Objective:Lamotrigine (LTG) is a first-line antiseizure medication, yet substantial interindividual variability in therapeutic response remains a major clinical challenge. This study integrated therapeutic drug monitoring (TDM) and metabolomics to explore pharmacokinetic and metabolic factors associated with LTG efficacy in patients with epilepsy. Methods:A total of 141 epilepsy patients receiving LTG therapy were retrospectively enrolled and classified as responders or non‑responders by 6-month seizure frequency reduction. LTG plasma concentrations were measured using HPLC-DAD. Untargeted UPLC-MS/MS metabolomics and multivariate analysis were performed on serum from 10 responders and 10 non-responders with LTG >5.15 mg/L, with differential amino acids quantified. Differential metabolites were further quantified, with a focus on amino acids. Results:Mean LTG plasma concentration was significantly higher in responders (7.55 ± 4.57 mg/L) than non‑responders (4.53 ± 2.94 mg/L; P = 0.001). Receiver operating characteristic (ROC) curve analysis suggested 5.15 mg/L as the optimal cut‑off for discriminating responders from non‑responders, within the conventional ILAE reference range (2.5-15 mg/L). Concomitant use of valproic acid (VPA) increased LTG exposure, was more prevalent among responders, and enabled seizure control at lower LTG doses. Metabolomic analysis in the high‑concentration subgroup revealed a distinct amino acid profile in responders, characterized by higher proline and lower lysine and cystine (P < 0.05). These changes suggest reduced oxidative stress and improved neuronal stability in responders. Conclusion:Higher LTG plasma concentrations were associated with better seizure control, and VPA co‑administration enhances LTG exposure and efficacy. The distinct amino acid profile in high-concentration responders suggests amino acid metabolism and redox balance may modulate LTG efficacy.

The early pathogenic mechanisms underlying human diquat (DQ) poisoning remain incompletely understood. This study integrated retrospective clinical analysis, exploratory serum metabolomics, and in vitro experiments to characterize early metabolic disturbances and investigate potential cell-death mechanisms associated with acute DQ poisoning. Clinical data from 95 patients with acute DQ poisoning were retrospectively analyzed to identify clinical variables associated with outcome. Guided by these findings, exploratory pretreatment serum metabolomics was performed in eight patients with acute DQ poisoning and eight age- and sex-matched healthy controls. Cellular experiments were subsequently conducted in DQ-exposed HepG2 cellsto examine oxidative stress, lipid peroxidation, and ferroptosis-related features using biochemical, molecular, morphological, and inhibitor-based approaches. DQ concentration and markers of multi-organ injury were among the variables most strongly associated with clinical outcome in the random-forest analysis. Serum metabolomic analysis revealed alterations in polyunsaturated fatty acid metabolism, including perturbations in linoleic acid- and α-linolenic acid-related pathways and increased levels of several ω-6 polyunsaturated fatty acids and their oxidation derivatives. Correlation analysis demonstrated associations between these lipid-related metabolites and clinical markers of renal, hepatic, and muscular injury. In HepG2 cells, DQ exposure was associated with ROS accumulation, GSH depletion, lipid peroxidation, increased intracellular Fe²⁺, time-dependent alterations in GPX4 and SLC7A11 expression, and mitochondrial ultrastructural injury, a combination of changes consistent with ferroptosis-related cellular injury. Ferrostatin-1 significantly improved cell viability, whereas Necrostatin-1 showed no comparable protective effect and Z-DEVD-FMK provided less protection than Ferrostatin-1 under the conditions tested. In conclusion, acute DQ poisoning is associated with distinct clinical and metabolic disturbances involving PUFA metabolism. The HepG2 experiments further suggested that ferroptosis-related lipid peroxidation may contribute to DQ-induced cellular injury. These findings support further investigation of lipid peroxidation and ferroptosis-related pathways as potential therapeutic targets in DQ poisoning.

