Bevacizumab 联合吉西他滨对荷肝癌裸鼠移植瘤生长的抑制作用

目的:探讨抗血管生成药物Bevacizumab联合吉西他滨对人肝癌裸鼠皮下移植瘤生长的抑制作用。方法:构建人肝癌细胞HepG2裸鼠皮下移植瘤模型,随机分为空白对照组、Bevacizumab组、吉西他滨组和联合用药组。观察用药前后肿瘤体积,绘制肿瘤生长曲线;应用免疫组化检测肿瘤微血管密度(MVD);Western Blot检测Bcl-2蛋白的表达。结果:Bevacizumab和吉西他滨单药均能抑制肿瘤生长,两药联合疗效明显增强(P=0.000)。与对照组和吉西他滨组相比,Bevacizumab组和联合用药组能明显抑制肿瘤血管生成,MVD值均明显降低,以联合用药组最为明显(P均0.000)。Bevacizumab和吉西他滨单药均能下调Bcl-2的表达,两药联合下调作用明显增强。结论:Bevacizumab联合吉西他滨能增强对人肝癌裸鼠移植瘤的生长及微血管生成的抑制作用,其机制可能与调控Bcl-2的表达有关。     

RNAi沉默SDF-1基因对人胃癌裸鼠原位移植瘤生物学行为的影响及机制

目的:探讨RNAi技术沉默基质细胞衍生因子-1(stromal cell-derived factor-1,SDF-1)对人胃癌裸鼠原位移植瘤生物学行为的影响及可能机制。方法:利用RNAi-SDF-1细胞及对照组细胞建立人胃癌裸鼠皮下移植瘤,Western Blot检测裸鼠皮下移植瘤SDF-1蛋白表达情况。利用裸鼠皮下移植瘤建立其原位移植瘤模型,8周后处死裸鼠解剖尸体,检测原位移植瘤生长、凋亡及远处脏器转移情况。Western Blot检测SDF-1沉默对通路蛋白Akt、p-Akt、NF-κB、p-NF-κB以及侵袭转移相关基因E-cadherin、MMP-7表达的影响。结果:成功建立RNAi-SDF-1裸鼠原位移植瘤模型。与空白及阴性对照组比较,RNAi-SDF-1组裸鼠原位移植瘤生长缓慢,体积与质量均明显降低,差异有统计学意义(P0.01)。流式细胞术结果显示,RNAi-SDF-1组肿瘤细胞凋亡率明显高于空白及阴性对照组(P0.01)。尸体解剖结果显示,RNAi-SDF-1组肿瘤腹腔淋巴结及肝脏转移率明显低于空白及阴性对照组(P0.05)。Western Blot结果显示,与空白及阴性对照组比较,RNAi-SDF-1组肿瘤组织中Akt、p-Akt、NF-κB、p-NF-κB、MMP-7表达水平明显降低,E-cadherin表达水平明显升高(P0.01)。结论:RNAi-SDF-1能够有效抑制人胃癌SGC7901细胞裸鼠原位移植瘤的生长,诱导肿瘤细胞凋亡,抑制其腹腔淋巴结、肝脏转移,其机制可能与抑制PI3K-Akt、NF-κB信号通路及MMP-7的表达并上调E-cadherin的表达相关。     

