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Functional implications of genetic variation in human mi RNA seed regions

Functional implications of genetic variation in human mi RNA...

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中华医学会第十五次全国医学遗传学学术会议暨中国医师协会医学遗传医师分会第一届全国学术会议暨2016年浙江省医学遗传学年会

作者： Zhang Jian Hou Yu Zhao Cunyou Department of Medical Genetics School of Basic Medical Sciences Southern Medical University

Objective Micro RNAs(mi RNAs) are a class of non-coding RNAs important in posttranscriptional repressors involved in the regulation of almost every biological process. Mi RNA-mediated regulati on depends on perfect matching between the seven nucleotides of its seed region and the target sequence usually located at the 3‘ untranslated region of the regulated gene. Hence, even single changes in seed regions may affect mi RNA binding capacity and specificity. Previous studies hav e proven that genetic variations in mi RNAs, especially those in the seed regions, have profound a nd broad impacts on gene expression, phenotypic variation and disease susceptibility in human. Method For the present study, we employed a comparative bioinformatic analysis to explore the f unctional significance of sequence variation within the seed region of human mi RNAs. Result A total of 530 SNPs were mapped to the mi RNA seeds(including the known SNPs in the seed region of mi R-1304, mi R-96 and mi R-184), which are located at position 2-8 of the mature mi RNA sequences. We found that the central positions of the mi RNA seeds(positions 4, 5 and 6) contain fewer genetic variants than the peripheral positions in the seeds(2, 3, 7 and 8), indicated more evolutionary conserved and greater importance of middle seed base positions 4 to 6 in mi R NA function. Analysis of the predicted targets for reference and derived mi RNA alleles indicates that a substitution of even a single nucleotide within the seed region changes the spectrum of m R NA targets by >50%. Additionally, except for most of the genetic variations in mi RNA seed region are involved in human diseases, some SNPs are located in non-coding human adaptive accelerat ed regions after we checked for the diseases association and adaptive evolution of all SNPs in mi RNA seed region. Conclusion We suggest that functional changes of genetic variations in mi RNA seed region ma y prove very useful for understanding the disease pathogenicity an

关键词： RNA Functional implications of genetic variation in human mi RNA seed regions

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Genetic variations at BCL11A, HBS1L-MYB and HBG2 associatied with fetal hemoglobin (HbF) in South China

Genetic variations at BCL11A, HBS1L-MYB and HBG2 associatied...

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中华医学会第十五次全国医学遗传学学术会议暨中国医师协会医学遗传医师分会第一届全国学术会议暨2016年浙江省医学遗传学年会

作者： Ma Jing Xiong Likuan Ma Duan Key Laboratory of Metabolism and Molecular Medicine Ministry of Education Department of Biochemistry and Molecular Biology Collaborative Innovation Center of Genetics and Development Institutes of Biomedical Sciences School of Basic Medical Sciences Fudan University Center Laboratory Bao‘an Maternal and Children Healthcare Hospital Key Laboratory of Birth Defects Research

Objective β-thalassemia is one of the most common monogenic diseases with phenotype hetero geneity in many populations worldwide, such as in south China. Persistence of fetal hemoglobin(Hb F) is a major contributor to ameliorating severity and modify the clinical course of β-thalassem ia. Adult level Hb F is influenced by genetic variation at loci of BCL11 A and HBS1 L-MYB intergenic region and HBG2. Method For this reason, we investigated the relationship between Hb F levels and the relevant ge netic loci in 476 patients with β-thalassemia from Guangdong province in China. 33 single-nucleotide polymorphism(SNP) analysis of these 3 principle genomic regions was performed following the previous genome-wide association study(GWAS) in different populations using improved multiple ligase detection reaction(im LDR). The data of blood cell parameters, Hb F level relative quantification and β-thalassemia genotype were collected and recorded. Statistical analysis was made with SPSS software(p<0.05, 95%CI of O R>1). Result In our study, we found elevated Hb F level improved the hematological indices of red cell c ount(RBC), hematocrit(HCT) and mean corpuscular volume(MCV) and was more common in th e patients with the mutation types of 17 M/N,-28 M/N, 43 M/N. There were 26 SNPs correlated sign ificantly with Hb F *** addition, 2 haplotypes of HBS1 L-MYB and 1 haplotype of BCL11 A wer e identified and showed association with Hb F production. Conclusion The BCL11 A locus, HBS1 L-MYB intergenic region, and HBG2 locus were associate d with Hb F levels in Guangdong population of South China.

