MMP14单克隆抗体对口腔鳞状细胞癌增殖、凋亡的影响

[目的]探究MMP14单克隆抗体对口腔鳞状细胞癌细胞增殖、凋亡的影响。[方法]构建MMP14敲低(Kd-TCa8113)、过表达(Kd-TCa8113)细胞株，与野生型TCa8113(Wt-TCa8113)对比，考察MMP14对TCa8113细胞增殖、凋亡的影响。用不同浓度的MMP14单克隆抗体与TCa8113共培养，以探讨MMP14单克隆抗体对TCa8113细胞增殖、凋亡的影响。[结果]以Wt-TCa8113为对照组，Kd-TCa8113组、Kd-TCa8113组的细胞培养48 h的增殖率分别为74.55%、114.79%,P均<0.05。流式细胞术结果显示，Wt-TCa8113、Kd-TCa8113和Oe-TCa8113培养48 h后的细胞凋亡率分别为3.47%±0.36%、33.68%±7.33%和1.60%±0.22%,P均<0.05。用50μg/mL、100μg/mL和200μg/mL浓度的抗体处理TCa8113细胞24 h后，以PBS处理组为对照，细胞增殖率分别为93.52%、90.75%和87.61%,P均<0.05。流式细胞术结果显示，PBS组和上述抗...     

喜树碱-氟尿苷纳米颗粒对口腔鳞癌细胞Tca-8113增殖与迁移的影响

目的:探究喜树碱-氟尿苷(CPT-FUDR)纳米颗粒对口腔鳞癌Tca-8113细胞增殖与迁移的影响。方法:制备喜树碱-氟尿苷纳米颗粒,通过丁达尔现象证明已组装完毕。将制备好的纳米颗粒组和喜树碱(CPT)单药组、氟尿苷(FUDR)单药组以及两种单药混合组(CPT/FUDR)作对比,采用MTT实验检测药物对口腔鳞癌细胞Tca-8113增殖的抑制作用,通过划痕实验探究CPT-FUDR纳米颗粒和CPT/FUDR混合药物对细胞迁移能力的影响。结果:MTT结果显示:在药物浓度大于0.1μM时,随着浓度的增加,四组细胞存活率均明显下降(P0.05),而CPT-FUDR纳米颗粒组Tca-8113细胞的存活率明显低于单药CPT、FUDR和CPT/FUDR混合物组(P0.05)。在划痕实验中,培养48 h后,CPT/FUDR混合物组和CPT-FUDR纳米颗粒组均显著低于空白组(P0.05),且CPT-FUDR纳米颗粒组显著低于CPT/FUDR混合物组(P0.05)。结论:在体外,CPT-FUDR纳米颗粒对口腔鳞癌Tca-8113细胞的增殖与迁移有较好的抑制作用,且抑制效果优于CPT/FUDR两种单药混合。     

三氧化二砷联合氨氯地平对肝癌HepG2细胞体外作用的实验研究

目的:探讨三氧化二砷与氨氯地平单独用药及联合用药对肝癌Hep G2细胞增殖和凋亡的作用。方法:用不同浓度的三氧化二砷(4.0、2.0、1.0μmol/L)和氨氯地平(27×103、18×103、12×103 mg/m L)处理体外培养的人肝癌Hep G-2细胞,观察细胞形态的变化,采用CCK-8法检测两种药物单独及联合应用对Hep G-2细胞生长增殖的影响,并通过流式细胞术观察其对细胞凋亡及细胞周期的影响。结果:三氧化二砷和氨氯地平均可以浓度依赖性方式显著增加Hep G2细胞的生长抑制率及其凋亡率,以联合用药的作用较单独用药更显著(P0.05)。与三氧化二砷和氨氯地平单独用药相比,联合用药可显著阻滞Hep G2细胞于G2期及S期。结论:三氧化二砷和氨氯地平均可显著抑制人肝癌Hep G2细胞的生长和增殖,并促进其凋亡,且二者联合应用时具有协同作用。     

