signals and facilitating multicharge MS in positive- and negative-ion modes. Notably, results show that the oxidation of fatty acids using a DC arc produces [M + O - H]- ions, which aid in identifying the location of C═C bonds in unsaturated fatty acids and distinguishing between isomers based on diagnostic ions observed during collision-induced dissociation tandem MS. This study presents an approach for identifying the sn-1 and sn-2 positions in phosphatidylcholine using phosphatidylcholine and nitrate adduct ions, accurately determining phosphatidylcholine molecular configurations via the Paternò-Büchi reaction. With all the advantages above, DCAI-nano-ESI holds significant promise for future analytical and bioanalytical applications.

Comprehensive analysis of phospholipid in milk and their biological roles as nutrients and biomarkers.

Phospholipids (PL) have garnered significant attention due to their physiological activities. Milk and other dairy products are important dietary sources for humans and have been extensively used to analyze the presence of PL by various analytical techniques. In this paper, the analysis techniques of PL were reviewed with the eight trigrams of phospholipidomics and a comprehensive fingerprint of 1295 PLs covering 8 subclasses in milk and other dairy products, especially. Technology is the primary productive force. Based on phospholipidomics technology, we further review the relationship between the composition of PL and factors that may be involved in processing and experimental operation, and emphasized the significance of the biological role played by PL in dietary supplements and biomarkers (production, processing and clinical research), and providing the future research directions.

Rapid analysis of the chemical constituents of traditional Tibetan medicine Sbyor-bzo-ghi-wang using ballpoint electrospray ionization technique.

RATIONALE: The chemical constituents of traditional Tibetan medicines (TTM) can be identified using high-performance liquid chromatography and high-resolution mass spectrometry (HPLC-MS/MS) technique. However, the HPLC-MS/MS technique requires the sample to be pretreated and then separated using the specific liquid chromatography method, which is time consuming. This study developed a ballpoint electrospray ionization (BPESI) technique for analyzing the chemical constituents of Sbyor-bzo-ghi-wang. This technique is a simple and inexpensive method for the rapid identification of the chemical constituents of TTMs. METHODS: After the important parameters of the homemade BPESI device were optimized, the chemical constituents of Sbyor-bzo-ghi-wang were quickly identified without sample pretreatment. The raw data were converted to mzML file using MSConvert and then identified using SIRIUS 5 software. RESULTS: The results showed that 30 compounds were identified from Sbyor-bzo-ghi-wang, namely eight bile acids, six flavonoids, four alkaloids, three amino acids, and nine others. Compared to the ultra-high-performance liquid chromatography-Q/Orbitrap and high-resolution mass spectrometry (UHPLC-Q/Orbitrap HRMS) technique, the BPESI technique identified almost similar types of compounds and also a comparable number of compounds. CONCLUSIONS: Compared with the traditional HPLC-MS/MS methods, the BPESI technique does not require complex sample pretreatment and subsequent chromatographic separation steps; also it consumes a small quantity of samples. Therefore, BPESI can be used for the qualitative analysis of the chemical constituents of Sbyor-bzo-ghi-wang.

An improved 3D-UNet-based brain hippocampus segmentation model based on MR images.