关键词： in South China Genetic variations at BCL11A HBS1L-MYB and HBG2 associatied with fetal hemoglobin HbF MYB

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Two Novel Mutations In PPIB Gene cause a Rare Pedigree With Osteogenesis Imperfecta Type Ⅸ

Two Novel Mutations In PPIB Gene cause a Rare Pedigree With ...

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中华医学会第十五次全国医学遗传学学术会议暨中国医师协会医学遗传医师分会第一届全国学术会议暨2016年浙江省医学遗传学年会

作者： JIANG Yu GUO Dongwei XIE Jie ZHANG Wei GUO Yibin Zhongshan School of Medicine Sun Yat-sen University Medical College Xiamen University Baylor College of Medicine One Baylor Plaza

Objective The aim of this research is to discover the pathogenic variations in an unknown type of genetic bone disease pedigree in which the healthy parents have consecutive two fetuses with s hort limb deformities so as to make an early genetic diagnosis for the family. Method Next-Generation Sequencing(NGS) was used to screen the whole exome of the parents. Other methods including the screening of variation frequency, evolutionary conservation compari son, website pathogenicity evaluation, and protein structure prediction were used to assess the pathogenicity of the discovered mutations. RT-q PCR was used to analyze the expression level of PPIB gene. Result All mutant genes screened out by NGS are excluded except PPIB gene. Two novel hetero zygous mutations(c.25 A>G from the father and c.509 G>A from the mother) in PPIB gene are ide ntified to relate to osteogenesis imperfecta(OI) IX type. Bioinformatics prediction suggests that both mutations are likely to be pathogenic. The results of RT-q PCR show that the expression of PPIB gene in the two carriers are down-regulated. ize:10.5 pt;font-family:"Times New Roman","serif";mso-fareast-font-family:"Adobe 宋体 Std L";mso-font-kerning:1.0 pt;ms o-ansi-language: EN-US;mso-fareast-language:ZH-CN;mso-bidi-language:AR-SA'> assess the p athogenicity of the discovered mutations. RT-q PCR was used to analyze the expression level of PPIB gene. Conclusion We reported a rare pedigree with recessive osteogenesis imperfecta(OI) type Ⅸ ca used by PPIB gene mutations in China for the first time. The current study expands our knowledg e about the mutations of PPIB gene and their associated phenotypes, and provides a new information on the genetic defects of this disease for clinical diagnosis.

关键词： Two Novel Mutations In PPIB Gene cause a Rare Pedigree With Osteogenesis Imperfecta Type

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Common AZFc structure may possess the optimal spermatogenesis efficiency relative to the rearranged structures mediated by non-allele homologous recombination

Common AZFc structure may possess the optimal spermatogenesi...

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中华医学会第十五次全国医学遗传学学术会议暨中国医师协会医学遗传医师分会第一届全国学术会议暨2016年浙江省医学遗传学年会

作者： Yang Yuan Yang Bo Ma Yongyi Liu Yunqiang Li Lei Yang Dong Tu Wenling Shen Ying Dong Qiang Department of Medical Genetics State Key Laboratory of Biotherapy West China HospitalSichuan University Department of Urology West China Hospital Sichuan University Reproductive Medicine Centre West China Second Hospital Sichuan University Reproductive Medicine Institute Chengdu Women's & Children's Central Hospital

