Xin Wang

Introduction: Juglone is a redox-active constituent of walnut green husks, but its effects on drug-metabolizing cytochrome P450 (CYP) enzymes remain incompletely characterized. Methods: We evaluated juglone against six CYP activities in human and rat liver microsomes (HLMs and RLMs) using cocktail probe assays, time-dependent inhibition experiments, kinetic analyses, and molecular docking. Results: In HLMs, CYP2C9 was the most susceptible activity (IC ₅₀ = 8.12 μM; K _i = 4.97 μM) and showed competitive inhibition. CYP2E1 was the next most susceptible activity (IC ₅₀ = 12.67 μM; K _i = 19.59 μM; αK _i = 8.74 μM) and showed mixed inhibition. In RLMs, CYP1A2 (IC ₅₀ = 12.03 μM) and CYP2E1 (IC ₅₀ = 18.50 μM) met the prespecified criterion for further kinetic analysis. Subsequent analyses identified mixed inhibition of CYP1A2 ( K _i = 12.31 μM) and competitive inhibition of CYP2E1 (Ki = 19.89 μM). No consistent progressive loss of activity was observed during preincubation for up to 20 min under the tested conditions. CYP2E1 showed numerically similar apparent K _i values but different fitted inhibition models in HLMs and RLMs. However, differing assay conditions precluded attributing these findings solely to species. Molecular docking provided qualitative structural insight into the potential interactions between juglone and CYP2C9. Conclusion: Juglone inhibited CYP activities in vitro , with CYP2C9 showing the greatest susceptibility among the six human CYP activities tested. These findings characterize biochemical inhibition and provide a basis for evaluating potential metabolic interactions involving juglone-containing products.

Over 800 million people globally suffer from chronic kidney disease (CKD). Maternal obesity has emerged as a risk factor for CKD in offspring, but the mechanisms remain unclear. In this study, we use mouse models of maternal obesity to investigate macrophage involvement in offspring kidneys. Multi-omics analyses revealed that maternal obesity accelerates kidney disease in male offspring through dysregulated crosstalk between proximal tubules (PT) and macrophages. PT-derived 20-hydroxyeicosatetraenoic acid (20-HETE) promotes mitochondrial hyperactivity and macrophage to myofibroblast transition (MMT) via Ffar1 in macrophages. Targeting 20-HETE or depleting Ffar1 in offspring of obese mothers markedly reduces kidney pathology. Hormone screening identified 3,3,5-Triiodo-L-thyronine (T3) as a factor that enhances Ffar1 expression in macrophages. Mechanistically, the T3 receptor (TRβ) binds to an enhancer we identify upstream of the gene, promoting Ffar1 transcription via the TRβ-P300-BRD4 regulatory axis. These findings highlight T3 and 20-HETE co-activated MMT as a central mechanism to kidney disease in offspring of obese mothers, offering potential therapeutic targets.

Background: Escitalopram (SCT), a highly effective and low-risk antidepressant, is widely used in the clinical treatment of depression. It undergoes two N-demethylation steps, producing its primary metabolites, S-demethylcitalopram (S-DCT) and S-didemethylcitalopram (S-DDCT). Purpose: This study aimed to develop and validate an LC-MS/MS method to simultaneously detect SCT, S-DCT, and S-DDCT, and then apply it to study metabolism in different microsomes, comparing SCT metabolism in rat liver microsomes (RLM), human liver microsomes (HLM), and human placenta microsomes (HPM). Methods: This study developed a reliable LC-MS/MS method for the simultaneous detection of escitalopram and its metabolites in various microsomal systems. The method was fully validated in rat liver microsomes and then was applied into human liver and placenta microsomes. The method was further applied to investigate escitalopram metabolism in different microsomal systems, notably for the first time in human placenta microsomes. Results: LC-MS/MS method provides high specificity, good stability, and excellent extraction recovery. Both intra-day and inter-day accuracy (% RE) and precision (% RSD) were within ±15%, with no significant matrix effects observed. The metabolic rates of escitalopram and its metabolite levels differed among rat liver, human liver, and human placenta microsomes, likely due to variations in the types and activities of drug-metabolizing enzymes present in these systems. Conclusion: A sensitive and reliable LC-MS/MS method was developed and validated for the quantitative analysis of SCT, S-DCT, and S-DDCT in RLM. Subsequently, the method was applied to the study of HLM and HPM. Incubation experiments using these microsomal systems showed that all three types of microsomes, including HPM, could metabolize SCT.

