Stable plasmid-based siRNA silencing of gene expression in human embryonic stem cells

RNA interference (RNAi) using short inhibitory RNAs (siRNAs) has been widely explored for the suppression of cellular mRNA levels to investigate the function of specific genes, including gene function in differentiation and development. The establishment of human embryonic stem cell (hESC) models for differentiation of selected lineages is an area of intense interest and activity. On the basis of our previous work with stable overexpression of enhanced green fluorescent protein (EGFP) in hESC, we used plasmid vector-based siRNA expression to silence EGFP expression in stably-transfected hESC. After hygromycin selection, we derived several cell lines in which EGFP expression was significantly reduced. At the genomic DNA level, there was no difference between the two cell lines and the parental H1EGFP cell line when analyzed with quantitative PCR; however, there were significant differences among the three cell lines at the RNA and protein levels as analyzed with real-time RT-PCR and Western blotting. From these data, we conclude that the decrease in EGFP expression was caused by RNAi, not by genomic DNA loss. Down-regulation of EGFP expression was sustained through multiple passages of both siEGFP cell lines. This simple silencing system will allow novel investigations of target gene function in hESC self-renewal or differentiation, as well as differentiated function in other cell types.     

Enhanced siRNA delivery into cells by exploiting the synergy between targeting ligands and cell-penetrating peptides

We have developed a polymer nanoparticle-based siRNA delivery system that exploits a cell surface binding synergism between targeting ligands and cell-penetrating peptides. Nanoparticles were coated with folate and penetratin via a PEGylated phospholipid linker (DSPE-PEG): the combination of both of these ligands represents a strategy for enhancing intracellular delivery of attached polymer nanoparticles. Nanoparticles were characterized for size, morphology, density of surface modification, and ligand association and retention. The surface coverage achieved on DSPE-PEG-coated nanoparticles is as high as (or higher than) obtained with other ligand-modified nano-scale particulate systems (～0.5-5 pmol ligand/cm2). Additionally, these nanoparticles were loaded with a high density of siRNA (～130-140 pmol siRNA/mg nanoparticles), which is slowly released upon incubation in water. Synergies between the activity of surface binding and cell internalizing ligands on these siRNA-loaded nanoparticles impart delivery enhancements that improve their gene silencing efficacy both in culture and in tumor models. Traditionally, targeting ligands function by binding to cell surface receptors, while cell-penetrating peptides function by nonspecifically transporting across cell membranes. Interestingly, we have observed that improved delivery of these dual-functionalized nanoparticles was in part, a result of increased cell surface avidity afforded by both ligands. This siRNA delivery system presents an approach to surface modification of nanovehicles, in which multiple ligands function in parallel to enhance cell binding and uptake.     

针对S基因的siRNA或shRNA表达框对HepG2.2.15细胞中HBV基因表达和病毒复制的抑制作用比较

目的 化学合成针对乙型肝炎病毒（HBV）S基因的siRNA，同时制备针对相同区段的shRNA表达框，并比较二者对HepG2．2．15细胞中HBV基因表达和病毒复制的抑制作用。方法 化学合成针对HBVS基因的带有FITC标记的siRNA，设计合成带有FTrc标记的引物，PCR扩增含有RNA聚合酶Ⅲ启动子H1序列和shRNA编码序列的表达框，并对扩增产物进行纯化，将等摩尔数siRNA和shRNA表达框分别用脂质体转染稳定表达HBV的HepG2．2．15细胞，转染1d后流式细胞仪检测转染效率，转染3d后RT-PCR检测靶基因mRNA的水平，用SDS-PAGE、Westem blot及间接免疫荧光染色检测HBsAg的表达。收集转染前和转染后1、3、5和7d的细胞培养上清，检测HBsAg水平，同时用荧光定量PCR方法检测HBV　DNA的含量。结果成功制备了针对HBVS基因的shRNA表达框，将其与等摩尔数siRNA分别转染HepG2．2．15细胞后，RT-PCR证实细胞中HBV　mRNA水平降低，SDS-PAGE、Westem blot及间接免疫荧光染色检测到HBsAg的表达受到抑制，细胞培养上清中HBsAg和HBV DNA含量下降。与siRNA相比，shRNA表达框对HBV的抑制作用虽然起效较慢，但持续时间更长。结论shRNA表达框和siRNA均可明显抑制HBV的转录和表达，与siRNA相比，shRNA表达框能够更持久地抑制靶基因的表达。     

