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1.
目的:探讨血清中IL-8对心肌缺血再灌注(MlR)组织中p53、Bcl-2基因的影响。方法建立大鼠心肌缺血再灌注模型,分为对照组和实验组,缺血30min后,再灌注2、4、8、12、24、48h.各时间点分别处死动物,取血备用。采用双抗体夹心酶联免疫吸附(ELISAj法,原位杂交法,原位末端标记染色等,检测IL-8,p53mRNA,bcl-2mRNA,和细胞凋亡。结果:血清中IL-8在IR2h即开始升高,12h达高峰。组织中p53在4h开始升高,24h达高峰。BU-2在8h开始升高-直持续到48h达高峰。血清JL-8早于组织p53表达,Bcl-2晚于p53表达,心肌细胞凋亡和p53mRNA表达时间相-致。结论:血清中IL-8增高介导了组织中p53的高表达,抑制了Bcr-2的作用,在MIR损伤中扮演了-个重要角色。  相似文献   

2.
凋亡相关新基因TFAR19的cDNA克隆、表达和功能研究   总被引:13,自引:0,他引:13  
利用RDA技术从人白血病细胞TF-1中克隆成功一个与凋亡相关的新基因TFA R19.序列分析表明,TFAR19 cDNA全长559bp,其中25-399bp编码125个氨基酸的蛋白质.mRNA斑点杂交分析表明TFAR19在50种组织中均有不同程度的表达.在大肠杆菌中表达并纯化了重组TFAR19蛋白质,制备了多克隆抗体,Western Blot发现TFAR19蛋白在凋亡的TF-1细胞中高表达.瞬时转染TFAR19正义基因后,TF-1细胞去细胞因子后凋亡速度增加.研究表明它能抑制胃癌细胞株803细胞和Hela细胞的生长,促进它们的去血清所致的凋亡.  相似文献   

3.
采用改良的溶胶-凝胶法制备系列的As2O3纳米粒,用透射电镜、扫描电镜、能谱仪、图像分析系统等进行表征及特性检测.应用MTT法研究As2O3纳米粒体外对细胞的增殖抑制作用;用流式细胞仪测定As2O3纳米粒及亚砷酸诱导细胞的凋亡率;用免疫组织化学半定量法检测As2O3纳米粒及亚砷酸处理细胞后Bcl-2、Bax、CD44v6和P53基因的表达改变.研究结果表明,制备的As2O3纳米粒在电镜下呈圆形或椭圆形,分散性较好,平均直径约为80nm、110nm、130nm、150nm和450nm;体外细胞实验证实As2O3纳米粒抗肺癌A-549细胞的效应强于亚砷酸溶液;免疫组织化学半定量法显示As2O3纳米粒有较强的诱导肺癌细胞凋亡的作用,可能与其改变Bcl-2和Bax基因表达(Bcl-2/Bax比值降低)及促进P53基因的表达、抑制CD44v6基因表达有关.  相似文献   

4.
采用乳化-溶剂挥发法制备了同时携载Mn3O4纳米粒和CuS白蛋白纳米粒的诊疗一体化多功能核壳纳米粒,可用于磁共振造影成像手段指引下的肿瘤光热消融,实现癌症的高效诊治.首先用高温热解法制备出粒径均一、分散性较好的Mn3O4纳米粒,并通过蛋白模板法制备CuS白蛋白纳米粒,之后采用可降解性聚乳酸-羟基乙酸共聚物(PLGA)作为载体,通过超声乳化法制备同时携载Mn3O4和CuS纳米粒的核壳纳米粒(Mn3O4/PLGA@CuS),并进行理化性质表征及诊疗一体化性能评价.结果表明:紫外-可见-近红外分光光度计测定结果显示该纳米粒被成功制备,其在近红外区具有较强吸收;核壳纳米粒的平均粒径为160 nm,粒径分布较为均匀;体外磁共振造影成像结果显示随着纳米粒中Mn离子浓度的升高,其造影增强效果也明显提升,显示其具有Mn离子浓度依赖的磁共振成像效果,纵向弛豫率为1.351 mM-1·s-1;在785及980 nm激光辐照下,该核壳纳米粒均显示出良好的光热升温效果.成功制备了具有良好的磁共振造影成像性能和显著的光热升温效应的核壳纳米粒,有望应用于肿瘤的诊疗一体化.  相似文献   

