Association of Blood Copper With the Subclinical Carotid Atherosclerosis: An Observational Study.

BACKGROUND: Copper exposure is reported to be associated with increased risk of stroke. However, the association of copper exposure with subclinical carotid atherosclerosis remains unclear. METHODS AND RESULTS: This observational study included consecutive participants from Xinqiao Hospital between May 2020 and August 2021. Blood metals were measured using inductively coupled plasma mass spectrometry and carotid atherosclerosis was assessed using ultrasound. Modified Poisson regression was performed to evaluate the associations of copper and other metals with subclinical carotid plaque presence. Blood metals were analyzed as categorical according to the quartiles. Multivariable models were adjusted for age, sex, body mass index, education, smoking, drinking, hypertension, diabetes, dyslipidemia, estimated glomerular filtration rate, and coronary artery disease history. Bayesian Kernel Machine Regression was conducted to evaluate the overall association of metal mixture with subclinical carotid plaque presence. One thousand five hundred eighty-five participants were finally enrolled in our study, and carotid plaque was found in 1091 subjects. After adjusting for potential confounders, metal-progressively-adjusted models showed that blood copper was positively associated with subclinical carotid plaque (relative risk according to comparing quartile 4 to quartile 1 was 1.124 [1.021-1.238], relative risk according to per interquartile increment was 1.039 [1.008-1.071]). Blood cadmium and lead were also significantly associated with subclinical carotid plaque. Bayesian Kernel Machine Regression analyses suggested a synergistic effect of copper-cadmium-lead mixture on subclinical carotid plaque presence. CONCLUSIONS: Our findings identify copper as a novel risk factor of subclinical carotid atherosclerosis and show the potential synergistic proatherogenic effect of copper, cadmium, and lead mixture.

Integrating metabolomics and network pharmacology analysis to explore mechanism of Pueraria lobata against pulmonary fibrosis: Involvement of arginine metabolism pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: Pueraria lobata (Willd.) Ohwi is a typical medicinal and edible plant with a long application history in China and Southeast Asia. As a widely used traditional medicine, P. lobata exhibits the properties of anti-inflammatory, antipyretic, antioxidant, relieving cough and asthma. Particularly, the increasing evidence indicates that the P. lobata has the therapeutic effect on fibrotic-related diseases in terms of metabolic regulation. However, the mechanisms of P. lobata on pulmonary fibrosis (PF) has not been thoroughly explored. AIM OF THE STUDY: This study aimed to explore the effect of arginine metabolites of P. lobata against PF model by integrating metabolomics and network pharmacology analysis. It might provide a new idea for the target finding of P. lobata anti-pulmonary fibrosis. MATERIALS AND METHODS: In this study, the Sprague Dawley (SD) rats were randomly divided into five experimental groups: saline-treated control group, bleomycin-induced fibrosis group, prednisolone acetate group, P. lobata 3.2 g/kg group and P. lobata 6.4 g/kg group. The therapeutic effect of P. lobata on bleomycin-induced PF in rats was evaluated by clinical symptoms such as lung function, body weight, hematoxylin eosin staining (HE), Masson staining and hydroxyproline assay. Next, the plasma metabolomics analysis was carried out by LC-MS to explore the pathological differences between the group of control, PF and P. lobata-treated rats. Then, the network pharmacology study coupled with experimental validation was conducted to analysis the results of metabolic research. We constructed the "component-target-disease" network of P. lobata in the treatment of PF. In addition, the molecular docking method was used to verify the interaction between potential active ingredients and core targets of P. lobata. Finally, we tested NOS2 and L-OT in arginine-related metabolic pathway in plasma of the rats by enzyme-linked immunosorbent assay (ELISA). Real-time PCR was performed to observe the level of TNF-α mRNA and MMP9 mRNA. And we tested the expression of TNF-α and MMP9 by Western blot analysis. RESULTS: Our findings revealed that P. lobata improved lung function and ameliorated the pathological symptoms, such as pathological damage, collagen deposition, and body weight loss in PF rats. Otherwise, the plasma metabolomics were employed to screen the differential metabolites of amino acids, lipids, flavonoids, arachidonic acid metabolites, glycoside, etc. Finally, we found that the arginine metabolism signaling mainly involved in the regulating of P. lobata on the treatment of PF rats. Furtherly, the network pharmacology predicted that the arginine metabolism pathway was contained in the top 20 pathways. Next, we integrated metabolomics and network pharmacology that identified NOS2, MMP9 and TNF-α as the P. lobata regulated hub genes by molecular docking. Importantly, it indicated a strong affinity between the puerarin and the NOS2. P. lobata attenuated TNF-α, MMP-9 and NOS2 levels, suppressed TNF-α and MMP-9 protein expression, and decreased L-OT and NOS2 content in PF rats. These results indicated that the effects of P. lobata may ameliorated PF via the arginine metabolism pathway in rats. Therefore, P. lobata may be a potential therapeutic agent to ameliorated PF. CONCLUSION: In this work, we used metabolomics and network pharmacology to explore the mechanisms of P. lobata in the treatment of PF. Finally, we confirmed that P. lobata alleviated BLM-induced PF in rats by regulating arginine metabolism pathway based on reducing the L-OT and NOS2-related signal molecular. The search for the biomarkers finding of arginine metabolism pathway revealed a new strategy for P. lobata in the treatment of PF.

