Natural killer (NK) and antibody-dependent cellular cytotoxicity (ADCC) activities can be differentiated by their different sensitivities to interferon and prostaglandin E1

Though purported to be identical cells (or in identical populations of cells), the natural killer (NK) cell mediating spontaneous natural cytotoxicity and the killer (K) cell mediating antibody-dependent cellular cytotoxicity (ADCC) may not be totally identical, at least in susceptibility to regulation by the immunomodulators prostaglandin E₁ (PGE₁) and interferon (IFN). We demonstrate here that NK cells are always enhanced by IFN, while K cells are inhibited from binding targets, resulting in fewer effectors at optimal concentrations of antibody. Only at 10- to 100-fold suboptimal concentrations of antibody is ADCC activity enhanced. As measured by magnitude of inhibition and dose-response titration, ADCC activity is less sensitive to the effects of PGE₁ than is NK activity in the⁵¹Cr release assay and single-cell assay. After overnight incubation with or without PGE₁, whatever sensitivity ADCC activity had to PGE₁ is lost. However, NK cells incubated in the presence of PGE₁ overnight are still sensitive to inhibition. Indomethacin boosts NK activity without having any effect on ADCC activity. Finally, NK activity is substantially reduced by overnight incubation of cells at room temperature, which has no effect on K cells.     

siRNA抑制ErbB2基因的表达及对乳腺癌细胞生长的影响

目的:设计并构建ErbB2的小干扰RNA,检测其对ErbB2蛋白表达的干扰效果,并检测其对乳腺癌细胞ZR75-1生长的影响。方法:设计2条针对ErbB2的siRNA,并克隆到siRNA表达载体pSliencer 2.1-U6 neo上。经酶切和测序证明构建成功后,将其与pcDNA3-FLAG-ErbB2共转染于293T细胞,以及单转siRNA于乳腺癌SKBR3和ZR75-1细胞,通过Western blot分别检测siRNA对外源和内源ErbB2的干扰效果。通过结晶紫实验研究siRNA对ZR75-1生长的影响。结果:Western blot证明构建的两条ErbB2 siRNA均能有效抑制外源和内源ErbB2蛋白的表达,并抑制乳腺癌ZR75-1细胞的生长。结论:构建的siRNA能有效地抑制ErbB2蛋白的表达并抑制ZR75-1细胞的生长。     

Azathioprine suppression of natural killer activity and antibody-dependent cellular cytotoxicity in renal transplant recipients

The relative effects of azathioprine (AZA) and prednisone (PRED) on natural killer (NK) activity and the antibody-dependent cellular cytotoxicity (ADCC) of K (killer) cells and the number of FcR and other lymphoid cells were examined in renal transplant recipients. In addition to both long-term (>6 months) and short-term (<6 months) transplant recipients receiving conventional AZA-PRED therapy, an important group of long-term recipients receiving PRED but not AZA was studied for the first time. Both NK activity and ADCC are profoundly reduced in the long-term AZA-PRED group but are normal in the long-term PRED-alone (no-AZA) group. The short-term AZA-PRED group exhibits NK and ADCC levels significantly lower than normal but not as low as those of the long-term AZA-PRED group. Patient groups with low NK and ADCC also have low circulating Fc receptor-bearing (FcR) cells. A single patient in the long-term AZA-PRED group was removed from AZA therapy, and approximately 3 months was required for the patient's suppressed NK and ADCC to return to normal. These findings indicate that AZA rather than PRED is the major drug important in suppressing ADCC and NK activity in renal transplant recipients. Several months are required for combination AZA-PRED therapy to reduce these cytotoxic activities. Similarly, several months are required for suppressed ADCC and NK activity to return to normal upon discontinuation of AZA.     

Snail plays an oncogenic role in glioblastoma by promoting epithelial mesenchymal transition

Background: The factors affecting glioblastoma progression are of great clinical importance since dismal outcomes have been observed for glioblastoma patients. The Snail gene is known to coordinate the regulation of tumor progression in diverse tumors through induction of epithelial mesenchymal transition (EMT); however, its role in glioblastoma is still uncertain. Therefore, we aimed to further define its role in vitro. Methods and results: The small interfering RNA (siRNA) technique was employed to knock down Snail expression in three glioblastoma cell lines (KNS42, U87, and U373). Specific inhibition of Snail expression increased E-cadherin expression but decreased vimentin expression in all cell lines. In addition, inhibition of the expression of Snail significantly reduced the proliferation, viability, invasion, and migration of glioblastoma cells as well as increased the number of cells in the G1 phase. Conclusions: Knockdown of Snail suppresses the proliferation, viability, migration, and invasion of cells as well as inhibits cell cycle progression by promoting EMT induction. The findings suggest that expression of this gene facilitates glioblastoma progression. Therefore, these results indicate the clinical significance of Snail for use as a potential therapeutic target for glioblastoma.     

