An Arabidopsis Retention and Splicing complex regulates root and embryo development through pre-mRNA splicing.

Pre-mRNA splicing is an important step in the posttranscriptional processing of transcripts and a key regulator of development. The heterotrimeric retention and splicing (RES) complex plays vital roles in the growth and development of yeast, zebrafish, and humans by mediating pre-mRNA splicing of multiple genes. However, whether the RES complex is conserved in plants and what specific functions it has remain unknown. In this study, we identified Arabidopsis (Arabidopsis thaliana) BUD13 (AtBUD13), GROWTH, DEVELOPMENT AND SPLICING 1 (GDS1), and DAWDLE (DDL) as the counterparts of the yeast RES complex subunits Bud site selection protein 13 (Bud13), U2 snRNP component Snu17 (Snu17), and Pre-mRNA leakage protein 1, respectively. Moreover, we showed that RES is an ancient complex evolutionarily conserved in eukaryotes. GDS1 directly interacts with both AtBUD13 and DDL in nuclear speckles. The BUD13 domain of AtBUD13 and the RNA recognition motif domain of GDS1 are necessary and sufficient for AtBUD13-GDS1 interaction. Mutants of AtBUD13, GDS1, and DDL failed to properly splice multiple genes involved in cell proliferation and showed defects in early embryogenesis and root development. In addition, we found that GDS1 and DDL interact, respectively, with the U2 small nuclear ribonucleoproteins auxiliary factor AtU2AF65B and the NineTeen Complex-related splicing factor SKIP, which are essential for early steps of spliceosome assembly and recognition of splice sites. Altogether, our work reveals that the Arabidopsis RES complex is important for root and early embryo development by modulating pre-mRNA splicing.

AtBUD13 affects pre-mRNA splicing and is essential for embryo development in Arabidopsis.

Pre-mRNA splicing is an important step for gene expression regulation. Yeast Bud13p (bud-site selection protein 13) regulates the budding pattern and pre-mRNA splicing in yeast cells; however, no Bud13p homologs have been identified in plants. Here, we isolated two mutants that carry T-DNA insertions at the At1g31870 locus and shows early embryo lethality and seed abortion. At1g31870 encodes an Arabidopsis homolog of yeast Bud13p, AtBUD13. Although AtBUD13 homologs are widely distributed in eukaryotic organisms, phylogenetic analysis revealed that their protein domain organization is more complex in multicellular species. AtBUD13 is expressed throughout plant development including embryogenesis and AtBUD13 proteins is localized in the nucleus in Arabidopsis. RNA-seq analysis revealed that AtBUD13 mutation predominantly results in the intron retention, especially for shorter introns (≤100 bases). Within this group of genes, we identified 52 genes involved in embryogenesis, out of which 22 are involved in nucleic acid metabolism. Our results demonstrate that AtBUD13 plays critical roles in early embryo development by effecting pre-mRNA splicing.

Effects of Horseradish Oil and Eight Isothiocyanates Vapour Treatment on Postharvest Disease Control and Their Efficacy as Preservatives of Mature Green Tomato.

This study evaluated the potential of horseradish (Armoracia rusticana) oil (ARO) and eight isothiocyanates (propyl ITC [ProITC], isopropyl ITC [IsoproITC], n-butyl ITC [n-BuITC], 3-butenyl ITC [3-BeITC], phenyl ITC [PhITC], benzyl ITC [BzITC], 2-phenylethyl ITC [PhEITC], and allyl ITC [AITC]) as preservatives and antifungal agents for postharvest tomato disease control. Results showed that ARO and eight ITCs demonstrated antifungal activities against Botrytis cinerea, Alternaria alternata, Rhizopus stolonifer, and Geotrichum candidum, which can cause the decay of mature green tomato during storage. Allyl-ITC (AITC) had the lowest EC50 values of mycelia growth suppression, with 0.18, 0.44, 0.29, and 0.43 µg/ml air for B. cinerea, A. alternata, R. stolonifer, and G. candidum, respectively. ARO, 2-PhEITC, BzITC, and AITC exhibited better efficacy as preservatives of mature green tomato than other ITCs on the basis of some parameters, such as low decay rate, slow reduction in weight loss, slight change in hardness, slow decrease in acidity, and total soluble solid content of treated tomatoes. GC-MS revealed that 2-PhEITC (77.78%) and AITC (15.87%) were the major components of ARO. These results can be used as a basis to develop preservative products composed of ITCs.