藻蓝蛋白对非小细胞肺癌细胞移植瘤生长的抑制及促凋亡作用的机制研究

摘要 目的：探讨藻蓝蛋白对非小细胞肺癌细胞移植瘤生长的抑制及促凋亡作用的机制研究。方法：4~6周龄30只无胸腺BALB/c裸鼠随机分为移植瘤组和藻蓝蛋白组。每5天用卡尺测量裸鼠肿瘤体积。通过RT-PCR检测裸鼠肿瘤中凋亡相关因子MMP-2、MMP-9、Bcl-2、Bax的mRNA表达。通过流式细胞术分析细胞周期。通过蛋白印迹分析EMT相关蛋白的表达。通过ELISA检测血浆细胞因子TNF-α、IL-6、IL-1β和TGF-β的浓度。通过蛋白印迹分析STAT3/NF-κB信号通路的表达。结果：第13天时,移植瘤组和蓝藻蛋白组肿瘤大小比较无差异（P>0.05）,第18天和第25天时,藻蓝蛋白组肿瘤体积较移植瘤组减小（P<0.05）。藻蓝蛋白组MMP-2、MMP-9和Bcl-2的mRNA表达较移植瘤组降低（P<0.05）,藻蓝蛋白组Bax mRNA表达较移植瘤组升高（P<0.05）。藻蓝蛋白组G1期占比较移植瘤组升高（P<0.05）,藻蓝蛋白组G1期占比较移植瘤组升高（P<0.05）,藻蓝蛋白组S期和G2期占比较移植瘤组降低（P<0.05）。藻蓝蛋白组N-钙粘蛋白和VEGF蛋白表达较移植瘤组降低（P<0.05）,藻蓝蛋白组E-钙粘蛋白表达较移植瘤组升高（P<0.05）。藻蓝蛋白组TNF-α、IL-6、IL-1β和TGF-β的浓度较移植瘤组降低（P<0.05）。藻蓝蛋白组p-STAT3、p-IκBα和p-NF-κB p65蛋白表达较移植瘤组降低（P<0.05）。结论：藻蓝蛋白通过调控STAT3/ NF-κB信号通路抑制炎性细胞因子的分泌和EMT发生,促进肿瘤细胞凋亡,抑制裸鼠体内移植瘤的生长。     

PDSW联合热疗可抑制胃癌NF-κB的表达

为探讨生理性深层海水(PDSW)联合热疗对人胃癌SGC-7901细胞体外和体内的抗癌作用,通过在体外培养人胃癌SGC-7901细胞株,随机分为实验组、对照组和空白组,实验组加PDSW,对照组加生理盐水,空白组不做其他处理,分别在37℃、40℃和43℃热疗温度下培养。同时,将SGC-7901细胞注射至裸鼠皮下,建立移植瘤模型,随机分为5组,至肿瘤长至直径为1 cm时,分别进行H_2O+RT、H_2O+40℃、H_2O+43℃、PDSW+40℃、PDSW+43℃处理,然后绘制裸鼠肿瘤生长曲线,观察裸鼠肿瘤组织切片,并用实时荧光定量PCR检测细胞和组织中NF-κB基因表达情况,Western blotting检测NF-κB蛋白的表达情况。研究发现,与其他处理相比,PDSW联合热疗处理后肿瘤生长缓慢,肿瘤细胞的增长明显受抑,细胞和裸鼠皮下移植瘤组织中NF-κB基因和蛋白的表达量较低。说明PDSW联合热疗能抑制胃癌SGC-7901细胞中NF-κB基因和蛋白的表达,从而有效抑制胃癌细胞的增殖。     

固本抑瘤Ⅲ号方联合吉西他滨对人胰腺癌裸鼠异位移植瘤的抑瘤作用

目的探讨固本抑瘤Ⅲ号方以及与吉西他滨联合治疗人胰腺癌裸鼠异位移植瘤的抑瘤作用。方法将40只荷瘤裸鼠随机分为对照组、吉西他滨组、固本抑瘤Ⅲ号联合吉西他滨组、固本抑瘤Ⅲ号组,每组10只。于接种后第8天开始给药,观察指标为瘤重、裸鼠体重、移植瘤体积。结果吉西他滨组、固本抑瘤Ⅲ号联合吉西他滨组、固本抑瘤Ⅲ号组的抑瘤率分别为49.2%、68.9%和28.0%。固本抑瘤Ⅲ号联合吉西他滨组较吉西他滨组抑瘤作用更强(P0.05)。吉西他滨组、固本抑瘤Ⅲ号联合吉西他滨组、固本抑瘤Ⅲ号组移植瘤体积均较对照组明显降低,差异有显著性。固本抑瘤Ⅲ号联合吉西他滨组体重下降明显,体重较对照组、吉西他滨组及固本抑瘤Ⅲ号组明显下降,差异有显著性。结论固本抑瘤Ⅲ号方具有增加吉西他滨化疗治疗人胰腺癌裸鼠腋下移植瘤疗效的作用。     

