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.

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.

Detections of mefA, ermB, and mphA Macrolides Resistant Genes in Bacteria Isolated from Covid-19 Patients from Selected Health Facilities in Ibadan, Nigeria

Background: COVID-19 is a disease caused by Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2). Epidemiological data indicated that bacterial complications in COVID-19 would decrease clearance rate of the infecting agent and increase mortality rate. Macrolides such as Azithromycin are usually administered to COVID-19 patients as palliative treatments. Currently, a considerable number of bacterial strains have developed resistance to various antibiotics, especially macrolides. Resistance is reported to be due to possession of mefA, ermB, and mphA genes by Gram positive and Gram negative bacteria. Therefore, this study determined antibiotic resistance patterns and identify mefA, ermB and mphA macrolide-resistant genes in bacterial pathogens isolated from COVID-19 cases in Ibadan, Nigeria. Methods: 400 Nasopharyngeal samples were collected from symptomatic cases before antibiotic medication;structured questionnaires were administered to collect socio-demographic data of participants. Samples were cultured on Blood, Chocolate, MacConkey and Mannitol salt agar at 37°C for 48 hrs. Bacterial identification was performed using VITEK 2.0 ID cards and API 20E for Gram positive and negative bacteria respectively. Antibiotic Susceptibility Testing was performed using Kirby Bauer disc diffusion methods and VITEK 2.0 AST card kits. DNA of multidrug resistant bacterial isolates was extracted;resistant genes were determined using a polymerase chain reaction with specific primers. Amplified genes were detected using agarose gel electrophoresis. Results: 240 (60%) had bacterial growth and 97 (22.2%) yielded no growth. From the 240 bacterial isolates, 38 (15.83%) were multi-drug resistant including resistance to macrolides (Azithromycin) 20 (52.63%) of which were positive for either mefA or ermB, and none (0.0%) possess mphA gene;14 (36.8%) isolates had mefA gene, 10 (26.3%) isolates carried ermB gene. Conclusion: Multi-drug bacterial resistance including macrolides and quinolones was detected. Only mefA and ermB genes were detected in the bacterial isolates, especially in Gram positive organisms. The detection of mefA and ermB genes in the MDR bacterial isolates raised concern on the use of azithromycin as palliative treatment for COVID-19 symptomatic patients.

Trifunctional Cu-Mesh/Cu_(2)O@FeO Nanoarrays for Highly Efficient Degradation of Antibiotic, Inactivation of Antibiotic-Resistant Bacteria, and Damage of Antibiotics Resistance Genes

Trifunctional Cu-mesh/Cu_(2)O@FeO nanoarrays heterostructure is designed and fabricated by integrating CuCu_(2)O@FeO nanoarrays onto Cu-mesh(CM)via an in situ growth and phase transformation process.It is successfully applied to efficiently mitigate the antibiotic pollution,including degradation of antibiotics,inactivation of antibiotic-resistant bacteria(ARB),and damage of antibiotics resistance genes(ARGs).Under visible-light irradiation,CM/CuCu_(2)O@FeO nanoarrays exhibit a superior degradation efficiency on antibiotics(e.g.,up to 99%in 25 min for tetracycline hydrochloride,TC),due to the generated reactive oxygen species(ROS),especially the dominant·O^(2−).It can fully inactivate E.coli(HB101)with initial number of~108 CFU mL^(−1) in 10 min,which is mainly attributed to the synergistic effects of 1D nanostructure,dissolved metal ions,and generated ROS.Meanwhile,it is able to damage ARGs after 180 min of photodegradation,including tetA(vs TC)of 3.3 log 10,aphA(vs kanamycin sulfate,KAN)of 3.4 log 10,and tnpA(vs ampicillin,AMP)of 4.4 log 10,respectively.This work explores a green way for treating antibiotic pollution under visible light.

