迷迭香酸类似物-11通过EGFR-JNK通路抑制人胃癌MGC-803细胞增殖和迁移

目的基于EGFR-JNK通路,探讨RAA-11对人胃癌MGC-803细胞增殖和迁移的抑制作用。方法 MTT法检测RAA-11对人胃黏膜上皮GES-1细胞和人胃癌MGC-803细胞活力的影响;划痕实验检测RAA-11对人胃癌MGC-803细胞迁移能力的影响;实时荧光定量PCR检测RAA-11对人胃癌MGC-803细胞EGFR mRNA表达的影响;Western blot检测RAA-11对人胃癌MGC-803细胞凋亡相关蛋白caspase-3、Bcl-2、Bax及通路蛋白EGFR、JNK、p-JNK表达的影响。结果 MTT结果表明,与人胃黏膜上皮GES-1细胞相比,RAA-11明显抑制人胃癌MGC-803细胞的增殖(P<0.01),提示RAA-11对MGC-803细胞有选择作用;划痕实验结果提示,RAA-11能抑制人胃癌MGC-803细胞的迁移能力;实时荧光定量PCR结果提示,RAA-11降低了人胃癌MGC-803细胞EGFR mRNA的表达;Western blot结果提示,RAA-11上调人胃癌MGC-803细胞促凋亡相关蛋白caspase-3、Bax,并促进通路蛋白JNK、p-JNK的表达(P<0.01),下调抑凋亡相关蛋白Bcl-2的表达,并降低了通路蛋白EGFR的表达水平(P<0.01)。结论 RAA-11通过作用于EGFR-JNK通路,抑制人胃癌MGC-803细胞的增殖和迁移,并能够诱导其凋亡。     

迷迭香酸衍生物RAD-9通过PI3K/Akt和p38 MAPK信号通路诱导胃癌MGC-803细胞凋亡

目的研究迷迭香酸衍生物RAD-9诱导胃癌MGC-803细胞凋亡的作用及其机制。方法 MTT法观察RAD-9对胃癌MGC-803细胞增殖的抑制作用;流式细胞术检测细胞的凋亡;Hoechst 33258染色法观察RAD-9对MGC-803细胞核凋亡形态学的影响;Western blot检测RAD-9干预MGC-803细胞36 h后,对Akt、p-Akt、p38 MAPK、p-p38 MAPK蛋白及凋亡相关蛋白Bcl-2、Bax、caspase-3的影响。结果MTT结果显示,RAD-9呈时间、浓度依赖性抑制胃癌MGC-803细胞增殖;流式细胞术结果显示,RAD-9对胃癌MGC-803细胞有明显的促凋亡作用(P<0.01);Hoechst 33258染色实验结果显示,RAD-9干预胃癌MGC-803细胞36 h后,细胞核呈现典型凋亡形态学改变;Western blot结果显示,RAD-9干预胃癌MGC-803细胞36 h后,Bcl-2蛋白表达水平明显降低,Bax、caspase-3蛋白表达水平明显提高,Akt、p-Akt蛋白表达水平明显下调,p38 MAPK、p-p38 MAPK蛋白表达水平明显上调(P<0.01)。结论 RAD-9能抑制胃癌MGC-803细胞生长,且能诱导其凋亡,其机制可能与抑制PI3K/Akt和激活p38 MAPK信号通路相关。     