Manual acupuncture ameliorates inflammatory pain by upregulating adenosine A3 receptor in complete Freund's adjuvant-induced arthritis rats.

BACKGROUND: Adenosine A3 receptor (A3R) exerts analgesic, anti-inflammatory, and anti-nociceptive effects. In this study, we determined the analgesic mechanism of manual acupuncture (MA) in rats with complete Freund's adjuvant (CFA)-induced arthritis and explored whether MA ameliorates inflammation in these rats by upregulating A3R. METHODS: Sixty Sprague Dawley (SD) rats were randomly divided into the following groups: Control, CFA, CFA + MA, CFA + sham MA, CFA + MA + DMSO, CFA + MA + IB-MECA, and CFA + MA + Reversine groups. The arthritis rat model was induced by injecting CFA into the left ankle joints. Thereafter, the rats were subjected to MA (ST36 acupoint) for 3 days. The clinical indicators paw withdrawal latency (PWL), paw withdrawal threshold (PWT), and open field test (OFT) were used to determine the analgesic effect of MA. In addition, to explore the effect of A3R on inflammation after subjecting arthritis rats to MA, IB-MECA (A3R agonist) and Reversine (A3R antagonist) were injected into ST36 before MA. RESULTS: MA ameliorated the pathological symptoms of CFA-induced arthritis, including the pain indicators PWL and PWT, number of rearing, total ambulatory distance, and activity trajectory. Furthermore, after MA, the mRNA and protein expression of A3R was upregulated in CFA-induced arthritis rats. In contrast, the protein levels of TNF-α, IL-1ß, Rap1, and p-p65 were downregulated after MA. Interestingly, the A3R agonist and antagonist further downregulated and upregulated inflammatory cytokine expression, respectively, after MA. Furthermore, the A3R antagonist increased the degree of ankle swelling after MA. CONCLUSION: MA can alleviate inflammatory pain by inhibiting the NF-κB signaling pathway via upregulating A3R expression of the superficial fascia of the ST36 acupoint site in CFA-induced arthritis rats.

Association of multiple blood metals and systemic atherosclerosis: A cross-sectional study in the CAD population.

BACKGROUND: Coronary atherosclerotic disease (CAD) is often accompanied by peripheral atherosclerosis, resulting in a higher risk of ischemia and cardiovascular death. Exposure to metals is associated with atherosclerotic plaques at specific sites. However, less is known about the effects of mixed metals on systemic atherosclerotic burden in CAD patients. OBJECTIVES: To investigate the association of metal mixtures with systemic atherosclerotic burden in a CAD population. METHODS: A cross-sectional study including 1562 CAD patients from Southwest China was conducted. The levels of 10 blood metals were measured via inductively coupled plasma spectrometry. More than one vessel with a stenosis ≥50% vessel diameter was defined as CAD. Carotid and lower limb atherosclerosis was assessed by using ultrasound, and coronary atherosclerosis was quantified via arterial angiography. Systemic atherosclerosis was scored according to the presence or absence of lesions at the three sites and the total number of lesions. To investigate the combined impacts and interaction effects of metals, Bayesian kernel machine regression was used. Weighted quantile regression was used to identify the contributions of the metals. RESULTS: Significant overall associations of mixed metals with systemic atherosclerotic burden were found. These positive overall associations were mainly driven by Cd, Cu and Pb in systemic atherosclerosis. The main contributing factors were As and Cu for coronary atherosclerosis as well as Cd, Cu and Pb for carotid and lower limb atherosclerosis. Cd and Pb or Cr can interact, and Pb interacts with age, sex and alcohol. CONCLUSIONS: In CAD patients, exposure to combinations of metals was highly positively associated with systemic atherosclerotic burden. These significant trends were more pronounced in the peripheral arteries and carotid arteries. Controlling environmental metal exposure can contribute to reducing systemic atherosclerosis in CAD patients.

Ultrasensitive quantitation of Paraquat based on a small molecule-induced dual-cycle amplification strategy.

