槐耳清膏对人肺腺癌A549细胞增殖及凋亡的影响

目的探讨槐耳清膏对人肺腺癌A549细胞增殖抑制及诱导凋亡的影响。方法制备不同浓度的槐耳清膏(0、3、6、9 mg/ml)和羟基喜树碱100μg/ml作用于A549细胞,分别于24、36和48 h后,采用流式细胞术检测细胞凋亡率。结果槐耳清膏与羟基喜树碱均可诱导A549细胞凋亡,槐耳清膏各浓度组与阴性对照组相比差异显著(P<0.05),且这种抑制存在浓度和时间依赖性,槐耳清膏浓度在3.0 mg/ml 36 h后抑制作用最强,羟基喜树碱组与之比较无显著差异(P>0.05),各浓度槐耳清膏组在与A549细胞作用36 h后达到抑制高峰。结论槐耳清膏在体外能抑制人肺腺癌细胞A549的增殖,诱导细胞凋亡作用。     

槐耳清膏对人肺腺癌A549细胞增殖及周期的影响

目的研究槐耳清膏对人肺腺癌A549细胞增殖及周期的影响。方法制备浓度为0、3、6、9 mg/ml的槐耳清膏与人肺腺癌A549细胞作用后,采用流式细胞检测观察细胞凋亡率,并分析药物干预36 h对细胞周期和细胞凋亡率的影响。结果流式细胞术检测结果显示,槐耳清膏各浓度组均可发生细胞凋亡,与对照组相比差异显著(P0.05)。各浓度组可使人肺腺癌A549细胞G0/G1、S期细胞比例减少,G2/M期细胞比例增多。结论槐耳清膏能诱导人肺腺癌A549细胞增殖,可能与细胞被阻滞于G2/M期有关。     

槐耳清膏抑制结肠癌切除后肿瘤复发机制探讨

目的 探讨槐耳清膏抑制结肠癌肿瘤术后复发的作用及其机制.方法 采用36只4～6周龄BALB/c裸鼠,随机分为槐耳清膏组[槐耳清膏4.5 g/（kg·d）灌胃]、5-FU组（5-FU 150 mg/d灌胃）、空白对照组（生理盐水0.1 mL/d灌胃）,每组各12只.在皮下种植结肠癌细胞SW480构建荷瘤模型,待肿瘤生长3周后,切除初发肿瘤,观察各组药物对肿瘤复发生长及转移的影响.半定量RT-PCR检测复发肿瘤组织VEGF和HIF-1 αmRNA表达水平,Westernblot检测两者蛋白表达水平,采用免疫组织化学对复发肿瘤微血管密度（MVD）检测.结果 槐耳清膏组、5-FU组、空白对照组MVD分别为3.98％、1.65％、6.43％,槐耳清膏组、5-FU组MVD均低于空白对照组（P＜0.01）,5-FU组MVD低于槐耳清膏组（P＜0.05）.槐耳清膏组及5-FU组复发肿瘤中VEGF及HIF-1α mRNA表达水平均低于空白对照组（P＜0.01）.Westernblot检测显示槐耳清膏组及5-FU组VEGF和HIF-1 α的蛋白表达均较空白对照组下降（P＜0.01）,5-FU组复发肿瘤中VEGF和HIF-1α mRNA及蛋白的表达均高于槐耳清膏组（P＜0.05）.结论 槐耳清膏能够降低手术切除结肠癌后肿瘤复发及转移,可能与其抗血管生成作用有关.     

槐耳对人胃癌细胞MGC803增殖和凋亡的影响

[目的]观察中药槐耳对胃癌细胞株MGC803增殖和凋亡的影响.[方法]以不同浓度的槐耳（4、8、16、32、64 mg/ml）分别作用于体外培养的人胃癌细胞MGC803,通过MTT法、划痕实验等检测槐耳对MGC803细胞增殖和迁移的作用,倒置显微镜下观察细胞的形态变化,流式细胞仪检测细胞凋亡情况.[结果]MTT实验显示槐耳可抑制MGC803细胞的生长,呈时间及浓度依赖性;显微镜下可观察到细胞肿胀、破裂、呈坏死状;划痕实验显示,4、8 mg/ml槐耳分别作用MGC803细胞,划痕愈合明显慢于对照组,呈剂量依赖关系;流式细胞仪可检测到MGC803细胞凋亡率明显增加,呈时间及浓度依赖性.[结论]槐耳可抑制人胃癌MGC803细胞的迁移,且能通过诱导细胞凋亡的方式有效地抑制人胃癌细胞MGC803的生长.     