伊立替康联合去甲斑蝥素对人胃癌细胞作用的考察及协同作用机制研究

研究了伊立替康(CPT-11)联合去甲斑蝥素(NCTD)对人胃癌细胞BGC-823增殖的影响及协同作用机制.分别利用CPT-11 30、60、90、120、150μmol/L,NCTD 30、60、90、120、150μmol/L和两药按上述浓度1∶1联用,以及两药上述浓度完全交叉组合作用BGC-823细胞24、48和72 h,并且以不同序贯方式作用BGC-823细胞24 h.MTT法检测BGC-823细胞的增殖,采用中效原理评价两药联合作用;流式细胞术测定CPT-11 60μmol/L、NCTD 60μmol/L和联合用药(60∶60)μmol/L,以及不同序贯方式作用BGC-823细胞24 h后细胞周期及凋亡的情况;Western blot法检测CPT-11 30、60μmol/L,NCTD 30,60μmol/L和联合用药(30∶30,60∶60)μmol/L作用BGC-823细胞24 h后Pdcd4和p53蛋白表达水平.结果显示:CPT-11及NCTD联合用药比单独用药对细胞增殖的抑制作用增强,IC50明显减小(P0.05),单独使用CPT-11或NCTD作用24、48和72 h时的IC50分别是联合使用时的2.83、3.15、2.19倍以及2.66、3.11、2.45倍,并且两药联合用药具有协同作用效应;两药合用24 h时先给CPT-11方案抑制作用优于先给NCTD方案,且优于同时给药方案(P0.05);CPT-11 60μmol/L作用24 h引起BGC-823细胞S和G2-M期阻滞(P0.01),NCTD 60μmol/L作用24 h引起细胞G2-M期阻滞(P0.05),并诱导细胞凋亡(P0.05),联合用药(60∶60)μmol/L作用24 h,主要引起细胞G2-M期阻滞(P0.01),与先给NCTD 6h方案及同时给药相比,先给CPT-11 6h方案主要引起细胞S期阻滞增加(P0.05)以及细胞凋亡增加;CPT-11 30、60μmol/L作用12 h后Pdcd4上调表达,p53下调表达(P0.05),NCTD 30、60μmol/L作用12 h后Pdcd4下调表达,p53上调表达(P0.05),联合用药(30∶30,60∶60)μmol/L作用24 h后Pdcd4及p53蛋白表达均上调(P0.05).上述结果表明:CPT-11联合NCTD对人胃癌BGC-823细胞有协同抑制作用,其机制主要是诱导细胞G2-M期阻滞,并与抑癌基因蛋白Pdcd4及p53上调表达有关;两种药物联用时序贯次序对联合作用有影响,先给CPT-11方案要优于先给NCTD方案及同时给药方案,其机制主要是引起S期阻滞增加以及细胞凋亡增加;抑癌基因蛋白Pdcd4与p53之间可能存在负调控关系.     

碳酸酐酶CA9改变胞内pH影响舌癌的化疗

平阳霉素PYM（pingyangmycin）是舌癌最常用的化疗药物之一,而碳酸酐酶（carbonic anhydrase IX precursor, CA9）参与舌癌细胞对PYM的耐受. 有迹象表明,这种耐受的产生与CA9调节胞内pH的功能有关. 本文拟从此现象出发,探讨CA9的胞内pH调节功能对舌癌细胞化疗的影响. 以BCECF AM探针法评估实验细胞胞内pH的变化,并通过对比PYM与CA9抑制剂乙酰唑胺（acetazolamide,Atz）处理Tca8113/PYM细胞、PYM处理的Tca8113细胞及PYM处理的Tca8113/PYM细胞各组细胞中细胞凋亡标记物caspase 3的活化状态,以评估CA9的胞内pH调节功能对PYM诱导舌癌细胞凋亡的影响. Hoechst染色观察表明,经PYM和Atz共处理Tca8113/PYM细胞及经PYM处理24 h后的Tca8113细胞均出现细胞染色质固缩、细胞核呈碎石状和线粒体空泡化现象,表现出细胞凋亡状态,而对照组未见明显变化. Western印迹检测发现,经PYM处理的Tca8113细胞和经PYM与Atz共处理的Tca8113/PYM细胞中凋亡标志物caspase 3均处于活化状态,BCECF-AM探针检测发现,其胞内pH明显降低,对照组不见上述诸状. 由此可见,CA9在胞内pH调节中发挥重要作用,使用Atz抑制CA9活性,将会引起胞内pH值降低,促使PYM耐受舌癌细胞对PYM化疗增敏,可能有助于提高化疗效果.     

