Pulsed rise and growth of the Tibetan Plateau to its northern margin since ca. 30 Ma.

The onset of mountain building along margins of the Tibetan Plateau provides a key constraint on the processes by which the high topography in Eurasia formed. Although progressive expansion of thickened crust underpins most models, several studies suggest that the northern extent of the plateau was established early, soon after the collision between India and Eurasia at ca. 50 Ma. This inference relies heavily on the age and provenance of Cenozoic sediments preserved in the Qaidam basin. Here, we present evidence in the northern plateau for a considerably younger inception and evolution of the Qaidam basin, based on magnetostratigraphies combined with detrital apatite fission-track ages that date the basin fills to be from ca. 30 to 4.8 Ma. Detrital zircon-provenance analyses coupled with paleocurrents reveal that two-stage growth of the Qilian Shan in the northeastern margin of the Tibetan Plateau began at ca. 30 and at 10 Ma, respectively. Evidence for ca. 30 and 10 to 15 Ma widespread synchronous deformation throughout the Tibetan Plateau and its margins suggests that these two stages of outward growth may have resulted from the removal of mantle lithosphere beneath different portions of the Tibetan Plateau.

Challenges with the Synthesis of a Macrocyclic Thioether Peptide: From Milligram to Multigram Using Solid Phase Peptide Synthesis (SPPS).

We describe an optimization and scale-up of the 45-membered macrocyclic thioether peptide BMS-986189 utilizing solid-phase peptide synthesis (SPPS). Improvements to linear peptide isolation, macrocyclization, and peptide purification were demonstrated to increase the throughput and purification of material on scale and enabled the synthesis and purification of >60 g of target peptide. Taken together, not only these improvements resulted in a 28-fold yield increase from the original SPPS approach, but also the generality of this newly developed SPPS purification sequence has found application in the synthesis and purification of other macrocyclic thioether peptides.

Validation of hyperthermia as an enhancer of chemotherapeutic efficacy: insights from a bladder cancer organoid model.

OBJECTIVE: This study aimed to evaluate the combined efficacy of hyperthermia and chemotherapy using a bladder cancer organoid model and to explore hyperthermia-related molecular pathways. METHOD: Tumor organoids were generated by embedding RT4 bladder cancer cells into Matrigel. The resulting organoids were treated with pirarubicin or gemcitabine at 37 °C or 42 °C. Proliferation was determined by Ki67 immunofluorescence staining, and apoptosis was assessed using a TdT-mediated dUTP nick end labeling (TUNEL) assay. RNA sequencing was used to identify the differentially expressed genes. RESULTS: Bladder cancer organoids were successfully established and exhibited robust proliferative abilities. Treatment with gemcitabine or pirarubicin under hyperthermic conditions caused pronounced structural damage to the organoids and increased cell death compared to that in the normothermically treated group. Furthermore, Ki67 labeling and TUNEL assays showed that the hyperthermia chemotherapy group showed a significantly reduced proliferation rate and high level of apoptosis. Finally, RNA sequencing revealed the IFN-γ signaling pathway to be associated with hyperthermia. CONCLUSION: Overall, hyperthermia combined with chemotherapy exerted better therapeutic effects than those of normothermic chemotherapy in grade 1-2 non-muscle-invasive bladder cancer, potentially through activation of the IFN-γ-JAK-STAT pathway.

Application of contrast-enhanced ultrasound in diagnosis and grading of bladder urothelial carcinoma.

PURPOSE: To explore the application of contrast-enhanced ultrasound (CEUS) for the diagnosis and grading of bladder urothelial carcinoma (BUC). METHODS: The results of a two-dimensional ultrasound, color Doppler ultrasound and CEUS, were analyzed in 173 bladder lesion cases. The ultrasound and surgical pathology results were compared, and their diagnostic efficacy was analyzed. RESULTS: There were statistically significant differences between BUC and benign lesions in terms of color blood flow distribution intensity and CEUS enhancement intensity (both P < 0.05). The area under the time-intensity curve (AUC), rising slope, and peak intensity of BUC were significantly higher than those of benign lesions (all P < 0.05). The H/T (height H / basal width T)value of 0.63 was the critical value for distinguishing high- and low-grade BUC, had a diagnostic sensitivity of 80.0% and a specificity of 60.0%. CONCLUSION: The combination of CEUS and TIC can help improve the diagnostic accuracy of BUC. There is a statistically significant difference between high- and low-grade BUC in contrast enhancement intensity (P < 0.05); The decrease of H/T value indicates the possible increase of the BUC grade.