AtU2AF65b functions in abscisic acid mediated flowering via regulating the precursor messenger RNA splicing of ABI5 and FLC in Arabidopsis.

In mammalians and yeast, the splicing factor U2AF65/Mud2p functions in precursor messenger RNA (pre-mRNA) processing. Arabidopsis AtU2AF65b encodes a putative U2AF65 but its specific functions in plants are unknown. This paper examines the function of AtU2AF65b as a negative regulator of flowering time in Arabidopsis. We investigated the expression and function of AtU2AF65b in abscisic acid (ABA)-regulated flowering as well as the transcript abundance and pre-mRNA splicing of flowering-related genes in the knock-out mutants of AtU2AF65b. The atu2af65b mutants show early-flowering phenotype under both long-day and short-day conditions. The transcript accumulation of the flowering repressor gene FLOWERING LOCUS C (FLC) is reduced in the shoot apex of atu2af65b, due to both increased intron retention and reduced transcription activation. Reduced transcription of FLC results, at least partially, from the abnormal splicing and reduced transcript abundance of ABSCISIC ACID-INSENSITIVE 5 (ABI5), which encodes an activator of FLC in ABA-regulated flowering signaling. Additionally, the expression of AtU2AF65b is promoted by ABA. Transition to flowering and splicing of FLC and ABI5 in the atu2af65b mutants are compromised during ABA-induced flowering. ABA-responsive AtU2AF65b functions in the pre-mRNA splicing of FLC and ABI5 in shoot apex, whereby AtU2AF65b is involved in ABA-mediated flowering transition in Arabidopsis.

Trichoderma harzianum Improves Defense Against Fusarium oxysporum by Regulating ROS and RNS Metabolism, Redox Balance, and Energy Flow in Cucumber Roots.

Plant survival in the terrestrial ecosystem is influenced by both beneficial and harmful microbes. Trichoderma spp. are a group of filamentous fungi that promote plant growth and resistance to harmful microbes. Previously, we showed that the genus Trichoderma could effectively suppress Fusarium wilt in cucumber. However, the mechanisms that underlie the effects of the genus Trichoderma on plant defense have not been fully substantiated. Two essential metabolic pathways, such as the ascorbate (AsA)-glutathione (GSH) cycle and the oxidative pentose phosphate pathway (OPPP), have been shown to participate in plant tolerance to biotic stressors; nevertheless, the involvement of these pathways in Trichoderma-induced enhanced defense remains elusive. Here, we show that Trichoderma harzianum could alleviate oxidative and nitrostative stress by minimizing reactive oxygen species (ROS; hydrogen peroxide and superoxide) and reactive nitrogen species (nitric oxide [NO]) accumulation, respectively, under Fusarium oxysporum infection in cucumber roots. The genus Trichoderma enhanced antioxidant potential to counterbalance the overproduced ROS and attenuated the transcript and activity of NO synthase and nitrate reductase. The genus Trichoderma also stimulated S-nitrosylated glutathione reductase activity and reduced S-nitrosothiol and S-nitrosylated glutathione content. Furthermore, the genus Trichoderma enhanced AsA and GSH concentrations and activated their biosynthetic enzymes, γ-GCS and l-galactono-1,4-lactone dehydrogenase. Interestingly, the genus Trichoderma alleviated Fusarium-inhibited activity of glucose-6-phosphate dehydrogenase and 6-phosphogluconate dehydrogenase, enzymes involved in the OPPP. Such positive regulation of the key enzymes indicates the adequate maintenance of the AsA-GSH pathway and the OPPP, which potentially contributed to improve redox balance, energy flow, and defense response. Our study advances the current knowledge of Trichoderma-induced enhanced defense against F. oxysporum in cucumber.

Effect of mouse cumulus cells on the in vitro maturation and developmental potential of bovine denuded germinal vesicle oocytes.

