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1.
培养上皮性卵巢癌细胞H08910PM、H08910及正常卵巢上皮细胞HOSEA 3种细胞.对3种细胞分别采用免疫细胞化学法检测RhoA的表达、Transwell小室体外侵袭试验测定体外侵袭能力、与人脐静脉内皮细胞HUVEC建立细胞共培养系统测定血管形成能力.结果表明,RhoA在H08910PM细胞中表达最强,在H08910中次之,在HOSEA中最弱;3种细胞的侵袭能力以H08910PM最强,H08910较弱,HOSEA无侵袭能力;H08910PM的血管形成能力最强,H08910其次,HOSEA无血管形成能力(P<0.01).Pearson相关分析结果显示,RhoA表达水平分别与细胞侵袭能力及血管形成能力呈正相关(P<0.05).RhoA在上皮性卵巢癌细胞中表达水平越高,细胞体外侵袭及血管形成能力随之更强.RhoA可能作为重要因子参与上皮性卵巢癌的侵袭转移过程. 相似文献
2.
《现代生物医学进展》2017,(34)
目的:探讨dbp A蛋白在卵巢癌组织中的表达及其对卵巢癌细胞增殖的影响。方法:通过荧光定量和蛋白质免疫印迹方法检测临床卵巢癌组织和正常癌旁组织中dbp A的表达量;设计合成针对dbp A基因的双链小干扰RNA转染人卵巢癌细胞系SKOV3和A2780细胞,用荧光定量和蛋白质免疫印迹方法检测细胞中dbp A的表达量,MTT法和克隆形成试验检测细胞的增殖能力和克隆形成能力,流式细胞术检测各组细胞周期和细胞凋亡的变化。结果:dbp A在卵巢癌组织、SKOV3和A2780细胞中表达较癌旁正常卵巢组织显著升高。沉默dbp A后,SKOV3和A2780细胞中dbp A蛋白表达量显著降低,SKOV3和A2780细胞增殖能力和克隆形成能力显著下降(P0.05),G0/G1期细胞百分比显著增加(P0.01),S期细胞百分比明显减少(P0.05),细胞凋亡率显著升高(P0.01)。结论:dbp A在卵巢癌组织和卵巢癌细胞SKOV3和A2780中过表达,沉默dbp A基因后可抑制SKOV3和A2780细胞的增殖能力和克隆形成能力。 相似文献
3.
目的:明确Cosmc基因在卵巢癌细胞中的生物学功能。方法:本研究采用慢病毒转染技术,在卵巢癌细胞A2780和SKOV3中过表达Cosmc基因,MTT实验、细胞凋亡实验以及Transwell侵袭实验对过表达Cosmc后卵巢癌细胞在生长、凋亡、侵袭等方面的影响。结果:慢病毒转染卵巢癌细胞A2780和SKOV-3后,Cosmc的蛋白表达显著提高;MTT结果显示,与空载体对照组相比,过表达Cosmc的卵巢癌细胞生长能力显著减弱;流式细胞术结果表明Cosmc过表达的卵巢癌细胞凋亡数较空载体对照组细胞显著增加;Transwell实验显示Cosmc过表达的细胞侵袭和迁移能力较空载体对照组显著减少。结论:卵巢癌细胞系中过表达Cosmc基因能抑制卵巢癌细胞的生长和侵袭并促进细胞凋亡。 相似文献
4.