Objective The azoopsermia factor c(AZFc) region of human Y-chromosome is an essential geno mic segment for spermatogenesis with frequent non-allele homologous recombination(NAHR). R ecent case-control studies on the association of the NAHR-based AZFc structural mutations with spermatogenic failure produced inconsistent results. Method To more precisely evaluate their spermatogenesis effects, we investigated the correlatio n between the subdivided AZFc mutations and sperm production in 3,439 Han Chinese males. Result Our results showed that both partial AZFc deletion-only and primary duplication mutation presented a significant risk for decreased sperm production. Restoration of the reduced dosage of the AZFc content to the normal level had a milder effect, whereas an overdose of the A ZFc content arising from multiple duplications of a partial AZFc-deleted structure produced a mor e serious consequence compared to the partial deletion-only mutation. Additionally, the AZFc-mut ated structures with excessive NAHR-substrate showed a notably negative effect on spermatoge nesis. Conclusion These results suggest that the recurrent NAHR-based AZFc mutations may be asso ciated with decreased spermatogenesis efficiency in present population. More significantly, our fin ding implies that the overdose of AZFc NAHR-substrate may produce an additional risk for the m assive AZFbc deletions during the multi-stage division process of germ cells and thus impair the g lobal spermatogenesis efficiency in the carriers.

关键词： Common AZFc structure may possess the optimal spermatogenesis efficiency relative to the rearranged structures mediated by non-allele homologous recombination

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Systemic Evaluation MALBAC and MDA through Sanger Sequencing and Next-Generation Sequencing in Preimplantation Genetic Diagnosis (PGD) for β-Thalassemia

Systemic Evaluation MALBAC and MDA through Sanger Sequencing...

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中华医学会第十五次全国医学遗传学学术会议暨中国医师协会医学遗传医师分会第一届全国学术会议暨2016年浙江省医学遗传学年会

作者： He Fei Zhou Wanjun Cai Ren Xu Xiangmin Southern Medical University Liuzhou Municipal Maternity and Child Healthcare Hospital

Objective To systematically evaluate multiple annealing and looping-based amplification cycle(MALBAC) and multiple displacement amplification(MDA) in the application of preimplantation genetic diagnosis(PGD) for β-thalassemia. Method We established the monitoring and evaluation system in single cell level and multiplex cells level. Single peripheral lymphocytes were isolated from whole blood which containing eight of the most common β-globin gene mutations and subsequently amplified with MALBAC and MDA. And then we assessed the performance based on the preliminary experiments using homozygous mutant or double heterozygous mutant cell lines. Further, we collected the abandoned embryos including cleavage stage and blastocyst stage requiring one of the spouses was β-thalassemia carriers at least to verify the results of previous studies and lay the foundation for clinical testing. We applied direct Sanger sequencing of the PCR products from a portion of the WGA products to detect mutations within the β-globin gene. The remaining WGA products would be tested CNV and SNP through Next-generation Sequencing. Statistical analysis the difference of the performance indexes between the two methods in different level, for example the amplificationefficiency/ADO/CV value and so on. Result In single-cell level, MDA is higher than MALBAC in SNP detection and lower in ADO(9.74% vs 14.1%) and there is statistically significant difference. However MALBAC has better uniformity and lower CV value(0.153). In multiplex cells level, MDA has better property than MALBAC in success rate for mutation detection(92.78% vs 80.21%),ADO(7.22% vs 19.79%), SNP detection for ADO(2.4% vs 5.87%) and other indicators and were significant differences. Conclusion In single-cell level, MALBAC has advantage in the performance of CNV detection. But in multiplex cells level, MDA has the strengths in the success rate of a single gene detection and low ADO rate. Regretful, we did not repeat the results in the d

关键词： MALBAC Systemic Evaluation MALBAC and MDA through Sanger Sequencing and Next-Generation Sequencing in Preimplantation Genetic Diagnosis Thalassemia PGD

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Downregulation of BRG1 due to the demethylation of intronic Cp G shore attenuated the expression of GATA4 in congenital heart diseases

Downregulation of BRG1 due to the demethylation of intronic ...