We investigated the effect mouse cumulus cells (mCCs) on the in vitro maturation (IVM) and developmental potential of bovine denuded germinal vesicle oocytes (DOs). Cumulus-oocyte complexes (COCs), DOs and DOs cocultured with either mCCs (DOs + mCCs) or bovine cumulus cells (bCCs; DOs + bCCs) were subjected to IVM. The meiosis II (MII) rates of DOs, glutathione (GSH) contents, zona pellucida (ZP) hardening and parthenogenetic blastocyst rates of MII oocytes were determined. The relative expression levels of bone morphogenetic protein 15 (BMP-15) and growth differentiation factor 9 (GDF-9) in MII oocytes were measured using quantitative real-time polymerase chain reaction (PCR). mCCs significantly increased the MII rate of DOs from 53.5 ± 3.58% to 69.67 ± 4.72% (p < 0.05) but had no effect on the GSH content (2.17 ± 0.31 pmol/oocyte with mCCs, 2.14 ± 0.53 pmol/oocyte without mCCs). For the DOs + mCCs group, the BMP-15 and GDF-9 expression levels were significantly higher and the ZP dissolution time was significantly lower (162.49 ± 12.51 s) than that of the DOs group (213.95 ± 18.87 s; p < 0.05). The blastocyst rate of the DOs + mCCs group (32.56 ± 4.94%) was similar to that of the DOs group (31.75 ± 3.65%) but was significantly lower than that of the COCs group (43.52 ± 5.37%; p < 0.05). In conclusion, mCCs increased the MII rate of DOs and expression of certain genes in MII oocytes, and decreased the ZP hardening of MII oocytes, but could not improve their GSH content or developmental potential.

Role of general practitioners in prevention and treatment of hepatitis B in China.

BACKGROUND: Hepatitis B virus (HBV) infection may impose an economic burden to patients or their families. The prevention and control of HBV could effectively reduce the burden. However, the management of HBV-related patients has not been well controlled in China. With the development of general practitioner (GP) system in this country, GPs may greatly improve the management of the patients with HBV infection. However, the role of GPs in controlling HBV infection has been rarely studied. DATA SOURCES: A literature search of PubMed, CNKI, Wanfang data and VIP was performed with the following key words: "general practitioner", "family physician", "community management", "community health care workers", "family practice", "hepatitis B virus", "HBV", "HBV vaccination", "HBV prevention", "HBV management", "HBV treatment", "antiviral therapy" and "chronic hepatitis B (CHB)". The information about the GPs-involved prevention, diagnosis and treatment of CHB was reviewed. RESULTS: The reports on the role of GPs in the prevention, diagnosis and treatment of HBV infection are few. But the experiences from Western countries demonstrated that GPs could play a significant role in the management of patients with CHB. The importance of GPs is obvious although there are some difficulties in China. GPs and health officials at different levels should work together in the management of patients with CHB. CONCLUSIONS: The involvement of GPs in the management of patients with HBV infection is effective in China. But GPs' knowledge and skills for the control of HBV infection have to be improved currently. GPs' involvement will enforce the management of CHB in China in the near future.

Folate deficiency promotes cervical squamous carcinoma SiHa cells progression by targeting miR-375/FZD4/ß-catenin signaling.

Epidemiological studies suggest an association between folate deficiency (FD) and cervical squamous cell carcinoma (SCC) progression. However, the underlying mechanism is unclear. Our study showed that FD-driven downregulation of miR-375 promoted proliferation of SCC SiHa cells and progression of xenograft tumors developed from SiHa; however, the exact mechanism of this process remained unclear. The current study aimed to elucidate the underlying mechanisms by which FD promotes the progression of SiHa cells by downregulating miR-375 expression. The results showed that miR-375 acted as a suppressor of SCC and inhibited the proliferation, migration, and invasion of SiHa cells. The FZD4 gene was identified as a target gene of miR-375, which can reverse the anti-onco effect of miR-375 and promote the proliferation and migration of SiHa cells. Furthermore, the regulatory effects of miR-375 and FZD4 on SiHa cells may be achieved by activating the ß-catenin signaling pathway. Moreover, FD may regulate the expression of miR-375 by regulating its DNA methylation level in the promoter region. In conclusion, our study reveals that FD regulates the miR-375/FZD4 axis by increasing the methylation of the miR-375 promoter, thereby activating ß-catenin signaling to promote SiHa cells progression. This study may provide new insights into the role of folic acid in the prevention and treatment of SCC.

