一磷酸腺苷激酶在肿瘤细胞增殖及自噬中的研究进展

一磷酸腺苷激酶(AMPK)以异质三联体形式存在于所有真核细胞内,是维持机体能量稳态的感受器。AMPK的激活主要来自avicinD、LKB1等,而AMPK主要激活底物是TSC2、p53和p27kip1,其中TSC2是mTOR的负性调控元件,它的激活可以间接地抑制细胞增殖;p53也可抑制mTOR通路,中断细胞增殖;活化的p27kip1可以诱导自噬,促进细胞死亡,抑制肿瘤生长。这些都表明AMPK在肿瘤细胞增殖和自噬中发挥作用。由于在许多肿瘤细胞中可以发现失活的AMPK,因而研究AMPK与肿瘤发展及细胞自噬的关系,将可能为肿瘤治疗找到新靶点。     

去甲斑蝥素通过调控miR-182-5p 抑制卵巢癌细胞增殖和侵袭

目的:研究去甲斑蝥素(ND)对卵巢癌细胞增殖和侵袭能力的影响及分子机制。方法:CCK-8和Transwell检测不同浓度的ND给药后SKOV3细胞增殖和侵袭能力变化;qRT-PCR和Western blot检测给药前后LKB1/AMPK信号通路蛋白和上皮间充质转化(EMT)相关蛋白表达;加入LKB1/AMPK通路激动剂GSK621后,CCK-8和Transwell分别检测SKOV3细胞增殖和侵袭能力变化;qRT-PCR和Western blot检测给药前后EMT相关蛋白和LKB1/AMPK信号通路蛋白变化;si-miR-182-5p转染SKOV3细胞后CCK-8和Transwell检测miR-182-5p对SKOV3细胞增殖和侵袭能力影响;qRT-PCR和Western blot检测转染前后LKB1/AMPK信号通路蛋白和EMT相关蛋白表达。结果:30μmol/L ND对SKOV3细胞增殖抑制效果最明显;ND显著抑制SKOV3细胞侵袭同时显著激活LKB/AMPK信号通路和EMT。加入GSK621可以部分消除ND对SKOV3细胞增殖和侵袭影响,ND抑制miR-182-5p表达而激活LKB1/AMPK磷酸化和EMT。结论:ND可能通过抑制miR-182-5p阻滞LKB1/AMPK信号通路和EMT发生抑制卵巢癌细胞增殖和侵袭。     

PI3K/Akt/mTOR信号转导通路与肿瘤

近年来,信号转导通路与肿瘤已成为了研究热点,而无论在生理还是病理条件下磷脂酰肌醇3激酶和哺乳动物雷帕霉素靶蛋白(PI3K/Akt/mTOR)信号转导通路在抑制凋亡、促进增殖方面都是一条重要的信号转导通路,并且PI3 K/Akt/mTOR信号传导通路在肿瘤治疗的研究中同样也发挥着重要作用.因而,研究PI3K/Akt/mTOR信号转导通路和主要组成部分以及PI3K、Akt、mTOR抑制剂具有重要意义.     

腺苷酸活化蛋白激酶与肿瘤的靶向治疗

腺苷酸活化蛋白激酶(AMP-activated protein kinase,AMPK)是一种多方面调节细胞功能的能量感受器.AMPK可以由多种途径被激活,主要通过LKB1-AMPK-mTOR及相关通路,调节代谢、调控细胞周期.AMPK存在于肿瘤细胞中,可以抑制细胞增殖,与肿瘤的靶向治疗有着密切的关系.     

AMPK/mTOR信号通路介导的自噬在血根碱抑制鼻咽癌细胞增殖中的作用研究北大核心CSCD

目的:研究血根碱(SAN)对人鼻咽癌5-8F细胞自噬和增殖的影响,并探讨5-8F细胞自噬与增殖的关系及作用机制。方法:MTT和实时无标记细胞分析技术(RTCA)检测细胞增殖;单丹磺酰戊二胺(MDC)染色和透射电镜观察自噬情况;Western blot检测蛋白表达。结果:与Control组相比,SAN可显著抑制鼻咽癌细胞增殖,降低PCNA表达,诱导自噬小体形成,降低P62表达,提高Beclin-1表达、LC3Ⅱ/Ⅰ;而抑制自噬后(3-MA预处理),SAN诱导自噬和抑制细胞增殖作用显著减弱,同时,SAN对Beclin-1、LC3Ⅱ/Ⅰ、p62、PCNA的影响改变;SAN可激活AMPK/mTOR信号通路关键蛋白AMPK、p-AMPK并抑制mTOR表达;而抑制AMPK/mTOR信号通路后(抑制剂Compound C预处理),SAN诱导自噬和抑制细胞增殖作用显著减弱。结论:SAN能够抑制人鼻咽癌5-8F细胞增殖并诱导自噬,可能通过诱导细胞自噬抑制细胞增殖,其机制可能与激活AMPK/mTOR信号通路有关。     