Paraquat (PQ) is a typical biotoxic small molecule. Knowledge of how to directly introduce it into cyclic amplification rather than transform it into a secondary target is lacking in current analytical methods. Considering the urgent need for trace pesticide residue detection and the inherent defects of small molecule analysis, a CRISPR/Cas12a-driven small molecule-induced dual-cycle strategy was developed based on the immune competition method. The key to signal amplification is the mutual activation and acceleration between Cycle 1 triggered by the small molecule and Cycle 2 driven by CRISPR/Cas12a. Impressively, small molecules have been successfully incorporated into the dual-cycle strategy, which achieves a low detection limit (3.1 pg/mL) and a wide linear range (from 10 pg/mL to 50 µg/mL). Moreover, the designed biosensor was successfully employed to evaluate the PQ residual level in real samples and showed effective implementation for the bioanalysis of small molecule targets and pesticide residue-related food safety.

Cadmium exposure promotes ferroptosis by upregulating Heat Shock Protein 70 in vascular endothelial damage of zebrafish.

Cadmium (Cd) exposure is a risk factor for endothelial dysfunction and cardiovascular disease. Ferroptosis is a type of cell death that relies on lipid peroxidation. Whether ferroptosis acts in Cd-induced vascular endothelial damage and the underlying mechanisms remain unclear. Herein, we found that Cd resulted in ferroptosis of vascular endothelial cells (ECs) in vivo and in vitro. In the visualized zebrafish embryos, Cd accumulated in vascular ECs, ROS and lipid peroxidation levels were increased, and the oxidoreductase system was disturbed after exposure. Moreover, Cd decreased Gpx4 in ECs and caused smaller mitochondria with increased membrane density. Accompanied by ferroptosis, the number of ECs and the area of the caudal venous plexus in zebrafish embryos were reduced, and the survival rate of HUVECs decreased. These effects were partially reversed by ferrostatin-1 and aggravated by erastin. Mechanistically, an excessive increase in Heat Shock Protein 70 (Hsp70) was identified by transcriptomics after Cd exposure. Inhibition of Hsp70 by VER-155008 or siRNA ameliorated Cd-induced ferroptosis, thereby alleviating endothelial injury. Furthermore, Hsp70 regulated Cd-induced ferroptosis by targeting multiple targets, including Gpx4, Fth1, Nrf2 and Acsl4. Our findings provide a new approach to investigating the endothelial damage of Cd and indicate that regulation of Hsp70 is an important target for alleviating this process.

Liver fibrosis-derived exosomal miR-106a-5p facilitates the malignancy by targeting SAMD12 and CADM2 in hepatocellular carcinoma.

The mechanism of hepatocellular carcinoma (HCC) development induced by liver fibrosis is obscure. The objective of this study is to establish miRNAs from exosomes associated with liver fibrosis, and to identify potential biomarkers for the prediction of personalized clinical management effectiveness in HCC. Our research focused on miRNAs from exosomes and mRNA from liver fibrosis, which we found in the gene expression omnibus (GEO) database. Weighted gene co-expression network analysis (WGCNA) evaluated miRNAs from exosomes associated with liver fibrosis, and Wilcoxon analysis assessed differentially expressed mRNAs (DEGs) across liver fibrosis/normal tissues. Following that, DEGs were assessed through gene set enrichment analysis (GSEA), gene ontology (GO), and the Kyoto Encyclopedia of Genes and Genomes (KEGG). In addition, based on the screened targeted genes, including SAMD12 and CADM2, we further elucidated their correlation in HCC patients from the BEST database. The Kaplan-Meier Plotter platform was applied to evaluate the prognostic values of miRNA in HCC. In vitro and vivo experiments validated our findings. Six miRNAs associated with liver fibrosis were evaluated in our investigation. In-depth research presented exosome-derived miR-106a-5p, SAMD12 and CADM2 could exert valuable predictive implications for HCC treatment and illness assessment. Serum miR-106a-5p derived from liver fibrosis was decreased compared with healthy individuals. SAMD12 and CADM2 were diminished in liver cancer cell lines, and their knockdown of them exacerbated the proliferation capacities of liver cells in vitro. Exosome-derived miRNA of liver fibrosis modulated tumorigenesis by targeting SAMD12 and CADM2 in HCC.

Low serum Klotho reflects senile inflammation in middle-aged and elderly patients with coronary atherosclerosis.