Primary sclerosing cholangitis (PSC) is an autoimmune cholangiopathy characterized by chronic inflammation of the biliary epithelium and periductal fibrosis, with no curative treatment available, and liver transplantation is inevitable for end-stage patients. Human placental mesenchymal stem cell (hpMSC)-derived exosomes have demonstrated the ability to prevent fibrosis, inhibit collagen production and possess immunomodulatory properties in autoimmune liver disease. Here, we prepared hpMSC-derived exosomes (ExoMSC) and further investigated the anti-fibrotic effects and detailed mechanism on PSC based on Mdr2−/− mice and multicellular organoids established from PSC patients. The results showed that ExoMSC ameliorated liver fibrosis in Mdr2−/− mice with significant collagen reduction in the preductal area where Th17 differentiation was inhibited as demonstrated by RNAseq analysis, and the percentage of CD4+IL-17A+T cells was reduced both in ExoMSC-treated Mdr2−/− mice (Mdr2−/−-Exo) in vivo and ExoMSC-treated Th17 differentiation progressed in vitro. Furthermore, ExoMSC improved the hypersecretory phenotype and intercellular interactions in the hepatic Th17 microenvironment by regulating PERK/CHOP signaling as supported by multicellular organoids. Thus, our data demonstrate the anti-fibrosis effect of ExoMSC in PSC disease by inhibiting Th17 differentiation, and ameliorating the Th17-induced microenvironment, indicating the promising potential therapeutic role of ExoMSC in liver fibrosis of PSC or Th17-related diseases.

As the mechanism of paraquat (PQ) poisoning is still not fully elucidated, and no specific treatment has been developed in medical practice, the management of PQ poisoning continues to present a medical challenge. In this study, the objective was to investigate the early metabolic changes in serum metabolism and identify the key metabolic pathways involved in patients with PQ poisoning. Quantitative analysis was conducted to determine the relevant metabolites. Additionally, experiments were carried out in both plasma and cell to elucidate the mechanisms underlying metabolic disorder and cell death in PQ poisoning. The study found that polyunsaturated fatty acids (PUFAs) and their metabolites, such as arachidonic acid (AA) and hydroxy eicosatetraenoic acids (HETEs), were significantly increased by non-enzymatic oxidative reaction. Reactive oxygen species (ROS) production increased rapidly at 2 h after PQ poisoning, followed by an increase in PUFAs at 12 h, and intracellular glutathione, cysteine (Cys), and Fe2+ at 24 h. However, at 36 h later, intracellular glutathione and Cys decreased, HETEs increased, and the expression of SLC7A11 and glutathione peroxidase 4 (GPX4) decreased. Ultrastructural examination revealed the absence of mitochondrial cristae. Deferoxamine was found to alleviate lipid oxidation, and increase the viability of PQ toxic cells in the low dose. In conclusion, unsaturated fatty acids metabolism was the key metabolic pathways in PQ poisoning. PQ caused cell death through the induction of ferroptosis. Inhibition of ferroptosis could be a novel strategy for the treatment of PQ poisoning.

Background. Hundreds of millions of people worldwide suffer from chronic hepatitis B (CHB), making it one of the most serious public health problems. CHB can lead to serious acute-on-chronic liver failure (ACLF). Since ACLF has a high mortality rate, predicting the risk of ACLF is a critical issue in clinical practice. Methods. To investigate the occurrence of ACLF in CHB, we used high-throughput RNA sequencing to detect the expression of a novel endogenous noncoding RNA (circRNA) in healthy controls (HC) as well as CHB and ACLF patients. Differentially expressed circRNAs were selected and analyzed by Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) biological pathway analyses, circRNA-miRNA coexpression networks, and statistical analyses. Results. A total of 21,101 circRNAs were identified in HC, CHB, and ACLF subjects. Compared with the HCs, 4 circRNAs were upregulated and 14 circRNAs were downregulated in ACLF subjects, with a consistent trend in all three groups. GO analysis revealed that regulation of lymphocyte activation and macrophage tolerance induction were the most important biological processes in ACLF progression. KEGG pathway analysis revealed that primary immunodeficiency and NOD-like receptor signaling pathways were the most enriched terms. The circRNA-miRNA coexpression network and statistical analyses showed that circRNA_07734, circRNA_08533, and circRNA_16083 were the most important circRNAs in the process of ACLF. Conclusions. Immune dysfunction and deficiency may play a key role in the development of ACLF, especially circRNA_07734, circRNA_08533, and circRNA_16083.