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中华医学会第十五次全国医学遗传学学术会议暨中国医师协会医学遗传医师分会第一届全国学术会议暨2016年浙江省医学遗传学年会

作者： Xiao Deyong Qian Yanyan Wang Huijun Ma Xiaojing Huang Guoying Ma Duan Key Laboratory of Metabolism and Molecular Medicine Ministry of Education Department of Biochemistry and Molecular Biology School of Basic Medical Sciences Fudan University Children Hospital of Fudan University Institute of Biomedical Sciences Collaborative Innovation Center of Genetics and Development School of Basic Medical Sciences Fudan University

Objective BRG1 as an ATPase subunit of SWI/SNF complex is tightly associated with the develo pment of heart. While there is rare reports on its abnormality in CHD patients. The aim of this stud y is to find the possible association between the etiology of CHD and the abnormality of BRG1 in CHD patients. Method The change of BRG1 m RNA level was sketched in normal fetal and postnatal heart. Sub sequently, BRG1 m RNA and protein expression were detected in heart tissue samples from thirt y CHD patients and eleven normal controls. Next, the methylation of BRG1 promoter in patients a nd normal subjects‘ heart was analyzed using Mass ARRAY platform and pyrosequencing. Moreo ver, the expression of GATA4, as a target gene of BRG1 was compared between patients and co ntrols. Result BRG1 showed a high expression in normal fetal heart in the second trimester, while it dec reased after birth. In an addition, in comparison to the normal subjects, BRG1 is marked lower in CHD patients‘ heart. The methylation assay showed that the Cp G shore of BRG1 in the second in tron was demethylated in patients‘ heart. Noteworthily, GATA4 regulated by BRG1 in cardiomyoc ytes also showed a significant decrease in those patients consistent with BRG1. Conclusion Early high expression of BRG1 in fetal heart is probably to maintain the normal early heart development. It is possible that The abnormal expression of BRG1 in human‘s heart further affects the expression of GATA4 and causes CHDs.

关键词： BRG Downregulation of BRG1 due to the demethylation of intronic Cp G shore attenuated the expression of GATA4 in congenital heart diseases

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LINC00651 promote cell migration and invasion involved in activating SNAI1 in gastric cancer

LINC00651 promote cell migration and invasion involved in ac...

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中华医学会第十五次全国医学遗传学学术会议暨中国医师协会医学遗传医师分会第一届全国学术会议暨2016年浙江省医学遗传学年会

作者： Xiufang Liu Huazhang Wu Ying Hu Pihai Gong Wei Song Xiaohui Shen Department of Medical Genetics and Developmental Biology Medical School of Southeast University

Objective Gastric cancer(GC) is the third leading cause of cancer-related deaths worldwide. Mor e than half of GC patients were diagnosed at an advanced stage accompanied by malignant prolif eration or lymphatic metastasis. Metastatic cancer is the main reason leading to death of GC pati ents. Epigenetic regulators and non-coding RNAs have recently gained significant attention in deli neating the complex mechanisms underlying malignant cancer cells metastasis, lots of evidence have shown that nc RNAs played a n important role in cancer progression, and could provide new insights into the malignant biologic al behavior of gastric *** To explore the epigenetic mechanism on cell migration related to GC metastasis, long no n-coding RNA expression profiling have been investigated through lnc RNA microarray. e RNA LIN C00651 is one of overexpression lnc RNA in lung cancer and breast cancer. In our present study, we detected the expression pattern of e RNA LINC00651 in GC cell lines and c ases. The relative expression of LINC00651 detected by q PCR in clinical tissue samples showed the expression levels of LINC00651 is related to the clinicopathological features as well. Increase d expression of LINC00651 was positively correlated with lymph node metastasis in gastric cance r. Employing wound healing assay, transwell migration assay, matrigel invasion assay and flow cy tometry, we analyzed the function of LINC00651 on the migration, invasion and proliferation of G C cells. LINC00651 promoted the cell invasion and migration instead of cell proliferation in GC in vitro. In order to unveil the molecular mechanisms underlying the migration and invation induced by LINC00651, we use Chr omation mmunopricipitation(Ch IP) assay, RNA immunoprecipitation(RIP) assay and molecular b iology technology such as q PCR and western blot in the further study. Result Our data suggested that LINC00651 activated its adjacent gene SNAI1, an EMT related tr anscript factor via enhancer-l

关键词： LINC00651 promote cell migration and invasion involved in activating SNAI1 in gastric cancer

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A screening in Piggy Bac transgenic mice for new candidate genes in neural tube and congenital heart defects

A screening in Piggy Bac transgenic mice for new candidate g...