BACKGROUND: Anti-aging protein Klotho has been reported to be associated with atherosclerosis, which was considered as a chronic inflammatory disease. However, the relationship between Klotho and senile inflammation remained unclear. The present study aims to ascertain the correlation of Klotho with inflammation in middle-aged and elderly coronary atherosclerotic disease (CAD). METHODS: A total of 302 patients with CAD were included in this study. Coronary atherosclerosis was confirmed and quantified for all patients by coronary angiography. Serum Klotho was detected by enzyme linked immunosorbent assay. Serum concentrations of IL-6 and IL-8 were quantified by chemiluminescence assay. T-lymphocyte subsets were measured using flow cytometry. RESULTS: Multivariate linear regression analysis showed that serum Klotho was an independent predictor for circulating monocytes (standard ß = -0.321, P < 0.001) and CD4+/CD8+ ratio (standard ß = -0.522, P < 0.001). After adjustment, serum Klotho was still independently associated with IL-6 (standard ß = -0.395, P < 0.001) and IL-8 (standard ß = -0.296, P < 0.001). Moreover, circulating monocytes, CD4+ and CD8+ lymphocytes were correlated with increased serum concentrations of IL-6 and IL-8, independent of CRP (P < 0.05). In receiver operating characteristic curve analysis, CD4+/CD8+ ratio (AUC = 0.863, P < 0.001), IL-6 (AUC = 0.893, P < 0.001) and IL-8 (AUC = 0.884, P < 0.001) presented the excellent predictive performance for significant CAD. CONCLUSIONS: Decreased concentrations in serum Klotho reflect senile inflammation, which is related to the severity of CAD in middle-aged and elderly patients.

Upregulation of mitochondrial calcium uniporter contributes to paraquat-induced neuropathology linked to Parkinson's disease via imbalanced OPA1 processing.

Paraquat (PQ) is the most widely used herbicide in agriculture worldwide and has been considered a high-risk environmental factor for Parkinson's disease (PD). Chronic PQ exposure selectively induces dopaminergic neuron loss, the hallmark pathologic feature of PD, resulting in Parkinson-like movement disorders. However, the underlying mechanisms remain unclear. Here, we demonstrated that repetitive PQ exposure caused dopaminergic neuron loss, dopamine deficiency and motor deficits dose-dependently in mice. Accordingly, mitochondrial calcium uniporter (MCU) was highly expressed in PQ-exposed mice and neuronal cells. Importantly, MCU knockout (KO) effectively rescued PQ-induced dopaminergic neuron loss and motor deficits in mice. Genetic and pharmacological inhibition of MCU alleviated PQ-induced mitochondrial dysfunction and neuronal death in vitro. Mechanistically, PQ exposure triggered mitochondrial fragmentation via imbalance of the optic atrophy 1 (OPA1) processing manifested by cleavage of L-OPA1 to S-OPA1, which was reversed by inhibition of MCU. Notably, the upregulation of MCU was mediated by miR-129-1-3p posttranscriptionally, and overexpression of miR-129-1-3p could rebalance OPA1 processing and attenuate mitochondrial dysfunction and neuronal death induced by PQ exposure. Consequently, our work uncovers an essential role of MCU and a novel molecular mechanism, miR-MCU-OPA1, in PQ-induced pathogenesis of PD, providing a potential target and strategy for environmental neurotoxins-induced PD treatment.

Klotho ameliorates angiotension-II-induced endothelial senescence via restoration of autophagy by inhibiting Wnt3a/GSK-3ß/mTOR signaling: A potential mechanism involved in prognostic performance of Klotho in coronary atherosclerotic disease.

OBJECTIVE: We aimed to evaluate the prognostic performance of circulating Klotho in coronary atherosclerotic disease (CAD), and to further explore the effect of Klotho on stress-mediated endothelial senescence and underlying mechanism. METHODS: A cohort of 295 patients had a 12-month follow-up for major adverse cardiovascular events (MACE). Serum Klotho was detected by enzyme linked immunosorbent assay. Cell viability, SA-ß-Gal staining, the expression of P53 and P16 were analyzed for endothelial senescence. Oxidative stress was evaluated by measurement of reactive oxygen species, superoxide dismutase and malondialdehyde. LC3, P62, Wnt3a, GSK-3ß and mTOR were analyzed by western blotting. Autophagosome formation was detected by adenovirus transfection. RESULTS: In epidemiological analysis, low Klotho (≤295.9 pg/ml) was significantly associated with MACE risk (HR=2.266, 95 %CI 1.229-4.176). In experimental analysis, Klotho alleviated endothelial senescence and oxidative stress caused by Ang-II exposure; Klotho restored impaired autophagic flux to ameliorate Ang-II induced endothelial senescence; Ang-II activated Wnt3a/GSK-3ß/mTOR signaling to inhibit autophagy, whereas Klotho restored autophagy through blockade of Wnt3a/GSK-3ß/mTOR signaling; Klotho ameliorated endothelial senescence by suppressing Wnt3a/GSK-3ß/mTOR pathway under Ang-II exposure. CONCLUSIONS: Prognostic significance of Klotho in CAD is potentially ascribed to its anti-endothelial senescence effect via autophagic flux restoration by inhibiting Wnt3a/ GSK-3ß/mTOR signaling.

