AAV2-PDE6B restores retinal structure and function in the retinal degeneration 10 mouse model of retinitis pigmentosa by promoting phototransduction and inhibiting apoptosis

Retinitis pigmentosa is a group of inherited diseases that lead to retinal degeneration and photoreceptor cell death.However,there is no effective treatment for retinitis pigmentosa caused by PDE6B mutation.Adeno-associated virus(AAV)-mediated gene therapy is a promising strategy for treating retinitis pigmentosa.The aim of this study was to explore the molecular mechanisms by which AAV2-PDE6B rescues retinal function.To do this,we injected retinal degeneration 10(rd10)mice subretinally with AAV2-PDE6B and assessed the therapeutic effects on retinal function and structure using dark-and light-adapted electroretinogram,optical coherence tomography,and immunofluorescence.Data-independent acquisition-mass spectrometry-based proteomic analysis was conducted to investigate protein expression levels and pathway enrichment,and the results from this analysis were verified by real-time polymerase chain reaction and western blotting.AAV2-PDE6B injection significantly upregulated PDE6βexpression,preserved electroretinogram responses,and preserved outer nuclear layer thickness in rd10 mice.Differentially expressed proteins between wild-type and rd10 mice were closely related to visual perception,and treating rd10 mice with AAV2-PDE6B restored differentially expressed protein expression to levels similar to those seen in wild-type mice.Kyoto Encyclopedia of Genes and Genome analysis showed that the differentially expressed proteins whose expression was most significantly altered by AAV2-PDE6B injection were enriched in phototransduction pathways.Furthermore,the phototransductionrelated proteins Pde6α,Rom1,Rho,Aldh1a1,and Rbp1 exhibited opposite expression patterns in rd10 mice with or without AAV2-PDE6B treatment.Finally,Bax/Bcl-2,p-ERK/ERK,and p-c-Fos/c-Fos expression levels decreased in rd10 mice following AAV2-PDE6B treatment.Our data suggest that AAV2-PDE6B-mediated gene therapy promotes phototransduction and inhibits apoptosis by inhibiting the ERK signaling pathway and upregulating Bcl-2/Bax expression in retinitis pigmentosa.

Identification of hub genes associated with Helicobacter pylori infection and type 2 diabetes mellitus:A pilot bioinformatics study

BACKGROUND Helicobacter pylori(H.pylori)infection is related to various extragastric diseases including type 2 diabetes mellitus(T2DM).However,the possible mechanisms connecting H.pylori infection and T2DM remain unknown.AIM To explore potential molecular connections between H.pylori infection and T2DM.METHODS We extracted gene expression arrays from three online datasets(GSE60427,GSE27411 and GSE115601).Differentially expressed genes(DEGs)commonly present in patients with H.pylori infection and T2DM were identified.Hub genes were validated using human gastric biopsy samples.Correlations between hub genes and immune cell infiltration,miRNAs,and transcription factors(TFs)were further analyzed.RESULTS A total of 67 DEGs were commonly presented in patients with H.pylori infection and T2DM.Five significantly upregulated hub genes,including TLR4,ITGAM,C5AR1,FCER1G,and FCGR2A,were finally identified,all of which are closely related to immune cell infiltration.The gene-miRNA analysis detected 13 miRNAs with at least two gene cross-links.TF-gene interaction networks showed that TLR4 was coregulated by 26 TFs,the largest number of TFs among the 5 hub genes.CONCLUSION We identified five hub genes that may have molecular connections between H.pylori infection and T2DM.This study provides new insights into the pathogenesis of H.pylori-induced onset of T2DM.

Gene expression analysis of cytokines and MMPs in melatonin and rhBMP-2 enhanced bone remodeling

BACKGROUND In the medical and dental fields,there is a need for studies of new therapeutic approaches for the treatment of bone defects that cause extensive bone loss.Melatonin may be an important endogenous biological factor for bone remodeling,and growth factors may enhance the repair process.AIM To evaluate the gene expression of cytokines(IL-1β,IL-6,IL-10 and TNF-α),markers of osteoclastogenesis(RANK,RANKL and OPG)and MMPs(MMP-1,MMP-2,MMP-8 and MMP-13)from the treatment of melatonin associated with an osteogenic membrane and rhBMP-2 on the recovery of a bone injury.METHODS Sixty-four rats were used and divided into 9 experimental groups and were formed according to the treatment carried out in the region of the bone lesion,which varied between the combination of 1,10 and 100μmol/L of melatonin.Gene Expression analysis was performed using real time-PCR by reading the concentration of total RNA and reverse transcription.RESULTS There were differences between groups when compared with clot or scaffold control,and improvement with a higher concentration of melatonin or rhBMP-2.The combination melatonin(1μg)with 5μg of rhBMP-2,using the guided bone regeneration technique,demonstrated some effects,albeit mild,on bone repair of critical bone defects.CONCLUSION This indicates that the approach for administering these substances needs to be reassessed,with the goal of ensuring their direct application to the affected area.Therefore,future research must be carried out,seeking to produce materials with these ideal characteristics.