5.
转拟南芥AtNHX1基因促进烟草对钾吸收的研究   总被引:1,自引:0,他引:1  
提取拟南芥总RNA,反转录合成Na /H 逆向转运蛋白基因AtNHX1,构建了植物高效表达载体,用叶盘法将其导入烟草,经筛选获得了抗卡那霉素和GUS染色阳性的转化子38株.应用荧光定量PCR方法对转化材料部分含AtNHX1基因的烟草内源钾通道基因、烟草K 转运蛋白基因和烟草焦磷酸酶基因的转录水平进行了分析.结果表明:在转化烟草中可以检测到AtNHX1基因mRNA,与未转化材料相比,烟草钾离子转运蛋白基因NtHAK1的转录降低,H -ATPase基因NHA1转录增加,钾通道基因NKT转录无显著变化,T1代转化烟叶中K 含量显著增加,Na 、Ca2 无显著变化,烟叶中总糖含量降低,证明了编码拟南芥AtNHX1基因与植物的K 吸收有关.  相似文献   

6.
检测新合成的用作口服胰岛素载体的纳米级PLA-P85-PLA两亲性共聚物的细胞相容性,为临床安全应用提供理论依据。根据所合成材料将来的应用领域,选用Caco-2细胞系为研究对象,采用快速评定细胞增殖率和细胞毒性的四甲基偶氮唑盐微量酶反应比色法(MTT)、中性红摄取(NRU)法和乳酸脱氢酶(LDH)释放法评价共聚物PLA-P85-PLA纳米粒的细胞毒性。结果表明,所合成的共聚物PLA-P85-PLA纳米粒在作用时间(12和24h)和浓度范围(5、10、20和50mg/L)内,对Caco-2细胞无毒性,且有一定的增殖促进作用。新合成的两亲性共聚物PLA-P85-PLA纳米粒无细胞毒性,具有良好的细胞相容性,可用作口服胰岛素载体,为进一步的动物实验提供基础数据。  相似文献   

7.
检测新合成的用作口服胰岛素载体的纳米级PLA-P85-PLA两亲性共聚物的细胞相容性,为临床安全应用提供理论依据。根据所合成材料将来的应用领域,选用Caco-2细胞系为研究对象,采用快速评定细胞增殖率和细胞毒性的四甲基偶氮唑盐微量酶反应比色法(MTT)、中性红摄取(NRU)法和乳酸脱氢酶(LDH)释放法评价共聚物PLA-P85-PLA纳米粒的细胞毒性。结果表明,所合成的共聚物PLA-P85-PLA纳米粒在作用时间(12和24h)和浓度范围(5、10、20和50mg/L)内,对Caco-2细胞无毒性,且有一定的增殖促进作用。新合成的两亲性共聚物PLA-P85-PLA纳米粒无细胞毒性,具有良好的细胞相容性,可用作口服胰岛素载体,为进一步的动物实验提供基础数据。  相似文献   

8.
研究不同浓度Mg-6Zn合金浸提液对大鼠肠上皮细胞IEC-6的凋亡及凋亡相关基因Caspase-3表达的影响。将Mg-6Zn合金制成不同浓度(20%和40%)的浸提液体外培养大鼠肠上皮细胞IEC-6,并以含10%胎牛血清的高糖DMEM(Dulbecco′s modified Eagle′s medium)培养基作为阴性对照。采用噻唑蓝(MTT)法通过计算细胞相对增殖率检测不同浓度合金浸提液对IEC-6细胞的毒性作用;采用流式细胞仪检测不同浓度合金浸提液对IEC-6细胞凋亡的影响;采用逆转录聚合酶链反应(RT-PCR)法检测不同浓度合金浸提液对IEC-6细胞凋亡相关基因Caspase-3mRNA表达的影响。实验结果表明,20%和40%两种浓度的镁合金浸提液在体外培养IEC-6细胞第七天时细胞毒性分级分别为0级和1级,属于无毒性,符合医用材料细胞毒性分级的合格标准。40%和20%浓度镁合金浸提液中,IEC-6细胞的凋亡比例及Caspase-3活化度相对于对照组都增高,差异有统计学意义(P0.05),40%浓度组中,IEC-6细胞的凋亡比例及Caspase-3活化度相对于20%浓度组增高,差异有统计学意义(P0.05),说明Mg-6Zn合金浸提液在一定范围内处于较高浓度时易诱导IEC-6细胞的凋亡及Caspase-3的表达,且随浓度升高影响越大。  相似文献   