BACKGROUND:Hepatorenal syndrome (HRS) is a severe complication of decompensated cirrhosis with high mortality. However, few prognostic factors have been identified and studies are urgently needed to facilitate precise treatment.METHODS:Patients with decompensated cirrhosis and acute kidney injury were enrolled from four general hospitals between January 2010 and March 2020. Demographic and laboratory data were compared between surviving and non-surviving patients and also among different levels of HRS severity. COX regression analysis was performed to determine the effect of mean corpuscular hemoglobin concentration (MCHC) on survival of patients with HRS.RESULTS:Out of a total of 1287 patients enrolled, 325 patients were analyzed. MCHC was significantly higher in non-survivors than in survivors, and in patients with more serious disease, defined as failure of organ systems. The hazard ratio (HR) of mortality was 1.17, 1.18 and 1.11, when adjusted by the crude model, model 1 and model 2, respectively. When MCHC was converted to a categorical variable based on the quartile of MCHC, the HR for the highest quartile of MCHC was 2.11 (95% CI: 1.45-3.06, P <0.05) compared to the lowest quartile of MCHC in the crude model, and when adjusted for age and sex (model 1), the HR was 2.20 (95% CI: 1.52-3.20, P <0.05). In model 2, which was adjusted for complex characteristics, the HR was 1.77 (95% CI: 1.17-2.68, P <0.05). The results of Kaplan-Meier curves were consistent with those from Cox regression analysis.CONCLUSIONS:Higher MCHC was associated with worse prognosis in HRS.

Mesenchymal stem cells (MSCs) have attracted interest for their potential to alleviate liver injury. Here, the protective effect of MSCs on carbon tetrachloride (CCl4)-induced acute liver injury (ALI) was investigated. In this study, we illustrated a novel mechanism that ferroptosis, a newly recognized form of regulated cell death, contributed to CCl4-induced ALI. Subsequently, based on the in vitro and in vivo evidence that MSCs and MSC-derived exosomes (MSC-Exo) treatment achieved pathological remission and inhibited the production of lipid peroxidation, we proposed an MSC-based therapy for CCl4-induced ALI. More intriguingly, treatment with MSCs and MSC-Exo downregulated the mRNA level of prostaglandin-endoperoxide synthase 2 (Ptgs2) and lipoxygenases (LOXs) while it restored the protein level of SLC7A11 in primary hepatocytes and mouse liver, indicating that the inhibition of ferroptosis partly accounted for the protective effect of MSCs and MSC-Exo on ALI. We further revealed that MSC-Exo-induced expression of SLC7A11 protein was accompanied by increasing of CD44 and OTUB1. The aberrant expression of ubiquitinated SLC7A11 triggered by CCl4 could be rescued with OTUB1-mediated deubiquitination, thus strengthening SLC7A11 stability and thereby leading to the activation of system XC- to prevent CCl4-induced hepatocyte ferroptosis. In conclusion, we showed that MSC-Exo had a protective role against ferroptosis by maintaining SLC7A11 function, thus proposing a novel therapeutic strategy for ferroptosis-induced ALI.

Background: Ferroptosis, a newly recognized form of regulated cell death, was recently identified as a novel therapeutic target in tissue injury. Various studies have shown that administration of mesenchymal stem cells (MSC) is a promising therapeutic approach to repair liver injury. However, the role of ferroptosis in acute liver injury (ALI) and MSC-based therapy is unknown. Results: Here we found that CCl4 induced elevated lipid reactive oxygen species (lipid-ROS) and mRNA levels of putative molecular markers of ferroptosis such as Ptgs2 and LOX genes. CCl4 also downregulated the xCT protein levels resulting in the accumulation of lipid peroxidation and ferroptosis. MSC transplantation largely abolished CCl4-induced ferroptosis. Furthermore, the protective effects of MSC against ferroptosis were closely correlated with exosome-mediated stabilization of xCT. Administration of MSC-Exo restored the xCT protein level, decreased the elevated lipid-ROS level and Ptgs2 and LOX mRNA levels, and promoted liver restoration by inhibiting ferroptosis. Interestingly, in ALI mouse livers after MSC-Exo treatment, exosome-induced recovery of xCT protein was accompanied by upregulation of CD44 and OTUB1. The level of ubiquitinated xCT upregulated by CCl4 was significantly downregulated by OTUB1-mediated deubiquitination, and strong interactions of xCT with OTUB1 and CD44 proteins were detected. Conclusions: Taken together, our data indicate that MSC-Exo has a protective role against ferroptosis by maintaining xCT function. This provides a novel therapeutic strategy for ferroptosis-induced ALI.