应用Minigene剪接变异体分析技术诊断PMM2基因非经典剪接位点新变异的致病性

目的:研究Minigene剪接变异体分析技术在诊断磷酸甘露糖变位酶2(PMM2)相关先天性糖基化障碍(PMM2-CDG)中的价值,探讨磷酸甘露糖变位酶2(PMM2)基因剪接位点新变异对其转录产物的影响。方法:通过对1例PMM2-CDG患儿进行高通量测序查找可能的遗传学病因,利用Minigene剪接变异体分析技术,研究PMM2基因新剪接位点变异的致病性。根据美国医学遗传学与基因组学学会(ACMG)指南,判断新变异的致病性。结果:遗传学分析发现患儿系PMM2基因母源性c.691G>A(p.Val231Met)变异和父源性c.447+5G>A变异复合杂合子。Minigene剪接变异体分析发现:变异c.447+5G>A导致PMM2基因转录产物形成r.348_447del转录本,为致病性PMM2基因变异。患儿的临床特征为皮肤巩膜黄染,血清总胆红素、非结合胆红素和总胆汁酸明显升高,白蛋白明显降低,甲胎蛋白、铁蛋白和促甲状腺素等升高,对症支持治疗效果欠佳。结论:Minigene剪接变异体分析可为PMM2-CDG确诊和家系遗传咨询提供新的分子标记物,扩展了PMM2基因变异谱,为该病的临床诊治提供新的参考依据。

Vanillylacetone attenuates cadmium chloride-induced hippocampal damage and memory loss through upregulation of nuclear factor erythroid 2-related factor 2 gene and protein expression

Memory loss and dementia are major public health concerns with a substantial economic burden.Oxidative stress has been shown to play a crucial role in the pathophysiology of hippocampal damage-induced memory impairment.To investigate whether the antioxidant and anti-inflammatory compound vanillyla cetone(zingerone) can protect against hippocampal damage and memory loss induced by cadmium chloride(CdCl_(2)) administration in rats,we explo red the potential involvement of the nuclear factor erythroid 2-related factor 2(Nrf2) signaling pathway,which is known to modulate oxidative stress and inflammation.Sixty healt hy male Wistar rats were divided into five groups:vehicle-treated(control),vanillylacetone,CdCl_(2),vanillylacetone+ CdCl_(2),vanillylacetone+ CdCl_(2)+ brusatol(a selective pharmacological N rf2inhibitor) groups.Vanillylacetone effectively attenuated CdCl_(2)-induced damage in the dental gyrus of the hippocampus and improved the memory function assessed by the Morris Water Maze test.Additionally,vanillylacetone markedly decreased the hippocampal tissue levels of inflammatory biomarkers(interleukin-6,tumor necrosis factor-α,intracellular cell adhesive molecules) and apoptosis biomarkers(Bax and cleaved caspase-3).The control and CdCl_(2)-treated groups treated with va nillylacetone showed reduced generation of reactive oxygen species,decreased malondialdehyde levels,and increased superoxide dismutase and glutathione activities,along with significant elevation of nuclear Nrf2 mRNA and protein expression in hippocampal tissue.All the protective effects of vanillylacetone we re substantially blocked by the co-administration of brusatol(a selective N rf2 inhibitor).Va nillylacetone mitigated hippocampal damage and memory loss induced by CdCl_(2),at least in part, by activating the nuclear transcription factor Nrf2.Additionally,vanillylacetone exerted its potent antioxidant and antiinflammatory actions.

