Measurements of leucocyte membrane elasticity based on the optical tweezers

A 1-μm in diameter polystyrene bead coated with lectin is trapped by optical tweezers whics is formed by a focused laser beam (740nm).A leucocyte adhered to the bottom of sample cell is chosen to close to the trapped bead.When their stable combination is confirmed,the leucocyte is displaced by moving the sample stage.A“tether”is then formed between the trapped bead and the membrane.The force acting on the tether is measured by a detector equipped on the optical tweezers system.The deformation of the membrane is diverse in different contact conditions.When the contact area is small,the tether is very thin and the force on it is 7.4pN(10^-12N),while the tether is much thicker under a larger contact area and its corresponding force is about 14pN.It is presumed that the latter is related to the deformation of cytoskeleton.     

Nelin的肌动蛋白结合特性及与其相互作用

为了研究人新基因nelin及其心肌表达产物Nelin蛋白的生物特性, 并寻找能够与其发生相互作用的蛋白质, 对Nelin蛋白的全长及其部分结构域进行了肌动蛋白结合共沉淀实验, 并在真核细胞内进行了免疫荧光共定位实验. 结果发现, 在体外共沉淀和细胞内共定位实验中Nelin均表现出同肌动蛋白相结合的能力. 进一步采用酵母双杂交的方法将Nelin与人心脏cDNA文库进行杂交筛选, 得到了3个阳性克隆分别为细丝蛋白(Filamin)C亚型、肌联蛋白(Titin)N2B亚型和a胰蛋白酶抑制物重链前体 (ITIH). 通过免疫共沉淀实验, 验证了Nelin能够同Filamin的羧基末端免疫球蛋白样结构发生相互作用. 通过上述实验证明了Nelin为一个新的肌动蛋白结合蛋白, 并且能够同Filamin相结合. 由此推论新基因nelin的心肌表达产物Nelin是一个细胞骨架相关蛋白并且很可能作为一个膜-细胞骨架连接蛋白, 起到了参与细胞黏着斑形成的过程.     

人肝癌细胞骨架网络系统的初步研究

利用ＴｒｉｔｏｎＸ－１００及其联合（ＮＨ４）２ＳＯ４的抽提技术与ＣｏｏｍａｓｉｅｂｌｕｅＲ２５０染色、免疫酶标技术相结合，体外实验比较研究人肝癌细胞系（ＨｅｐＧ２）细胞骨架网络的分布构像及其中间纤维蛋白构型。结果：人肝癌细胞系（ＨｅｐＧ２）显示其细胞骨架网络的分布构像及其中间纤维对Ｖｉｍｅｎｔｉｎ、Ｋｅｒａｔｉｎ两种抗体均呈现阳性反应。结论：恶性肿瘤细胞的中间纤维蛋白构型可能具有异质性的蛋白分子共表达，这对仅以ＴｒｉｔｏｎＸ－１００加（ＮＨ４）２ＳＯ４的抽提技术进行恶性肿瘤细胞中间纤维蛋白构型分析，是一个必需考虑的问题。     

细胞骨架与早期基因mRNA定位的相关性研究

近年来利用高压电子显微镜、组化、生化和冰冻蚀刻等技术发现了一些mRNA,多聚核糖体及蛋白转译延伸因子在细胞骨架上的定位。蛋白转译机构在细胞骨架上的定位,提示细胞骨架可能与基因表达有关。早期基因c-jun,c-fos,c-mys的产物都是转录因子,对Go至S期有重要调节作用。Zambetti等发现微丝解聚剂——细胞松弛素D（Cytochalasin D,CD）可刺激Hela细胞。c-fos的转录活性,并发现问期细胞中80％的c-fos mRNA与细胞骨架相结合,而CD处理使微丝解聚后,70％以上的mRNA出现在可溶相中, 但微丝解聚剂对C-jun,C- myc的转录活性是否有影响,其mRNA是否与细胞骨架相结合,迄今尚未见报道,本文利用     