BACKGROUND Hepatorenal syndrome (HRS) is a severe complication of cirrhosis with high mortality, which necessitates accurate clinical decision. However, studies on prognostic factors and scoring systems to predict overall survival of HRS are not enough. Meanwhile, a multicenter cohort study with a long span of time could be more convincing. AIM To develop a novel and effective prognostic model for patients with HRS and clarify new prognostic factors. METHODS We retrospectively enrolled 1667 patients from four hospitals, and 371 eligible patients were finally analyzed to develop and validate a novel prognostic model for patients with HRS. Characteristics were compared between survivors and non-survivors, and potential prognostic factors were selected according to the impact on 28-d mortality. Accuracy in predicting 28-d mortality was compared between the novel and other scoring systems, including Model for End-Stage Liver Disease (MELD), Chronic Liver Failure-Sequential Organ Failure Assessment (CLIF-SOFA), and Chinese Group on the Study of Severe Hepatitis B-Acute-on-Chronic Liver Failure (COSSH-ACLF). RESULTS Five prognostic factors, comprised of gender, international normalized ratio, mean corpuscular hemoglobin concentration, neutrophil percentage, and stage, were integrated into a new score, GIMNS; stage is a binary variable defined by the number of failed organs. GIMNS was positively correlated with MELD, CLIF-SOFA, and COSSH-ACLF. Additionally, it had better accuracy [area under the receiver operating characteristic curve (AUROC): 0.830] than MELD (AUROC: 0.759), CLIF-SOFA (AUROC: 0.767), and COSSH-ACLF (AUROC: 0.759) in the derivation cohort (P < 0.05). It performed better than MELD and CLIF-SOFA in the validation cohort (P < 0.050) and had a higher AUROC than COSSH-ACLF (P = 0.122). CONCLUSION We have developed a new scoring system, GIMNS, to predict 28-d mortality of HRS patients. Mean corpuscular hemoglobin concentration and stage were first proposed and found to be related to the mortality of HRS. Additionally, the GIMNS score showed better accuracy than MELD and CLIF-SOFA, and the AUROC was higher than that of COSSH-ACLF.

OBJECTIVES:Acute lung injury (ALI) not only affects pulmonary function but also leads to intestinal dysfunction, which in turn contributes to ALI. Mesenchymal stem cell (MSC) transplantation can be a potential strategy in the treatment of ALI. However, the mechanisms of synergistic regulatory effects by MSCs on the lung and intestine in ALI need more in-depth study. MATERIALS AND METHODS:We evaluated the therapeutic effects of MSCs on the murine model of lipopolysaccharide (LPS)-induced ALI through survival rate, histopathology and bronchoalveolar lavage fluid. Metagenomic sequencing was performed to assess the gut microbiota. The levels of pulmonary and intestinal inflammation and immune response were assessed by analysing cytokine expression and flow cytometry. RESULTS:Mesenchymal stem cells significantly improved the survival rate of mice with ALI, alleviated histopathological lung damage, improved intestinal barrier integrity, and reduced the levels of inflammatory cytokines in the lung and gut. Furthermore, MSCs inhibited the inflammatory response by decreasing the infiltration of CD8+ T cells in both small-intestinal lymphocytes and Peyer's patches. The gut bacterial community diversity was significantly altered by MSC transplantation. Furthermore, depletion of intestinal bacterial communities with antibiotics resulted in more severe lung and gut damages and mortality, while MSCs significantly alleviated lung injury due to their immunosuppressive effect. CONCLUSIONS:The present research indicates that MSCs attenuate lung and gut injury partly via regulation of the immune response in the lungs and intestines and gut microbiota, providing new insights into the mechanisms underlying the therapeutic effects of MSC treatment for LPS-induced ALI.