Inhibition of West Nile Virus replication by retrovirus-delivered small interfering RNA in human neuroblastoma cells

West Nile virus (WNV) has been responsible for the largest outbreaks of arboviral encephalitis in U.S. history. No specific drug is currently available for the effective treatment of WNV infection. To exploit RNA interference as a potential therapeutic approach, a Moloney murine leukemia virus-based retrovirus vector was used to effectively deliver WNV-specific small interfering RNA (siRNA) into human neuroblastoma HTB-11 cells. Viral plaque assays demonstrated that transduced cells were significantly refractory to WNV replication, as compared to untransduced control cells (P < 0.05), which correlated with the reduced expression of target viral genes and respective viral proteins. Therefore, retrovirus-mediated delivery of siRNA for gene silencing can be used to study the specific functions of viral genes associated with replication and may have potential therapeutic applications.     

Natural Killer Cytotoxicity and Antibody-Dependent Cell Cytotoxicity to Herpes Simplex Virus-Infected Target Cells in Murine Pregnancy

ABSTRACT: In vitro natural killer cytotoxicity (NKC) and antibody-dependent cell cytotoxicity (ADCC) activity against herpes simplex virus (HSV)-infected cells were evaluated in a pregnant murine model (C₅₇B1₆inbred strain). Virgin (n = 16) and pregnant (late gestation) mice (n = 15) were infected intraperitoneally with HSV, type 1. After 18 hr, a 0.5-ml aliquot of the peritoneal wash was frozen for virus plaque assay, and the cells were cultured in the ⁵¹chromium release assay for NKC and ADCC. %NKC (mean ± S.E.) to HSV-infected targets was significantly suppressed (P < 0.05) in pregnant mice, 10.3% ± 1.9, compared to that of virgin mice, 32.5% ± 2.5. This suppression was abrogated with HSV-specific antisera (%ADCC); 53.9% ± 4.4 (pregnant) compared to 49.1% ± 3.6 (virgin). The diminished NKC activity in pregnant mice was reflected in an increased mean number of virus particles in the peritoneal wash, 266 + 66 PFU/ml, compared to 38 ± 11 PFU/ml in virgin mice (P < 0.05). We concluded that NKC, but not ADCC, to HSV-infected targets was suppressed and that HSV elimination was impaired in pregnant mice.     

RNA interference-mediated inhibition of brain-derived neurotrophic factor expression increases cocaine's cytotoxicity in cultured cells

Previous studies showed that cocaine exposure decreased brain-derived neurotrophic factor (BDNF) function and resulted in neuronal cell death. To investigate a role of BDNF in cocaine's cytotoxicity, an RNA interference (RNAi) approach was used. Transfection of neuroblastoma SK-N-AS cells or primary rat hippocampal neurons with the small double-stranded interfering RNA (siRNA) targeting BDNF mRNA, but not the scrambled siRNA, resulted in reductions in levels of BDNF mRNA and proteins by more than 70% in the transfected cells as compared with the control group, suggesting an RNAi-mediated, sequence-specific gene silencing. The results also showed that cocaine-induced cytotoxicity, assessed by the MTT (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazodium bromide) assay, was more pronounced in the cells transfected with the siRNA than in the cells transfected with the scrambled siRNA or in the cells treated with Lipofectamine 2000 alone (the control group), suggesting that inhibition of BDNF expression enhances cocaine's cytotoxicity. Together with previous studies showing that cocaine suppresses BDNF expression, the present data suggest that the drug-induced reduction of BDNF productions may make neurons more vulnerable to cocaine's toxic effects and precipitate cocaine-induced central nervous system damages.     