五步蛇毒AAVC-I对人口腔鳞癌细胞的增殖抑制作用

目的观察祁门五步蛇毒抑瘤组分I(Agkistrodon acutus venom component-I, AAVC-I)对人口腔鳞癌Tca8113细胞的增殖抑制及凋亡的影响。方法以离子交换和凝胶过滤法从安徽祁门五步蛇粗毒中分离纯化五步蛇毒AAVC-I。常规培养口腔鳞癌Tca8113细胞,观察正常细胞形态学。实验分为对照组(加入等量细胞培养液)和实验组(分别加入AAVC-I 2.5、5.0、10.0μg/ml)。采用MTT法检测不同浓度(2.5、5.0、10.0μg/ml)的AAVC-I对Tca8113细胞24、48及72 h的增殖抑制率,倒置显微镜观察不同浓度AAVC-I处理后的细胞形态变化,透射电镜及AO/EB双荧光染色观察AAVC-I对细胞凋亡的影响,并与对照组进行比较。结果与对照组比较,实验组AAVC-I对Tca8113细胞增殖具有明显抑制作用(P0.05);AAVC-I处理后,倒置显微镜下可见明显细胞凋亡现象,电镜下可见细胞核染色质、胞质浓缩及凋亡小体;AO/EB双荧光染色可见细胞膜完整,细胞核致密浓染,染成黄绿色荧光的早期凋亡细胞及细胞膜破损,细胞核浓聚及偏移,呈橘红色荧光的晚期凋亡细胞。结论祁门五步蛇毒AAVC-I能抑制人口腔鳞癌Tca8113细胞生长增殖,诱导其凋亡。     

康莱特联合顺铂对宫颈癌SiHa细胞增殖和凋亡的影响

目的:通过康莱特联合顺铂对宫颈癌SiHa细胞增殖和凋亡的影响,探讨其作用机制。方法:体外培养宫颈癌Siha细胞,分别将康莱特(浓度为1,2,4,6,8 mg/mL),顺铂(浓度梯度为1.5,3,6,9,12μg/mL),单独作用于宫颈癌SiHa细胞,加药24h、48h用噻唑蓝(MTT法)检测细胞增殖情况。用流式细胞术检测康莱特组和顺铂组细胞24h凋亡率,选取合适的药物浓度(康莱特6 mg/mL,顺铂3μg/mL),进行联合用药,加药24h、48h用MTT法检测细胞增殖情况,用流式细胞术检测24h细胞凋亡率。结果:①MTT法显示加药后两组的24h、48h,宫颈癌SiHa细胞的抑制率均高于对照组(P0.05),并且在一定程度上呈浓度和时间依赖性。②联合用药时,细胞的抑制率和凋亡率要显著高于单独用药(P0.01)。结论:康莱特、顺铂单独或联合作用均能抑制SiHa细胞的增殖,促进其凋亡,且康莱特联合顺铂的作用要显著高于单独用药,康莱特与化疗药物联合使用可提高肿瘤细胞对化疗药物的敏感性。     