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中华医学会第十五次全国医学遗传学学术会议暨中国医师协会医学遗传医师分会第一届全国学术会议暨2016年浙江省医学遗传学年会

作者： Zheng Yufang Jing Yingchun Chen Zhongzhong Shi Zhiwen Liu Weiqi Liu Jiaojiao Wang Chunyan Chen Shuxia Xu Hong Wu Xiaohui Wang Hongyan State Key Laboratory of Genetic Engineering MOE Key Laboratory of Contemporary Anthropology and Coll aborative Innovation Center for Genetics & Development School of Life Sciences Fudan University The Institute of Developmental Biology and Molecular Medicine of Fudan University Key Lab of Reproduction Regulation of NPFPC in SIPPR Institute of Reproduction & Development in Obstetrics & Gynecology Hospital Fudan University The children hospital of Fudan University

Objective Neural tube defects(NTDs) and congenital heart disease(CHDs) are the top two birth defects in China and among the world. Genetic factors play an important role in the progression o f these diseases as more than 70% NTDs can be attributed to genetic factors. Identifying more ca usative genes and further understanding the genetic mechanism behind those birth defects can h elp to establish a prenatal genetic screening list, which is a key step in reducing the incidence of NTDs and CHDs. Method It is very hard to find relative pedigree due to high mortality and reproductive fitness. The refore, animal models are very important resource to sorting the candidate genes. By using the es tablished Piggy Bac transposon inserted transgenic mice, we performed a large-scale screening o n embryonic lethal lines for NTDs or/and CHDs phenotypes. A Chinese sporadic NTDs cohort wa s whole-genome sequenced and the top candidated genes were analyzed. Result Our screening covers 342 PB lines which are embryonic or perinatal lethal. According to MGI database, 295 lines had not been reported for knockout mouse. Within those 295 lines, there are 29.2%(86/295) lines showing NTD and 6.4%(19/295) showing CHD phenotype. Whole-genome sequencing was performed on 100 sporad ic Chinese NTDs samples. The mutation rate within those candidate genes are higher than other non-NTD related house keeping genes. Conclusion Our work successfully identified large number of new candidate genes for birth defec ts, which could provide new windows to understand the genetic and molecular mechanisms behind those birth defects.

关键词： A screening in Piggy Bac transgenic mice for new candidate genes in neural tube and congenital heart defects

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Combined anti-HBV and anti-PD1 gRNA/cas9 produces a synergistic antiviral effect in HBV infection

Combined anti-HBV and anti-PD1 gRNA/cas9 produces a synergis...

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中华医学会第十五次全国医学遗传学学术会议暨中国医师协会医学遗传医师分会第一届全国学术会议暨2016年浙江省医学遗传学年会

作者： Zhen Shuai Hua Ling Liu Yunhui Lu Jiaojiao Chen Wei Zhao Le Li Xu the First Affiliated Hospital Xi'an Jiaotong University Rongchang Campus southwest University Department of Pharmacology and Toxicology Beijing Institute of Radiation Medicine

