A novel microRNA miR-4433a-3p as a potential diagnostic biomarker for lung adenocarcinoma.

Background: Lung adenocarcinoma is one of the leading causes of cancer-related deaths because of the lack of early specific clinical indicators. MicroRNAs (miRNAs) have become the focus in lung cancer diagnosis. Further studies are required to explore miRNA expression in the serum of lung adenocarcinoma patients and their correlation with therapy and analyse specific messenger RNA targets to improve the specificity and sensitivity of early diagnosis. Methods: The Toray 3D-Gene miRNA array was used to compare the expression levels of various miRNAs in the sera of patients with lung adenocarcinoma and healthy volunteers. Highly expressed miRNAs were selected for further analysis. To verify the screening results, serum and pleural fluid samples were analysed using qRT-PCR. Serum levels of the miRNAs and their correlation with the clinical information of patients with lung adenocarcinoma were analysed. The functions of miRNAs were further analysed using the Kyoto Encyclopedia of Gene and Genomes and Gene Ontology databases. Results: Microarray analysis identified 60 and 50 miRNAs with upregulated and downregulated expressions, respectively, in the serum of patients with lung adenocarcinoma compared to those in healthy individuals. Using qRT-qPCR to detection of miRNAs expression in the serum or pleural effusion of patients with early and advanced lung adenocarcinoma, we found that miR-4433a-3p could be used as a diagnostic marker and therapeutic evaluation indicator for lung adenocarcinoma. Serum of miR-4433a-3p levels significantly correlated with the clinical stage. miR-4433a-3p may be more suitable than other tumour markers for the early diagnosis and evaluation of therapeutic effects in lung adenocarcinoma. miR-4433a-3p may affect tumour growth and metastasis by acting on target genes (PIK3CD, UBE2J2, ICMT, PRDM16 and others) and regulating tumour-related signalling pathways (MAPK signal pathway, Ras signalling pathway and others). Conclusion: miR-4433a-3p may serve as a biomarker for the early diagnosis of lung adenocarcinoma and monitoring of therapeutic effects.

Fatigue Behavior and Fracture Surface Analysis of Corroded High-Strength Bridge Cable Wires.

Bridge cable wires suffer from alternating stress and environmental erosion, leading to premature failure prior to its design life. This paper investigates the fatigue and mechanical behaviors of corroded bridge cable wires with a zinc-aluminum (Zn-Al) alloy coating. Based on the salt spray corrosion test and microstructure analysis, the anti-corrosion resistance and corrosion appearance characteristics of the Zn-Al alloy coating and galvanized coating were investigated. The Zn-Al alloy coating was superior in resistance to corrosion fatigue for the improvement in toughness and the generation of anti-corrosion Zn-Al and Fe-Zn-Al phases. Equations of the accelerated corrosion depth of the steel wires had been regressed to roughly estimate the corrosion life of the Zn-Al alloy coating, which can reach 29.1 years with a thickness of 70 µm. The fatigue and mechanical properties of the bare wires after the salt spray test were further studied based on tensile tests and fatigue tests. The fatigue properties of the bridge cable wire would decrease with the corrosion degree due to the deterioration and embrittlement of materials, where ductility characterized by the elongation rate was the most affected. Fracture surfaces of the wires were captured and analyzed based on a method for recognizing graphical contours. Insufficient fatigue life may occur in the steel wires after corrosion and increase with the degree of corrosion. The pit depth logarithmically weakened the fatigue life of steel wires for the weakening of fatigue toughness and the bearing area. The flat fracture was more common with a single fatigue source, while multiple fatigue sources led to step-like fractures for the generation of multiple dispersed crack propagation regions. Corrosion fatigue was more sensitive to the existence of fatigue sources than the reduction. Multiple initiation sources significantly reduced the fatigue life due to the cracking facilitation of the joint effect of multiple pits. The electrochemical reactions of corrosion can lead to material embrittlement and a reducing effect on the fracture toughness of the steel wires.

PERP May Affect the Prognosis of Lung Adenocarcinoma by Inhibiting Apoptosis.

