Impact analysis of behavior of front-line managers on employee safety behavior by integrating interpretive structural modeling and Bayesian network.

Employee safety behavior is a basic element of enterprise work safety. The results of accident investigations and risk assessments in enterprises indicate that management factors are some of the most important factors that affect employee safety behavior. The purpose of this study is to explore the relationship between the behavior of front-line managers (FLMs) and employee safety behavior by integrating a qualitative method, i.e., the interpretive structural model (ISM), and a quantitative method, i.e., the Bayesian network (BN). The results of the BN analysis showed that safety incentives and safety communication were the best predictors of safety participation, while safety supervision and safety control were the best predictors of safety compliance. Moreover, the results revealed that an instantaneous improvement of safety communication, safety incentives, safety supervision and safety guidance was the most effective joint measure to reach a high-level of safety behavior of employees in the workplace.

Variations of forage yield and forage-livestock balance in grasslands over the Tibetan Pla-teau, China.

An in-depth understanding of variations in grassland productivity and forage-livestock balance is the basis of ecological barrier construction and ecosystem conservation in the Qinghai-Tibetan Plateau. Using an ecohydrological process-based model VIP with remotely sensed vegetation index and leaf area index, we simulated the spatial and temporal variations of grassland productivity in the Tibetan Plateau in 2000-2018. The variations in the status of forage-livestock balance at the county level were analyzed, combining with agriculture and animal husbandry statistics in the same period. The results showed that the mean annual net primary productivity (NPP) of grassland in the Tibetan Plateau was 158.4 g C·m-2·a-1, which had increased significantly in the past 20 years, with a significant increase in 44.7% of the total area. Climate warming, increased precipitation, prolonged growing season, and elevated carbon dioxide concentration were main driving forces for grassland productivity. The mean theoretical livestock carrying capacity estimated based on pasture yield was 1.17 SU·hm-2, with a growth rate of 0.011 SU·hm-2. The situation of overgrazing in the Tibetan Plateau had generally improved since 2000. The proportion of counties with severe overgrazing had dropped to less than 20%. In areas with more severe overgrazing, animal husbandry's maintenance and development mainly relied on supplementation of crop straw. The results could provide a scientific basis for regional agricultural and animal husbandry structural adjustment and environmental protection.

Expression of tumor necrosis factor-alpha-induced protein 8 in pancreas tissues and its correlation with epithelial growth factor receptor levels.

Tumor necrosis factor (TNF)-alpha-induced protein 8 (TNFAIP8 or TIPE) is a recently identified protein considered to be associated with carcinogenesis. To investigate its expression pattern in pancreatic cancer patients and to analyse its correlation with clinicopathological significance and the expression levels of epithelial growth factor receptor (EGFR), immunohistochemistry was performed to detect the TNFAIP8 and EGFR proteins in pancreatic cancers, pancreatitis tissues, and healthy controls. The results showed stronger staining of TNFAIP8 protein in pancreatic cancer tissues compared with normal pancreas tissue. Furthermore, in 56 patients with pancreatic cancer, the expression levels of TNFAIP8 in patients with low tumor stage was higher than that with high tumor stage, and correlated with tumor staging and lymph node metastasis (P<0.05). Furthermore, TNFAIP8 expression positively correlated with EGFR levels (r=0.671135, P<0.05). These results indicate that TNFAIP8 may play important roles in the progression of pancreatic cancer.

Hepcidin destabilizes atherosclerotic plaque via overactivating macrophages after erythrophagocytosis.

OBJECTIVE: To explore a direct and causal relationship between vascular hepcidin and atherosclerotic plaque stability. METHODS AND RESULTS: Accelerated atherosclerotic lesions were established by perivascular collar placement in apolipoprotein E-deficient (ApoE(-/-)) mice. Adenoviral overexpression of hepcidin in the carotid artery during plaque formation enhanced intraplaque macrophage infiltration and suppressed the contents of collagen and vascular smooth muscle cells, whereas hepcidin shRNA treatment exerts opposite effects. The overexpression or knockdown of hepcidin did not affect plaque lipid deposition but increased or decreased oxidized low-density lipoprotein (ox-LDL) levels within intraplaque macrophages. In cultured macrophages, ox-LDL not only increased reactive oxygen species formation, inflammatory cytokine production, and apoptosis but also upregulated hepcidin expression. However, hepcidin did not exaggerate the ox-LDL-induced activation of macrophages until an onset of erythrophagocytosis. Whereas hepcidin was critical for the upregulation of L-ferritin and H-ferritin in both ox-LDL-treated erythrophagocytosed macrophages and atherosclerotic plaques, the adding of iron chelators suppressed the intracellular lipid accumulation, reactive oxygen species formation, inflammatory cytokine expression, and apoptosis in erythrophagocytosed macrophages. CONCLUSIONS: Hepcidin promotes plaque destabilization partly by exaggerating inflammatory cytokine release, intracellular lipid accumulation, oxidative stress, and apoptosis in the macrophages with iron retention.