Childhood maltreatment and association with trajectories of depressive symptoms among older adults: a longitudinal study in China.

OBJECTIVES: Childhood maltreatment has long-lasting effects on mental health. Existing evidence suggests that trajectories of depressive symptoms vary among individuals; however, little is known about how childhood maltreatment shapes these trajectory patterns. Therefore, this study investigated the impacts of childhood maltreatment on eight-year depressive trajectories among Chinese older adults. METHOD: Five waves of longitudinal data from the China Health and Retirement Longitudinal Study were utilized. Growth Mixture Modelling was performed to identify distinct trajectories of depressive symptoms, and multinomial logistic regression was conducted to explore the associations between these trajectories and childhood maltreatment. RESULTS: Four trajectories of depressive symptoms were identified: the 'no symptoms' class (61.83%), the 'increasing symptoms' class (14.49%), the 'decreasing symptoms' class (16.44%), and the 'chronic symptoms' class (7.24%). Older adults who experienced childhood physical abuse were more likely to be in the 'chronic symptoms' class than in the 'no symptoms' class, whereas emotional neglect did not show a significant association with three problematic trajectories. CONCLUSION: This study provides empirical evidence that childhood physical abuse increases the likelihood of developing chronic depressive symptoms in later life. To mitigate this risk, it is crucial to institute comprehensive treatment plans that incorporate trauma-informed care principles, employ evidence-based therapies specifically designed to address the long-term effects of abuse, and prioritize regular screening and assessment of mental health among older adults.

Platinum Species on Oxygen Vacancy-Rich Titania for Efficient Basic Electrocatalytic Hydrogen Evolution.

Oxygen vacancy-rich titania is a promising support for enhancing the hydrogen evolution reaction (HER). This work innovatively loaded Pt nanoparticles on oxygen vacancy-rich TiO2 (Pt/Vo-TiO2) in situ by using a photocatalytic device. The synthesis conditions are mild, do not require high temperatures and strong reducing agents, and can avoid the accumulation of platinum species. X-ray photoelectron spectroscopy (XPS) and X-ray absorption spectrometry (XAS) verified the synergistic effect of Pt species and oxygen vacancies on the progress of the reaction kinetics, where the Pt particles exposed by the in situ synthesis functioned as reaction sites in the electrocatalytic hydrogen evolution. Based on this, Pt/Vo-TiO2 exhibits excellent electrocatalytic performance with an overpotential of only 56 mV at a current density of 10 mA cm-2 and a Tafel slope of only 73.5 mV dec-1. This work provides a new strategy for designing highly efficient HER catalysts.

Study on an Animal Model of Seawater Immersion Injury Following Hemorrhagic Shock.

