Association of single nucleotide polymorphisms in the nuclear respiratory factor-2 beta subunit-encoding the GABPB1 gene within the occupational environment.

GABPB1, known as nuclear respiratory factor 2 (Nrf2), activates the mitochondrial genes that are responsible for antioxidant action and detoxification. Two single nucleotide polymorphisms (SNPs) of GABPB1, such as rs7181866 and rs8031031, were reported to be associated with the prevention of the increasing cancer risk caused by environmental deterioration. Between March 1 and May 1, 2018, human peripheral blood mononuclear cells (PBMCs) from a cohort of 300 volunteers working in adverse occupational environments were genotyped for the two SNPs in the present study. The SNP rs7181866 was found to be significantly greater in the male group than in the female group. Frequencies of SNP rs7181866 and bi-allele SNPs (rs7181866 + rs8031031) were significantly different between the <35-year-old group and the ≥35-year-old group. Further, multinomial logistic regression analysis of the occupational environments revealed the highest predictive frequency of SNPs for four environmental factors, of which chemical factors accounted for 15.33% rs7181866, physical factors accounted for 34.79% rs7181866 + rs8031031, physical + chemical factors accounted for 39.5% rs8031031, and unknown factors accounted for 26.5% rs7181866 + rs8031031. In conclusion, the G allele of rs7181866 was found to be significantly more susceptible than the rs8031031 allele under adverse occupational environmental factors, and physical factors such as noise, which appear to play vital roles in causing SNP mutations.

Investigating the adsorption behavior and quantitative contribution of Pb2+ adsorption mechanisms on biochars by different feedstocks from a fluidized bed pyrolysis system.

The aim of this study was to examine the qualitative and quantitative analysis of Pb2+ adsorption mechanisms performed with biochars derived from rice straw (RSBs), rice husk (RHBs) and saw dust (SDBs) at several pyrolysis temperatures (400-600 °C) in a fluidized bed system. Adsorption isotherms, kinetics, and desorption analysis were determined, and biochars were analyzed by X-ray Photoelectron Spectroscopy (XPS), Fourier Transform Infrared Spectroscopy (FTIR), Scanning Electron Microscope with Energy Dispersive Spectrometer (SEM-EDS) and Boehm titration method. The effect of minerals on Pb2+ adsorption, including precipitation and cation exchange, revealed increasing contribution of precipitation from a range of 4.13%-38.83% to a range of 34.08%-79.94% and decreasing effect of cation exchange from a range of 50.17%-69.75% to a range of 9.57%-43.47% with increasing pyrolysis temperature. However, it remained the dominant adsorption mechanism of all biochars (accounted for 69.49-89.52%). Especially, RSBs with quite high maximum adsorption capacity (qm) values (116-127.57 mgg-1) were mainly due to precipitation mechanism of Pb2+ adsorption, which exhibited better adsorption capacities than RHBs (25.15-30.40 mgg-1) and SDBs (21.81-24.05 mgg-1). Only with the fluidized bed shown in this study, 2.00t RSBs could be produced and the corresponding Pb2+ adsorption may reach 255.50kg per year depending on its maximum adsorption capacity under 500 °C pyrolysis temperature. The results suggest that RSBs produced in a fluidized bed reactor is a promising, cost-effective, engineered biochar for application of Pb2+ remediation in aqueous solutions.

Development of a novel isolation unit for entomophthoralean fungi.

Entomophthoralean fungi are important natural enemies of pests and highly co-evolved with their hosts. However, successful isolation of entomophthoralean fungi can be difficult due to their fastidious culture requirements; this is an important obstacle to research on Entomophthorales. In this study, we designed an isolation unit and evaluated it against the conventional 'descending conidia' isolation method. There was no difference in contamination rate between the methods (78% and 76% clean isolations) despite the isolation unit not requiring laminar-flow facilities. Furthermore, more conidia were collected in the new isolation unit than using a standard method. The isolation unit is efficient, convenient and is operational in the field.

Effects of miR-9 and tetramethylpyrazine on activation of hepatic stellate cells.