BACKGROUND As human placenta-derived mesenchymal stem cells (hP-MSCs) exist in a physiologically hypoxic microenvironment, various studies have focused on the influence of hypoxia. However, the underlying mechanisms remain to be further explored. AIM The aim was to reveal the possible mechanisms by which hypoxia enhances the proliferation of hP-MSCs. METHODS A hypoxic cell incubator (2.5% O2) was used to mimic a hypoxic microenvironment. Cell counting kit-8 and 5-ethynyl-20-deoxyuridine incorporation assays were used to assay the proliferation of hP-MSCs. The cell cycle was profiled by flow cytometry. Transcriptome profiling of hP-MSCs under hypoxia was performed by RNA sequencing. CD99 mRNA expression was assayed by reverse transcription-polymerase chain reaction. Small interfering RNA-mediated hypoxia-inducible factor 1α (HIF-1α) or CD99 knockdown of hP-MSCs, luciferase reporter assays, and the ERK1/2 signaling inhibitor PD98059 were used in the mechanistic analysis. Protein expression was assayed by western blotting; immunofluorescence assays were conducted to evaluate changes in expression levels. RESULTS Hypoxia enhanced hP-MSC proliferation, increased the expression of cyclin E1, cyclin-dependent kinase 2, and cyclin A2, and decreased the expression of p21. Under hypoxia, CD99 expression was increased by HIF-1α. CD99-specific small interfering RNA or the ERK1/2 signaling inhibitor PD98059 abrogated the hypoxia-induced increase in cell proliferation. CONCLUSION Hypoxia promoted hP-MSCs proliferation in a manner dependent on CD99 regulation of the MAPK/ERK signaling pathway in vitro.

Exosomal microRNA (miRNA) secretion has been characterized as a vital factor in intercellular communication among cancer cells. However, little is known about cancer-secreted miRNAs specifically involved in metastasis of colorectal cancer (CRC). Here, we found that exosomes derived from metastatic CRC cell line SW620 promoted migration, invasion, and epithelial-mesenchymal transition (EMT) of CRC cells. The profiling of exosome miRNAs revealed that microRNA (miR)-335-5p was highly expressed in exosomes from metastatic SW620 cells compared to those derived from primary SW480 cells. miR-335-5p was transmitted from metastatic SW620 cells to CRC cells via exosomes and promoted migration, invasion, and EMT of CRC cells. Moreover, exosome-transmitted miRNA-335-5p promotes CRC cell invasion and metastasis by facilitating EMT via targeting RAS p21 protein activator 1 (RASA1). Overexpression of RASA1 abolished the promotive effects of exosomal miR-335-5p on CRC cell migration, invasion, and EMT. Collectively, our data revealed that exosomal miR-335-5p derived from metastatic CRC cells promotes CRC cell invasion and metastasis by facilitating EMT via targeting RASA1, which may serve as a potential therapeutic target for CRC metastasis.

Rationale: Acute lung injury (ALI)-recruited mononuclear phagocytes play a pivotal role in lung injury and repair. This study investigated the types of recruited mononuclear phagocytes and the immunotherapeutic effects of allograft mesenchymal stem cells (MSCs) in a mouse model of lipopolysaccharide (LPS)-induced ALI. Methods: C57BL/6 mice were orotracheally instilled with LPS (20 mg/kg). Compact bone-derived MSCs were administered orotracheally 4 h after LPS inhalation. Mononuclear phagocytes recruited in the lung tissues were characterized at different timepoints by high-dimensional analysis including flow cytometry, mass cytometry, and single-cell RNA sequencing. Results: Eight mononuclear phagocyte subsets recruited to LPS-challenged lungs were precisely identified. On day 3 after LPS administration, both Ly6ChiCD38+ and Ly6ClowCD38+ monocytes were recruited into acutely injured lungs, which was associated with increased secretion of neutrophil chemokines. Ly6ChiCD38+ monocytes differentiated into M1 macrophages on day 3, and subsequently differentiated into CD38+ monocyte-derived dendritic cells (mo-DCs) on day 7, while Ly6ClowCD38+ monocytes differentiated into CD11b+CD38+ DCs on day 7. When ALI mice were treated with MSCs, the mortality significantly reduced. Notably, MSCs reduced the amount of M1 macrophages and reduced the secretion of neutrophil chemokines on day 3. Furthermore, MSCs reduced the number of CD38+ mo-DCs and CD11b+CD38+ DCs on day 7, suppressing the antigen presentation process. Recruited mononuclear phagocyte subsets with a high level of CD38 exhibited an activated phenotype and could secrete higher levels of cytokines and chemokines. Conclusions: This study characterized the dynamic functions and phenotypes of recruited mononuclear phagocytes in ALI mice and MSC-treated ALI mice.

