SK4在甲状腺乳头状癌中的表达及其功能机制研究

采用Real-time PCR、Western blot及免疫组化检测中电导钙激活钾离子通道蛋白(intermediate-conductance calcium-activated potassium channel protein, SK4/Kca3.1/KCNN4)在甲状腺乳头状癌(papillary thyroid carcinoma, PTC)组织及细胞株中的表达情况,同时,对比研究SK4蛋白表达与PTC临床病理特征的关系。通过SK4特异性抑制剂(TRAM-34)下调SK4的表达后,采用CCK-8、平板克隆实验及Transwell实验分别检测SK4对PTC细胞株CGTHW-3增殖、集落形成及迁移能力的影响并分析其可能的分子机制。Real-time PCR、Western blot结果显示, SK4在PTC组织中的mRNA及蛋白表达量均明显高于癌旁组织(P0.01)。免疫组化结果显示, SK4的阳性率明显高于正常甲状腺组织(P0.01),而SK4蛋白表达阳性率与PTC患者性别、年龄、肿瘤大小、淋巴结转移等临床病理特征之间并无显著统计学意义(P0.05)。TRAM-34可通过阻断SK4通道抑制CGTHW-3细胞的增殖、集落形成能力及迁移能力。以上结果提示, SK4通道在PTC组织及细胞中高表达,参与调控PTC细胞的增殖及迁移过程,可能与PTC发病的分子机制有关。SK4有可能成为PTC新的治疗靶点。     

Notch-1信号通路与甲状腺乳头状癌的关系

研究Notch-1信号通路中Notch-1、NICD、Hes1、c-Myc与人甲状腺乳头状癌的关系,探讨Notch-1信号通路在甲状腺乳头状癌中的分子机制。对照人甲状腺乳头状癌及正常甲状腺组织标本各35例,采用Real Time PCR检测Notch-1、Hes1、c-Myc的mRNA表达情况,采用免疫组织化学、Western blot方法检测组织标本中Notch-1、NICD、Hes1、c-Myc蛋白的表达情况。Notch-1、Hes1在甲状腺乳头状癌中的mRNA表达水平明显降低,c-Myc mRNA的表达在甲状腺乳头状癌中升高,Notch-1、NICD、Hes1蛋白在人甲状腺乳头状癌中的表达低于正常甲状腺组织（P〈0.05）,c-Myc蛋白在甲状腺乳头状癌中的表达高于正常甲状腺组织（P〈0.05）。甲状腺乳头状癌组织标本中,Notch-1信号通路中Notch-1、NICD、Hes1在基因转录和蛋白质表达水平上均明显下调,而c-Myc的基因转录和蛋白质表达水平均升高,提示Notch-1信号通路在甲状腺乳头状癌的发生发展中起抑癌作用,c-Myc基因表达升高与Notch-1信号通路可能无关。     

DcR3在甲状腺乳头状癌中的表达及临床意义

应用RT-PCR、Westem blot、免疫组化分别检测甲状腺乳头状癌组织与癌旁正常甲状腺组织标本中DcR3mRNA及蛋白的表达情况,探讨DcR3在甲状腺乳头状癌组织中的表达及,临床意义。RT-PCR检测显示,甲状腺乳头状癌中DcR3 mRNA的表达明显高于正常甲状腺组织（P〈0．05）：Western blot提示,DcR3蛋白在甲状腺乳头状癌中表达比正常甲状腺组织高（P〈0．05）;免疫组化显示,DcR3蛋白在甲状腺乳头状癌中高表达（P〈0．05）。DcR3mRNA及蛋白质在甲状腺乳头状癌及正常甲状腺组织间的表达差异有统计学意义（P〈0．05）。DcR3基因及蛋白在甲状腺乳头状癌中高表达,提示DcR3可能促进了甲状腺乳头状癌的发生发展。     

TLR4 对人骨骼肌细胞炎症因子表达及胰岛素抵抗的影响

目的:研究TLR4对脂多糖(LPS)及Polymymin B(PMB)作用下的人骨骼肌细胞的炎症因子表达的影响及其在细胞胰岛素抵抗中的作用。方法:通过脂多糖(LPS)及Polymymin B(PMB)干预骨骼肌细胞24h,再用胰岛素刺激1h后,Real-time PCR检测检测骨骼肌细胞TLR4、MyD88、TNF-αmRNA的表达;Western blot检测TLR4,Myd88和CRP的表达;葡萄糖氧化酶法(GOD-POD法)检测细胞培养液中葡萄糖浓度。结果:TLR4高表达可以使炎症因子的表达增高,细胞培养液中的葡萄糖浓度增高;TLR4低表达可使炎症因子的表达降低,细胞培养液中的葡萄糖浓度没有明显变化。结论:TLR4调控了炎症因子的表达,继而可以引起胰岛素敏感性的改变,影响了胰岛素抵抗的发生。     