To Analyze the Sensitivity of RT-PCR Assays Employing S Gene Target Failure with Whole Genome Sequencing Data during Third Wave by SARS-CoV-2 Omicron Variant

Introduction: Omicron is a highly divergent variant of concern (VOCs) of a severe acute respiratory syndrome SARS-CoV-2. It carries a high number of mutations in its spike protein hence;it is more transmissible in the community by immune evasion mechanisms. Due to mutation within S gene, most Omicron variants have reported S gene target failure (SGTF) with some commercially available PCR kits. Such diagnostic features can be used as markers to screen Omicron. However, Whole Genome Sequencing (WGS) is the only gold standard approach to confirm novel microorganisms at genetically level as similar mutations can also be found in other variants that are circulating at low frequencies worldwide. This Retrospective study is aimed to assess RT-PCR sensitivity in the detection of S gene target failure in comparison with whole genome sequencing to detect variants of Omicron. Methods: We have analysed retrospective data of SARS-CoV-2 positive RT-PCR samples for S gene target failure (SGTF) with TaqPath COVID-19 RT-PCR Combo Kit (ThermoFisher) and combined with sequencing technologies to study the emerged pattern of SARS-CoV-2 variants during third wave at the tertiary care centre, Surat. Results: From the first day of December 2021 till the end of February 2022, a total of 321,803 diagnostic RT-PCR tests for SARS-CoV-2 were performed, of which 20,566 positive cases were reported at our tertiary care centre with an average cumulative positivity of 6.39% over a period of three months. In the month of December 21 samples characterized by the SGTF (70/129) were suggestive of being infected by the Omicron variant and identified as Omicron (B.1.1.529 lineage) when sequence. In the month of January, we analysed a subset of samples (n = 618) with SGTF (24%) and without SGTF (76%) with Ct values Conclusions: During the COVID-19 pandemic, it took almost more than 15 days to diagnose infection and identify pathogen by sequencing technology. In contrast to that molecular assay provided quick identification with the help of SGTF phenomenon within 5 hours of duration. This strategy helps scientists and health policymakers for the quick isolation and identification of clusters. That ultimately results in a decreased transmission of pathogen among the community.

Analysis of SMOC2 gene variants in familial and nonfamilial primary open angle glaucoma Pakistani patients

AIM:To find out the association of secreted protein acidic and rich in cysteine(SPARC)-related modular calcium binding 2(SMOC2)gene variants rs2255680 and rs13208776 with genotypic and phenotypic characteristics in both familial and non-familial primary open angle glaucoma(POAG)patients.METHODS:A total of 212 POAG patients,comprising 124 familial and 88 non-familial,were enrolled.For genotyping the SMOC2 variant rs2255680,amplification refractory mutation system(ARMS)-polymerase chain reaction(PCR)method and PCR-restriction fragment length polymorphism(PCR-RFLP)were utilized for analyzing rs13208776 variant.RESULTS:The mean age of familial POAG patients was 50.92±9.12y,with 78 males and 46 females.The mean age of non-familial POAG patients was 53.14±13.44y,with 52 males and 36 females.The SMOC2 gene variant rs13208776 showed the significant association with POAG between familial and non-familial groups.The homozygous G/G variant was frequent among non-familial(60.2%)whereas the heterozygous G/A variant was more frequent in familial POAG patients(46%).There were significant differences in G/A variant between familial and non-familial glaucoma patients,and the risk was decreased to 0.53-fold in non-familial glaucoma patients[odds ratio(OR):0.53;95%confidence interval(CI):0.29-0.94;P=0.033]in codominant model.The risk was further reduced to 0.49-fold(95%CI:0.28-0.86;P=0.012)in dominant model for non-familial patients.No significant association of SMOC2 gene variant rs2255680 between familial and non-familial glaucoma patients was found in our population.The haplotype analysis showed the decreased risk for TA[OR:0.48(95%CI:0.29-0.79);P=0.004]and an increased risk for TG[OR=2.28(95%CI:1.22-4.25);P=0.01]haplotypes.CONCLUSION:Current findings show significant association of SMOC2 gene variant rs13208776 with POAG between familial and non-familial Pakistani patients.

