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1.
结肠腺瘤息肉蛋白(APC)是一个肿瘤抑制因子,它不仅参与Wnt信号通路的传导,而且对细胞粘附、细胞骨架的组织和迁移等都有影响.APC突变发生于大多数结肠癌中.为了探讨APC突变对细胞粘附的影响及机制,本研究利用细胞粘附实验分析了MDCK-APC-N1和对照MDCK-GFP稳定表达细胞株系的细胞粘附情况.实验结果显示,在MDCK细胞中过表达APC-N1导致细胞-细胞间的粘附减少,细胞-基质间的粘附增加.荧光定量PCR和Western印迹实验表明,在MDCK-APC+N1细胞中,E-cadherin表达水平降低,CD29、P-FAK (Y397)、β-catenin和 P-AKT (T308)表达水平升高. 在MDCK-APC-N1细胞中,敲减β-catenin导致E-cadherin表达量升高,而CD29表达没有明显变化.进一步利用PI3K抑制剂LY294002处理MDCK-APC-N1细胞,结果发现,E-cadherin表达量明显升高,CD29表达量明显降低.这些结果揭示,APC-N1可活化 PI3K/AKT 信号通路,进而改变粘附蛋白E-cadherin和CD29影响细胞粘附.  相似文献   

2.
结肠腺瘤息肉蛋白(APC)是一个肿瘤抑制因子,它不仅参与Wnt信号通路的传导,而且对细胞粘附、细胞骨架的组织和迁移等都有影响.APC突变发生于大多数结肠癌中.为了探讨APC突变对细胞粘附的影响及机制,本研究利用细胞粘附实验分析了MDCK-APC-N1和对照MDCK-GFP稳定表达细胞株系的细胞粘附情况.实验结果显示,在MDCK细胞中过表达APC-N1导致细胞-细胞间的粘附减少,细胞-基质间的粘附增加.荧光定量PCR和Western印迹实验表明,在MDCKAPC-N1细胞中,E-cadherin表达水平降低,CD29、P-FAK(Y397)、β-catenin和P-AKT(T308)表达水平升高.在MDCK-APC-N1细胞中,敲减β-catenin导致E-cadherin表达量升高,而CD29表达没有明显变化.进一步利用PI3K抑制剂LY294002处理MDCK-APC-N1细胞,结果发现,E-cadherin表达量明显升高,CD29表达量明显降低.这些结果揭示,APC-N1可活化PI3K/AKT信号通路,进而改变粘附蛋白E-cadherin和CD29影响细胞粘附.  相似文献   

3.
用Factin 特异性FITCphalloidin荧光染料,观察肺炎链球菌(Streptococcus pneumoniae)作用A549细胞前后的Factin细胞骨架重排情况;用细胞松弛素D预处理A49细胞,观察肺炎链球菌对A549细胞的侵袭率;使用Datrolene预处理A549细胞,观察其与Factin细胞骨架重排百分率间是否存在剂量依赖关系;用Fura2/AM荧光探针负载A549细胞后测定肺炎链球菌粘附A549细胞后的胞内Ca2+浓度。结果发现肺炎链球菌作用A549细胞后,Factin细胞骨架呈块状、丝状聚集;而松弛素D可明显降低肺炎链球菌对A549细胞的侵袭率;肺炎链球菌粘附A549细胞后胞内Ca2+高于对照;Datrolene可部分抑制A549细胞Factin细胞骨架重排,且与Factin细胞骨架重排百分率间存在量效关系。以上结果提示肺炎链球菌可通过Ca2+细胞信号转导途径触发A549细胞Factin细胞骨架重排,进而导致肺炎链球菌侵袭A549细胞。  相似文献   

