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1.
该研究探讨氰酸盐(cyanate)诱导肾小管上皮细胞氧化应激损伤和促进肾纤维化的作用。氰酸盐作用HK-2肾小管上皮细胞后, CCK8法检测其对细胞活力的影响;倒置显微镜观察细胞形态的改变; DCFH-DA法检测细胞ROS水平;细胞免疫荧光和Western blot分别检测E-cadherin、Fibronectin、α-SMA的表达; Western blot检测TGF-β的表达水平。结果显示, 2 mmol/L氰酸盐明显下调HK-2细胞的活力(P<0.05),细胞形态变为长梭形。氰酸盐作用24 h后, HK-2细胞内ROS水平呈浓度依赖性升高。免疫荧光和Western blot结果均显示,氰酸盐作用24 h后, HK-2的Fibronectin、α-SMA表达升高, E-cadherin表达下降; TGF-β的表达水平随氰酸盐浓度升高而上调(P<0.05)。以上结果表明,氰酸盐诱导肾小管上皮细胞产生过量ROS,上调TGF-β水平促进细胞上皮–间充质细胞转化(epithelia-mesenchymal transition, EMT)。  相似文献   

2.
目的: 观察丙泊酚对转化生长因子-β1(TGF-β1)诱导的肝星状细胞系HSC2-T6细胞激活的影响并探讨其可能的作用机制。方法: 实验分为对照组、TGF-β1组、丙泊酚组、TGF-β1+丙泊酚组、雷帕霉素组、TGF-β1+丙泊酚+雷帕霉素组。先用含雷帕霉素(5 μmol/L)DMEM培养液培养1 h,用丙泊酚(100 μmol/L)处理1 h,然后再加入TGF-β1(5 ng/ml)继续共同培养24 h。细胞的增殖水平通过MTT法测量,细胞培养液上清中透明质酸(HA)、IV型胶原(IV-C)和层粘连蛋白(LN)的浓度采用ELISA法测量,细胞的超微结构采用透射电镜观测,α-平滑肌肌动蛋白(α-SMA)、哺乳动物雷帕霉素靶蛋白(mTOR)、磷酸化mTOR(p-mTOR)及自噬相关基因Beclin 1、微管相关蛋白1轻链3(LC3)和p62的表达通过Western blot测量。结果: 与对照组比较,TGF-β1组细胞增殖、α-SMA蛋白的表达、培养液上清中HA、IV-C和LN的浓度、自噬体数量、Beclin-1和LC3-Ⅱ的蛋白表达及LC3-Ⅱ/LC3-Ⅰ比值显著性增加(P均<0.05),p-mTOR蛋白的表达和p-mTOR/mTOR比值及p62的蛋白表达显著性降低(P均<0.05)。与TGF-β1组比较,丙泊酚+TGF-β1组细胞增殖、α-SMA蛋白的表达、培养液上清中HA、IV-C和LN的浓度、自噬体数量、Beclin-1和LC3-Ⅱ的蛋白表达及LC3-Ⅱ/LC3-Ⅰ比值均显著性降低(P均<0.05),p-mTOR蛋白表达和p-mTOR/TOR比值及p62的蛋白表达均显著性增加(P均<0.05)。mTOR抑制剂雷帕霉素部分逆转丙泊酚的作用。结论: 丙泊酚抑制TGFβ1诱导的肝星状细胞激活,其机制涉及mTOR-自噬途径。  相似文献   

3.
4.
目的:研究上调大鼠肾小管上皮细胞中trip-1蛋白的表达量对TGF-β1诱导的上皮细胞转分化的影响.方法:包装人TRIP-1基因重组腺病毒,用其感染NRK52E细胞36h上调内源性trip-1蛋白表达量,之后用10 ng/ml的TGF-β1对细胞进行刺激诱导,72h后做Western Blot检测细胞中E-cadherin蛋白和α-SMA蛋白表达量.结果:①包装的人TRIP-1基因重组腺病毒感染细胞能够有效上调细胞中trip-1蛋白的表达量.②上调细胞中trip-1蛋白表达量,对TGF-β1引起的NRK52E细胞中E-cadherin蛋白表达水平降低有所抑制,但对TGF-β1引起的NRK52E细胞中α-SMA蛋白表达水平升高没有明显调节作用.结论:上调大鼠肾小管上皮细胞中trip-1蛋白的表达量在一定程度上抑制了TGF-β1诱导的NRK52E细胞转分化.  相似文献   