Modeling long-term persistence after 8 years of hepatitis B booster vaccination in 5- to 15-year-old children.

Vaccination remains the most effective and cost-saving measure to protect against hepatitis B, a global health problem. It is crucial to characterize the persistence of the immune response after booster vaccination. This study aimed to quantify the persistence through mathematical modeling. Booster vaccination against hepatitis B was conducted in children 5-15 years in 2009-10 in Zhejiang Province. There were four dosage formulations of hepatitis B vaccines [Shenzhenkangtai Biotechnology Co. Ltd. Dalianhanxin Biotechnology Co. Ltd. NCPC GeneTech Biotechnology Pharmaceutical Co. Ltd. Sinovac Biotech Co. LTD. China]: 5, 10, and 20 µg hepatitis B vaccines or 5 µg hepatitis A and B (HAB) combination vaccine with a 0-1-6-month schedule. These were randomly administered to children negative for all hepatitis B markers, named as the schedule 2 group. Anti-HBs positive subjects were given one dose of booster, named as the schedule 1 group. Anti-HBs antibody was measured 1, 7, 18, 66, and 102 months after the first booster dose. A linear mixed-effects model was proposed to predict long-term persistence. One hundred two months after the booster dose, the mean anti-HBs levels were 33.8 mIU/mL, with 73.7 mIU/mL for the schedule 1 group and 20.2 mIU/mL for the schedule 2 group. The model predicted that 99.5% of subjects would remain seropositive (≥10mIU/mL) at year 20 post booster vaccination, with 100.0% and 98.8% for the schedule 1 group and the schedule 2 group, respectively, whereas at year 30, the seropositivity rates would decrease to 76.8%, with 99.4% for the schedule 1 group and 62.5% for the schedule 2 group. The immunogenicity of the booster vaccination could persist for at least 8 years. Mathematical modeling may predict even longer, up to 30 years of protection.

Plant-Specific AtS40.4 Acts as a Negative Regulator in Abscisic Acid Signaling During Seed Germination and Seedling Growth in Arabidopsis.

Abscisic acid (ABA) is an important phytohormone regulating plant growth, development and stress responses. A multitude of key factors implicated in ABA signaling have been identified; however, the regulation network of these factors needs for further information. AtS40.4, a plant-specific DUF584 domain-containing protein, was identified previously as a senescence regulator in Arabidopsis. In this study, our finding showed that AtS40.4 was negatively involved in ABA signaling during seed germination and early seedling growth. AtS40.4 was highly expressed in seeds and seedlings, and the expression level was promoted by ABA. AtS40.4 was localized both in the nucleus and the cytoplasm. Moreover, the subcellular localization pattern of AtS40.4 was affected by ABA. The knockdown mutants of AtS40.4 exhibited an increased sensitivity to ABA, whereas the overexpression of AtS40.4 decreased the ABA response during seed germination and seedling growth of Arabidopsis. Furthermore, AtS40.4 was involved in ABRE-dependent ABA signaling and influenced the expression levels of ABA INSENTIVE (ABI)1-5 and SnRK2.6. Further genetic evidence demonstrated that AtS40.4 functioned upstream of ABI4. These findings support the notion that AtS40.4 is a novel negative regulator of the ABA response network during seed germination and early seedling growth.

A novel zinc finger protein Zfp637 behaves as a repressive regulator in myogenic cellular differentiation.