沉默c-myc基因对Jiyoye细胞增殖与凋亡的影响

目的 运用RNA干扰（RNAi）技术,探讨经慢病毒载体介导的小干扰RNA（siRNA）转染Jiyoye细胞对肿瘤细胞增殖及凋亡的影响,以期为靶向c-myc基因的白血病和恶性淋巴瘤基因治疗提供体外实验依据。方法 设计、合成一对针对c-myc基因的c-myc-3寡聚核苷酸链和一对阴性对照c-myc- neg寡聚核苷酸链。退火,然后连接到pLVX干扰载体上,包装,慢病毒介导转染人Jiyoye细胞株。转染后培养72 h,流式细胞术检测各组细胞转染率,pLVX-c-myc-3转染Jiyoye细胞,MTT法检测细胞增殖;Annexin V-异硫氰酸荧光素/碘化吡啶（FITC/PI）双染流式细胞术检测细胞凋亡。结果 ①转染72 h后,流式细胞术鉴定阳性细胞比例,c-myc-neg组与c-myc-3组比较,差异无统计学意义（P 〉 0.05）[（54.3 ± 4.2） % vs（52.8 ± 2.9） %];两组细胞平均荧光强度差异、细胞转染率差异无统计学意义。②转染168 h生长曲线显示,转染pLVX-c-myc实验组细胞生长缓慢,与阴性对照组和空白对照组比较,差异有统计学意义（P 〈 0.05）。转染pLVX-c-myc 72 h后,实验组细胞抑制率为（46.40 ± 1.41） %,阴性对照组细胞抑制率为（17.03 ± 1.32） %,差异均有统计学意义（P 〈 0.05）。③转染pLVX-c-myc 72 h后,实验组早期凋亡细胞比例（25.6 ± 4.2） %,阴性对照组早期凋亡细胞（15.1 ± 4.2） %,空白对照组早期凋亡细胞为（12.7 ± 1.8） %,差异有统计学意义（P 〈 0.05）。实验组晚期凋亡细胞比例（11.5 ± 4.7） %,阴性对照组为（9.3 ± 4.7） %,空白对照组为（8.14 ± 2.8） %,差异有统计学意义（P 〈 0.05）。结论 设计合成构建的干扰序列pLVX-c-myc-3,可沉默c-myc基因,抑制Jiyoye细胞增殖,诱导细胞凋亡,为进一步探讨沉默c-myc基因在白血病和淋巴瘤靶向治疗中的作用提供了实验依据。     

Antibody-dependent cellular cytotoxicity (ADCC)-mediated destruction of human immunodeficiency virus (HIV)-coated CD4+ T lymphocytes by acquired immunodeficiency syndrome (AIDS) effector cells

The acquired immunodeficiency syndrome (AIDS) is defined in clinical terms by the development of Kaposi's sarcoma and/or severe opportunistic infections in persons without predisposing conditions. A hallmark of the syndrome has been a decrease in the number of CD4⁺ T helper cells. The reduction in the frequency of the CD4⁺ lymphocytes has been postulated to be primarily the result of human immunodeficiency virus (HIV) tropism and cytophathogenicity for the T-cell subset. Yet only a small percentage of cells is actually infected with HIV. Recently, we provided evidence indicating that AIDS patients' natural killer cells can mediate normal levels of antibody-dependent cellular cytotoxicity (ADCC) despite exhibiting a defect in natural killer (NK) effector function (J Immunol 139:55, 1987). This finding prompted us to investigate whether AIDS patients' effector cells could mediate ADCC against circulating CD4⁺ T cells infected with or expressing HIV antigen. The findings reported herein demonstrate that AIDS effector cells can mediate lysis of CEM (CD4⁺ T-cell line) coated with HIV protein in the presence of HIV-specific antibody. Lysis was specific, as non-HIV-coated CEM or the addition of HIV-negative serum resulted in no lysis. We then examined HIV-coated peripheral blood-derived CD4⁺ T lymphocytes as targets in ADCC. We demonstrate that in the presence of HIV-specific antibody, HIV-coated CD4⁺ T lymphocytes serve as targets for ADCC by AIDS effector cells. The lytic activity obtained with AIDS effector cells was comparable to that obtained with normal effector cells. These results demonstrate that AIDS effector cells can mediate ADCC against HIV-coated CD4⁺ T lymphocytes and suggest that ADCC may play a rolein vivo in the pathogenesis of AIDS.     