4.
中子属于高传能线密度电离辐射,能产生比κ射线更为严重的放射损伤,肠上皮对中子辐射高度敏感,迄今未见有关中子辐射致肠上皮细胞损伤中PI3K对NF-κB信号通路调控的研究报道.本研究旨在探讨中子照射后肠上皮细胞中PI3K对NF-κB信号通路的调控及其在中子辐射致肠上皮细胞损伤中的作用.选取肠上皮细胞系-6(intestinal epithelial cell No.6,IEC-6)进行传代培养,随机分为对照组、4Gy中子照射组和4Gy中子照射+LY294002处理组,照射组和LY294002处理组细胞采用4Gy中子均匀照射,LY294002处理组细胞在照前24h给予终浓度为10κmol/L的LY294002,各组于照射后6和24h采用MTT比色法、流式细胞术和免疫印迹(Western blot)方法检测IEC-6细胞增殖活力、凋亡与坏死率以及NF-κB信号通路相关分子NF-κB(p65),IKKκ和IκBκ的表达变化.研究发现,4Gy中子照射后6和24h,IEC-6细胞增殖活力下降,凋亡和坏死率增加;应用LY294002后IEC-6细胞增殖活力较照射组明显下降,IEC-6细胞凋亡和坏死率较照射组增加.4Gy中子照射后6和24h,IEC-6细胞NF-κB(p65)和IKKκ表达升高,IκBκ表达降低;应用LY294002后NF-κB(p65)和IKKκ表达降低,IκBκ表达升高,表明4Gy中子照射可引起IEC-6细胞增殖活力下降,凋亡和坏死率增加;PI3K可激活NF-κB信号通路,对中子辐射IEC-6细胞损伤发挥保护作用.  相似文献   

5.
大肠杆菌感染后家蝇幼虫的细胞免疫反应   总被引:1,自引:0,他引:1  
目的探讨家蝇3龄幼虫被大肠杆菌感染后的细胞免疫反应.方法 (1)观察家蝇3龄幼虫感染大肠杆菌后不同时间血淋巴细胞总数(THC)、各类血细胞比例及形态的变化.(2)用比色法分别测定感染后不同时间家蝇3龄幼虫血清中酸性磷酸酶(ACP)及过氧化氢酶(CAT)活性的变化.(3)用聚丙烯酰胺凝胶电泳法测定感染后不同时间家蝇3龄幼虫血清中二酚氧化酶(DPO)活性的变化.结果 (1)感染后4 h、8 h、16 h、24 h THC均有显著增加(P<0.01),(2)感染后4 h、8 h、16 h组的浆血胞和粒血胞的比例明显增加(P<0.01),而珠血胞在感染后各时间组的比例均明显减少(P<0.01),各时间组原血胞和类绛血胞的比例均无明显变化(P>0.05).(3)浆血胞出现细胞变形、空泡等形态变化.(4)大肠杆菌感染后各时间组血清中ACP及CAT活性显著高于对照组(P<0.01),活性分别在感染后8 h、16 h达高峰后逐渐下降.(5)家蝇幼虫血清中二酚氧化酶(DPO)活性在感染后4 h上升,16 h达高峰后下降.结论家蝇幼虫感染大肠杆菌后诱发体内细胞免疫反应,不仅出现血淋巴细胞形态和数量的变化,而且还有血细胞合成、释放多种参与微生物杀灭、清除的酶.  相似文献   

6.
目的探讨温热对人胃癌SGC790l细胞增殖、迁移的影响。方法对照组常温(37℃)下培养人胃癌SGC790l细胞,实验组43℃水浴加热0.5h、1h、2h、3h后培养24h,采用倒置显微镜观察胃癌细胞的形态结构变化;Hoechst-33258荧光染色观察细胞核的变化;四甲基偶氮唑盐比色法(MTT)检测细胞增殖抑制;细胞划痕愈合实验观察温热对胃癌细胞的运动迁移能力的影响;体外细胞侵袭实验(Transwell实验)观察温热对胃癌细胞侵袭能力的影响。结果温热后细胞明显皱缩、变圆及细胞漂浮,3h大部分细胞漂浮;荧光染色显示温热后部分细胞核内出现浓染致密的颗粒块状荧光,胞核固缩、染色质高度凝聚和碎裂;MTT实验提示温热可明显抑制SGC790l细胞生长;细胞划痕实验发现SGC790l细胞温热1h、2 h后细胞迁移距离均明显小于对照组,温热3h后细胞基本未发生迁移;Transwell实验提示SGC790l细胞温热后细胞侵袭能力明显下降。结论温热对胃癌SGC790l细胞具有明显的杀伤作用,温热可明显抑制胃癌SGC790l细胞增殖和侵袭迁移能力。  相似文献   

