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Three new alternative splicing variants of human cytochrome P450 2D6 mRNA in human extratumoral liver tissue

World Journal of Gastroenterology 2004年第22期10卷 3356-3360页

作者： JianZhuge ying-nianyu DepartmentofPathophysiology EnvironmentalGenomicsCenterSchoolofMedicineZhejiangUniversityHangzhou310031ZhejiangProvinceChina

AIM: To identify the new alternative splicing variants of human CYP2D6 in human extratumoral liver tissue with RT-PCR and sequencing. METHODS: Full length of human CYP2D6 cDNAs was amplificated by reverse transcription-polymerase chain reaction (RT-PCR) from a human extratumoral liver tissue and cloned into pGEM-T vector. The cDNA was sequenced. Exons from 1 to 4 of human CYP2D6 cDNAs were also amplificated by RT-PCR from extratumoral liver tissues of 17 human hepatocellular carcinomas. Some RT-PCR products were sequenced. Exons 1 to 4 of CYP2D6 gene were amplified by PCR from extratumoral liver tissue DNA. Two PCR products from extratumoral liver tissues expressing skipped mRNA were partially sequenced. RESULTS: One of the CYP2D6 cDNAs had 470 nucleotides from 79 to 548 (3' portion of exons 1 to 5' portion of exon 4), and was skipped. Exons 1 to 4 of CYP2D6 cDNA were assayed with RT-PCR in 17 extratumoral liver tissues. Both wild type and skipped mRNAs were expressed in 4 samples, only wild type mRNA was expressed in 5 samples, and only skipped mRNA was expressed in 8 samples. Two more variants were identified by sequencing the RT-PCR products of exons 1 to 4 of CYP2D6 cDNA. The second variant skipped 411 nucleotides from 175 to 585. This variant was identified in 4 different liver tissues by sequencing the RT-PCR products. We sequenced partially 2 of the PCR products amplified of CYP2D6 exon 1 to exon 4 from extratumoral liver tissue genomic DNA that only expressed skipped mRNA by RT-PCR. No point mutations around exon 1, intron 1, and exon 4, and no deletion in CYP2D6 gene were detected. The third variant was the skipped exon 3, and 153 bp was lost. CONCLUSION: Three new alternative splicing variants of CYP2D6 mRNA have been identified. They may not be caused by gene mutation and may lose CYP2D6 activity and act as a down-regulator of CYP2D6.

关键词： Alternative Splicing Base Sequence Carcinoma, Hepatocellular Cytochrome P-450 CYP2D6 DNA, Complementary Exons Humans Liver Liver Neoplasms Molecular Sequence Data Mutation RNA, Messenger Research Support, Non-U.S. Gov't Reverse Transcriptase Polymerase Chain Reaction

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Establishment of a transgenic cell line stably expressing human cytochrome P450 2C18 and identification of a CYP2C18 clone with exon 5 missing

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World Journal of Gastroenterology 2002年第5期8卷 888-892页

作者： JianZhu-GE ying-nianyu DepartmentofPathophysiologyandLaboratoryofMedicalMolecularBiology ZhejiangUniversitySchoolofMedicineHangzhou310031 DepartmentofPathophysiologyandLaboratoryofMedicalMolecularBiology ZhejiangUniversity