Micro-RNAs (miRNAs) are involved in regulation of the incidence and development of several hepatic diseases. Thus manipulating miRNAs may be a promising therapeutic strategy against these entities. In this study hepatic stellate cells (HSCs) were transfected with hsa-miR-9 or anti-hsa-miR-9, treated with tetramethylpyrazine (TMP), or subjected to treatment with TMP and hsa-miR-9 transfection (combined treatment group). Then, real-time polymerase chain reaction (PCR) was performed to measure mRNA levels of hsa-miR-9. Expression of hsa-miR-9 was highest in the combination treatment group compared with other groups, and significantly higher than TMP-treated and hsa-miR-9-transfected groups (both p<0.05). The anti-hsa-miR-9-transfected group expressed the lowest mRNA level of hsa-miR-9 with marked decrease versus control (p<0.05). Downstream factors that may be affected by miR-9 such as leptin, α-smooth muscle actin (SMA), and collagen I, as well as phosphorylation levels of Janus kinase 1 (JAK1)/signal transducer and activator of transcription 3 (STAT3) were investigated at the protein level. All these factors were regulated contrariwise to expression trends of hsa-miR-9, showing the lowest level in the combination treatment group and highest level in anti-hsa-miR-9-transfected group. These results suggest that both transfection of hsa-miR-9 and TMP can lead to upregulated endogenous expression of hsa-miR-9, inhibit activation of JAK1/STAT3 signal pathway induced by leptin, and lead to reduction of α-SMA and collagen I-thus impeding activation of HSC.

[Effect of echinacoside on replication and antigen expression of hepatitis B virus].

To verify the effect of echinacoside on replication and antigen expression of hepatitis B virus (HBV) by using HBV-transfected HepG2. 2. 15 cells as the in vitro model. The ELISA method was used to determine HBeAg and HBsAg levels in cellular supernatants. The effect of echinacoside on HBV replication was studied by using HBV transgenic mice as the in vivo model. First of all, the HBV DNA level in hepatic tissues was quantified with PCR method. Meanwhile, the serum transaminase levels and hepatic pathological changes were also evaluated. Subsequently, HBV transgenic mice were divided into five groups: the control group, the lamivudine group (50 mg · kg(-1)) and echinacoside high, medium and low dose group (50, 25 and 12.5 mg · kg(-1)). The mice were orally administered with drugs once per day for 30 days. At the 31st day, the mice serum was separated to measure HBsAg, HBeAg and HBV DNA. Additionally, the liver HBV DNA level and histopathological change were detected. The results indicated that echinacoside at 50 and 100 mg · L(-1) suppressed significantly HBsAg and HBeAg expressions on the sixth day, with the maximum inhibition ratios of 42.68% and 46.29%; And echinacoside at 100 mg · L(-1) also showed an inhibitory effect on HBV DNA. Besides, echinacoside at 50 mg · kg(-1) inhibited significantly HBsAg and HBeAg expressions of HBV transgenic mice, with the inhibition ratios of 42.82% and 29.12%, and reduced markedly the serum HBV DNA level in HBV transgenic mice. In conclusion, the study suggested that echinacoside has a strong effect against HBV replication and antigen expression.

MicroRNA-193b modulates proliferation, migration, and invasion of non-small cell lung cancer cells.

MicroRNAs have been reported to be closely related to the development of human lung cancers. However, the functions of microRNAs in non-small cell lung cancer (NSCLC) remain largely undefined. Here, we investigated the role of microRNA-193b (miR-193b) in NSCLC. Our data showed that miR-193b was markedly down-regulated in NSCLC cancer tissues compared with adjacent normal tissues. The NSCLC cell line (A549) transfected with the miR-193b exhibited significantly decreased proliferation, migration, and invasion capacities when compared with the control cells. In contrast, inhibition of miR-193b increased the proliferation, migration, and invasion of A549 cells. Moreover, miR-193b repressed the expressions of cyclin D1 and urokinase-type plasminogen activator in A549 cells. These data suggest that miR-193b is a tumor suppressor in NSCLC.

[Detection of the antigenic peptides in pHLA complexes by ultrafiltration-high performance liquid chromatography].