Prenatal exposure to pesticides and domain-specific neurodevelopment at age 12 and 18 months in Nanjing, China.

BACKGROUND: The extensive usage of pesticides has led to a ubiquitous exposure in the Chinese general population. Previous studies have demonstrated developmental neurotoxicity associated with prenatal exposure to pesticides. OBJECTIVES: We aimed to delineate the landscape of internal pesticides exposure levels from pregnant women's blood serum samples, and to identify the specific pesticides associated with the domain-specific neuropsychological development. METHODS: Participants included 710 mother-child pairs in a prospective cohort study initiated and maintained in Nanjing Maternity and Child Health Care Hospital. Maternal spot blood samples were collected at enrollment. Leveraging on an accurate, sensitive and reproducible analysis method for 88 pesticides, a total of 49 pesticides were measured simultaneously using gas chromatography-triple quadrupole tandem mass spectrometry (GC-MS/MS). After implementing a strict quality control (QC) management, 29 pesticides were reported. We assessed neuropsychological development in 12-month-old (n = 172) and 18-month-old (n = 138) children using the Ages and Stages Questionnaire (ASQ), Third Edition. Negative binomial regression models were used to investigate the associations between prenatal exposure to pesticides and ASQ domain-specific scores at age 12 and 18 months. Restricted cubic spline (RCS) analysis and generalized additive models (GAMs) were fitted to evaluate non-linear patterns. Longitudinal models with generalized estimating equations (GEE) were conducted to account for correlations among repeated observations. Weighted quantile sum (WQS) regression and Bayesian kernel machine regression (BKMR) were applied to examining the joint effect of the mixture of pesticides. Several sensitivity analyses were performed to assess the robustness of the results. RESULTS: We observed that prenatal exposure to chlorpyrifos was significantly associated with a 4 % decrease in the ASQ communication scores both at age 12 months (RR, 0.96; 95 % CI, 0.94-0.98; P < 0.001) and 18 months (RR, 0.96; 95 % CI, 0.93-0.99; P < 0.01). In the ASQ gross motor domain, higher concentrations of mirex (RR, 0.96; 95 % CI, 0.94-0.99, P < 0.01 for 12-month-old children; RR, 0.98; 95 % CI, 0.97-1.00, P = 0.01 for 18-month-old children), and atrazine (RR, 0.97; 95 % CI, 0.95-0.99, P < 0.01 for 12-month-old children; RR, 0.99; 95 % CI, 0.97-1.00, P = 0.03 for 18-month-old children) were associated with decreased scores. In the ASQ fine motor domain, higher concentrations of mirex (RR, 0.98; 95 % CI, 0.96-1.00, P = 0.04 for 12-month-old children; RR, 0.98; 95 % CI, 0.96-0.99, P < 0.01 for 18-month-old children), atrazine (RR, 0.97; 95 % CI, 0.95-0.99, P < 0.001 for 12-month-old children; RR, 0.98; 95 % CI, 0.97-1.00, P = 0.01 for 18-month-old children), and dimethipin (RR, 0.94; 95 % CI, 0.89-1.00, P = 0.04 for 12-month-old children; RR, 0.93; 95 % CI, 0.88-0.98, P < 0.01 for 18-month-old children) were associated with decreased scores. The associations were not modified by child sex. There was no evidence of statistically significant nonlinear relationships between pesticides exposure and RRs of delayed neurodevelopment (Pnonlinearity > 0.05). Longitudinal analyses implicated the consistent findings. CONCLUSION: This study gave an integrated picture of pesticides exposure in Chinese pregnant women. We found significant inverse associations between prenatal exposure to chlorpyrifos, mirex, atrazine, dimethipin and the domain-specific neuropsychological development (i.e., communication, gross motor and fine motor) of children at 12 and 18 months of age. These findings identified specific pesticides with high risk of neurotoxicity, and highlighted the need for priority regulation of them.

Metabolomics of the anti-inflammatory effect of Pueraria lobata and Pueraria lobata var. Thomsonii in rats.