INTRODUCTION: To establish an animal model of delayed intravenous resuscitation following seawater immersion after hemorrhagic shock (HS). METHODS: Adult male SD rats were randomly divided into three groups: group NI (HS with no immersion), group SI (HS with skin immersion), and group VI (HS with visceral immersion). Controlled HS in rats was induced by withdrawing 45% of the calculated total blood volume within 30 min. In SI group, immediately after blood loss, 0.5 cm below the xiphoid process was immersed in artificial seawater, at (23 ± 1) °C, for 30 min. In VI group, the rats were performed by laparotomy and the abdominal organs were immersed in (23 ± 1) °C seawater for 30 min. Two hours after seawater immersion, the extractive blood and lactated Ringer's solution were delivered intravenously. The mean arterial pressure (MAP), lactate, and other biological parameters were investigated in different time points. The survival rate of 24 h after HS was recorded. RESULTS: After seawater immersion following HS, MAP and abdominal viscera blood flow decreased significantly, and the plasma levels of lactate and the organ function parameters were increased than the baseline. The above changes in VI group were more serious than those in SI and NI group, especially in myocardial and small intestine damage. The hypothermia, hypercoagulation, and metabolic acidosis were also observed after seawater immersion; the injury was more severely in VI group than that of SI group. However, the plasma levels of sodium, potassium, chlorine, and calcium in VI group were significantly higher than those before injury and in the other two groups. In the VI group, the level of plasma osmolality in instant, 2 h, and 5 h after immersion was 111%, 109%, and 108% of the SI group, respectively, all P < 0.01. The 24-h survival rate of VI group was 25%, which was significantly lower than that of SI group (50%) and NI group (70%), P < 0.05. CONCLUSIONS: The model fully simulated the key damage factors and field treatment conditions, reflected the effects of low temperature and hypertonic damage caused by seawater immersion on the severity and prognosis of naval combat wounds, and provided a practical and reliable animal model for the study of field treatment technology of marine combat shock.

Association between increased serum interleukin-8 levels and improved cognition in major depressive patients with SSRIs.

BACKGROUND: The effect of neuroinflammatory cytokines on cognitive deficits in patients with major depressive disorder (MDD) can be altered by selective serotonin reuptake inhibitors (SSRIs). This study aimed to examine serum interleukin-8 (IL-8) levels, cognitive function, and their associations in MDD patients with SSRIs. METHODS: Thirty SSRI-treated MDD patients and 101 healthy controls were recruited for this study. We examined cognitive performance using the Repeatable Battery for the Assessment of Neuropsychological Status (RBANS) and serum IL-8 levels using the Human Inflammatory Cytokine Cytometric Bead Array in both cases and controls. RESULTS: The RBANS test scores were significantly lower in MDD patients with SSRIs than in healthy controls after controlling for covariates (all p < 0.001). Serum levels of IL-8 were higher in MDD patients with SSRIs than in healthy controls after adjusting for covariates (F = 3.82, p = 0.05). Serum IL-8 levels were positively correlated with sub-scores of delayed memory (r = 0.37, p = 0.04) and visuospatial/constructional (r = 0.43, p = 0.02) in MDD patients with SSRIs but not in in healthy controls (delayed memory score: r = -0.12, p = 0.24; visuospatial/constructional score: r = 0.02, p = 0.81). CONCLUSIONS: Our findings suggested that increased serum IL-8 level might not only be involved in the MDD psychopathology or the use of SSRIs but also correspond to improving MDD delayed memory and visuospatial/constructional function.

Evaluation of the stiffness of normal cervix and its change with different factors using transvaginal two-dimensional shear wave elastography under strict quality control.

BACKGROUND: The usefulness of transvaginal two-dimensional shear wave elastography (2D SWE) for cervical lesions is still uncertain. This study was to explore the value of transvaginal 2D SWE in the evaluation of the stiffness of normal cervix and its change with different factors under strict quality control (QC). METHODS: Two hundred patients with normal cervix were included in this study and were examined using quantitative 2D SWE to evaluate cervical stiffness and its change with different factors under strict QC. RESULTS: Intra-observer concordance of transvaginal 2D SWE parameters in midsagittal planes were acceptable with intraclass correlation coefficients higher than 0.5. Transvaginal 2D SWE parameters were significantly higher than the corresponding transabdominal parameters. 2D SWE parameters of internal cervical os were significantly higher than the corresponding parameters of external cervical os in a transvaginal midsagittal plane. 2D SWE parameters of external cervical os increased significantly over 50 years old, while these parameters of internal cervical os didn't change significantly with increasing age. 2D SWE parameters of internal cervical os of horizontal position cervix were significantly higher than those of vertical position cervix. SWE parameters of normal cervix did not change according to different menstrual cycles, parities and human papilloma virus test results. CONCLUSIONS: Transvaginal 2D SWE under strict QC could provide quantitative, repeatable and reliable cervical stiffness information. Internal cervical os was stiffer than external cervical os. Menstrual cycles, parities and human papilloma virus test results wouldn't affect cervical stiffness. However, age and cervical positions should be taken into condition while interpreting 2D SWE results of cervical stiffness.