Heart failure (HF), characterized by maladaptive cardiac fibrosis and progressive functional deterioration, remains a therapeutic challenge. In this study, we established a cardiac organoid HF model derived from human‐induced pluripotent stem cells (hiPSCs) and observed a significant downregulation of the desmosomal protein plakophilin‐2 (PKP2) in this model. Reduced PKP2 expression was detected in both HF rat and mouse. Subsequent in vivo studies on Pkp2‐ knockout ( Pkp2 ‐KO) rats demonstrated that adeno‐associated virus serotype 9 (AAV9)‐mediated restoration of PKP2 not only restored cardiac PKP2 expression but also attenuated the progression of fibrosis. Administration of AAV9‐PKP2 could also inhibit myocardial fibrosis and slow down disease progression in HF mouse. Single‐cell RNA sequencing analysis in rats revealed enriched pathological profibrotic cardiac fibroblasts (CFs) in PKP2‐deficient myocardium. Mechanistically, AAV9‐PKP2 administration induced the phenotypic conversion of activated CFs into quiescent antifibrotic states. Integrated bioinformatics identified that protein tyrosine phosphatase receptor type C ( Ptprc ) was a pivotal regulator orchestrating this cellular reprogramming. Our findings thus unveil PKP2 as a master regulator of fibroblast activation and propose AAV9‐PKP2 gene therapy as a promising novel therapeutic strategy targeting pathological fibrosis in HF.

BACKGROUND: Preeclampsia is a life-threatening pregnancy disorder characterized by hypertension and multiorgan dysfunction, posing significant risks to both maternal and fetal health. Although low-dose aspirin is widely recommended for preventing preeclampsia, the underlying mechanisms of action are still poorly understood, which hinders the optimization of therapeutic strategies. METHODS: We developed an in vitro hypoxia-induced preeclampsia model using human trophoblast organoids to replicate key pathological features. RNA sequencing identified dysregulated pathways and molecular targets. Functional assays assessed the effects of aspirin on trophoblast proliferation, mitochondrial activity, and hormonal regulation, focusing on the PI3K (phosphatidylinositol 3-kinase)-AKT (protein kinase B) pathway and CYP (cytochrome P450) enzymes. We also analyzed the effects of aspirin in the N’-nitro-L-arginine-methyl ester hydrochloride rat model. RESULTS: The hypoxia-induced preeclampsia model successfully mimicked clinical hallmarks, including elevated sFLT-1 (soluble fms-like tyrosine kinase 1)/PlGF (placental growth factor) ratios and oxidative damage. RNA sequencing revealed significant suppression of the PI3K-AKT-mTOR (mammalian target of rapamycin) pathway and dysregulation of CYP enzymes. Aspirin treatment restored the sFLT-1/PlGF balance, reactivated the PI3K-AKT-mTOR pathway, and improved mitochondrial function, enhancing trophoblast proliferation. Furthermore, aspirin regulated CYP by increasing CYP19A1 (cytochrome P450 1A1) and inhibiting CYP1A1 (cytochrome P450 1A1), thereby improving placental hormonal homeostasis. CONCLUSIONS: This study clarifies aspirin’s multitarget mechanisms in alleviating preeclampsia, which include restoring the sFLT-1/PlGF balance, activating the PI3K-AKT-mTOR signaling pathway, optimizing mitochondrial function, and regulating CYP-mediated hormonal metabolism. These findings provide a mechanistic basis for aspirin’s clinical effectiveness in preventing preeclampsia.