Recombinant HLA-Ⅰ molecules/antigenic peptide complexes (pHLA complexes) are applied in the research of human T cell-specific immune responses. The preparation of pHLA complex is based on genetic engineering and protein in vitro dilution and folding-refolding technology. In an in vitro refolding system, recombinant HLA-Ⅰ molecules correctly fold and bind with antigenic peptides to form complexes. In this study, ultrafiltration-high performance liquid chromatography (ultrafiltration-HPLC) was used for quantitative determination of the antigenic peptides in recombinant pHLA complexes, especially for those in a small amount of prepared products. By adding the recombinant HLA-Ⅰ molecules and antigenic peptides into the refolding buffer, the heavy chain (HC) and light chain (ß2m) of recombinant HLA-Ⅰ molecules were refolded and bond with the VYF antigenic peptide containing anchor residues to form a pHLA complex. The unbound free antigenic peptide VYF was removed by ultrafiltration to retain the complex. Finally, the pHLA complex was treated by acid to destroy its interaction, thus releasing the antigenic peptide. The results showed that the prepared recombinant pHLA complex was recognized by HLA-Ⅰ molecule specific antibody W6/32, which indicated that the recombinant HLA-Ⅰ class molecule had correct folding and was identified as pHLA complex. The antigen peptide VYF contained in the pHLA complex was also detected by ultrafiltration-HPLC, so it is feasible to apply ultrafiltration-HPLC for determination of pHLA complex. Compared with Western blotting, the concentration of antigenic peptides detected by ultrafiltration-HPLC was 0-9 µg/mL. The binding conditions can be optimized according to the amount of antigenic peptides bound in the complex in order to improve the folding efficiency of HLA-Ⅰ molecules and promote the binding of HLA-Ⅰ molecules to antigenic peptides. The production rate of pHLA complexes in the refolding system can also be calculated according to the content of antigenic peptides bound by pHLA complexes. Therefore, ultrafiltration-HPLC in this study can be used for the quality control of the preparation process of pHLA complexes, and may facilitate the research of T cell-specific immunity, artificial antigen-presenting cells, and development of specific tetramer probe applications.

Identification of fatty acid synthase from the Pacific white shrimp, Litopenaeus vannamei and its specific expression profiles during white spot syndrome virus infection.

Fatty acid synthase (FAS) in animal tissues consists of two identical monomers and is known to be a complex multi-functional enzyme that plays an important role in energy homeostasis. However, there are few reports of studies focused on the relationship between FAS and virus infection in invertebrates. In the present study, we cloned the FAS gene from an economically important invertebrate, the Pacific white shrimp Litopenaeus vannamei. The full-length FAS cDNA is 8268 bp, including a 5'-terminal untranslated region of 137 bp, a 3'-terminal untranslated region of 601 bp and an open reading frame of 7530 bp. FAS cDNA encodes a polypeptide of 2509 amino acid residues that contains a typical ß-ketoacyl synthase (KS) domain at the N-terminus, next to a malonyl/acetyltransferase (MAT) domain, a dehydrase domain, an enoyl reductase domain, a ketoacyl reductase domain, a phosphopantetheine attachment site domain and a thioesterase domain at the C-terminus. Quantitative real-time RT-PCR revealed the up-regulated expression of FAS in L. vannamei hepatopancreas and muscle after white spot syndrome virus (WSSV) infection. The expression of FAS in muscle was 13.03-fold greater than that in the control (p<0.05) and 2.93-fold greater in hepatopancreas (p>0.05). Meanwhile, expression of the fatty acid-binding protein (FABP), another important factor in lipid metabolism, was increased in muscle to 19.20-fold greater than that in the control (p<0.05) but only 0.76-fold in hepatopancreas (p>0.05). These results implied that WSSV infected body surface tissues, but there was very little infection of internal organs. We suggest that the increase of FAS expression is induced in WSSV-infected shrimps, and the virus changes the lipid metabolism of the host, which directly affects virus assembly or defense against virus infection.

Aqueous synthesis and fluorescence-imaging application of CdTe/ZnSe core/shell quantum dots with high stability and low cytotoxicity.

CdTe/ZnSe core/shell quantum dots were directly synthesized in an aqueous condition by heating a mixed solution of ZnCI2, NaHSe and CdTe QDs in the presence of mercaptosuccinic acid as a stabilizer. By controlling the size and composition, the CdTe/ZnSe QDs with emission wavelength ranging from 540 to 630 nm, high quantum yield (44%) and narrow full width at half maximum (FWHM) could be obtained. Characterization results with HRTEM, XRD and EDX have shown that the synthesized CdTe/ZnSe QDs have good monodispersity and a nice crystal structure, and exhibited better stability and less cytotoxicity as compared with CdTe QDs. Furthermore, luminescent QD-IgG bioprobes were produced to detect the breast cancer marker Her2 on the surface of fixed MCF-7 cancer cells for their optical imaging.