影响洋葱表皮细胞骨架制片效果的探讨

以洋葱表皮细胞为材料,探究其经卡诺氏液固定后进行细胞骨架制片观察时,1%Triton X-100抽提非骨架蛋白的最佳时间,并研究洋葱不同表皮细胞及经不同理化因素处理的材料,经卡诺氏液固定后进行细胞骨架制片的步骤及制片效果。结果表明:经卡诺氏液固定后,1%Triton X-100抽提非骨架蛋白内表皮30min较佳,外表皮25min较佳;不同成熟度的鳞片叶细胞骨架形态及稳定性有差异;经紫外线、温度、秋水仙素处理后增加固定步骤再制片对制片观察结果基本无影响。     

2013-02 科技期刊亮点

微切削去除法可测量细胞内部结构的力学特性哈尔滨工业大学机器人技术与系统国家重点实验室刘利等利用原子力显微镜单晶硅针尖对固定的卵巢癌细胞和结肠癌细胞进行微切削逐层去除,测量细胞内部细胞骨架细胞器等各结构的力学特性,为疾病的诊断和治疗提供更精确的实验数据。     

Rearrangements of microtubule cytoskeleton in stomatal closure of <Emphasis Type="Italic">Arabidopsis</Emphasis> induced by nitric oxide

NO （nitric oxide）, known as a key signal molecule in plant, plays important roles in regulation of stomatal movement. In this study, microtubule dynamics and its possible mechanism in the NO signal pathway were investigated. The results were as follows： （i） In vivo stomatsl aperture assays revealed that both vinblastine （microtubule-disrupUng drug） and SNP （exogenous NO donor） prevented stomatsl opening in the light, and vinblastine even could enhance the inhibitory effect of SNP, whereas tsxol （a microtubule-stsbilizing agent） was able to reduce this effect; （ii） microtubules in the opening Arabi-dopsis guard cells expressing GFP：a-tubulin-6 （AtGFP：a-tubulin-6） were organized in parallel, straight and dense bundles, radiating from the ventral side to the dorsal side, and most of them were localized perpendicularly to the ventral wall; （iii） under the same environmental conditions, treated with SNP for 30 min, the radial arrays of microtubules in guard cells began to break down, twisted partially and be- came oblique or exhibited a random pattern; （iv） furthermore, the involvement of cytosolic Ca^2＋ in this event was tested. Stomatal aperture assays revealed that BAPTA-AM （a chelator of Ca^2＋） greatly suppressed the effect of NO on stomatal closure; however, it did not show the same function on stomatal closure induced by vinblastine. When BAPTA-AM was added to the SNP-pretreated solution, the SNP-induced disordered microtubulue cytoskeleton in guard cells underwent rearrangement in a time-dependent manner. After 30 min of treatment with BAPTA-AM, the cortical microtubules resumed the original radial distribution, almost the same as the control. All this indicates that NO may promote rearrangement of microtubule cytoskeleton via elevation of [Ca^2＋]cyt （free Ca^2＋ concentration in the cytoplasm）, finally leading to stomatsl closure.     

La3+通过重组细胞骨架影响大鼠成骨细胞增殖、分化

研究了稀土离子La3 对体外培养的成骨细胞增殖、分化及细胞骨架的影响,并初步探讨了相关机理.用细胞计数法检测了成骨细胞的增殖.用RT-PCR技术测定了碱性磷酸酶(ALP)、骨钙素(OC)、骨桥蛋白(OPN),骨涎蛋白(BSP)以及cbfa-1 mRNA水平.采用激光扫描共聚焦显微镜观察了细胞中F肌动蛋白(F-actin)的变化.结果显示:La3 在48h促进了成骨细胞增殖;在4 d促进了早期分化指标ALP,BSP和cbfa-1 mRNA的表达;在21d促进了晚期分化指标OC和OPN mRNA的表达.与此同时,La3 使成骨细胞骨架发生重组.另外,Western Blot分析证实La3 作用于成骨细胞短时间即可激活粘着斑激酶(FAK)酪氨酸磷酸化.结果提示,La3 通过提高FAK酪氨酸的磷酸化水平,改变细胞骨架的分布和聚合,从而促进成骨细胞的增殖和分化.     

植物细胞骨架在橡胶树产胶和排胶中的可能作用

目前我们对乳管细胞的细胞骨架还了解得很少。根据乳管细胞的结构特点和橡胶生产中乳管被切割的情况,我们推测乳管细胞的细胞骨架在橡胶树产胶和排胶、在乳管伤口的堵塞和愈合中可能起重要作用。