LncRNA-NEF suppressed oxaliplatin resistance and epithelial-mesenchymal transition in colorectal cancer through epigenetically inactivating MEK/ERK signaling.

A major cause of oxaliplatin chemoresistance in colorectal cancer (CRC) is acquired epithelial-mesenchymal transition (EMT) in cancer cells, making the cancer cells easy to metastasis and recurrence. LncRNA Neighboring Enhancer of FOXA2 (lncRNA-NEF) has been characterized as a tumor suppressor to mediate cancer metastasis in multiple cancer types. However, whether it mediated the drug resistance remains unknown. In the present study, an oxaliplatin-resistant CRC cell line (SW620R) was established and lncRNA-NEF was obviously down-regulated in this resistant cell line. The further loss and gain-of-function studies demonstrated that this lncRNA suppressed oxaliplatin resistance as well as EMT programme in vitro and inhibited metastasis in vivo. Mechanistically, lncRNA-NEF epigenetically promoted the expression of DOK1 (Downstream of Tyrosine kinase 1), a negative regulator of MEK/ERK signaling, by disrupting DNA methyltransferases (DNMTs)-mediated DNA methylation. DOK1, in turn, induced the inactivation of MEK/ERK signaling, forming the lncRNA-NEF/DOK1/MEK/ERK regulatory axis to mediate oxaliplatin resistance in CRC. Collectively, our work reveals the critical function of lncRNA-NEF in mediating the oxaliplatin chemotherapy resistance in CRC, and provides a promising therapeutic strategy for CRC patients with oxaliplatin resistance.

Baicalein mediates the anti-tumor activity in Osteosarcoma through lncRNA-NEF driven Wnt/ß-catenin signaling regulatory axis.

Background: Osteosarcoma (OS) is a common type of malignant bone tumor in adolescents with high risk of metastasis. However, the clinical management still remains unsatisfactory. Traditional Chinese medicine (TCM) has been widely considered as an alternative treatment, and their extracts have proved to possess great potential for drug discovery. Baicalein (BA), the active pharmaceutical ingredient of rhizoma coptidis, was proved to have anti-tumor properties in OS, but the mechanism remains poorly understood. Methods: The potential anti-cancer effects on cell growth, cell cycle, apoptosis and migration were examined in OS cells. Moreover, the lncRNA-Neighboring Enhancer of FOXA2 (lncRNA-NEF) and Wnt/ß-catenin signaling were detected by qPCR and Western blotting assays. The in vivo effect of GA on tumor growth was investigated using a xenograft mice model. Results: In the present study, BA was found to significantly suppress tumor growth in vitro and in vivo. And it was also found to inhibit the invasion and metastasis as well. As for the mechanism investigation, lncRNA-NEF was obviously upregulated by BA in OS cells, and thus induced the inactivation of Wnt/ß-catenin signaling. Moreover, lncRNA-NEF knockdown partially reversed the BA-induced anti-cancer activities; and successfully compensated the suppressive effect on Wnt/ß-catenin signaling. We therefore suggested that BA induced the inactivation of Wnt/ß-catenin signaling through promoting lncRNA-NEF expression. Conclusions: In conclude, our results demonstrated that BA suppressed tumor growth and metastasis in vitro and in vivo through an lncRNA-NEF driven Wnt/ß-catenin regulatory axis, in which lncRNA-NEF was upregulated by BA, and thus induced the inactivation of Wnt/ß-catenin signaling. The Translational potential of this article: The findings derived from this study validates the anti-cancer activity of BA in OS and provides a novel underlying mechanism, which suggest that BA may be a potential candidate to develop the effective drug for OS patients.

Genome-wide association study identifies quantitative trait loci affecting cattle temperament.

