细胞

Some key techniques related to the close and continuous process were investigated by the application of H9N2 avian influenza virus with Vero cells, such as the susceptibility of cell to influenza virus, virus production with cell microcarrier culture method, cell bead-to-bead transfer, virus production through bead-to-bead transfer, cell culture and virus production with serum free medium, metabolism analysis, and repetitiously intermittent bead-to-bead transfer of cell for virus production to simulate the close and continuous process.

通过使用Vero细胞增殖禽流感病毒H9N2，本文针对封闭连续工艺过程的一些关键技术开展研究，包括细胞对流感病毒敏感性分析、细胞微载体培养生产病毒工艺、细胞珠到珠转移、转移后细胞对病毒增殖敏感性验证、细胞无血清培养生产流感病毒、代谢分析、可模拟连续操作的多次间歇式珠到珠转移培养细胞生产流感病毒等方面。

Objective: To investigate the effects of sulindac on the growth and motility of ovarian cancer cell SKOV3 and its involved mechanism. Methods: In order to detect the effects of sulindac on cultured SKOV3, sulindac with different concentration and untreated control were set up. The changes in morphology of SKOV3 were observed by phase contrast microscopy. MTT colorimetric assay was used to study the effects of sulindac on SKOV3 cell growth. Motility of SKOV3 was determined by scarification assay.

目的：研究舒林酸对卵巢癌SKOV3细胞的生长和细胞运动的影响以及对基质金属蛋白酶2（MMP2）、环氧合酶-2(COX-2)表达的影响：方法：将不同浓度的舒林酸加入培养液中观察其对SKOV3细胞形态的影响，采用MTT法观察舒林酸对SKOV3细胞生长的影响，通过划痕试验观察舒林酸对SKOV3细胞运动的影响，利用免疫细胞化学方法研究舒林酸对SKOV3细胞MMP2、COX-2表达的影响。

Methods: in order to detect the effects of sulindac on cultured skov3, sulindac with different concentration and untreated control were set up. the changes in morphology of skov3 were observed by phase contrast microscopy. mtt colorimetric assay was used to study the effects of sulindac on skov3 cell growth. motility of skov3 was determined by scarification assay.

目的：研究舒林酸对卵巢癌skov3细胞的生长和细胞运动的影响以及对基质金属蛋白酶2（mmp2）、环氧合酶-2(cox-2)表达的影响：方法：将不同浓度的舒林酸加入培养液中观察其对skov3细胞形态的影响，采用mtt法观察舒林酸对skov3细胞生长的影响，通过划痕试验观察舒林酸对skov3细胞运动的影响，利用免疫细胞化学方法研究舒林酸对skov3细胞mmp2、cox-2表达的影响。

Results:(1)NSCs form typical neurospheres under adequate concentration in vitro, which are immunoreactive to Vimentin. Typically and terminally differentiated mature neural cells could not be found without the stimulus of mitogen or only under NSCs self-regulation and self-induction;(2)NSCs derived from hippocampus maintain the character of stem cells much longer with better biological behavior; NSCs passed to the 2-3 passage are the best to graft since they have not differentiated;(3)NSCs cultured in vitro could self-regulate and differentiate into neurospheres and progenitors positively immunoreactive to specific antibodies representing neurons, astrocytes, oligodendrocytes and Schwann cells;(4)There are widespread synaptic contacts between various kinds of descendent clones and cells;(5)Neurospheres could be formed without the stimulus of mitogen when NSCs and OECs are cocultured. Many neurospheres and cells immunoreactive to Vimentin, GFAP, MAP2, 02, p75NGFR, GFAP, S-100, Synaptosis, Vimentin, Tau (Tau is only positive in cocultureof HNSCs+HOECs) could be found;(6)The supernatant fluid triturated from adult rat spinal cord stimulates NSCs to differentiate into neurons, but do not terminally differentiate;(7)Fibroblasts and O4 oligodendrocytes are not supported to grow under this culture medium.Part II: Isolation, culture and identification of rat and human olfactory ensheathing cellsOlfactory ensheathing cells/glials are the most powerful cells to enable the regeneration of axons in the central nervous system.