5.
该研究旨在探讨Hedgehog-Gli1(HH)信号在肾小管上皮细胞表型转化(epithelial-mesenchymal transition,EMT)和胶原累积中的作用及与TGF-β1信号的对话机制。该实验通过体外培养大鼠肾小管上皮细胞NRK-52E,以溶剂作为对照组,以1~50 ng/m L重组蛋白sonic hedgehog(Shh)或5 ng/m L TGF-β1作为诱导组,以加入或不加入HH信号特异性阻断剂环靶明(cyclopamine,Cyp)5μmol/L为干预组。细胞培养24 h,采用ELISA、q RT-PCR、免疫细胞荧光染色和Western blot等方法检测HH信号相关分子(Ptch1、Smo和Gli1)、TGF-β1、EMT相关分子(Rac1蛋白、肌成纤维细胞标志物α-SMA和上皮细胞标志物E-cadherin)、III型胶原m RNA或蛋白的表达。结果发现,外源性Shh上调Smo和Gli1表达,抑制Ptch1表达,继而激活HH信号;HH信号活化抑制肾小管上皮细胞E-cadherin的表达,上调α-SMA、III型胶原和TGF-β1的表达。环靶明干预后,Smo表达下调,进而抑制HH信号、EMT和胶原累积,并下调TGF-β1的表达。应用TGF-β1诱导小管上皮细胞EMT,同时也上调HH信号分子Smo和Gli1的表达,下调Ptch1的表达,提示TGF-β1可诱导HH信号活化。综上所述,HH信号和TGF-β1均参与了肾小管上皮细胞EMT和胶原累积过程。HH信号活化可促进TGF-β1的表达,同时TGF-β1能激活HH信号,推测TGF-β1与HH信号可能存在交叉对话以调控EMT和胶原累积。  相似文献   

6.
目的:探讨大黄素对TGF-β1诱导的人肾小管上皮细胞(HK-2)间质转分化的影响。方法:不同浓度大黄素分别作用于TGF-β1诱导HK-2细胞24 h和48 h,通过细胞增殖实验确定最佳大黄素最佳给药浓度。TGF-β1诱导HK-2细胞24 h后收集细胞用于免疫印迹Western blot和实时荧光定量PCR(RT-PCR)分析。Western印迹法分别检测纤维化相关蛋白Collagen IV的表达,和肾小管上皮细胞向间充质细胞转分化关键蛋白α-SMA和E-Cadherin的表达;RT-PCR法检测肾小管上皮细胞向间充质细胞转分化关键蛋白α-SMA的表达。结果:由细胞增殖实验结果表明40μM大黄素是最佳给药浓度。Western结果表明,与模型组相比,大黄素组下调纤维化相关蛋白Collagen IV的表达,大黄素组与模型组蛋白差异有统计学意义(P0.05)。与模型组相比,大黄素组下调α-SMA蛋白表达水平,而上调E-Cadherin蛋白表达,差异有统计学意义(P0.05)。RT-PCR结果表明,与模型组相比,大黄素组降低α-SMA mRNA的含量,大黄素组与模型组α-SMA mRNA含量差异有统计学意义(P0.05)。结论:大黄素可通过抑制TGF-β1诱导的HK-2细胞间质转分化,从而发挥延缓肾间质纤维化的过程。  相似文献   