乙肝病毒X蛋白通过NF-κB通路调节肝癌细胞迁移、侵袭的研究

慢性乙肝病毒感染是亚洲国家肝癌的常见病因,乙肝病毒所表达的乙肝病毒X蛋白(Hepatitis B virus X protein,HBx)是肝癌发生发展的重要推动因子。核因子-κB(Nuclear factorκB,NF-κB)是模式识别受体下游重要的转录因子,肝细胞生长因子(Hepatocyte growth factor,HGF)/c-Met途径能够促进NF-κB活化,并通过活化的NF-κB来调节肝癌细胞的迁移、侵袭等生物学环节。但HBx是否直接通过NF-κB通路来调节肝癌细胞的迁移、侵袭尚未明确。为探究HBx通过NF-κB通路调节肝癌细胞迁移、侵袭的作用,本研究采用培养肝癌HepG2细胞并分组,空白对照组用不含药物及质粒的DMEM处理,空白质粒组转染1.2μg的pcDNA3.1空白质粒,HBx质粒组转染不同浓度的HBx表达质粒pcDNA3.1-HBx,HBx+PDTC组转染1.2μg的HBx表达质粒pcDNA3.1-HBx并用含有50μmol/L PDTC的DMEM进行处理;皮下注射HepG2细胞建立移植瘤小鼠模型,称量移植瘤的质量。检测细胞迁移及侵袭活力、细胞及移植瘤中NF-κB通路分子、迁移基因、侵袭基因的表达。结果显示,与空白对照组、空白质粒组比较,HBx质粒组细胞中NF-κB、HGF、c-Met、N-cadherin、Vimentin、MMP2、MMP9的蛋白表达水平及相对愈合面积、侵袭数目均明显增多(P0.05);与HBx质粒组比较,HBx+PDTC组细胞中NF-κB、HGF、c-Met、N-cadherin、Vimentin、MMP2、MMP9的蛋白表达水平及相对愈合面积、侵袭数目均明显减少(P0.05);与空白对照组、空白质粒组比较,HBx质粒组移植瘤的质量及移植瘤中NF-κB、HGF、c-Met的蛋白表达水平明显增加(P0.05)。本研究得出结论,HBx能够促进肝癌细胞的迁移、侵袭,且该作用与激活NF-κB通路有关。     

双歧杆菌预防大肠癌生长的信号转导机制

目的:从信号转导这一层次探索青春型双歧杆菌预防大肠癌生长的机制.方法:以大肠癌裸鼠移植瘤为动物模型,用膜结合法和免疫组化分别测定了大肠癌组织蛋白酪氨酸激酶(PTK)的含量以及NF-κB的表达.结果:双歧杆菌预防组大肠癌组织PTK的活性以及NF-κB的阳性细胞密度均明显低于肿瘤对照组(P＜0.01).结论:青春型双歧杆菌体内预防大肠癌生长的途径之一为降低其PTK的活性,同进抑制NF-κB的活化.     

西施舌多糖对人食管鳞癌裸鼠移植瘤作用的研究北大核心CSCD

构建人食管鳞癌EC-9706细胞裸鼠移植瘤模型,研究西施舌多糖对人食管鳞癌EC-9706细胞裸鼠移植瘤生长的影响。将移植瘤裸鼠随机分成5组,每天分别腹腔注射生理盐水(NS)、环磷酰胺(CTX,5 mg/kg)、西施舌多糖3个剂量(100、200、300 mg/kg),2周后,每3 d用游标卡尺测量肿瘤体积。实验持续26 d,实验结束后,处死裸鼠,称瘤重,计算抑瘤率。结果表明,不同浓度西施舌多糖对人食管鳞癌裸鼠移植瘤生长均有抑制作用,且西施舌多糖各剂量组中抑瘤率呈现剂量-效应关系,其中西施舌多糖300 mg/kg实验组效果最佳,抑瘤率达28.85%。     