Adsorption characteristics and mechanism of Pb(II) by agricultural waste-derived biochars produced from a pilot-scale pyrolysis system.

The objective of this study was to investigate the feasibility of removing Pb2+ by pilot-scale fluidized bed biochar, and then to put forward an industrial-scale fluidized bed pyrolysis progress of cogeneration of biochar and high-temperature gas. Corn stalk biochars (CSBs) were prepared at 400-600 °C, in which the maximum Pb2+adsorption capacity (Qm) of CSB450 is 49.70 mg⋅g-1 at the optimal condition. Adsorption isotherms, kinetics, and thermodynamics were determined, and Pb2+-loaded biochar was analyzed by fourier transform infrared spectroscopy (FTIR), x-ray photoelectron spectroscopy (XPS), x-ray diffraction (XRD) and scanning electron microscope with energy dispersive spectrometer (SEM-EDS). Ion exchange, complexation and mineral precipitation together contributed to Pb2+ adsorption on CSBs. For high-temperature CSBs with fewer oxygen functional groups (OFGs) and stronger aromatization, Pb2+ adsorption by ion exchange and functional group complexation was reduced. The mineral precipitationwas formed during the adsorption process. Using the pilot-scale fluidized bed in this study, the carbon yield per year would achieve 31.79 t, and about 1.58 t of Pb2+ would be adsorbed according to the adsorption capacity at the pyrolytic temperature of 450 °C.The results are beneficial to screen for effective biochar as a cost-effective industrial adsorbent to remove Pb2+ in contaminated water.

Use of electrical penetration graphs (EPG) and quantitative PCR to evaluate the relationship between feeding behaviour and Pandora neoaphidis infection levels in green peach aphid, Myzus persicae.

A real-time qPCR method was developed, validated, and used to quantity the fungal pathogen, P. neoaphidis, within aphids at different times during infection; colonization rate fitted the Gompertz model well (R2 = 0.9356). Feeding behaviour of P. neoaphidis-infected and uninfected M. persicae were investigated, for the first time, using DC-electrical penetration graphs (DC-EPG) that characterized the waveforms made during different aphid stylet probing periods corresponding to epidermis penetration, salivation and ingestion. In the 6 h following the 12-h incubation period (to achieve infection), there were significant differences in the number of events of Np (non-probing) and C (stylet pathway) between infected and uninfected aphids. However, the difference between total duration of Np and C were not significantly different between infected and uninfected aphids. There were no significant differences in the number of events or total duration of E1 (phloem salivation) or E2 (phloem ingestion) between infected and uninfected aphids. There were significant differences in mean number of events and total duration of the pd waveform (intracellular punctures) in infected and uninfected aphids. In the 16 h prior to death, the same differences in behaviour were observed but they were even more obvious. Furthermore, the total duration time of E2 was significantly greater in uninfected aphids than infected aphids, a change that had not been observed in the first 6 h observation period. In conclusion, qPCR quantification demonstrated 'molecular' colonization levels throughout infection, and EPG data analysis during the two periods (during early infection and then during late infection just prior to death) demonstrated the actual physical effects of fungal infection on feeding behaviour of M. persicae; this has the potential to decrease the aphid's capacity of transmission and dispersal. These studies increase our understanding of the interaction between P. neoaphidis and its host aphid.

Identification and characterization of follistatin as a novel angiogenin-binding protein.

Angiogenin enhances tumorigenesis. However, the mechanisms of angiogenin-induced angiogenesis and cancer cell proliferation remain elusive. In this study, follistatin was identified as a binding partner of angiogenin by a yeast two-hybrid screen and confirmed by a pull-down experiment. The interaction of fluorescently tagged angiogenin and follistatin was monitored in real time by a laser confocal microscope and shown to localize at the sub-nuclear region of HeLa cells. Additional yeast two-hybrid analysis revealed that domains 2 and 3 of follistatin were the minimal structure requirement for angiogenin binding. These findings provide new clues for further studies on the mechanisms of angiogenin-induced angiogenesis or cancer cell growth.