Toll样受体4通过Akt/FoxO3a/Bim信号通路参与海马神经元凋亡

为了探讨海马神经元中是否有Toll样受体4(Toll-like receptor 4,TLR4)介导的Akt/FoxO3a/Bim信号通路,及该通路在海马神经元凋亡中的作用和机制,本实验运用脂多糖(lipopolysaccharide,LPS)作用于原代培养的大鼠海马神经元,利用不同的工具药,以减弱或加强TLR4/Akt/FoxO3a/Bim通路的作用。应用CCK-8法检测细胞活力,Western blot法检测神经元p-Akt(Ser473)、Akt、p-FoxO3a(Thr32)、FoxO3a、Bim、活化的Caspase-3蛋白的表达变化,real-time PCR法检测海马神经元Bim的mRNA表达变化,免疫荧光法观察FoxO3a核易位情况,流式细胞术检测细胞凋亡率。结果显示:LPS作用于海马神经元不同时间后,各组细胞活力均下降(P0.05),且具有一定的时间依赖性;LPS作用后p-Akt(Ser473)、p-FoxO3a(Thr32)表达减少,FoxO3a易位进入胞核,而促凋亡蛋白Bim及活化的Caspase-3表达增加,且海马神经元的凋亡率也增加(P0.05);PI3K特异性抑制剂LY294002预处理后p-Akt(Ser473)、p-FoxO3a(Thr32)表达较LPS组降低,而促凋亡蛋白Bim及活化的Caspase-3表达升高,细胞凋亡率也明显升高(P0.05);TLR4抗体预处理后p-Akt(Ser473)、p-FoxO3a(Thr32)较LPS组增多,FoxO3a易位进入胞核的活动减弱,而促凋亡蛋白Bim、活化的Caspase-3及细胞的凋亡率均减少(P0.05)。以上结果表明,海马神经元中有TLR4介导的Akt/FoxO3a/Bim通路;神经元可通过该通路引起自身的凋亡。     

甲状腺乳头状癌相关致病基因及microRNAs的筛选和生物信息学分析

本研究利用美国国家生物技术信息中心(NCBI)基因表达综合数据库(GEO)下载获取甲状腺乳头状癌(papillary thyroid carcinoma,PTC)m RNA表达谱芯片(7例PTC样本VS 7例正常组样本,登录号为GSM85-215~GSM85228)及miro RNA高通量测序数据(3例PTC样本VS 3例正常组样本,GSM1388420~GSM138-8425)。通过Qlucore Omics Explorer(QOE)3.2、DAVID 6.7、STING 9.1、mi Records等分析软件对PTC差异基因及micro RNA进行综合生物信息学分析。筛选出来497个甲状腺乳头状癌相关致病基因及86个相关micro RNAs;其中,致病基因中上调表达的有257个,下调表达的有240个;micro RNAs中上调表达的有47个,下调表达的有39个。对其进行生物信息学分析发现,ERBB3、TNNT1、MYL1、POSTN基因及hsa-mi R-183-5p(ERBB4)、hsa-mi R-139-5p(SUV39H1)、hsa-mi R-152(MAML1)、hsa-mi R-1(FOXD3)、hsa-mi R-146b-5p(MEF2C)、hsa-mi R-152(DOT1L)、hsa-mi R-493-5p(FBXW7)、hsa-mi R-876-5p(PIK3R1)、hsa-mi R-183-3p(MDM2)、hsa-mi R-382-3p(CD80)、hsa-mi R-181b-5p(SIRT1)、hsa-mi R-874-5p(MTOR)、hsa-mi R-876-5p(GPRC6A)等micro RNAs-靶基因参与涉及Erb B信号通路、细胞凋亡信号通路、B细胞受体信号通路、磷脂酰肌醇代谢信号通路等,在PTC疾病发生发展中可能起着重要作用。从分子水平上分析甲状腺乳头状癌相关致病基因及micro RNAs,为揭示PTC发病机制、药物研发及临床诊断治疗提供新的思路和依据。     