槐耳清膏对A549细胞增殖和自噬的影响

目的 观察槐耳清膏对人肺腺癌细胞系A549细胞增殖和自噬的影响,探讨其作用机制.方法 以A549细胞为研究对象,采用CCK-8法检测槐耳清膏对A549的增殖抑制率,用Western blot 方法检测自噬相关蛋白P62、LC3B表达的变化和mTOR信号通路分子表达的变化.结果 0.5 g/L及以上的槐耳清膏可明显抑制A549细胞的增殖,并且具有时间和浓度依赖性(P＜0.05).用不同浓度(1、2、4、6、8 g/L)的槐耳清膏作用A549细胞48 h可以降低P62表达水平,增加LC3B的表达,降低mTOR信号通路关键分子的表达水平.联合使用自噬抑制剂3-MA可以加强槐耳清膏对A549细胞的增殖抑制率.结论 槐耳清膏通过下调mTOR信号通路抑制A549细胞增殖,并且诱导A549发生自噬.联合使用槐耳和自噬抑制剂可能会更有效发挥槐耳抗肿瘤增殖的作用.     

细胞自噬与结肠癌Lovo细胞迁移和侵袭的相关性研究

目的探讨不同自噬状态对结直肠癌Lovo细胞迁移和侵袭能力的影响。 方法将Lovo细胞分为自噬增强组、正常细胞组和3-甲基嘌呤自噬抑制组。用激光共聚焦显微镜观察绿色荧光颗粒,用Q-PCR检测Beclin1 mRNA表达水平,透射电镜观察自噬溶酶体,Transwell实验评价Lovo细胞迁移与侵袭能力。 结果绿色荧光颗粒数以自噬增强组最多,其次为正常细胞组,而自噬抑制组最少。Beclin1 mRNA表达量自噬增强组为(1.23±0.12)个,正常细胞组为(1±0.13)个,自噬抑制组为(0.98±0.1)个。自噬增强组可见大量的自噬溶酶体,明显多于正常细胞组和自噬抑制组。迁移实验细胞计数：自噬增强组高于正常细胞组(138.0±16.7与90.7±12.9,P＝0.026)和自噬抑制组(138.0±16.7与92.7±26.7,P＝0.030)。侵袭实验细胞计数：自噬增强组高于正常细胞组(147.0±13.0与99.0±20.5,P＝0.028)和自噬抑制组(147.0±13.0与95.7±25.6,P＝0.021)。 结论结肠癌Lovo细胞自噬增强促进肿瘤细胞迁移和浸润,可能是局部浸润和远处转移的机制之一。     

中药槐耳诱导人胃癌细胞MGC803凋亡的实验研究

[目的]探讨槐耳、顺铂对人胃癌细胞株MGC803细胞增殖和凋亡的影响以及槐耳诱导人胃癌MGC803细胞凋亡的机制。[方法]以不同浓度的槐耳、顺铂以及联合用药分别作用于体外培养的人胃癌细胞MGC803,MTT法检测MGC803凋亡的情况,倒置显微镜观察其形态的变化,流式细胞术检测早期细胞凋亡情况;Western blot法检测槐耳处理24h后基质金属蛋白酶(MMP)-2的表达情况。[结果]MTT实验显示槐耳及顺铂均可抑制MGC803细胞的生长,呈时间及浓度依赖性;流式细胞仪可检测到MGC803细胞凋亡率明显增加,呈时间及浓度依赖性;槐耳能降低MGC803细胞中MMP-2蛋白的表达水平,并呈浓度依赖性。[结论]槐耳可在体外抑制人胃癌细胞MGC803的生长,并促进它的凋亡,且机制可能与抑制MMP-2的表达有关。联合使用槐耳和化疗药物可能会更有效发挥槐耳抗肿瘤增值的作用。     

槐耳清膏诱导人肺腺癌细胞A549凋亡的实验研究

槐耳清膏是药用真菌槐耳菌质的热水提取物 ,临床上证实有一定的抗癌效果。为进一步探讨其抗癌机制 ,我们研究了槐耳清膏在体外诱导人肺腺癌细胞系Ａ5 49凋亡的作用。材料与方法　引进并培养人肺腺癌细胞系Ａ5 49细胞株。制备不同浓度的槐耳清膏ＰＲＭＩ 16 40含药培养液与接种成功、呈对数生长的Ａ5 49细胞共同培养 2 4ｈ后 ,每 6ｈ进行以下观察及测定 (设阳性对照组羟基喜树碱 10 0 μｇ/ｍｌ,共同孵育 10～ 12ｈ后进行测定 ) [1] 。 (1)荧光染色 :收集培养细胞 ,清洗并均匀涂片自然风干 2 4ｈ ,置于 0 0 5ｍｇ/Ｌ的Ｈｏｅｃｈｓｔ332 5 8…     