Background: PERP, a member of the peripheral myelin protein gene family, is a new therapeutic target in cancer. The relationships between PERP and immune cell infiltration in lung cancer have not been studied. Therefore, the role of PERP in the tumour microenvironment (TME) of lung cancer needs to be further explored. Methods: In this study, we explored the association between PERP expression and clinical characteristics by analysing data from the TCGA database. Cox regression and Kaplan‒Meier methods were used to investigate the relationship between the expression of PERP and overall survival in patients with lung adenocarcinoma (LUAD). The relationship between PERP expression and the degree of infiltration of specific immune cell subsets in LUAD was evaluated using the TIMER database and GEPIA. We also performed GO enrichment analysis and KEGG enrichment analysis to reveal genes coexpressed with PERP using the Coexpedia database. Finally, we verified the expression and function of PERP in LUAD tissues and the A549 cell line by RT‒PCR, Western blot, CCK-8, IHC, and wound healing assays. The mouse model was used to study the in vivo effects of PERP. Results: According to our results, PERP expression was significantly higher in LUAD tissues and associated with the clinical characteristics of the disease. Survival was independently associated with PERP in LUAD patients. We further verified that PERP might regulate B-cell infiltration in LUAD to affect the prognosis of LUAD. To identify PERP-related signalling pathways in LUAD, we performed a genome-aggregation analysis (GSEA) between low and high PERP expression datasets. LUAD cells express higher levels of PERP than paracarcinoma cells, and PERP inhibits the proliferation and metastasis of A549 cells through apoptosis. Conclusion: PERP may affect the prognosis of lung adenocarcinoma by inhibiting apoptosis and is associated with immune cell infiltration.

Development and validation of a prospective study to predict the risk of readmission within 365 days of respiratory failure: based on a random survival forest algorithm combined with COX regression modeling.

BACKGROUND: There is a need to develop and validate a widely applicable nomogram for predicting readmission of respiratory failure patients within 365 days. METHODS: We recruited patients with respiratory failure at the First People's Hospital of Yancheng and the People's Hospital of Jiangsu. We used the least absolute shrinkage and selection operator regression to select significant features for multivariate Cox proportional hazard analysis. The Random Survival Forest algorithm was employed to construct a model for the variables that obtained a coefficient of 0 following LASSO regression, and subsequently determine the prediction score. Independent risk factors and the score were used to develop a multivariate COX regression for creating the line graph. We used the Harrell concordance index to quantify the predictive accuracy and the receiver operating characteristic curve to evaluate model performance. Additionally, we used decision curve analysiso assess clinical usefulness. RESULTS: The LASSO regression and multivariate Cox regression were used to screen hemoglobin, diabetes and pneumonia as risk variables combined with Score to develop a column chart model. The C index is 0.927 in the development queue, 0.924 in the internal validation queue, and 0.922 in the external validation queue. At the same time, the predictive model also showed excellent calibration and higher clinical value. CONCLUSIONS: A nomogram predicting readmission of patients with respiratory failure within 365 days based on three independent risk factors and a jointly developed random survival forest algorithm has been developed and validated. This improves the accuracy of predicting patient readmission and provides practical information for individualized treatment decisions.

Experimental Study on the Impact of Using FRP Sheets on the Axial Compressive Performance of Short-Circular Composite Columns.

This paper conducts an experimental study on the axial compressive performance of FRP-steel-concrete composite columns. Nine short columns were produced and evaluated in the study, comprising of three concrete-filled steel tube reference columns and six FRP-steel-concrete composite columns, respectively denoted as "reference columns" and "composite columns". Two categories of failure modes, including shear failure and waist drum, were observed from the experiments. The failure mode may trend toward waist drum from shear failure as more FRP layers were used. The number of FRP layers had a direct effect on the level of compressive strength attained, with a greater number of layers resulting in a greater increase in compressive strength. Moreover, a greater tensile strength and higher elastic modulus of CFRP sheets are more effective at improving the compressive stiffness of the columns. Finally, a four-stage confinement mechanism for FRP-wrapped steel tube concrete composite columns is proposed and discussed, through which the damage mechanisms of the composite structures are more rationally characterized.