AIM: The human cytochrome P-450 2C18(CYP2C18) hasbeen characterized. However, the protein has not beenpurified from liver and very little is known regarding thespecific substrate of CYP2C18. In order to study its enzymaticactivity for drug metabolism, the CYP2C18cDNA was clonedand a stable CHL cell line expressing recombinant CYP 2C18was ***: The human CYP2C18cDNA was amplified withreverse transcription-polymerase chain reaction (RT-PCR)from total RNAs extracted from human liver and cloned intopGEM-T vector. The cDNA segment was identified by DNAsequencing and subcloned into a mammalian expressionvector pREP9. A transgenic cell line was established bytransfecting the recombinant plasmid of pREPg-CYP2C18toChinese hamster lung (CHL) cell. The enzyme activity ofCYP2C18 catalyzing oxidation of tolbutamide tohydroxytolbutamide in postmitochondrial supernant(Sg)fraction of the cell was determined by high performanceliquid chromatography(HPLC).RESULTS: The amino acid sequence predicted from thecloned cDNA segment was identical to that of reported byRomkes et al(GenBank accession number: M61856,J05326).The S9 fraction of the established cell line metabolizestolbutamide to hydroxytolbutamide. Tolbutamide hydroxylaseactivity was found to be 0.509±0.052 μ***-1.g-1 S9protein or 8.82±0.90 ***-1 CYP, but wasundetectable in parental CHL cell. In addition, we haveidentified a CYP2C18cDNA clone with exon 5 ***: The cDNA of human CYP2C18 wassuccessfully cloned and a cell line, CHL-CYP2C18, efficientlyexpressing the protein of CYP2C18, was established. Aspliced variant of CYP2C18 with exon 5 missing was identifiedin the cloning process.

关键词：转基因细胞 P450细胞色素第5外显子丢失 CYP2C18 鉴定克隆

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Stable expression of human cytochrome P450 2D6*10 in HepG2 cells

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World Journal of Gastroenterology 2004年第2期10卷 234-237页

作者： JianZhuge ying-nianyu Xiao-DanWu DepartmentofPathologyandPathophysiology SchoolofMedicineZhejiangUniversityHangzhou310031ZhejiangProvinceChina CenterofAnalysis ZhejiangUniversityHangzhou310031ZhejiangProvinceChina

AIM:Over 90% of drugs are metabolized by the cytochrome P-450 (CYP) family of liver *** most important enzymes are CYPIA2, 3A4,2C9/19,2D6 and *** CYP2D6 accounts for<2% of the total CYP liver enzyme content,it mediates metabolism in almost 25% of *** order to study its enzymatic activity for drug metabolism, its cDNA was cloned and a HepG2 cell line stably expressing CYP2D6 was ***:Human CYP2D6 cDNA was amplified with reverse transcription-polymerase chain reaction (RT-PCR) from total RNA extracted from human liver tissue and cloned into pGEM-T vector,cDNA segment was identified by DNA sequencing and subcloned into a mammalian expression vector pREP9.A cell line was established by transfecting the recombinant plasmid of pREP9-CYP2D6 to hepatoma HepG2 *** of mRNA was validated by *** activity of catalyzing dextromethorphan Odemethylation in postmitochondrial supernant (S9) fraction of the cells was determined by high performance liquid chromatography (HPLC).RESULTS:The cloned cDNA had 4 base differences, e.g.100 C→T, 336 T→C,408 C→G and 1 457 G→C,which resulted in P34S,and S486T amino acid substitutions, and two samesense mutations were 112F and 136V compared with that reported by Kimura et al (GenBank accession number: M33388).P34S and S486T amino acid substitutions were the characteristics of CYP2D6*10 *** relative activity of S9 fraction of HepG2-CYP2D6*10 metabolized detromethorphan O-demethylation was found to be 2.31±0.19nmol·min^-1·mg^-1 S9 protein (n=3),but was undetectable in parental HepG2 ***:cDNA of human CYP2D6*IOcan be successfully doned.A cell line,HepG2-CYP2D6*10,expressing CYP2D6*10 mRNA and having metabolic activity,has been established.

关键词： P-450细胞色素 HepG2细胞逆转录聚合酶链反应高效液相色谱法肝脏同功酶 DNA序列

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Response of human REV3gene to gastric cancer inducing carcinogen N-methyl-N'-nitro-N-nitrosoguanidine and its role in mutagenesis

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World Journal of Gastroenterology 2003年第5期9卷 888-893页

作者： FengZhu Cai-XiaJin JunYang LeiGuo ying-nianyu DepartmentofPathophysiology ZhejiangUniversitySchoolofMedicineHangzhou31003lZhejiangProvinceChina DepartmentofBiology NingxiaMedicalCollegeYingchuan750004NingxiaSelf-governingRegionChina