目的:探讨TGF-β1对卵巢癌细胞A2780增殖、迁移及侵袭能力的影响。方法:以体外培养的卵巢癌细胞A2780为研究对象,给予不同浓度(0、2、4…20 ng/m L)TGF-β1处理不同时间(12、24…72 h)。采用CCK-8法检测不同的浓度TGF-β1处理不同时间对卵巢癌细胞A2780增殖的影响。根据增殖实验结果选择合适的TGF-β1作用浓度及处理时间,采用细胞划痕实验测定细胞的迁移能力,Transwell实验检测细胞的侵袭及迁移能力。结果:相较于空白对照组,TGF-β1可以剂量和时间依赖性显著促进卵巢癌细胞A2780的增殖(P0.05)。细胞划痕实验结果显示TGF-β1处理组ΔS%/h明显高于空白对照组(P0.05);Transwell迁移实验结果显示:TGF-β1处理组OD570明显高于空白对照组,差异具有统计学意义(P0.05)。Transwell细胞侵袭实验结果显示:与空白对照组相比,TGF-β1处理组OD570明显升高(P0.05)。结论:TGF-β1可以明显促进卵巢癌细胞系A2780的增殖、迁移及侵袭能力,其促增殖效应呈剂量/时间依赖效应。 相似文献
5.
《中国细胞生物学学报》2019,(11)
该文探讨了幽门螺杆菌(Helicobacter pylori, HP)炎症微环境对结肠癌SW620细胞发生上皮-间质转化(epithelial mesenchymal transformation, EMT)的影响。使用ELISA方法检测HP干预U937细胞后上清中炎症因子MIF、IL-1β等的变化;采用RT-PCR方法检测MIF、IL-1β、NF-κB基因水平的变化;细胞划痕实验检测炎症上清处理SW620细胞后SW620细胞的侵袭迁移能力; CCK-8方法检测HP处理U937细胞后炎症上清对SW620细胞增殖活性的影响;免疫荧光实验检测炎症上清处理SW620细胞后SW620细胞EMT相关蛋白Vimentin的荧光变化; Western blot实验检测脂多糖(LPS)上清干预SW620细胞后EMT相关蛋白Vimentin、E-cadherin和N-cadherin的变化; Western blot实验检测HP干预U937细胞后NF-κB蛋白,炎症上清处理SW620细胞后EMT相关蛋白Vimentin、E-cadherin和N-cadherin的变化。结果显示, HP干预U937细胞后引起巨噬细胞移动抑制因子(MIF)、白细胞介素-1β(IL-1β)和NF-κB等相关炎症因子及基因表达升高; CCK-8结果表明HP干预U937细胞后, U937细胞毒性增强;划痕实验结果表明在12 h、24 h、36 h时,随着时间点的推移, SW620细胞的侵袭迁移能力有所增强;免疫荧光实验结果显示SW620细胞Vimentin绿色点状聚集显著增加; Western blot结果显示在16 h、24 h时间点U937细胞NF-κB蛋白表达明显升高; LPS上清干预SW620细胞后Vimentin蛋白表达无明显变化,N-cadherin蛋白24 h组表达减少, E-cadherin蛋白增多, HP炎症微环境上清干预SW620细胞后12 h、24 h、36 h时,随着时间点推移SW620细胞Vimentin蛋白表达显著增加, E-cadherin和N-cadherin蛋白表达显著减少。该实验研究结果表明,幽门螺杆菌致炎症微环境可以促进SW620细胞发生EMT。 相似文献
6.
目的:探讨人卵巢癌细胞A2780中的HES1表达与人卵巢癌侵袭及转移的关系。方法:分别运用实时RT-PCR、WesternBlotting及细胞免疫荧光法检测γ分泌酶抑制剂作用前后A2780细胞中HES1的表达水平,Transwell小室试验及MTT法分别用于检测细胞的侵袭、迁移及黏附能力。结果:下调A2780细胞中HES1的表达水平后,A2780细胞的黏附、侵袭及转移能力下降,差异有统计学意义(P<0.01)。结论:HES1的表达与人卵巢癌细胞的侵袭及转移密切相关,下调其表达可抑制卵巢癌细胞的黏附、侵袭及转移能力。 相似文献
7.