γ-生育三烯酚联合亚砷酸对急性早幼粒细胞NB4生长的抑制作用及机制

目的:研究γ-生育三烯酚联合亚砷酸对急性早幼粒细胞NB4生长的抑制作用及可能的分子机制。方法:采用CCK-8、细胞周期实验检测细胞增殖、利用激光共聚焦显微镜、Annexin V/PI染色、Caspase活性检测、Western Blot等方法测定细胞凋亡。采用1μmol/L ATO及不同浓度(0、15、30、45μmol/L)的γ-生育三烯酚处理NB4细胞24、48和72小时,通过CCK-8检测细胞的增殖情况,流式细胞术、激光共聚焦显微镜检测细胞周期和凋亡情况,检测Caspase3,8,9的活性,Western Blot检测细胞中c-caspase-3、Bcl-2和survivin的蛋白表达。结果:γ-生育三烯酚联合亚砷酸显著抑制NB4细胞增殖(P0.01),且随着作用时间延长和γ-生育三烯酚浓度的增加,其增殖抑制作用增强;细胞周期阻滞在S期,S期的比例由38.21%±2.99上升到50.31%±5.03;γ-生育三烯酚联合亚砷酸诱导NB4细胞凋亡,1μmol/L ATO联合15、30μmol/L的γ-生育三烯酚处理48h后,细胞活率分别为82.27%±3.16、66.97%±3.17、12.63%±2.66;1μmol/L ATO联合30μmol/L的γ-生育三烯酚处理后,NB4细胞caspase-3,-8,-9的活性均较ATO单独用药组显著增高,c-caspase-3表达增高而Bcl-2和survivin蛋白的表达无明显变化。结论:生育三烯酚联合亚砷酸对急性早幼粒细胞NB4的生长具有抑制作用,此作用可能与抑制增殖并诱导凋亡相关,其作用靶点可能与促进Cas-pase诱导的凋亡有关。     

染料木素抑制人胰腺癌细胞株PANC-1细胞增殖机制的研究

目的探讨不同浓度染料木素对胰腺癌细胞作用及其抑癌基因表达的研究。方法本实验共分实验组(80,120,160,200μmol/L)的染料木素、空白对照组(PBS)和标准对照(吉西他滨)。各组培养基处理细胞12,24,36,48,60 h后,细胞计数试剂盒(CCK-8)法检测人胰腺癌细胞株PANC-1在染料木素作用下的增殖活性;流式细胞术检测人胰腺癌细胞株PANC-1在染料木素木素作用下的凋亡细胞百分率;用逆转录-聚合酶连反应(RTPCR)方法测定P53及P21抑癌基因的表达;Western-blot法检测抑癌基因P53和P21的蛋白表达水平。细胞凋亡与细胞基因mRNA采用的是LSD法,细胞增殖数据采用的是配对t检验。结果流式细胞术显示,人胰腺癌细胞株PANC-1处理后60 h时在不同浓度(80,120,160,200μmol/L)的染料木素作用下细胞凋亡率分别为(3.72±1.58)﹪、(35.98±3.76)﹪、(51.36±4.32)﹪和(64.10±2.09)﹪。吉西他滨(gemcitabine,GEM)组为(43.45±5.28)﹪,不同浓度染料木素组与标准对照组间比较差异有统计学意义(均P<0.01);细胞增殖效果:160μmol/L与200μmol/L、120μmol/L与200μmol/L、200μmol/L与吉西他滨组之间比较,均有统计学意义(P<0.01),而160μmol/L与吉西他滨组比较无统计学意义(P>0.05);Western-blot检测蛋白水平显示160μmol/L和200μmol/L染料木素相对于空白对照能够明显的增强抑癌基因P21和P53的蛋白水平的表达,相对于标准组没有明显的差异。结论染料木素可以抑制人胰腺癌细胞株PANC-1增殖,并且初步探索了这种抑制作用表现出浓度及时间依赖性。     