ETHNOPHARMACOLOGICAL RELEVANCE: Pueraria lobata (Willd.) Ohwi and Pueraria lobata var. Thomsonii (Benth.) Maesen are essential medicinal and edible homologous plants widely cultivated in Asian countries. Therefore, P. lobata and P. thomsonii are widely used in the food, health products and pharmaceutical industries and have significant domestic and international market potential and research value. P. lobata and P. thomsonii have pharmacological effects in the clinic, such as antipyretic, analgesic, anti-inflammatory and antioxidant effects. These plants are commonly used in the treatment of inflammatory diseases and other related diseases. However, the potential mechanisms of the anti-inflammatory effects of P. lobata and P. thomsonii have not been elucidated. AIM OF THE STUDY: This study aimed to confirm the anti-inflammatory effects of P. lobata and P. thomsonii on inflammatory model diseases and to investigate the mechanism of their anti-inflammatory effects from the perspective of plasma metabolomics. MATERIALS AND METHODS: First, P. lobata and P. thomsonii were identified by high‒performance liquid chromatography (HPLC). Second, we established the following three inflammation models: an acute inflammation model of auricular swelling in mice induced by xylene, an acute inflammation model of foot swelling in rats induced by carrageenan gum, and a chronic inflammation model of cotton ball granuloma in rats. Then we examined the weight and swelling rate of auricular swelling in mice; the residence time, contact area, and mean contact pressure in rats on the gait meter; and the weight of granulomas in rats and the content of IL-1ß and TNF-α in plasma to investigate the anti-inflammatory pharmacodynamics of P. lobata and P. thomsonii. Third, we used LC‒MS‒based plasma metabolomics techniques to obtain potential biomarkers of P. lobata and P. thomsonii related to inflammation. Then, the potential biomarkers were enriched by MetaboAnalyst and KEGG metabolomics analysis tools to obtain metabolic pathways related to inflammation. Finally, we tested the indicators of COX-2, 5-LOX, GSH, GSSG and γ⁃GCL in rat plasma from the granuloma model by enzyme-linked immunosorbent assays (ELISAs) to verify the inflammation-related metabolic pathway. RESULTS: The experimental results showed that P. lobata and P. thomsonii could reduce the swollen weight and swelling rate of the auricle in mice, and could increase the residence time, contact area and mean contact pressure in rats on the gait meter. Moreover, P. lobata and P. thomsonii could inhibit the growth of granulomas and reduce the content of IL-1ß and TNF-α in plasma in rats. The above results preliminarily verified that P. lobata and P. thomsonii have different anti-inflammatory effects. We identified eighteen plasma biomarkers associated with P. lobata and sixteen plasma biomarkers related to P. thomsonii in regulating inflammation by a plasma metabolomics analysis. The following two major metabolic pathways were further screened and enriched: arachidonic acid metabolism and glutathione metabolism. Then we noted that P. lobata and P. thomsonii could reduce the COX-2, 5-LOX and GSSG levels and increase the GSH, GSH/GSSG and γ⁃GCL levels based on the ELISA results, which demonstrated that P. lobata and P. thomsonii affect the anti-inflammatory mechanism through arachidonic acid metabolism and glutathione metabolism. CONCLUSIONS: The results of this study further elucidate the anti-inflammatory mechanism of action of P. lobata and P. thomsonii, providing a scientific basis for developing new drugs for the treatment of inflammation-related diseases and laying a foundation for the development of herbal resources, such as P. lobata and P. thomsonii.

MCU Upregulation Overactivates Mitophagy by Promoting VDAC1 Dimerization and Ubiquitination in the Hepatotoxicity of Cadmium.

Cadmium (Cd) is a high-risk pathogenic toxin for hepatic diseases. Excessive mitophagy is a hallmark in Cd-induced hepatotoxicity. However, the underlying mechanism remains obscure. Mitochondrial calcium uniporter (MCU) is a key regulator for mitochondrial and cellular homeostasis. Here, Cd exposure upregulated MCU expression and increased mitochondrial Ca2+ uptake are found. MCU inhibition through siRNA or by Ru360 significantly attenuates Cd-induced excessive mitophagy, thereby rescues mitochondrial dysfunction and increases hepatocyte viability. Heterozygous MCU knockout mice exhibit improved liver function, ameliorated pathological damage, less mitochondrial fragmentation, and mitophagy after Cd exposure. Mechanistically, Cd upregulates MCU expression through phosphorylation activation of cAMP-response element binding protein at Ser133(CREBS133 ) and subsequent binding of MCU promoter at the TGAGGTCT, ACGTCA, and CTCCGTGATGTA regions, leading to increased MCU gene transcription. The upregulated MCU intensively interacts with voltage-dependent anion-selective channel protein 1 (VDAC1), enhances its dimerization and ubiquitination, resulting in excessive mitophagy. This study reveals a novel mechanism, through which Cd upregulates MCU to enhance mitophagy and hepatotoxicity.