目的:探讨赖氨酸脱甲基酶2A(Lysine-specific demethylase 2A,KDM2A)对顺铂(cisplatin,DDP)耐药的人卵巢癌细胞A2780细胞增殖和凋亡的影响及其可能作用机制。方法:通过构建慢病毒载体转染A2780/DDP,分为A2780、A2780/DDP、A2780/DDP/KDM2A(转染KDM2A)、A2780/DDP/Jagged1(转染Jagged1)以及A2780/DDP/NC(转染病毒载体)组。采用Western blot检测3组KDM2A、Jagged1、Bcl2和BAX蛋白表达,CCK8和平板克隆形成实验检测细胞对顺铂的敏感性,流式细胞术检测细胞凋亡情况。结果:A2780/DDP细胞KDM2A和Jagged1的蛋白表达水平均显著高于A2780细胞(P0.05),且A2780/DDP/KDM2A细胞中KDM2A和Jagged1的蛋白表达均低于A2780/DDP以及阴性对照组A2780/DDP/NC(P0.05);A2780/DDP/Jagged1细胞的Jagged1蛋白表达低于A2780/DDP以及A2780/DDP/NC(P0.05),而其KDM2A蛋白的表达比较差异无统计学意义(P0.05)。不同浓度DDP处理的A2780/DDP/KDM2A细胞的生长抑制率均显著高于A2780/DDP/NC和A2780/DDP细胞(P0.05),A2780/DDP/KDM2A细胞克隆形成数量亦明显高于A2780/DDP/NC和A2780/DDP细胞(P0.05)。A2780/DDP/KDM2A细胞凋亡率为(25.84±3.27)%,明显高于A2780/DDP细胞[(14.29±1.96)%](P0.05)和A2780/DDP/NC细胞[(12.46±2.15)%](P0.05)。A2780/DDP/KDM2A细胞中Bcl-2蛋白表达明显低于A2780/DDP细胞(P0.05),而A2780/DDP/KDM2A细胞Bax的表达水平却高于A2780/DDP细胞(P0.05)。结论:KDM2A可能通过上调Jagged1的表达,促进人卵巢癌细胞A2780的增殖并抑制其凋亡,进而降低人卵巢癌耐药细胞A2780的顺铂敏感性。 相似文献
8.
《中国生物工程杂志》2020,(Z1)
目的:探讨S100A9在具核梭杆菌(Fusobacterium nucleatum,Fn)促结肠癌HCT116和SW480细胞增殖与迁移中的作用。方法:Fn感染HCT116和SW480细胞后,分别采用CCK8实验及Transwell实验检测结肠癌细胞的增殖及迁移能力的变化,采用Real time PCR及Western blot分别检测S100A9基因及蛋白质水平的变化;用S100A9 siRNA转染HCT116和SW480细胞后感染Fn,采用CCK8实验及Transwell实验分别检测两株细胞增殖及迁移能力的变化;利用NF-κB抑制剂BAY 11-7082预处理HCT116和SW480细胞后再感染Fn,采用Real time PCR及Western blot分别检测S100A9基因及蛋白质水平的变化。结果:(1) Fn可促进HCT116和SW480细胞增殖及迁移(P 0.001);(2) Fn感染的HCT116和SW480细胞中S100A9基因及蛋白质水平均较相应对照组明显升高,差异均具有高度显著性(P 0. 01或P 0. 001),即Fn上调S100A9的表达;(3)用S100A9 siRNA下调HCT116和SW480细胞中S100A9表达后,Fn促进这两株细胞增殖及迁移的作用则明显减弱,差异均具有高度显著性(P 0. 001),表明S100A9参与介导Fn的促HCT116和SW480细胞增殖及迁移的作用;(4)在NF-κB通路抑制剂BAY 11-7082预处理的HCT116和SW480细胞组,Fn上调S100A9基因及蛋白质水平的作用明显被逆转,差异均具有高度显著性(P 0. 01或P 0. 001),表明激活NF-κB转录活性是Fn上调S100A9的机制之一。结论:Fn可上调S100A9表达且与NF-κB活化有关,上调S100A9是Fn促结肠癌细胞增殖及迁移的机制之一。 相似文献
9.