锰超氧化物歧化酶模拟化合物对人胃癌MGC-803细胞增殖和凋亡的影响

目的观察Mn SOD模拟化合物(Mn SODm)对人胃癌MGC-803细胞增殖和凋亡的影响。方法体外培养人胃癌MGC-803细胞,MTT法观察Mn SODm对MGC-803细胞增殖的影响;用Hoechst33258染色观察MGC-803细胞形态学的变化;Annexin V-FITC/PI检测MGC-803细胞凋亡;Western blot法检测p53、cleaved caspase-3、cleaved caspase-9、Bcl-2、Bax蛋白的表达。结果 MTT法检测结果显示1~20μg·m L~(-1)Mn SODm对MGC-803细胞有显著的抑制作用,24、48、72 h的IC50分别为10.18、6.93和5.05μg·m L~(-1);20μg·m L~(-1)Mn SODm作用细胞48 h后,细胞凋亡率为(69.33±4.07)%(P<0.01);Western blot结果显示,用5、10、20μg·m L~(-1) Mn SODm处理细胞48 h后,Bcl-2表达显著降低,同时p53、cleaved caspase-3、cleaved caspase-9和Bax表达显著增加(P<0.05或P<0.01)。结论 Mn SODm对人胃癌MGC-803细胞有明显的抑制作用,可能是通过下调Bcl-2表达,增加p53、cleaved caspase-3、cleaved caspase-9和Bax表达来诱导MGC-803细胞凋亡。     

重楼皂苷Ⅱ诱导人胃癌MGC-803细胞凋亡的体外研究

目的初步探讨重楼皂苷Ⅱ抑制人胃癌MGC-803细胞增殖并诱导其凋亡的作用及机制。方法体外培养人胃癌MGC-803细胞,CCK-8法检测不同浓度的重楼皂苷Ⅱ作用于细胞后其存活率;DAPI染色观察细胞凋亡;AV/PI双染检测细胞凋亡率;比色法检测天冬氨酸蛋白水解酶caspase-3的活性;Western blot法检测Cyt-c的蛋白表达水平。结果与正常对照组相比,重楼皂苷Ⅱ可降低MGC-803细胞的存活率,且呈剂量与时间依赖性;镜下可见细胞核破碎,具有明显凋亡特征,凋亡率随着浓度的增大而增加(P <0.01);细胞内caspase-3的活性增加(P <0.01),Cyt-c蛋白的表达量明显增加(P<0.01)。结论重楼皂苷Ⅱ可明显抑制人胃癌MGC-803细胞的增殖,并诱导细胞发生凋亡。     

金雀异黄酮对三阴乳腺癌MDA-MB-231细胞凋亡及EGFR/PI3K/Akt通路的影响

目的研究金雀异黄酮(genistein)诱导三阴性乳腺癌MDA-MB-231细胞凋亡及其机制。方法 MTT法观察金雀异黄酮对乳腺癌MDA-MB-231细胞增殖的抑制作用;Hoechst 33258染色观察金雀异黄酮对MDA-MB-231细胞核凋亡形态学的影响;qRT-PCR法观察金雀异黄酮干预MDAMB-231细胞36 h后,EGFR mRNA表达水平的变化;金雀异黄酮干预MDA-MB-231细胞36 h后,Western blot检测凋亡相关蛋白Bcl-2、Bax、caspase-3,EGFR、Akt、p-Akt蛋白的变化;Akt激活剂胰岛素(insulin)、金雀异黄酮单独及联合胰岛素干预乳腺癌MDA-MB-231细胞后,Western blot检测Akt和p-Akt蛋白表达量的变化。结果 MTT结果显示,金雀异黄酮呈时间浓度依赖性抑制乳腺癌MDA-MB-231细胞增殖;Hoechst 33258染色结果显示,金雀异黄酮干预乳腺癌MDAMB-231细胞36 h后细胞核呈现典型凋亡形态学改变;qRTPCR结果显示,经金雀异黄酮干预MDA-MB-231细胞36 h后,EGFR的mRNA表达水平明显下降(P<0.01);Western blot结果显示金雀异黄酮干预乳腺癌MDA-MB-231细胞36 h后,与对照组对比,Bcl-2、EGFR、Akt、p-Akt蛋白表达水平明显下调(P<0.01),Bax、caspase-3蛋白表达水平明显上调(P<0.01),Akt激活剂胰岛素可以明显激活p-Akt(P<0.01),金雀异黄酮可以明显下调被激活的p-Akt(P<0.01)。结论金雀异黄酮能抑制三阴乳腺癌MDA-MB-231细胞生长并诱导其凋亡,其机制可能与抑制EGFR/PI3K/Akt信号通路有关。     