Toll样受体激动剂对小鼠脾脏细胞中SARM蛋白表达的影响

目的探讨多种Toll样受体(toll-like receptor, TLR)激动剂分别刺激小鼠脾脏细胞后对SARM蛋白(sterile-α and armadillo motif-containing protein)表达的影响。方法无菌条件下收集小鼠脾脏并制成单细胞悬液,分别加入TLR1/2激动剂Pam3Cys、TLR2/6激动剂LTA、TLR3激动剂poly(I∶C)、TLR4激动剂LPS或TLR9激动剂CpG,终质量浓度均为20μg/mL,刺激时间分别为2、4、8、20和24 h,然后利用蛋白质印迹法(Western blot)检测SARM的表达。结果小鼠脾脏细胞中SARM的表达,Pam3Cys刺激后4~20 h持续上调,24 h后回落接近初始水平;LTA刺激后8~24 h持续下调;poly(I∶C)刺激后4~20 h持续上调,24 h后回落接近初始水平;LPS刺激后20~24 h持续上调;CpG刺激后24 h开始下调。结论小鼠脾脏细胞中SARM可能参与对TLR信号传导通路的调控。     

Toll样受体4介导内毒素对内皮细胞NF-κB的激活

为探讨Toll样受体4(Toll-like receptor 4,TLR4)在内毒素(LPS)对内皮细胞NF-κB激活中的作用,以LPS刺激培养的ECV-304细胞为模型,运用RT-PCR和蛋白质印迹技术检测了内皮细胞TLR4的表达及LPS对其表达的影响.同时利用基因转染和抗体阻断方法进一步观察了TLR4在LPS对内皮细胞NF-κB激活中的作用.研究发现,LPS能明显上调内皮细胞TLR4的表达,呈一定的时间和剂量依赖性.转染TLR4的功能突变体和运用抗TLR4单抗能明显抑制LPS对内皮细胞NF-κB的激活.提示TLR4介导了LPS对内皮细胞NF-κB激活,可能在LPS对内皮细胞激活/损伤效应中具有重要的地位.     

脂多糖通过核因子-κB途径下调泡沫细胞ATP结合盒转运体A1的表达

脂多糖(LPS)介导的免疫炎症反应与动脉粥样硬化(As)的发生密切相关,ATP结合盒转运体A1(ABCA1)促进细胞内胆固醇流出,具有抗As作用.观察了LPS对THP-1巨噬细胞源性泡沫细胞ABCA1表达及胆固醇流出的影响,并探讨TLR4/NF-κB信号途径和LXRs在此过程中的作用.THP-1巨噬细胞源性泡沫细胞经不同浓度LPS处理或者用LPS作用不同时间,以LPS单独或用NF-κB抑制剂对甲苯磺酰-L-苯丙氨酸氯甲基甲酮(TPCK)预处理细胞后再加入LPS处理.RT-PCR检测ABCA1、TLR4和LXRα mRNA的表达,Western blot检测ABCA1、LXRα及核内NF-κBp65蛋白的表达,液体闪烁计数器检测细胞内胆固醇流出,高效液相色谱分析细胞内总胆固醇、游离胆固醇和胆固醇酯含量.结果表明,LPS呈浓度和时间依赖性抑制ABCA1的表达,而增加TLR4 mRNA和核内NF-κBp65蛋白的表达,LPS使泡沫细胞内胆固醇流出减少,细胞总胆固醇、游离胆固醇与胆固醇酯增加,TPCK预处理后,LPS的这种作用被部分抑制,LXRα的表达不受LPS和TPCK的影响.这一结果提示,TLR4/NF-κB信号...     

EGCG通过PI3-K/Akt信号通路诱导人甲状腺乳头状癌细胞K1增殖和凋亡的影响

研究表没食子儿茶素没食子酸酯(EGCG)通过PI3-K/Akt信号通路对人甲状腺乳头状癌细胞K1增殖和凋亡的影响。利用MTT法研究不同剂量EGCG对K1细胞的增殖作用;采用流式细胞术分析EGCG对K1细胞周期和凋亡影响;Westernblot方法检测分析EGCG对人甲状腺乳头状癌K1细胞PI3-K、Akt/p-Akt、mTOR、cyclin D1、CDK4、Bcl-2/Bad、cleaved-caspase-3蛋白表达影响。MTT结果显示EGCG作用后K1细胞增殖显著受到抑制,且表现出明显的剂量依赖性和时间依赖性(P0.001);流式细胞术结果显示EGCG能够将K1细胞阻滞于G1期,并且产生剂量依赖性诱导K1细胞凋亡(P0.001);Westernblot结果显示EGCG能够上调K1细胞促凋亡蛋白Bad及cleaved-caspase-3的表达,下调PI3-K、mTOR、cyclin D1、CDK4蛋白表达,降低AKT蛋白磷酸化及抑凋亡蛋白Bcl-2表达(P0.05)。结果证明,EGCG能够通过PI3-K/Akt信号通路,诱导G1阻滞及细胞凋亡,抑制人甲状腺乳头状癌细胞K1增殖。     