AIM: To understand the response of human REV3gene to gastric cancer inducing carcinogen N-methyl-N'-nitro-Nnitrosoguanidine (MNNG) and its role in human ***: The response of the human REV3 gene to MNNG was measured in human 293 cells and FL cells by RT-PCR. By using antisense technology, mutation analysis at HPRTlocus (on which lesion-targeted mutation usually occurs) was conducted in human transgenic cell line FLREV3- by 8-azaguanine screening, and mutation occurred on undamaged DNA template was detected by using a shuttle plasmid pZ189 as the probe in human transgenic cell lines 293-REV3- and FL-REV3-. The blockage effect of REV3 was measured by combination of reverse transcriptionpolymerase chain reaction to detect the expression of antisense REV3 RNA and Western blotting to detect the REV3protein ***: The human REV3 gene was significantly activated by MNNG treatment, as indicated by the upregulation of REV3 gene expression at the transcriptional level in MNNG-treated human cells, with significant increase of REV3 expression level by 0.38 fold, 0.33 fold and 0.27fold respectively at 6 h, 12 h and 24 h in MNNG-treated 293 cells (P＜0.05); and to 0.77 fold and 0.65 fold at 12 h and 24 h respectively in MNNG-treated FL cells (P＜0.05).In transgenic cell line (in which REV3 was blocked by antisense REV3 RNA), high level of antisense REV3 RNA was detected, with a decreased level of REV3 *** treatment significantly increased the mutation frequencies on undamaged DNA template (untargeted mutation), and also at HPRT locus (lesion-targeted mutation). However, when REV3 gene was blocked by antisense REV3 RNA, the MNNG-induced mutation frequency on undamaged DNA templates was significantly decreased by 3.8 fold (P＜0.05) and 5.8 fold (P＜0.01)respectively both in MNNG-pretreated transgenic 293 cells and FL cells in which REV3was blocked by antisense RNA,and almost recovered to their spontaneous mutation *** spontaneous HPRT mutation was

关键词： N-甲基-N'-硝基-N-亚硝基胍致癌物胃癌 REV3基因基因突变肿瘤生成作用机制

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ATM and ATR:Sensing DNA damage

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World Journal of Gastroenterology 2004年第2期10卷 155-160页

作者： JunYang Zheng-PingXu YunHuang HopeE.Hamrick PenelopeJ.Duerksen-Hughes ying-nianyu DepartmentofPathologyandPathophysiology andDepartmentofPublicHealthSchoolofMedicineZhejiangUniversityHangzhou310031ZhejiangProvinceChina DepartmentofPsychology WellesleyCollegeWellesleyMA02481.USA CenterforMolecularBiologyandGeneTherapy SchoolofMedicineLomaLindaUniversityLomaLindaCA92354USA

Cellular response to genotoxic stress is a very complex process, and it usually starts with the “sensing” or “detection” of the DNA damage,followed by a series of events that include signal transduction and activation of transcription *** activated transcription factors induce expressions of many genes which are involved in cellular functions such as DNA repair,cell cycle arrest,and cell death. There havebeen extensive studies from multiple disciplines exploring the mechanisms of cellular genotoxic responses, which have resulted in the identification of many cellular components involved in this process,including the mitogen-activated protein kinases (MAPKs) cascade. Although the initial activation of protein kinase cascade is not fully understood,human protein kinases ATM (ataxia-telangiectasia, mutated) and ATR (ATM and Rad3-related) are emerging as potential sensors of DNA damage. Current progresses in ATM/ATR research and related signaling pathways are discussed in this review, in an effort to facilitate a better understanding of genotoxic stress response.

关键词： DNA损伤遗传毒性信号转导促分裂蛋白激酶 p53基因细胞应答

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Genomic DNA hypomethylation could not be detected in the survivor cells after exposure to the genotoxic carcinogens

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癌变．畸变．突变 1993年第3期5卷 116-119页

作者： ying-nianyu

Enzymatic methylation of DNA in eukaryocytes is thought to be impli-cated in the regulation of gene *** of theregulatory control region of gene is in general correlated with itstranscriptional *** evidence of inapprop... 详细信息

关键词：基因毒致癌物存活细胞基因组DNA亚甲基化检测敏感性

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Interplay between creeping fat and gut microbiota: A brand-new perspective on fecal microbiota transplantation in Crohn's disease