Cattle temperament is an interesting trait due to its correlation with production efficiency, labor safety, and animal welfare. To date, however, its genetic basis is not clearly understood. Here, we performed a genome-wide association study for a series of temperament traits in cattle, assessed with via open field and novel object tests, using autosomal single nucleotide polymorphisms (SNPs) derived from the whole-genome sequence. We identified 37 and 29 genome-wide significant loci in the open field and novel object tests, respectively. Gene set analysis revealed the most significant pathway was the neuroactive ligand-receptor interaction pathway, which may be essential for emotional control in cattle. Analysis of the expression levels of 18 tissue-specific genes based on transcriptomic data showed enrichment in the brain, with some candidate genes involved in psychiatric and neurodegenerative diseases in humans. Based on principal component analysis, the first principal component explained the largest variance in the open field and novel object test data, and the most significant loci were assigned to SORCS3 and SESTD1, respectively. Our findings should help facilitate cattle breeding for sound temperament by pyramiding favorable alleles to further improve cattle production.

Gallic Acid Suppressed Tumorigenesis by an LncRNA MALAT1-Wnt/ß-Catenin Axis in Hepatocellular Carcinoma.

Gallic acid (3,4,5-trihydroxybenzoic acid; GA), a natural phenolic acid, is abundantly found in numerous natural products. Increasing evidence have demonstrated that GA plays anti-cancer roles in multiple cancers. However, its anti-tumor effects on hepatocellular carcinoma (HCC) and the underlying mechanism remain obscure. In the present study, we found that GA suppressed the in vitro cell viability and metastasis and inhibited the in vivo tumor growth of HCC cells. The underlying mechanism was further to investigate and it was showed that GA suppressed the expression of ß-catenin and led to the functional inactivation of Wnt/ß-catenin signaling. As a kind of significant regulators, the long noncoding RNA molecules (lncRNAs) have attracted widespread attentions for their critical roles in diverse biological process and human diseases. To further identify which lncRNA participated this GA-mediated process, several lncRNAs related to Wnt/ß-catenin signaling were chosen for examination of their expression profiling in the GA-treated HCC cells. Of which, Metastasis-Associated Lung Adenocarcinoma Transcript 1 (MALAT1) was the most promising candidate. And moreover, MALAT1 was significantly down-regulated by GA. Its overexpression partially reversed the GA-induced the inhibitory effects on cell proliferation and metastasis; and successfully abolished the suppressive effect of GA on Wnt/ß-catenin signaling. In conclusion, our results indicated that GA suppressed tumorigenesis in vitro and in vivo by the MALAT1-Wnt/ß-catenin signaling axis, suggesting that GA has great potential to be developed as a chemo-prevention and chemotherapy agent for HCC patients.

Histone methylatic modification mediates the tumor-suppressive activity of curcumol in hepatocellular carcinoma via an Hotair/EZH2 regulatory axis.

ETHNOPHARMACOLOGICAL RELEVANCE: Curcuma kwangsiensis S. G. Lee & C. F. Liang (Guangxi ezhu, in Chinese) has been used as a traditional Chinese medicine (TCM) for approximately 2000 years. Curcumol is one of the major bioactive components of this herb, which has been demonstrated possesses anti-cancer properties, and was recorded in the Chinese Pharmacopoeia 2020 edition. However, most studies mainly focused on the superficial anti-cancer activity, the underlying mechanism remains poorly understood. AIM OF THE STUDY: In the present study, we aimed to investigate the anti-tumor effect of Curcumol on hepatocellular carcinoma (HCC), and elucidate its underlying mechanism from the perspective of epigenetic modification. MATERIALS AND METHODS: The potential anti-cancer properties of Curcumol were evaluated in HepG2 and SMMC-7721 cells. Its effects on cell growth, cell cycle, apoptosis and migration were examined in these HCC cells. Moreover, the lncRNA HOX transcript antisense intergenic RNA (Hotair) and histone methylatic modification were detected by qPCR and Western blotting assays. RESULTS: In the present study, Curcumol was illustrated to suppress cell growth in HCC cells via inducing apoptosis and cell cycle arrest. And it was also found that Curcumol inhibited the invasion and metastasis of HCC as well. As for the mechanism investigation, it was showed that lncRNA Hotair was significantly downregulated by Curcumol in HCC cells. As is well known, Hotair recruited histone methyltransferase enhancer of zeste homolog 2 (EZH2) to exert transcriptional regulation. Our results showed that EZH2 were downregulated by Curcumol in HCC cells, and thus disrupted the trimethylation of H3K9 and H3K27 which were specifically catalyzed by EZH2. CONCLUSIONS: In conclude, our results demonstrated that Curcumol suppressed tumor growth and metastasis via an Hotair/EZH2/histone modification regulatory axis.