Hepatitis E virus chimeric DNA vaccine elicits immunologic response in mice.

AIM: To construct the plasmid pcHEV23 containing fragments of HEV ORF2 and ORF3 chimeric gene and to assess its ability to elicit specific immunologic response in mice. METHODS: The gene encoding the structural protein of HEV ORF2 fragment and full-length ORF3 was amplified by PCR. The PCR products were cloned into an eucaryotic expression plasmid pcDNA3. The resulting plasmid pcHEV23 was used as a DNA vaccine to inoculate BALB/c mice intramuscularly thrice at a dose of 100 or 200 microg. Mice injected with empty pcDNA3 DNA or saline served as control and then specific immune responses in the mice were detected. RESULTS: After 2-3 times of inoculation, all mice injected with pcHEV23 had anti-HEV IgG seroconversion and specific T lymphocyte proliferation. The lymphocyte stimulation index in the group immunized with pcHEV23 (3.1+/-0.49) was higher than that in the control group (0.787+/-0.12, P<0.01). None in the control group had a detectable level of anti-HEV IgG. CONCLUSION: DNA vaccine containing HEV ORF2 and ORF3 chimeric gene can successfully induce specific humoral and cellular immune response in mice.

Epigallocatechin-3-O-Gallate Protects Against Hepatic Damage and Testicular Toxicity in Male Mice Exposed to Di-(2-Ethylhexyl) Phthalate.

The aim of this study was to examine the effects of epigallocatechin-3-O-gallate (EGCG) on hepatic damage and testicular toxicity in male mice exposed to daily oral administration of di-(2-ethylhexyl) phthalate (DEHP). A mouse model was used to assess the effects of daily intraperitoneal EGCG injection on hepatic and testicular damage. Histological and mitochondrial membrane potential results revealed that EGCG treatment significantly arrested the progression of hepatic damage. EGCG treatment resulted in significant suppression of liver injury (i.e., reduced activities of alanine aminotransferase [ALT] and aspartate aminotransferase [AST]). The development of DEHP-induced hepatic and testicular damage altered the testosterone concentration in mouse serum, which could affect the reproductive ability of male mice. Moreover, EGCG treatment markedly attenuated testes lesions, sperm deformity, and spermatogenic cell apoptosis. At the molecular level, hepatic CYP3A4 expression was substantially reduced by EGCG treatment in mice exposed to DEHP compounds, whereas testicular aromatase expression was increased significantly in testes. Thus, these results demonstrate that EGCG administration may protect against liver damage and reproductive toxicity in males exposed to DEHP.

Dynamic correlation between induction of the expression of heme oxygenase-1 and hepatitis B viral replication.

Heme oxygenase­1 (HO­1) possesses significant potential as a drug target for hepatitis B, which may be transferable to patient therapy. The aim of the present study was to clarify the dynamic correlation between the hepatitis B virus (HBV) and HO­1. The levels of HBV replication and expression of HO­1 were investigated in HepG2.2.15 hepatoma cells following exposure to 5­50 µM hemin for 1­6 days. The mRNA expression levels of HO­1 were then detected using reverse transcription­quantitative polymerase chain reaction (RT­qPCR). HBV replication levels were determined by enzyme­immunoassay and a PCR­fluorescence quantitation assay. The results of the present study demonstrated that the mRNA expression levels of HO­1 increased in a dose­dependent manner in the HepG2.2.15 cells, following exposure to 5­50 µM hemin. The mRNA expression levels of HO­1 reached a peak following exposure of the cells to hemin for three days, subsequently the expression of HO­1 decreased. Following exposure to hemin at an optimal concentration of 50 µM for 1­6 days, the levels of the hepatitis B surface antigen (HBsAg) and hepatitis B e antigen (HBeAg) in the cells were significantly reduced. This marked reduction in the expression of HBsAg and HBeAg reached its peak on the first day, following which the inhibition weakened as the duration of exposure increased. In addition, the inhibition of HBV DNA replication increased with the a longer duration of exposure. Furthermore, HBV DNA levels were significantly decreased following exposure to hemin for 3­6 days. In conclusion, the present study demonstrated that induced expression of HO­1 interfered with HBV replication in a dose and time­dependent manner, implying that a reduction of the HBV viral load may contribute to upregulation in the expression of HO­1.