Independent association of blood cadmium with subclinical lower extremity atherosclerosis: An observational study based on dose-response analysis.

BACKGROUND: Atherosclerosis is an increasingly public health issue globally. Previous studies have showed a causal link between heavy metal exposure and atherosclerosis. However, the association of cadmium concentration with subclinical lower extremity atherosclerosis (SLEA) remains unclear. AIMS: To investigate the association of blood cadmium with SLEA and its extent, and further analyze the potential dose-response relationship. METHODS: Blood cadmium concentration was measured using inductively coupled plasma mass spectrometry. SLEA and its extent were assessed by ultrasound diagnosis system. Multivariate models were applied to evaluate the association of blood cadmium with SLEA and its extent. Restricted cubic splines were performed to explore the potential dose-response relationship. RESULTS: This observational study consisted of 1664 participants from cardiovascular outpatient, with an average age of 62.4 years and 1218 (73.2%) men. When blood cadmium was included as a categorical variable in multivariate models, logistic regression analysis showed that high quartile in blood cadmium was an independent risk factor of SLEA (OR = 2.704, 95%CI 1.866-3.919). After log-transformed for SLEA extent parameters, linear regression analysis indicated that high quartile in blood cadmium was significantly associated with higher Crouse score (GMR = 1.21, 95%CI 1.15-1.28), plaque maximum thickness (GMR = 1.13, 95%CI 1.09-1.18) and diseased vessel count (GMR = 1.14, 95%CI 1.10-1.19), respectively. When blood cadmium was used as a continuous variable in restricted cubic splines, the dose-response relationship presented a positive progression in SLEA (P = 0.302), plaque maximum thickness (P = 0.145) and diseased vessel count (P = 0.055) apparently that did not deviate from linearity. CONCLUSIONS: Blood cadmium exhibited an independent association with SLEA, and this dose-response relationship was progressive without significant departure from linearity.

Klotho Ameliorates Vascular Calcification via Promoting Autophagy.

Vascular calcification (VC) is regarded as a common feature of vascular aging. Klotho deficiency reportedly contributes to VC, which can be ameliorated by restoration of Klotho expression. However, the specific mechanisms involved remain unclear. Here, we investigated the role of autophagy in the process of Klotho-inhibiting VC. The clinical study results indicated that, based on Agatston score, serum Klotho level was negatively associated with aortic calcification. Then, Klotho-deficient mice exhibited aortic VC, which could be alleviated with the supplementation of Klotho protein. Moreover, autophagy increased in the aorta of Klotho-deficient mice and protected against VC. Finally, we found that Klotho ameliorated calcification by promoting autophagy both in the aorta of Klotho-deficient mice and in mouse vascular smooth muscle cells (MOVAS) under calcifying conditions. These findings indicate that Klotho deficiency induces increased autophagy to protect against VC and that Klotho expression further enhances autophagy to ameliorate calcification. This study is beneficial to exploring the underlying mechanisms of Klotho regulating VC, which has important guiding significance for future clinical studies in the treatment of VC.

A rapid and reliable immunochromatographic strip for detecting paraquat poinsoning in domestic water and real human samples.

Paraquat (PQ) is one of the most commonly used herbicides, but it has polluted the environment and threatened human health through extensive and improper usage. Here, a new naked-eye PQ immunochromatographic strip was developed to recognize PQ in domestic water and real human samples within 10 min based on a novel custom-designed anti-PQ antibody. The PQ test strip could recognize PQ at a concentration as low as 10 ng/ml, reaching the high-efficiency time-of-flight mass spectrometry detection level and identifying trace amounts of PQ in samples treated with a diquat (DQ) and PQ mixture. Notably, both the performance evaluation and clinical trial of the proposed PQ strips were validated in multiple hospitals and public health agencies. Taken together, our study firstly provide the clinical PQ-targeted colloidal gold immunochromatographic test strip designed both for environment water and human sample detection with multiple advantages, which are ready for environmental monitoring and clinical practice.

Prenatal exposure to titanium dioxide nanoparticles induces persistent neurobehavioral impairments in maternal mice that is associated with microbiota-gut-brain axis.

