紫草素对人肝癌HepG2细胞增殖的抑制及诱导凋亡作用

目的:观察紫草素抑制人肝癌HepG2细胞增殖及凋亡诱导的作用。方法:用不同浓度的紫草素处理HepG2细胞,MTT检测紫草素对HepG2细胞生长增殖的抑制作用;比色法测定Caspase-3酶活性;Western blot法检测磷酸化Akt蛋白(pAkt)的表达。结果:紫草素能够抑制人肝癌HepG2细胞的增殖,并呈浓度、时间依赖性,紫草素与HepG2细胞作用24小时后Caspase-3酶活性显著增强,显示紫草素诱导的调亡作用随时间的延长而增加;同时,紫草素处理HepG2细胞后,随着药物浓度的增加,磷酸化Akt蛋白表达下降。结论:紫草素可抑制人肝癌细胞HepG2的增殖,诱导HepG2细胞凋亡,凋亡机制可能与紫草素抑制PI3K/Akt信号途径有关。     

雷氏大疣蛛毒液对人肝癌HepG2细胞p21基因表达的影响

本文研究了雷氏大疣蛛毒液对人肝癌细胞株HepG2增殖抑制作用及其分子机制。采用XTT法观察到雷氏大疣蛛毒液剂量依赖抑制HepG2细胞增殖;流式细胞仪检测发现,经过雷氏大疣蛛毒液作用的HepG2细胞周期发生明显的选择性改变;RT-PCR方法检测到p21基因表达增强;Western blot检测发现,p21蛋白表达增加。结果提示,雷氏大疣蛛毒液抑制人肝癌细胞HepG2增殖的可能机制之一是使p21基因和蛋白表达增加,G2IM细胞周期被阻滞,从而诱导细胞凋亡。     

龙牙百合多糖对人肝癌细胞HepG2凋亡的影响

为研究龙牙百合多糖(LP)对人肝癌HepG2细胞抑制和凋亡的影响,采用MTT法考察了3组LP不同浓度对人肝癌HepG2细胞抑制作用,采用Annexin-V/PI双染法和Western blotting法探究了LP1和LP42个组分对人肝癌HepG2细胞凋亡作用机制。LP对人肝癌HepG2细胞抑制呈剂量依赖关系,LP4组1 mg/mL对HepG2细胞的抑制率达46.36%,效果最佳;LP1组8 mg/mL最高抑制率达43.23%;LP2组4 mg/mL最高抑制率达24.35%;LP1和LP4对HepG2细胞的晚期凋亡率显著高于早期。激活Caspase-3、Bax、蛋白表达显著升高(p0.05),Bcl-2蛋白表达水平明显降低(p0.05)。研究表明,LP通过激活死亡受体凋亡途径和线粒体途径,诱导人肝癌HepG2细胞凋亡。     

二烯丙基三硫通过线粒体依赖性途径诱导人肝癌HepG2细胞凋亡

二烯丙基三硫(diallyl trisulfide,DATS)对多种肿瘤有抗癌作用,但机制尚不完全清楚．为探讨DATS对人肝癌细胞系HepG2细胞凋亡的影响,用丫啶橙/溴化乙锭(AO/EB)法观察细胞凋亡情况,在显微镜下计数凋亡细胞数．JC-1荧光染色观察线粒体膜电势变化．Western blot法检测细胞色素c蛋白分布,ELISA法检测caspase-3活性． 结果显示,DATS诱导HepG2细胞凋亡,用 50 μmol/L 与100 μmol/L DATS处理48 h,细胞凋亡率分别达到60.33%和93.67%,并引起HepG2细胞线粒体膜电势降低．Western blot显示,DATS能诱导胞浆细胞色素c增加,与此同时,线粒体细胞色素c减少,诱导HepG2细胞caspase-3活化．提示：DATS可通过降低线粒体膜电势,促进细胞色素c由线粒体膜释放到胞浆中,激活caspase-3途径诱导人肝癌细胞系HepG2细胞凋亡．     