特异siRNA下调卵巢癌H08910细胞NAC-1基因表达并抑制生长

 目的：研究小RNA干扰NAC-1 (Nucleus accumbens-1,Nac1 or NAC-1)基因表达对卵巢癌HO8910细胞增殖的影响。方法：设计、合成针对NAC-1基因的特异性小干扰RNA（small interfering RNA,siRNA）及阴性siRNA,并同时设立空白对照,然后用脂质体转染HO8910细胞。通过实时荧光定量RT-PCR 和Western印迹法分别检测转染前后HO8910细胞中NAC-1 mRNA和蛋白表达水平,MTT法和克隆形成实验检测细胞增殖率,FCM法检测细胞周期分布。结果：针对NAC-1基因的特异性siRNA均能抑制NAC-1 mRNA和蛋白的表达,其中NAC-1-siRNA-1的沉默效率最高。转染NAC-1-siRNA-1 48h后,HO8910细胞中NAC-1 mRNA和蛋白表达水平分别下调74%和81%,而且细胞增殖明显受到抑制,细胞周期被阻滞于G1期,与转染阴性siRNA 及未转染组比较,有显著性差异(P?0.05)。结论：NAC-1特异性siRNA能够有效沉默NAC-1基因表达,并显著抑制卵巢癌HO8910细胞增殖。     

Cryopreservation of human mononuclear cells for quality control in clinical immunology. I. Correlations in recovery of K- and NK-cell functions,surface markers,and morphology

Cryopreserved human peripheral blood mononuclear cells (PBMC) were tested for natural killer (NK) and antibody-dependent cellular cytotoxicity (ADCC) and for high-affinity (29°C) and total (4°C) rosette formation with sheep erythrocytes. PBMC produced variable NK activity following freezing and thawing, but consistently reacted well in ADCC. A significant correlation was found between low NK activity and a decreased percentage of low-affinity rosette-forming cells. On the contrary, the number of large granular lymphocytes (LGL), among which NK cells are restricted, and the reactivity with the monoclonal antibody OKT10, which recognizes the majority of LGL in the peripheral blood, were not significantly altered by cryopreservation. Cryopreserved cells proved to be excellent controls for determining the day-to-day variability of the NK assay and for selecting optimum conditions for this test in the clinical immunology laboratory.     

Gene therapy for head and neck squamous cell carcinoma using KITENIN (KAI1 COOH-Terminal Interacting Tetraspanin)-antisense therapy

Purpose

KAI1 COOH-terminal interacting tetraspanin (KITENIN) has been found to act as a promoter of metastasis in murine models of colon cancer and squamous cell carcinoma (SCC). The suppression of tumor progression and metastasis of established colon cancer in mice was observed after intravenous delivery of small interfering RNA (siRNA) targeting KITENIN. The purpose of this study was to investigate the efficacy of gene therapy targeting KITENIN in human head and neck SCC.

Materials and Methods

SNU-1041, a well-established human hypopharyngeal SCC cell line, was used. KITENIN expression in SNU-1041 was measured by Western blot analysis. The cells were prepared, maintained in culture dishes with media, and divided into two groups: the si-KITENIN group and the scrambled group (control). The siRNA targeting KITENIN (si-KITENIN) and scrambled DNA were transfected into the SNU-1041 cells in each group. The effect of gene therapy was compared by in vitro experiments to evaluate invasion, migration, and proliferation.

Results

KITENIN was strongly expressed in the SNU-1041 cells, and the number of invaded cells was reduced more in the si-KITENIN group than in the scrambled group (p<0.001). The speed for the narrowing gap, made through adherent cells, was lower in the si-KITENIN group (p<0.001), and the number of viable proliferating cells was reduced in the si-KITENIN group compared to the scrambled group (p<0.001, the third day). KITENIN protein expression was no longer identified in the si-KITENIN group.

Conclusion

Gene therapy using an anti-KITENIN strategy might be effective for head and neck squamous carcinoma.     

以腺相关病毒为载体的小干扰RNA对肝星状细胞中金属蛋白酶组织抑制因子的抑制作用

目的 观察以腺相关病毒（AAV）为载体含有针对大鼠金属蛋白酶组织抑制因子（TIMP）-1具有较强抑制作用的小干扰RNA（siRNA）感染大鼠星状细胞系HSC-T6后TIMP-1的表达抑制作用。方法 针对大鼠TIMP-1mRNA基因序列挑选一段22bp片段，在体外构建为短发夹siRNA（short hairpin siRNA，shRNA）表达载体后，将其包装为重组AAV并感染大鼠肝星状细胞系HSC-T6后，于感染后30d及90d应用荧光定量PCR方法及Westem blot方法分别检测TIMP-1 mRNA及蛋白质表达情况，同时通过PCR技术以感染后细胞的基因组DNA为模板扩增外源基因验证其长效表达。结果 经PCR、酶切及序列测定证实含有siRNA-TIMP-1基因的重组AAV载体质粒已成功克隆。将重组质粒包装成病毒后感染HSC-T6细胞，与对照组细胞相比，感染后30d及90d细胞TIMP-1 mRNA水平明显降低（P〈0．01），感染后30d TIMP-1蛋白表达水平较对照组细胞相比下降约60％，而感染后90d，TIMP-1蛋白表达几乎下降90％。PCR结果显示在重组病毒感染后90d细胞基因组DNA中仍可扩增出外源基因，证实外源基因可长期表达。结论 重组病毒rAAV／siRNA-TIMP-1／neo可长期有效地抑制TIMP-1基因的表达。     