Objective Currently, no licensed therapy can thoroughly eradicate hepatitis B virus(HBV) from th e body, including interferon αand inhibitors of HBV reverse-transcription. Recently, clustered regu larly interspaced short palindromic repeat(CRISPR) systems, originally identified in bacteria and archaea, have been found to have effect on the pre-existing HBV covalently closed circular DN A. Targeted HBV with unique CRISPR/Cas9 therapy results in objective responses in HBV treatm ent, however the responses may be short. In contrast, treatment with immune checkpoint inhibitor s results in a lower responses rate, but the responses tend to be more durable. Based on these fi ndings, we hypothesized that surface antigen(HBs Ag)-encoding region of HBV-targeted therapy may synergize with the PD-1 pathway blockade to enhance anti-HBV activity. Method To test hypothesis, we described the effects of the CRISPR/Cas9 that was targeted to th e surface antigen(HBs Ag)-encoding region of HBV and PD1 to explore the response of transgeni c mice. HBs Ag in the media of the sera of mice were analyzed by a quantitative enzyme-linked immunosorbent assay;HBV DNA levels were assessed by fluorogenic quantitative polymerase c hain reaction, immune phenotypes and cytokines expression of human dendritic cells was detecte d by flow cytometry and PCR, and HBs Ag expression in the livers of the mice was assessed by a n immunohistochemical assay. Result The CRISPR/Cas9 system efficiently produced mutations in HBV DNA. HBs Ag secreted i n mouse serum was reduced by CRISPR/Cas9 treatment;Immunohistochemistry showed almo st no HBs Ag-positive cells in the liver tissues of CRISPR/Cas9-treated mice, especially in the co mbined groups The CRISPR/Cas9 system efficiently produced mutations in HBV *** For the first time, our studies demonstrate potent synergistic effects of combination t herapy using HBV-targeted therapy and immune checkpoint blockade PD1 could strengthen the i mmune response and

关键词： HBV DNA CRISPR Combined anti-HBV and anti-PD1 gRNA/cas9 produces a synergistic antiviral effect in HBV infection

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The function and mechanism of CUL4B in obesity and adipogenesis

The function and mechanism of CUL4B in obesity and adipogene...

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中华医学会第十五次全国医学遗传学学术会议暨中国医师协会医学遗传医师分会第一届全国学术会议暨2016年浙江省医学遗传学年会

作者： Li Peishan Song Yu Zan Wenying Shao Changshun Gong Yaoqin The Key Laboratory of Experimental Teratology Ministry of Education and Department of Molecular Medicine and Genetics Shandong University School of Medicine

Objective CULLIN-RING E3 ligases represent the largest mammalian E3 ligase family, in which each of Cullin family members including CUL4 B serves as a molecular scaffold. Mutations in hum an CUL4 B have been found to be a common cause of X-linked mental retardation. In addition to b eing mentally retarded, patients with CUL4 B mutations also manifest short stature, abnormal gait, absence of speech, central obesity, and other developmental defects. However, the role of CUL4 B in adipogenesis and obesity remains unknown. Method In this study, we investigate the role of CUL4 B in obesity and adipogenesis using adipoc yte-specific Cul4 b knockout mouse(referred to AKO). Result CUL4 B expression in adipose tissue was significantly down-regulated in obesity mice and during adipocyte differentiation, which indicating that the expression of CUL4 B appears to be neg atively correlated with obesity. Using 3 T3-L1 preadipocytes, we found CUL4 B functions as a nega tive regulator of adipogenesis. Next, we generated adipocyte-specific Cul4 b knockout mice. Whe n placed on a HFD, the weight gain and ing WAT mass in AKO mice was significantly accelerated compared to their WT littermates. Adipoctyes from AKO mice appeared to be smaller than WT adi pocytes, especially in ing WAT, indicating that hyperplasia may play a major role in adipose tissue expansion in AKO mice. This result was corrected with the data that knockdown Cul4 b in 3 T3-L1 cells increase adipogenesis. Both glucose intolerance and insulin resistance and adipose tissue i nflammation induced by HFD were significantly attenuated in AKO mice when compared to WT m ice, these results suggest that adipocyte-specific Cul4 b deficiency confers an overall improvemen t in the metabolic phenotypes associated with HFD-induced *** The results that CUL4 B negatively regulate obesity in animal model were consistentl y with the phenotype found in humans with CUL4 B mutations. As the AKO mice with smaller adip ocytes a

关键词： The function and mechanism of CUL4B in obesity and adipogenesis

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