目的:探讨RNA干扰(RNAi)双向沉默Survivin和RhoA基因表达对人卵巢癌H08910PM细胞增殖、凋亡及侵袭能力的影响。方法:构建Survivin和RhoA基因的串联microRNA干扰载体(Survivin-RhoA-microRNA),通过脂质体介导转染人卵巢癌H08910PM细胞作为实验组,对照组为空载体转染组。转染48小时后,RT—PCR检测Survivin和RhoA的mRNA表达,Western.Blot检测Survivin和RhoA的蛋白质表达;利用MTT法、Transwell小室体外侵袭实验及流式细胞术检测转染后卵巢癌细胞的增殖、侵袭及凋亡情况。结果:Survivin—RhoA—microRNA明显抑制了H08910PM细胞中Survivin和RhoA基因mRNA和蛋白的表达:转染48小时后,实验组SurvivinmRNA和RhoAmRNA表达抑制率分别为68.82%、90.23%,Survivin和RhoA蛋白表达抑制率分别为63.91%、88.47%;MTT分析显示实验组细胞OD490的值为0.706±0.177,较对照组(1.172±0.241)明显降低,差异有统计学意义(P〈0.05);流式细胞术检测实验组细胞凋亡率为36.41±3.34%,较对照组(2.92±0.78%)明显升高(P〈0.01);实验组细胞侵袭百分比为12.56±6.17%,较对照组(32.96±5.14%)明显降低(P〈0.05)。结论:成功构建Survivin和RhoA串联microRNA干扰载体(Survivin.RhoA.microRNA)。Survivin和RhoA双基因沉默显著抑制了卵巢癌H08910PM细胞的增殖和侵袭能力,并增加其凋亡率,两者具有协同作用,为双靶点治疗卵巢癌提供了新的思路和策略。 相似文献
10.
目的:观察异丙酚对大鼠心肌缺血/再灌注时核因子-κB(NF-κB)的活化和细胞凋亡的影响,以探讨异丙酚的心肌保护作用机制。方法:采用阻断大鼠左冠状动脉前降支30min,再灌注2h心肌缺血/再灌注损伤模型。60只SD大鼠随机分为假手术组(Sham)、缺血/再灌注组(I/R)和异丙酚3、6、12mg/(kg.h)组。光、电镜观察心肌组织的形态学变化。免疫组化染色分析心肌组织中NF-κB的核移位,Western blot检测心肌组织NF-κB和caspase-3的表达。原位末端标记(TUNEL)检测心肌细胞凋亡。结果:I/R组心肌纤维排列紊乱,心肌细胞水肿;线粒体膜肿胀,嵴排列紊乱甚至溶解消失。与I/R组相比,6,12mg/(kg·h)组异丙酚组心肌损伤明显减轻。与Sham组相比,I/R组NF-κB活化,明显从细胞浆移位于细胞核,表达量也显著增加(P0.05);心肌caspase-3表达增强(P0.01),心肌细胞凋亡指数升高(P0.05)。而异丙酚6mg/(kg·h)、12mg/(kg·h)组,NF-κB从细胞浆向细胞核的移位被明显限制,NF-κB的表达量也明显低于I/R组(P均0.05);心肌caspase-3表达减弱,心肌细胞凋亡指数减少(与I/R组相比,P0.05)。结论:异丙酚的心肌保护作用可能与其抑制NF-κB的活化,下调caspase-3的表达,从而抑制心肌细胞凋亡有关。 相似文献
11.
目的:克隆壳聚糖酶基因于大肠杆菌中实现高表达,制备壳寡糖。方法:以枯草芽孢杆菌总DNA为模板扩增壳聚糖酶基因(CSN),克隆至载体pET23a(+)上,转化菌株BL21(DE3)。重组子经0.5 mmol/L IPTG诱导后,SDS-PAGE和质谱检测与鉴定重组酶。酶纯化后水解壳聚糖,薄层色谱分析其水解产物。结果:质谱证明壳聚糖酶(31.5kDa)成功表达,表达量占菌体总蛋白的45%左右。纯化后重组酶浓度为900 mg/L,纯度95%、回收率85%,酶活力为10 000 U/mg。壳聚糖降解产物为壳二糖至壳四糖。结论:原核表达载体pET23a(+)-CSN构建正确,壳聚糖酶表达量与活性高,适用于水解壳聚糖制备壳寡糖。 相似文献
12.