9.
藻蓝蛋白对Hela细胞CD59基因表达调控作用的研究   总被引:3,自引:0,他引:3  
探讨了钝顶螺旋藻藻蓝蛋白(PC)对Hela细胞CD59基因表达的调控作用.以正常人CD59cDNA基因为模板,经PCR扩增后重组入真核表达质粒载体pALTER-MAX,然后利用阳离子脂质体(Lipfectamine-2000)将重组质粒和PcDNA共转染人子宫颈癌细胞(Hela)和对照用正常中国仓鼠卵巢细胞(Chinese hamster ovary,CHO)进行表达.用不同浓度的钝顶螺旋藻藻蓝蛋白作用于转染细胞,通过核酸分子杂交技术、免疫荧光标记法和ELISA法对细胞中CD59分子的表达进行检测.结果表明:成功构建了重组质粒pALTER-MAX-CD59,并将其导入真核细胞(Hela,CHO),经G418筛选获得了CD59分子高效表达的细胞克隆.用藻蓝蛋白作用于筛选出的转基因细胞,证实藻蓝蛋白可促进Hela细胞表面CD59蛋白的表达并抑制Hela细胞的增殖,而对于正常CHO细胞无明显作用.  相似文献   

10.
在哈维氏弧菌TS-628菌株鞭毛丝蛋白FlaA基因末端加上一段编码Flag短肽的核苷酸序列作为检测标记后,将该基因克隆到真核表达载体pcDNA3.1( ),酶切、PCR扩增及重组质粒测序证实基因片段插入正确,将该重组质粒命名为pcFlaA.将pcFlaA以肌肉注射方式免疫青石斑鱼.免疫后第7天开始检测鞭毛丝蛋白在石斑鱼肌肉中的表达状况,之后每隔1周检测1次,共检测4次.首先采用PCR技术在DNA水平检测重组质粒转染石斑鱼肌肉细胞的情况,再以RT-PCR法在mRNA水平上检测转染质粒在鱼肌肉中的转录,最后以免疫组化染色技术在蛋白质水平上检测目的蛋白的表达.结果在DNA及mRNA水平上均可检测到目的条带,在蛋白质水平上可检测到明显阳性位点,由此证实pcFlaA可以转染石斑鱼肌肉细胞并可在其中进行表达,而且质粒在鱼体内持续表达的时间至少1个月.  相似文献   

11.
An investigation has been made into the antitumor effect on K562 cells of orpiment nanoparticles which were prepared chemically and analyzed by transmission electron microscope and energy dispersive spectrometry (EDS). Methyl thiazolyl tetrazolium and flow cytometry assays were performed to examine their antitumor effect compared with that of traditional orpiment at various concentrations. The average diameters of the two types of orpiment nanoparticles prepared were 60 nm and 140 nm, respectively, and EDS identified that only orpiment was present. Orpiment nanoparticles greatly inhibited the proliferation of K562 cells by apoptosis, in a concentration-and time-dependent manner, much more effectively than traditional orpiment (p < 0.001). The survival ratio of cells treated with orpiment nanoparticles at 2, 4, 8, and 16 micromol/l after 72 h was 23.0%, 10.1%, 3.2%, and 0.5%, respectively, much lower than 80.0%, 69.0%, 52.3%, and 31.7% of cells treated with traditional orpiment at the corresponding concentration for 72 h. The IC50 of orpiment nanoparticles in K562 cells for 48 h was only 1.27 micromol/l, in comparison with 13.0 micromol/l of traditional orpiment. After treated with orpiment nanoparticles at 4, 8, and 16 micromol/l for 48 h, the apoptotic rate of cells was 11.55%, 20.70%, and 26.45%, respectively, but that in cells treated with traditional orpiment at the same concentration for 48 h was only 3.16%, 3.86%, and 6.46%, respectively. Thus, orpiment nanoparticles can produce a much better cytotoxic effect on cancer cells than that of traditional orpiment.  相似文献   