Zinc finger proteins have been implicated as transcription factors in the differentiation and development of cells and tissues in higher organisms. The classical C2H2 zinc finger motif is one main type of motif of zinc finger proteins. Our previous studies have shown that Zfp637, which comprises six consecutively typical and one atypical C2H2 zinc finger motifs, is highly expressed in undifferentiated or poorly differentiated cell lines, but is moderately or slightly expressed in normal tissues. We have also demonstrated that Zfp637 can promote cell proliferation. However, its role in the regulation of cell differentiation remains unknown. We report here that endogenous Zfp637 as well as mTERT is expressed in proliferating C2C12 myoblasts and that their expression is downregulated during myogenic differentiation. Constitutive expression of Zfp637 in C2C12 myoblasts increased mTERT expression and telomerase activity, and promoted the progression of the cell cycle and cell proliferation. By contrast, endogenous repression of Zfp637 expression by RNA interference downregulated the mTERT gene and the activity of telomerase, and markedly reduced cell proliferation. Overexpression of Zfp637 also inhibited the expression of myogenic differentiation-specific genes such as MyoD and myogenin, and prevented C2C12 myoblast differentiation. Our results suggest that Zfp637 inhibits muscle differentiation through a defect in the cell cycle exit by potentially regulating mTERT expression in C2C12 myoblasts. This may provide a new research line for studying muscle differentiation.

Screening of the Repellent Activity of 12 Essential Oils Against Adult German Cockroach (Dictyoptera: Blattellidae): Preparation of a Sustained Release Repellent Agent of Binary Oil-γ-CD and its Repellency in a Small Container.

Cockroaches are important sanitary pests and very difficult to control worldwide. With public concern about traditional insecticides, cockroach control agents should be environmentally friendly, highly efficient, and economical. In this article, 12 essential oils were screened to test their repellent effect against Blattella germanica. To develop essential oils as repellent agents, the oils were further examined in binary synergistic combinations. Ilex chinensis Sims (Sapindales: Aquifoliaceae) oil, Lavandula spp (Tubiflorae: Labiatae) oil, and Elsholtzia ciliata (Thunb.) Hyland (Tubiflorae: Labiatae) oil showed excellent repellent activities with lower RD50 (repellency dose for 50% of treated adults) values of 218.634, 154.590, and 223.989 µg/cm2, respectively, compared to those of other oils and the positive control. The I. chinensis oil and E. ciliata oil (weight ratio of 1:1.41) combination also displayed a remarkable synergistic effect against B. germanica. Their cotoxicity coefficient was 214.4. The major chemical constituents in E. ciliata and I. chinensis oils were respectively 3,7-dimethyl-1, 6-octadien-3-ol and methyl salicylate. The binary oil mixtures were formulated as a sustained release agent with γ-CD. The optimal preparation should be an 8:1 ratio of γ-CD to oils, with a 1 h stirring time, 50°C stirring temperature, and 1:12 ratio of γ-CD to ddH2O. The results of this study suggest that sustained release of binary oil-γ-CD exhibited a prolonged repellent activity (10 h) against B. germanica. This sustained-release agent could be further investigated and developed as a novel repellent preparation.

The Cannabinoid WIN 55,212-2 Reduces Delayed Neurologic Sequelae After Carbon Monoxide Poisoning by Promoting Microglial M2 Polarization Through ST2 Signaling.

Delayed neurologic sequelae (DNS) are among the most serious complications of carbon monoxide (CO) poisoning caused partly by elevated neuroinflammation. WIN 55,212-2, a non-selective agonist of cannabinoid receptors, has been demonstrated to have anti-inflammatory properties in various brain disorders. The anti-inflammatory action of WIN 55,212-2 is potentially associated with driving microglial M2 polarization. ST2 signaling is important in regulating inflammatory responses and microglial polarization. Therefore, we aimed to investigate the neuroprotective effect of WIN 55,212-2 on DNS after CO poisoning and elucidate its relationship with ST2-mediated microglial M2 polarization. The behavioral tests showed that treatment with WIN 55,212-2 significantly ameliorates the cognitive impairment induced by CO poisoning. This behavioral improvement was accompanied by reduced neuron loss, decreased production of pro-inflammatory cytokines, and a limited number of microglia in the hippocampus. Moreover, WIN 55,212-2 elevated the protein expression of IL-33 (the ligand of ST2) and ST2, increased the ratio of CD206-positive (M2 phenotype) and ST2-positive microglia, and augmented production of M2 microglia-associated cytokines in the hippocampus of CO-exposed rats. Furthermore, we observed that the WIN 55,212-2-mediated increases in ST2 protein expression, CD206-positive and ST2-positive microglia, and microglia-associated cytokines were blocked by the cannabinoid receptor 2 (CB2R) antagonist AM630 but not by the cannabinoid receptor 1 (CB1R) antagonist AM251. In contrast, the WIN 55,212-2-induced upregulation of the IL-33 protein expression was inhibited by AM251 but not by AM630. Altogether, these findings reveal cannabinoid receptors as promising therapeutic agents for CO poisoning and identify ST2 signaling-related microglial M2 polarization as a new mechanism of cannabinoid-induced neuroprotection.