Treg / Th17 代谢特点及相关通路对Treg / Th17 平衡的影响

调节性T细胞(Treg)与辅助性T细胞Th17(Th17)的动态平衡是维持免疫稳定的重要机制,与炎症、肿瘤及自身免疫疾病密切相关,近些年,越来越多的研究表明Treg与Th17具有不同的代谢方式,mTOR信号通路、AMPK信号通路及一些小分子蛋白可通过调节细胞代谢影响Treg/Th17平衡。本文作者查阅近年文献,就细胞代谢及其调控机制与Treg/Th17平衡的相互关系作一综述。     

胰升糖素样肽1对肥胖大鼠肝脏组织中Sesn2/AMPK/mTOR信号通路的干预效应

目的:研究胰升糖素样肽1受体激动剂利拉鲁肽对肥胖大鼠肝脏组织Sesn2/AMPK/mTOR信号通路的影响。方法:雄性SD大鼠随机分为正常饮食组(NC组)与高脂饮食组(HF组),喂养12周后,每组取5只评估肥胖大鼠模型建立。HF组再随机分为HF组、低剂量利拉鲁肽组(LG组)、中剂量利拉鲁肽组(MG组)与高剂量利拉鲁肽组(HG组),各组分别给予生理盐水及不同剂量利拉鲁肽(50、100和200μg/kg,皮下注射,每天2次) 4周。16周时测定体重及附睾脂肪指数;取大鼠肝脏组织HE染色观察肝脏脂肪变性情况; Western blot法检测肝脏组织中Sesn2、AMPK、p-AMPK、mTOR和p-mTOR的蛋白水平。结果:HF组大鼠的体重明显高于NC组(P 0. 01)。HF组肝脏细胞出现脂肪变性。与NC组相比,HF组的Sesn2蛋白水平明显降低(P 0. 01),p-AMPK/AMPK水平明显降低(P 0. 01),而p-mTOR/mTOR水平与NC组相比差异没有统计学显著性。利拉鲁肽干预4周后,药物处理组大鼠体重明显低于HF组(P 0. 01),MG与HG组大鼠的附睾脂肪指数较HF组降低(P 0. 01),肝组织脂肪变性较HF组减轻; HG组的Sesn2蛋白水平明显高于HF组(P 0. 01),MG与HG组的p-AMPK/AMPK水平较HF组明显升高(P 0. 01),LG组、MG与HG组p-mTOR/mTOR水平较HF组明显降低(P 0. 01)。结论:胰升糖素样肽1受体激动剂可能通过Sesn2/AMPK/mTOR信号通路影响机体能量代谢,改善肥胖状态。     

过表达LKB1基因增加子宫内膜癌对顺铂敏感性的作用及机制研究

目的肝激酶基因B1(The liver kinase B1,LKB1)对子宫内膜癌(endometrial cancer,EC)细胞顺铂(Cisplatin,DDP)化疗敏感性的影响及作用机制。方法 qRT-PCR检测LKB1在DDP耐药EC细胞株Ishikawa/DDP及子EC细胞株Ishikawa中的表达情况。过表达质粒转染Ishikawa/DDP细胞,分为oe-NC组和oe-LKB1组,MTS检测各组细胞对DDP的敏感性;Hoechst 33342凋亡染色检测DDP处理各组细胞后细胞发生凋亡变化;流式细胞仪检测DDP处理各组细胞后细胞凋亡率变化;LKB1过表达重组慢病毒及其阴性对照空载以MOI=(病毒滴度×病毒体积)/细胞数目为20分别感染Ishikawa/DDP细胞,经1.0μg/mL G418(遗传霉素)筛选,成功构建稳定过表达LKB1(oe-LKB1)或对照(oe-NC)的Ishikawa/DDP细胞,注射到BALB/c裸鼠中构建裸鼠移植瘤模型,每天给予DDP 3mg/kg灌胃,观察各组肿瘤体积变化;Western-blot法检测DDP处理各组细胞后细胞中p-AMPK、p-mTOR和Bcl-2蛋白的表达。结果 qRT-PCR结果LKB1 mRNA在DDP耐药EC细胞株Ishikawa/DDP中的表达为0.32±0.06,显著低于EC细胞株Ishikawa中的表达1.00±0.10;MTS检测结果显示过表达LKB1后DDP作用于Ishikawa/DDP的IC50值降低,对DDP的敏感性增加;Hoechst 33342凋亡染色结果显示过表达LKB1后DDP处理的细胞Hoechst 33342染色阳性细胞增多;流式细胞仪结果显示过表达LKB1后DDP处理的细胞凋亡率增加;裸鼠移植瘤模型结果显示过表达LKB1后DDP灌胃的裸鼠肿瘤体积减小;Western-blot检测发现过表达LKB1后DDP处理的细胞中p-AMPK表达增多、p-mTOR和Bcl-2蛋白表达降低(P均0.05)。结论 LKB1基因可能通过抑制AMPK/mTOR信号通路作用于Bcl-2凋亡蛋白,增加EC对DDP的化疗敏感性。     