Background Millions of adults have been reported with hyperlipemia in the world. It is still unclear whether the plasma level of essential amino acids (AAs) will be influenced by the hyperlipemia. This study was aimed to investigate the AAs levels and the underlying metabolic relationship in hyperlipidemic subjects. Methods An ultra-high performance liquid chromatography-tandem mass spectrometric (UPLC-MS/MS) method was developed for the determination of phenylalanine (Phe), valine (Val), histidine (His), tryptophan (Trp), and methionine (Met). Plasma samples (100 μL) were precipitated by acetonitrile (300 μL) and analyzed on a BEH C18 (2.1 mm × 100 mm, 1.7 μm) column at 40 °C by gradient elution. The mobile phase composed of 0.1% formic acid and acetonitrile was used with flow rate at 0.2–0.4 ml/0-3 min. Five AAs were determined at positive electrospray ionization (ESI+) at m/z 118.1/72.1 (Val), 150.12/104.02(Met), 156.06/110.05(His), 166.1/120.1(Phe), and 205.2/188.02 (Trp). A total of 75 healthy subjects and 83 hyperlipidemic subjects, who had blood routine test and plasma lipid test were determined by developed UPLC-MS/MS. Results It was shown that there was good linearity for Val, Met, His, Phe, and Trp within 1–100 μg/mL. The relative standard deviations of precision and accuracy were all within 15%. The level of Val, Phe, Trp, His, and Met were 35.34 ± 15.64, 22.72 ± 9.13, 17.23 ± 4.94, 16.78 ± 13.64, and 6.24 ± 1.97 μg/mL in healthy subjects, while they were 38.04 ± 16.70, 22.41 ± 8.45, 15.62 ± 5.77, 18.35 ± 14.49, and 6.21 ± 1.97 μg/mL in hyperlipidemic subjects respectively. The Spearman’s correlations analysis showed that there were high correlations between Val, Phe, Trp, His, Met and triglyceride in healthy subjects. While, those correlations decreased in hyperlipemia cases. Conclusion A convenient and sensitive method for simultaneous determination of Val, Phe, Trp, His, and Met in human plasma was developed. There was a high correlation between Val, Phe, Trp, His, Met and triglyceride. Hyperlipemia influences the metabolic balance of His, Phe, Trp, Met and Val.

Background: Hundreds of millions of people worldwide suffer from chronic hepatitis B (CHB), which is thus among the most serious public health problems. CHB can lead to serious acute chronic liver failure (ACLF). Since ACLF has a very high mortality rate, prediction of ACLF risk is a critical issue in clinical practice. Methods: To investigate the development of ACLF from CHB, circular RNA (circRNA), a novel type of endogenous non-coding RNA, was examined in healthy control (HC) subjects and CHB and ACLF patients using high-throughput RNA sequencing technology. Differentially expressed circRNAs were selected and analyzed by Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) biological pathway analyses, circRNA-miRNA co-expression networks, and statistical analyses. Results: A total of 21,101 circRNAs were identified in HC, CHB and ACLF subjects. Compared with the HCs, 4 up-regulated and 14 down-regulated circRNAs were identified in ACLF subjects, which exhibited a consistent trend of change in the three groups. GO analysis revealed that regulation of lymphocyte activation and macrophage tolerance induction were the most important biological processes in ACLF progression. KEGG pathway analysis revealed that primary immunodeficiency and NOD-like receptor signaling pathways were the most enriched terms. The circRNA-miRNA co-expression network and statistical analyses showed that circRNA_07734, circRNA_08533, and circRNA_16083 were the most important circRNAs in the process of ACLF. Conclusions: Immune dysfunction and immunodeficiency may be critical in the development of ACLF, which results in aberrant expression of circRNAs, especially circRNA_07734, circRNA_08533, and circRNA_16083.