7.
探讨转化生长因子-β1(TGF-β1)对人子宫颈癌细胞Siha侵袭、迁移能力的影响及其可能的分子机制。用5 ng/m L TGF-β1作用Siha细胞72 h,通过倒置显微镜、CCK-8实验、单细胞克隆形成实验、细胞黏附实验和Transwell小室实验分别观察TGF-β1作用前后Siha细胞形态、增殖能力、克隆形成能力、黏附能力、迁移及侵袭能力的改变;Western blot检测TGF-β1作用前后基质金属蛋白酶-2(MMP-2)、MMP-9、VEGF、CFTR、P50、P65、E-cadherin及Vimentin蛋白表达的变化。结果表明,5 ng/m L TGF-β1作用Siha细胞72 h后,Siha细胞出现上皮细胞向间质细胞形态转变、黏附能力减弱、迁移和侵袭能力增强,伴随MMP-2、MMP-9、VEGF、CFTR、P50、P65和Vimentin表达上调,E-cadherin表达下调。该研究表明,TGF-β1可能通过诱导Siha细胞发生上皮–间质转化及上调MMP-2、MMP-9和VEGF的表达,促进Siha细胞的侵袭转移。  相似文献   

8.
采用定量Real-time PCR(q RT-PCR)、Western blot法检测NP-69和5-8F细胞中FHL2表达情况及转染后5-8F各组细胞中FHL2、c-myc、β-catenin的m RNA与蛋白质水平;利用CCK-8法、细胞平板克隆形成实验、划痕实验及Transwell实验分别检测转染后5-8F各组细胞增殖、迁移及侵袭能力。实验结果显示,鼻咽癌细胞株5-8F的FHL2蛋白质水平高于NP-69,转染si RNA的5-8F细胞中FHL2表达水平显著低于无关序列组和空白对照组(P0.01)。下调FHL2基因表达后5-8F细胞增殖、侵袭、迁移能力均受到抑制(P0.05),且转染组5-8F细胞中c-myc、β-catenin的m RNA及相应蛋白质水平均有下降(P0.01)。综上所述,在5-8F细胞中FHL2高表达。FHL2基因敲低后可以抑制5-8F细胞增殖、侵袭、迁移能力,并可能通过Wnt信号通路影响鼻咽癌5-8F细胞的恶性生物学行为。  相似文献   

9.
p38 MAPK介导高糖诱导的肾小管上皮细胞向间充质细胞转变   总被引:2,自引:0,他引:2  
本文旨在观察p38MAPK与高糖诱导的肾小管上皮细胞向间充质细胞转变之间的关系。将雄性Sprague—Dawley(SD)大鼠随机分为对照组、糖尿病组、胰岛素治疗组,用免疫组织化学、Western blot检测p38MAPK和磷酸化p38MAPK(P—p38MAPK)蛋白表达。采用机械分离和酶消化获取SD大鼠肾小管节段,进行肾小管上皮细胞培养,将肾小管上皮细胞分为对照组、高渗组(20mmol/L D—mannitol)、高糖组(20mmol/L D—glucose)和SB202190(p38MAPK特异性抑制剂)+高糖组,处理72h后收集细胞,用免疫细胞化学检测α-平滑肌肌动蛋白(α—smooth muscleactin,α-SMA)、p-p38MAPK和Snaill蛋白表达,Western blot检测p38MAPK、p-p38MAPK、Snaill、转化生长因子β1(transforming growth factor—β1,TGF-β1)、α-SMA和E-cadherin的表达,RT-PCR检测α-SMA和E-cadherin mRNA的表达。体内和体外结果均显示,高糖状态激活了p38MAPK,这种活化作用在体内可因胰岛素控制血糖而被消除,在体外可被p38MAPK特异性抑制剂SB202190显著抑制;高糖组α-SMA蛋白和mRNA在原代培养肾小管上皮细胞的表达较对照组分别增加12倍和8倍(P〈0.01),SB202190处理组其表达则较高糖组分别减少67%和50%(P〈0.01)。SB202190不影响TGF—β1蛋白表达,但下调Snaill蛋白表达,并部分恢复高糖组E—cadherin蛋白和mRNA的表达。上述结果提示,p38MAPK可能通过转录因子Snaill介导高糖诱导的肾小管上皮细胞向间充质细胞转变。  相似文献   