Identification of key genes and biological pathways in lung adenocarcinoma by integrated bioinformatics analysis

BACKGROUND The objectives of this study were to identify hub genes and biological pathways involved in lung adenocarcinoma(LUAD)via bioinformatics analysis,and investigate potential therapeutic targets.AIM To determine reliable prognostic biomarkers for early diagnosis and treatment of LUAD.METHODS To identify potential therapeutic targets for LUAD,two microarray datasets derived from the Gene Expression Omnibus(GEO)database were analyzed,GSE3116959 and GSE118370.Differentially expressed genes(DEGs)in LUAD and normal tissues were identified using the GEO2R tool.The Hiplot database was then used to generate a volcanic map of the DEGs.Weighted gene co-expression network analysis was conducted to cluster the genes in GSE116959 and GSE-118370 into different modules,and identify immune genes shared between them.A protein-protein interaction network was established using the Search Tool for the Retrieval of Interacting Genes database,then the CytoNCA and CytoHubba components of Cytoscape software were used to visualize the genes.Hub genes with high scores and co-expression were identified,and the Database for Annotation,Visualization and Integrated Discovery was used to perform enrichment analysis of these genes.The diagnostic and prognostic values of the hub genes were calculated using receiver operating characteristic curves and Kaplan-Meier survival analysis,and gene-set enrichment analysis was conducted.The University of Alabama at Birmingham Cancer data analysis portal was used to analyze relationships between the hub genes and normal specimens,as well as their expression during tumor progression.Lastly,validation of protein expression was conducted on the identified hub genes via the Human Protein Atlas database.RESULTS Three hub genes with high connectivity were identified;cellular retinoic acid binding protein 2(CRABP2),matrix metallopeptidase 12(MMP12),and DNA topoisomerase II alpha(TOP2A).High expression of these genes was associated with a poor LUAD prognosis,and the genes exhibited high diagnostic value.CONCLUSION Expression levels of CRABP2,MMP12,and TOP2A in LUAD were higher than those in normal lung tissue.This observation has diagnostic value,and is linked to poor LUAD prognosis.These genes may be biomarkers and therapeutic targets in LUAD,but further research is warranted to investigate their usefulness in these respects.

Role of IL-2/IL-2 receptor in pathogenesis of autoimmune disorders:Genetic and therapeutic aspects

Interleukin-2(IL-2)is an important cytokine that plays a key role in the immune response.The IL-2 receptor(IL-2R)is composed of three subunits,alpha,beta,and gamma,with the alpha subunit having the highest affinity for IL-2.Several studies reported that immune dysregulation of IL-2 may cause tissue injury as well as damage leading to the pathogenesis of various autoimmune diseases such as acute necrotizing vasculitis in systemic lupus erythematosus(SLE),inflammatory synovitis in rheumatoid arthritis(RA),salivary and lacrimal gland dysfunction in Sjogren syndrome(SS),obliterative vasculopathy fibrosis in systemic sclerosis(SSc),and inflammatory demyelination in multiple sclerosis(MS).The aim of this review paper was to examine the role of IL-2/IL-2R in various autoimmune disorders,taking into account recent advancements and discoveries,gaps in the current literature,ongoing debates,and potential avenues for future research.The focus of this review is on systemic lupus erythematosus,rheumatoid arthritis,systemic sclerosis,sjogren syndrome,and multiple sclerosis,which are all linked to the malfunctioning of IL-2/IL-2R.In genetic studies,gene polymorphisms of IL-2 such as IL-2330/T,IL-2330/G,and rs2069763 are involved in increasing the risk of SLE.Furthermore,genetic associations of IL-2/IL-2R such as rs791588,rs2281089,rs2104286,rs11594656,and rs35285258 are significantly associated with RA susceptibility.The IL-2 polymorphism including rs2069762A,rs6822844T,rs6835457G,and rs907715T are significant connections with systemic sclerosis.In addition,rs2104286(IL-2),rs11594656(IL-2RA),rs35285258(IL-2RB)gene polymorphism significant increases the risk of multiple sclerosis.In therapeutic approaches,low-dose IL-2 therapy could regulate Tfr and Tfh cells,resulting in a reduction in disease activity in the SLE patients.In addition,elevated sIL-2R levels in the peripheral blood of SLE patients could be linked to an immunoregulatory imbalance,which may contribute to the onset and progression of SLE.Consequently,sIL-2R could potentially be a target for future SLE therapy.Moreover,Low dose-IL2 was well-tolerated,and low levels of Treg and high levels of IL-21 wereassociated with positive responses to Ld-IL2 suggested to be a safe and effective treatment for RA.Additionally,low-dose IL-2 treatment improves the exocrine glands'ability to secrete saliva in SS-affected mice.Whereas,Basiliximab targets the alpha chain of the IL-2 receptor suggested as a potential treatment for SSc.Also,pre-andpost-treatment with Tregs,MDSCs,and IL-2 may have the potential to prevent EAE induction in patients with MS.It is suggested that further studies should be conducted on IL-2 polymorphism in Sjogren syndrome.