针对S基因的siRNA抑制HepG2 2.2.15细胞中HBV基因的表达

目的 构建针对乙型肝炎病毒（HBV）S基因的siRNA（short interfering RNA）表达载体pSUPER-S1和pSUPER-S2,观察其对HepG2 2.2.15细胞中的HBV基因表达的影响.方法 设计并合成针对HBV S区基因的siRNA寡核苷酸,经退火形成双链后克隆入pSUPER载体,构建成功的siRNA表达载体与pTK-Hyg质粒共转染稳定表达HBV的HepG2 2.2.15细胞,经200μg/ml潮霉素抗性筛选,4周后获得稳定细胞克隆,对所得细胞培养上清中的HBsAg和HBeAg进行定量检测,免疫荧光染色检测细胞内抗原的表达,同时用RT-PCR检测靶基因mRNA的抑制效果.结果 成功构建了针对HBV S基因的siRNA表达载体pSUPER-S1和pSUPER-S2,两种siRNA均能明显抑制HepG2 2.2.15细胞的HBsAg和HBeAg分泌,抑制率分别为83%和78%,免疫荧光染色显示siRNA能抑制细胞内抗原的合成,RT-PCR结果证实HBV的mRNA表达降低,而无关序列的siRNA和对照则无此作用.结论 载体产生的针对HBV S基因的siRNA能稳定、高效、特异地抑制HBV基因的表达.     

人胚胎干细胞外源基因的瞬时表达效率

背景：人类胚胎干细胞的修饰和操作技术多种多样,它们在效率、可靠性及安全性方面各有不同。 目的：对比观察两种化学转染试剂转染含不同启动子的增强型绿色荧光蛋白表达载体在人胚胎干细胞中的表达效率。 方法：采用Fugene HD和lipofectamine2000分别转染无滋养层培养的H9细胞,荧光显微镜下观察阳性克隆,流式细胞术分析不同载体pCAG-EGFP、pEGFP-N3在H9细胞中的表达效率。 结果与结论：Fugene HD转染的pCAG-EGFP、pEGFP-N3两种载体在H9细胞中的表达效率分别为(42.45±3.32)%、 (25.95±1.91)%,差异有显著性意义(P < 0.05)。lipofectamine2000转染的pCAG-EGFP、pEGFP-N3两种载体在H9细胞中的表达效率分别为(1.94±0.18)%、(1.49±0.33)%。采用Fugene HD转染的表达效率均高于lipofectamine2000转染的表达效率(P < 0.05)。结果显示在人胚胎干细胞的H9细胞系中,Fugene HD的转染效率显著高于lipofectamine2000;CAG启动子驱动的增强型绿色荧光蛋白表达效率显著高于CMV启动子。     

VEGF基因特异性siRNA转染后对人乳腺癌细胞增殖和凋亡的影响

目的：探讨利用干扰RNA（RNAi）抑制血管内皮生长因子（VEGF）基因表达对乳腺癌细胞MCF-7增殖和凋亡的影响。方法：设计针对VEGF基因的小干扰RNA（siRNA）,合成DNA模板,体外转录siRNA。以脂质体转染法将双链siRNA导入MCF-7细胞后,用MTT比色法检测siRNA对MCF-7细胞增殖的影响。通过Hoechst33258染色观察MCF-7细胞的凋亡。用流式细胞术检测细胞周期的改变,RT-PCR检测VEGF mRNA表达的变化,免疫细胞化学法检测VEGF蛋白的表达。结果：所设计的两个靶位点siRNA,均能有效地抑制MCF-7细胞的增殖,诱导细胞凋亡,使细胞周期阻滞于G0/G1期,VEGFmRNA及其蛋白的表达明显减少;而作为阴性对照的错义序列组siRNASCR则没有上述效应。结论：体外转录合成的siRNA可抑制MCF-7细胞中VEGF基因的表达,抑制细胞增殖,促进细胞凋亡。     

