Study of Huperzine A derivatives with extended protection against soman intoxication.

Pre-administration of huperzine A (Hup A) was validated to prevent poisoning from exposure to nerve agents (NAs) by reversibly inhibiting acetylcholinesterase (AChE). However, like the currently commonly used reversible inhibitors, Hup A has a short half-life and is unable to produce a long-term preventative effect. To extend the protective time of Hup A against NAs, 42 derivatives with a CN bond were designed based on the structure of Hup A in this study. All designed derivatives showed good binding capability with AChE via molecular docking. Six compounds (H3, H4, H11, H14, H16, and H25) with representative structures were selected for synthesis by Schiff base reaction, and their structures were stable. The modified Ellman's method showed the six compounds concentration-dependently inhibited AChE, and the half maximal inhibitory concentration (IC50) were higher than that of Hup A. Pretreatment of AChE with the derivatives significantly increased the IC50 of soman. In vivo experiments demonstrated H3, H4, H14, H16, and H25 had longer protective capacities against 1 × LD95 soman-induced death in mice than Hup A. The 12 h protective index showed that the protective ratios of H3, H4, H14 and H16 were 2.31, 1.85, 2.23 and 1.99 respectively, better than that of Hup A. The extended protection of the derivatives against soman may be explained by their transformation to Hup A in vivo. Furthermore, all six compounds showed lower acute oral toxicity than Hup A. Overall, our study provided an optional strategy to acquire pretreatment agents for NAs with extended action and low toxicity.

PGC 1α-Mediates Mitochondrial Damage in the Liver by Inhibiting the Mitochondrial Respiratory Chain as a Non-cholinergic Mechanism of Repeated Low-Level Soman Exposure.

This work aimed to assess whether mitochondrial damage in the liver induced by subacute soman exposure is caused by peroxisome proliferator-activated receptor-gamma coactivator 1 alpha (PGC-1α) and whether PGC-1α regulates mitochondrial respiratory chain damage. Toxicity mechanism research may provide theoretical support for developing anti-toxic drugs in the future. First, a soman animal model was established in male Sprague-Dawley (SD) rats by subcutaneous soman injection. Then, liver damage was biochemically evaluated, and acetylcholinesterase (AChE) activity was also determined. Transmission electron microscopy (TEM) was performed to examine liver mitochondrial damage, and high-resolution respirometry was carried out for assessing mitochondrial respiration function. In addition, complex I-IV levels were quantitatively evaluated in isolated liver mitochondria by enzyme-linked immunosorbent assay (ELISA). PGC-1α levels were detected with a Jess capillary-based immunoassay device. Finally, oxidative stress was analyzed by quantifying superoxide dismutase (SOD), malondialdehyde (MDA), glutathione (GSH), oxidized glutathione (GSSG), and reactive oxygen species (ROS) levels. Repeated low-level soman exposure did not alter AChE activity, while increasing morphological damage of liver mitochondria and liver enzyme levels in rat homogenates. Complex I, II and I + II activities were 2.33, 4.95, and 5.22 times lower after treatment compared with the control group, respectively. Among complexes I-IV, I-III decreased significantly (p < 0.05), and PGC-1α levels were 1.82 times lower after soman exposure than in the control group. Subacute soman exposure significantly increased mitochondrial ROS production, which may cause oxidate stress. These findings indicated dysregulated mitochondrial energy metabolism involves PGC-1α protein expression imbalance, revealing non-cholinergic mechanisms for soman toxicity.

Regulation of H9C2 cell hypertrophy by 14-3-3η via inhibiting glycolysis.

It has been reported that Ywhah (14-3-3η) reduces glycolysis. However, it remains unclear about the downstream mechanism by which glycolysis is regulated by 14-3-3η in cardiac hypertrophy. As an important regulator, Yes-associated protein (YAP) interacts with 14-3-3η to participate in the initiation and progression of various diseases in vivo. In this study, the model of H9C2 cardiomyocyte hypertrophy was established by triiodothyronine (T3) or rotenone stimulation to probe into the action mechanism of 14-3-3η. Interestingly, the overexpression of 14-3-3η attenuated T3 or rotenone induced cardiomyocyte hypertrophy and decreased glycolysis in H9C2 cardiomyocytes, whereas the knockdown of 14-3-3η had an opposite effect. Mechanistically, 14-3-3η can reduce the expression level of YAP and bind to it to reduce its nuclear translocation. In addition, changing YAP may affect the expression of lactate dehydrogenase A (LDHA), a glycolysis-related protein. Meanwhile, LDHA is also a possible target for 14-3-3η to mediate glycolysis based on changes in pyruvate, a substrate of LDHA. Collectively, 14-3-3η can suppress cardiomyocyte hypertrophy via decreasing the nucleus translocation of YAP and glycolysis, which indicates that 14-3-3η could be a promising target for inhibiting cardiac hypertrophy.

The mechanism of 14-3-3η in thyroxine induced mitophagy in cardiomyocytes.