An Eco-Friendly Adsorbent Based on Bacterial Cellulose and Vermiculite Composite for Efficient Removal of Methylene Blue and Sulfanilamide.

In this work, a novel composite of bacterial cellulose (BC) and expanded vermiculite (EVMT) composite was used to adsorb dyes and antibiotics. The pure BC and BC/EVMT composite were characterized using SEM, FTIR, XRD, XPS and TGA. The BC/EVMT composite exhibited a microporous structure, providing abundant adsorption sites for target pollutants. The adsorption performance of the BC/EVMT composite was investigated for the removal of methylene blue (MB) and sulfanilamide (SA) from an aqueous solution. The adsorption capacity of BC/ENVMT for MB increased with increasing pH, while the adsorption capacity for SA decreased with increasing pH. The equilibrium data were analyzed using the Langmuir and Freundlich isotherms. As a result, the adsorption of MB and SA by the BC/EVMT composite was found to follow the Langmuir isotherm well, indicating a monolayer adsorption process on a homogeneous surface. The maximum adsorption capacity of the BC/EVMT composite was found to be 92.16 mg/g for MB and 71.53 mg/g for SA, respectively. The adsorption kinetics of both MB and SA on the BC/EVMT composite showed significant characteristics of a pseudo-second-order model. Considering the low cost and high efficiency of BC/EVMT, it is expected to be a promising adsorbent for the removal of dyes and antibiotics from wastewater. Thus, it can serve as a valuable tool in sewage treatment to improve water quality and reduce environmental pollution.

Parameter Study of Interfacial Capacities for FRP-Steel Bonded Joints Based on 3D FE Modeling.

This paper investigated the stress distribution of an adhesive layer for GFRP-steel bonded joints under 22.48 kN tensile loading using a three-dimensional numerical simulation. Firstly, a stress analysis of three paths was conducted, and after comparison, path II (through the middle layer of the bonding layer) was adopted as the analyzing path. Furthermore, a systemically parametric study of the effects of the FRP stiffness (i.e., elastic modulus and thickness), bonding length, adhesive thickness, and adhesive modulus was conducted. For the joints with different FRP elastic moduli, the minimum value of normal peeling stress was calculated as -3.80 MPa by the FRP for 10 GPa, showing a significantly severe stress concentration of FRP for 10 GPa. An analysis of the von Mises stresses proved that the increase in FRP stiffness could reduce the stress concentration of the adhesive layer effectively. The study of the effect of bonding lengths indicated that a more uniform peeling stress distribution could result from the longest bonding size; the largest peeling stress of 6.54 MPa was calculated for a bonding length of 30 mm. Further parameter analysis showed that the stress concentration of the adhesive layer could be influenced by the FRP thickness, bonding thickness, and elastic modulus of the adhesive layer.

Prognosis-related autophagy genes in female lung adenocarcinoma.