国内外蝗害治理技术现状与展望 总被引:3,自引:0,他引:3
本文首先概述了国内外蝗虫发生与为害的态势,总结了现阶段我国蝗虫发生与为害的主要特点:即农田飞蝗暴发频繁而且严重,草原土蝗的发生时常造成严重的经济损失,而且侵入城市干扰市民生活,我国与周边国家之间蝗虫过境迁移频繁,使用化学农药污染环境和农产品;分析了国内外蝗虫防治对策与技术的发展现状,重点介绍了应急防治和可持续治理对策、... 相似文献
13.
Juozas Kulys Kastis Krikstopaitis Arturas Ziemys 《Journal of biological inorganic chemistry》2000,5(3):333-340
N -substituted phenothiazines (PTs) and phenoxazines (POs) catalyzed by fungal Coprinus cinereus peroxidase and Polyporus pinsitus laccase were investigated at pH 4–10. In the case of peroxidase, an apparent bimolecular rate constant (expressed as k
cat/K
m) varied from 1 ×107 M−1 s−1to 2.6×108 M−1 s−1 at pH 7.0. The constants for PO oxidation were higher in comparison to PT. pH dependence revealed two or three ionizable
groups with pK
a values of 4.9–5.7 and 7.7–9.7 that significantly affected the activity of peroxidase. Single-turnover experiments showed
that the limiting step of PT oxidation was reduction of compound II and second-order rate constants were obtained which were
consistent with the constants at steady-state conditions. Laccase-catalyzed PT and PO oxidation rates were lower; apparent
bimolecular rate constants varied from 1.8×105 M−1 s−1 to 2.0×107 M−1 s−1 at pH 5.3. PO constants were higher in comparison to PT, as was the case with peroxidase. The dependence of the apparent
bimolecular constants of compound II or copper type 1 reduction, in the case of peroxidase or laccase, respectively, was analyzed
in the framework of the Marcus outer-sphere electron-transfer theory. Peroxidase-catalyzed reactions with PT, as well as PO,
fitted the same hyperbolic dependence with a maximal oxidation rate of 1.6×108 M−1 s−1 and a reorganization energy of 0.30 eV. The respective parameters for laccase were 5.0×107 M−1 s−1 and 0.29 eV.
Received: 20 September 1999 / Accepted: 24 February 2000 相似文献
14.
Coiled-coil nanomechanics and uncoiling and unfolding of the superhelix and alpha-helices of myosin
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The nanomechanical properties of the coiled-coils of myosin are fundamentally important in understanding muscle assembly and contraction. Force spectra of single molecules of double-headed myosin, single-headed myosin, and coiled-coil tail fragments were acquired with an atomic force microscope and displayed characteristic triphasic force-distance responses to stretch: a rise phase (R) and a plateau phase (P) and an exponential phase (E). The R and P phases arise mainly from the stretching of the coiled-coils, with the hinge region being the main contributor to the rise phase at low force. Only the E phase was analyzable by the worm-like chain model of polymer elasticity. Restrained molecular mechanics simulations on an existing x-ray structure of scallop S2 yielded force spectra with either two or three phases, depending on the mode of stretch. It revealed that coiled-coil chains separate completely near the end of the P phase and the stretching of the unfolded chains gives rise to the E phase. Extensive conformational searching yielded a P phase force near 40 pN that agreed well with the experimental value. We suggest that the flexible and elastic S2 region, particularly the hinge region, may undergo force-induced unfolding and extend reversibly during actomyosin powerstroke. 相似文献
15.