芥菜籽提取物对人结肠癌细胞系SW480凋亡的影响

目的观察芥菜籽提取物（MSE）对人结肠癌细胞系SW480细胞凋亡的影响并初步探讨其可能的机制。方法采用体外细胞培养技术,用CCK-8法观察MSE对人大肠癌SW480细胞生长的影响;Hoechest3325荧光染色显微镜下观察凋亡细胞形态学改变,流式细胞仪检测细胞凋亡;Caspase-3活性检测试剂盒分析其凋亡机制。结果在浓度0．2—1．0mg／ml作用24—72h范围内,MSE对SW480细胞增殖具有明显抑制作用（P〈0．01）。24h内其凋亡率随着药物浓度的增加而增加,且逐渐从早期凋亡走向晚期凋亡,呈浓度依赖性;24h内Caspase-3活性较对照组显著提高（P〈0．05）。结论MSE能有效抑制体外培养的人结肠癌细胞SW480的生长,这种抑制作用与MSE促进肿瘤细胞凋亡、激活凋亡蛋白酶Caspase-3的活性有关。     

缺氧诱导BCL9促进结肠癌细胞的增殖与迁移

目的探索缺氧对B-淋巴细胞瘤蛋白9（BCL9）的调控关系及BCL9对结肠癌细胞的生物学作用。 方法收取20例结肠癌标本及同一患者的正常结肠标本,利用免疫组化技术、实时荧光定量PCR技术检测BCL9的表达;利用常氧或缺氧培养HCT116细胞,实时荧光定量PCR及蛋白印迹法（western-blot）检测BCL9的表达;构建包含BCL9启动子序列的荧光素酶报告载体,通过缺氧处理或过表达HIF1α,检测荧光素酶活性变化;通过过表达质粒或siRNA干扰BCL9的表达,随后检测HCT116细胞的增殖活性及迁移能力。 结果免疫组化及实时荧光定量PCR检测显示肿瘤组织中BCL9表达明显高于正常组织［（3.25±0.53）vs.（1.03±0.12）,P<0.05］;缺氧培养HCT116细胞后BCL9的表达显著提高,［（6.71±0.83）vs.（1.54±0.21）,P<0.05］;同时,缺氧培养或过表达HIF1α能显著提高荧光素酶的活性［（3.53±0.75）vs.（0.96±0.15）,（4.83±0.62）vs.（1.02±0.14）;P<0.05］;干扰BCL9的表达抑制HCT116细胞的增殖及迁移能力［（1.23±0.12）vs.（1.87±0.15）,P<0.05］;提高BCL9表达能促进HCT116细胞的增殖及迁移能力［（2.43±0.16）vs.（1.81±0.14）,P<0.05］。 结论缺氧可诱导BCL9在结肠癌中高表达,并通过BCL9促进结肠癌细胞增殖、迁移。     

塞莱昔布对结肠癌细胞生长及肝转移瘤血管生长因子表达的影响

目的:探讨塞莱昔布对体外培养的结肠癌细胞生长及肝转移瘤模型血管生长因子(VEGF)表达的影响.方法:以人结肠癌细胞株HT-29,HCT-116为对象,体外药物敏感实验(MTT)法检测塞莱昔布对肿瘤细胞的增殖抑制效应,流式细胞术检测肿瘤细胞各细胞周期分布变化情况,肿瘤细胞接种裸鼠,观察肝转移瘤VEGF表达情况.结果:塞莱昔布对人结肠癌细胞株生长的抑制作用呈时间、剂量依赖性效应,且对HT-29细胞作用强于HCT-116细胞(P<0.01);塞莱昔布可改变结肠癌细胞株细胞周期的分布,明显降低增殖指数(P<0.05).塞莱昔布具有明显的抑制肝转移瘤VEGF表达的作用(P=0.00).结论:塞莱昔布可通过抑制COX-2酶活性而抑制肿瘤细胞的分裂和增殖,诱导其凋亡,并抑制肿瘤血管生成,干预结肠癌的转移与复发.     