Tea Polyphenols Prevent Sepsis-Induced Lung Injury via Promoting Translocation of DJ-1 to Mitochondria.

BACKGROUND: Sepsis is the systemic inflammatory response syndrome caused by infection, which commonly targets on the lung. Tea polyphenols (TP) have many pharmacological activities, but their role in sepsis induced lung injury remains unclear. RESULTS: Injection of TP after cecal ligation and puncture (CLP) operation elevated the survival rate in a concentration dependent manner. TP treatment improved alveoli structure injury under CLP operation. CLP surgery increased the expression of inflammatory factors IL1ß, IL6, and TNFα expression, which was reversed by TP injection. In addition, CLP operation promoted apoptosis and senescence in tissues and cells during lung injury, while TP administration removed the damaged role of CLP on lung tissues and cells. Furthermore, CLP operation or LPS (lipopolysaccharide) treatment induced dysfunction of mitochondria in lung tissues and cells, but TP contributed to recover mitochondria function, which exhibited as inhibition of ROS production inhibition and increase of ATP content and Mitochondrial membrane potential (MMP). Interestingly, DJ-1 was inhibited by CLP operation but promoted by TP treatment. Overexpression of DJ-1 reversed the injury of LPS on L2 cells and recovered mitochondria normal function. And silencing of DJ-1 in rats or alveolar epithelial cells blocked the protection effect of TP. CONCLUSION: Our research revealed that TP protected against lung injury via upregulating of DJ-1 to improve mitochondria function, which contributed to the prevention and treatment of sepsis induced lung injury.

Effect of selective lymph node dissection on immune function in patients with T1 stage non-small cell lung cancer: a randomized controlled trial.

BACKGROUND: Many lymph nodes resected from early-stage non-small cell lung cancer (NSCLC) patients haven't metastasis. Selective lymph node dissection (SLND) can reduce surgical trauma by retaining non-metastatic lymph nodes, we aimed to investigate whether SLND could reduce immune impairment compared with systematic lymph node dissection. METHODS: According to the selection criteria, patients with no metastasis in hilar and regional lymph nodes were selected as the research subjects. The patients were randomly divided into 2 groups: the SLND group (Group SD) and the systematic lymph node dissection group (Group CD). At 24 hours before surgery and on the 1st and 3rd postoperative days (POD), fasting venous blood was sampled to detect cytokine indicators [interleukin-6 (IL-6), C-reactive protein (CRP)], cellular immune indicators [lymphocytes, natural killer cells (NK cells), CD4+, CD8+, CD4+/CD8+], and humoral immune indicators (IgG, IgA, IgM). At the same time, clinically indicators of the patients were recorded. All indicators between the 2 groups were compared. RESULTS: The comparison of clinical indicators between the two groups showed that SLND is more conducive to patients' rapid recovery after surgery. CRP and IL-6 levels in Group CD were significantly higher than those in Group SD after surgery (P<0.05). There were no statistical differences between the 2 groups in the proportions of lymphocytes and NK cells after surgery (P>0.05). The proportions of CD4+ cells and CD4+/CD8+ in Group CD were significantly lower than those in Group SD at POD1 (P<0.05). The proportion of CD8+ cells was significantly higher in Group CD than in Group SD at POD3 (P<0.05). There were no significant differences in IgG, IgA, and IgM levels between the 2 groups at the same point in time (P>0.05). CONCLUSIONS: Compared with systematic lymph node dissection, SLND has the following advantages: (I) it is more conducive to patients' rapid recovery after surgery; (II) it can reduce the body's acute inflammatory response and non-specific immune damage; (III) it can reduce the damage to cellular immune function caused by surgery. TRIAL REGISTRATION: Chinese Clinical Trial Registry ChiCTR2100045893.