[Expression and purification of T7 bacteriophage capsid protein P11 and preparation of monoclonal antibody against P11].

AIM: To express capsid tail protein P11 of T7 bacteriophage and produce mouse monoclonal antibody(mAb) against the protein. METHODS: P11 protein was cloned and recombinant P11 protein was expressed as a fusion protein with an N-terminal 6-His tag. The purified protein was used to immunize BALB/c mouse. The specificity of mAb was analyzed by ELISA and Western blot. RESULTS: P11 protein was successfully expressed and purified. SDS-PAGE analysis showed that the molecular weight of the expressed protein was approximately 27 kd. One hybridoma cell (2G11) secreting mAb against P11 was developed. The isotype of the mAb was IgG(2b). ELISA detection showed that titers of mAb was 1:8. 1 x 10(5) in ascites.Western blot analysis proved mAb obtained could react specifically to the recombinant p11 protein. CONCLUSION: Recombinant P11 protein and mAbs were successfully prepared.

Blockade of TRAIL pathway ameliorates HBV-induced hepatocyte apoptosis in an acute hepatitis model.

Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) may play important roles during hepatitis B virus (HBV) infection. In this study, we used a recombinant human soluble death receptor 5 (sDR5) to explore its effect in a mouse model of HBV-induced acute hepatitis. By measuring blood transaminase activity and hepatocyte apoptosis, we found that sDR5 could alleviate liver damage by blocking TRAIL-induced apoptosis of HBV-transfected hepatocytes. sDR5 injection at 16 mg/kg 24h before HBV transfection was the most effective. Additionally, we showed that sDR5 was equally effective in protecting liver injury as the Stronger Neo-Minophagen C (SNMC), a commonly used drug for patients with liver diseases. Thus, sDR5 represents a potential novel therapeutic drug for patients with fulminant hepatitis.

Human endostatin gene transfer, either naked or with liposome, has the same inhibitory effect on growth of mouse liver tumor cells in vivo.

AIM: To explore a safe and efficient strategy of tumor therapy using anti-angiogenetic agents. METHODS: Endostatin gene with a signal sequence of human IgGgamma chain was amplified by PCR and cloned into pVAX1 plasmid which was the only vector authorized by FDA in clinical trial to construct a recombinant plasmid named as pVAX-sEN. The recombinant plasmid was detected with EcoRI/KpnI and DNA sequencing. BALB/c mice bearing hepatocarcinoma cell line H22 were treated with naked pVAX-sEN or liposome-DNA complex in which the dose of DNA and the ratio of DNA and liposome were different from each other. To compare the efficiency of gene transfection, expression of endostatin at the treated tumor site was assayed with ELISA. To investigate the effect of pVAX1-sEN on hepatocellular carcinoma, pVAX-sEN either naked or in liposome-DNA complex was injected into BALB/c mice bearing H22, then the diameter of tumors was measured, microvessel density was detected by immunohistochemistry, endostatin expression in vivo was assayed at different time points. RESULTS: DNA sequencing showed the endostatin gene with the signal peptide was correctly cloned. In situ gene expression assay indicated that both the ratio of DNA and liposome and the dose of DNA could affect the gene transfection efficiency. Interestingly, naked pVAX-sEN had a similar in situ endostatin expression to pVAX-sEN with liposome. Animal experiments showed that pVAX-sEN together with pVAX-sEN-liposome complex could efficiently suppress the growth of mouse hepatoma cells. CONCLUSION: Naked endostatin plasmid intratumoral injection can get a similar gene transfection efficiency to liposome-DNA complex when used in situ.

Establishment of mice model with human viral hepatitis B.

