Maternal Lead Exposure Impairs Offspring Learning and Memory via Decreased GLUT4 Membrane Translocation.

Lead (Pb) can cause a significant neurotoxicity in both adults and children, leading to the impairment to brain function. Pb exposure plays a key role in the impairment of learning and memory through synaptic neurotoxicity, resulting in the cognitive function. Researches have demonstrated that Pb exposure plays an important role in the etiology and pathogenesis of neurodegenerative diseases, such as Alzheimer's disease. However, the underlying mechanisms remain unclear. In the current study, a gestational Pb exposure (GLE) rat model was established to investigate the underlying mechanisms of Pb-induced cognitive impairment. We demonstrated that low-level gestational Pb exposure impaired spatial learning and memory as well as hippocampal synaptic plasticity at postnatal day 30 (PND 30) when the blood concentration of Pb had already recovered to normal levels. Pb exposure induced a decrease in hippocampal glucose metabolism by reducing glucose transporter 4 (GLUT4) levels in the cell membrane through the phosphatidylinositol 3 kinase-protein kinase B (PI3K-Akt) pathway. In vivo and in vitro GLUT4 over-expression increased the membrane translocation of GLUT4 and glucose uptake, and reversed the Pb-induced impairment to synaptic plasticity and cognition. These findings indicate that Pb exposure impairs synaptic plasticity by reducing the level of GLUT4 in the cell membrane as well as glucose uptake via the PI3K-Akt signaling pathway, demonstrating a novel mechanism for Pb exposure-induced neurotoxicity.

The Alexipharmic Mechanisms of Five Licorice Ingredients Involved in CYP450 and Nrf2 Pathways in Paraquat-Induced Mice Acute Lung Injury.

Oxidative stress is an important mechanism in acute lung injury (ALI) induced by paraquat (PQ), one of the most widely used herbicides in developing countries. In clinical prophylaxis and treatment, licorice is a widely used herbal medicine in China due to its strong alexipharmic characteristics. However, the corresponding biochemical mechanism of antioxidation and detoxification enzymes induced by licorice's ingredients is still not fully demonstrated. In this study, the detoxification effect of licorice was evaluated in vivo and in vitro. The detoxification and antioxidation effect of its active ingredients involved in the treatment was screened systematically according to Absorption, Distribution, Metabolism, and Excretion (ADME): predictions and evidence-based literature mining methods in silico approach. Data shows that licorice alleviate pulmonary edema and fibrosis, decrease Malondialdehyde (MDA) contents and increase Superoxide Dismutase (SOD) activity in PQ-induced ALI mice, protect the morphologic appearance of lung tissues, induce cytochrome 3A4 (CYA3A4) and Nuclear factor erythroid 2-related factor 2 (Nrf2) expression to active detoxification pathways, reduce the accumulation of PQ in vivo, protect or improve the liver and renal function of mice, and increase the survival rate. The 104 genes of PPI network contained all targets of licorice ingredients and PQ, which displayed the two redox regulatory enzymatic group modules cytochrome P450 (CYP450) and Nrf2 via a score-related graphic theoretic clustering algorithm in silico. According to ADME properties, glycyrol, isolicoflavonol, licochalcone A, 18beta-glycyrrhetinic acid, and licoisoflavone A were employed due to their oral bioavailability (OB) ≥ 30%, drug-likeness (DL) ≥ 0.1, and being highly associated with CYP450 and Nrf2 pathways, as potential activators to halt PQ-induced cells death in vitro. Both 3A4 inhibitor and silenced Nrf2 gene decreased the alexipharmic effects of those ingredients significantly. All these disclosed the detoxification and antioxidation effects of licorice on acute lung injury induced by PQ, and glycyrol, isolicoflavonol, licochalcone A, 18beta-glycyrrhetinic acid, and licoisoflavone A upregulated CYP450 and Nrf2 pathways underlying the alexipharmic mechanisms of licorice.

Time-resolved fluorescence and chemometrics-assisted excitation-emission fluorescence for qualitative and quantitative analysis of scopoletin and scopolin in Erycibe obtusifolia Benth.

This paper presents a new strategy for qualitative identification of scopoletin and scopolin in Erycibe obtusifolia Benth using time-resolved (lifetimes) fluorescence and quantitative analysis with chemometrics-assisted excitation-emission matrix (EEM) fluorescence. Due to the significant spectral overlapping among analytes and interference, the use of the more selective time-resolved fluorescence is proposed for qualitative identification in quality control of traditional Chinese medicine (TCM) for the first time. Using the strategy of combining EEM fluorescence with second-order calibration method, i.e. parallel factor analysis (PARAFAC), the simultaneous quantification of scopoletin and scopolin in the complex system of Erycibe obtusifolia Benth was achieved successfully. The predicted concentrations were compared with the values obtained using high performance liquid chromatography-coupled to fluorimetric detector (HPLC-FLD), and no significant differences between them were observed. Therefore, the proposed methods using time-resolved fluorescence for qualitative analysis and EEMs coupled with second-order calibration for quantitative analysis in TCM are comparable and provide a suitable alternative to the chromatography-based method.