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World Journal of Gastroenterology 2025年第2期31卷 138-143页

作者： ying Wang Jie Liu Department of Life Sciences and Medicine South District of Endoscopic CenterThe First Affiliated Hospital of University of Science and Technology of ChinaUniversity of Science and Technology of ChinaHefei 230001Anhui ProvinceChina Department of Gastroenterology The First Affiliated Hospital of University of Science and Technology of ChinaHefei 230001Anhui ProvinceChina

Inflammatory bowel disease,particularly Crohn's disease(CD),has been linked to modifications in mesenteric adipose tissue(MAT)and the phenomenon known as"creeping fat"(CrF).The presence of CrF is believed to serve as a predictor for early clinical recurrence following surgical intervention in patients with ***,the incorporation of the mesentery during ileocolic resection for CD has been correlated with a decrease in surgical recurrence,indicating the significant role of MAT in the pathogenesis of *** numerous studies have indicated that dysbiosis of the gut microbiota is a critical factor in the development of CD,the functional implications of translocated microbiota within the MAT of CD patients remain *** manuscript commentary discusses a recent basic research conducted by Wu et *** their study,intestinal bacteria from individuals were transplanted into CD model mice,revealing that fecal microbiota trans-plantation(FMT)from healthy donors alleviated CD symptoms,whereas FMT from CD patients exacerbated these ***,FMT was found to affect intestinal permeability,barrier function,and the levels of proinflammatory factors and ***,these findings suggest that targeting MAT and CrF may hold therapeutic potential for patients with ***,the study did not evaluate the composition of the intestinal microbiota of the donors or the subsequent alterations in the gut ***,the gut microbiota plays a crucial role in the histopathology of CD,and thus,targeting MAT and CrF may represent a promising avenue for treatment in this patient population.

关键词： Crohn's disease Mesenteric adipose tissue Creeping fat Fecal microbiota transplantation Gut microbiota

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Effects of Bifidobacterium lactis BLa80 on fecal and mucosal flora and stem cell factor/c-kit signaling pathway in simulated microgravity rats

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World Journal of Gastroenterology 2025年第1期31卷 93-109页

作者： Ping Zhang ying Zhu Pu Chen Tong Zhou Zhe-Yi Han Jun Xiao Jian-Feng Ma Wen Ma Peng Zang ying Chen Graduate School Hebei North UniversityZhangjiakou 075000Hebei ProvinceChina Key Laboratory of Aerospace Nutrition and Food Engineering China Astronaut Research and Training CenterBeijing 100094China Department of Gastroenterology The Air Force Medical CenterBeijing 100142China Department of Blood Transfusion Air Force Medical CenterBeijing 100142China

BACKGROUND Simulated microgravity environment can lead to gastrointestinal motility *** pathogenesis of gastrointestinal motility disorders is closely related to the stem cell factor(SCF)/c-kit signaling pathway associated with intestinal flora and Cajal stromal ***,intestinal flora can also affect the regulation of SCF/c-kit signaling pathway,thus affecting the expression of Cajal stromal *** cells are the pacemakers of gastrointestinal *** To investigate the effects of Bifidobacterium lactis(***)BLa80 on the intestinal flora of rats in simulated microgravity and on the gastrointestinal motility-related SCF/c-kit *** The internationally recognized tail suspension animal model was used to simulate the microgravity environment,and 30 rats were randomly divided into control group,tail suspension group and drug administration tail suspension group with 10 rats in each group for a total of 28 *** tail group was given *** BLa80 by intragastric administration,and the other two groups were given water intragastric administration,the concentration of intragastric administration was 0.1 g/mL,and each rat was 1 mL/***&eosin staining was used to observe the histopathological changes in each segment of the intestine of each group,and the expression levels of SCF,c-kit,extracellular signal-regulated kinase(ERK)and p-ERK in the gastric antrum of each group were detected by Western blotting and *** fecal flora and mucosal flora of rats in each group were detected by 16S *** Simulated microgravity resulted in severe exfoliation of villi of duodenum,jejunum and ileum in rats,marked damage,increased space between villi,loose arrangement,shortened columnar epithelium of colon,less folds,narrower mucosal thickness,reduced goblet cell number and crypts,and significant improvement after probiotic *** microgravity reduced the expressions of SCF and c-kit,and increased the express