10.
目的:观察miR-21在转化生长因子β1(TGF-β1)诱导的人肾小管上皮细胞(HK-2细胞)上皮间质转分化(EMT)中的作用,并探讨miR-21参与调控HK-2细胞EMT的可能靶点。方法:体外培养的HK-2细胞分为6组:正常对照组、转化生长因子β1(TGF-β1)模型组、miR-21 mimic阴性组、miR-21 mimic组、miR-21 inhibitor阴性组和miR-21 inhibitor组。细胞经4 ng/ml TGF-β1处理建立EMT模型,检测miR-21和EMT相关指标的表达变化,利用基因转染技术,将miR-21 mimic质粒或miR-21 inhibitor质粒转染经TGF-β1处理的HK-2细胞,使细胞过表达或抑制表达miR-21,在此基础上观察细胞EMT相关指标的变化以及磷酸酯酶(PTEN)基因的影响。结果:(1)与正常组相比,模型组的miR-21含量显著升高(P0.05),上皮表型标志物E-cadherin的mRNA和蛋白表达水平均显著降低(P0.01),间质表型标志物α平滑肌肌动蛋白(α-SMA)mRNA和蛋白水平也显著升高(P0.05,P0.01);(2)转染miR-21 mimic后,与miR-21 mimic阴性对照组相比,miR-21含量显著升高(P0.01),PTEN、E-cadherin的mRNA和蛋白水平显著降低(P0.05,P0.01),α-SMA mRNA和蛋白水平显著升高(P0.05,P0.01);转染miR-21 inhibitor后,与miR-21 inhibitor阴性对照组相比,miR-21含量显著降低(P0.01),PTEN、E-cadherin的mRNA和蛋白含量显著升高(P0.05,P0.01),α-SMA mRNA、蛋白水平显著降低(P0.05,P0.01)。结论:miR-21在TGF-β1诱导的HK-2细胞EMT发生中具有重要作用,并且可能通过靶基因PTEN参与EMT相关分子的表达调控。  相似文献   

11.
The goal of this article was to establish reference ranges of the concentration of trace elements in human serum and to compare these results with those reported by other authors. We describe the sample preparation and measurement conditions that allow the rapid, precise, and accurate determination of Al, As, B, Be, Cd, Co, Cu, Fe, Li, Mn, Mo, Ni, Rb, Se, Sr, and Zn in human serum samples (n=110) by inductively coupled plasma-mass spectrometry (ICP-MS). Accuracy and precision were determined by analyzing three reconstituted reference serum samples by comparison with other methods and by the standard addition procedure. The advantages of the ICP-MS method include short time of analysis of the elements mentioned, low detection limit, high precision, and high accuracy. Disadventages include a high risk of contamination due to the presence of some of the elements of interest in the environment, the relatively delicate sample handling, and the high cost of the equipment.  相似文献   

12.
Summary Concentration of N, P, K, Ca, Mg and S in summer groundnut crop was higher than in kharif while Zn, Fe, Mn and Cu contents were higher in summer crop. Kernel's N, P and Zn; Leaflet's Ca and Mn; Stem's K and Fe; Root's S and Cu and Petiole's Mg contents were highest. Shell's N, P, K, Mg, S, Zn and Cu; Kernel's Ca, Fe and Mn contents were the least. N, P, K, S, Zn and Cu concentrations decreased linearly as the crop grew. Ca, Mg, Fe and Mn concentrations did not display any distinct pattern. Ca concentration was positively correlated with pod yield in both the seasons.  相似文献   

13.
A new polynitro cage compound 2, 4, 6, 8, 10, 12, 13, 14, 15-nonanitro-2, 4, 6, 8, 10, 12, 13, 14, 15-nonaazaheptcyclo [5.5.1.1(3,11).1(5,9)] pentadecane (NNNAHP) was designed in the present work. Its molecular structure was optimized at the B3LYP/6-31 G(d,p) level of density functional theory (DFT) and crystal structure was predicted using the Compass and Dreiding force fields and refined by DFT GGA-RPBE method. The obtained crystal structure of NNNAHP belongs to the P-1 space group and the lattice parameters are a = 9.99 ?, b = 10.78 ?, c = 9.99 ?, α = 90.01°, β = 120.01°, γ = 90.00°, and Z = 2, respectively. Based on the optimized crystal structure, the band gap, density of state, thermodynamic properties, infrared spectrum, strain energy, detonation characteristics, and thermal stability were predicted. Calculation results show that NNNAHP has detonation properties close to those of CL-20 and is a high energy density compound with moderate stability.  相似文献   