姜黄素联合顺铂对人骨肉瘤细胞增殖和凋亡的影响

目的:观察姜黄素联合顺铂对人骨肉瘤细胞MG-63增殖和凋亡的影响。方法:采用不同浓度的姜黄素、顺铂和姜黄素联合顺铂处理人骨肉瘤细胞MG-63不同时间,通过MTT法检测其对MG-63细胞生长的抑制作用;平板克隆实验检测其对MG-63细胞克隆形成能力的影响;流式细胞仪检测其对MG-63细胞凋亡的影响。结果:单用姜黄素或顺铂均可以时间和浓度依赖性方式抑制骨肉瘤细胞MG-63的增殖。与空白对照组相比,单用10μmol/L姜黄素和2、4μmol/L顺铂可抑制MG-63细胞的增殖,降低其克隆形成率并提高细胞凋亡率(P<0.05);而与姜黄素和顺铂单用处理相比,10μmol/L姜黄素分别与2、4μmol/L顺铂联合应用,可更显著抑制MG-63细胞的增殖,降低其克隆形成率并提高细胞凋亡率(P<0.01)。结论:姜黄素联合顺铂与单药相比能够显著抑制人骨肉瘤细胞MG-63细胞的增值,促进其凋亡,两药对骨肉瘤细胞的杀伤效应具有一定的协同性。     

A Disintegrin and Metalloprotease (ADAM) 10 and ADAM17 Are Major Sheddases of T Cell Immunoglobulin and Mucin Domain 3 (Tim-3)

T cell immunoglobulin and mucin domain 3 (Tim-3) dampens the response of CD4⁺ and CD8⁺ effector T cells via induction of cell death and/or T cell exhaustion and enhances the ability of macrophages to clear pathogens via binding to galectin 9. Here we provide evidence that human Tim-3 is a target of A disintegrin and metalloprotease (ADAM)-mediated ectodomain shedding resulting in a soluble form of Tim-3. We identified ADAM10 and ADAM17 as major sheddases of Tim-3 as shown by ADAM-specific inhibitors and the ADAM10 pro-domain in HEK293 cells and ADAM10/ADAM17-deficient murine embryonic fibroblasts. PMA-induced shedding of Tim-3 was abrogated by deletion of amino acids Glu¹⁸¹–Asp¹⁹⁰ of the stalk region and Tim-3 lacking the intracellular domain was not efficiently cleaved after PMA stimulation. Surprisingly, a single lysine residue within the intracellular domain rescues shedding of Tim-3. Shedding of endogenous Tim-3 was found in primary human CD14⁺ monocytes after PMA and ionomycin stimulation. Importantly, the recently described down-regulation of Tim-3 from Toll-like receptor-activated CD14⁺ monocytes was caused by ADAM10- and ADAM17-mediated shedding. Inhibition of Tim-3 shedding from lipopolysaccharide-induced monocytes did not influence lipopolysaccharide-induced TNFα and IL-6 but increases IL-12 expression. In summary, we describe Tim-3 as novel target for ADAM-mediated ectodomain shedding and suggest a role of Tim-3 shedding in TLR-mediated immune responses of CD14⁺ monocytes.     

Adenomatous Polyposis Coli (APC) in cell migration

Adenomatous Polyposis Coli (APC) protein is mostly known as a tumor suppressor that regulates Wnt signaling, but is also an important cytoskeletal protein. Mutations in the APC gene are linked to colorectal cancer and various neurological disorders and intellectual disabilities. Cytoskeletal functions of APC appear to have significant contributions to both types of these disorders. As a cytoskeletal protein, APC can regulate both actin and microtubule cytoskeletons, which together form the main machinery for cell migration. As APC is a multifunctional protein with numerous interaction partners, the complete picture of how APC regulates cell motility is still unavailable. However, some molecular mechanisms begin to emerge. Here, we review available information about roles of APC in cell migration and propose a model explaining how microtubules, using APC as an intermediate, can initiate leading edge protrusion in response to external signals by stimulating Arp2/3 complex-dependent nucleation of branched actin filament networks via a series of intermediate events.     