Hyperthyroidism is becoming increasingly important as an independent risk factor for cardiovascular disease, eventually resulting in cardiac hypertrophy and heart failure. The 14-3-3 protein family subtypes regulate many cellular processes in eukaryotes by interacting with a diverse array of client proteins. Considering that the 14-3-3η protein protects cardiomyocytes by affecting mitochondrial function, exploring the biological influence and molecular mechanisms by which 14-3-3η alleviates the cardiac hypertrophy of hyperthyroidism is imperative. In vivo and in vitro, RT-PCR, Western blot, and Mitochondrial tracking assay were performed to understand the molecular mechanism of thyroxine-induced cardiomyocyte hypertrophy. HE staining, transmission electron microscopy, and immunofluorescence were used to observe intuitively changes of hearts and cardiomyocytes. The in vivo and in vitro results indicated that overexpression of the 14-3-3η ameliorated thyroxine-induced cardiomyocyte hypertrophy, whereas knockdown of the 14-3-3η protein aggravated thyroxine-induced cardiomyocyte hypertrophy. Additionally, overexpression of the 14-3-3η protein reduces thyroxine-induced mitochondrial damage and mitophagy in cardiomyocytes. Overexpression of 14-3-3η protein improves excessive mitophagy in the myocardium caused by thyroxine and thus prevents cardiac hypertrophy.

Subacute sarin exposure disrupted the homeostasis of purine and pyrimidine metabolism in guinea pig striatum studied by integrated metabolomic, lipidomic and proteomic analysis.

Sarin was used as a chemical weapon due to its high neurotoxicity and mortality. Subacute sarin induced cognitive and behavioral disorder. However, the underlying mechanism is still unclear. Here we offered a multi-omic approach for the analysis of altered metabolites, lipids, and proteins to explore the neurotoxicity of subacute sarin. Guinea pigs were administered between the shoulder blades 16.8 µg/kg of sarin in a volume of 1.0 ml/kg body weight by subcutaneous injection once daily for 14 days. At the end of the final injection, guinea pigs were sacrificed, and striatum were dissected for analysis. A total of 138 different metabolites were identified in the metabolome analysis. Lipids and lipid-like molecules is the largest group (38.41%). For lipidomic analysis, a total of 216 lipids were identified. In proteomic study, over 4300 proteins were identified and quantified. By integrating these enriched components, we demonstrated that the joint pathways disturbed by subacute sarin mainly involving lipid, purine and pyrimidine metabolism in guinea pig striatum. Overall, this study highlights the powerfulness of omics platforms to deepen the understanding of nerve agents caused neurotoxicity.

Serum 14-3-3ß protein: a new biomarker in asthmatic patients with acute exacerbation in an observational study.

BACKGROUND: The determination of systemic inflammatory markers is one of the important directions to study the pathogenesis of asthma and improve the diagnosis of asthma. Current studies have found that the 14-3-3 protein family subtypes interact with target proteins to participate in the pathogenesis of a variety of immune inflammatory diseases. However, studies on serum tyrosine3-monooxygenase/tryptophan5-monooxygenase activation protein ß (14-3-3ß) in asthma are scarce. This study aimed to assess the clinical significance of 14-3-3ß in asthmatic patients. METHODS: We recruited 54 asthmatic patients with acute exacerbation and 50 asthmatic patients with chronic persistent. The normal control group included 54 healthy individuals. Clinical characteristics, clinical indicators [fractional expiratory nitric oxide (FeNO), eosinophil count, forced vital capacity (FVC), percent of predicted FVC (FVC% predicted), forced expiratory volume in one second (FEV1), percent of predicted FEV1 (FEV1% predicted), the ratio of forced expiratory volume in one second to forced vital capacity (FEV1/FVC) and serum 14-3-3ß levels were measured to compare among each group. Spearman's rank correlation coefficient was used to evaluate the correlation between 14-3-3ß and clinical indicators. Finally, Receiver-operating characteristic (ROC) curves analysis was used to determine the sensitivity and specificity of 14-3-3ß. RESULTS: Our results showed that median (interquartile range) of serum 14-3-3ß concentration (ng/mL) in acute exacerbation group of asthma (41.18 [33.06-51.76]) was much higher than that in normal control group (24.99 [17.43-29.91]; P < 0.001) and chronic persistent group of asthma (25.88 [21.03-34.55]; P < 0.001). Spearman's correlation coefficient shows that the serum 14-3-3ß level was positively correlated with FeNO (r = - 0.292, P = 0.032) and peripheral blood eosinophil count (r = 0.328, P = 0.016), and was negatively related to FEV1/FVC (r = - 0.293, P = 0.031) in the acute exacerbation group of asthma. At the same time, the serum 14-3-3ß level was also negatively associated with FEV1 (r = - 0.297, P = 0.036) in the chronic persistent group of asthma. ROC curve analysis comparing acute exacerbation group of asthma with normal control group demonstrated a significant (P < 0.001) AUC of 0.90 (95% CI 0.85-0.96). CONCLUSION: The serum 14-3-3ß protein may become a potential biomarker in asthmatic patients with acute exacerbation.