Role of p53 suppression in the pathogenesis of hepatocellular carcinoma

In the world,hepatocellular carcinoma(HCC)is among the top 10 most prevalent malignancies.HCC formation has indeed been linked to numerous etiological factors,including alcohol usage,hepatitis viruses and liver cirrhosis.Among the most prevalent defects in a wide range of tumours,notably HCC,is the silencing of the p53 tumour suppressor gene.The control of the cell cycle and the preservation of gene function are both critically important functions of p53.In order to pinpoint the core mechanisms of HCC and find more efficient treatments,molecular research employing HCC tissues has been the main focus.Stimulated p53 triggers necessary reactions that achieve cell cycle arrest,genetic stability,DNA repair and the elimination of DNA-damaged cells’responses to biological stressors(like oncogenes or DNA damage).To the contrary hand,the oncogene protein of the murine double minute 2(MDM2)is a significant biological inhibitor of p53.MDM2 causes p53 protein degradation,which in turn adversely controls p53 function.Despite carrying wt-p53,the majority of HCCs show abnormalities in the p53-expressed apoptotic pathway.High p53 in-vivo expression might have two clinical impacts on HCC:(1)Increased levels of exogenous p53 protein cause tumour cells to undergo apoptosis by preventing cell growth through a number of biological pathways;and(2)Exogenous p53 makes HCC susceptible to various anticancer drugs.This review describes the functions and primary mechanisms of p53 in pathological mechanism,chemoresistance and therapeutic mechanisms of HCC.

Identification and Expression Analysis of Abscisic Acid Signal Transduction Genes in Hemp Seeds

Abscisic acid(ABA)is involved in regulating diverse biological processes,but its signal transduction genes and roles in hemp seed germination are not well known.Here,the ABA signaling pathway members,PYL,PP2C and SnRK2 gene families,were identified from the hemp reference genome,including 7 CsPYL(pyrab-actin resistance1-like,ABA receptor),8 CsPP2CA(group A protein phosphatase 2c),and 7 CsSnRK2(sucrose nonfermenting1-related protein kinase 2).The content of ABA in hemp seeds in germination stage is lower than that in non-germination stage.Exogenous ABA(1 or 10μM)treatment had a significant regulatory effect on the selected PYL,PP2C,SnRK2 gene families.CsAHG3 and CsHAI1 were most significantly affected by exogenous ABA treatment.Yeast two-hybrid experiments were performed to reveal that CsPYL5,CsSnRK2.2,and CsSnRK2.3 could interact with CsPP2CA7 and demonstrate that this interaction was ABA-independent.Our results indicated that CsPYL5,CsSnRK2.2,CsSnRK2.3 and CsPP2CA7 might involve in the ABA signaling transduction pathway of hemp seeds during the hemp seed germination stages.This study suggested that novel genetic views can be brought into investigation of ABA signaling pathway in hemp seeds and lay the foundation for further exploration of the mechanism of hemp seed germination.

Study of pathogenic genes in a pedigree with familial dilated cardiomyopathy

BACKGROUND Dilated cardiomyopathy(DCM)is a genetically heterogeneous cardiac disorder characterized by left ventricular dilation and contractile dysfunction.The substantial genetic heterogeneity evident in patients with DCM contributes to variable disease severity and complicates overall prognosis,which can be very poor.AIM To identify pathogenic genes in DCM through pedigree analysis.METHODS Our research team identified a patient with DCM in the clinic.Through invest-igation,we found that the family of this patient has a typical DCM pedigree.High-throughput sequencing technology,next-generation sequencing,was used to sequence the whole exomes of seven samples in the pedigree.RESULTS A novel and potentially pathogenic gene mutation-ANK2p.F3067L-was discovered.The mutation was completely consistent with the clinical information for this DCM pedigree.Sanger sequencing was used to further verify the locus of the mutation in pedigree samples.These results were consistent with those of high-throughput sequencing.CONCLUSIONS ANK2p.F3067L is considered a novel and potentially pathogenic gene mutation in DCM.