结果表明：①在适宜的浓度体外培养条件下，NSCs能形成典型的神经干细胞克隆球，Vimentin免疫荧光染色阳性，单靠丝裂原刺激或NSCs自我调节和分化诱导，不会产生典型的终末分化的成熟神经细胞；②海马源性的NSCs维持干细胞特性的时间更长，生物学特性更优；③传至第2～3代的NSCs尚未分化时移植最佳；④体外培养的NSCs能自我调控分化为神经元、星形胶质细胞、O2少突胶质细胞、雪旺氏细胞染色阳性克隆球和前体细胞；⑤各种子代克隆球和细胞存在广泛的突触联系；⑥NSCs与OECs联合培养时，不需丝裂原刺激即能形成克隆球，获得大量Vimentin、GFAP、MAP2、O2、p75NGFR、GFAP、S-100、Synaptosis、Vimentin、Tau(Tau只有人HNSCs+HOECs联合培养时出现阳染)染色阳性的克隆球和细胞；⑦脊髓研磨后的上清液刺激神经干细胞向神经元方向分化，但并不出现终末分化；⑧本研究培养条件不利于成纤维细胞、O4生长。

Inaddition,we extract total RNA from cultural U251 cell of humanglioma,make coding gene extron amplification of TfR through ReverseTranscription-Polymerase Chain Reaction,construct expressionplasmid of pCDNA3.1-TfR after digestion of enzyme andconjunction,do transfection to U251 cell by liposome afteridentification,cause excess expression,detect the efficiency of transientexpression of TfR protein by pEGFPN1-TfR, sieve positive cell ofpCDNA3.1-TfR through G418, choose positive cell clone, detect theexpression of TfR in anteroposterior period through Western blot,FCM,cell immunochemistry and mRNA through RT-PCR, identify the effect of transfection.

另外，培养人脑胶质瘤U251细胞，进行细胞总RNA抽提，运用RT-PCR方法进行TfR基因的完整编码区外显子的扩增，经过酶切、连接后构建pCDNA3.1-TfR的表达质粒，鉴定正确后按脂质体转染方法转染人胶质瘤U251细胞，使其过量表达，通过pEGFPN1-TfR检测其瞬时转染的效率；用G418对稳定表达pCDNA3.1-TfR的细胞进行筛选，挑选阳性细胞克隆，运用Western blot、流式细胞检测技术和细胞免疫化学以及mRNA半定量的方法检测TfR在转染前后的表达水平，鉴定转染的效果。

METHODS: A series of N-octyl-N'-succinyl chitosan labeled with fluorescein, were incubated with HepG2 liver cancer cells, K562 leukemia cells, A549 human lung cancer cells, and BGC cancer cells respectively and followed by using flow cytometry and enzyme-linked immunosorbent assay to assess the affinity and inhibition of N-octyl-N'-succinyl chitosan for the four kinds of tumor cells already mentioned.

用异硫氰酸荧光素对N-正辛基－N'-琥珀酰基壳聚糖进行标记，再分别与人肝癌细胞(HepG2)、人白血病细胞(K562)、人非小细胞肺癌细胞(A549)及人胃癌细胞共培养，通过流式细胞仪及酶联免疫检测仪测定N-正辛基－N'-琥珀酰基壳聚糖对肿瘤细胞的亲和性及抑制力。

Then we transfected transitorily the recombinant of green fluorescent protein gene and middle molecular weight neurofilament cDNA into wide type N2a (N2a/wt) and N2a/tau40 to observe the effect of tau accumulation on GFP-NFM fusion protein transport in cellular processes in living cells. At last we used an apoptotic inducer, camptothecin (an inhibitor of topoisomerase-1) to treat N2a/wt and N2a/tau40 cell lines, and compared their apoptotic response.