H19-Wnt/ß-catenin regulatory axis mediates the suppressive effects of apigenin on tumor growth in hepatocellular carcinoma.

Hepatocellular Carcinoma (HCC) is one of the leading causes of cancer-related deaths in the world. However, the effective pharmacological approaches remain scanty in clinical practice. As a bioactive flavonoid, apigenin (API) is enriched in common fruits and vegetables. Although pharmacological activities of API have been widely investigated, its biological function in HCC remains obscure. In the present study, we found that API strongly suppressed cell growth and induced apoptosis in HCC cells. Using a xenograft mice model, API was demonstrated to inhibit the in vivo tumor growth. It is known that the long non-coding RNA H19, which is frequently elevated in HCC, plays a vital role in mediating tumorigenesis and cancer progression. Our results demonstrated that H19 was down-regulated by API, and thereby induced the inactivation of the canonical Wnt/ß-catenin signaling. In conclusion, our results demonstrated that API was able to suppress tumor growth of HCC through H19-mediated Wnt/ß-catenin signaling regulatory axis, suggesting that API may be a promising candidate for developing novel therapeutic approaches against liver cancer.

LincROR Mediates the Suppressive Effects of Curcumin on Hepatocellular Carcinoma Through Inactivating Wnt/ß-Catenin Signaling.

As one of the leading causes of cancer-related death in the world, hepatocellular carcinoma (HCC) has continued to attract growing attention in recent decades. The use of traditional Chinese herbs in medicine has been practiced for thousands of years, and holds the potential of being a possible treatment for HCC. Curcumin, a bioactive ingredient derived from Curcuma longa, exhibits anti-tumor activity in various cancers. Although the effects of Curcumin on HCC have been elucidated, the underlying mechanism remains unclear. In the present study, Curcumin was demonstrated to inhibit the proliferation of HCC cells via inducing cell cycle arrest and apoptosis. Several previously reported lncRNAs related to tumorigenesis were chosen for examination of their expression profiles, and lincROR was found to be the most down-regulated in the Curcumin-treated HCC cells. Furthermore, Curcumin was found to decrease ß-catenin expression and induce the inactivation of Wnt/ß-catenin signaling. Therefore, Curcumin suppressed tumor growth through a lincROR/ß-catenin regulatory pattern. In conclusion, our results demonstrated that Curcumin suppressed the cell proliferation via the down-regulation of lincROR and inactivation of Wnt/ß-catenin signaling, suggesting that it may be a potential anti-cancer candidate for HCC patients with activated Wnt/ß-catenin signaling.

Base excess in predicting the prognosis of patients with paraquat poisoning: A meta-analysis.

BACKGROUND: Although the prognostic significance of base excess (BE) in patients with paraquat (PQ) poisoning has been investigated for several years, the results remain controversial. Thus, we performed for the first time a comprehensive meta-analysis to explore the value of BE in predicting the prognosis of patients with PQ poisoning. METHODS: We searched PubMed, EMBase, Web of Science, ScienceDirect, Cochrane Library, and the Chinese National Knowledge Infrastructure to identify all relevant papers that were published up to August 2018. The data were extracted for pooled analysis, heterogeneity testing, sensitivity analysis, publication bias analysis, and subgroup analysis. RESULTS: Pooled analysis revealed that a decreased BE is correlated with poor mortality (pooled OR = 21.358, 95% CI: 12.716-35.873, P < .001). Pooled sensitivity, specificity, positive likelihood ratio, negative likelihood ratio, and diagnostic odds ratio were 78% (95% CI: 0.66-0.86), 88% (95% CI: 0.66-0.97), 6.6 (95% CI: 2.2-19.9), 0.25 (95% CI: 0.18-0.36), and 26 (10-69), respectively. No publication bias was detected by Egger test (P = .263) and Begg test (P = .462). Sensitivity analyses indicated no important differences among the estimates of effects. CONCLUSION: Our findings show that BE is useful for predicting the prognosis of PQ poisoning.