12.
A novel strategy of enhancing the drug uptake by cancer cells through the combination of anticancer drug doxorubicin with cadium sulfide (CdS) nanoparticles has been explored by using confocal fluorescence scanning microscopy as well as electrochemical studies, which demonstrates that CdS nanoparticles can readily conjugate with doxorubicin on the targeted cancer cells and facilitate the uptake of drug molecules in the human leukemia K562 cells. Besides, our observations also indicate that the aggregation of the leukemia cells occured when CdS nanoparticles were introduced into the relative target system together with doxorubicin, suggesting that the specific association of CdS nanoparticles with biologically active molecules on the surface of leukemia K562 cells may change some biorecognition or signal transfer pathway among cancer cells. It is suggested that the competitive binding of CdS nanoparticles with accompanying anticancer drug to the membrane of leukemia K562 cells could efficiently prevent the drug release by the drug resistant leukemia cells and thus inhibit the relative multidrug resistance (MDR) of targeted cancer cells.  相似文献   

13.
Nickel nanoparticles (Ni NPs) have been applied in a wide range of areas because of their unique structure and properties such as catalysts, high-density magnetic recording media and others. However, little effort has been paid to their biological application and the concrete effect of Ni NPs on biological systems is still unknown. In this study, the possibility of the utilization of the magnetic Ni NPs in cancer cell studies was explored and the effects of the Ni NPs capped with positively charged tetraheptylammonium on leukemia K562 cells in vitro were investigated. Our observations of optical microscopy, atomic force microscopy (AFM) and scanning electron microscopy (SEM) studies indicate that the morphological changes of cancer cells induced by Ni NPs could be apparently observed. The results of 3-(4,5-dimethylthiazol-2-yl)2,5-diphenyl-tetrazolium bromide (MTT) assay, DNA fragmentation and flow cytometry studies demonstrate that the Ni NPs could exert cytotoxicity to leukemia K562 cells at high concentration, and subsequently induce both apoptosis and necrosis of target cancer cells, whilst it had little impact on target cells when at low concentration. Meanwhile, functionalized Ni NPs with positively charged groups could enhance the permeability of cell membrane and facilitate the cellular uptake of outer target molecules into cancer cells. These findings reveal the potential mechanism of Ni NPs to target cancer cells which could induce the cytotoxicity to leukemia cancer cells and suggest the possibility for applications of the Ni NPs in related clinical and biomedical areas.  相似文献   

14.
Quantum dots (QDs) have been receiving a lot of attention recently for their unique fluorescence properties that can be used in drug discovery and bioimaging applications. We have in this article focused particularly on QDs and used it as a transfection vector as well as a fluorescence label for the RNA interference research. The siRNAs were designed to knock down the bcr/abl oncogene in leukaemia K562 cells. EDAC used as a cross-linker, COOH-functionalized QDs were conjugated with NH2-modified siRNAs to generate QD-siRNA conjugates. We also demonstrated their application to the K562 cells. Using such constructs, the delivery and transfection of siRNAs could be monitored by the presence of fluorescent QDs in the conjugates. QDs not only exhibited superior photostability for labeling cells but also worked as a good vector that remarkably increased the transfection efficiency of siRNAs into the cells. Cell proliferation was examined by the MTT assay and cell apoptosis by FACS. Our data have shown that the QD-siRNA conjugates could efficiently inhibit the viability of K562 cells and induced their apoptosis. In summary, QDs can be considered strong tools for the functional analysis of RNAi.  相似文献   

15.
The respiratory activity of collagen-embedded living cells was imaged by scanning electrochemical microscopy (SECM) with the objective to study anticancer drug sensitivity. Two kinds of cancer cells, the human erythroleukemia cell line (K562) and its adriamycin-resistant subline (K562/ADM), were immobilized at the array of microholes micromachined on a silicon wafer for comparative characterization of their sensitivity to the anticancer drug, ADM. The results obtained by the SECM method showed correspondence to a conventional colorimetric assay (SDI assay). Furthermore, since the SECM assay is based on the noninvasive measurement of the respiration activity, continuous monitoring of a dose response was possible.  相似文献   