Regulatory potential of soil available carbon,nitrogen,and functional genes on N_(2)O emissions in two upland plantation systems

Dynamic nitrification and denitrification processes are affected by changes in soil redox conditions,and they play a vital role in regulating soil N_(2)O emissions in rice-based cultivation.It is imperative to understand the influences of different upland crop planting systems on soil N_(2)O emissions.In this study,we focused on two representative rotation systems in Central China:rapeseed–rice(RR)and wheat–rice(WR).We examined the biotic and abiotic processes underlying the impacts of these upland plantings on soil N_(2)O emissions.The results revealed that during the rapeseed-cultivated seasons in the RR rotation system,the average N_(2)O emissions were 1.24±0.20 and 0.81±0.11 kg N ha^(–1)for the first and second seasons,respectively.These values were comparable to the N_(2)O emissions observed during the first and second wheat-cultivated seasons in the WR rotation system(0.98±0.25 and 0.70±0.04 kg N ha^(–1),respectively).This suggests that upland cultivation has minimal impacts on soil N_(2)O emissions in the two rotation systems.Strong positive correlations were found between N_(2)O fluxes and soil ammonium(NH_(4)^(+)),nitrate(NO_(3)^(–)),microbial biomass nitrogen(MBN),and the ratio of soil dissolved organic carbon(DOC)to NO_(3)^(–)in both RR and WR rotation systems.Moreover,the presence of the AOA-amoA and nirK genes were positively associated with soil N_(2)O fluxes in the RR and WR systems,respectively.This implies that these genes may have different potential roles in facilitating microbial N_(2)O production in various upland plantation models.By using a structural equation model,we found that soil moisture,mineral N,MBN,and the AOA-amoA gene accounted for over 50%of the effects on N_(2)O emissions in the RR rotation system.In the WR rotation system,soil moisture,mineral N,MBN,and the AOA-amoA and nirK genes had a combined impact of over 70%on N_(2)O emissions.These findings demonstrate the interactive effects of functional genes and soil factors,including soil physical characteristics,available carbon and nitrogen,and their ratio,on soil N_(2)O emissions during upland cultivation seasons under rice-upland rotations.

Pathogenesis of chronic enteropathy associated with the SLCO2A1 gene:Hypotheses and conundrums

Chronic enteropathy associated with the SLCO2A1 gene(CEAS)is a complex gastroenterological condition characterized by multiple ulcers in the small intestine with chronic bleeding and protein loss.This review explores the potential mechanisms underlying the pathogenesis of CEAS,focusing on the role of SLCO2A1-encoded prostaglandin transporter OATP2A1 and its impact on prostaglandin E2(PGE2)levels.Studies have suggested that elevated PGE2 levels contribute to mucosal damage,inflammation,and disruption of the intestinal barrier.The effects of PGE2 on macrophage activation and Maxi-Cl channel functionality,as well as its interaction with nonsteroidal anti-inflammatory drugs play crucial roles in the progression of CEAS.Understanding the balance between its protective and pro-inflammatory effects and the complex interactions within the gastrointestinal tract can shed light on potential therapeutic targets for CEAS and guide the development of novel,targeted therapies.

Transglutaminase 2 serves as a pathogenic hub gene of KRAS mutant colon cancer based on integrated analysis

BACKGROUND Colon cancer is acknowledged as one of the most common malignancies worldwide,ranking third in United States regarding incidence and mortality.Notably,approximately 40%of colon cancer cases harbor oncogenic KRAS mutations,resulting in the continuous activation of epidermal growth factor receptor signaling.AIM To investigate the key pathogenic genes in KRAS mutant colon cancer holds considerable importance.METHODS Weighted gene co-expression network analysis,in combination with additional bioinformatics analysis,were conducted to screen the key factors driving the progression of KRAS mutant colon cancer.Meanwhile,various in vitro experiments were also conducted to explore the biological function of transglutaminase 2(TGM2).RESULTS Integrated analysis demonstrated that TGM2 acted as an independent prognostic factor for progression-free survival.Immunohistochemical analysis on tissue microarrays revealed that TGM2 was associated with an elevated probability of perineural invasion in patients with KRAS mutant colon cancer.Additionally,biological roles of the key gene TGM2 was also assessed,suggesting that the downregulation of TGM2 attenuated the proliferation,invasion,and migration of the KRAS mutant colon cancer cell line.CONCLUSION This study underscores the potential significance of TGM2 in the progression of KRAS mutant colon cancer.This insight not only offers a theoretical foundation for therapeutic approaches but also highlights the need for additional clinical trials and fundamental research to support our preliminary findings.