虫草素对人胃癌细胞系HGC-27的影响及分子机制

目的 探讨虫草素对HGC-27细胞增殖、运动性和凋亡的影响及相关机制。方法 用不同浓度虫草素处理人胃癌细胞系HGC-27、AGS和MGC-803,利用CCK-8法检测细胞活力,集落形成试验检测细胞增殖能力,流式细胞术检测细胞凋亡情况,体外划痕试验检测细胞运动性。Western blot检测Bcl-2、Bax、ERK、磷酸化ERK（p-ERK）和糖原合成酶激酶-3β（GSK-3β）蛋白表达水平。结果 与对照组相比,虫草素处理组明显抑制细胞增殖、集落形成率、细胞运动性并诱导细胞凋亡,下调Bcl-2、GSK-3β和p-ERK蛋白表达水平,且呈浓度和时间依赖性,但对Bax蛋白表达水平没有影响。结论 虫草素通过调节ERK/GSK-3β信号通路起到抗肿瘤作用,有望成为临床干预和治疗胃癌的潜在药物。     

NF-κB和MAPK信号通路参与金雀异黄酮诱导乳腺癌MDA-MB-231细胞凋亡的体外研究

目的探讨金雀异黄酮(genistein)诱导人乳腺癌MDA-MB-231细胞凋亡的可能机制。方法采用MTT法观察金雀异黄酮对乳腺癌MDA-MB-231细胞增殖的抑制作用;集落形成法观察金雀异黄酮对MDA-MB-231细胞集落形成能力的影响;金雀异黄酮干预MDA-MB-231细胞36 h后,Western blot检测凋亡相关蛋白Bcl-2、Bax、caspase-3及NF-κB、ERK、p-ERK、JNK、p-JNK蛋白的表达。结果 MTT结果显示,金雀异黄酮呈时间、浓度依赖性抑制乳腺癌MDAMB-231细胞增殖;集落形成实验结果显示,金雀异黄酮能明显抑制乳腺癌MDA-MB-231细胞的集落形成(P<0.05);Western blot结果显示,金雀异黄酮干预乳腺癌MDA-MB-231细胞36 h后,与对照组相比,Bcl-2、NF-κB、p-ERK蛋白表达水平明显下调(P<0.05),Bax、caspase-3、p-JNK蛋白表达水平明显上调(P<0.05)。结论金雀异黄酮能抑制乳腺癌MDA-MB-231细胞生长,且能诱导其凋亡,其机制可能与抑制NF-κB、ERK,激活JNK信号转导通路有关。     

MAPK/ERK信号通路在益气解毒方水提物诱导鼻咽癌细胞凋亡中的作用

目的研究益气解毒方水提物对鼻咽癌细胞凋亡的影响,并从MAPK/ERK信号通路探讨其诱导凋亡的作用机制。方法 CCK-8法检测益气解毒方水提物对CNE1、CNE2细胞增殖的影响;Hoechst 33342染色法、JC-10染色法、荧光双染流式细胞仪检测其对CNE1、CNE2细胞凋亡的影响;Western blot法检测其对CNE1、CNE2细胞蛋白表达的影响。结果益气解毒方水提物能抑制CNE1、CNE2细胞增殖(P<0.05)、诱导凋亡(P<0.05);药物作用48 h后,Survivin、XIAP、Bcl-2表达下降,Bax表达上升,MAPK/ERK信号通路关键蛋白p-c-Raf、p-MEK、p-ERK表达下降(P<0.05);在此基础上,加入激活剂ISO和益气解毒方水提物后,与单用益气解毒方水提物相比,p-c-Raf、p-MEK、p-ERK1/2表达上调,Survivin、XIAP、Bcl-2表达增加,Bax表达下降,促凋亡效应也降低(P<0.05)。结论益气解毒方水提物可诱导鼻咽癌细胞凋亡,该效应与其抑制MAPK/ERK信号通路关键蛋白p-c-Raf、p-MEK、p-ERK1/2表达,进而下调Survivin、XIAP、Bcl-2表达,上调Bax表达有关。     