Amino acids (AAs) are abundant in seminal fluid and play an important role in the reproduction process. However, the functional significance of AAs in seminal is not clear. This study was designed to profile the AAs in seminal fluid of asthenozoospermia (AS) and oligospermia (OL) patients and evaluate its potential diagnostic value. To determine the true function of AAs in seminal fluid, an ultra high performance liquid chromatography-mass spectrometry (UPLC-MS/MS) was developed and validated. The seminal fluid samples collected from 76 AS patients, 58 OL patients, and 76 healthy subjects were determined by UPLC-MS/MS and analyzed by fisher discriminant and receiver operating characteristic (ROC). The results showed that, the AAs levels were dramatically decreased in AS and OL patients compared with healthy subjects. What's more there were significant variation in the AAs profile of AS and OL patients. Specifically, some polar AAs decreased, while nonpolar AAs increased in OL patients. Based on the level of AAs, 93.4 % of original and 90.9 % of cross -validated grouped cases were correctly classified between AS patients, OL patients, and healthy subjects. The further ROC analysis shows that glutamic acid, tyrosine, aspartic acid, proline, and tryptophan have the diagnostic value (P < 0.01) in the three groups. Moreover, aspartic acid and glutamic acid exhibit a high correlation (Correlation Coefficient >0.9) in AS, OL, and healthy subjects. In conclusion, there are significant variation in AAs profile in seminal plasma of AS and OL patients with high accuracy in fisher classification. The associations between the levels of AAs in seminal fluid of AS, OL, and healthy subjects may have clinical applications in providing valuable diagnostic indicator. (C) 2020 Published by Elsevier B.V.

A new compound, bis(4-hydroxybenzyl)ether mono-β-L-galactopyranoside (1), was isolated from the rhizome of Gastrodia elata Blume. Its structure was elucidated using extensive spectroscopic analysis, including 1D and 2D NMR, HR-ESI-TOF-MS, and chemical derivatization. Compound 1 extended the replicative lifespan of K6001 and the chronological lifespan of YOM36 yeast strains. To understand the mechanism of action, oxidative stress assessment, reactive oxygen species (ROS) and malondialdehyde (MDA) levels, catalase (CAT) and total glutathione peroxidase (GPx) activity assays, and replicative lifespan assay of sod1, sod2, uth1, and skn7 yeast mutant strains were performed. Results indicated the significant increase in the survival rate of yeast under oxidative stress after treatment with 1. ROS and MDA levels were reduced significantly. Meanwhile, the activity of CAT and GPx was significantly increased. The lifespan of sod1, sod2, uth1, and skn7 mutants of K6001 was not affected by 1. Furthermore, we investigated the gene expression related to longevity after administrating 1. The significant increase of Sir2 and reduction of Uth1 gene expression in the 1-treated group were observed. These results indicated that antioxidative stress played an important role in the antiaging effect of 1; Sir2 and Uth1 genes were involved in antiaging effects of 1.