四逆散对人肝癌HepG2细胞增殖、凋亡的影响及其机制*

目的: 探讨含四逆散药液血清对人肝癌HepG2细胞增殖、凋亡的影响及机制。方法: 将人肝癌HepG2细胞分为5组,每组3个复孔。实验组细胞用五氟尿嘧啶(5-FU)或不同浓度的含四逆散药液血清处理48 h后,用倒置显微镜观察含四逆散药液血清处理后人肝癌HepG2细胞形态的变化;MTT法检测含四逆散药液血清对HepG2细胞生长的抑制作用;荧光染色和流式细胞术分别分析含四逆散药液血清对HepG2细胞凋亡的影响。Rho123染色法检测线粒体膜电位变化,Western blot检测细胞凋亡相关蛋白的表达。结果: 与对照组比较,含四逆散药液血清处理人肝癌HepG2细胞后,细胞数量显著减少(P＜0.01),形态发生改变,呈现典型的凋亡细胞形态;G1期细胞数明显增加,而G2 期细胞数量显著减少(P＜0.05);Bax、Caspase-3、-9和Cyt-c的表达显著升高,而Bcl-2的表达显著降低(P＜0.05);随着含四逆散药液血清浓度增大,HepG2细胞线粒体膜电位显著下降(P＜0.05)。结论: 四逆散可以抑制HepG2细胞增殖,并通过线粒体途径诱导细胞凋亡。     

NF-kB 拮抗剂PDTC 对HepG2 细胞的抑制及对caspase-3 表达的影响

目的:研究NF-kB拮抗剂PDTC诱导人肝癌HepG2细胞凋亡及其对凋亡相关基因caspase-3表达的影响,初步探讨其诱导凋亡的可能机制。方法:以不同浓度的PDTC处理人肝癌HepG2细胞,利用MTT法检测对人肝癌HepG2细胞凋亡的影响;利用RT-PCR和Western-blot检测caspase-3mRNA和蛋白的表达。结果:不同浓度PDTC作用人肝癌HepG2细胞不同时间后,能够显著抑制HepG2细胞的生长增殖,存在剂量和时间依赖性(P<0.05);PDTC能够上调HepG2细胞中caspase-3mRNA和蛋白的表达。结论:NF-kB拮抗剂PDTC对人肝癌HepG2细胞产生显著抑制,并能上调HepG2细胞中caspase-3mRNA和蛋白的表达。     

半边旗二萜类成分PAG通过ROS诱导肝癌细胞凋亡的作用研究

探讨半边旗二萜类成分Pteisolic acid G(PAG)对人肝癌细胞HepG2增殖和凋亡的影响及作用机制。用不同浓度的PAG处理HepG2细胞后,采用MTT法检测细胞存活率;采用PI单染法检测细胞周期分布;采用Annexin V-FITC/PI双染法检测细胞凋亡率;采用RT-PCR和Western Blotting检测细胞内mRNA和蛋白表达情况;采用DCFH-DA法检测细胞内ROS水平,采用ROS抑制剂乙酰半胱氨酸(NAC)评价PAG细胞增殖抑制作用对ROS的依赖性。结果表明,在24 h、48 h和72 h时,PAG可剂量依赖性地抑制HepG2细胞的增殖(p0.05),IC_(50)分别为64.8μmol/L,38.5μmol/L和24.8μmol/L;用药24 h时PAG可剂量依赖性地使HepG2细胞阻滞在G_2/M期,同时增加HepG2细胞凋亡率(p0.05);PAG可剂量依赖性地降低HepG2细胞内Bcl-2 mRNA和caspase 3、PARP、Bcl-2蛋白的表达(p0.05),增加Bax mRNA和actived-caspase 3、cleaved-PARP、Bax蛋白的表达(p0.05)。当使用1 mmol/L的ROS抑制剂NAC预处理HepG2细胞时,PAG对HepG2细胞增殖抑制作用被显著阻断。上述结果表明,半边旗二萜类成分PAG可提高Bax/Bcl-2的基因和蛋白表达比值,从而诱导肝癌细胞HepG2凋亡,该作用可能是通过升高细胞内ROS水平来实现的。     

NF-kB拮抗剂PDTC对HepG2细胞的抑制及对caspase-3表达的影响

目的：研究NF-kB拮抗剂PDTC诱导人肝癌HepG2细胞凋亡及其对凋亡相关基因caspase-3表达的影响,初步探讨其诱导凋亡的可能机制。方法：以不同浓度的PDTC处理人肝癌HepG2细胞,利用MTT法检测对人肝癌HepG2细胞凋亡的影响;利用RT-PCR和Western-blot检测caspase-3mRNA和蛋白的表达。结果：不同浓度PDTC作用人肝癌HepG2细胞不同时间后,能够显著抑制HepG2细胞的生长增殖,存在剂量和时间依赖性（P〈0.05）;PDTC能够上调HepG2细胞中caspase-3mRNA和蛋白的表达。结论：NF-kB拮抗剂PDTC对人肝癌HepG2细胞产生显著抑制,并能上调HepG2细胞中caspase-3mRNA和蛋白的表达。     