7.
大肠杆菌K88体外黏附Caco-2细胞及其对细胞膜的影响   总被引:1,自引:0,他引:1  
采用体外Caco-2细胞培养模型,研究大肠杆菌K88黏附Caco-2肠上皮细胞后对其存活率及增殖活力、细胞膜磷脂酶A2、细胞内Ca^2 浓度及膜流动性的影响。结果表明,细菌黏附3h后细胞活力明显下降,PLA2活性升高,细胞内Ca^2 浓度增加,细胞膜流动性降低,从而导致肠上皮细胞膜结构和功能的损害。  相似文献   

8.
机械拉伸对血管平滑肌细胞粘附及生长的影响   总被引:7,自引:0,他引:7  
通过自行研制的“四点弯曲梁”实验装置对血管平滑肌细胞(VSMC)加载培养,并结合显微形态观察和计算机图像处理系统测量细胞铺展投影面积、微管吸吮实验系统检测细胞与表面的粘附力、α-肌动蛋白(actin)免疫组化试验,了解细胞骨架发育和排列取向、流式细胞仪检测细胞动力学以及细胞生长行为等认识VSMC对应变刺激的响应.发现VSMC粘附铺展与实验时间正相关,细胞粘附力、铺展面积、单位面积粘附力4 h后实验组与对照组无显著性差异.VSMC内α-actin发育随加载时间延长呈增加趋势.细胞动力学检测实验组加载24 h后VSMC增殖活动受到抑制.VSMC可能通过调节细胞铺展行为、胞内应力纤维发育等主动机制,实现对机械拉伸的适应性改建.应变刺激有利于体外培养的VSMC维持收缩表型.  相似文献   

9.
钙依赖性粘附素及其信号转导   总被引:1,自引:0,他引:1  
Yan WS  Jiang Y 《生理科学进展》2000,31(4):353-356
钙依赖性粘附素介导的粘附连接在决定和维持发育及成年机体的组织结构中起着重要作用。钙依赖性粘附素结合的特异性取决于其细胞外段,但完整的生理性粘附还需其胞质尾段与胞质相关蛋白以及细胞骨架的相互作用和联系。粘附连接的调节涉及到钙依赖性粘附素基因表达、聚集和磷酸化以及缝隙连接通讯等;此外,钙依赖性粘附素-连环素复合体还参与信号转导过程,从而影响组织的结构和功能。  相似文献   

10.
用荧光染料FIuo-3标记人肝癌细胞株H_(7402)细胞内游离钙,在粘附式细胞仪观察检测单个细胞内游离钙水平的动态变化,细胞在无钙环境中,直接溶解因子(DLF)刺激下细胞内游离钙迅速升高,达到峰值后下降;在细胞培养皿中加入1mmol/L CaCl_2,DLF使胞浆游离钙持续升高;加入10mmol/L CaCl_2,DLF刺激后胞浆游离钙水平无明显变化,表明DLF能引起胞内Ca~(2 )释放和胞外Ca~(2 )内流,细胞外高浓度Ca~(2 )能阻断DLF升高细胞内Ca~(2 )浓度的 作用。  相似文献   