Genistein inhibits invasive potential of human hepatocellular carcinoma by altering cell cycle, apoptosis, and angiogenesis

AIM: To study the in vitro and in vivo inhibitory effects of genistein on invasive potential of Bel 7402 hepatocellular carcinoma (HCC) cells and to explore the underlying mechanism. METHODS: Bel 7402 HCC cells were exposed to genistein. The invasive activity of tumor cells was assayed in transwell cell culture chamber. p125FAK expression and cell cycle were evaluated by a functional assay. Cell apoptosis analysis was performed with TUNEL method. In addition, bilateral subrenal capsule xenograft transplantation of HCC was performed in 10 nude mice. Genistein was injected and the invasion of HCC into the renal parenchyma was observed. Microvessels with immunohistochemical staining were detected. RESULTS: Genistein significantly inhibited the growth of Bel 7402 cells, the inhibitory rate of tumor cells was 26 -42%. The invasive potential of Bel 7402 cells in vitro was significantly inhibited, the inhibitory rate was 11-28%. Genistein caused G2/M cell cycle arrest, S phase decreased significantly. The occurrence of apoptosis in genistein group increased significantly. The expression of p125FAK in 5 μg/mL genistein group (15.26±0.16%) and 10 μg/mL genistein group (12.89±0.36%) was significantly lower than that in the control group (19.75±1.12%,P<0.05). Tumor growth in genistein-treated nude mice was significantly retarded in comparison to control mice, the inhibitory rate of tumor growth was about 20%. Genistein also significantly inhibited the invasion of Bel 7402 cells into the renal parenchyma of nude mice with xenograft transplant. The positive unit value of microvessels in genistein-treated group (10.422±0.807) was significantly lower than that in control group (22.330±5.696, P< 0.01). CONCLUSION: Genistein can effectively inhibit the invasive potential of Bel 7402 HCC cells by altering cell cycle, apoptosis and angiogenesis, inhibition of focal adhesion kinase may play a significant role in this process.     

抑制MET对RAS突变型结肠癌细胞的抗肿瘤作用

目的探讨抑制MET对于RAS突变结肠癌的体内体外抗肿瘤作用。 方法选取4种常见RAS突变型结肠癌细胞（HCT-116、DLD-1、Lovo和HCT-15）,采用siRNA敲除MET蛋白表达,或者加入特异性MET抑制剂PHA-665752,采用MTT及集落形成实验观察抑制MET对结肠癌细胞增殖抑制的作用,应用Western Blotting检测PHA-665752对MET/AKT/ERK信号通路的作用。另外,应用HCT-116细胞系构建裸鼠皮下成瘤模型,给予PHA-665752腹腔注射,观察MET抑制剂对体内肿瘤的抑制作用。 结果RAS突变的4种结肠癌细胞系均有MET蛋白表达。应用siRNA敲除MET蛋白表达对HCT-116和Lovo细胞的增殖抑制作用分别为19.6±4.5%和27.8±5.8%,而应用MET特异性抑制剂PHA-665752对两种细胞的增殖抑制作用呈剂量依赖性,细胞克隆形成实验证实PHA-665752可有效抑制细胞克隆形成。另外,应用HCT-116细胞构建裸鼠皮下成瘤模型,给予PHA-665752单药证实MET抑制剂可明显抑制HCT-116细胞的皮下移植瘤（第四周肿瘤体积：PHA-665752组为300±72 mm³,对照组为608±59 mm³,t=5.731,P=0.005）。Western Blotting证实应用PHA-665752预处理能明显抑制HGF激活的p-MET、p-AKT和p-ERK。 结论抑制MET对于RAS突变型结肠癌具有抗肿瘤作用,靶向MET可能成为RAS突变型结肠癌的有效治疗。     

2-Methoxyestradiol: New perspectives in colon carcinoma treatment

Colon carcinoma represents a major problem in oncology, since this type of cancer responds poorly to conventional chemotherapy. Many groups are actively involved in the search of new experimental strategies to bypass this problem. We investigated the effects of 2-methoxyestradiol (2-ME), which derives from the NADPH-dependent cytochrome P450 metabolism of 17β-estradiol. This compound has raised much interest in the past few decades for its inhibitory effects on the growth of cancer cells of different origin; however, little is known about its use on colon carcinoma-derived cell lines. In the present study, we investigated the effects of 2-ME on cell proliferation and cell cycle of two human colon carcinoma cell lines, namely HCT116 and SW613-B3. Our results showed a net anti-proliferative effect of 2-ME on both cell lines, which is accompanied by cell cycle arrest; moreover, we demonstrated that 2-ME is able to induce apoptosis as well as autophagy. This body of evidence points out that 2-ME could be considered as a promising tool against colon carcinoma.     