ABSTRACT: To screen the prognosis-related autophagy genes of female lung adenocarcinoma by the transcriptome data and clinical data from The Cancer Genome Atlas (TCGA) database.In this study, screen meaningful female lung adenocarcinoma differential genes in TCGA, use univariate Cox proportional regression model to select genes related to prognosis, and establish the best risk model. In this study, Gene Ontology and Kyoto Encyclopedia of Genes and Genomes were applied for carrying out bioinformatics analysis of gene function.The gene expression and clinical data of 264 female lung adenocarcinoma patient samples were downloaded from TCGA. Twelve down-regulated genes: NRG3, DLC1, NLRC4, DAPK2, HSPB8, PPP1R15A, FOS, NRG1, PRKCQ, GRID1, MAP1LC3C, GABARAPL1. Up-regulated 15 genes: PARP1, BNIP3, P4HB, ATIC, IKBKE, ITGB4, VMP1, PTK6, EIF4EBP1, GAPDH, ATG9B, ERO1A, TMEM74, CDKN2A, BIRC5. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes analysis showed that these genes were significantly associated with autophagy and mitochondria (animals). Multifactor Cox analysis of autophagy-related genes showed that ITGA6, ERO1A, FKBP1A, BAK1, CCR2, FADD, EDEM1, ATG10, ATG4A, DLC1, VAMP7, ST13 were identified as independent prognostic indicators. According to the multivariate Cox proportional hazard regression model, there was a significant difference in the survival rate observed between the high-risk group (n = 124) and the low-risk group (n = 126) during the 10-year follow-up (P < .05). Univariate Cox analysis showed that tumor stage, T, M, and N stages, and risk score were all related to the survival rate of female lung adenocarcinoma patients. Multivariate Cox analysis found that autophagy-related risk scores were independent predictors, with an area under curve (AUC) value of 0.842. At last, there is autophagy genes differentially expressed among various clinicopathological parameters: ATG4A, BAK1, CCR2, DLC1, ERO1A, FKBP1A, ITGA6.The risk score can be used as an independent prognostic indicator for female patients with lung adenocarcinoma. The autophagy genes ITGA6, ERO1A, FKBP1A, BAK1, CCR2, FADD, EDEM1, ATG10, ATG4A, DLC1, VAMP7, ST13 were identified as prognostic genes in female lung adenocarcinoma, which may be the targets of treatment in the future.

Effect of the particle size and surface area on Escherichia coli attachment to mineral particles in fresh water.

The objective of this study was to assess the effects of the particle size and specific surface area (SSA) on the attachment of Escherichia coli to sediment particles. To exclude the effect of different sediment mineral compositions, pure minerals were used, and three typical suspended sediment (<62 µm) components, quartz, K-feldspar and calcite, were separated into four groups with different grain size distributions. Equilibrium attachment experiments covering common E. coli concentrations in surface water were conducted for each group. The results show that the finer fractions of each pure mineral had the greatest attachment capacity. Different mineral properties were measured, as well as an author-defined parameter (SSA_a), which was calculated by integrating the particle size distribution and only reflected the microscopic surface areas accessible to E. coli cells (∼1 µm) while excluding the effects of nanoscopic pores (5-10 nm). Pearson correlation and partial correlation analyses suggested that the partition coefficient (Kd) was positively correlated with the clay content (CC) and SSA_a (P < 0.01). Stepwise multiple regression analysis suggested that SSA_a was the dominant factor (P < 0.01) and was a better explanatory variable than CC. Moreover, in addition to SSA_a, the zeta potential and SSA also partially explained the results (P < 0.05).

Blockade of Plasmodium falciparum erythrocyte invasion: New assessment of anti-Plasmodium falciparum reticulocyte-binding protein homolog 5 antibodies.

There is great interest in any new discoveries in malaria vaccine research. Plasmodium falciparum reticulocyte-binding protein homolog 5 (PfRH5) shows promise in this area and may be used together with other merozoite antigens as a potential vaccine. In the present study, a bioinformatics prediction approach was applied to a PfRH5 B-cell epitope, and two B-cell epitope distributions were selected. Antibodies against the two PfRH5 distributions were obtained and the growth activity inhibition was measured. No inhibition of the P. falciparum CY strain was found, but the growth of the P. falciparum 3D7 strain was inhibited by all of the antibodies, in contrast to the results of other studies. It was additionally found that certain quantities of protein led to the inhibition of the parasitic invasion. Equally noteworthy was that the survival time of the group immunized with a portion of PfRH5 was significantly longer than that of the group immunized with the full-length protein, following infection by P. berghei ANKA. The present study produced conflicting results in in vitro and in vivo experiments, although the accuracy of the evaluation may be lessened due to the use of a murine malaria model. The findings of the present study may indicate that PfRH5 may not be suitable in malaria vaccine research.

In vivo and in vitro antimalarial activity of bergenin.