西施舌多糖对人食管鳞癌裸鼠移植瘤作用的研究

构建人食管鳞癌EC-9706细胞裸鼠移植瘤模型,研究西施舌多糖对人食管鳞癌EC-9706细胞裸鼠移植瘤生长的影响。将移植瘤裸鼠随机分成5组,每天分别腹腔注射生理盐水(NS)、环磷酰胺(CTX,5 mg/kg)、西施舌多糖3个剂量(100、200、300 mg/kg),2周后,每3 d用游标卡尺测量肿瘤体积。实验持续26 d,实验结束后,处死裸鼠,称瘤重,计算抑瘤率。结果表明,不同浓度西施舌多糖对人食管鳞癌裸鼠移植瘤生长均有抑制作用,且西施舌多糖各剂量组中抑瘤率呈现剂量-效应关系,其中西施舌多糖300 mg/kg实验组效果最佳,抑瘤率达28.85%。     

PHD1通过下调NF-κB介导的cyclinD1的表达抑制肺癌细胞的生长和增殖

目的:探讨PHD1在肺癌中的功能,并进一步研究其分子机制,为肺癌的治疗寻找新的靶点。方法:选取人肺癌细胞A549,以脂质体为载体一方面过表达PHD1,另一方面合成设计靶向PHD1的siRNA沉默PHD1,利用荧光素酶检测NF-κB的活性,分别用western blot和real-time PCR检测cyclinD1的表达水平。在A549细胞中过量稳定表达带GFP标记的PHD1,流式细胞仪检测细胞周期变化,测量细胞的生长曲线,并将细胞注射到裸鼠皮下观察其成瘤情况。结果:过表达PHD1可明显抑制NF-κB的活性和IκBα的降解,降低cyclin D1的mRNA和蛋白表达水平;而干扰PHD1的表达可显著增加NF-κB的活性,并上调cyclin D1的mRNA和蛋白表达水平,而不影响cyclinE1。过表达IκBαSR可以阻止干扰PHD1引起的cyclinD1 mRNA水平的上调。过表达PHD1可引起细胞周期的停滞,显著抑制细胞的增殖和移植瘤的生长。结论:PHD1可能通过下调NF-κB介导的cyclinD1的表达抑制肺癌细胞的生长和增殖。     

Enhanced antitumor efficacy of gemcitabine by evodiamine on pancreatic cancer via regulating PI3K/Akt pathway

Evodiamine has therapeutic potential against cancers. This study was designed to investigate whether combination therapy with gemcitabine and evodiamine enhanced antitumor efficacy in pancreatic cancer. In vitro application of the combination therapy triggered significantly higher frequency of pancreatic cancer cells apoptosis, inhibited the activities of PI3K, Akt, PKA, mTOR and PTEN, and decreased the activation of NF-κB and expression of NF-κB-regulated products. In vivo application of the combination therapy induced significant enhancement of tumor cell apoptosis, reductions in tumor volume, and inhibited activation of mTOR and PTEN. In conclusion, evodiamine can augment the therapeutic effect of gemcitabine in pancreatic cancer through direct or indirect negative regulation of the PI3K/Akt pathway.     

Tumor-suppressing function of caspase-2 requires catalytic site Cys-320 and site Ser-139 in mice