16.
Twenty specimens of bone-implant interface membrane from THR/TKR were used for in situ localization of apoptotic changes. A panel of antibodies was used to label leukocyte antigens (CD68 and CD3) cytokines (IL-1 and IL-1) and apoptosis inhibiting and promoting proteins (bcl-2 and bax) by means of immunohistochemical techniques. A DNA fragment test on the tissue sections was also carried out to confirm actual cell death using the enzyme terminal deoxy nucleotidyl transferase (TdT) to incorporate biotinylated nucleotide with the 3-OH DNA ends. Leukocyte antigen staining showed that there were large numbers of CD68 positive macrophages as well as multinucleate giant cells (MNGC) but that CD3 positive lymphocytes were also present in the interface membrane. The leukocyte surface antigen staining pattern corresponded to previous findings [1]. Immunostaining with bcl-2 and bax antibodies revealed that both of these proteins were expressed in the cytoplasm of the cells in the interface membrane but they showed different cellular patterns. Bcl-2 was localized in a small number of lymphocyte-like cells while bax was expressed by large numbers of cells, mainly macrophages. The number of cells which expressed bcl-2 was significantly lower than that of bax (P < 0.01). DNA fragment localization occurred mostly in a layer of cells (13 cells deep) next to the implant surface. Again the level of DNA fragment-containing cells was significantly lower than that of bax positive cells (P < 0.01). The results, for the first time, indicate that there is an apoptotic activity occurring in cells in the interface membrane, but not all the cells which express apoptosis-promoting protein (i.e. bax) will enter into the phase of cell death.  相似文献   

17.
The therapeutic effect of Fe2O3 nanoparticles combined with magnetic fluid hyperthermia (MFH) on human hepatocarcinoma SMMC-7721 cells in vitro and xenograft liver cancer in nude mice is studied. We examined growth and apoptosis of SMMC-7721 cells treated with MFH containing Fe2O3 nanoparticles at various concentrations (2, 4, 6, and 8 g/liter) for 30-60 min by using MTT, flow cytometry (FCM), and transmission electron microscopy (TEM). We also observed weight and volume inhibitory rates of the tumors of SMMC-7721-bearing nude mice by using animal experiments. The results showed that Fe2O3 nanoparticles combined with MFH could significantly inhibit the proliferation and increase the ratio of apoptosis of SMMC-7721 cells, and the effect was dose-dependent. The inhibitory rate was 26.5%, 33.53%, 54.4%, and 81.2%, respectively, and the apoptosis rate was 30.26%, 38.65%, 50.28%, and 69.33%, respectively. Animal experiments showed that tumors became small. The weight inhibitory ratio was 42.10%, 66.34%, 78.5%, and 91.46%, and the volume inhibitory ratio was 58.77%, 80.44%, 93.40%, and 98.30%, respectively. Compared with the control and experimental groups, each group had statistically significant difference (p < 0.05). So, Fe2O3 nanoparticles combined with MFH could inhibit the proliferation and induce apoptosis of SMMC-7721 cells and also has a significant inhibitory effect on the weight and volume of xenograft liver cancer. However, the mechanism remains to be further investigated.  相似文献   