5,7,3′-三乙酰橙皮素对AA大鼠成纤维样滑膜细胞Jak2/Stat3信号通路及凋亡相关蛋白的影响

目的 研究5,7,3′-三乙酰橙皮素(5,7,3′-triacetyl hesperetin,TAHP)对佐剂性关节炎大鼠成纤维样滑膜细胞(FLS)Jak2/Stat3信号通路及凋亡相关蛋白的影响.方法 用弗氏完全佐剂诱导大鼠AA模型;MTT法检测FLS的增殖反应;Hoechst 33258染色法检测FLS的凋亡;RT-PCR法检测FLS中Jak2、Stat3、Bcl-2、Bax及Caspase-3的基因表达;Western blot法检测FLS中p-Stat3及Caspase-3的蛋白表达情况.结果 TAHP呈剂量和时间依赖性抑制FLS的增殖(P<0.05);Hoechst 33258染色结果提示TAHP可以明显地促进FLS的凋亡;同时TAHP(50,250 μmol·L-1)可以明显降低Jak2、Stat3及Bcl-2表达,而上调Bax及Caspase-3表达.Western blot结果显示,TAHP可降低p-Stat3的表达、上调Caspase-3表达.结论 TAHP可以抑制FLS增殖,其作用机制可能与抑制FLS Jak2/Stat3信号通路、促进Bax及Caspase-3表达、抑制Bcl-2的表达有关.     

血管生成素1与血管内皮生长因子165对人胃癌细胞增殖与凋亡的影响

目的研究人血管生成素1(Ang-1)、血管内皮生长因子165(VEGF165)以及两种因子相互作用对人胃癌细胞株MGC-803增殖与凋亡的影响。方法应用MTT法测定腺病毒绿色荧光蛋白(Ad-GFP)(B组)、Ad-Ang-1(C组)、Ad-VEGF165(D组)以及Ad-Ang-1+Ad-VEGF165/2(E组)对MGC-803细胞增殖的影响;另设对照组(A组)。采用流式细胞术分析血清饥饿时其对凋亡的影响;运用Western blot方法检测Bcl-2和Bax蛋白的表达。结果 24、48、72 h时,C、D、E组吸光度均明显高于B、A组(P<0.05);E组吸光度明显高于C、D组(P<0.05)。与A组或B组相比,C、D、E组均能够抑制细胞凋亡(P<0.01),其中E组抑制凋亡作用最强。C、D、E组Bcl-2蛋白表达均比B、A组明显增高(P<0.05),Bax蛋白的表达则明显降低(P<0.05)。结论 Ang-1、VEGF165和Ang-1+VEGF165/2能够明显促进MGC-803细胞体外增殖,抑制血清饥饿时的凋亡;Ang-1与VEGF165联合作用要显著强于单用Ang-1或VEGF165。     

Analgesia induced by brief footshock is inhibited by 5-hydroxytryptamine but unaffected by antagonists of 5-hydroxytryptamine or by naloxone

Exposure to footshock (1 mA) for 30 sec induced a marked analgesia that was enhanced by pretreatment with the 5HT synthesis inhibitor, p-chlorophenylalanine, and attenuated by the 5HT releasing drugs p-chloroamphetamine and fenfluramine, by the 5HT re-uptake inhibitor, fluoxetine and by the 5HT agonists, 5-methoxy-N,N-dimethyltryptamine and MK212. However, agonists, quipazine and trifluoromethylphenylpiperazine, with greated reported affinities for 5HT binding sites on rat brain membranes than MK212 were without effect as were the antagonists metergoline, methysergide, cyproheptadine, mianserine and methiothepin. The specific opioid antagonist naloxone was also without effect. The results in general indicate that analgesia induced by brief footshock (1 mA, 30 sec) is inversely related to 5HT availability but thereis little evidence of involvement of known 5HT receptors.     

Synergism by caffeine and by cocaine of the motor control deficit produced by midazolam

To evaluate the effects of caffeine and cocaine on the impairment of discriminative motor control produced by midazolam, rats were trained to hold a force transducer operated with a paw so that it remained between upper and lower limits of a force band for a continuous 1.5-s period to deliver each food pellet. Acute doses of 3 mg/kg midazolam SC impaired motor performance. Except for one animal, caffeine (10-40 mg/kg IP) had little or no effect on performance, while cocaine (3.75-22.5 mg/kg IP) produced dose-related impairment. When each dose of caffeine was combined with 3 mg/kg midazolam, a marked synergism in motor performance impairment occurred. Cocaine plus midazolam produced mainly an additive synergism. The conspicuous synergistic action of caffeine on the motor control deficit produced by midazolam contrasts with the typical antagonism found between the benzodiazepines and methylxanthines when performance is evaluated by psychomotor tests not requiring fine motor control.     