用小双链RNA抑制转化入HUVEC中绿色荧光蛋白基因的表达

目的：建立稳定表达绿色荧光蛋白(GFP)的人脐静脉血管内皮细胞（HUVECs），研究小干扰RNA(siRNA)对HUVECs中GFP表达的抑制作用。 方法： 用lipofectamine2000将编码GFP的质粒pN3-EGFP转入HUVECs中。用G418筛选并维持已转化pN3-EGFP的HUVEC(HUVEC-GFP)。利用T7 RNA转录试剂盒，制备可抑制GFP基因表达的siRNA（GFPsiRNA）和无关对照的RNA（control siRNA）。用oligofectamine将siRNA转入HUVEC-GFP中。继续培养48 h后，检测HUVEC-GFP中GFP蛋白和mRNA表达水平。 结果： 用G418筛选获得了HUVEC-GFP细胞株，可以观察到GFP的稳定表达。HUVEC-GFP转化siRNA后48 h，GFP的荧光强度明显下降，而对照组的荧光强度无明显下降。半定量RT-PCR检测显示，GFPsiRNA对GFP mRNA表达有较强的抑制作用，抑制率达40%，而control siRNA对GFP mRNA表达水平无明显的抑制作用。 结论： 利用体外转录合成的siRNA能有效地抑制HUVECs中GFP的表达。     

Expression of BLM (the causative gene for Bloom syndrome) and screening of Bloom syndrome

Bloom syndrome (BS) is a rare autosomal recessive genetic disorder characterized by growth deficiency, unusual facies, sun-sensitive telangiectatic erythema, immunodeficiency and predisposition to cancer. The causative gene for BS is the BLM gene which encodes the BLM RecQ helicase protein. The BLM gene has 4437 bp and encodes 1417 amino acids. The detection of BLM gene mutations for laboratory diagnosis of BS is laborious and impractical, unless there are common mutations in a population. Here we describe the immunoblot and immunohistochemical analyses for the detection of the BLM protein using a polyclonal BLM antibody. The BLM gene and protein were consistently and clearly detected in Epstein-Barr virus (EBV)-transformed or phytohemagglutinin (PHA)-stimulated lymphoblasts from control and various human hematopoietic cell lines. In a 7-week old human fetal brain, the BLM gene expression was strongly detected in contrast to an adult human brain. The BLM protein was not detected in EBV-transformed lymphoblasts from three BS patients. By immunohistochemistry, nuclear dots of the BLM protein were detected in both EBV-transformed lymphoblasts and PHA-stimulated lymphoblasts from the control. However, in lymphoblasts from BS patients no nuclear dots of the BLM protein were detected. These results indicate that the combinational analysis of immunoblotting and immunohistochemistry is a useful approach to screening of BS, although a mutation analysis is necessary for a definitive diagnosis of BS.     

Modeling for Lesch-Nyhan disease by gene targeting in human embryonic stem cells

Human embryonic stem (ES) cells are pluripotent cells derived from blastocyst-stage embryos. It has been suggested that these cells should play a major role in transplantation medicine and be able to advance our knowledge in human embryology. We propose that these cells should also play a vital role in the creation of models of human disorders. This aspect would be most valuable where animal models failed to faithfully recapitulate the human phenotype. Lesch-Nyhan disease is caused by a mutation in the HPRT1 gene that triggers an overproduction of uric acid, causing gout-like symptoms and urinary stones, in addition to neurological disorders. Due to biochemical differences between humans and rodents, a mouse lacking the HPRT expression will fail to accumulate uric acid. In this research we demonstrate a model for Lesch-Nyhan disease by mutating the HPRT1 gene in human ES cells using homologous recombination. We have verified the mutation in the HPRT1 allele at the DNA and RNA levels. By using selection media, we show that HPRT1 activity is abolished in the mutant cells, and the HPRT1-cells show a higher rate of uric acid accumulation than the wild-type cells. Therefore, these cells recapitulate to some extent the characteristics of Lesch-Nyhan syndrome and can help researchers further investigate this genetic disease and analyze drugs that will prevent the onset of its symptoms. We therefore suggest that human diseases may be modeled using human ES cells.     