酸性磷酸酶法检测体外培养细胞数

利用小鼠成纤维细胞系(NIH3T3)、小鼠骨髓瘤细胞系(SP2/0)、人大肠癌细胞系(LO-VO)和人白血病细胞系(K562),评价酸性磷酸酶(APA)法用于检测体外各类型细胞的增殖和杀伤作用。用直线回归分析光吸收度与每孔活细胞数的关系。结果表明,APA法能准确地反映检测的活细胞数(相关系数均>0.99)。本方法不仅能很好地检测表皮生长因子对细胞的增殖作用,也能够检测顺铂对体外细胞的杀伤作用。结果表明APA法简单、灵敏,可以用于上皮和间质等贴壁和悬浮生长的细胞计数。     

SEC14 and Spectrin Domains 1 (Sestd1) and Dapper Antagonist of Catenin 1 (Dact1) Scaffold Proteins Cooperatively Regulate the Van Gogh-like 2 (Vangl2) Four-pass Transmembrane Protein and Planar Cell Polarity (PCP) Pathway during Embryonic Development in Mice

The planar cell polarity (PCP) pathway is a conserved non-canonical (β-catenin-independent) branch of Wnt signaling crucial to embryogenesis, during which it regulates cell polarity and polarized cell movements. Disruption of PCP components in mice, including Vangl2 and Dact1, results in defective neural tube closure and other developmental defects. Here, we show that Sestd1 is a novel binding partner of Vangl2 and Dact1. The Sestd1-Dact1 interface is formed by circumscribed regions of Sestd1 (the carboxyl-terminal region) and Dact1 (the amino-terminal region). Remarkably, we show that loss of Sestd1 precisely phenocopies loss of Dact1 during embryogenesis in mice, leading to a spectrum of birth malformations, including neural tube defects, a shortened and/or curly tail, no genital tubercle, blind-ended colons, hydronephrotic kidneys, and no bladder. Moreover, as with Dact1, a knock-out mutation at the Sestd1 locus exhibits reciprocal genetic rescue interactions during development with a semidominant mutation at the Vangl2 locus. Consistent with this, examination of Wnt pathway activities in Sestd1 mutant mouse embryonic tissue reveals disrupted PCP pathway biochemistry similar to that characterized in Dact1 mutant embryos. The Sestd1 protein is a divergent member of the Trio family of GTPase regulatory proteins that lacks a guanine nucleotide exchange factor domain. Nonetheless, in cell-based assays the Sestd1-Dact1 interaction can induce Rho GTPase activation. Together, our data indicate that Sestd1 cooperates with Dact1 in Vangl2 regulation and in the PCP pathway during mammalian embryonic development.     

Histone Deacetylase (HDAC) 10 Suppresses Cervical Cancer Metastasis through Inhibition of Matrix Metalloproteinase (MMP) 2 and 9 Expression

Aberrant expression of histone deacetylases (HDACs) is associated with carcinogenesis. Some HDAC inhibitors are widely considered as promising anticancer therapeutics. A major obstacle for development of HDAC inhibitors as highly safe and effective anticancer therapeutics is that our current knowledge on the contributions of different HDACs in various cancer types remains scant. Here we report that the expression level of HDAC10 was significantly lower in patients exhibiting lymph node metastasis compared with that in patients lacking lymph node metastasis in human cervical squamous cell carcinoma. Forced expression of HDAC10 in cervical cancer cells significantly inhibited cell motility and invasiveness in vitro and metastasis in vivo. Mechanistically, HDAC10 suppresses expression of matrix metalloproteinase (MMP) 2 and 9 genes, which are known to be critical for cancer cell invasion and metastasis. At the molecular level, HDAC10 binds to MMP2 and -9 promoter regions, reduces the histone acetylation level, and inhibits the binding of RNA polymerase II to these regions. Furthermore, an HDAC10 mutant lacking histone deacetylase activity failed to mimic the functions of full-length protein. These results identify a critical role of HDAC10 in suppression of cervical cancer metastasis, underscoring the importance of developing isoform-specific HDAC inhibitors for treatment of certain cancer types such as cervical squamous cell carcinoma.     