Lapcík O Stursa J Kleinová T Vítková M Dvoráková H Klejdus B Moravcová J 《Steroids》2003,68(14):1147-1155
3-O-Carboxymethylcoumestrol was prepared as the hapten for immunoassay by a partial alkylation of coumestrol with ethyl chloroacetate in acetone alkalized with potassium carbonate. 3-O-Ethoxycarbonylmethylcoumestrol was separated by column chromatography and finally was hydrolyzed with formic acid. 1H and 13C NMR data (APT, COSY, HMQC, and HMBC) revealed that the reaction was regioselective, as 3-O-ethoxycarboxymethylcoumestrol was the only monosubstituted derivative. The hapten was then conjugated to bovine serum albumin and used for immunization of rabbits. A radioimmunoassay (RIA) system was established based on the polyclonal antiserum and a 125I-labeled hapten-tyrosine methyl ester conjugate as the radioligand. Parameters of the RIA: sensitivity: 12 pg per tube, 50% intercept: 140 pg per tube, working range: 20-4000 pg per tube. The cross-reactivity of a panel isoflavonoid and lignan phytoestrogens was either negligible (e.g. formononetin 0.07%; biochanin A 0.06%) or not detectable at all. The major immunoreactive peak in HPLC fractions from an alfalfa extract had the same retention time as coumestrol standard and represented 94.8% of the signal. The remaining 5.2% of immunoreactivity was distributed between five minor peaks. We conclude that after the validation for particular matrices, the method will be a useful tool for analysis of coumestrol, especially in low volume and low concentration samples. 相似文献
16.
白术同源四倍体的诱导和鉴定及其与二倍体过氧化物酶的比较 总被引:18,自引:0,他引:18
以白术(Atractylodes macrooephala Koidz.)二倍体组培苗为材料,对其四倍体诱导方法进行研究,共获得45个白术同源四倍体株系,为优良株系的选育提供了材料。此外,还分析比较了其中8个白术四倍体株系与二倍体的过氧化物酶同工酶(POD)的酶谱差异,发现四倍体各株系过氧化物酶同工酶谱比二倍体的均多了Rf0.310的谱带,且总过氧化物酶比活力也发生了很大改变,对探讨白术四倍体优良株系的生理生化机理具有一定的参考价值。 相似文献
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放牧对牧草光合作用、呼吸作用和氮、碳吸收与转运的影响 总被引:9,自引:1,他引:8
研究放牧对草地植物生理活动的影响,对于揭示草地放牧演替的生理机制有重要意义.大量研究表明,家畜放牧对牧草光合作用、呼吸作用以及C和N吸收与转运的影响,可以分为生理伤害和生理恢复2个阶段.放牧通过改变草地冠层结构影响牧草光合作用,净光合作用速率短期内迅速下降,随着叶面积指数增加又逐渐上升,呼吸作用有相似的变化趋势.牧草放牧后再生长所需的C和N最初主要来自根系和留茬中的贮藏物质,此后随着牧草生长恢复逐渐由同化作用供给,C代谢与土壤N水平负相关.放牧后牧草生理活动变化与牧草遗传特性、种间竞争、家畜放牧特征、非生物环境等因素密切相关. 相似文献
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Summary Three wheats and one triticale were grown, up to flowering stage, in pots on calcareous soil adjusted to a range of salinities
(S1=3.5, S2=6, S3=8.5, and S4=11 mmhos/cm, 20°C, soilpaste extract) by adding solution consisting of 3∶2∶1 of Na-, Ca- and Mg chlorides in chemical equivalent
amounts. Moisture in the pots was kept at 100% (W1), 40% (W2) and 20% (W3) of the available water.
The vegetative growth, nitrogen and phosphate were affected by S and W treatments, chloride was affected only by S. The interaction
S×W affected only dry weight. Varietal effect was observed between wheat as a group and triticale. Multiple quadratic regression
equations of these properties on salinity and water revealed that the higher the available water the wider the range of tolerable
salinity. Triticale was relatively more tolerant to water stress. Salinity increases Cl and decreases N, whereas water stress
enhances N accumulation to a certain extent. However, in triticale at S3 and S4 the effect of water stress on N was overshadowed by the excessive salinity. This did not occur for the wheat (Florence).
P trends were described. R2 for P was low (0.7435–0.3603) which made interpretations rather difficult. 相似文献