[The network pharmacological mechanisms of four anti-vitiligo Uyghur medicines based on Phlegmatic temperament theory].

To explore the network pharmacological mechanisms of four anti-vitiligo Uyghur medicines based on the Phlegmatic temperament theory. First, The anti-vitiligo Uyghur medicine formulas based on Phlegmatic temperament theory were collected. The pharmacokinetic characteristic of main compounds in four anti-vitiligo Uyghur medicines were obtained by using admetSAR. The targets of active compounds were predicted via bSDTNBI (balanced substructure-drug-target network-based inference model) method. Then, biological process (BP) and molecular function (MF) enrichment analysis of targets were analysed via DAVID database. Constructing anti-vitiligo Uyghur medicine formula-Uyghur medicines network model (FMI network) and Uyghur medicines-active compounds-targets-BP-Hilit network model (MCTBHI network), we utilized closeness centrality to analyse key Uyghur medicines, active compounds, key targets as well as Hilit. Finally, the in vitro melanin production model of C57BL/6 mice was used to verify the ability of the active compounds to improve melanogenesis. The results showed that Psoralea corylifolia, Vernonia anthelmintica, Syzygium aromaticum and Anacyclus pyrethrum were the key Uyghur medicines in the FMI network. There were 22 active compounds with a relatively higher bioavailability interacted with 58 therapeutic targets. These active compounds were mainly composed of coumarins and flavonoids. In the MCTBHI network, the MF of 58 therapeutic targets was related to steroid hormone receptor activity, heme binding and enzyme binding functhon. Classification of the Hilit according to the BP of 58 therapeutic targets, the first place was the blood, followed by the lymph, the cerebrospinal fluid and digestive juice. It was found that the expression of some targets located in the skin was closed to the heart muscle, lymph node, spleen, cerebral cortex and so on, which were the main places for Hilit. In particular, ESR1, PTGS2, PPARA, PPARG, PTGS1 and CA2 were regulated by the flavonoids (kaempferide and isorhamnetin). The in vitro melanin production model showed that kaempferide and isorhamnetin could promote the melanin production in C57BL/6 mice ear skin. Based on admetSAR and bSDTNBI, we used network pharmacological method to construct a systematic means of studying anti-vitiligo Uyghur medicines, providing clues for the further study of the modern molecular mechanisms of Phlegmatic temperament.

Network pharmacological mechanisms of Vernonia anthelmintica (L.) in the treatment of vitiligo: Isorhamnetin induction of melanogenesis via up-regulation of melanin-biosynthetic genes.

BACKGROUND: Vitiligo is a long-term skin disease characterized by the loss of pigment in the skin. The current therapeutic approaches are limited. Although the anti-vitiligo mechanisms of Vernonia anthelmintica (L.) remain ambiguous, the herb has been broadly used in Uyghur hospitals to treat vitiligo. The overall objective of the present study aims to identify the potential lead compounds from Vernonia anthelmintica (L.) in the treatment of vitiligo via an oral route as well as the melanogenic mechanisms in the systematic approaches in silico of admetSAR and substructure-drug-target network-based inference (SDTNBI). RESULTS: The results showed that the top 5 active compounds with a relatively higher bioavailability that interacted with 23 therapeutic targets were identified in Vernonia anthelmintica (L.) using admetSAR and SDTNBI methods. Among these compounds, Isorhamnetin and Kaempferide, which are methyl-flavonoids, performed 1st and 2nd. Isorhamnetin and Kaempferide significantly increased the expression of melanin-biosynthetic genes (MC1R, MITF, TYR, TYRP1 and DCT) and the tyrosinase activity in B16F10 cells. Isorhamnetin and Kaempferide significantly increased the mRNA-expression of melanin-biosynthetic genes (MC1R, MITF, TYR, TYRP1 and DCT), the protein level of MITF and the tyrosinase activity. Based on the SDTNBI method and experimental verification, Isorhamnetin and Kaempferide effectively increased melanogenesis by targeting the MC1R-MITF signaling pathway, MAPK signaling pathway, PPAR signaling pathway (PPARA, PPARD, PPARG), arachidonic acid metabolism pathway (ALOX12, ALOX15, CBR1) and serotonergic synapses (ALOX12, ALOX15) in the treatment of vitiligo from a network perspective. CONCLUSION: We identified the melanogenic activity of the methyl-flavonoids Isorhamnetin and Kaempferide, which were successfully predicted in a network pharmacological analysis of Vernonia anthelmintica (L.) by admetSAR and SDTNBI methods.