基于AMPK/mTOR信号通路介导的自噬途径研究黄芪多糖对急性放射性肠炎大鼠肠黏膜的保护作用

目的:探究基于AMP活化蛋白激酶(AMPK)/哺乳动物雷帕霉素靶蛋白(mTOR)信号通路介导的自噬途径研究黄芪多糖(APS)对急性放射性肠炎大鼠肠黏膜的保护作用.方法:对SD大鼠进行12 Gy单次照射,制备急性放射性肠炎模型,随机分为模型组、APS(2 g/kg)组、AMPK抑制剂compound C(CC,0.2 m...     

枸杞多糖对胰岛素抵抗型多囊卵巢综合征大鼠的改善作用及LKB1/AMPK通路的调节作用研究

目的 研究枸杞多糖对多囊卵巢综合征大鼠胰岛素抵抗的改善作用及对LKB1/AMPK通路的调控效果。方法 72只大鼠随机分为空白对照组,模型组,枸杞多糖25 mg/kg、50 mg/kg、100 mg/kg组及二甲双胍组（12只/组）,采用来曲唑联合高脂饲料喂养方法建立多囊卵巢综合征大鼠模型,枸杞多糖各剂量组分别灌胃给予25 mg/kg、50 mg/kg、100mg/kg的枸杞多糖,二甲双胍组灌胃200 mg/kg的二甲双胍,1次/d,连续28 d。全自动生化分析仪测定大鼠空腹血糖、空腹胰岛素水平,并计算胰岛素抵抗指数;使用酶联免疫吸附（ELISA）试剂盒测定血清睾酮、雌二醇、促卵泡生成素及促黄体生成素水平;苏木精-伊红（H-E）染色法检查卵巢组织病理改变;实时定量聚合酶链反应（qRT-PCR）测定卵巢组织LKB1、AMPK mRNA水平;免疫印迹法（Western blot）测定卵巢组织LKB1、AMPK蛋白水平。结果 与模型组比较,枸杞多糖各剂量组及二甲双胍组空腹血糖、空腹胰岛素、胰岛素抵抗指数、血清睾酮及促黄体生成素水平均显著降低（P<0.05）,血清雌二醇及促卵泡生成素水平、...     

Heterotypic interference between influenza viruses A/Aichi/2/68 and B/Massachusetts/1/71.

Virus particles produced by MDCK cells mixedly infected with 3 PFU/cell each of A/Aichi/2/68 (H3N2) (Aichi) and B/Massachusetts/1/71 (Mass) influenza viruses exclusively possessed haemagglutinin (HA) of Mass, although approximately one-fifth of the mixed yield had coding potential for Aichi serotype. Synthesis of major viral proteins of Aichi was markedly suppressed by co-infecting Mass. By increasing the multiplicity of co-infecting Aichi to 30 PFU/cell, interference became reciprocal. Aichi interfered with replication of Mass more severely than Mass did with replication of Aichi. All the major viral proteins of both Aichi and Mass were expressed within the infected cells.     