靶向着丝粒蛋白A的小RNA干扰对肝癌细胞株HepG2细胞生物学行为的影响

目的 研究靶向着丝粒蛋白A(CENP-A)的siRNA对肝癌细胞株HepG2细胞生物学行为的影响.方法 设计并合成三对CENP-A编码基因的反向重复序列,中间间隔9个核苷酸序列,通过定向克隆至载体pSilencerTM 2.1-U6 neo,构建siRNA真核表达质粒,经稳定转染HepG2细胞后检测肝癌细胞中CENP-A基因mRNA及蛋白质表达的抑制情况;通过观察HepG2细胞生长、凋亡、细胞周期和平板克隆形成能力评价CENP-A基因干扰对细胞生物学行为的影响;通过检测bcl-2、Bax、p21waf1、mdm2、p53蛋白表达水平初步探讨CENP-A生物学作用的可能机制.结果 三对靶向CENP-A的siRNA干扰片段中有二对抑制效果明显.与未转染组及空载体组相比,转染CENP-A干扰片段的细胞生长减慢,平板克隆形成率下降.细胞周期检测显示,G1期阻滞(P＜0.01),S期细胞比例减少(P＜0.001).细胞凋亡比例增加(P=0.003),并伴随bcl-2蛋白表达明显下降(P=0.000),Bax蛋白表达明显增高(P=0.001).还可致p21waf1表达增高,mdm2表达下降,但对野生型p53表达未显示明显影响.结论 CENP-A通过参与细胞周期调控而密切相关于细胞恶性增殖和凋亡抑制,其机制可能涉及野生型p53非依赖性通路和凋亡相关基因bcl-2Bax的表达异常.     

Enhanced docetaxel-mediated cytotoxicity in human prostate cancer cells through knockdown of cofilin-1 by carbon nanohorn delivered siRNA

We synthesized a non-viral delivery system (f-CNH3) for small interfering RNA (siRNA) by anchoring a fourth-generation polyamidoamine dendrimer (G4-PAMAM) to carbon nanohorns (CNHs). Using this new compound, we delivered a specific siRNA designed to knockdown cofilin-1, a key protein in the regulation of cellular cytoskeleton, to human prostate cancer (PCa) cells. The carbon nanohorn (CNH) derivative was able to bind siRNA and release it in the presence of an excess of the polyanion heparin. Moreover, this hybrid nanomaterial protected the siRNA from RNAse-mediated degradation. Synthetic siRNA delivered to PCa cells by f-CNH3 decreased the cofilin-1 mRNA and protein levels to about 20% of control values. Docetaxel, the drug of choice for the treatment of PCa, produced a concentration-dependent activation of caspase-3, an increase in cell death assessed by lactate dehydrogenase release to the culture medium, cell cycle arrest and inhibition of tumor cell proliferation. All of these toxic effects were potentiated when cofilin-1 was down regulated in these cells by a siRNA delivered by the nanoparticle. This suggests that knocking down certain proteins involved in cancer cell survival and/or proliferation may potentiate the cytotoxic actions of anticancer drugs and it might be a new therapeutic approach to treat tumors.     

靶向性表皮生长因子受体siRNA表达载体的构建和表达

目的构建靶向性表皮生长因子受体(epidermal growth factor receptor，EGFR)的小分子干扰RNA(small interfering RNA，siRNA)表达载体并在TJ905人脑恶性胶质瘤细胞系表达。方法在人EGFR细胞外区受体结合结构域和细胞内区酪氨酸激酶结构域各选择1个siRNA靶序列、进行了psiRNA—NeoG2表达载体的构建，进而以反义EGFR表达载体P—anti—hEGFR为对照进行了脂质体介导的TJ905人脑恶性胶质瘤细胞系表达。应用免疫荧光和蛋白印迹检测EGFR的表达。结果成功构建以psiRNA—NeoG2为载体的siRNA表达质粒，并分别使其在稳定转染TJ905细胞后EGFR表达下调90％和92％；反义EGFR表达载体P—anti—hEGFR使EGFR表达下调82％。结论靶向EGFR的siRNA表达载体可以特异性地抑制EGFR的表达，可以成为胶质瘤靶向性EGFR基因治疗的一种新策略。     

Relatively common mutations of the Bloom syndrome gene in the Japanese population

Bloom syndrome (BS) is a rare autosomal recessive genetic disorder characterized by lupus-like erythematous facial telangiectasia, sun sensitivity, infertility, stunted growth and a high predisposition to various types of cancer. Chromosomal abnormalities are hallmarks of this disorder, and high frequencies of sister chromatid exchanges and quadriradial configurations in lymphocytes and fibroblasts are diagnostic features. BLM is the causative gene for BS. We investigated the mutation in the BLM gene in 4 Japanese BS kindreds. Taken together with previously documented mutations, 2 kindreds were homozygous for 631delCAA and 2 were compound heterozygous for 631delCAA. The silent mutation of A1055C (Thr to Thr) was detected in control Japanese individuals. The 6-bp deletion/7-bp insertion at position 2,281, which most Askenazi Jewish BS patients carry, was not detected in 200 Japanese alleles. These results suggest that 631delCAA is a relatively common mutation among the Japanese BS patients.     