The impact of pyrvinium pamoate on colon cancer cell viability

Purpose

The in vitro and in vivo effects of pyrvinium pamoate (PP), a newly identified WNT signaling inhibitor, were evaluated against colon cancer cell lines and primary colon cancer samples.

Experimental design

Antiproliferative activity of PP and its effects on protein and RNA levels of WNT targets were evaluated on adenomatous polyposis coli (APC ^mut) and β-catenin^mut cell lines, one WNT^wt colon cancer cell line, as well as six primary colon cancer samples with mutant APC in vitro. In addition, the effect of PP on the growth of liver metastasis was examined.

Results

PP blocked colon cancer cell growth in vitro in a dose-dependent manner with great differences in the inhibitory concentration (IC₅₀), ranging from 0.6?×?10^?6 to 65?×?10^?6 mol/L for colon cancer cells with mutations in WNT signaling. In addition, PP demonstrated a cytotoxic effect on primary colon cancer samples. A combined cytotoxic effect of PP with 5-fluorouracil (5-FU) was observed for two cell lines. PP decreased messenger RNA (mRNA) and protein levels of known WNT target genes as c-MYC and thereby led to the induction of p21. PP inhibited the migration of HCT116 colon cancer cells in vitro and decreased tumor growth in vivo after intraportal injection of HCT116 cells in nude mice.

Conclusions

PP displays promising anticancer activity against a broad panel of human colon cancer cell lines, as well as primary colon cancer samples. However, our findings do not demonstrate a predominant cytotoxic effect of PP on colon cancer cells with mutations in WNT signaling.     

In vivo and in vitro antitumor effect of a unique nutrient mixture on lung cancer cell line A-549

The high incidence of lung cancer and ineffective toxic action of current mono and doublet chemotherapy approaches result in poor patient survival. Further, matrix metalloproteinases (MMPs) are implicated in neoplastic invasion and metastasis. Based on this, the authors investigated the effect of a dietary micronutrient mixture (NM) containing lysine, proline, arginine, ascorbic acid, and green tea extract on the tumor growth of human lung carcinoma cell A-549 xenografts in athymic nude mice. Additionally, the authors tested the in vitro antitumor effect of NM on lung carcinoma A-549 cells by measuring cell proliferation by MTT assay, MMP-2 and -9 secretion by gelatinase zymography, and cell invasion through Matrigel. Nutrient supplementation strongly suppressed the growth of tumors without adverse effects in nude mice; tumor weight was reduced by 44% (P = .0001) and tumor burden was reduced by 47% (P < .0001) with supplementation. Zymography demonstrated in vitro secretion of MMP-2 by uninduced human lung carcinoma cells and both MMP-2 and -9 by phorbol 12-mysristate 13-acetate (PMA) (200 ng/mL)-treated cells. NM inhibited the secretion of both MMPs in a dose-dependent fashion, with virtual total inhibition at 500 microg/mL concentration. The invasion of human lung carcinoma cells through Matrigel was significantly reduced at 100 microg/mL (64%) and totally inhibited at 500 microg/mL concentration of NM (P = .01). Suppression of lung tumor growth in nude mice and inhibition of MMP secretion and Matrigel invasion suggest NM may act as an anticancer agent and as such warrants further investigation.     

Nicotine enhances migration and invasion of human esophageal squamous carcinoma cells which is inhibited by nimesulide

AIM： To study the effect of nicotine on the migration and invasion of human esophageal squamous carcinoma cells and to investigate whether nimesulide can inhibit the effect of nicotine.METHODS： The esophageal squamous carcinoma cell line （TE-13） was treated with different concentrations of nicotine （100 μg/mL and 200 μg/mL） or 200 μg/mL nicotine plus 100 μmol/L nimesulide. Cell migration and invasion were measured using migration and invasion chamber systems. COX-2 expression was determined by Western blotting. Matrix metalloproteinase-2 （MMP-2） was analyzed by zymography and ELISA.RESULTS： Nicotine （100 μg/mL, 200 μg/mL） enhanced TE-13 cells migration and invasion, and increased the protein expression of COX-2 and the activity of MMP-2. Nicotine （200 μ/mL） stimulated TE-13 cells migration and invasion which were partly blocked by nimesulide. This was associated with decreased protein expression of COX-2 and decreased activity and protein expression of MMP-2. CONCLUSION： Nicotine enhances the migration and invasion of the esophageal squamous carcinoma cell line, and nimesulide partly blocks the effect ofnicotine-enhanced esophageal squamous carcinoma cell migration and invasion.