Malaria is a potentially life-threatening protozoal parasitic disease transmitted by female Anopheles mosquitoes. Drug therapy is currently the most widely used method for the control and treatment of this disease. Several plants were found to contain substances possessing antimalarial properties. In this study, we investigated the antimalarial activity of bergenin, a sesquiterpene lactone compound derived from Rodgersia aesculifolia Batal. The results indicated that bergenin effectively inhibited Plasmodium falciparum growth in vitro (IC50, 14.1 µg/ml, with ~100% inhibition at 50 µg/ml), without apparent cytotoxicity to erythrocytes or to mammalian HeLa and HepG2 cells. Bergenin exhibited less cytotoxic activity and the selectivity index (SI) was 887 and 1,355 for HeLa and HepG2 cells, respectively. The administration of bergenin to Plasmodium berghei-infected mice for 6 days significantly inhibited the growth of the parasites. Taken together, these findings provide evidence that bergenin may be a promising novel drug for antimalarial treatment.

Genetic polymorphisms of XRCC1 gene and susceptibility to hepatocellular carcinoma in Chinese population.

Hepatocellular carcinoma (HCC) is a common cancer in the worldwide. Accumulated evidences indicate that genetic polymorphisms of human X-ray repair complementing group 1 gene (XRCC1) are associated with the susceptibility to HCC. This study aims to investigate the potential association between XRCC1 c.482C>T and c.1178G>A genetic polymorphisms and the susceptibility to HCC. A total of 1,069 Chinese Han subjects consisting of 530 HCC patients and 539 cancer-free controls were recruited in this case-control study. The created restriction site-polymerase chain reaction and directly DNA sequencing methods were utilized to analyze the genotyping of XRCC1 genetic polymorphisms. Our data suggested that the XRCC1 c.482C>T and c.1178G>A genetic polymorphisms were statistically associated with the increased risks of HCC [for c.482C>T, TT vs. CC: OR 2.05, 95% CI 1.26-3.32, P = 0.003; T vs. C: OR 1.26, 95% CI 1.04-1.51, P = 0.017; for c.1178G>A, AA vs. GG: OR 2.15, 95% CI 1.26-3.67, P = 0.004; A vs. G: OR 1.33, 95% CI 1.10-1.61, P = 0.004]. The allele-T and genotype-TT of c.482C>T and allele-A and genotype-AA of c.1178G>A genetic polymorphisms may enhance the susceptibility to HCC. Our findings indicate that the studied XRCC1 genetic polymorphisms may influence the risk of HCC in Chinese populations and might be used as molecular markers for assessing the risk of HCC.

Immune responses induced in HHD mice by multiepitope HIV vaccine based on cryptic epitope modification.

CD8+ T cells play an important role in early HIV infection. However, HIV has the capacity to avoid specific CTL responses due to a high rate of mutation under selection pressure. Although the HIV proteins, gag and pol, are relatively conserved, these sequences generate low-affinity MHC-associated epitopes that are poorly immunogenic. Here, we applied an approach that enhanced the immunogenicity of low-affinity HLA-A2.1-binding peptides. The first position with tyrosine (P1Y) substitution enhanced the affinity of HLA-A2.1-associated peptides without altering their antigenic specificity. More importantly, P1Y variants efficiently stimulated in vivo native peptide-specific CTL that also recognized the corresponding naturally processed epitope. The potential to generate CTL against any low-affinity HLA-A2.1-associated peptide provides us with the necessary technique for identification of virus cryptic epitopes for development of peptide-based immunotherapy. Therefore, identification and modification of the cryptic epitopes of gal and pol provides promising candidates for HIV immunotherapy dependent upon efficient presentation by virus cells. Furthermore, this may be a breakthrough that overcomes the obstacle of immune escape caused by high rates of mutation. In this study, bioinformatics analysis was used to predict six low-affinity cryptic HIV gag and pol epitopes presented by HLA-A*0201. A HIV compound multi-CTL epitope gene was constructed comprising the gene encoding the modified cryptic epitope and the HIV p24 antigen, which induced a strong CD8+ T cell immune response regardless of the mutation. This approach represents a novel strategy for the development of safe and effective HIV prophylactic and therapeutic vaccines.