主要结果如下：一、tau蛋白过度表达和聚积对细胞形态的影响：倒置显微镜下观察两种细胞的形态，发现N2a/wt细胞的突起多而长，而N2a/tau40细胞胞体变圆，突起明显缩短；免疫印迹结果显示转染了tau40的细胞内tau的免疫反应约增加14倍，免疫荧光结果显示N2a/tau40细胞胞体内呈现出较强的红色荧光，tau主要分布在核周和突起起始部分的胞质内，而N2a/wt细胞内的荧光很弱。

The results showed that (1) the positive staining of ChAT was obviously observed in the cells of the three kinds of human pituitary adenoma, however, it was lower than that in normal human pituitary gland;(2) ACh had a similar effect on the proliferation of the three kinds of human pituitary adenoma cells. ACh at 0.1～10 μmol/L decreased the ［3H］TdR incorporation and the MTT A value in a dose-dependent manner. At the same time, ACh decreased the ratio of S or G2 phase pituitary adenoma cells significantly, but increased the ratio of G1 phase pituitary tumour cells markedly;(3) the effect of acetylcholine on the proliferation of human pituitary adenoma cells was inhibited by atropine, but not by tubocurarine;(4) ACh had no effect on the apoptosis of human pituitary adenoma cells cultured in vitro.

结果发现：(1)三种垂体腺瘤细胞中均有胆碱酯酶的表达，但明显少于正常垂体；(2)ACh对三种类型的人垂体腺瘤细胞增殖代谢的影响相类似，不同浓度的ACh能明显抑制体外培养的三种人垂体腺瘤细胞的增殖，呈明显的剂量效应关系，同时ACh能减少垂体腺瘤细胞进入S、G2期的细胞比例，而使处于G1期的细胞比例增加；(3)ACh的这种作用可被阿托品阻断，但不受筒箭毒的影响；(4)ACh对体外培养的三种人垂体腺瘤细胞凋亡无明显影响。

Cases indicated middle cell unimodal distribution and expanded from middle to small as well as large cell areas, and 27.0% cases indicated abnormally large cell unimodal distribution） while remaining 18.9% cases showed bimodal distribution.

急性淋巴细胞白血病，白细胞直方图均表现为小细胞峰极度增高，呈单峰分布并向中间细胞区扩展变宽；急性髓系细胞白血病，81.1％白细胞直方图呈单峰分布（其中54.1％表现为中间细胞单峰增高并向小细胞区和大细胞区扩展，27.0％以大细胞单峰极度增高），18.9％呈双峰分布。

But in 2007, scientists made a revolutionary breakthrough. Three different groups simultaneously announced that they had converted unipotent, mature skin cells back into an undifferentiated state. In other words, they had created cells that behaved like embryonic stem cells without using any embryo derived materials. These new cells, dubbed "induced pluripotent stem cells" or IPS's could then be converted into a variety of different cell types, such as pancreas, muscle or brain cells. The IPS's were created by inserting certain genes into the DNA of the adult cells.

不过，在2007年，科学家已经取得了重大突破，三个不同组织同时发表声明称：已经能将成熟皮细胞转化回心肌细胞，也就是说不通过胚胎组织就能让细胞具有胚胎干细胞的属性，能够让多种不同类型的细胞细胞如胰腺细胞，肌肉细胞或脑细胞重新转化为人造胚胎干细胞，将某种组织注入成体细胞DNA中便能产生IPS'S。

With Death guitarist Schuldiner adopting vocal duties, the band made a major impact on the scene.

随着死亡的吉他手Schuldiner接受主唱的职务，乐队在现实中树立了重要的影响。

But he could still end up breakfasting on Swiss-government issue muesli because all six are accused of nicking around 45 million pounds they should have paid to FIFA.

不过他最后仍有可能沦为瑞士政府&议事餐桌&上的一道早餐，因为这所有六个人都被指控把本应支付给国际足联的大约4500万英镑骗了个精光。