Disease burden of inflammatory bowel disease in China from 1990 to 2017: Findings from the global burden of diseases 2017.

BACKGROUND: A comprehensive evaluation of the burden of inflammation bowel disease (IBD) is important for identifying potential strategies to control the disease. We present results from the Global Burden of Diseases (GBD) 2017 of IBD at the national level, the trends in disease burden and its epidemiological features in China. METHODS: Using the methods and results from GBD 2017, we describe the IBD burden based on the prevalence, incidence, mortality, years of lost (YLLs), the years of life lived with disability (YLDs) and disability-adjusted life years (DALYs) in China estimated using DisMod-MR 2·1. We additionally evaluated the rate of DALYs at national locations in 2017. FINDINGS: From 1990 to 2017, the cases, deaths, YLLs, YLDs and DALYs for IBD in China were from 1,047,991 to 2,665,081, from 5701 to 5198, from 188,814 to 107,373, from 157,581 to 394,887, from 346,396 to 502,260, respectively. Increasing trends were observed in prevalence (APC: 2·9%), incidence (APC: 1·1%), DALYs (APC: 0·8%) and YLDs (APC:2·9%). There were decreasing trends in mortality (APC: -1·0%) and YLLs (APC: -2·7%). As to the age-standardized rates of DALYs, it observed a decreasing trend (APC: -0·78%). Similar trends were observed in men and women. The age-standardized APCs in incidence, mortality and rate of YLLs among women were higher than those among men. The age-standardized rate of DALYs was 27·51 per 100,000 in 2017. INTERPRETATION: Between 1990 and 2017, China experienced a decrease in the age-standardized DALYs, mortality rates and YLLs due to IBD, despite an increase in the age-standardized rate of prevalence, incidence and YLDs. China is still one of the low endemic areas. FUNDING: This work had no supporting funding.

MicroRNA-144 represses gliomas progression and elevates susceptibility to Temozolomide by targeting CAV2 and FGF7.

Malignant gliomas are the most common tumor in central nervous system with poor prognosis. Due to the limitation of histological classification in earlier diagnosis and individualized medicine, it is necessary to combine the molecular signatures and the pathological characteristics of gliomas. Lots of microRNAs presented abnormal expression in gliomas and modulated gliomas development. Exploration the miRNAs profile is helpful for the diagnosis, therapy and prognosis of gliomas. It has been demonstrated that miR-144 plays important roles in solid tumors. However, the detail mechanisms remained unrevealed. In this study, we have demonstrated the level of miR-144 decreased in glioma tissues from patients, especially in gliomas with higher grades. MiR-144 was also validated have lower expression in glioma cell lines compared with cortical neuron cell by using qRT-PCR. The in vitro functional experiment indicated miR-144 improved gliomas progression through repressing proliferation, sensitizing to chemotherapeutics and inhibiting metastasis. We further identified fibroblast growth factor 7 (FGF7) and Caveolin 2 (CAV2) were target genes of miR-144 by luciferase reporter assay and western blotting. The mechanisms study suggested forced FGF7 expression elevated Akt activation and decreased reactive oxygen species (ROS) generation. The MTT and cell cycle assay indicated miR-144 suppressed glioma cells proliferation through modulating FGF mediated Akt signaling pathway. Meanwhile, miR-144 promoted Temozolomide (TMZ) induced apoptosis in glioma cells via increasing ROS production by using FACS. On the other hand, CAV2, as another target of miR-144, accelerated glioma cells migration and invasion via promoting glioma cells EMT progress. Retrieved expression of FGF7 or CAV2 rescued the proliferation and migration function mediated by miR-144. Furthermore, the in vivo experiments in PDX models displayed the anti-tumor function of miR-144, which could be retrieved by overexpression of FGF7 and CAV2. Taken together, these findings indicated miR-144 acted as a potential target against gliomas progression and uncovered a novel regulatory mechanism, which may provide a new therapeutic strategy and prognostic indicator for gliomas.