OBJECTIVE: Accurate delineation of the hippocampal region via magnetic resonance imaging (MRI) is crucial for the prevention and early diagnosis of neurosystemic diseases. Determining how to accurately and quickly delineate the hippocampus from MRI results has become a serious issue. In this study, a pixel-level semantic segmentation method using 3D-UNet is proposed to realize the automatic segmentation of the brain hippocampus from MRI results. METHODS: Two hundred three-dimensional T1-weighted (3D-T1) nongadolinium contrast-enhanced magnetic resonance (MR) images were acquired at Hangzhou Cancer Hospital from June 2020 to December 2022. These samples were divided into two groups, containing 175 and 25 samples. In the first group, 145 cases were used to train the hippocampus segmentation model, and the remaining 30 cases were used to fine-tune the hyperparameters of the model. Images for twenty-five patients in the second group were used as the test set to evaluate the performance of the model. The training set of images was processed via rotation, scaling, grey value augmentation and transformation with a smooth dense deformation field for both image data and ground truth labels. A filling technique was introduced into the segmentation network to establish the hippocampus segmentation model. In addition, the performance of models established with the original network, such as VNet, SegResNet, UNetR and 3D-UNet, was compared with that of models constructed by combining the filling technique with the original segmentation network. RESULTS: The results showed that the performance of the segmentation model improved after the filling technique was introduced. Specifically, when the filling technique was introduced into VNet, SegResNet, 3D-UNet and UNetR, the segmentation performance of the models trained with an input image size of 48 × 48 × 48 improved. Among them, the 3D-UNet-based model with the filling technique achieved the best performance, with a Dice score (Dice score) of 0.7989 ± 0.0398 and a mean intersection over union (mIoU) of 0.6669 ± 0.0540, which were greater than those of the original 3D-UNet-based model. In addition, the oversegmentation ratio (OSR), average surface distance (ASD) and Hausdorff distance (HD) were 0.0666 ± 0.0351, 0.5733 ± 0.1018 and 5.1235 ± 1.4397, respectively, which were better than those of the other models. In addition, when the size of the input image was set to 48 × 48 × 48, 64 × 64 × 64 and 96 × 96 × 96, the model performance gradually improved, and the Dice scores of the proposed model reached 0.7989 ± 0.0398, 0.8371 ± 0.0254 and 0.8674 ± 0.0257, respectively. In addition, the mIoUs reached 0.6669 ± 0.0540, 0.7207 ± 0.0370 and 0.7668 ± 0.0392, respectively. CONCLUSION: The proposed hippocampus segmentation model constructed by introducing the filling technique into a segmentation network performed better than models built solely on the original network and can improve the efficiency of diagnostic analysis.

Evidence for chromium crosses blood brain barrier from the hypothalamus in chromium mice model.

It has been shown that exposure to hexavalent Chromium, Cr (Ⅵ), via nasal cavity can have neurotoxicological effects and induces behavioral impairment due to the fact that blood brain barrier (BBB) does not cover olfactory bulb. But whether Cr (Ⅵ) can cross the BBB and have a toxicological effects in central nervous system (CNS) remains unclear. Therefore, we investigated the effects of Cr (Ⅵ) on mice treated with different concentrations and exposure time (14 days and 28 days) of Cr (Ⅵ) via intraperitoneal injection. Results revealed that Cr accumulated in hypothalamus (HY) in a timely dependent manner. Much more severer neuropathologies was observed in the group of mice exposed to Cr (Ⅵ) for 28 days than that for 14 days. Gliosis, neuronal morphological abnormalities, synaptic degeneration, BBB disruption and neuronal number loss were observed in HY. In terms of mechanism, the Nrf2 related antioxidant stress signaling dysfunction and activated NF-κB related inflammatory pathway were observed in HY of Cr (Ⅵ) intoxication mice. And these neuropathologies and signaling defects appeared in a timely dependent manner. Taking together, we proved that Cr (Ⅵ) can enter HY due to weaker BBB in HY and HY is the most vulnerable CNS region to Cr (Ⅵ) exposure. The concentration of Cr in HY increased along with time. The accumulated Cr in HY can cause BBB disruption, neuronal morphological abnormalities, synaptic degeneration and gliosis through Nrf2 and NF-κB signaling pathway. This finding improves our understanding of the neurological dysfunctions observed in individuals who have occupational exposure to Cr (Ⅵ), and provided potential therapeutic targets to treat neurotoxicological pathologies induced by Cr (Ⅵ).

Alginate Oligosaccharides Enhance Antioxidant Status and Intestinal Health by Modulating the Gut Microbiota in Weaned Piglets.