AIM: To establish a mice model of hepatitis B by using HBV-transgenic mice, and to transfer HBV-specific cytotoxic T lymphocytes (CTL) induced from syngeneic BALB/c mice immunized by a eukaryotic expression vector containing HBV complete genome DNA. METHODS: HBV DNA was obtained from digested pBR322-2HBV and ligated with the vector pcDNA3. Recombinant pcDNA3-HBV was identified by restriction endonuclease assay and transfected into human hepatoma cell line HepG2 with lipofectin. ELISA was used to detect the expression of HBsAg in culture supernatant, and RT-PCR to determine the existence of HBV PreS1 mRNA. BALB/c mice were immunized with pcDNA3-HBV or pcDNA3 by intramuscular injection. ELISA was used to detect the expression of HBsAb in serum. MTT assay was used to measure non-specific or specific proliferation ability and specific killing activity of spleen lymphocytes. Lymphocytes from immunized mice were transferred into HBV-transgenic mice (2.5X10(7) per mouse). Forty-eight hours later, the level of serum protein and transaminase was detected with biochemical method, liver and kidney were sectioned and stained by HE to observe the pathological changes. RESULTS: By enzyme digestion with Eco RI, Xho I and Hind III, the recombinant pcDNA3-HBV was verified to contain a single copy of HBV genome, which was inserted in the positive direction. HepG2 cells transfected with the recombinant could stably express PreS1 mRNA and HBsAg. After immunized by pcDNA3-HBV for 4 weeks, HBsAb was detected in the serum of BALB/c mice. The potential of spleen lymphocytes for both non-specific and specific proliferation and the specific killing activity against target cells were enhanced. The transgenic mice in model group had no significant changes in the level of serum protein but had an obvious increase of ALT and AST. The liver had obvious pathological changes, while the kidney had no evident damage. CONCLUSION: A eukaryotic expression vector pcDNA3-HBV containing HBV complete genome is constructed successfully. HepG2 cells transfected with the recombinant can express PreS1 mRNA and HBsAg stably. Specific cellular immune response can be induced in mice immunized by pcDNA3-HBV. A mice model of acute hepatitis with HBV has been established.

Antisense oligonucleotide targeting at the initiator of hTERT arrests growth of hepatoma cells.

AIM: To evaluate the inhibitory effect of antisense phosphorothioate oligonucleotide (asON) complementary to the initiator of human telomerase catalytic subunit (hTERT) on the growth of hepatoma cells. METHODS: The as-hTERT was synthesized by using a DNA synthesizer. HepG2.2.15 cells were treated with as-hTERT at the concentration of 10 micromol/L. After 72 h, these cells were obtained for detecting growth inhibition, telomerase activity using the methods of MTT, TRAP-PCR-ELISA, respectively. BALB/c(nu/nu) mice were injected HepG2.2.15 cells and a human-nude mice model was obtained. There were three groups for anti-tumor activity study. Once tumors were established, these animals in the first group were administered as-hTERT and saline. Apoptosis of tumor cells was detected by FCM. In the 2nd group, the animals were injected HepG2.2.15 cells together with as-hTERT. In the third group, the animals were given as-hTERT 24 hours postinjection of HepG2.2.15 cells. The anti-HBV effects were assayed with ELISA in vitro and in vivo. RESULTS: Growth inhibition was observed in cells treated with as-hTERT in vitro. A significant different in the value of A570-A630 was found between cells treated with as-hTERT and control (P<0.01) by MTT method. The telomerase activity of tumor cells treated with as-hTERT was reduced, the value of A450 nm was 0.42 compared to control (1.49) with TRAP-PCR-ELISA. The peak of apoptosis in tumor cells given as-hTERT was 21.12%, but not seen in saline-treated control. A prolonged period of carcinogenesis was observed in the second and third group animals. There was inhibitory effect on the expression of HBsAg and HBeAg in vivo and in vitro. CONCLUSION: As-hTERT has an anti-tumor activity, which may be useful for gene therapy of tumors.

[Inhibition of HBV DNA replication and expression in 2.2.15 hepatoma cells infected with AFP-mediated HBX antisense RNA].