Deamination of beta-phenylethylamine by monoamine oxidase--inhibition by imipramine

The oxidative deamination of tyramine (Tyr), 5-hydroxytryptamine (5-HT), and β-phenylethylamine (PEA) by mitochondrial preparations of rabbit lung and brain was inhibited by imipramine. This tricyclic iminodibenzyl antidepressant drug was most effective in decreasing the deamination of PEA: at 1 × 10^?4M imipramine, deamination of PEA, Tyr and 5-HT was inhibited by approximately 70, 45 and 45 per cent, respectively, when either lung or brain mitochondrial monoamine oxidase (MAO) preparations were used. Imipramine-induced inhibition of MAO was shown to be of a mixed type based on Lineweaver-Burk plots, but was found to be completely reversible. The desmcthyl and didesmethyl derivatives of imipramine were equally as effective as the parent drug in inhibiting the deamination of PEA, whereas the N-oxide analog of imipramine was less effective as an inhibitor of this reaction. These results support the premise that the action of imipramine as a clinically effective antidepressive agent may be related to its inhibitory effect on the specific form of MAO which deaminates PEA.     

Augmentation by serrapeptase of tissue permeation by cefotiam

Cefotiam (CTM) is a new cephalosporin with a broad spectrum of activity against both Gram-positive and Gram-negative microorganisms. Cephalosporins are widely used for prophylaxis of infections in patients undergoing thoracotomy. Augmentation by serrapeptase on tissue permeation of CTM was examined in 35 thoracotomy patients with lung cancer. The subjects were divided into two groups according to the method of the administration of CTM. Group I consisted of 17 subjects, each of whom received a single dose of 2 g of CTM alone by an instillation for 30 minutes. Group II consisted of 18 subjects, each of whom received a combination of CTM and serrapeptase; serrapeptase was given 2 tablets (10 mg) each time for three times/day until the day before surgery, and then CTM was administered by the same procedure. The following results were obtained: Individual difference was observed for the permeation of CTM into tissues. Pathologic differences also affected the permeation. Nevertheless, the CTM levels in pulmonary tissues reached about a half of those in the blood in both the single dose group and the combination group, hence sufficient concentrations exceeding MIC80 for main microorganisms that caused infections in the lung were obtained. The concentrations of CTM in inflammatory tissues have showed lower levels than those of normal tissues in both CTM single dose and the combination groups. Decrease of blood flow volume may have contributed to the reduction in levels of CTM in the inflammatory tissues. The ratio of the concentration of the drug in pulmonary tissues to that in the blood was 29.1 +/- 2.5% in the single dose group, and 44.2 +/- 6.0% in the combination group, the latter showing quite a significant increase (P less than 0.05). Combined administrations of CTM and serrapeptase deserves more trials in the case when surgical treatments of the lung are performed. An antiinflammatory effect of serrapeptase in the respiratory system is expected, and in addition, the combined use of CTM and serrapeptase should stimulate permeation of the antibiotic into tissues.     

Immunomodulation by non-absorbable antibiotics given by the intragastric route

To test the role of bacterial fractions released from intestinal flora during immunomodulation by antimicrobial agents, BALB/c mice were treated with the non-absorbable antibiotics polymyxin B or teicoplanin by the intragastric route. The composition of faecal microbiota and the capacity of spleen cells to proliferate in response to B-cell and T-cell mitogens were assessed at several times during the treatment. Both antibiotics lowered the count of some bacteria of the intestinal flora and induced significant modifications in spleen cell ability to proliferate in response to mitogens. Thus, the active fractions released from intestinal bacteria during antibiotic treatments may be able to induce immunomodulating effects.