Alginate oligosaccharides (AOSs), which are an attractive feed additive for animal production, exhibit pleiotropic bioactivities. In the present study, we investigated graded doses of AOS-mediated alterations in the physiological responses of piglets by determining the intestinal architecture, barrier function, and microbiota. A total of 144 weaned piglets were allocated into four dietary treatments in a completely random design, which included a control diet (CON) and three treated diets formulated with 250 mg/kg (AOS250), 500 mg/kg (AOS500), and 1000 mg/kg AOS (AOS1000), respectively. The trial was carried out for 28 days. Our results showed that AOS treatment reinforced the intestinal barrier function by increasing the ileal villus height, density, and fold, as well as the expression of tight junction proteins, especially at the dose of 500 mg/kg AOS. Meanwhile, supplementations with AOSs showed positive effects on enhancing antioxidant capacity and alleviating intestinal inflammation by elevating the levels of antioxidant enzymes and inhibiting excessive inflammatory cytokines. The DESeq2 analysis showed that AOS supplementation inhibited the growth of harmful bacteria Helicobacter and Escherichia_Shigella and enhanced the relative abundance of Faecalibacterium and Veillonella. Collectively, these findings suggested that AOSs have beneficial effects on growth performance, antioxidant capacity, and gut health in piglets.

T790M mutation sensitizes non-small cell lung cancer cells to radiation via suppressing SPOCK1.

Background: Approximately 50% of patients harbor the T790M mutation after developing first-generation epidermal growth factor receptor tyrosine kinase inhibitor (EGFR-TKI) resistance. Evidence has showed the major treatment failure is local relapses and limited metastases. Several studies have demonstrated the value of radiotherapy in metastatic non-small cell lung cancer (NSCLC) with the EGFR T790M mutation after the development of TKI resistance. The aim of this study was to explore the role of radiation in T790M-mutant NSCLC and the value of early radiotherapy for NSCLC with T790M-mediated EGFR-TKI resistance. Methods: Gefitinib-resistant NSCLC cell lines were established via stepwise exposure to increasing concentrations of gefitinib (PC-9-GR). Droplet digital PCR was used to determine the relative T790M subclone abundance. In vitro and in vivo models were established using different mixtures of PC-9-GR and PC-9 cells. Differentially expressed genes were identified using RNA sequencing. Two research models were constructed (salvage and prophylactic radiotherapy) to determine the effects of early radiotherapy on gefitinib-resistant cells. Results: PC-9-GR cells exhibited higher radiosensitivity than PC-9 cells (sensitivity enhancement ratio = 1.5). Salvage radiation reduced the number of T790M-mutant subclones, and the relative T790M abundance was significantly lower than that without radiation at 90 days (10.94% vs. 21.54%). Prophylactic radiation prevented the development of T790M subclones. These results were also confirmed in vivo. qRT-PCR revealed threefold elevation of miR-1243 in PC-9-GR cells, and the increased radiosensitivity of PC-9-GR cells was inhibited when miR-1243 was knocked down. RNA sequencing revealed that SPOCK1 was downregulated in PC-9-GR cells. Interestingly, bioinformatic analysis showed that SPOCK1 was a target gene of miR-1243. SPOCK1 knockdown markedly increased the radiosensitivity of PC-9 cells. Conclusion: Gefitinib-resistant NSCLC with the T790M mutation had higher radiosensitivity than that without the mutation, possibly mediated by SPOCK1. Early radiotherapy can eliminate T790M subclones, providing evidence for the benefit of early local treatment in patients with TKI-resistant NSCLC.

Chain electrospray ionization mass spectrometry for ultra-low volume sample analysis.

In this work, chain electrospray ionization (chain-ESI) was developed to efficiently ionize trace samples for mass spectrometry analysis. The primary ion source was found to have the ability to induce secondary electrospray ionization with an extraordinarily low sample consumption rate in the picoliters per minute (pLs/min). This allows low volume sample to generate substantial tandem mass spectrum (MS2) data for metabolite annotations. Notably, chain-ESI can effectively prevent the electro-redox reaction in the process of electrospray, so as to reflect the native state of the analytes. Furthermore, from a single Broussonetia papyrifera (B. papyrifera) trichome and a single A549 cancer cell, 1426 and 617 metabolites were detected respectively. All of those observations demonstrated that chain-ESI offers the advantages of direct, rapid analysis with extreme-low volumes and high coverage, enabling the measurement of bio-information in low volume samples.

Cell-membrane engineering strategies for clinic-guided design of nanomedicine.