11.
Vitamin K1 (phylloquinone) intestinal absorption is thought to be mediated by a carrier protein that still remains to be identified. Apical transport of vitamin K1 was examined using Caco-2 TC-7 cell monolayers as a model of human intestinal epithelium and in transfected HEK cells. Phylloquinone uptake was then measured ex vivo using mouse intestinal explants. Finally, vitamin K1 absorption was compared between wild-type mice and mice overexpressing scavenger receptor class B type I (SR-BI) in the intestine and mice deficient in cluster determinant 36 (CD36). Phylloquinone uptake by Caco-2 cells was saturable and was significantly impaired by co-incubation with α-tocopherol (and vice versa). Anti-human SR-BI antibodies and BLT1 (a chemical inhibitor of lipid transport via SR-BI) blocked up to 85% of vitamin K1 uptake. BLT1 also decreased phylloquinone apical efflux by ∼80%. Transfection of HEK cells with SR-BI and CD36 significantly enhanced vitamin K1 uptake, which was subsequently decreased by the addition of BLT1 or sulfo-N-succinimidyl oleate (CD36 inhibitor), respectively. Similar results were obtained in mouse intestinal explants. In vivo, the phylloquinone postprandial response was significantly higher, and the proximal intestine mucosa phylloquinone content 4 h after gavage was increased in mice overexpressing SR-BI compared with controls. Phylloquinone postprandial response was also significantly increased in CD36-deficient mice compared with wild-type mice, but their vitamin K1 intestinal content remained unchanged. Overall, the present data demonstrate for the first time that intestinal scavenger receptors participate in the absorption of dietary phylloquinone.  相似文献   

12.
Thiouracil and thyroxine (T4) were injected onto the chick chorioallantois at various developmental stages to study their effects on corneal cellularity, dehydration and structure. Corneas were excised 2–9 days after treatment for histological examination and for analyses of water content, sodium concentration [Na+] and potassium concentration [K+]. Untreated chick corneas showed that water content and [Na+] decreased with advancing embryonic age, while [K+] increased up to stage 42 and then rapidly declined. Corneas from embryos injected with 10 mg thiouracil at stage 36 had a significantly reduced [K+] at stages 40 and 42. Corneas at stages 40, 42 and 45 had a significantly elevated water content when compared with controls. Injection of 15 μg of T4 prior to stage 36 or at or after stage 40 did not produce significant changes in corneal water and ionic content compared with controls; injection of 15 μg of T4 during the cell proliferation period of corneal development (stages 36–40) produced a significant increase in [K+]. Similar results were obtained in corneas from embryos injected with 1 μg of T4. Total corneal thickness was increased in thiouracil treated corneas, and decreased in T4 treated corneas. Epithelial growth was markedly decreased with thiouracil treatment, while T4 had little effect. It is likely that thiouracil treatment decreases cell division in the cornea, and prevents formation of the epithelial barrier, whereas T4 accelerates these processes.  相似文献   

13.
The characteristics of dopamine uptake after acute and subacute cocaine administration were determined in striata from WKY and SHR. In acutely-treated (40 mg/kg, s.c.) rats, significant increases in the Vmax of dopamine uptake were observed 30 min after the cocaine injection in both strains, without changes in Km values. The in vitro IC50 for cocaine was significantly decreased at 30 min in WKY and at 2 h in SHR. However, the in vitro IC50 for GBR-12909 was significantly increased at 30 min and at 2 h in both strains following cocaine administration. In both strains, the density (Bmax) of the [3H]GBR-12935 binding site was significantly increased at 30 min and at 2 h with no charges in Kd. In subacutely-treated (20 mg/kg, twice daily for 3 or 7 days) rats, a significant increase in the Km for dopamine uptake was observed in 7 day treated SHR. The in vitro IC50 for GBR-12909 was significantly increased in 3 day treated WKY. The results suggest that cocaine administration alters dopamine uptake and characteristics of dopamine uptake sites in the rat brain.  相似文献   

14.
The aim of the present work was to study the effects of an unilateral ischaemic-reperfusion injury on Na+, K+-ATPase activity, α1 and β1 subunits protein and mRNA abundance and ATP content in cortical and medullary tissues from postischaemic and contralateral kidneys. Right renal artery was clamped for 40 min followed by 24 and 48 h of reperfusion. Postischaemic and contralateral renal function was studied cannulating the ureter of each kidney. Postischaemic kidneys after 24 (IR24) and 48 (IR48) hours of reperfusion presented a significant dysfunction. Na+, K+-ATPase α1 subunit abundance increased in IR24 and IR48 cortical tissue and β1 subunit decreased in IR48. In IR24 medullary tissue, α1 abundance increased and returned to control values in IR48 while β1 abundance was decreased in both periods. Forty minutes of ischaemia without reperfusion (I40) promoted an increment in α1 mRNA in cortex and medulla that normalised after 24 h of reperfusion. β1 mRNA was decreased in IR24 medullas. No changes were observed in contralateral kidneys. This work provides evidences that after an ischaemic insult α1 and β1 protein subunit abundance and mRNA levels are independently regulated. After ischaemic-reperfusion injury, cortical and medullary tissue showed a different pattern of response. Although ATP and Na+, K+-ATPase activity returned to control values, postischemic kidney showed an abnormal function after 48 h of reflow.  相似文献   