Association between disability in activities of daily living and phase angle in hemodialysis patients.

BACKGROUND: Disability in activities of daily living (ADL) significantly increases the risk of mortality among patients undergoing hemodialysis. Malnutrition and decreased exercise capacity are closely correlated with ADL disability. Phase angle (PhA) has been proposed as a measure of nutritional status and exercise capacity. This study aims to investigate the prevalence of ADL disability in hemodialysis patients and its association with PhA. METHODS: A prospective, observational study was conducted, involving hemodialysis patients treated between November 2019 and January 2020 in an affiliated hospital of Chinese university. ADL was measured using both basic ADL (BADL) scales and instrumental ADL (IADL) scales. PhA measurements were obtained using a BIA device while the patients were in the supine position after dialysis. RESULTS: A total of 237 hemodialysis patients with a mean age of 60.01 ± 13.55 years were included in this study. The prevalence of disability in ADL was 43.5%. Multivariable analysis results showed a robust association between low PhA and disability in both BADL and IADL (for each unit decrease in PhA: odds ratio 4.83 [95% CI: 2.56-9.0], and 3.57 [95% CI: 2.14-5.95], respectively). The optimal cut-off values of PhA for disability in BADL and IADL were 4.8 and 5.4, with the area under the ROC curve (AUC) were 0.783 (0.727, 0.835) and 0.799 (0.743, 0.848), respectively. CONCLUSIONS: Low PhA is strongly associated with disability in ADL in hemodialysis patients. These findings suggest that PhA may serve as a potentially objective measure of ADL disability in hemodialysis patients.

Homocysteine accelerates hepatocyte autophagy by upregulating TFEB via DNMT3b-mediated DNA hypomethylation.

Autophagy plays a critical role in the physiology and pathophysiology of hepatocytes. High level of homocysteine (Hcy) promotes autophagy in hepatocytes, but the underlying mechanism is still unknown. Here, we investigate the relationship between Hcy-induced autophagy level and the expression of nuclear transcription factor EB (TFEB). The results show that Hcy-induced autophagy level is mediated by upregulation of TFEB. Silencing of TFEB decreases the level of autophagy-related protein LC3BII/I and increases p62 expression level in hepatocytes after exposure to Hcy. Moreover, the effect of Hcy on the expression of TFEB is regulated by hypomethylation of the TFEB promoter catalyzed by DNA methyltransferase 3b (DNMT3b). In summary, this study shows that Hcy can activate autophagy by inhibiting DNMT3b-mediated DNA methylation and upregulating TFEB expression. These findings provide another new mechanism for Hcy-induced autophagy in hepatocytes.

Cooperation between NSPc1 and DNA methylation represses HOXA11 expression and promotes apoptosis of trophoblast cells during preeclampsia.

Accumulating evidence has shown that the apoptosis of trophoblast cells plays an important role in the pathogenesis of preeclampsia, and an intricate interplay between DNA methylation and polycomb group (PcG) protein-mediated gene silencing has been highlighted recently. Here, we provide evidence that the expression of nervous system polycomb 1 (NSPc1), a BMI1 homologous polycomb protein, is significantly elevated in trophoblast cells during preeclampsia, which accelerates trophoblast cell apoptosis. Since NSPc1 acts predominantly as a transcriptional inactivator that specifically represses HOXA11 expression in trophoblast cells during preeclampsia, we further show that NSPc1 is required for DNMT3a recruitment and maintenance of the DNA methylation in the HOXA11 promoter in trophoblast cells during preeclampsia. In addition, we find that the interplay of DNMT3a and NSPc1 represses the expression of HOXA11 and promotes trophoblast cell apoptosis. Taken together, these results indicate that the cooperation between NSPc1 and DNMT3a reduces HOXA11 expression in preeclampsia pathophysiology, which provides novel therapeutic approaches for targeted inhibition of trophoblast cell apoptosis during preeclampsia pathogenesis.