Cells are the fundamental units of life. The cell membrane primarily composed of two layers of phospholipids (a bilayer) structurally defines the boundary of a cell, which can protect its interior from external disturbances and also selectively exchange substances and conduct signals from the extracellular environment. The complexity and particularity of transmembrane proteins provide the foundation for versatile cellular functions. Nanomedicine as an emerging therapeutic strategy holds tremendous potential in the healthcare field. However, it is susceptible to recognition and clearance by the immune system. To overcome this bottleneck, the technology of cell membrane coating has been extensively used in nanomedicines for their enhanced therapeutic efficacy, attributed to the favorable fluidity and biocompatibility of cell membranes with various membrane-anchored proteins. Meanwhile, some engineering strategies of cell membranes through various chemical, physical and biological ways have been progressively developed to enable their versatile therapeutic functions against complex diseases. In this review, we summarized the potential clinical applications of four typical cell membranes, elucidated their underlying therapeutic mechanisms, and outlined their current engineering approaches. In addition, we further discussed the limitation of this technology of cell membrane coating in clinical applications, and possible solutions to address these challenges.

Rational design of ROS generation nanosystems to regulate innate immunity of macrophages, dendrtical and natural killing cells for immunotherapy.

Innate immunity serves as the first line of host defense in the body against pathogenic infections or malignant diseases. Reactive oxygen species (ROS), as vital signaling mediators, can efficiently elicit innate immune responses to oxidative-related stress or damage. In the era of nanomedicine, various immunostimulatory nanosystems have been extensively designed and synthesized to elicit immune responses for the immunotherapy of cancer or infectious diseases. In this review, we emphasize that ROS derived from nanosystems regulates innate immune cells to potentiate immunotherapeutic efficacy, such as primarily dendritic cells, macrophages, or natural killer cells. Meanwhile, we also summarize the pathway of ROS generation triggered by exogenous nanosystems in innate immune cells of DCs, macrophages, and NK cells.

Multifunctional nanozymes for disease diagnosis and therapy.

The development of nanotechnology has brought about groundbreaking advancements in diseases' diagnostics and therapeutics. Among them, multifunctional nanomaterials with enzyme-like activities (i.e., nanozymes) featured with high stability, large surface area for bioconjugation, and easy storage, offer unprecedented opportunities for disease diagnostics and treatment. Recent years have witnessed the great progress of nanozyme-based theranostics. To highlight these achievements, this review first introduces the recent advancements on nanozymes in biosensing and diagnostics. Then, it summarizes the applications of nanozymes in therapeutics including anti-tumor and antibacterial treatment, anti-inflammatory treatment, and other diseases treatment. In addition, several targeted strategies to improve the therapeutic efficacy of nanozyme are discussed. Finally, the opportunities and challenges in the field of diagnosis and therapy are summarized.

Analysis of therapeutic effects of congenital kyphosis in children due to anterior vertebral bone bridges.

Objective: To investigate the choice of treatment options and long-term orthopedic results of congenital kyphosis in children due to anterior vertebral bone bridges. Methods: The clinical data of children with congenital kyphosis due to anterior vertebral bridges treated at our center from May 2005 to May 2020 were retrospectively analyzed. We evaluated the clinical features of the deformity, the choice of treatment plan, the change in the Cobb angle of the kyphosis and the improvement of the sagittal trunk deviation before and after treatment and at the final follow-up visit by means of pre-treatment and post-treatment imaging, physical examination and analysis of the case data. Results: A total of 35 children were included. Clinical follow-up was conducted on a cohort of 5 children, all of whom presented with type Ⅱ congenital kyphosis caused by less than three thoracic anterior bone bridges. The study findings revealed no noteworthy advancement in segmental kyphosis, thoracic kyphosis, lumbar lordosis, and sagittal vertical axis during the final follow-up assessment (p > 0.05). In a cohort of 30 pediatric patients who underwent surgical intervention, segmental kyphosis was corrected, with a decrease from an average angle of (40.1 ± 20.5)° to (15.6 ± 9.5)°. Furthermore, significant improvements were noted in segmental kyphosis, thoracic kyphosis, lumbar lordosis, and sagittal vertical axis at the postoperative stage compared to the preoperative stage (p < 0.05). Notably, improvements in thoracic kyphosis and lumbar lordosis persisted at the final follow-up visit compared to postoperative (p < 0.05). Conclusion: Type Ⅱ congenital kyphosis in children caused by anterior bony bridges of less than three vertebrae in the thoracic segment can be followed up for a long period, and type Ⅱ/Ⅲ congenital kyphosis caused by anterior bony bridges of the vertebrae in the thoracolumbar, lumbar, and lumbosacral segments requires early surgery.