14.
This report attempts to formulate reference ranges of elemental concentrations for 15 trace elements in selected human tissues and body fluids. A set of samples consisting of whole blood, blood serum, urine, milk, liver, and hair were chosen and considered for 15 elements of biological significance: As, Cd, Co, Cr, Cu, F, Fe, I, Hg, Mn, Mo, Ni, Pb, Se, and Zn. The results represent wholly or partially data received from 40 countries of the global regions of Africa, Asia, Europe, North, South, and Central America, Australia, and New Zealand. This survey, even if qualitative, has been useful in demonstrating certain trends of trace-element scenarios around the world. It is of course recognized that both diet and environment exert a strong influence on the distribution pattern of several elements, such as As, Cd, Mn, Pb, Se, and Zn. A limited comparison of the available information on soil status of different countries reflected some interesting associations for elements, such as Mn and Zn. Importantly, this study revealed that only a few countries were in a position to identify a reasonable amount of data on samples requested for this project. Regretably, for a number of countries, any dependable data for even such essential elements as Cu, Fe, and Zn were not available. In view of the nutritional importance of many elements, the time is ripe for concerted efforts by intergovernmental agencies to initiate investigations or commission task forces/projects to generate reliable reference data for selected global regions, which sadly lack data of any kind at present.  相似文献   

15.
16.
The unitary conductances and permeability sequences of the rat connexin40 (rCx40) gap junction channels to seven monovalent cations and anions were studied in rCx40-transfected neuroblastoma 2A (N2A) cell pairs using the dual whole cell recording technique. Chloride salt cation substitutions (115 mM principal salt) resulted in the following junctional maximal single channel current-voltage relationship slope conductances (γj in pS): CsCl (153), RbCl (148), KCl (142), NaCl (115), LiCl (86), TMACl (71), TEACl (63). Reversible block of the rCx40 channel was observed with TBA. Potassium anion salt γj are: Kglutamate (160), Kacetate (160), Kaspartate (158), KNO3 (157), KF (148), KCl (142), and KBr (132). Ion selectivity was verified by measuring reversal potentials for current in rCx40 gap junction channels with asymmetric salt solutions in the two electrodes and using the Goldman-Hodgkin-Katz equation to calculate relative permeabilities. The permeabilities relative to Li+ are: Cs+ (1.38), Rb+ (1.32), K+ (1.31), Na+ (1.16), TMA+ (0.53), TEA+ (0.45), TBA+ (0.03), Cl (0.19), glutamate (0.04), and NO3− (0.14), assuming that the monovalent anions permeate the channel by forming ion pairs with permeant monovalent cations within the pore thereby causing proportionate decreases in the channel conductance. This hypothesis can account for why the predicted increasing conductances with increasing ion mobilities in an essentially aqueous channel were not observed for anions in the rCx40 channel. The rCx40 effective channel radius is estimated to be 6.6 Å from a theoretical fit of the relationship of relative permeability and cation radius.  相似文献   