[Simulation of AquaCrop model and management practice optimization for dryland maize production under whole plastic-film mulching on double ridges]. / æ—±åœ°å ¨è†œåŒåž„æ²ŸçŽ‰ç±³ç”Ÿäº§çš„AquaCrop模型模拟及管理措施优化.

In order to study the applicability of AquaCrop model for simulating dryland whole plastic-film mulching in double ridges cultivation mode and to find the best agronomic management measures, the data of nitrogen gradient test in 2014 and 2015 were selected to validate the variety and stress parameters in the model. The change trends of yield were simulated under different mana-gement measures. The results showed that the root mean square error (RMSE), normalized root mean square error (NRMSE) and the compliance index (d) of the measured and simulated production for all treatments were 717 kg·hm-2, 10.0% and 0.96, respectively, the RMSE, NRMSE and d of the total biomass were 951 kg·hm-2, 6.5% and 0.98, respectively, which indicated that the cultivation characteristics of the whole plastic-film mulching on double ridges maize in the dryland could be well reflected. The best fitting degree was 270 kg N·hm-2 from dynamic simulation analysis of canopy cover degrees and biomass, and with the increase of N stress, the simulation accuracy gradually declined. The best sowing time of the whole plastic-film mulching on double ridges maize in the middle part of Gansu Province was from late April to early May, the seeding density was 45000-65000 plants·hm-2, the growth period was 130-145 days, and the nitrogen application rate was 240-280 kg·hm-2. The results of this study had a certain reference value for the application of AcquaCrop model in arid region of Gansu, and would contribute to the transformation and popularization of agricultural cultivation techniques.

DNA methylation dynamics of a maternally methylated DMR in the mouse Dlk1-Dio3 domain.

The mouse delta-like homolog 1 and type III iodothyronine deiodinase (Dlk1-Dio3) imprinted domain contains three known paternally methylated differentially methylated regions (DMRs): intergenic DMR (IG-DMR), maternally expressed 3-DMR (Gtl2-DMR), and Dlk1-DMR. Here, we report the first maternally methylated DMR, CpG island 2 (CGI-2), is located approximately 800 bp downstream of miR-1188. CGI-2 is highly methylated in sperm and oocytes, de-methylated in pre-implantation embryos, and differentially re-methylated during post-implantation development. CGI-2, similarly to Gtl2-DMR and Dlk1-DMR, acquires differential methylation prior to embryonic day 7.5 (E7.5). Both H3K4me3 and H3K9me3 histone modifications are enriched at CGI-2. Furthermore, CCCTC-binding factor (CTCF) binds to both alleles of CGI-2 in vivo. These results contribute to the investigation of imprinting regulation in this domain.

Expression analysis of AK003491, an imprinted noncoding RNA, during mouse development.

The Dlk1-Dio3 imprinted domain on mouse chromosome 12qF1 contains three paternally expressed protein-coding genes and multiple maternally expressed long or short noncoding RNA genes. All these imprinted genes are regulated by IG-DMR located between Dlk1 and Meg3/Gtl2. Recently, several novel imprinted noncoding RNAs were identified in the intergenic region of this domain, although the exact number of imprinted genes within the region is unclear. Here, we report that a novel noncoding RNA, AK003491, located between Meg3/Gtl2 and Meg8, is maternally expressed in E15.5 brain, tongue, heart, lung, liver and kidney tissues. In situ hybridization analysis at E15.5 shows AK003491 is predominantly expressed in forebrain, tongue, thymus, somites, lung and liver. Quantitative real-time RT-PCR analysis confirms this expression pattern and detects highest expression in tongue. While the AK003491 expression pattern at postnatal day 1 is similar to E15.5, AK003491 expression at postnatal day 30 is mainly restricted to the brain. These results expand the number of known imprinted long noncoding RNAs in this domain, and contribute to further investigation of their regulatory mechanism and function.