siRNA干扰 BMI-1 基因对膀胱癌EJ细胞增殖的调控作用及其机制

目的: 对比观察膀胱癌EJ细胞及人膀胱移行上皮细胞中 BMI-1 基因及下游 p16^INK4a、p14^ARF 基因的mRNA及蛋白表达水平,探讨siRNA干扰 BMI-1 基因后对EJ细胞增殖的影响及其调控机制。方法: 细胞免疫荧光观察BMI-1、p16^INK4a和p14^ARF蛋白在EJ细胞中表达情况及定位。Real-time RT-PCR检测EJ细胞及膀胱正常移行上皮细胞中3种基因的表达水平,Western blotting检测蛋白水平。构建干扰 BMI-1 基因siRNA,通过脂质体转染导入EJ细胞中,设立空白对照和阴性干扰对照组,检测 BMI-1 及下游 p16、p14 基因表达水平及蛋白表达水平的变化;以CCK-8检测细胞生长观察siRNA干扰 BMI-1 表达对EJ细胞增殖的抑制作用;用流式细胞术分析细胞凋亡的改变。结果: BMI-1mRNA及蛋白水平在EJ细胞表达高于膀胱移行上皮细胞,而p16^INK4a和p14^ARFmRNA及蛋白在EJ细胞表达水平稍低于膀胱移行上皮细胞。siRNA干扰 BMI-1 后可以上调EJ细胞中其下游p16和p14 mRNA及蛋白水平,使EJ细胞增殖能力下降,细胞凋亡增多。结论: siRNA干扰 BMI-1 基因表达对EJ细胞的体外生长具有明显的抑制作用,其作用机制与上调其下游 p16^INK4a、p14^ARF 基因的表达有关。     

Stable and uniform gene suppression by site-specific integration of siRNA expression cassette in murine embryonic stem cells

We developed a simple system to introduce small interfering RNA (siRNA) into murine embryonic stem cells (ESCs) and then showed its stable and uniform expression. Using hypoxanthine guanine phosphoribosyl transferase 1 (Hprt)-deficient ESCs as a recipient, we efficiently introduced an siRNA expression cassette into the Hprt locus by homologous recombination, which was easily detected by HAT selection. Nearly all of the HAT-resistant clones exhibited a silenced expression of the exogenous target gene (enhanced green fluorescent protein [EGFP]) or the endogenous target gene (Flk1). Flow cytometry profiles demonstrated that there were no significant differences in level of suppression among individual clones and cells. The suppressing effect by siRNA was maintained for more than 1 month in both undifferentiated and differentiated ESCs, while its persistent expression did not disturb their growth or differentiation potential. The stable and uniform suppression capability of this system will help to screen genes and provide important information regarding cell differentiation in ESCs.     

针对HLA-G的siRNA的载体构建与表达及效应检测

目的构建针对HLA-G的siRNA的表达载体,研究其对NK细胞杀伤效应的影响。方法构建针对HLA-G的siRNA真核表达质粒pSuppressor-U6-neo-HLA-G,转染JEG-3滋养细胞系。采用RT-PCR、Western blot检测转染的JEG-3细胞中HLA-G的表达水平。将NK-92MI细胞(效应细胞)与滋养细胞(靶细胞)共同培养,以LDH释放法观察不同效靶比例时NK-92MI细胞对靶细胞的杀伤效应。结果NK-92MI细胞对转染针对HLA-G的siRNA的JEG-3细胞的杀伤作用较未转染细胞明显升高(P<0.001)。结论针对HLA-G的SiRNA增加NK细胞对滋养细胞的杀伤,HLA-G具有保护滋养细胞免受NK细胞杀伤的作用。     

Suppression of inflammatory gene expression in T cells by Porphyromonas gingivalis is mediated by targeting MAPK signaling