Strategies for hepatitis B booster vaccination among children: an 8-year prospective cohort study.

Debate continues regarding the need for a booster vaccination in children who received a universal infant hepatitis B virus (HBV) vaccination. The aim was to explore the need and the strategies for the booster HBV vaccination. 8-year prospective cohort study was conducted among children aged 5-15 years in 2009-2010 in Zhejiang Province. The participants were divided into groups A (<0.1 mIU/mL), B (0.1 to < 1 mIU/mL) and C (1 to <10 mIU/mL) according to the pre-booster anti-HBs antibody levels. 5 µg (group I), 10 µg (group II), 20 µg hepatitis B vaccines (group III) or 5 µg hepatitis A and B (HAB) vaccines (group IV) with 0-1-6-month schedule were randomly administered to children negative for all markers. Blood samples were collected at baseline HBV marker testing, 1 month after the first dose, 1 month, 1 year, 5 years and 8 years after the third dose. Among 4170 children, 2326 (55.8%) were negative for all HBV markers. Group II showed the highest seropositive rates of 92.8%, 99.7%, 97.6%, 90.3% and 83.4% with GMTs of 4194.5 mIU/ml, 4163.9 mIU/ml, 466.9 mIU/ml, 190.6 mIU/ml, 122.6 mIU/ml from 1 month after dose 1 to 8 years after dose 3, respectively (P < .01). Participants in group C showed seropositive rates of 98.9%, 99.9%, 99.5%, 95.5%, 92.8% after the revaccination with GMTs of 6519.6 mIU/ml, 5267.4 mIU/ml, 547.1 mIU/ml, 249.5 mIU/ml, 155.3 mIU/ml, respectively, higher than group A and B (P < .001), except 1 month after the third dose. The 10 µg of HBV vaccine with a 0-1-6-month booster regimen may elicit robust responses and persist for 8 years or longer. Additionally, 1-dose revaccination maybe suitable for children with 1 to < 10 mIU/ml anti-HBs titers.

[Construction of eukaryotic expression plasmid for a novel zinc finger protein gene Zfp637 and its effect on breast carcinoma cell EMT6].

OBJECTIVE: To construct the eukaryotic expression plasmid and establish stably transfected cell line, which contained the code gene of zinc finger protein637 (Zfp637), and to observe the effect of Zfp637 gene to the proliferation of tumor cells. METHODS: The Zfp637 DNA was amplified from the template of normal spleen tissue cDNA and cloned into the eukaryotic expression vector pEGFP-C3. The recombinant plasmid, named as pEGFP-Zfp637, was determined by restriction enzyme and sequencing analyses. Next the pEGFP-Zfp637 recombinant plasmid was transferred into mouse breast carcinoma EMT6 cells with lipofectamine 2000, and the stably transfected cells were selected by G418 (named Zfp637-EMT6). The growth condition of cells was observed, and subcellular localization of Zfp637 gene was located by fluorescence microscope at the same time. The Zfp637 mRNA expression in the transfected cells was detected by RT-PCR, and the proliferation of such cells was measured by MTT. RESULTS: The analysis confirmed that the recombinant pEGFP-Zfp637 contained the Zfp637 full-length cDNA. The Zfp637 mRNA was over-expressed stably in Zfp637-EMT6. The growth of Zfp637-EMT6 was increased obviously when compared with the negative control group and blank group. CONCLUSION: The recombinant pEGFP-Zfp637 has been constructed successfully, and the expression of the Zfp637 gene can promote the proliferation of cells. This recombinant eukaryotic expression plasmid can be used in advanced studies in the biological effects of Zfp637 and anti-tumor gene therapy.

Suitable hepatitis B vaccine for adult immunization in China: a systematic review and meta-analysis.