15.
Effects of ammonium on the photosynthetic recovery of Nostoc flagelliforme Berk. et M. A. Curtis were assayed when being rehydrated in low‐K+ or high‐K+ medium. Its photosynthetic recovery was K+ limited after 3 years of dry storage. The potassium absorption of N. flagelliforme reached the maximum after 3 h rehydration in low‐K+ medium but at 5 min in high‐K+ medium. The K+ content of N. flagelliforme rehydrated in high‐K+ medium was much higher than that in low‐K+ medium. The maximal PSII quantum yield (Fv/Fm) value of N. flagelliforme decreased significantly when samples were rehydrated in low‐K+ medium treated with 5 mM NH4Cl. However, the treatment of 20 mM NH4Cl had little effect on its Fv/Fm value in high‐K+ medium. The relative Fv/Fm 24 h EC50 (concentration at which 50% inhibition occurred) value of NH4+ in high‐K+ medium (64.35 mM) was much higher than that in low‐K+ medium (22.17 mM). This finding indicated that high K+ could alleviate the inhibitory action of NH4+ upon the photosynthetic recovery of N. flagelliforme during rehydration. In the presence of 10 mM tetraethylammonium chloride (TEACl), the relative Fv/Fm 24 h EC50 value of NH4+ was increased to 46.34 and 70.78 mM, respectively, in low‐K+ and high‐K+ media. This observation suggested that NH4+ entered into N. flagelliforme cells via the K+ channel. Furthermore, NH4+ could decrease K+ absorption in high‐K+ medium.  相似文献   

16.

Background

Drought is a common stress limiting crops growth and productivities worldwide. Water deficit may increase cellular membrane permeability, resulting in K outflow. Internal K starvation may disorder plant metabolism and limit plant growth. However, it is seldom reported about the effects of external K on drought tolerance of contrasting wheat cultivars.

Methodology/Principal Findings

A hydroponics experiment was carried out in a non-controlled greenhouse. Seedlings of drought-tolerant SN16 and intolerant JM22 were simultaneously treated by five levels of K2CO3 (0, 2.5, 5, 7.5, 10 mM) and two levels of PEG6000 (0, 20%) for 7 days. External K2CO3 significantly increased shoot K+ content, water potential, chlorophyll content as well as gas exchange, but decreased electrolyte leakage (EL) and MDA content in both cultivars under PEG6000 stress. Antioxidant enzymes activities were up-regulated by PEG6000 while external K2CO3 reduced those changes. Molecular basis was explained by measuring the expression levels of antioxidant enzymes related genes. Shoot and root biomass were also increased by K2CO3 supply under drought stress. Although adequate K2CO3 application enhanced plant growth for both cultivars under drought stress, SN16 was better than JM22 due to its high drought tolerance.

Conclusions/Significance

Adequate external K may effectively protect winter wheat from drought injuries. We conclude that drought-tolerant wheat combined with adequate external K supply may be a promising strategy for better growth in arid and semi-arid regions.  相似文献   

17.
18.

Background

Recently electroporation using biphasic pulses was successfully applied in clinical developments for treating tumours in humans and animals. We evaluated the effects of electrical treatment on cell adhesion behaviour of breast cancer cells and fibroblasts. By applying bipolar electrical pulses we studied short- and long-lived effects on cell adhesion and survival, actin cytoskeleton and cell adhesion contacts in adherent cancer cells and fibroblasts.

Methods

Two cancer cell lines (MDA-MB-231 and MCF-7) and one fibroblast cell line 3T3 were used. Cells were exposed to high field intensity (200 - 1000 V/cm). Cell adhesion and survival after electrical exposure were studied by crystal violet assay and MTS assay. Cytoskeleton rearrangement and cell adhesion contacts were visualized by actin staining and fluorescent microscope.