Conservation of genomic imprinting at the NDN, MAGEL2 and MEST loci in pigs.

Imprinted genes have important effects on the regulation of fetal growth, development, and postnatal behavior. However, the study of imprinted genes has been limited in mammalian species other than human and mouse. Therefore, the study of porcine imprinted genes is useful for defining the extent of conservation of genomic imprinting among different species. In this study, the imprinting status of porcine NDN, MAGEL2 and MEST genes was determined by direct sequencing of the cDNAs and detection of single nucleotide polymorphisms (SNPs) identified in individuals from reciprocal crosses between Meishan and Large White pigs for allele discrimination. The analysis was carried out in 13 different tissues (skeletal muscle, fat, pituitary gland, heart, lung, liver, kidney, spleen, stomach, small intestine, uterus, ovary and testis) from 12 two-month-old piglets. Imprinting analysis showed that NDN and MAGEL2 were paternally expressed in all tissues where the genes were expressed as in human and mouse. Interestingly, MEST showed tissue-specific imprinting, being paternally expressed in skeletal muscle, fat, pituitary gland, heart, kidney, lung, stomach and uterus, and maternally expressed in spleen and liver.

[Analysis of several key problems about the two linkage gene mapping of Neurospora crassa in genetics teaching].

Ordered tetrad analysis is important content in the genetics teaching. In particular, the two linkage gene mapping is not only a key point, but also a difficult one. How to explain the content better is a hard nut for the many genetics teachers or editors of the teaching material to crack. Here, based on teaching practice of many years we summarized several key problems, which are difficult to understand by students and frequently neglected by the teachers and the editors of genetics. Furthermore, we deeply analyzed these problems and presented some opinions and suggestions relative to them so as to provide a reference for the teachers of genetics and the editors of teaching materials.

Imprinting and expression analysis of a non-coding RNA gene in the mouse Dlk1-Dio3 domain.

The Dlk1-Dio3 imprinted domain not only is implicated growth and development of the embryo and placenta, but also affects adult metabolism and brain function. In this study, we identified the imprinting status of a mouse non-coding RNA gene, B830012L14Rik, mapped to the Dlk1-Dio3 domain by the polymorphism- and sequencing-based approach. Imprinting analysis showed that the gene was expressed maternally at E15.5, E18.5 and postnatal day 1 mice. Two transcripts of approximately 1.9 and 3.5 kb were detected by northern blot. Furthermore, we examined the spatiotemporal expression patterns of the gene during the mouse development. In situ hybridization analysis showed that B830012L14Rik was mainly expressed in forebrain, pituitary, cartilage primordium of spinal column, lung and liver at E13.5 and E15.5. The results of real-time quantitative RT-PCR showed that the B830012L14Rik expression in brain, heart, lung and liver was higher at E15.5 than at E12.5 and E18.5. Furthermore, the gene expression increased progressively in brain from E12.5 to E15.5 whereas decreased from E15.5 to E19.5. This study may provide further insights into the imprinting, genomic features and expression regulation of the Dlk1-Dio3 imprinted cluster.

Molecular cloning, mRNA expression and imprinting status of PEG3, NAP1L5 and PPP1R9A genes in pig.

Imprinted genes are expressed monoallelically depending on their parental origin, and play important roles in the regulation of fetal growth, development, and postnatal behavior. Most genes known to be imprinted have been identified and studied in the human and the mouse. However, there are only a small number of reported imprinted genes in pigs. Therefore, identification and characterization of more imprinted genes in pigs is useful for comparative analysis of genomic imprinting across species. In this study, we cloned the porcine PEG3, NAP1L5 and PPP1R9A genes. The imprinting status of these genes was determined using sequencing directly and single nucleotide polymorphisms (SNPs) identified in individuals from reciprocal cross of Meishan and Large White pigs. Imprinting analysis was carried out in 13 different tissues (skeletal muscle, fat, pituitary gland, heart, lung, liver, kidney, spleen, stomach, small intestine, uterus, ovary and testis) from twelve 2-month-old piglets. Imprinting analysis showed that PEG3 and NAP1L5 were exclusively expressed from the paternal allele whereas PPP1R9A was biallelically expressed in all tissues tested where the genes were expressed. The study is of interest to understand the conservation of genomic imprinting among mammals at the 3 loci.