A promising alternative anti-HBV agent: the targeted ribonuclease.

HBV-targeted ribonuclease (TR) is a fusion of HBV core protein (HBVc) and human eosinophil-derived neurotoxin (hEDN). Introduction of TR by transfection or transduction into HepG2.2.15 cells (a cell model of HBV infection) revealed that it significantly reduces serological markers of HBV replication (including HBsAg, HBeAg and HBV DNA) in cell supernatants, suggesting that the targeted ribonuclease inhibits HBV replication. To further our understanding of the molecular mechanism of the anti-HBV effect of TR, we expressed TR in E. coli and found that purified TR possesses RNase activity and targeting activity. Furthermore, the antiviral effect of TR depends both on an enzymatically active hEDN and on the core domain. In or out of HepG2.2.15 cells, TR coassembles with the wild-type capsid protein into particles with internal hEDN domains. Our data suggest an intracellular ribonuclease activation mechanism that, owing to the characteristics of HBV morphogenesis, is highly virus specific. HBV may therefore be particularly vulnerable to the capsid-targeted viral inactivation approach.

[The influences of Tet system on activity and tissue specificity of HBV Cp.].

AIM: Analyze the effect of Tetracycline-controlled system on Hepatitis B virus core promoter (Cp) activity and tissue specificity. METHODS: The 7 Teto sequence was amplified from plasmid pTL-8 using polymerase chain reaction (PCR) and cloned into T vector pMD19-T Simple. After sequencing, the 7 teto sequence was subcloned into upstream of Cp in pGL3-Basic/Cp. In order to observe the effect of Tetracycline-controlled system on Cp activity and tissue specificity, the plasmid pTL-8 which luciferase encoding sequence was excised and pGL3-Basic/7t/Cp were cotransfected into different tissue-derived cell lines including HepG2, Hela, COS-7, MDA-MB-231 and HT-29. The dual luciferase activities were determined by Dual Luciferase Report (DLR) assay system. RESULTS: Have successfully constructed plasmid pGL3-Basic/7t/Cp. The transcriptional activity of Cp was increased significantly after the teto sequence was cloned upstream of Cp. CONCLUSION: The transcriptional activity of Cp is augmented significantly by Tetracycline-controlled system. But the tissue specificity of Cp lost at the same time.

[Biosynthesis, catabolism and related signal regulations of plant chlorophyll].

Plant photosynthesis is determined by chlorophyll (Chl) metabolism, which is an important factor of determining crop yield. The genes involved in Chl biosynthesis, catabolism, and related signal regulations are numerous, and the mutation of any of them may change the pigment level, causing abnormalities in leaf color and even inducing individual death. Spontaneous or artificial mutants are necessary for functional analysis of Chl related genes. At present, Chl mutants are widely used in foundational research and production practice. This article reviews the latest research progress in this field.

[Prediction and modification analysis of low-affinity HLA-A*0201 restricted CTL epitopes derived from HIV-1 pol antigen].

AIM: To screen the possible HLA-A*0201 restricted low-affinity CTL epitopes derived from HIV-1 pol antigen and to predict and identify the possible change of the affinity between epitope and the HLA-A*0201 molecule when the epitope is modified. METHODS: HLA-A*0201 restricted low-affinity CTL epitopes were predicted by CTL epitope prediction software based on super motif, proteasome cleavage probability, HLA affinity and so on. The candidates were modified acid substitution and analyzed by computer. T2 cells were used to determine the peptide by amino binding affinity and HLA-A*0201-peptide complex stability. RESULTS: Among the three predicted peptides by softer ware, YVSLSFPQI (pol52-60Y1), YVSQIIEQL pol(673-681Y1), YIQKETWEA(pol548-556Y1)could bind to HLA-A*0201 with high affinity, and the dissociation time of 50% HLA-A*0201 peptide complex was over to 8 h. CONCLUSION: Our results suggest that the three predicted peptides, as modification, might be HLA-A*0201 restricted CTL epitopes.