OBJECTIVE: To study the specific expression of the antisense RNA against hepatitis B virus X (HBX) gene in hepatoblastoma cell line and its anti -HBV activity. METHODS: HBX gene (nt.1370-1827) was amplified by PCR, then cloned into EB virus vector pEBAF which contained human alpha-fetoprotein promoter and enhancer. After transfected into 2.2.15 hepatoma cells and ECV304 human endothelial cells by lipofectin, northern blot, ELISA and real-time qualitative PCR were carried out to assay the expression of HBX mRNA, HBV antigens and HBV DNA level, respectively. RESULTS: The HBX antisense RNA expression vector pEBAF-as-HBX which could be expressed specifically in 2.2.15 hepatoblastoma cells was successfully constructed. Both HBV DNA level and the expressions of hepatitis B virus surface antigen (HBsAg) and e antigen (HBeAg) in 2.2.15 hepatoblastoma cells were inhibited by pEBAF-as-HBX. Compared with those in sense control (pEBAF-s-HBX), the inhibitory rates of HBsAg, HBeAg, and HBV DNA were 37.9%, 36.8%, and 25%, respectively. CONCLUSIONS: The pEBAF-as-HBX expression vector may lead to targeted-expression of HBX antisense RNA in hepatoma cells and shows great inhibition effect on HBV.

A novel HBV antisense RNA gene delivery system targeting hepatocellular carcinoma.

AIM: To construct a novel HBV antisense RNA delivery system targeting hapatocellular carcinoma and study its inhibitory effect in vitro and in vivo. METHODS: GE7,a 16-peptide specific to EGFR, and HA20,a homologue of N-terminus of haemagglutinin of influenza viral envelope protein, were synthesized and conjugated with polylysin. The above conjugates were organized into the pEBAF-as-preS2, a hepatocarcinoma specific HBV antisense expression vector, to construct a novel HBV antisense RNA delivery system, named AFP-enhancing 4-element complex. Hepatocelluar carcinoma HepG2.2.15 cells was used to assay the in vitro inhibition of the complex on HBV. Expression of HBV antigen was assayed by ELISA. BALB/c nude mice bearing HepG2.2.15 cells were injected with AFP-enhancing 4-element complex. The expression of HBV antisense RNA was examined by RT-PCR and the size of tumor in nude mice were measured. RESULTS: The AFP-enhancing 4-element complex was constructed and DNA was completely trapped at the slot with no DNA migration when the ratio of polypeptide to plasmid was 1:1. The expression of HBsAg and HBeAg of HepG2.2.15 cells was greatly decreased after being transfected by AFP-enhancing 4-element complex. The inhibitory rates were 33.4 % and 58.5 % respectively. RT-PCR showed HBV antisense RNA expressed specifically in liver tumor cells of tumor-bearing nude mice. After 4 injections of AFP-enhancing 4-element complex containing 0.2 micro g DNA, the diameter of the tumor was 0.995 cm+/-0.35, which was significantly smaller than that of the control groups(2.215 cm+/-0.25, P<0.05). CONCLUSION: AFP-enhancing 4-element complex could deliver HBV antisense RNA targeting on hepatocarcinoma and inhibit both HBV and liver tumor cells in vitro and in vivo.

Detection of soluble TRAIL in HBV infected patients and its clinical implications.

AIM: To detect the expression of soluble TRAIL (TNF-related apoptosis inducing ligand, TRAIL) in the peripheral blood of HBV infected patients and try to elucidate whether the expression level of sTRAIL have any correlativity with the clinical staging, the expression level of HBV markers and the degree of liver damage. METHODS: 52 cases of HBV infected patients were investigated, including 8 HBV carriers, 30 chronic hepatitis B, 11 cirrhotics and 3 HBV infection related hepatocellular carcinoma. Expression of soluble TRAIL and markers of the hepatitis B were measured by enzyme-linked immunosorbent assay. RESULTS: The expression level of sTRAIL in the peripheral blood of the HBV infected patients was significantly higher than that of healthy controls (1378.35+/-540.23 pg/ml vs 613.75+/-175.80 pg/ml, P<0.001). In the group of chronic hepatitis, the expression level of sTRAIL was coincident with the status of the disease and was significantly correlated with the level of ALT. In the group of cirrhosis and liver cancer, its expression level was significantly higher than that of the healthy persons and HBV carriers, but lower than that of the hepatitis B patients; meanwhile, the expression of sTRAIL did not have any correlativity with the functional indexes of the liver. CONCLUSION: The soluble TRAIL in the HBV infected people may participate in the liver damage. Our results indicated that the expression level of soluble TRAIL may reflect the ravage of liver caused by host immune reaction to a certain degree.