关键词： Simulated microgravity Rat Intestinal flora Gastrointestinal motility Stem cell factor/c-kit signaling pathway

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NOX4 exacerbates Parkinson's disease pathology by promoting neuronal ferroptosis and neuroinflammation

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Neural Regeneration Research 2025年第7期20卷 2038-2052页

作者： Zhihao Lin Changzhou ying Xiaoli Si Naijia Xue Yi Liu Ran Zheng ying Chen Jiali Pu Baorong Zhang Department of Neurology The Second Affiliated HospitalZhejiang University School of MedicineHangzhouZhejiang ProvinceChina

Parkinson's disease is primarily caused by the loss of dopaminergic neurons in the substantia nigra ***,a novel form of regulated cell death characterized by iron accumulation and lipid peroxidation,plays a vital role in the death of dopaminergic ***,the molecular mechanisms underlying ferroptosis in dopaminergic neurons have not yet been completely *** oxidase 4 is related to oxidative stress,however,whether it regulates dopaminergic neuronal ferroptosis remains *** aim of this study was to determine whether NADPH oxidase 4 is involved in dopaminergic neuronal ferroptosis,and if so,by what *** found that the transcriptional regulator activating transcription factor 3 increased NADPH oxidase 4 expression in dopaminergic neurons and astrocytes in an 1-methyl-4-phenyl-1,2,3,6 tetrahydropyridine-induced Parkinson's disease *** oxidase 4 inhibition improved the behavioral impairments observed in the Parkinson's disease model animals and reduced the death of dopaminergic ***,NADPH oxidase 4 inhibition reduced lipid peroxidation and iron accumulation in the substantia nigra of the Parkinson's disease model ***,we found that NADPH oxidase 4 interacted with activated protein kinase Cαto prevent ferroptosis of dopaminergic ***,by lowering the astrocytic lipocalin-2 expression,NADPH oxidase 4 inhibition reduced 1-methyl-4-phenyl-1,2,3,6 tetrahydropyridine-induced *** findings demonstrate that NADPH oxidase 4 promotes ferroptosis of dopaminergic neurons and neuroinflammation,which contribute to dopaminergic neuron death,suggesting that NADPH oxidase 4 is a possible therapeutic target for Parkinson's disease.

关键词： dopaminergic neuron ferroptosis NADPH oxidase 4(NOX4) neuroinflammation Parkinson's disease

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Multiple liver metastases of unknown origin:A case report

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World Journal of Gastrointestinal Oncology 2025年第1期17卷 233-238页

作者： ying-Jin Wang Ze-Chuan Liu Jian Wang Yin-Mo Yang Department of Hepatobiliary Pancreatic Surgery Peking University First HospitalBeijing 100034China Department of Interventional and Vascular Surgery Peking University First HospitalBeijing 100034China

BACKGROUND The liver is the most common site of digestive system tumor metastasis,but not all liver metastases can be traced back to the primary *** it is unusual,syphilis can impact the liver,manifesting as syphilitic hepatitis with inflammatory nodules,which might be misdiagnosed as *** SUMMARY This case report involves a 46-year-old female who developed right upper abdominal pain and intermittent low fever that persisted for more than three *** definitive diagnosis of a tumor had been made in the past decades,but signs of multiple liver metastases were recognized after a computed tomo-graphy scan without evidence of primary *** positive serological tests for syphilis and a biopsy of the liver nodules,a diagnosis of hepatic syphilis was made and confirmed with follow-up nodule reduction after anti-syphilis *** Clinicians must be aware of the possibility that syphilis can cause hepatic inflam-matory masses,especially when liver metastasis is suspected without evidence of primary lesions.A definitive diagnosis should be established in conjunction with a review of the patient’s medical history for accurate therapeutic intervention.

关键词： Syphilitic hepatitis Liver nodules Liver metastasis Treponema pallidum Case report

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