Results

The degree of electropermeabilization of the adherent cells elevated steadily with the increasing of the field intensity. Adhesion behaviour of fibroblasts and MCF-7 was not significantly affected by electrotreatment. Interestingly, treating the loosely adhesive cancer cell line MDA-MB-231 with 200 V/cm and 500 V/cm resulted in increased cell adhesion. Cell replication of both studied cancer cell lines was disturbed after electropermeabilization. Electroporation influenced the actin cytoskeleton in cancer cells and fibroblasts in different ways. Since it disturbed temporarily the actin cytoskeleton in 3T3 cells, in cancer cells treated with lower and middle field intensity actin cytoskeleton was well presented in stress fibers, filopodia and lamellipodia. The electrotreatment for cancer cells provoked preferentially cell-cell adhesion contacts for MCF-7 and cell-ECM contacts for MDA-MB- 231.

Conclusions

Cell adhesion and survival as well as the type of cell adhesion (cell-ECM or cell-cell adhesion) induced by the electroporation process is cell specific. The application of suitable electric pulses can provoke changes in the cytoskeleton organization and cell adhesiveness, which could contribute to the restriction of tumour invasion and thus leads to the amplification of anti-tumour effect of electroporation-based tumour therapy.  相似文献   

19.
海拔和郁闭度对祁连山青海云杉林叶凋落物分解的影响   总被引:4,自引:0,他引:4  
李娜  赵传燕  郝虎  臧飞  常亚鹏  汪红  杨建红 《生态学报》2021,41(11):4493-4502
为了探究海拔和郁闭度对青海云杉林叶凋落物分解的影响,本文选择海拔为2850 m,3050 m,3250 m和3450 m四个梯度和高、中、低三个林分郁闭度,采用分解网袋法,研究青海云杉叶凋落物分解速率及分解过程中N、P元素变化。结果表明,质量损失率随时间在波动增大。分解速率先减小后增大,不同海拔下分解速率为K3450 > K3050 > K3250 > K2850,不同郁闭度下分解速率为K > K > K,青海云杉叶枯落物分解50%和95%所需时间约为5.3 a和22.7 a。枯落物分解过程中,N、P含量和累积系数在不同海拔和郁闭度下的变化不同,与季节变化有关。研究结果为祁连山森林生态系统地球化学循环奠定基础。  相似文献   

20.

Background

The intratracheal instillation of Pseudomonas aeruginosa entrapped in agar beads in the mouse lung leads to chronic lung infection in susceptible mouse strains. As the infection generates a strong inflammatory response with some lung edema, we tested if it could modulate the expression of genes involved in lung liquid clearance, such as the α, β and γ subunits of the epithelial sodium channel (ENaC) and the catalytic subunit of Na+-K+-ATPase.

Methods

Pseudomonas aeruginosa entrapped in agar beads were instilled in the lung of resistant (BalB/c) and susceptible (DBA/2, C57BL/6 and A/J) mouse strains. The mRNA expression of ENaC and Na+-K+-ATPase subunits was tested in the lung by Northern blot following a 3 hours to 14 days infection.

Results

The infection of the different mouse strains evoked regulation of α and β ENaC mRNA. Following Pseudomonas instillation, the expression of αENaC mRNA decreased to a median of 43% on days 3 and 7 after infection and was still decreased to a median of 45% 14 days after infection (p < 0.05). The relative expression of βENaC mRNA was transiently increased to a median of 241%, 24 h post-infection before decreasing to a median of 43% and 54% of control on days 3 and 7 post-infection (p < 0.05). No significant modulation of γENaC mRNA was detected although the general pattern of expression of the subunit was similar to α and β subunits. No modulation of α1Na+-K+-ATPase mRNA, the catalytic subunit of the sodium pump, was recorded. The distinctive expression profiles of the three subunits were not different, between the susceptible and resistant mouse strains.

Conclusions

These results show that Pseudomonas infection, by modulating ENaC subunit expression, could influence edema formation and clearance in infected lungs.  相似文献   

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