The multifunctional caspase-2 protein is involved in apoptosis, NF-κB regulation, and tumor suppression in mice. However, the mechanisms of caspase-2 responsible for tumor suppression remain unclear. Here we identified two sites of caspase-2, the catalytic Cys-320 site and the Ser-139 site, to be important for suppression of cellular transformation and tumorigenesis. Using SV40- and K-Ras-transformed caspase-2 KO mouse embryonic fibroblast cells reconstituted with expression of wild-type, catalytic dead (C320A), or Ser-139 (S139A) mutant caspase-2, we demonstrated that similar to caspase-2 deficiency, when Cys-320 and Ser-139 were mutated, caspase-2 lost its ability to inhibit cellular transformation and tumorigenesis. These mutant cells exhibited enhanced cell proliferation, elevated clonogenic activity, accelerated anchorage-independent growth, and transformation and were highly tumorigenic, rapidly producing large tumors in athymic nude mice. Investigation into the underlying mechanism showed that these two residues are needed for caspase-2 to suppress NF-κB activity, promote apoptosis, and sustain the G(2)/M checkpoint following DNA damage induction. In addition, tumors in nude mice derived from the two mutant cell lines had higher constitutive NF-κB activity and elevated expression of NF-κB targets of antiapoptotic proteins Bcl-xL, XIAP, and cIAP2. A reduction in caspase-2 mRNA was associated with multiple types of cancers in patients. Together, these observations suggest the combined functions of caspase-2 in suppressing NF-κB activation, promoting apoptosis, and sustaining G(2)/M checkpoint contribute to caspase-2 tumor-suppressing function and that caspase-2 may also impact tumor suppression in humans. These findings provide insight into tumor suppression at the cross-roads of apoptosis, cell cycle checkpoint, and NF-κB pathways.     

Silencing of PTGS2 exerts promoting effects on angiogenesis endothelial progenitor cells in mice with ischemic stroke via repression of the NF-κB signaling pathway

The objective of the current study is to investigate the effect of PTGS2 on proliferation, migration, angiogenesis and apoptosis of endothelial progenitor cells (EPCs) in mice with ischemic stroke through the NF-κB signaling pathway. Middle cerebral artery occlusion (MCAO) model was established in mice. EPCs were identified, in which ectopic expression and depletion experiments were conducted. The mRNA and protein expression of related factors in tissues and cells were measured. Besides, proliferation, migration, angiogenesis, and apoptosis, as well as cell cycle distribution, of cells were determined. MCAO mice showed overexpression of interleukin-6 (IL-6), IL-17, and IL-23, and increased positive protein expression of PTGS2, as well as expression of PTGS2, nuclear factor-κB (NF-κB), tumor suppressor region 1 (TSP-1) and Bcl-2-associated X protein (Bax), but underexpression of vascular endothelial growth factor (VEGF), S-phase kinase associated protein 2 (Skp2), and B-cell lymphoma 2 (Bcl-2). Moreover, ectopic expression of tumor necrosis factor-α significantly elevated the expression of PTGS2, NF-κB, TSP-1, and Bax, as well as cell apoptosis and cell cycle arrest, but decreased the expression of VEGF, Skp2, and Bcl-2, as well as proliferation, migration and angiogenesis of EPCs, and the PTGS2-siRNA group showed an opposite trend. Taken together, we conclude that the specific knockdown of PTGS2 expression could repress the NF-κB signaling pathway, thereby inhibits apoptosis and promotes proliferation, migration and angiogenesis of EPCs, providing protective effect on mice with ischemic stroke.     

An experiment of the combined treatment of traditional Lei-huo-jiu therapy with Chinese medicine for the lacrimal gland of Sjögren’s syndrome

This experiment chooses nonobese diabetic (NOD) mouse as the animal model of Sjögren’s syndrome and investigates the morphologic changes, the expression of inflammatory factors and growth factors of this mouse’s lacrimal gland in response to a combined treatment of traditional Lei-huo-jiu therapy alone and in combination with Chinese medicine. The methods were to (1) use a morphological approach to directly observe pathological changes of the lacrimal gland in response to combined treatment and (2) to detect the level of tumor necrosis factor (TNF)-α, interleukin (IL)-1, and nuclear factor kappa B (NF-κB) in lacrimal gland tissue caused by the combined treatments using a immunohistochemical approach. There is a reduction of the mast cell’s degranulation and modulation of the level of cytokines in TNF-α, IL-1, and NF-κB in the combined therapy group. The combined treatment of traditional Lei-huo-jiu therapy with Chinese medicine can improve the pathological changes of the lacrimal gland tissue of the NOD mouse through modulating the level of TNF-α, IL-1, and NF-κB which results in improved tear secretion and function of the lacrimal gland.     