Analytical application of poly{methyl[3-(2-hydroxy-3,4-difluoro)phenyl]propyl siloxane} as a QCM coating for DMMP detection.

A new material-poly{methyl[3-(2-hydroxy-3,4-difluoro)phenyl]propyl siloxane}(PMDFPS) sensitive to toxic organophosphate vapor was synthesized with 2,3-difluorophenol, allyl bromide and poly (methyl hydrosiloxane) as raw materials, via O-alkylation, Claisen rearrange reaction and hydrosilylation reaction. This novel material was then coated on a quartz crystal microbalance (QCM) to investigate its gas sensitive properties to the nerve agent simulant dimethyl methylphosphonate (DMMP) vapor, as well as known interfering vapors. When tested with competing vapors, the sensor was more than 10 times sensitive to DMMP than to other interfering vapors. Thus, high selectivity of poly{methyl[3-(2-hydroxy-3,4-difluoro)phenyl]propyl siloxane} to DMMP was demonstrated. The poly{methyl[3-(2-hydroxy-3,4-difluoro)phenyl]propyl siloxane}-QCM sensor responded linearly to DMMP vapor with a slope of 14 Hz/ppm in the 1-50 ppm range with a detection limit of 0.21 ppm (S/N=3).

Adenoviral-vector mediated transfer of HBV-targeted ribonuclease can inhibit HBV replication in vivo.

Hepatitis B virus (HBV)-targeted ribonuclease (HBV-TR) is a fused protein of HBV core protein and a ribonuclease, human eosinophil-derived neurotoxin (hEDN). Our previous results showed that HBV-TR could effectively inhibit HBV replication in vitro. To test whether HBV-TR can inhibit HBV replication in vivo, we constructed a recombinant adenoviral vector expressing HBV-TR (Ad-TR) and used it to treat HBV-transgenic mice. Immunohistochemical staining showed that TR was expressed at varied levels in different tissues of Ad-TR-treated mice. Serum HBsAg concentration was decreased by 64.8% for the Ad-TR-treated mice compared with empty adenoviral vector-treated control mice. The amount of HBV-DNA in the livers of the Ad-TR-treated mice was 0.74 x 10(7) copies/mug of genomic DNA while the amount of HBV-DNA in the livers of the empty adenoviral vector-treated control mice was 2.86 x 10(7) copies/mug of genomic DNA. Serum HBV-DNA of Ad-TR-treated mice was also decreased by 71.4% compared with empty adenoviral vector-treated control mice. In addition, for some Ad-TR-treated mice, the expression of HBsAg in the liver cells turned negative. No discernible adverse effects were observed for Ad-TR-treated mice. Taken together, our results indicated that adenovirus mediated transfer of HBV-TR can inhibit HBV replication in vivo.

[Cloning and expression of PbTIP analogous gene from Plasmodium berghei ANKA and preparation of its polyclonal antibody].

OBJECTIVE: To clone and express a novel protein analogous to TIP (T cell immunomodulatory protein) of Plasmodium berghei ANKA and prepare its polyclonal antibody. METHODS: The PbTIP encoding nucleotide sequence was searched from the Plasmodium berghei genomic database and amplified by PCR. The gene was sub-cloned into prokaryotic expression vector pGEX4T-1 and expressed in E.coli BL21 (DE3). After induction by IPTG, the expression of PbTIP-GST fusion protein was characterized by SDS-PAGE and Western blotting. The inclusion bodies of GST-PbTIP fusion protein were injected into BALB/c mouse. Anti-sera were identified by indirect fluorescent antibody test and western blotting. RESULTS: The PbTIP-GST fusion protein was successfully expressed in the form of inclusion bodies, by controlling the temperature and concentration of IPTG. Anti-PbTIP-GST sera were acquired with high titer. The sera specifically recognized the PbTIP with a band of 60 000 in P.berghei infected erythrocyte protein. CONCLUSION: PbTIP/GST fusion protein and polyclonal antibody have been obtained.