AT2R基因在体转染抑制大鼠颈动脉新生内膜增生

目的:探讨AngⅡ2型受体(AT2R)基因在体转染对大鼠颈动脉球囊损伤后新生内膜增生的抑制作用。方法:大鼠颈动脉球囊损伤后,局部转染AT2R重组腺病毒载体(pAdCMV/AT2R)或空病毒载体(pAd-GFP),于术后7、14和21 d用RT-PCR、免疫组织化学及HE染色方法,进行AT2R、AngⅡ1型受体(AT1R)、PCNA在颈动脉壁中表达的变化及定量组织形态学分析;免疫荧光双标染色和激光共聚焦技术检测血管中AT2R与PC-NA表达的关系。结果:pAdCMV/AT2R转染后,大鼠颈动脉AT2R的表达水平显著高于未转染组和pAd-GFP组(P<0.01),21 d时仍维持较强表达。在14 d时pAdCMV/AT2R组PCNA阳性表达率显著低于未转染组和pAd-GFP组[(27.29±5.81)%vs(72.25±4.47)%、(68.43±9.12)%,P<0.01],在AT2R表达阳性的部位PCNA表达阴性。在21 d时,pAdCMV/AT2R组的内膜面积与中膜面积比显著低于未转染组和pAd-GFP组(0.78±0.06vs1.44±0.22、1.36±0.21,P<0.01),pAd-GFP组和未转染组间无显著差异(P>0.05);各组颈动脉AT1R表达水平无显著差异(P>0.05)。结论:AT2R基因在体转染可抑制球囊损伤后大鼠颈动脉平滑肌细胞增殖和新生内膜增生,AT2R基因转染后表达并发挥其生物学作用时,AT1R和AT2R之间不存在表达量上此起彼伏的关系,可能是建立在信号转导基础上的功能调节关系。

强力霉素可调控的双重稳定表达AT2R的骨髓间充质干细胞的建立

目的构建四环素类抗生素可调控的双重稳定表达AT2R的骨髓间充质干细胞。方法将四环素可调控系统（Tetracycline-on，Tet-on）的4种质粒用脂质体转染法连续两个回合转染体外培养的大鼠骨髓间充质干细胞并抗性筛选，分别采用发光计检测不同细胞克隆荧光素酶活性改变以及RT-PCR法检测AT2R目的基因表达情况，根据各个细胞克隆受强力霉素调控表达的程度，筛选出高诱导、低背景表达目的基因AT2R的骨髓间充质细胞系，并检测在不同浓度（0～10^4ng／ml）强力霉素干预下以及转染后不同时间（0～8周）目的基因的蛋白表达情况。结果连续两个回合的转染及筛选后获得的引入Tet-on系统的细胞系，高诱导低背景表达AT2R，在强力霉素诱导下48h内可以使AT2R表达显著增加，强力霉素在一定浓度范围内可以诱导AT2R的表达呈剂量依赖性的增加，且至少在8周内保持稳定。结论构建的含四环素调控系统的骨髓间充质干细胞系诱导活性可靠，使外源AT2R基因的表达处于有效的主动控制下。