An experiment of the combined treatment of traditional Lei-huo-jiu therapy with Chinese medicine for the lacrimal gland of Sjögren’s syndrome

This experiment chooses nonobese diabetic (NOD) mouse as the animal model of Sjögren’s syndrome and investigates the morphologic changes, the expression of inflammatory factors and growth factors of this mouse’s lacrimal gland in response to a combined treatment of traditional Lei-huo-jiu therapy alone and in combination with Chinese medicine. The methods were to (1) use a morphological approach to directly observe pathological changes of the lacrimal gland in response to combined treatment and (2) to detect the level of tumor necrosis factor (TNF)-α, interleukin (IL)-1, and nuclear factor kappa B (NF-κB) in lacrimal gland tissue caused by the combined treatments using a immunohistochemical approach. There is a reduction of the mast cell’s degranulation and modulation of the level of cytokines in TNF-α, IL-1, and NF-κB in the combined therapy group. The combined treatment of traditional Lei-huo-jiu therapy with Chinese medicine can improve the pathological changes of the lacrimal gland tissue of the NOD mouse through modulating the level of TNF-α, IL-1, and NF-κB which results in improved tear secretion and function of the lacrimal gland.     

The effect of tripeptide tyroserleutide (YSL) on animal models of hepatocarcinoma

This study aimed to observe the effects of tyroserleutide (tyrosyl-seryl-leucine, YSL) on the survival time of mice transplanted with the ascitic fluid-type hepatocarcinoma H22, as well as the inhibitory effect of tyroserleutide on the human hepatocarcinoma Bel-7402 that was transplanted into nude mice. At doses of 80, 20 and 5 microg/kg/d, tyroserleutide significantly prolonged the survival of mice transplanted with H22 tumor cells, producing survival rates of 89%, 39% and 49%, respectively, which were statistically significantly different from the saline group (P < 0.05). YSL, at doses of 80, 160 and 320 microg/kg/d significantly inhibited the growth of the human hepatocarcinoma Bel-7402 tumor in nude mice, producing inhibition of 40%, 64% and 59%, respectively; this inhibition was significantly greater than that by saline (P < 0.05). HE staining and electron microscopy of the pathological changes of the tumor in nude mice showed that YSL changed the structure Bel-7402 tumor cells that were transplanted into nude mice, and also induced tumor cell apoptosis and necrosis, which could be a mechanism by which YSL inhibits the tumor growth in animal models.     

Clinical case report of patients with osteosarcoma and anticancer benefit of calycosin against human osteosarcoma cells

Osteosarcoma (OS) is a malignant neoplasia in bone, characterized with main occurrence in teenagers. Calycosin (CC), a bioactive compound, is found to play potent pharmacological effects against cancer. Our previous study indicates CC-exerted benefits for anti-OS effect. However, further molecular mechanism behind this action needs to be investigated. In this study, human OS samples and clinical data were collected and used for further test and analysis. In addition, human osteosarcoma cell line (143B) and tumor-xenograft nude mice were used to evaluate antineoplastic activities of CC through a series of biochemical methods and immunoassays, respectively. Compared with non-OS controls, human OS samples showed increased levels of neoplastic microRNA-223 (miR-223), and elevated expressions of NF-κBp65, IκBα proteins in tumor cells. In cell culture study, CC-treated 143B cells showed reduced cell growth, increased lactic dehydrogenase (LD) content, and downregulated cellular miR-223 level. Immunolabeled cells of proliferating cell nuclear antigen, B-cell lymphoma 2 (Bcl-2), poly(ADP-ribose) polymerase (PARP) in CC treatments were decreased dose-dependently, while caspase-3 positive cells were elevated. Further, protein expressions of NF-κBp65, IκBα in CC-treated cells were downregulated. In addition, tumor-xenograft nude mice followed by CC treatments exhibited reductions of tumor mass, miR-223 levels, and Bcl-2, PARP-positive cells, as well as downregulations of NF-κBp65, IκBα protein expressions in OS samples. Taken together, these experimental findings reveal that CC exhibits potential pharmacological activities against OS through inducing apoptosis and inhibiting miR-223-IκBα signaling pathway in neoplastic cells.