Hepatitis B virus (HBV) infection remains an important public health problem in China, and adults need to be vaccinated. This systematic review and meta-analysis assessed the appropriate immunization of adults in China. Only randomized controlled trials (RCTs) were eligible, and seroprotection was defined as anti-HBs≥ 10 mIU/ml; 18,308 participants in 27 studies were included. Relative risk (RR) and random effects models were used. Twenty micrograms of HBV vaccine resulted in a better response than 10 µg (RR: 1.05, 95% confidence interval (CI): 1.02 to 1.08), and the 0-, 1-, and 6-month schedule was more effective than the 0-, 1-, and 2 - or 3-month schedule (RR: 0.98, 95% CI: 0.96 to 1.00). No significant differences were observed between 10 µg and 5 µg (RR: 1.05, 95% CI: 0.88 to 1.01); (yeast-derived hepatitis B vaccines) YDV and recombinant Chinese hamster ovary cell (CHO) hepatitis B vaccine (RR: 1.01, 95% CI: 0.98 to 1.04); domestic and imported (RR: 1.02, 95% CI: 0.99 to 1.05); or 0-, 1-, and 6-month and 0-, 1-, and 12-month schedules (RR: 1.02, 95% CI: 0.89 to 1.08). In conclusion, 20 µg of vaccine is recommended for adults in China, and the 0-, 1-, and 12-month immunization program schedule is also worth choosing when it is not possible to complete the 0-, 1-, and 6-month schedule.

Partial biological characterization of cancer stem-like cell line (WJ(2)) of human glioblastoma multiforme.

To provide suitable models for human GBM cancer stem cells in vitro and in vivo, and investigate their biological characteristics, a new human GBM cancer stem-like cell line, WJ2, was established in this experiment through serial passages from adherent monolayer culture to nonadherent tumor sphere culture in turns; Its partial biological characteristics were studied through cell proliferation and tumor sphere assay; cell cycle distribution, side population, and CD133 phenotype were analyzed with FCM. The expressions of CD133, Nestin, and GFAP of cancer stem-like cells and xenograft tumor cells were detected with RT-PCR and immunohistochemistry. Biological characterization, side population, CD133 phenotype and CD133 Nestin, BCRP-1, Wnt-1 gene expression revealed the stemness of this cancer stem-like cell line. Tumorigenicity heterotransplanted in nude mice; histopathological characteristics of xenograft tumor, and expressions of CD133, Nestin, and GFAP of xenograft tumor cells indicated that xenograft tumors recapitulated the phenotype and biological characterization of human primary GBM. All findings of this experimental study suggested that WJ(2) cancer stem-like cell line could accurately mimic human GBM cancer stem cell in vitro and in vivo; it would be useful in the cellular and molecular studies as well as in testing novel therapies of CSC-based anti-cancer therapies for human GBM.

[Construction of eukaryotic expression plasmid pEGFP-ATP1B1 and its effect on gastric adenocarcinoma cell SGC-7901].

OBJECTIVE: To establish the eukaryotic expression plasmid containing the code gene of Na+, K+-ATPase beta1-subunit (ATP1B1) and the basis of ATP1B1 applied to antitumor gene therapy. METHODS: The ATP1B1 cDNA was amplified from leukocyte gene library and then cloned into the eukaryotic expression vector pEGFP-C3. The recombinant plasmid, named as pEGFP-ATP1B1, was determined with restriction enzyme and sequencing analyses. Next pEGFP-ATP1B1 was transferred into gastric adenocarcinoma SGC-7901 cells by lipofectamine, then ATP1B1 mRNA expression in transfected cells was detected by real-time PCR, and also ATPase was detected after cell transfection, as well as the proliferation of such cells was measured by MTT. RESULTS: The analysis confirmed that the recombinant pEGFP-ATP1B1 contained the ATP1B1 cDNA. After cell transfection, the expression of ATP1B1 mRNA(129.2%) and the activity of ATPase [(2.95+/-0.210)%] were higher, and the growth of the SGC-7901 cells transfected with ATP1B1 was inhibited obviously when compared with the control group. CONCLUSION: The recombinant pEGFP-ATP1B1 is constructed successfully, and this recombinant eukaryotic expression vector could be used in additional studies on the biological effect of ATP1B1 and its use in anti-tumor gene therapy.