Network analysis reveals crosstalk between autophagy genes and disease genes.

Autophagy is a protective and life-sustaining process in which cytoplasmic components are packaged into double-membrane vesicles and targeted to lysosomes for degradation. Accumulating evidence supports that autophagy is associated with several pathological conditions. However, research on the functional cross-links between autophagy and disease genes remains in its early stages. In this study, we constructed a disease-autophagy network (DAN) by integrating known disease genes, known autophagy genes and protein-protein interactions (PPI). Dissecting the topological properties of the DAN suggested that nodes that both autophagy and disease genes (inter-genes), are topologically important in the DAN structure. Next, a core network from the DAN was extracted to analyze the functional links between disease and autophagy genes. The genes in the core network were significantly enriched in multiple disease-related pathways, suggesting that autophagy genes may function in various disease processes. Of 17 disease classes, 11 significantly overlapped with autophagy genes, including cancer diseases, metabolic diseases and hematological diseases, a finding that is supported by the literatures. We also found that autophagy genes have a bridging role in the connections between pairs of disease classes. Altogether, our study provides a better understanding of the molecular mechanisms underlying human diseases and the autophagy process.

Adult lead exposure increases blood-retinal permeability: A risk factor for retinal vascular disease.

Low-to-moderate level developmental and adult lead exposure produces retinal dysfunction and/or degeneration in humans and experimental animals. Although high level in vivo or in vitro lead disrupts blood-brain-barrier tight junctions and increases its permeability, the blood-retinal-barrier (BRB) has not been examined. There were four overall goals. First, generate environmentally relevant dose-response models of short-term lead exposure in adult rats. Second, assess retinal histology and functional integrity of the BRB. Third, investigate the transmembrane proteins occludin and claudin-5 as targets mediating the increased BRB permeability. Fourth, examine the contribution of the PI3K-Akt signaling pathway as a mechanism underlying increased BRB permeability. Young adult rats were given water, 0.01% or 0.02% lead drinking solutions for six weeks. In control, 0.01% and 0.02% groups the six week mean blood [Pb] were 1, 12.5 and 19µg/dl, respectively. We employed histology, stereology, quantitative image analysis, immunoblots and densitometry, and pharmacology techniques. Major findings were that adult lead exposure produced dose-dependent 1) decreases in outer and inner nuclear layer thickness, 2) increases in BRB permeability, 3) decreases in occludin and claudin-5 expression, 4) increases in pAkt (Ser473), but not pAkt (Thr308), expression, and 5) wortmannin partially or completely blocked the increased BRB permeability and changes in protein expression. These results indicate that lead-induced increases in PI3K-Akt signaling partially underlie the increased BRB permeability and advance our knowledge about lead-induced retinotoxicity. Furthermore, they suggest that environmental and occupational lead exposures are risk factors for increased BRB permeability in diseases such as age-related macular degeneration, diabetes and stroke.

[Effect of rapamycin on proliferation of rat heart valve interstitial cells in vitro].

OBJECTIVE: To investigate the effect of rapamycin on the proliferation of rat valvular interstitial cells in primary culture. METHODS: The interstitial cells isolated from rat aortic valves were cultured and treated with rapamycin, and the cell growth and cell cycle changes were analyzed using MTT assay and flow cytometry, respectively. RT-PCR was used to detect mRNA expression levels of S6 and P70S6K in cells, and the protein expressions level of S6, P70S6K, P-S6, and P-P70S6K were detected using Western blotting. RESULTS: Rat aortic valvular interstitial cells was isolated successfully. The rapamycin-treated cells showed a suppressed proliferative activity (P<0.05), but the cell cycle distribution remained unaffected. Rapamycin treatment resulted in significantly decreased S6 and P70S6K protein phosphorylation level in the cells (P<0.05). CONCLUSION: The mechanism by which rapamycin inhibits the proliferation of valvular interstitial cells probably involves suppression of mTOR to lower S6 and P70S6K phosphorylation level but not direct regulation of the cell cycle.

[Genotypes distribution among hepatitis B virus infected patients with different immune statuses in Guangxi north region].