survivin在二烯丙基二硫诱导HepG2细胞凋亡中的作用

目的:研究survivin在二烯丙基二硫(diallyl disulfide,DADS)诱导HepG2细胞凋亡中的作用。方法:MTT法检测细胞的生长活性;流式细胞仪检测细胞周期;RT-PCR法检测survivin mRNA水平;Western blot法检测survivin蛋白水平。结果:用药组HepG2细胞活性与正常组相比,随着药物浓度的增加(25,50,100,200μmol/l),分别下降了9．3%、10．4%、21．6%、31．2%,HepG2细胞凋亡率分别增加0．83%、1．97%、6．0%、9．9%,低浓度(25,50μmol/l)的DADS可以诱导survivin mRNA和蛋白表达升高,而高浓度的(100,200μmol/l)DADS可以降低survivin mRNA和蛋白表达。结论:DADS诱导HepG2细胞survivin表达增加,抵抗DADS诱导HepG2细胞的凋亡作用。     

维甲酸对肝癌细胞HepG2糖基化磷脂酰肌醇特异性磷酯酶D表达及细胞生物学特性的影响

应用MTT、流式细胞仪、免疫印迹法检测全反式维甲酸(ATRA)单独或联合糖基化磷脂酰肌醇特异性磷酯酶D(GPI-PLD)特异性抑制剂1,10-二氮杂菲对肝癌细胞HepG2生物学特性的改变.ATRA使肝癌细胞HepG2 GPI-PLD基因表达及酶活性上调,并呈现剂量和时间依赖性.ATRA可抑制肝癌细胞HepG2增殖,使肝癌细胞Caspase-3表达水平显著增加,Bcl-2表达水平下调,促进肝癌细胞凋亡(P<0.05).ATRA联合1,10-二氮杂菲诱导组细胞,Bcl-2、细胞增殖活性较ATRA单独诱导组显著增强,Caspase-3、凋亡率显著下降.维甲酸可促进肝癌细胞HepG2 GPI-PLD基因表达上调,高活性的GPI-PLD有助于维甲酸抑制肝癌细胞增殖,促进肝癌细胞凋亡.     

Investigation of the mechanism of temperature sensitivity of the electroreceptors of ampullae of lorenzini

In experiments on Black Sea skates (Raja clavata), the potential of the receptor epithelium of the ampullae of Lorenzini and spike activity of single nerve fibers connected to them were investigated during electrical and temperature stimulation. Usually the potential within the canal was between 0 and –2 mV, and the input resistance of the ampulla 250–400 k. Heating of the region of the receptor epithelium was accompanied by a negative wave of potential, an increase in input resistance, and inhibition of spike activity. With worsening of the animal's condition the transepithelial potential became positive (up to +10 mV) but the input resistance of the ampulla during stimulation with a positive current was nonlinear in some cases: a regenerative spike of positive polarity appeared in the channel. During heating, the spike response was sometimes reversed in sign. It is suggested that fluctuations of the transepithelial potential and spike responses to temperature stimulation reflect changes in the potential difference on the basal membrane of the receptor cells, which is described by a relationship of the Nernst's or Goldman's equation type.I. P. Pavlov Institute of Physiology, Academy of Sciences of the USSR, Leningrad. I. M. Sechenov, Institute of Evolutionary Physiology and Biochemistry, Academy of Sciences of the USSR, Leningrad. Pacific Institute of Oceanology, Far Eastern Scientific Center, Academy of Sciences of the USSR, Vladivostok. Translated from Neirofiziologiya, Vol. 12, No. 1, pp. 67–74, January–February, 1980.     

Determination of stoichiometry of radioiodination of proteins

A sensitive method for the detection of small quantities of hydrophobic antioxidant free radical scavengers such as butylatedhydroxytoluene (BHT) and butylatedhydroxyanisole (BHA) in aqueous samples is described. The procedure involves extraction of the hydrophobic free radical scavenger into an organic solvent phase, followed by the subsequent reaction of an aliquot of this extract with the stable cation radical tris(p-bromophenyl)amminium hexachloroantimonate (TBACA). In experiments with BHT and BHA, the loss of TBACA absorbance at 730 nm was found to be linearly proportional to the amount of antioxidant added, with quantities of BHT as small as 200 pmol being easily detectable. In aqueous suspensions of dimyristoylphosphatidylcholine vesicles, assays of the aqueous BHT concentration showed that BHT partitioned strongly into the membrane phase, achieving very high BHT/phospholipid ratios. For a given concentration of BHT, partitioning into the membrane phase was greater in large, multilamellar liposomes than in either small, single-walled vesicles or in purified rat brain synaptic vesicle membranes. Direct assay of BHT and BHA in phospholipid membranes, however, was complicated by a nonspecific interaction between TBACA and the phospholipid.