Repeated low-dose exposures to sarin disrupted the homeostasis of phospholipid and sphingolipid metabolism in guinea pig hippocampus.

Repeated low-level exposure to sarin results to hippocampus dysfunction. Metabonomics involves a holistic analysis of a set of metabolites in an organism in the search for a relationship between these metabolites and physiological or pathological changes. The objective of the present study was to evaluate the effects of repeated exposure to low-level sarin on the metabonomics in hippocampus of a guinea pig model. Guinea pigs were divided randomly into control and sarin treated groups (n = 14). Guinea pigs in the control group received saline; while the sarin-treated group received 0.4×LD50 (16.8 µg/kg) sarin. Daily injections (a total of 14 days) were administered sc between the shoulder blades in a volume of 1.0 ml/kg body weight. At the end of the final injection, 6 animals in each group were chosen for Morris water maze test. The rest guinea pigs (n = 8 for each group) were sacrificed by decapitation, and hippocampus were dissected for analysis. Compared with the control-group, the escape latency in sarin-group was significantly (p < 0.05) longer while the crossing times were significantly decreased in the Morris water task (p < 0.05). Sarin inhibited activities of acetylcholinesterase (AChE) and neuropathy target esterase (NTE) in hippocampus. The AChE activity of hippocampus from sarin-treated groups is equivalent to 59.9 ± 6.4 %, and the NTE activity of hippocampus from sarin-groups is equivalent to 78.1 ± 8.3 % of that from control-group. Metabolites were identified and validated. A total of 14 variables were selected as potential biomarkers. Phospholipids [phosphatidylcholine (PC), phosphatidylethanolamine (PE), phosphatidic acid (PA), phosphatidylglycerol (PG), phosphatidylinositol (PI), Lysophosphatidylethanolamine (LysoPE or LPE)] and sphingolipids (SPs) [sphinganine (SA), phytosphingosine (PSO) and sphinganine-1-phosphate (SA1P)] were clearly modified. In conclusion, repeated low-dose exposures to sarin disrupted the homeostasis of phospholipid and sphingolipid metabolism in guinea pig hippocampus and may lead to a neuronal-specific function disorders. Identified metabolites such as SA1P need to be studied more deeply on their biological function that against sarin lesions. In future research, we should pay more attention to characterize the physiological roles of lipid metabolism enzymes as well as their involvement in pathologies induced by repeated low-level sarin exposure.

Diagnostic accuracy of 14-3-3 η protein in rheumatoid arthritis: A meta-analysis.

AIM: To evaluate the overall diagnostic performance of 14-3-3 η protein in patients with rheumatoid arthritis (RA). METHODS: PubMed, EMBASE, and Web of Science were searched to acquire eligible studies. Articles published in English before 20 February 2020 were included. Quality Assessment of Diagnostic Accuracy Studies 2 was used to evaluate the risk of bias and application concern of the included articles. Pooled analysis of diagnostic indicators of 14-3-3 η protein for RA was conducted by using a random effects model. Subgroup analysis was used to explore the sources of heterogeneity. Deeks' funnel plot asymmetry test was used to evaluate for the presence of publication bias. RESULTS: A total of 13 studies (1554 positive and 1934 negative participants) were included. The pooled sensitivity and specificity were 0.73 (95% CI 0.71-0.75) and 0.88 (95% CI 0.87-0.90), respectively. The pooled positive/negative likelihood were 5.98 (95% CI 4.39-8.14) and 0.28 (95% CI 0.21-0.37), respectively. In addition, the pooled diagnostic odds ratio was 23.48 (95% CI 13.76-40.08) and the area under curve was 0.9245. The results of subgroup analysis indicated that ethnicity and control group might be the source of heterogeneity. The results of sensitivity analysis were stable. No significant publication bias was found. CONCLUSIONS: The current evidence indicated that 14-3-3 η protein has moderate accuracy for the diagnosis of RA.

Nitrogen Fertilizer and Straw Applications Affect Uptake of 13C,15N-Glycine by Soil Microorganisms in Wheat Growth Stages.

This study investigated the influence of nitrogen (N) fertilizer and straw on intact amino acid N uptake by soil microorganisms and the relationship between amino acid turnover and soil properties during the wheat growing season. A wheat pot experiment was carried out with three treatments: control (CK), N fertilizer (NF) and N fertilizer plus rice straw (NS). We used stable isotope compound-specific analysis to determine the uptake of 13C,15N-glycine by soil microorganisms. In the NF treatment, microbial 13C,15N-glycine uptake was lower compared with CK, suggesting that inorganic N was the preferred N source for soil microorganisms. However, The application of straw with N fertilizer (in NS treatment) increased microbial 13C,15N-glycine uptake even with the same amount of N fertilizer application. In this treatment, enzyme activities, soil microbial biomass C and microbial biomass N increased simultaneously because more C was available. Soil mineral N and plant N contents all decreased substantially. The increased uptake of intact 13C,15N-glycine in the NS treatment can be attributed to direct assimilation by soil microorganisms to satisfy the demand for N when inorganic N was consumed.