Predicting radiation pneumonitis in lung cancer: a EUD-based machine learning approach for volumetric modulated arc therapy patients.

Purpose: This study aims to develop an optimal machine learning model that uses lung equivalent uniform dose (lung EUD to predict radiation pneumonitis (RP) occurrence in lung cancer patients treated with volumetric modulated arc therapy (VMAT). Methods: We analyzed a cohort of 77 patients diagnosed with locally advanced squamous cell lung cancer (LASCLC) receiving concurrent chemoradiotherapy with VMAT. Patients were categorized based on the onset of grade II or higher radiation pneumonitis (RP 2+). Dose volume histogram data, extracted from the treatment planning system, were used to compute the lung EUD values for both groups using a specialized numerical analysis code. We identified the parameter α, representing the most significant relative difference in lung EUD between the two groups. The predictive potential of variables for RP2+, including physical dose metrics, lung EUD, normal tissue complication probability (NTCP) from the Lyman-Kutcher-Burman (LKB) model, and lung EUD-calibrated NTCP for affected and whole lung, underwent both univariate and multivariate analyses. Relevant variables were then employed as inputs for machine learning models: multiple logistic regression (MLR), support vector machine (SVM), decision tree (DT), and K-nearest neighbor (KNN). Each model's performance was gauged using the area under the curve (AUC), determining the best-performing model. Results: The optimal α-value for lung EUD was 0.3, maximizing the relative lung EUD difference between the RP 2+ and non-RP 2+ groups. A strong correlation coefficient of 0.929 (P< 0.01) was observed between lung EUD (α = 0.3) and physical dose metrics. When examining predictive capabilities, lung EUD-based NTCP for the affected lung (AUC: 0.862) and whole lung (AUC: 0.815) surpassed LKB-based NTCP for the respective lungs. The decision tree (DT) model using lung EUD-based predictors emerged as the superior model, achieving an AUC of 0.98 in both training and validation datasets. Discussions: The likelihood of developing RP 2+ has shown a significant correlation with the advancements in RT technology. From traditional 3-D conformal RT, lung cancer treatment methodologies have transitioned to sophisticated techniques like static IMRT. Accurately deriving such a dose-effect relationship through NTCP modeling of RP incidence is statistically challenging due to the increased number of degrees-of-freedom. To the best of our knowledge, many studies have not clarified the rationale behind setting the α-value to 0.99 or 1, despite the closely aligned calculated lung EUD and lung mean dose MLD. Perfect independence among variables is rarely achievable in real-world scenarios. Four prominent machine learning algorithms were used to devise our prediction models. The inclusion of lung EUD-based factors substantially enhanced their predictive performance for RP 2+. Our results advocate for the decision tree model with lung EUD-based predictors as the optimal prediction tool for VMAT-treated lung cancer patients. Which could replace conventional dosimetric parameters, potentially simplifying complex neural network structures in prediction models.

Hypofractionated stereotactic radiotherapy for brain metastases in lung cancer patients: dose‒response effect and toxicity.