Hepatoprotective and antiviral properties of isochlorogenic acid A from Laggera alata against hepatitis B virus infection.

ETHNOPHARMACOLOGICAL RELEVANCE: The aim of this study was to determine the anti-hepatitis B effect of isochlorogenic acid A isolated from Laggera alata (Asteraceae), a traditional Chinese herbal medicine. MATERIALS AND METHODS: The anti-hepatitis B activity of isochlorogenic acid A was evaluated by the D-galactosamine (D-GalN)-induced HL-7702 hepatocyte damage model and the HBV-transfected HepG2.2.15 cells. RESULTS: Isochlorogenic acid A significantly improved HL-7702 hepatocyte viability and markedly inhibited the productions of HBsAg and HBeAg. The inhibitory rates of isochlorogenic acid A on the HBsAg and HBeAg expressions were 86.9% and 72.9%, respectively. In addition, isochlorogenic acid A declined markedly the content of hepatitis B virus covalently closed circular DNA (HBV cccDNA) and induced significantly the heme oxygenase-1 (HO-1) expression in HepG2.2.15 cells. CONCLUSIONS: Isochlorogenic acid A was verified to possess the potent anti-hepatitis B activity. The anti-HBV target of isochlorogenic acid A is probably associated with blocking the translation step of the HBV replication. Overexpression of HO-1 may contribute to the anti-HBV activity of isochlorogenic acid A by reducing the stability of the HBV core protein and thus blocking the refill of nuclear HBV cccDNA. Additionally, the hepatoprotective effect of isochlorogenic acid A could be achieved by its antioxidative property and induction of HO-1.

Identification of two hsp90 genes from the marine crab, Portunus trituberculatus and their specific expression profiles under different environmental conditions.

Hsp90, a molecular chaperone, was generally thought to be a unique cytoplasmic form in invertebrates, playing important roles in multiple cellular stress responses. Now, two cytoplasmic Hsp90 cDNAs (ptHSP90-1 and ptHSP90-2 genes) were isolated from an invertebrate - crab Portunus trituberculatus. Main features, sequence identities and phylogenetic analysis with other species were described. Expression profiles in tissues and under stressful conditions were analyzed using semi-quantitative RT-PCR method. ptHSP90-1 and ptHSP90-2 were constitutively expressed with higher transcript levels in ovary and muscle, respectively. A cold treatment rapidly activated both ptHSP90s transcription in hepatopancreas and gill, but caused the ptHSP90-2 mRNA decrease in muscle and ovary. Under heat treatment ptHSP90-1 mRNA was accumulated in hepatopancreas and muscle (but down-regulated in ovary), while ptHSP90-2's transcription tendency in each tissue was the same as that in cold shock. Moreover, the transcriptional levels of both ptHSP90 genes under Cu(2+) stress were evaluated. This crab exposed to the low and high salinity exhibited either lower expression levels of both ptHSP90s or no changes in four tissues except the up-regulation of ptHSP90-2 transcription in hepatopancreas. These results suggested there were at least two Hsp90s in P. trituberculatus, which played differing roles in physiological and stressful conditions.

Alpha-actinin-2, a cytoskeletal protein, binds to angiogenin.

Angiogenin is an angiogenic factor which is involved in tumorigenesis. However, no particular intracellular protein is known to interact directly with angiogenin. In the present study, we reported the identification of alpha-actinin-2, an actin-crosslinking protein, as a potential angiogenin-interacting partner by yeast two-hybrid screening. This interaction was confirmed by different approaches. First, angiogenin was pulled down together with His-tagged alpha-actinin-2 by Ni(2+)-agarose resins. Second, alpha-actinin-2 was coimmunoprecipitated with angiogenin by anti-angiogenin monoclonal antibody. Third, the in vivo interaction of these two proteins was revealed by fluorescence resonance energy transfer analysis. Since members of alpha-actinin family play pivotal roles in cell proliferation, migration, and invasion, the interaction between alpha-actinin-2 and angiogenin may underline one possible mechanism of angiogenin in angiogenesis. Our finding presents the first evidence of an interaction of a cytosolic protein with angiogenin, which might be a novel interference target for anti-angiogenesis and anti-tumor therapy.