18.
In this study, the authors have successfully prepared the polyethylene glycol (PEG)‐coated zinc oxide nanoparticles (ZNPs) and studied its effect in pancreatic cancer cells. The authors have observed a nanosized particle with spherical shape. In this study, the authors have demonstrated the cytotoxic effect of ZNP and PZNP in PANC1 cells. To be specific, PZNP was more cytotoxic compared to that of ZNP in PANC1 cancer cells. The authors have further showed that apoptosis is the main mode of cytotoxic activity. It is worth noting that PEGylation of ZNP did not decrease the cell killing activity of zinc particles, whereas it further increases its anticancer effect in the pancreatic cancer cells. The authors have observed a significant upregulation of proapoptotic BAX while expression of antiapoptotic Bcl‐2 was significantly downregulated indicating the potent anticancer effect of zinc nanoparticles. Overall, PEGylation of ZNP could be an effective strategy to improve the stability, while at the same time, its anticancer activity could be enhanced for better therapeutic response.Inspec keywords: biomedical materials, drug delivery systems, tumours, toxicology, nanoparticles, cellular biophysics, drugs, nanomedicine, cancer, nanofabrication, zinc compounds, II‐VI semiconductorsOther keywords: pancreatic cancer cells, reactive oxygen species, polyethylene glycol‐coated zinc oxide nanoparticles, cytotoxic effect, cytotoxic activity, PEGylation, anticancer effect, PEGylated zinc oxide nanoparticle induce apoptosis, proapoptotic BAX upregulation, ZnO  相似文献   

19.
One of the most important challenges in treating cancer is the invasion and the angiogenesis of cancer cells. The synthesis of green nanoparticles (NPs) and their use in therapeutic fields is one of the most effective methods with minimal side effects in cancer treatment. In this study, cytotoxic and anti‐angiogenic effects of silver NPs (AgNPs) coated with palm pollen extract [Ag–PP(NPs)] were evaluated. For this purpose, the cells were treated with NPs and then were subjected to trypan blue testing (48 h). Then, the cancer invasion was evaluated by the scratch procedure and the expressions of the vascular endothelial growth factor (VEGF) and its receptor (VEGF‐R) genes were estimated using real‐time PCR assay. Also, the angiogenesis effect of the NPs was investigated with chick chorioallantoic membrane (CAM) assay. The Ag–PP(NPs) induced cytotoxicity on MCF7 cells. The findings also showed that Ag–PP(NPs) inhibit invasive cancer cells and reduce the expression of VEGF and VEGF‐R and significantly reduced the number and vessels lengths and the lengths and weights of the embryos in CAM assay. Ag–PP(NPs) with the induction of cytotoxic effects, metastatic inhibition and anti‐angiogenesis properties should be considered as an appropriate option for treatment of cancerInspec keywords: nanomedicine, genetics, cellular biophysics, toxicology, patient treatment, silver, cancer, biochemistry, biomedical materials, nanoparticles, nanofabrication, membranesOther keywords: minimal side effects, cancer treatment, silver NPs, cancer invasion, vascular endothelial growth factor, receptor genes, VEGF‐R, real‐time polymerase chain reaction assay, angiogenesis effect, chick chorioallantoic membrane assay, MCF7 cells, invasive cancer cells, cytotoxic effects, putative mechanism, anticancer properties, antiangiogenic effects, antiangiogenesis properties, Ag–PP‐induced cytotoxicity, metastatic inhibition, palm pollen extraction, trypan blue testing, time 48.0 hour, Ag  相似文献   

20.
The goal of this research is to structure a hyaluronic acid modified nanostructured lipid carrier (HA-NLCs) for vincristine sulfate (VCR) delivery, and detect its efficiency to improve the oral bioavailability. Emulsion solvent evaporation method was used to prepare the HA-NLCs nanoparticles. The particle size, zeta potential and entrapment efficiency of VCR-NLCs and HA-NLCs were 187?±?3.52?nm, ?8.61?±?1.29?mV, 33.12?±?1.16% and 192?±?4.41?nm, ?32.82?±?2.64?mV, 34.41?±?2.21%, respectively. HA-NLCs could significantly improve the cellular uptake efficiency and cytotoxicity in MCF-7 cells than other VCR formulations. The expressions of apoptosis related protein Caspase-3, Caspase-9, Bax and Bcl-2 were estimated by western blot assay in MCF-7 cells, and HA-NLCs exhibited the strongest effect in promoting cell apoptosis. The pharmacokinetics of HA-NLCs was evaluated in Sprague–Dawley male rats and the relative oral bioavailability of HA-NLCs and VCR-NLCs was improved about 1.8-fold and two-fold compared with VCR solution, respectively. Therefore, these results indicated that HA-NLCs could significantly improve the oral bioavailability and was a promising vehicle for the oral delivery of VCR.  相似文献   

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