Background: Omega-3 polyunsaturated fatty acids (PUFAs), including alpha-linolenic acid (ALA), eicosapentaenoic acid (EPA), docosahexaenoic acid (DHA), and docosapentaenoic acid (DPA), play critical roles in numerous biochemical reactions. Our aim is to develop a rapid and sensitive method for simultaneous determination of ALA, EPA, DHA and DPA in the plasma of hyperlipidemic and normolipidemic subjects. Methods: An ultra-high-performance liquid chromatography-tandem mass spectrometric (UPLC-MS/MS) method of ALA, EPA, DHA, and DPA was developed with chlorzoxazone as the internal standard (IS). The analytes were separated on an Acquity BEH C18 column (2.1 mm x 100 mm, 1.7 mu m) with gradient elution by acetonitrile and 0.1% ammonia water. ALA, EPA, DHA, DPA, and IS were determined by negative electrospray ionization (ESI-) with multiple reaction monitoring (MRM) at m/z 277.42/259.05, 301.20/257.00, 327.30/283.40, 329.24/285.32, and 168.03/132.02. A total of 80 normolipidemic subjects and 83 hyperlipidemic subjects, who underwent testing for plasma lipids, liver and kidney functions, and blood routine blood test (BRT), were enrolled. Results: There was good linearity for ALA within 1-10 mu g/mL, and EPA, DHA and DPA were within 0.125-10 mu g/mL. The relative standard deviation (RSD) of precision was below 15%. The concentrations of ALA, EPA, DHA and DPA were 3.47 +/- 2.58, 0.41 +/- 0.26, 2.93 +/- 1.39 and 0.25 +/- 0.21 mu g/mL, respectively, in normolipidemic subjects, increasing to 4.14 +/- 3.71, 0.57 +/- 0.46, 3.43 +/- 2.13, 0.27 +/- 0.25 mu g/mL, respectively in hyperlipidemic subjects. Among them, only the EPA concentration was significantly different between two groups. There was a high correlation between ALA, EPA, DHA and DPA. Conclusion: We developed a rapid and sensitive method for simultaneously determination of ALA, EPA, DHA and DPA in hyperlipidemic and normolipidemic subjects. In hyperlipidemic and normolipidemic subjects, concentrations of ALA were highest, followed by DHA, EPA and DPA; there were high degrees of correlation between each value.

The budding yeast Saccharomyces cerevisiae has been used as a model organism for the basic mechanism of aging, which provides useful assay systems for measuring both replicative and chronological lifespans. In the course of our screening program for substances that extend replicative lifespan, cucurbitacin B (CuB) was found as a hit compound from a compound library, which contains cerebrosides, phenols, sesquiterpenoid, triterpenoids, and sterols isolated from natural products by our research group. Importantly, it prolonged not only the replicative lifespan but also the chronological lifespan in yeast. CuB increased ATG32 gene expression, suggesting that CuB induces autophagy. Indeed, the GFP signal generated from the cleavage of GFP-Atg8, which is a signature of autophagy, was increased upon CuB treatment. On the other hand, CuB failed to increase the chronological lifespans when either ATG2 or ATG32, essential autophagy genes, was deleted, indicating that the lifespan extension by CuB depends on autophagy induction. Furthermore, CuB significantly increased superoxide dismutase (Sod) activity and the survival rate of yeast under oxidative stress, while it decreased the amount of reactive oxygen species (ROS) and malondialdehyde (MDA) production, indicating that CuB has activity to antagonize oxidative stress. Additionally, CuB did not affect replicative lifespans of sod1, sod2, uth1, and skn7 mutants with the K6001 background, indicating that aging-related genes including SOD1, SOD2, UTH1, and SKN7 participate in the antiaging effect of CuB. These results suggest that CuB exerts antiaging activity by regulating autophagy, ROS, antioxidative ability, and aging-related genes. Finally, we discuss the possible intracellular targets of CuB based on the phenotypic comparison between the CuB and global gene deletion databases.

The biochemical indexes are used to assess the hepatic and renal function of paraquat (PQ) poisoning patients. However, these indexes correlated with the prognosis of patients are unidentified. This paper aims to explore useful indexes from biochemical tests and to identify their predictive value. A total of 101 PQ poisoning patients including 51 dead patients and 50 survived patients is involved in this study. The biochemical indexes of PQ poisoning patients in different status are collected and analyzed by the independent-sample test. After that, Fisher scores feature selection is applied to screen prognostic factors from biochemical indexes. Based on the results of Fisher selection, an effective extreme learning machine (ELM) is applied to diagnose the prognosis status of PQ poisoning patients. The created ELM method is rigorously evaluated for accuracy, sensitivity, and specificity. The results show that there is statistical significance between dead and survived people in biochemical indexes (P<;0.01). Feature selection revealed that direct bilirubin, alanine aminotransferase (ALT), total bilirubin, aspartate aminotransferase (AST), the ratio of ALT/AST, and creatinine are the most crucial indexes, which correlated with the prognosis of PQ poisoning. The maximal classification accuracy is 79.6% when these six indexes are selected as the dataset. In conclusion, the biochemical test is related to the prognosis of PQ poisoning patients. It provides a new method for prognosis of PQ poisoning with feature selection and ELM model.