Estradiol protects adipose tissue-derived stem cells against H(2) O(2) -induced toxicity

Oxidative stress is associated with various pathophysiological processes, including cell survival, adhesion, apoptosis, and cancer. In the present study, we aimed to evaluate the effects of H₂O₂‐induced toxicity on adipose tissue–derived stem cells (ADSCs) and whether 17β‐estradiol (E₂) has protective effects on these cells. ADSCs derived from adult Sprague–Dawley rats were pretreated with different doses of E₂ for 24 h and then exposed to 200 µM H₂O₂ for 4 h. Incubation of ADSCs with H₂O₂‐decreased cell viability in a concentration‐dependent fashion (p < 0.0001), whereas pretreatment of these cells with E₂ significantly reversed toxicity (p < 0.05), inhibited apoptotic changes, and decreased lipid peroxidation (p < 0.0005). Our findings suggest that E₂ protects ADSCs from oxidative‐induced cell death, and therefore, it may be used to improve the survival rate and regenerative capacity of stem cells. © 2012 Wiley Periodicals, Inc. J Biochem Mol Toxicol 26:301–307, 2012; View this article online at wileyonlinelibrary.com . DOI 10.1002/jbt.21421     

Elevated CO(2) levels cause mitochondrial dysfunction and impair cell proliferation

Elevated CO(2) concentrations (hypercapnia) occur in patients with severe lung diseases. Here, we provide evidence that high CO(2) levels decrease O(2) consumption and ATP production and impair cell proliferation independently of acidosis and hypoxia in fibroblasts (N12) and alveolar epithelial cells (A549). Cells exposed to elevated CO(2) died in galactose medium as well as when glucose-6-phosphate isomerase was knocked down, suggesting mitochondrial dysfunction. High CO(2) levels led to increased levels of microRNA-183 (miR-183), which in turn decreased expression of IDH2 (isocitrate dehydrogenase 2). The high CO(2)-induced decrease in cell proliferation was rescued by α-ketoglutarate and overexpression of IDH2, whereas proliferation decreased in normocapnic cells transfected with siRNA for IDH2. Also, overexpression of miR-183 decreased IDH2 (mRNA and protein) as well as cell proliferation under normocapnic conditions, whereas inhibition of miR-183 rescued the normal proliferation phenotype in cells exposed to elevated levels of CO(2). Accordingly, we provide evidence that high CO(2) induces miR-183, which down-regulates IDH2, thus impairing mitochondrial function and cell proliferation. These results are of relevance to patients with hypercapnia such as those with chronic obstructive pulmonary disease, asthma, cystic fibrosis, bronchopulmonary dysplasia, and muscular dystrophies.     

Effects of sodium tungstate on the ultrastructure and growth of pea (Pisum sativum) and cotton (Gossypium hirsutum) seedlings

Pea (Pisum sativum L. cv. Onmard) and cotton (Gossypium hirsutum L. cv. Campo) seedlings were treated with two concentrations (200 and 500 mg/l) of sodium tungstate (Na₂WO₄) and the developmental effects were investigated. Tungstate retarded seedling growth rate and stopped root elongation in both species. Seedling growth recovered when tungstate was removed, but primary roots continued to be stunted, while lateral root initiation and growth were stimulated. Tungstate induced premature vacuolation in cells of the root apical meristem, with vacuoles having an unusual semi-circular or cap-like shape around the nucleus. In control roots, the nuclei were spherical with prominent nucleoli bearing several randomly distributed fibrillar centres. In the tungstate-treated cells nuclei contained spherical nucleoli with a big nucleolar vacuole. Occasionally, cytoplasmic components, such as mitochondria, were entrapped in the nucleoplasm of interphasic cells of the treated roots. In these roots, most cell plates were fused to only one lateral parental wall suggesting a non-uniform centrifugal extension. The vesicles in these cell plates were dark and fused to each other at a much lower rate than in the dividing cells of the untreated seedlings. Phragmoplast and cortical microtubules were abundant in the untreated cells, but scarcely detected in the treated ones. All these observations are consistent with the view that tungstate causes considerable toxic effects to pea and cotton seedlings.