AT2R基因在体电穿孔转染抑制大鼠颈总动脉新生内膜增生

目的探讨AngⅡ2型受体(AT2R)基因局部电穿孔转染对大鼠颈总动脉球囊损伤后新生内膜增生的影响。方法建立大鼠颈总动脉球囊损伤模型,用局部电穿孔方法转染AT2R真核表达载体(pEGFP-AT2R)或空载体(pEGFP-N2),分别于术后7、14 d和21 d采用HE染色及RT-PCR方法进行AT2R、AngⅡ1型受体(AT1R)在颈动脉壁中表达的变化和组织形态学分析,检测其对在体血管新生内膜的影响作用。结果球囊损伤21 d后,pEGFP-AT2R组大鼠颈动脉AT2R mRNA表达为(1.262±0.317),pEGFP-N2组为(0.396±0.100),单纯损伤组为(0.410±0.053),pEGFP-AT2R组与pEGFP-N2组和单纯损伤组相比差异有统计学意义(P<0.01);球囊损伤后21 d时AT1R的表达,pEGFP-AT2R组为(0.469±0.065)、pEGFP-N2组为(0.363±0.046)、单纯损伤组为(0.373±0.045),pEGFP-AT2R组较pEGFP-N2组和单纯损伤组相比差异有统计学意义(P<0.05),pEGFP-N2组和单纯损伤组间无统计学差异(P>0.05);球囊损伤后21 d,pEGFP-AT2R组的内膜面积与中膜面积比(I/M)为(0.828±0.101),pEGFP-N2组为(1.432±0.086),单纯损伤组为(1.515±0.078),pEGFP-AT2R组较pEGFP-N2组和单纯损伤组相比差异有统计学意义(P<0.01)。pEGFP-AT2R组与单纯损伤组相比,使实验动物新生内膜增生平均受抑制率达45.35%。结论在体血管局部电穿孔转染AT2R基因可使AT2R在损伤血管组织表达较单纯损伤组明显增加。AT2R基因与AT1R基因之间在表达量上不存在此消彼长的关系。

AT2R基因可调控表达对体外培养VSMC纤维黏连蛋白表达的影响

目的利用Dox-on可调控哺乳动物表达系统,建立起了受四环素类似物Doxycycline(Dox)紧密调控、表达AT2R基因的双重稳定血管平滑肌细胞(vascular smooth muscle cells,VSMC),在此基础上对纤维黏连蛋白(fibronectin,FN)的表达受AngⅡ及其受体拮抗剂的影响进行研究。方法建立Dox可调控表达AT2R基因的双重稳定大鼠VSMC细胞,观察该VSMC细胞中AT2R受调控表达情况,以及血管紧张素Ⅱ(angiotensinⅡ,AngⅡ)及其1型、2型受体拮抗剂干预上述细胞后FN的mRNA及蛋白表达情况变化。结果Dox-on可调控哺乳动物表达系统可成功介导AT2R基因在原代培养大鼠主动脉VSMC的表达,该表达受到Dox给予/去除的紧密调控;Dox干预可在48h内迅速诱导该VSMC细胞表达AT2R,AT2R表达在Dox干预后72h进一步增强(P<0.01)。AT2R基因的可调控表达抑制由于AngⅡ干预VSMC后引起FN表达的增强(P<0.01)。这一作用被AT1R拮抗剂CV-11974进一步增强(P<0.01);而被加入AT2R拮抗剂干预而取消;同时给予AT1R拮抗剂和AT2R拮抗剂时FN的表达与基础状态时的情况一致。结论AngⅡ干预增强FN的表达,该作用是通过AT1R介导的;经Dox诱导表达AT2R基因可以明显抑制这一生物学作用,说明在这一生物学效应上,AT2R具有与AT1R相拮抗的生物学功能。

AT1R、AT2R、RAGE在糖尿病大鼠脑组织中的表达及意义

目的探讨血管紧张素Ⅱ受体-1(AT1R)、血管紧张素Ⅱ受体-2(AT2R)、高级糖基化终末产物受体(RAGE)在糖尿病大鼠脑组织损伤中的可能作用。方法 40只SD大鼠随机均分为糖尿病组和对照组。链脲佐菌素注射后12周末处死大鼠,留取脑组织标本,行HE染色观察其病理学改变,采用免疫组化Evision二步法检测脑组织中AT1R、AT2R、RAGE及基质金属酶9(MMP9)的表达,并将糖尿病组AT1R、AT2R、RAGE、MMP9进行相关性分析。结果糖尿病组较对照组大鼠脑组织病理学显示脑小血管周隙明显增宽,但两组脑组织均无炎细胞浸润;糖尿病组大鼠脑组织AT1R、AT2R、RAGE、MMP9阳性细胞数较对照组明显升高,差异有统计学意义(P<0.05);糖尿病组大鼠AT2R与MMP9、RAGE、AT1R均呈正相关(P<0.05),AT1R与MMP9、RAGE呈正相关(P<0.05),MMP9与RAGE呈正相关(P=0.001)。结论糖尿病大鼠脑组织存在水肿,提示有血脑屏障异常,其机制可能与AT1R、RAGE、MMP9的过度表达或AT2R的激活以及它们之间的相互作用有关。