BACKGROUND: Lung cancer is a common cause of brain metastases, approximately 40% of patients with lung cancer will develop brain metastases at some point during their disease. Hypofractionated stereotactic radiotherapy (HSRT) has been demonstrated to be effective in controlling limited brain metastases. However, there is still no conclusive on the optimal segmentation of HSRT. The aim of our study was to explore the correlation between the HSRT dosage and its treatment effect and toxicity. METHODS: A retrospective analysis was conducted on patients with non-small cell lung cancer (NSCLC) brain metastasis at Hangzhou Cancer Hospital from 1 January 2019 to 1 January 2021. The number of brain metastases did not exceed 10 in all patients and the number of fractions of HSRT was 5. The prescription dose ranges from 25 to 40 Gy. The Kaplan-Meier method was used for estimation of the localised intracranial control rate (iLC). Adverse radiation effects (AREs) were evaluated according to CTCAE 5.0. This study was approved by the Institutional Ethics Review Board of the Hangzhou Cancer Hospital (#73/HZCH-2022). RESULTS: Forty eligible patients with a total of 70 brain metastases were included in this study. The 1-year iLC was 76% and 89% in the prescribed dose ≤ 30 Gy and > 30 Gy group, respectively (P < 0.05). For patients treated with HSRT combined with targeted therapy, immunotherapy and chemotherapy, the 1-year iLC was 89%, 100%, and 45%, respectively. No significant associations were observed between the number, maximum diameter, location, and type of pathology of brain metastases. The rate of all-grade AREs was 33%. Two patients who received a total dose of 40 Gy developed grade 3 headache, the rest of the AREs were grade 1-2. CONCLUSIONS: Increasing the prescription dose of HSRT improves treatment effect but may also exacerbate the side effects. Systemic therapy might impact the iLC rate, and individualized treatment regimens need to be developed.

Dietary lipoic acid alleviates autism-like behavior induced by acrylamide in adolescent mice: the potential involvement of the gut-brain axis.

Consuming fried foods has been associated with an increased susceptibility to mental health disorders. Nevertheless, the impact of alpha-lipoic acid (α-LA, LA) on fried food-induced autism-like behavior remains unclear. This study aimed to explore how LA affects autism-related behavior and cognitive deficits caused by acrylamide in mice, a representative food hazard found in fried foods. This improvement was accomplished by enhanced synaptic plasticity, increased neurotrophin expression, elevated calcium-binding protein D28k, and restored serotonin. Additionally, LA substantially influenced the abundance of bacteria linked to autism and depression, simultaneously boosted short-chain fatty acid (SCFA) levels in fecal samples, and induced changes in serum amino acid concentrations. In summary, these findings suggested that exposure to acrylamide in adolescent mice could induce the development of social disorders in adulthood. LA showed promise as a nutritional intervention strategy to tackle emotional disorders during adolescence.

Association of GAL-8 promoter methylation levels with coronary plaque inflammation.

BACKGROUND AND AIMS: Coronary heart disease (CHD) is a condition that carries a high risk of mortality and is associated with aging. CHD is characterized by the chronic inflammatory response of the coronary intima. Recent studies have shown that the methylation level of blood mononuclear cell DNA is closely associated with adverse events in CHD, but the roles and mechanisms of DNA methylation in CHD remain elusive. METHODS AND RESULTS: In this study, the DNA methylation status within the epigenome of human coronary tissue in the sudden coronary death (SCD) group and control (CON) group of coronary heart disease was analyzed using the Illumina® Infinium Methylation EPIC BeadChip (850 K chip), resulting in the identification of a total of 2553 differentially methylated genes (DMGs). The differentially methylated genes were then subjected to Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis, and significant differential DNA methylation was found. Among the differentially hypomethylated genes were GAL-8, LTF, and RFPL3, while the highly methylated genes were TMEM9B, ANK3, and C6orF48. These genes were mainly enriched in 10 significantly enriched pathways, such as cell adhesion junctions, among which the differentially methylated gene GAL-8 was involved in inflammatory pathway signaling. For functional analysis of GAL-8, we first examined the differences in GAL-8 promoter methylation levels among different subgroups of human coronary tissue in the CON, CHD, and SCD groups using pyrophosphate sequencing. The results revealed reduced GAL-8 promoter methylation levels in the SCD group, while the difference between the CHD and CON groups was not statistically significant (P > 0.05). The reduced GAL-8 promoter methylation level was associated with upregulated GAL-8 expression, which led to increased expression of the inflammatory markers TNF-α, IL-1ß, MCP-1, MIP-2, MMP-2, and MMP-9. This enhanced inflammatory response contributed to the accumulation of foam cells, thickening of the intima of human coronary arteries, and increased luminal stenosis, which promoted the occurrence of sudden coronary death. Next, we found that GAL-8 promoter methylation levels in PBMC were consistent with human coronary tissue. The unstable angina group (UAP) had significantly lower GAL-8 promoter methylation levels than stable angina (SAP) and healthy controls (CON) (P < 0.05), and there was a significant correlation between reduced GAL-8 promoter methylation levels and risk factors for coronary heart disease. These findings highlight the association between decreased GAL-8 promoter methylation and the presence of coronary heart disease risk factors. ROC curve analysis suggests that methylation of the GAL 8 promoter region is an independent risk factor for CHD. In conclusion, our study confirmed differential expression of GAL-8, LTF, MUC4D, TMEM9B, MYOM2, and ANK3 genes due to DNA methylation in the SCD group. We also established the consistency of GAL-8 promoter methylation alterations between human coronary tissue and patient peripheral blood monocytes. The decreased methylation level of the GAL-8 promoter may be related to the increased expression of GAL-8 and the coronary risk factors. CONCLUSIONS: Accordingly, we hypothesized that reduced levels of GAL-8 promoter methylation may be an independent risk factor for adverse events in coronary heart disease.