There is increasing awareness of the effects of Porphyromonas gingivalis on host immune responses. Degradation of cytokines and chemokines by cysteine proteinases has previously been reported. However, the precise mechanisms by which P. gingivalis is able to alter intracellular signaling, and thus proliferation and inflammation, have not been described. We have previously reported suppression of activator protein-1 (AP-1) and degradation of IL-2 by proteinases from P. gingivalis. In the present study, we have analyzed the effects of P. gingivalis on Jurkat T-cell signal transduction and subsequent IL-2 and CXCL8 expression. We found that CXCL8, but not IL-2, gene expression levels were significantly suppressed by viable P. gingivalis. Analysis of intracellular signaling revealed an inhibitory effect of P. gingivalis on c-Jun and c-Fos, but not NFκB (p50 and p65), NFAT or STAT5 expression. This inhibitory effect was not due to suppression of mitogen-activated protein kinase (MAPK) (p38, erk and JNK) gene expression, but was rather due to prevention of protein kinase C (PKC) and p38 phosphorylation, as demonstrated by western blot analysis. Furthermore, SOCS1 and SOCS3 expression levels decreased following treatment of Jurkat T cells with viable P. gingivalis. The results indicate that P. gingivalis is able to suppress inflammatory gene expression by targeting the activity of MAPK pathways in T cells, which was confirmed by using specific inhibitors of NF-κB, PKC, ERK, p38 and JNK.     

High transfection efficiency, gene expression, and viability of monocyte-derived human dendritic cells after nonviral gene transfer

Dendritic cells (DCs) are bone marrow-originated, professional antigen-capturing cells and APCs, which can function as vaccine carriers. Although efficient transfection of human DCs has been achieved with viral vectors, viral gene products may influence cellular functions. In contrast, nonviral methods have generally resulted in inefficient gene transfer, low levels of gene expression, and/or low cell viability. Monocyte-derived DCs are the most common source of DCs for in vitro studies and for in vivo applications. We hypothesized that reduction of the time to generate immature DCs (iDCs) might result in higher viability after transfection. Therefore, we established a protocol to generate human iDCs from CD14(+) monocytes within 3 days. These "fast" iDCs were phenotypically and functionally indistinguishable from conventional iDCs, showing high endocytic ability and low antigen-presenting capacity. Furthermore, the fast iDCs matured normally and had similar antigen-presenting capacity to conventional mature DCs. To optimize transfection of iDCs, we compared nonviral transfection of plasmid DNA and in vitro-transcribed (IVT) RNA with transfection reagents, electroporation, and nucleofection. Nucleofection of IVT RNA with the X1 program of an Amaxa Co. Nucleofector resulted in the most efficient transfection, with an average of 93% transfected iDCs, excellent long-term viability, and strong protein expression. Furthermore, the IVT RNA-transfected iDCs retained all phenotypic and functional characteristics of iDCs. This method is applicable to most purposes, including in vitro functional assays, in vivo DC immunotherapy, and DC-based vaccines.     

siRNA沉默转酮醇酶样基因1下调人宫颈癌细胞HIF-1α表达及糖酵解关键酶活性

 目的：通过siRNA技术沉默宫颈癌细胞转酮醇酶样基因-1（TKTL1）后观察其在缺氧条件下低氧诱导因子1α（HIF-1α）表达水平及糖酵解途径中2个关键酶己糖激酶Ⅱ（HK-Ⅱ）和乳酸脱氢酶（LDH）活性的变化情况,旨在探讨肿瘤细胞TKTL1表达与糖酵解途径的关系。方法：用构建的靶向TKTL1基因siRNA干扰质粒载体转染HeLa细胞株,RT-PCR检测TKTL1 mRNA表达并结合转酮醇酶（TKT）活性测定判断其干扰效率;以未转染HeLa细胞为对照,进一步观察TKTL1沉默的癌细胞在缺氧条件下HIF-1α和HK-Ⅱ表达水平,以及HK-Ⅱ和LDH活性的变化。结果：siRNA沉默HeLa细胞TKTL1基因后TKTL1 mRNA表达下降,TKT活性显著降低（P<0.01）,HIF-1α和HK-Ⅱ表达水平及HK-Ⅱ和LDH活性较对照组细胞均显著降低（P<0.01）。结论：沉默肿瘤细胞TKTL1基因能下调HIF-1α表达并降低糖酵解代谢关键酶活性,从而影响肿瘤细胞的恶性表型。