老年性痴呆患者载脂蛋白E基因多态性与淀粉样β蛋白前体16-17外显子表达的相关性及意义

目的探讨老年性痴呆(AD)患者血清载脂蛋白E(ApoE)基因多态性和淀粉样β蛋白前体16-17(β-APP16-17)外显因子表达相关性及其临床意义。方法选取2018-02—2019-08在郑州市第二人民医院诊治的32例AD患者纳入研究组,随机选取同期门诊或健康体检的老年自愿者32例纳入正常组,均接受聚合酶链反应-限制性片段长度多态性(PCR-RFLP)检测和相对定量竞争RNA扩增聚合酶链反应(RT-PCR)检测,对比分析2组检测结果。结果研究组ApoE基因型频率整体高于正常组(P<0.05);ApoEε4等位基因携带者的β-APP16-17表达水平高于ε2、ε3等位基因,但差异无统计学意义(P>0.05);2组有无ApoEε4等位基因间App16-17水平无显著性差异(P>0.05)。结论ApoEε4基因是AD发生的重要危险因子,其等位基因对患者的β-APP16-17表达影响不明显,但在目前发现的基因片段中,该基因的表达可为下一步的临床诊治提供新方向。

Cloning and Sequence Analysis of 5′ Flanking Region and Exon of Inhibin α Precursor Gene in Goats

[Objective] The aim of this study was to reveal the relationship between inihibin （INH） α precursor gene and seasonal reproduction of goats, and investigate the evolutionary conservation of INHα precursor gene. [ Method] Cloning and sequence analysis of 5＇ flanking region and exon of inihibinα （INHE） precursor gene in twenty ewes between non-seasonal estrous breed （Haimen goats） and seasonal estrous breed （Anhui white goats） was analyzed in this study. [ Result] Compared with Anhui white goats, INHα precursor gene in Haimen goats had three SNP but no amino acid change, while its nucleotide homology was 99.7% and amino acid homology was 100%. The nucleotide homology of INHα precursor gene in goat, cattle, pig, person, chicken, horse, rat and dog ranged from 12.7% to 96.5%. [ Conclusion] INHα precursor gene tends to be highly conserved in species, and any change of nucleotide and amino acid maybe directly influence the function of the whole gene coding and regulation.

Polymorphism Detection of the Twelfth Exon of Equine MxA Gene

[Objective] The study aimed to establish a fast and accurate method to detect the polymorphism of the 12^th exon of equine MxA gene. [Method] The 12^th exon of MxA gene was amplified by mismatch PCR and the products were analyzed by restriction fragment length polymorphism （RFLP） to determine the point mutation at the 1 790 nt of MxA cDNA. The sequence of the PCR products was also analyzed. [Result] There were three genotypes （AA, AB and BB） in the 12^th exon of equine MxA gene; the 2 081 nt of MxA cDNA mutated from G to C, correspondingly changing the 562^th amino acid of the coding region of MxA protein from tryptophan to cysteine; the specific sequence of the PCR products amplified by mismatch PCR-RFLP was consistent with the analysis results of RFLP. [ Conclusion] The mismatch PCR-RFLP was an easy method with accurate results to detect the polymorphism of the 12^th exon of equine MxA gene.

KIT exon 11 codon 557/558 deletion/insertion mutations define a subset of gastrointestinal stromal tumors with malignant potential

AIM: To study the association of the frequency and pattern of KIT and PDGFRA mutations and clinicopathological factors in a group of patients with gastrointestinal stromal tumors (GIST). METHODS: Thirty patients with GIST were examined. Exons 9, 11, 13, and 17 of the KIT and exons 12 and 18 of the PDGFRA gene were analyzed for the presence of mutations by PCR amplification and direct sequencing. RESULTS: KIT or PDGFRA mutations were detected in 21 of the 30 patients (70%). Sixteen patients had mutations within KIT exon 11, three within KIT exon 9, and two within PDGFRA exon 18. GISTs with KIT exon 9 mutations were predominantly located in the small intestine, showed a spindle cell phenotype, and were assessed as potentially malignant. GISTs with KIT exon 11 mutations were located in the stomach and intestine, showed mainly a spindle cell phenotype, and were scored as potentially malignant (P < 0.05). Tumors with KIT exon 11 codon 557/558 deletion/insertion mutations were found to be associated with a potentially malignant clinical behaviour (P < 0.003). GISTs with PDGFRA mutations located in stomach showed a mixedcell phenotype and were classified as of very low or low moderate malignant potential. CONCLUSION: Determination of KIT and PDGFRA mutations should be additional parameters for the better prediction of GISTs clinical behaviour. Tumors with deletion/insertion mutations affecting codons 557/558 of the KIT gene seem to represent a distinct subset of malignant GISTs.