17.
The variation with age of the 18 trace element mass fractions and some histological characteristics of intact prostate glands of 50 subjects aged 0–30 years was investigated by instrumental neutron activation analysis, inductively coupled plasma atomic emission spectrometry, and a quantitative morphometric analysis. Mean values ± standard error of the mean (M ± SΕΜ) for the mass fractions (in milligrams per kilogram wet tissue) of these trace elements in pre-puberty were: Al 28.5 ± 9.0, B 0.40 ± 0.11, Ba 1.48 ± 0.44, Br 10.5 ± 1.5, Ca 241 ± 30, Cl 3,203 ± 278, Cu 3.51 ± 0.89, Fe 33.7 ± 4.1, K 2,364 ± 145, Li 0.020 ± 0.004, Mg 153 ± 23, Mn 0.46 ± 0.06, Na 2,286 ± 130, P 1,391 ± 100, S 1,698 ± 132, Si 62 ± 11, Sr 0.38 ± 0.08, and Zn 27.6 ± 2.3. During puberty and postpuberty, when there is a significant increase in circulating androgens, the mean values were: Al 7.2 ± 1.4, B 0.21 ± 0.05, Ba 0.25 ± 0.06, Br 5.8 ± 1.0, Ca 433 ± 81, Cl 2,314 ± 201, Cu 1.77 ± 0.13, Fe 20.9 ± 1.6, K 2,585 ± 118, Li 0.0088 ± 0.0014, Mg 232 ± 27, Mn 0.34 ± 0.04, Na 1,875 ± 107, P 1,403 ± 98, S 1,673 ± 73, Si 22.2 ± 3.1, Sr 0.22 ± 0.03, and Zn 93.3 ± 8.9. Mean values (M ± SΕΜ) of percent volumes (%) of the stroma, epithelium and lumen in the prostate before puberty were 73.4 ± 2.6, 20.4 ± 1.7, and 4.45 ± 0.94, respectively, versus 46.5 ± 2.5, 38.5 ± 1.9, and 14.9 ± 1.2 during puberty and postpuberty. This work’s results confirm that the Zn mass fraction in prostate tissue is an androgen-dependent parameter. For the first time it has been demonstrated that the glandular lumen is a main pool of Ca, Mg, and Zn accumulation and that the stroma is a main pool of Al, B, Ba, Br, Cl, Cu, Fe, Mn, Na, and Si accumulation in the normal human prostate, for the age range 0–30 years. It was concluded that the Ca, Mg, and Zn binds tightly within the prostatic fluid, because the volume of glandular lumen reflects the volume of prostatic fluid.  相似文献   

18.
The effect of age and gender on major, minor, and trace element contents in the intact rib bone of 80 relatively healthy 15–55-year-old women and men was investigated. Contents or upper limit of contents of 16 chemical elements in the rib bone were determined by inductively coupled plasma atomic emission spectrometry (ICP-AES). Mean values (M?±?SΕΜ) for the mass fraction of Ba, Ca, Cu, Fe, K, Li, Mg, Na, P, S, Sr, and Zn (milligram per kilogram of dry bone) were as follows: 2.54?±?0.16, 171,400?±?4,050, 1.35?±?0.22, 140?±?11, 1,874?±?71, 0.049?±?0.011, 2,139?±?38, 5,378?±?88, 75,140?±?1,660, 1,881?±?51, 291?±?20, and 92.8?±?1.5, respectively. The upper limits of contents of Al, B, Mn, and V were <7.20, <0.65, <0.36, and <0.03, respectively. Statistically significant tendency for the Ca, Mg, and P content to decrease with age was found in the human rib bone, regardless of gender. The mass fraction of Fe in the male rib bone increases with age. It was shown that higher Ca, Mg, Na, P, and Sr mass fractions as well as lower Fe content were typical of female ribs as compared to those in male ribs.  相似文献   

19.
Nine blood group systems of goats were identified using 12 caprine reagents produced by absorption of alloimmune antisera. The caprine C blood group system, possibly homologous to the ovine C blood group system, was characterized by two reagents and shown to be controlled by three alleles,C 12,C 25, andC . A more complex blood group system of goats, designated G, was identified using three reagents and shown to be controlled by six codominant alleles (G 10.19.20,G 10.19,G 10.20,G 10,G 19,G 20) and a recessive allele (G ). A further seven one-factor two-allelic systems were identified by seven reagents. The nine genetic systems provided exclusion probabilities of 0.479, 0.492, 0.548, and 0.572 in Australian Angora, Dairy, Cashmere, and Texan Angora goat breeds, respectively. This work was supported by a grant from the Australian Stud Book, Alison Road, Randwick, New South Wales 2031, Australia.  相似文献   

20.
《Ibis》1894,36(3):447-460
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