Gestational exposure to titanium dioxide nanoparticles (TiO2NPs) has been widely reported to have deleterious effects on the brain functions of offspring. However, little attention has been paid to the neurotoxic effects of TiO2NPs on maternal body after parturition. The pregnant mice were orally administrated with TiO2NPs at 150 mg/kg from gestational day 8-21. The potential effects of TiO2NPs on the neurobehaviors were evaluated at postnatal day 60. The gut microbiota, morphological alterations of intestine and brain, and other indicators that involved in gut-brain axis were all assessed to investigate the underlying mechanisms. The results demonstrated that exposure to TiO2NPs during pregnancy caused the persistent neurobehavioral impairments of maternal mice after delivery for 60 days, mainly including behavioural changes, pathological changes in hippocampus, cortex and intestine. Our data also showed that persistent dysfunction and tissue injuries were probably associated with the disruption of gut-brain axis, manifested by the shift in the composition of gut microbial community, alteration of Sstr1, inhibition of enteric neurons and reduction of diamine oxidase contents in maternal mice. These findings provide a novel insight that regulation of gut microecology may be an alternative strategy for the protection against the neurotoxicity of TiO2NPs in pregnant women.

Polystyrene nanoparticles enhance the adverse effects of di-(2-ethylhexyl) phthalate on male reproductive system in mice.

Coexposure of nanoplastics (NPs) with other pollutants adsorbed from the surroundings has received extensive attention. Currently, the combined effects of NPs and plasticizers remain unclear. Di-(2-ethylhexyl) phthalate (DEHP) is a commonly used plasticizer that has raised much concern owing to its ubiquitous pollution and endocrine-disrupting potential. This study aimed to investigate the toxic effects on the male reproductive system upon coexposure to NPs and DEHP. The C57BL/6J mice were orally administrated with polystyrene nanoparticles (PSNPs), DEHP or both for 35 days to evaluate their effects on sperm quality, histology of testes and epididymides, testicular transcriptomic characteristics as well as expression of some important genes in the epididymides. The low-dose PSNPs used here did not induce significant changes in sperm quality, while DEHP alone or cotreatment with DEHP and PSNPs caused notable impairment, mainly manifesting as decreased sperm quality and aberrant structure of the testis and epididymis. Moreover, enhanced toxic effects were found in the cotreatment group when compared with the individual DEHP treatment group, as manifested by more obvious alterations in the sperm parameters as well as histological changes in the testis and epididymis. Testicular transcriptomic analysis revealed differential regulation of genes involved in immune response, cytoplasmic pattern recognition receptor signaling pathways, protein ubiquitination, oxidative stress, necrotic cell death, ATP synthesis and the cellular respiratory chain. RT-qPCR verified that the expression patterns of Cenpb, Crisp1 and Mars were changed in testes, and genes relevant to epididymal function including Aqp9 and Octn2 were downregulated in epididymides, particularly in the cotreatment group. Collectively, our results emphasize that DEHP at an environmentally relevant dose can induce male reproductive toxicity, and PSNPs may aggravate the toxic effects.

Paraquat exposure induces Parkinsonism by altering lipid profile and evoking neuroinflammation in the midbrain.

Paraquat (PQ) is the most widely used herbicide in the world and a well-known potent neurotoxin for humans. PQ exposure has been linked to increase the risk of Parkinson's disease (PD). However, the mechanism underlying its neurotoxic effects in PD pathogenesis is unclear. In our present study, C57BL/6J mice treated with PQ manifested severe motor deficits indicated by the significant reductions in suspension score, latency to fall from rotarod, and grip strength at 8 weeks after PQ exposure. Pathological hallmarks of Parkinsonism in the midbrain such as dopaminergic neuron loss, increased α-synuclein protein, and dysregulated PD-related genes were observed. Non-targeted lipidome analysis demonstrated that PQ exposure alters lipid profile and abundance, increases pro-inflammatory lipids.27 significantly altered subclasses of lipids belonged to 6 different lipid categories. Glycerophospholipids, sphingolipids, and glycerides were the most abundant lipids. Abundance of pro-inflammatory lipids such as Cer, LPC, LPS, and LPI was significantly increased in the midbrain. mRNA expressions of genes regulating ceramide biosynthesis in the midbrain were markedly up-regulated. Moreover, PQ exposure increased serum pro-inflammatory cytokines and provoked neuroinflammation in the midbrain. Pro-inflammatory lipids and cytokines in the midbrain were positively correlated with motor deficits. PQ poisoning in humans significantly also elevated serum pro-inflammatory cytokines and induced an intense systemic inflammation. In summary, we presented initial investigations of PQ induced molecular events related to the PD pathogenesis, capturing aspects of disturbed lipid metabolism, neuroinflammation, impairment of dopaminergic neurons in the midbrain, and an intense systemic inflammation. These neurotoxic effects of PQ exposure may mechanistically contribute to the pathogenesis of PQ induced Parkinsonism. Results of this study also strongly support the hypothesis that ever-increasing prevalence of Parkinson's disease is etiologically linked to the health risk of exposure to neurotoxic environmental pollutants.