11.4 T ultra-high static magnetic field has no effect on morphology but induces upregulation of TNF signaling pathway based on transcriptome analysis in zebrafish embryos.

As magnetic resonance imaging (MRI) scanners with ultra-high field (UHF) have optimal performance, scientists have been working to develop high-performance devices with strong magnetic fields to improve their diagnostic potential. However, whether an MRI scanner with UHF poses a risk to the safety of the organism require further evaluation. This study evaluated the effects of 11.4 Tesla (T) UHF on embryonic development using a zebrafish model. Multiple approaches, including morphological parameters, physiological behaviors, and analyses of the transcriptome at the molecular level, were determined during 5 days after laboratory-controlled exposure from 6 hour post fertilization (hpf) to 24 hpf. No significant effects were observed in embryo mortality, hatching rate, body length, Left-Right patterning, locomotor behavior, etc. RNA-sequencing analysis revealed up-regulated tumor necrosis factor (TNF) inflammatory factors and activated TNF signaling pathways in the 11.4 T exposure group. The results were further validated using qPCR. Our findings indicate that although UHF exposure under 11.4 T has no effect on the development of zebrafish embryos, it has specific effects on the immune response that require further investigation.

Immune Regulation of Seminal Plasma on the Endometrial Microenvironment: Physiological and Pathological Conditions.

Seminal plasma (SP) accounts for more than 90% of semen volume. It induces inflammation, regulates immune tolerance, and facilitates embryonic development and implantation in the female reproductive tract. In the physiological state, SP promotes endometrial decidualization and causes changes in immune cells such as macrophages, natural killer cells, regulatory T cells, and dendritic cells. This leads to the secretion of cytokines and chemokines and also results in the alteration of miRNA profiles and the expression of genes related to endometrial tolerance and angiogenesis. Together, these changes modulate the endometrial immune microenvironment and contribute to implantation and pregnancy. However, in pathological situations, abnormal alterations in SP due to advanced age or poor diet in men can interfere with a woman's immune adaptation to pregnancy, negatively affecting embryo implantation and even the health of the offspring. Uterine pathologies such as endometriosis and endometritis can cause the endometrium to respond negatively to SP, which can further contribute to pathological progress and interfere with conception. The research on the mechanism of SP in the endometrium is conducive to the development of new targets for intervention to improve reproductive outcomes and may also provide new ideas for semen-assisted treatment of clinical infertility.

Studying the mechanism of sperm DNA damage caused by folate deficiency.

Sperm DNA injury is one of the common causes of male infertility. Folic acid deficiency would increase the methylation level of the important genes, including those involved in DNA double-strand break (DSB) repair pathway. In the early stages, we analysed the correlation between seminal plasma folic acid concentration and semen parameters in 157 infertility patients and 91 sperm donor volunteers, and found that there was a significant negative correlation between seminal folic acid concentration and sperm DNA Fragmentation Index (DFI; r = -0.495, p < 0.01). Then through reduced representation bisulphite sequencing, global DNA methylation of sperm of patients in the low folic acid group and the high folic acid group was analysed, it was found that the methylation level in Rad54 promoter region increased in the folic acid deficiency group compared with the normal folic acid group. Meanwhile, the results of animal model and spermatocyte line (GC-2) also found that folic acid deficiency can increase the methylation level in Rad54 promoter region, increased sperm DFI in mice, increased the expression of γ-H2AX, that is, DNA injury marker protein, and increased sensitivity of GC-2 to external damage and stimulation. The study indicates that the expression of Rad54 is downregulated by folic acid deficiency via DNA methylation. This may be one of the mechanisms of sperm DNA damage caused by folate deficiency.

FABP4 activates the JAK2/STAT2 pathway via Rap1a in the homocysteine-induced macrophage inflammatory response in ApoE-/- mice atherosclerosis.