Stevioside Ameliorates Prenatal Obesity Induced Postpartum Depression: The Potential Role of Gut Barrier Homeostasis.

SCOPE: Postpartum depression and cognitive impairment are the common complications of prenatal obesity. Stevioside is a non-nutritive natural sweetener with antioxidant and anti-inflammatory. However, its effects on depression behaviors and cognitive impairment induced by a high-fat diet (HFD) remain unclear. METHODS AND RESULTS: An 8-week HFD is used to establish a prenatal obesity model in female C57BL/6J mice to explore the improvement effects of stevioside (0.5 mg mL-1 in drinking water) on maternal depression and cognitive dysfunction after weaning. The results demonstrated that stevioside improves behavioral performance of obese maternal mice, and inhibits neuronal damage and 5-hydroxytryptamine (5-HT) abnormality induced by HFD. In addition, stevioside inhibits oxidative stress by reducing malondialdehyde (MDA) and increasing superoxide dismutase (SOD) and glutathione (GSH) activities in the brains of obese maternal mice. Additionally, stevioside improves gut barrier integrity and prevented lipopolysaccharide (LPS) extravasation, and alleviates neuroinflammation. Correlation analysis shows that gut barrier and serum LPS are closely related to behavioral performance and brain biochemical indicators. CONCLUSION: Stevioside is capable to prevent prenatal obesity-induced cognitive and mood disorders by restoring intestinal barrier damage and inhibiting inflammation.

Automated online solid-phase extraction-tandem mass spectrometry detection for simultaneous analysis of acidic and alkaline catecholamines and their metabolites in human urine.

Metabolic dysregulation of catecholamines (CAs) is implicated in various human diseases. Simultaneously analyzing these acidic and alkaline CAs and their metabolites poses a significant challenge for clinical detection. This study introduces an efficient method employing automated online solid-phase extraction coupled with tandem mass spectrometry (aoSPE-MS/MS). The method employs weak cation exchange (WCX) and mixed-mode anion exchange (MAX) adsorbents to fabricate an on-line solid-phase extraction (SPE) column, along with an automated injection and multi-valve switching capabilities. The setup allows for automated extraction and analysis of urine samples in 15 minutes while retaining a wide range of acidic and basic CAs and their metabolites. The applicability of this method was demonstrated by optimising the adsorbent dosage volume, extraction solvent, and extraction rate. The limits of detection (LODs) and limits of quantitation (LOQs) for the 8 CAs and their metabolites were determined using the aoSPE-MS/MS approach, with ranges of 0.0625 ∼ 62.5 ng/mL and 0.125 ∼ 125 ng/mL, respectively. Additionally, assessments were made on the linearity, accuracy, and precision within and between batches, as well as matrix and ionic effects, and spiked recoveries. The study discovered that the aoSPE-MS/MS technique simplifies operation, increases efficiency, saves time, and has low detection and quantification limits when detecting a wide range of acid and alkaline CAs and their metabolites in urine. The study successfully demonstrated the high-throughput and automated detection of the 8 CAs and their metabolites with varying acidity and alkalinity in human urine samples. This method is expected to be a potential powerful tool for clinical detection.