Correlation of p53 over-expression and alteration in p53 gene detected by polymerase chain reaction-single strand conformation polymorphism in adenocarcinoma of gastric cancer patients from India

AIM： To study the alterations in p53 gene among Indian gastric cancer patients and to correlate them with the various clinicopathological parameters. METHODS： A total of 103 gastric cancer patients were included in this study. The p53 alterations were studied by both immunohistochemical method as well as polymerase chain reaction （PCR）-single strand conformation polymorphism （SSCP） analysis. We only studied four （exon 5, 6, 7, and 8） of the 11 ,p53 exons. The alterations in p53 were also correlated with respect to various clinicopathological parameters. RESULTS： Among 103 cases, p53 over-expression and alteration were detected in 37 （35.92%） and 19 （18.44%） cases, respectively. Most of the ,p53 alterations were found at exon 5 （31.54%）, followed by exon 6 （26.31%）, exon 7 （21.04%） and exon 8 （21.04%）. A significant correlation of p53 overexpression was found with p53 alteration （P = 0.000）. Concordance between ,p53 alteration （as detected by SSCP） and over-expression [as detected by immunohistochemistry （IHC）] was found in 75% cases. We found that IHC-positive/SSCP-negative cases accounted for 21% of cases and IHC-negative/SSCP- positive cases accounted for remaining 4% cases. CONCLUSION： Our results show that p53 gene mutations are significantly correlated with p53 protein over-expression, with 75% concordance in over-expression and alteration in the p53 gene, but 25% disconcordance also cautions against the assumption that p53 over-expression is always associated with a gene mutation. There may be other mechanisms responsible for stabilization and accumulation of p53 protein with no evidence of gene mutation that reflect an accumulation of a non-mutated protein, or a false negative SSCP result.

MUTATIONS IDENTIFIED IN EXON 7 OFPHENYLALANINE HYDROXYLASE GENE IN CHINESE

Exon 7 of the l’henylalan1ne hydroxylase (PAH) gene was analyzed in 15 chlldren affected wlth classicphenylketonL1rla (PKU) from northern Chlna by uslng PCRxsingle strand conformation polymorphism(PCR-SSCP) technique and DNA direct sequencing. Six missense mutatlons (l. e. R2413Q. R 241H, G247V,1,2 19H, F2541;lnd G257V )and one silent rnutatlon (V245v ) were identified. The latter three missense mu-tations were demonstrated as novel mltations in comparison with the PAH mutation database. one missense mt1tation (R241 H) was flrst dowumeTlted in Chinese. our results showed populatlon ancl reglon tllffer-ences in the PAH mutation clistribution. and suggest that there is more thfln one founding population forPKU in China. The fincling of novel mutations will enhence the molecular diagnosis of PKU.

Mutations in Exons of the CYP17-Ⅱ Gene Affect Sex Steroid Concentration in Male Japanese Flounder(Paralichthys olivaceus)

As a specific gene of fish,cytochrome P450c17-Ⅱ(CYP17-Ⅱ) gene plays a key role in the growth,development and reproduction level of fish.In this study,the single-stranded conformational polymorphism(SSCP) technique was used to characterize polymorphisms within the coding region of CYP17-Ⅱ gene in a population of 75 male Japanese flounder(Paralichthys olivaceus).Three single nucleotide polymorphisms(SNPs) were identified in CYP17-Ⅱ gene of Japanese flounder.They were c.G594A(p.G188R),c.G939A and c.G1502A(p.G490D).SNP1(c.G594A),located in exon 4 of CYP17-Ⅱ gene,was significantly associated with gonadosomatic index(GSI).Individuals with genotype GG of SNP1 had significantly lower GSI(P < 0.05) than those with geno-type AA or AG.SNP2(c.G939A) located at the CpG island of CYP17-Ⅱ gene.The mutation changed the methylation of exon 6.Indi-viduals with genotype AA of SNP2 had significantly lower serum testosterone(T) level and hepatosomatic index(HSI) compared to those with genotype GG.The results suggested that SNP2 could influence the reproductive endocrine of male Japanese flounder.How-ever,the SNP3(c.G1502A) located in exon 9 did not affect the four measured reproductive traits.This study showed that CYP17-Ⅱ gene could be a potentially useful candidate gene for the research of genetic breeding and physiological aspects of Japanese flounder.

The expressions and interrelation of p130Cas and PTEN in gastric cancer

Objective： To research the contributions of p130Cas and PTEN signal molecules to the carcinogenesis of gastric carcinoma and the relationship between them. Methods： Detecting proteins of p130Cas, PTEN and PTEN mRNA of 76 cases normal gastric mucosa and 112 cases gastric carcinoma by immunohistochemistry EnVision method and molecular hybridization in situ method respectively. Detecting PTEN genetic mutation of 30 cases normal gastric mucosa, 7 cases early gastric cancer and 30 cases progressive gastric cancer by PCR-SSCP. Results： The expression of p130Cas protein of gastric carcinoma increased significantly than that of normal gastric mucosa （P 〈 0.05）. Opposite to above, the expression of PTEN protein of gastric carcinoma group was significantly lower than that of normal gastric rnucosa group （P 〈 0.05）. The expression of PTEN mRNA of gastric carcinoma group decreased obviously than normal gastric mucosa group （P 〈 0.001）. Only one case exon 5 and one case exon 8 of PTEN appeared gene mutation of progressive gastric carcinoma group, the difference has no significance compared with normal gastric mucosa group and early gastric cancer group. Conclusion： The signaling molecules p130Cas and PTEN play an important role in the carcinogenesis of gastric carcinoma, and p130Cas plays the part of promoter, oppositely, maybe PTEN can inhibit it.