Main Agronomic Characters and Grain Quality of Rice Blast Resistance Gene Pi-d2 Transgenic Rice

[Objective] The aim of this study was to provide metabolic evidence for the analysis of the ecological and safety assessment of Pi-d2-transgenic rice.[Method] The main agronomic characters of Pi-d2-transgenic rice were observed in field experiment and the grain chemical characters and amino acid content were measured.[Results] Introduction of foreign gene Pi-d2 resulted in stably hereditable variation in agronomic characteristics in the descents.Most of the transgenic lines grew normally and orderly.Compared with the control（wild type plants）,about half of transgenic plants showed an increased or reduced plant height.There was no observable difference between transgenic plants and controls in tiller number,length of panicle,panicles per plant,seed-setting rate and 1 000-grain weight.Total amino acid content in transgenic rice was reduced,while the starch content,GC and GT were not altered in comparison with the control.[Conclusion] Introduction of foreign gene Pi-d2 has remarkable influence on plant height,while little on grain chemical characters.

Determination and Analysis of Mitochondrial ND2 Gene Sequence of Anas platyrhynchos

[Objective] The study was to analyze the phylogenesis of Anas platyrhynchos. [Method] Complete sequence of mitochondrial ND2 gene of 4 Anas platyrhynchos was determined by direct DNA sequencing based on PCR products. Combined with ND2 gene sequences of the Anas Linnaeus accessed in GenBank, phylogenetic tree was constructed by Neighbor-joining and maximum parsimony methods. [Result] The ND2 gene sequences of 4 Anas platyrhynchos were identical(1 041 bp in length; the nucleotide contents of A, G, T, and C were 28.91%, 13.35%, 20.75% and 36.98% respectively; A+T content approximated to that of C+G). Sequences of ND2 gene of mallard were same as spotbill duck, and had high homology with others. The phylogenetic trees indicated mallard and spotbilled duck were close in genetic relationship, both shared a haplotype; then Philippine duck, green-winged teal and northern pintail fell into branch ''A". [Conclusion] The domestic duck may be domesticated from mallard and spotbilled duck.

PTEN knockdown with the Y444F mutant AAV2 vector promotes axonal regeneration in the adult optic nerve

The lack of axonal regeneration is the major cause of vision loss after optic nerve injury in adult mammals. Activating the PI3K/AKT/mTOR signaling pathway has been shown to enhance the intrinsic growth capacity of neurons and to facilitate axonal regeneration in the central nervous system after injury. The deletion of the mTOR negative regulator phosphatase and tensin homolog （PTEN） enhances regeneration of adult corticospinal neurons and ganglion cells. In the present study, we used a tyrosine-mutated （Y444F） AAV2 vector to efficiently express a short hairpin RNA （shRNA） for silencing PTEN expression in retinal ganglion cells. We evaluated cell survival and axonal regeneration in a rat model of optic nerve axotomy. The rats received an intravitreal injection of wildtype AAV2 or Y444F mutant AAV2 （both carrying shRNA to PTEN） 4 weeks before optic nerve axotomy. Compared with the wildtype AAV2 vector, the Y444F mutant AAV2 vector enhanced retinal ganglia cell survival and stimulated axonal regeneration to a greater extent 6 weeks after axotomy. Moreover,post-axotomy injection of the Y444F AAV2 vector expressing the shRNA to PTEN rescued ～19% of retinal ganglion cells and induced axons to regenerate near to the optic chiasm. Taken together, our results demonstrate that PTEN knockdown with the Y444F AAV2 vector promotes retinal ganglion cell survival and stimulates long-distance axonal regeneration after optic nerve axotomy. Therefore, the Y444F AAV2 vector might be a promising gene therapy tool for treating optic nerve injury.

Sequence Analysis of Type III Effector tccP and tccP2 Genes in Escherichia coli O157:H7 from Chinese Water-chestnut

[Objective] This study aimed to analyze the type III effector tccP and tccP2 genes in Escherichia coli O157：H7 from Chinese water-chestnut. [Method] Gene-specific and locus-specific primers were utilized to amplify tccP/tccP2 and their flanking regions for sequence analysis. [Result] E. coli O157：H7 CWN11 harbored intact tccP and tccP2 genes, however, the number of proline-rich repeats in tccP gene was only one that probably resulted in biological incapability, whereas, the tccP2 gene consisted of five and half proline-rich repeats and could encode functional protein. [Conclusion] Here, we reported the first sequence of tccP gene that consisted of only one proline-rich repeat and tccP2 was assumed to play a crucial role in colonization and subsequent signaling cascades.