[Establishment of hepatitis B virus (HBV) chronic infection mouse model by in vivo transduction with a recombinant adeno-associated virus 8 carrying 1. 3 copies of HBV genome (rAAN8-1. 3HBV)].

In this report, we developed a HBV infection model in C57BL/6 mouse line by in vivo injection of a recombinant adeno-associated virus 8 vector carrying 1. 3 copies of HBV genome (ayw subtype) (rAAV8-1. 3HBV). We firstly prepared and purified the rAAV8-1. 3HBV and then injected it into three C57BL/6 mice with the dose of 2 x 10e11vg, respectively. HBsAg and HBeAg were assayed in sera collected at different time points post injection. Ten weeks post injection, the three mice were sacrificed and blood and liver tissue were taken for assay. Copies of HBV DNA were detected by real time PCR and the way of HBV DNA replication was identified by PCR. Subsequently, detection of HBV antigen by immunohistochemistry and pathology analysis of liver tissue of mice were performed. The results suggested that expression of HBsAg and HBeAg lasted for at least 10 weeks in mice sera. Among mice injected with rAAV8-1. 3HBV, HBsAg levels were showed an 'increasing-decreasing-increasing' pattern (the lowest level at the 4th week post injection), while HBeAg levels were kept high and relatively stable. HBV DNA copies were 4.2 x 10(3), 3.6 x 10(3), 2.5 x 10(3) copies/mL in sera and 8.0 x 10(6), 5.7 x 10(6), 2.6 x 10(6) copies/g in hepatic tissues of three mice, respectively. We found that the linear 1. 3HBV DNA in the rAAV8-1. 3HBV could self form into circular HBV genome and replicate in livers of HBV transfected mice. HBsAg and HBcAg were both positive in liver tissue of mice injected with rAAV8-1. 3HBV and no obvious pathological characters were found in liver of mice injected with rAAV8-1. 3HBV. In conclusion, we successfully developed a HBV chronic infection model in C57BL/6 mouse line by in vivo transduction with the recombinant virus rAAV8-1. 3HBV, in which HBV genes could be continuously expressed and replicated over 10 weeks, and paved a way for further characterization of the human chronic hepatitis B virus infection and evaluation of vaccine and anti-HBV agents.

[The expression analysis of Grb10 during mouse embryonic development].

Grb10 gene expression was analyzed in fetus and four organs during gestation process using the real-time RT-PCR and in situ hybridization. The real-time RT-PCR analysis result showed that the expression of Grb10 in whole embryos is throughout E8.5 to E19.5. The Grb10 gene expression level was gradually increased from E8.5 to E13.5, and then gradually reduced. The tissues specific expression levels were decreased in brain, heart, and lung from E12.5 to E19.5, but a peak expression was observed in liver at E18.5. Grb10 gene expression was also investigated at E13.5, E15.5, E16.5 and E18.5 sections by using in situ hybridization analysis. Strong signals of Grb10 were detected in brain, spine, kidney, and muscle tissues. These results suggest that Grb10 may play an important role during the development process in mouse.

NNAT and DIRAS3 genes are paternally expressed in pigs.

Although expression and epigenetic differences of imprinted genes have been extensively characterised in man and the mouse, little is known on livestock species. In this study, the polymorphism-based approach was used to detect the imprinting status of NNAT and DIRAS3 genes in five heterozygous pigs (based on SNP) of Large White and Meishan F(1) hybrids. The results show that both genes were paternally expressed in all the tested tissues (heart, liver, spleen, lung, kidney, stomach, small intestine, skeletal muscle, fat, uterus, ovary and pituitary). In addition, the NNAT gene had two transcripts in all tested tissues, which is consistent with its counterpart in man and cattle.