AT2R转染表达促进人肾间质纤维母细胞凋亡

目的 探讨血管紧张素Ⅱ (AngⅡ ) 2型受体 (AT2R)基因表达对人肾间质纤维母细胞凋亡的影响。方法 构建带AT2R基因的重组复制缺陷型腺病毒载体 (AdCMV -AT2R) ,转染培养的人肾间质纤维母细胞 ,用流式细胞仪检测AT2R细胞表达率 ,RT -PCR方法检测AT2R ,bcl -2和baxmRNA表达 ,肾间质纤维母细胞凋亡用流式细胞仪、原位末端标记法检测。结果 构建的AdCMV -AT2R转染培养肾间质纤维母细胞表达率为 90 6 %。AT2R峰值表达时 ,其凋亡发生率较未转染组增加 3 2倍 (P <0 0 1) ,bax表达增加 76 3 % (P <0 0 5 ) ,bcl-2则无明显变化 ,TUNEL检测结果表明转染组出现大量凋亡细胞。结论 AT2R转染表达可显著增加体外培养肾间质纤维母细胞的凋亡 ,这对延缓肾间质纤维化是有益的。

AT2R基因可调控表达在血管紧张素Ⅱ介导的体外培养VSMC增殖中作用

目的 研究血管紧张素Ⅱ 2型受体 (AT2R)基因可调控表达对体外培养血管平滑肌细胞(VSMC)表达细胞周期蛋白依赖性激酶 2 (CDK2 )、增殖细胞核抗原 (PCNA)以及 p2 1的影响 ,旨在探讨AT2R基因在VSMC上条件表达在再狭窄防治中的潜在意义。方法 本研究通过常规分子生物学方法 ,建立四环素类似物Doxycycline可调控表达AT2 R基因的双重稳定VSMC细胞系。应用RT PCR和免疫细胞化学染色技术 ,观察该双重稳定表达AT2R的VSMC细胞系中CDK2、PCNA及 p2 1的mRNA及蛋白表达情况 ;采用血管紧张素Ⅱ (AngⅡ )及其Ⅰ型、Ⅱ型其受体拮抗剂干预上述细胞 ,观测上述指标的变化。 结果 ①加入Doxycycline前转染组低水平表达CDK2、PCNA ;AngⅡ 10 - 7mol/L干预VSMC后CDK2、PCNA表达显著增加 (P <0 .0 1) ;在AngⅡ 10 - 7mol/L干预的同时加入Doxycycline后 ,CDK2、PCNA的表达受到显著抑制 (P <0 .0 1) ,但仍高于基础水平 (P <0 .0 1) ;ATIR拮抗剂CV 11974可进一步下调CDK2、PCNA的表达 ;AT2R拮抗剂PD12 3319干预组CDK2、PCNA强表达 (与AngⅡ组比较 ,P >0 .0 5 )。②加入Doxycy cline前转染组 p2 1表达处于高水平 ;AngⅡ 10 - 7mol/L干预VSMC后 p2 1表达显著降低 (P <0 .0 1) ;在AngⅡ 10 - 7mol/L干预的同时加入Doxycycline后 。