Cloning and Phylogenetic Analysis of NS1 Genes from Different Isolates of H9N2 Subtype Duck Influenza Virus

[ Objective] The study aimed to lay a foundation for the further studies on function mechanism of NS1 protein in the interspecies transmission of waterfowl influenza virus. [Method] Using the serologic assay and the specific RT-PCR method, some strains of H9 subtype waterfowl influenza virus were isolated from the 12 to 20 day-old muscovy duck flocks without any clinical symptoms in different areas of Guangdong Province. Four of these strains, including A/duck/ZQ/303/2007（H9N2） （A3 for short）, A/Duck/FJ/301/2007 （H9N2） （C1 for short）, A/Duck/NH/306/2007（H9N2） （ D6 for short）, A/duck/SS/402/2007（H9N2） （ E2 for short）, and a strain named A/duck/ZC/2007（H9N2） （L1 for short） from a muscovy duck died of avian influenza virus （AIV）, were used for NSl gene cloning and sequencing. Subsequently, the obtained NSl gene sequences were compared with other NS1 sequences registered in GenBank, and the phylogenetic analysis was also conducted. [Result] When compared with the H9N2 AIV NS1 sequences in GenBank, the NSl genes of the four AIV strains A3, C1, 136 and E2 displayed homologies ranging from 99% to 100% at nucleotide level, and 95% to 100% at amino acid level; while the NSl gene of L1 strain displayed homology ranging from 94% to 97% at nucleotide level, and 93% to 98% at amino acid level. The phylogenetic tree demonstrated that A3, C1, D6 and E2 were highly resemblant, and L1 was closest to AY66473 （chicken, 2003）. By comparison with the NS1 gene sequences of L1, AF523514 （duck）, AY664743 （chicken） and EF155262.1 （quail） using DNAstar, A3, C1, D6 and E.2 presented nucleotide variations at site 21 （ R→Q）, 70, 71 （ KE→EG）, 86 （ A→S）, 124 （V→M） and 225 （ S→N）, and amino acid variations at site 21,70, 71 and 86 in dsRNA- dependent protein kinase （PKR） binding domain of NSl gene, which induced the evident variations of antigenic determinant and surface proba- bility plot of NS1 protein. [ Conclusion] This study suggested that the amino acid sequence variation in PKR binding domain of NS1 protein had something to do with the virus pathogenicity.

Cytochrome P450 2E1 genetic polymorphism and gastric cancer in Changle,Fujian Province

AIM: Genetic polymorphism in enzymes of carcinogen metabolism has been found to have the influence on the susceptibility to cancer. Cytochrome P450 2E1 (CYP2E1) is considered to play an important role in the metabolic activation of procarcinogens such as N-nitrosoamines and low molecular weight organic compounds. The purpose of this study is to determine whether CYP450 2E1 polymorphisms are associated with risks of gastric cancer. METHODS: We conducted a population based case-control study in Changle county, Fujian Province, a high-risk region of gastric cancer in China. Ninety-one incident gastric cancer patients and ninety-four healthy controls were included in our study. Datas including demographic characteristics, diet intake, and alcohol and tobacco consumption of individuals in our study were completed by a standardized questionnaire.PCR-RFLP revealed three genotypes:heterozygote (C1/C2) and two homozygotes (C1/C1 and C2/C2) in CYP2E1. RESULTS: The frequency of variant genotypes (C1/C2 and C2/C2) in gastric cancer cases and controls was 36.3% and 24.5%, respectively. The rare homozygous C2/C2 genotype was found in 6 individuals in gastric cancer group(6.6%), whereas there was only one in the control group (1.1%). However, there was no statistically significant difference between the two groups (two-tailed Fisher's exact test P=0.066). Individuals in gastric cancer group were more likely to carry genotype C1/C2 (odds ratio, OR=1.50) and C2/C2 (OR=7.34) than individuals in control group (chi(2) =4.597, for trend P=0.032). The frequencies of genotypes with the C2 allele (C1/C2 and C2/C2 genotypes) were compared with those of genotypes without C2 allele (C1/C1 genotype) among individuals in gastric cancer group and control group according to the pattern of gastric cancer risk factors. The results show that individuals who exposed to these gastric cancer risk factors and carry the C2 allele seemed to have a higher risk of developing gastric cancer. CONCLUSION: Polymorphism of CYP2E1 gene may have some effect in the development of gastric cancer in Changle county, Fujian Province.