In order to find out the distribution of Genotype of those people infected with HBV (hepatitis B virus) from north Guangxi and the relationship between different immune status of HBV infected people and their genotypes, the HBV infected people are classified into three types according to immune tolerance, immune clearance ( response) and immune incompetence (residues). 150 cases from each type, a total of 450 cases are chosen to be tested with real time fluorescence quantitative PCR assay for detection of HBV infection in three kinds of different immune state of the HBV genotype. In the 450 cases, 323 cases belong to type B, 94 cases belong to type B, 23 cases belong to mixed type B+C and 10 cases belong to none B and none C type. Type B are the majority in all the three HBV immune status, made up to 70%, 78%, 67.33% of each type. The different immune state genotype proportion difference don't have statistical significance; immune state and genotypic correlation isn't statistically significant; type B HBV-DNA load is higher than that of type C, groups of persons aged 30 years or older with type C are significantly higher than that of < 30 years of age, the difference was statistically significant; among the genotypes of alanine aminotransferase (ALT), aspartate aminotransferase (AST), total bilirubin (TBil) positive rate showed no significant difference between male and female; there was no significant difference in genotype distribution. The results show that, in North Guangxi HBV genotypes B, C accounts for the proportion, a small amount of B+C hybrid, occasionally fails to type HBV infection; among immune tolerance, immune clearance (response) and immune incompetence (residues) type B are in majority in these three kinds of immune state, chronic HBV infection immunity with the HBV genotype correlations were not statistically significant.

Agrobacterium tumefaciens-mediated genetic transformation of the phytopathogenic fungus Penicillium digitatum.

Agrobacterium tumefaciens-mediated transformation (ATMT) system was assessed for conducting insertional mutagenesis in Penicillium digitatum, a major fungal pathogen infecting post-harvest citrus fruits. A transformation efficiency of up to 60 transformants per 10(6) conidia was achieved by this system. The integration of the hph gene into the fungal genome was verified by polymerase chain reaction (PCR) amplification and sequencing. These transformants tested were also shown to be mitotically stable. Southern blot analysis of 14 randomly selected transformants showed that the hph gene was randomly integrated as single copy into the fungal genome of P. digitatum. Thus, we conclude that ATMT of P. digitatum could be used as an alternatively practical genetic tool for conducting insertional mutagenesis in P. digitatum to study functional genomics.

[Synthesis and fluorescence characteristics of Eu-TB complexes with trimesic acid].

A series of mixed rare earth complexes Eu(1-x)Tb(x) (BTC) x nH2O(H3BTC=1,3,5-H3BTC, x = 0.1, 0.3, 0.5, 0.7, 0.9, 1.0) were synthesized by the reaction of trivalent rare earth chlorides with trimesic acid. The contents of rare earth were mearsured by using EDTA titration method, the element analyses of C and H were performed by Vario EL elemental analyzer. Their infrared spectra were recorded by FTIR-8900 infrared spectrophotometer with KBr pellet, and the fluorescence spectra were recorded by using a Hitachi F-4500 fluorescence spectrophotometer at room temperature. These results show that these complexes Eu(1-x) Tb(x) (BTC) x nH2O can emit intense fluorescence from europium ion, the intensity of the fluorescence of the series complexes is greater in (5)D0-->7F(1,2), and both the positions ((5)D0-->7F(1,2)) show splitting emission peaks, but the peak positions of their emission spectra basically remain unchanged. These may be caused by the different coordinating pattern of the carboxylate groups. Additionally, the fluorescence intensity of europium ions was sensitized by terbium ions and that of terbium ions was quenched by europium ions. in these complexes. It was attributed to efficient energy transfer from the strongly absorbing ligand to the rather poorly absorbing lanthanide ion. and from Tb3+ to Eu3+. Moreover, the fluorescence emission of terbium ions increased markedly in these complexes.

[Synthesis and fluorescence properties of the Tb(Ln) complexes with trimesic acid (Ln = Y, Gd)].

In the present paper two series of mixed rare earth complexes Tb(1-x)LnxBTC x nH2O (Ln = Y, Gd, x = 0, 0.1, 0.3, 0.5, 0.7, 0.9) were synthesized by hydrothermal method. The contents of rare earth were measured by using EDTA titration method; the element analyses of C and H were performed by using Vario EL III elemental analyzer, and the IR spectra were recorded by FTIR-8900 infrared spectroscopy with KBr pellet. Then the molecular formula of the complexes were determined to be Tb(1-x)LnxBTC x 0.5H2O (Ln = Y, Gd). The fluorescence spectrum of all the complexes were recorded by using a Hitachi F-4500 fluorescence spectrophotometer at room temperature. The results indicated that all the complexes emitted characteristic fluorescence of Tb3+, and the fluorescence intensities of the complexes were obviously enhanced with doping rare earth ion of Y3+ or Gd3+. It was showed that Tb3+ was sensitized by doping rare earth ions. This may be because there was intra molecular energy transfer in the complexes. Being doped with the rare earth ion the emission peak positions did not change. Among the four emission peaks, 5D4 --> 7F5 (544 nm) was the strongest one, and it was found when the proportion is Y3+ : Tb3+ = 0.5 : 0.5 or Gd3+ : Tb3+ = 0.3 : 0.7, the fluorescence intensity of Tb(1-x)YxBTC x 0.5H2O or Tb(1-x)GdxBTC x 0.5H2O was the greatest, meanwhile the sensitization degree was bigger with doping Gd3+ than doping Y3+ at 544 nm.