AT2R基因可调控表达对体外培养VSMC基质金属蛋白酶2表达的影响

目的 研究血管紧张素Ⅱ 2型受体 (AT2R)基因可调控表达对体外培养血管平滑肌细胞(VSMC)基质金属蛋白酶 2形成的影响 ,旨在探讨AT2R基因在VSMC上条件表达在再狭窄防治中的潜在意义。方法 本研究通过常规分子生物学方法 ,建立四环素类似物Doxycycline可调控表达AT2 R基因的双重稳定VSMC细胞系。应用RT PCR和免疫细胞化学染色技术 ,观察该双重稳定表达AT2R的VSMC细胞系中基质金属蛋白酶 2的mRNA及蛋白表达情况 ;采用血管紧张素Ⅱ (AngⅡ )及其Ⅰ型、Ⅱ型其受体拮抗剂干预上述细胞 ,观测上述指标的变化。结果 Doxycycline干预前 ,AT2R的表达处于静止状态 ,通过Doxycycline(1μg/ml)的干预 ,可以使该VSMC系稳定表达AT2R。加入Doxycycline前转染组低水平基质金属蛋白酶 2 ;AngⅡ 10 - 7mol/L干预VSMC后基质金属蛋白酶 2的mRNA及蛋白表达均显著增加 (P <0 .0 1) ;在AngⅡ干预的同时加入Doxycycline后 ,基质金属蛋白酶 2的mRNA及蛋白表达受到显著抑制 (P<0 .0 1) ;ATIR拮抗剂CV 11974可进一步下调上述mRNA及蛋白的表达 ;AT2R拮抗剂PD12 3319干预组基质金属蛋白酶 2mRNA及蛋白仍旧强表达 (与AngⅡ组比较 ,P >0 .0 5 )。 结论 AngⅡ干预VSMC后通过ATIR介导引起基质金属蛋白酶 2表达增强 ;

AT2R转染表达抑制人肾间质成纤维细胞迁移活性

目的 探讨血管紧张素Ⅱ (AngⅡ ) 2型受体 (AT2R)基因表达对人肾间质成纤维细胞 (hRIF)迁移活性的影响。方法 构建带AT2R基因的重组复制型腺病毒载体 (AdCMV -AT2R) ,转染培养的hRIF ,用流式细胞仪检测AT2R细胞表达率 ,RT -PCR方法检测AT2RmRNA表达。用改良Boyden’s趋化小室和激光共聚焦显微镜检测hRIF迁移活性。结果 构建的AdCMV -AT2R转染培养hRIF表达率为 90 6%。改良Boyden’s趋化小室检测表明 ,迁移细胞数抑制比例最高可达 62 2 % (P <0 0 1) ,激光共聚焦显微镜检测表明 ,转染组F -actin表达明显受抑制。结论 AT2R转染表达可显著抑制hRIF迁移活性 。

VS MC转染表达AT2R对AT1R表达的影响

目的 探讨血管平滑肌细胞 (VSMC)转染表达血管紧张素Ⅱ (AngⅡ ) 2型受体 (AT2R)后其 1型受体 (AT1R)表达所受的影响。方法 用同源重建方法构建带AT2R基因的重组复制缺陷型腺病毒载体(AdCMV AT2R) ,体外转染VSMC ,分别用流式细胞仪检测AT1R、AT2R细胞转染表达率、免疫组织化学法和免疫荧光法检测其膜表达、RT PCR法和蛋白印迹法检测其mRNA和蛋白表达。结果 AdCMV AT2R转染后 ,在不同浓度AngⅡ刺激下AT1R、AT2R在VSMC的细胞转染表达率如下表。如果表明随着转染表达时间延长 ,AT2R细胞表达率呈显著增加趋势 ,4 8小时最高表达率达 89.5 1% ,AngⅡ作用与否及不同浓度AngⅡ作用对AT2R表达无显著影响。而转染前后AT1R表达相对较稳定 ,受不同浓度AngⅡ作用 ,其表达明显呈增加趋势。AT2R峰值表达时 ,免疫组织化学法和免疫荧光法检测其膜表达结果也提示AT2R表达随转染表达时间延长显著增加 ,转染前后AT1R表达无明显变化 ,在一定浓度范围内 ,AngⅡ刺激对AT2R表达无显著影响 ,却显著增加AT1R的膜表达。RT PCR法和蛋白印迹法检测AT1R和AT2R的mRNA和蛋白表达结果与其细胞表达率和膜表达的检测结果相一致。结论 生理状态下 ,VSMC主要表达AT1R ,AT2R表达较少或不表达。AT1R表达受一系列因素影响 。