Atherosclerosis is a chronic inflammatory vascular disease, and inflammation plays a critical role in its formation and progression. Elevated serum homocysteine (Hcy) is an independent risk factor for atherosclerosis. Previous studies have shown that fatty acid binding protein 4 (FABP4) plays an important role in macrophage inflammation and lipid metabolism in atherosclerosis induced by Hcy. However, the underlying molecular mechanism of FABP4 in Hcy-induced macrophage inflammation remains unknown. In this study, we found that FABP4 activated the Janus kinase 2/signal transducer and activator of transcription 2 (JAK2/STAT2) pathway in macrophage inflammation induced by Hcy. Of note, we further observed that ras-related protein Rap-1a (Rap1a) induced the Tyr416 phosphorylation and membrane translocation of non-receptor tyrosine kinase (c-Src) to activate the JAK2/STAT2 pathway. In addition, the suppressor of cytokine signaling 1 (SOCS1)-a transcriptional target of signal transducer and activator of transcription (STATs) inhibited the JAK2/STAT2 pathway and Rap1a expression via a negative feedback loop. In summary, these results demonstrated that FABP4 promotes c-Src phosphorylation and membrane translocation via Rap1a to activate the JAK2/STAT2 pathway, contributing to Hcy-accelerated macrophage inflammation in ApoE-/- mice.

Comparative Phosphoproteomic Profiling in the Prefrontal Cortex of Prenatally Stressed Male Offspring Rats.

Many investigations have indicated that prenatal stress (PS) causes depressive-like disturbances in offspring rats. However, the underlying pathogenic mechanisms have not yet been fully elucidated. The prefrontal cortex (PFC) has been shown to play a role in susceptibility to stress during fetal development; thus, we focused our attention on differential protein phosphorylation in this region of PS-S (susceptibility to PS) offspring rats. The sucrose preference test was used to screen for susceptibility to PS. The validity of the prenatally stressed model was verified by other common depression-like behaviors. We used MS-based TMT quantitative proteomics in combination with the phosphopeptide enrichment method to compare phosphoproteomic profiling in the PFC of PS-S and control male offspring rats. In total, 3,418 phosphoproteins, 8,404 phosphopeptides, and 12,175 phosphosites were identified in this analysis. According to the screening criteria, 902 phosphopeptides increased and 609 decreased in the PFC of the PS-S group compared to the control rats. Gene ontology enrichment analysis indicated that the main enriched terms in the cellular component category were "synapse part," "myelin sheath," "synapse," "neuron part," and "axon." The phosphoproteins enriched in the molecular function and biological process categories were mainly related to cytoskeleton- and projection morphogenesis-associated proteins. KEGG pathway enrichment analyses identified 30 significant KEGG pathways; the top five pathways included salivary secretion, endocrine and other factor-regulated calcium reabsorption, pancreatic secretion, and insulin secretion. Motifs such as ……_S_P…RR, ……S_PE……, ……_S_PV……, ……_S_P.H……, and …S…_S_PT…. were the top five motifs enriched in phosphorylated sites. PS may induce depressive-like behaviors in male offspring rats by regulating the phosphorylation of proteins mainly related to synapses, myelin sheaths, neurons, and the cytoskeleton. The phosphorylation of related proteins may act as key pathogenic hits. Data are available via ProteomeXchange with identifier PXD026563.

N-Cadherin Regulates GluA1-Mediated Depressive-Like Behavior in Adolescent Female Rat Offspring following Prenatal Stress.