Associations Between G/A1229, A/G3944, T/C30875, C/T48200 and C/T65013 Genotypes and Haplotypes in the Vitamin D Receptor Gene, Ultraviolet Radiation and Susceptibility to Prostate Cancer

Ultraviolet radiation (UVR) may protect against prostate cancer via a mechanism involving vitamin D. Thus, the vitamin D receptor (VDR) gene is a susceptibility candidate, though published data are discrepant. We studied the association of prostate cancer risk with five VDR single nucleotide polymorphisms (SNPs): G/A¹²²⁹ (SNP 1), A/G³⁹⁴⁴ (SNP 2), T/C³⁰⁸⁷⁵ (SNP 3), C/T⁴⁸²⁰⁰ (SNP 4) and C/T⁶⁵⁰¹³ (SNP 5), in 430 cancer and 310 benign prostatic hypertrophy (BPH) patients. The SNP 2 GG genotype frequency was lower in cancer than BPH patients (odds ratio = 0.63, 95% CI = 0.41–0.98, p = 0.039). SNPs 1 and 2, and SNPs 4 and 5, were in linkage disequilibrium. Two copies of haplotypes comprising SNPs 1‐2, G‐G (odds ratio = 0.63, p = 0.039), SNPs 2‐3 G‐C (odds ratio = 0.45, p = 0.008) and SNPs 1‐2‐3 G‐G‐C (odds ratio = 0.44, p = 0.006), but not SNPs 1‐3, G‐C (odds ratio = 0.81, p = 0.34), were associated with reduced risk (reference, no copies of the haplotypes) . These associations were observed after stratification of subjects by extent of UVR exposure. These data show that SNP 2 GG genotype mediates prostate cancer risk, complementing studies reporting this allele is protective in malignant melanoma pathogenesis. They further suggest that published associations of risk with SNP 1 may result from linkage disequilibrium with SNP 2.     

Pathogenicity of PMV-3/parakeet/Netherlands/449/75 for chickens

Infection of 1-day-old chicks with PMV-3/parakeet/Netherlands/449/75 (449) by intramuscular, intranasal or contact routes resulted in severe impairment of growth in all groups compared to uninfected control birds. In the group infected intramuscularly with 449 virus 16/22 birds died within 14 days of infection. No clinical signs were seen in 6-week-old chickens infected with 449 by intramuscular, intranasal or contact routes. One-day-old chicks infected with a large dose of NDV-B(1) and one-day-old chicks placed in contact with these birds also showed significant impairment of growth compared to uninfected controls.     

Immunogenic and isotype-specific responses to Russian and US cold-adapted influenza a vaccine donor strains A/Leningrad/134/17/57, A/Leningrad/134/47/57, and A/Ann Arbor/6/60 (H2N2) in mice

The immunogenicity of the Russian cold-adapted (ca) donor stains, A/Leningrad (Len)/134/17/57 and A/Leningrad/134/47/57, and the US strain A/Ann Arbor (AA)/6/60-ca, were compared in BALB/c mice with their respective wild-type parental viruses. Each ca donor strain was less immunogenic than its wild-type parent. The vaccinating dose, when administered twice, which prevented multiplication of a standard challenge of parental wild-type virus in 50% of mice (the 50% protective dose or PD(50)), was shown for A/Len/134/17/57-ca, A/Len/134/47/57-ca, and A/AA/6/60-ca to be 10(3.77), 10(4.32), and 10(4.70), respectively. These findings were extended by measuring the number of antibody secreting cells induced in the lungs and mediastinal lymph nodes of mice infected with the same ca donors using an ELISPOT assay. When each donor strain was administered twice at a dose of 100 PD(50) over a 3-week interval, the overall immunoglobulin isotype antibody secreting cell profiles were shown to be similar. However, A/Len/134/17/57-ca and A/Len/134/47/57-ca induced significantly higher total immunoglobulin responses in the lungs than A/AA/6/60-ca (P < 0.05). A/Len/134/17/57-ca also induced a significantly greater IgA response in the lungs than A/AA/6/60-ca (P < 0.05). These results suggest that A/Len/134/17/57-ca is a superior immunogen to A/Len/134/47/57-ca which in turn is more immunogenic than A/AA/6/60-ca.     

Recent H1N1 viruses (A/USSR/90/77, A/Fiji/15899/83, A/Firenze/13/83) replicate poorly in ferret bronchial epithelium

Summary Three recent wild-type H1N1 influenza virus isolates (A/USSR/90/77, A/Fiji/15899/83 and A/Firenze/13/83) replicated poorly in organ cultures of ferret bronchial tissue compared with the replication of an H3N2 wild-type virus (A/England/939/69). All four viruses replicated well in nasal turbinate tissue. Examination of one H1N1 virus (A/USSR/90/77)in vivo showed heavy infection in the upper respiratory tract of ferrets but little in the lower respiratory tract. These results raise the possibility that the mildness of human influenza arising from the H1N1 strains may be due to lack of capacity to attack the lower respiratory tract as well as the presence of antibody in previously exposed persons.With 1 Figure