A Unique Protease Cleavage Site Predicted in the Spike Protein of the Novel Pneumonia Coronavirus (2019-nCoV) Potentially Related to Viral Transmissibility

正Dear Editor,In December 2019, a novel human coronavirus caused an epidemic of severe pneumonia(Coronavirus Disease 2019,COVID-19) in Wuhan, Hubei, China(Wu et al. 2020; Zhu et al. 2020). So far, this virus has spread to all areas of China and even to other countries. The epidemic has caused 67,102 confirmed infections with 1526 fatal cases     

Curcuminoids as inhibitors of thioredoxin reductase: A receptor based pharmacophore study with distance mapping of the active site

Curcumin is the yellow pigment of turmeric that interacts irreversibly forming an adduct with thioredoxin reductase (TrxR), an enzyme responsible for redox control of cell and defence against oxidative stress. Docking at both the active sites of TrxR was performed to compare the potency of three naturally occurring curcuminoids, namely curcumin, demethoxy curcumin and bis-demethoxy curcumin. Results show that active sites of TrxR occur at the junction of E and F chains. Volume and area of both cavities is predicted. It has been concluded by distance mapping of the most active conformations that Se atom of catalytic residue SeCYS498, is at a distance of 3.56 from C13 of demethoxy curcumin at the E chain active site, whereas C13 carbon atom forms adduct with Se atom of SeCys 498. We report that at least one methoxy group in curcuminoids is necessary for interation with catalytic residues of thioredoxin. Pharmacophore of both active sites of the TrxR receptor for curcumin and demethoxy curcumin molecules has been drawn and proposed for design and synthesis of most probable potent antiproliferative synthetic drugs.     

Comparative morphology of the carpel in the Liliaceae: Hewardieae, Petrosavieae, and Tricyrteae

The young pistils in the melanthioid tribes, Hewardieae, Petrosavieae and Tricyrteae, are uniformly tricarpellate and syncarpous. They lack raphide idioblasts. All are multiovulate, with bitegmic ovules. The Petrosavieae are marked by the presence of septal glands and incomplete syncarpy. Tepals and stamens adhere to the ovary in the Hewardieae and the Petrosavieae but not in the Tricyrteae. Two vascular bundles occur in the stamens of the Hewartlieae and Tricyrtis latifolia. Ventral bundles in the upper part of the ovary of the Hewardieae are continuous with compound septal bundles and placental bundles in the lower part. Putative ventral bundles occur in the alternate position in the Tricyrteae and putative placental bundles in the opposite. position in the Petrosavieae. The dichtomously branched stigma in each carpel of the Tricyrteae is supplied by a bifurcated dorsal bundle.     

Selection with two alleles and complete dominance

For a plant selection model with frequency-independent viabilities, fertilities and selfing rates, it is shown that apart from global fixation, for certain parameter combinations a protected polymorphism and facultative fixation (either allele may become fixed according to initial frequencies) may both occur. Facultative fixation requires different selling rates for the dominant and recessive type. Protection of the polymorphism requires resource allocation for male and female function. In this connection the problem of purely genetically caused population extinction is discussed.
For general frequency dependence and regular segregation, the chances for establishment of a completely recessive gene are compared to those of a completely dominant gene. It is proven that the process of establishment of the recessive gene, despite a fitness advantage, may be considerably endangered by drift effects if random mating prevails. The recessive gene may reach the same effectivity in establishment as a dominant gene, only if the recessive homozygote mates exclusively with its own type during the period of establishment.     

Perinatal Vertical Transmission of Chikungunya Virus in Ruili,a Town on the Border between China and Myanmar

正Dear Editor,Chikungunya virus (CHIKV), an arbovirus in the family of Togaviridae, genus Alphavirus, is transmitted by the A.aegyptii or A. albopictus mosquito, and causes disease in humans characterized by fever, rash, and arthralgia (Silva and Dermody 2017; Suhrbier 2019). It was first reported in 1953 in Tanzania, and caused only a few outbreaks and sporadic cases in Africa and Asia in last century. However, in the epidemic in 2004, CHIKV acquired mutations that conferred enhanced transmission by the A. albopictus mosquito(Schuffenecker et al. 2006). Since then, it has successively caused outbreaks in Africa, the Indian Ocean, South East Asia, the South America, and Europe (Zeller et al. 2016).