抑癌基因DOC-2对卵巢癌细胞增殖的影响

目的:探讨抑癌基因DOC2对卵巢癌细胞系HO8910在生长代谢、超微结构及细胞周期等方面的影响及其作用机制。方法:实验分3组:卵巢癌细胞系HO8910、8910P93(转染并表达DOC2基因)、8910pcDNA3.1(转染空载体pcDNA3.1),通过细胞消化计数法和3HTdR掺入法分别测定3组细胞的生长曲线及DNA合成代谢情况;利用透射电镜对细胞的超微结构进行观察比较;最后利用流式细胞仪观察卵巢癌细胞转染前后细胞周期的变化。结果:8910P93在生长增殖及DNA合成方面均明显低于HO8910和8910pcDNA3.1(P<0.05),而后两者之间无明显差异;电镜结果显示,8910P93存在内质网扩张,线粒体空泡化,溶酶体增生,部分细胞可见染色质边集等凋亡前期改变;细胞周期检测结果显示,处于G1和G2期的8910P93细胞明显增多而S期细胞明显减少。结论:抑癌基因DOC2可通过改变卵巢癌细胞的形态及细胞周期而明显抑制细胞的增殖。

人DOC-2氨基端PID结构域的克隆与原核表达

目的 :克隆人DOC 2氨基端PID结构域 (暂命名为nDOC 2 ) ,并在原核细胞中表达。方法 :用RT PCR从正常人卵巢组织中扩增nDOC 2基因 ,并克隆到载体 pUC 19中。经测序证实后 ,用BamHI/EcoRI双酶切 ,亚克隆到原核表达载体 pGEX 4T 1,并转化E .coliDH5α菌株。取工程菌 ,用IPTG诱导表达 ,对表达产物进行SDS PAGE鉴定。结果 :①经RT PCR、测序和酶切鉴定 ,成功地克隆了人nDOC 2基因。②经IPTG诱导的重组质粒pGEX 4T nDOC 2表达出相对分子质量 (Mr)约为 5 0 0 0 0的融合蛋白 ,与预期的结果相符。结论 :成功克隆到人DOC 2氨基端PID结构域 ,并在E .coliDH5α中表达出GST nDOC

兔抗人DOC-2抗血清的制备与鉴定

目的 利用大肠杆菌中表达的融合蛋白GST nDOC 2 ,制备兔抗DOC 2的多克隆抗体。方法 将克隆有人DOC 2氨基端PID结构域 (暂命名为nDOC 2 )的原核表达载体pGEX 4T nDOC 2转化大肠杆菌 ,IPTG诱导表达融合蛋白GST nDOC 2 ,经包涵体洗涤后 ,用其免疫兔 ,制备兔抗人DOC 2抗血清。并以ELISA、Westernblot和免疫组化的方法对抗血清进行鉴定。结果 用ELISA法测定抗血清的效价为 1∶32 0 0 0 ;Westernblot结果显示对应于GST nDOC 2融合蛋白位置上有阳性条带 ;免疫组化结果显示人正常卵巢上皮细胞胞浆呈阳性染色 ,人卵巢浆液性囊腺瘤上皮细胞染色呈弱阳性 ,而人卵巢癌浆液性囊腺癌上皮细胞呈阴性染色。结论 成功制备了兔抗人DOC 2抗血清 ,为研究DOC

The human δ2 glutamate receptor gene is not mutated in patients with spinocerebellar ataxia

The human glutamate receptor delta 2 gene （GRID2） shares 90%homology with the orthologous mouse gene. The mouse Grid2 gene is involved with functions of the cerebellum and sponta-neous mutation of Grid2 leads to a spinocerebellar ataxia-like phenotype. To investigate whether such mutations occur in humans, we screened for mutations in the coding sequence of GRID2 in 24 patients with familial or sporadic spinocerebellar ataxia and in 52 normal controls. We de-tected no point mutations or insertion/deletion mutations in the 16 exons of GRID2. However, a polymorphic 4 nucleotide deletion （IVS5-121_-118 GAGT） and two single nucleotide polymor-phisms （c.1251G〉T and IVS14-63C〉G） were identiifed. The frequency of these polymorphisms was similar between spinocerebellar ataxia patients and normal controls. These data indicate that spontaneous mutations do not occur in GRID2 and that the incidence of spinocerebellar ataxia in humans is not associated with GRID2 mutation or polymorphisms.