BACKGROUND: The incidence of depression is twice higher in women than in men, and gender differences in the prevalence rates first emerge around puberty. Prenatal stress (PS) induces gender-dependent depressive-like behavior in adolescent offspring, but the neuro-physiological mechanisms remain unclear. Our study aimed to investigate the possible neuro-physiological mechanisms of gender-dependent depressive-like behavior in PS adolescent offspring and further explored the possibility of treating depression in adolescent female rats. METHODS: The pregnant rats were exposed to restraint stress in the third trimester for 7 days. The depressive-like behavior and the expression of N-cadherin and AMPARs in the hippocampus of adolescent offspring rats were assessed. 10 mg/kg AMPAR antagonist CNQX and 10 mg/kg N-cadherin antagonist ADH-1 were intraperitoneally injected into female adolescent offspring, respectively; 0.2 µg AMPAR agonist CX546 was administered to the dentate gyrus of male adolescent offspring to determine the role of N-cadherin-AMPARs in depressive-like behavior of the offspring following PS. RESULTS: We found that PS increased N-cadherin expression, which upregulated GluA1 expression in the dentate gyrus, mediating depressive-like behavior in adolescent female rat offspring by reducing PSD-95. In addition, ADH-1 and CNQX improved depressive-like behavior in adolescent female offspring following PS. Furthermore, injection of the CX546 into the dentate gyrus induced depressive-like behavior in PS male offspring. CONCLUSION: The gender-dependent expression of N-cadherin-GluA1 pathway in adolescent offspring in the dentate gyrus was the key factor in gender differences of depressive-like behavior following PS.

Homocysteine facilitates endoplasmic reticulum stress and apoptosis of hepatocytes by suppressing ERO1α expression via cooperation between DNMT1 and G9a.

Endoplasmic reticulum (ER) stress and apoptosis play a critical role in liver injury. Endoplasmic reticulum oxidoreductase 1α (ERO1α) is an oxidase that exists in the luminal side of the ER membrane, participating in protein folding and secretion and inhibiting apoptosis, but the underlying mechanism on liver injury induced by homocysteine (Hcy) remains obscure. In this study, hyperhomocysteinemia (HHcy) mice model was established in cbs+/- mice by feeding a high-methionine diet for 12 weeks; and cbs+/- mice fed with high-methionine diet exhibited more severe liver injury compared to cbs+/+ mice. Mechanistically, we found that Hcy promoted ER stress and apoptosis of hepatocytes and thereby aggravated liver injury through inhibiting ERO1α expression; accordingly, overexpression of ERO1α remarkably alleviated ER stress and apoptosis of hepatocytes induced by Hcy. Epigenetic modification analysis revealed that Hcy significantly increased levels of DNA methylation and H3 lysine 9 dimethylation (H3K9me2) on ERO1α promoter, which attributed to upregulated DNA methyltransferase 1 (DNMT1) and G9a, respectively. Further study showed that DNMT1 and G9a cooperatively regulated ERO1α expression in hepatocytes exposed to Hcy. Taken together, our work demonstrates that Hcy activates ER stress and apoptosis of hepatocytes by downregulating ERO1α expression via cooperation between DNMT1 and G9a, which provides new insight into the mechanism of Hcy-induced ER stress and apoptosis of hepatocytes in liver injury.

Characterization of the metabolites of tirabrutinib generated from rat, dog and human liver microsomes using ultra-high-performance liquid chromatography combined with high-resolution mass spectrometry.

RATIONALE: Tirabrutinib is an orally administered Bruton's tyrosine kinase (BTK) inhibitor developed for the treatment of autoimmune disorders and haematological malignancies. The goals of this study were to identify the metabolites of tirabrutinib and to propose the metabolic pathways. METHODS: Tirabrutinib was individually incubated with rat, dog and human liver microsomes at 37°C for 1 h. To trap the potential reactive metabolites, glutathione (GSH) was incorporated into the incubation samples. The incubation samples were analysed using ultra-high-performance liquid chromatography combined with high-resolution mass spectrometry (UHPLC-HRMS). The metabolites were identified and characterized by exact masses, product ions and retention times. RESULTS: A total of 18 metabolites, including four GSH conjugates, were identified and characterized in terms of elemental compositions and product ions. The metabolic pathways of tirabrutinib included amide hydrolysis, O-dealkylation, mono-oxygenation, di-oxygenation and GSH conjugation. Among these metabolites, M10 was the most abundant metabolite. Compared with dog, rat has the closer metabolic profiles to humans, and thus it would be more suitable for toxicity study. CONCLUSIONS: This study provides valuable data regarding the in vitro metabolism of tirabrutinib, which may be helpful for further safety assessment of this drug.