Increased insulin-like growth factor 1 receptor (IGF1R) expression in small cell lung cancer and the effect of inhibition of IGF1R expression by RNAi on growth of human small cell lung cancer NCI-H446 cell

Abstract

Insulin-like growth factor 1 receptor (IGF1R) is a tyrosine kinase receptor implicated in tumourigenesis that may be an attractive target for anti-cancer treatment. In this study, the expression and clinical significance of IGF1R were investigated in serum and lung cancer tissues from small cell lung cancinoma (SCLC). We also compared the effect of IGF1R up-regulation and IGF1R inhibition on viability and apoptosis of NCI-H446 cells. We found the concentration of IGF1R in blood serum was significantly increased and positive IGF1R protein in cancer tissue was more prevalent in SCLC. A statistically significant correlation among IGF1R-positve tumors, lymph node metastasis and local invasion was discussed. Furthermore, IGF1R overexpression lead to an increase of cell survival and suppressed cell apoptosis, IGF1R silencing mediated by RNAi abrogate this response of NCI-H446 cells. Our results further demonstrated that the effects of these treatments may be assigned to the effective inhibition of lung cancer cells from Akt/P27^Kip1 pathway in IGF-1R signaling. These features may have important implications for future anti-IGF1R therapeutic approaches.     

Targeting the Twist and Wnt signaling pathways in metastatic breast cancer

Breast cancer is the leading cause of cancer-related deaths in the United States with over 232,000 new diagnoses expected in 2014 and almost 40,000 deaths. While treatment of primary breast cancer is often well-managed with surgery and radiation, metastatic breast cancer (MBC) that has spread to the brain, bones, liver, and lungs is often incurable. One of the major challenges in the treatment of breast cancer is the presence of a subpopulation of cancer cells that are chemoresistant and metastatic. Given that metastasis is the driving force behind mortality for breast cancer patients, it is essential to identify the characteristics of these aberrant cancer cells that allow them to spread to distant sites in the body and develop into metastatic tumors. Understanding the metastatic mechanisms driving cancer cell dispersal will open the door to developing novel therapies that prevent metastasis and improve long-term outcomes for patients. In this review we assess the feasibility of targeting the Twist and Wnt signaling pathways in breast cancer. These pathways mediate epithelial-mesenchymal transition (EMT), a process that can give rise to chemoresistance. We review potential treatment strategies for targeting EMT and drug resistance as well as the problems that may arise with these targeted delivery therapeutic approaches. Finally, we examine recent advances in the field, including nanoparticle delivery and small interfering RNA (siRNA) technology, and discuss the impact that these approaches may have on translating much needed therapeutic approaches into the clinic, for the benefit of patients battling MBC.     

A novel long non-coding RNA NFIA-AS1 is down-regulated in gastric cancer and inhibits proliferation of gastric cancer cells

Gastric cancer is one of the most common malignant gastrointestinal tumors whose morbidity and mortality account for the second and third place respectively in malignant tumors in China. As an important participant in tumor biology, the abnormal expression of long non-coding RNA (lncRNAs) in cancer cells is closely related to the occurrence and development of tumors and plays the role of oncogenes or tumor suppressor genes. In this study, we identified a novel lncRNA NFIA antisense RNA 1 (NFIA-AS1) and explored its role and clinical significance in gastric cancer. Real-time quantitative PCR was performed to detect the expression of NFIA-AS1 in tumor tissues and corresponding normal tissues from 42 pairs of gastric cancer samples. The lower expression of NFIA-AS1 was significantly associated with larger tumor size, lower histological grade, and advanced TNM stage. Kaplan-meier analysis showed that NFIA-AS1 expression could be used as an independent predictor of overall survival. We also demonstrated that overexpression of NFIA-AS1 significantly inhibited the proliferation of gastric cancer cells through affecting p16 levels. In conclusion, our results suggest that the lncRNA NFIA-AS1 may play the role of tumor suppressor gene, and serve as a biomarker for prognosis or progression of gastric cancer.     

Selection of the expressed B cell repertoire by infusion of normal immunoglobulin G in a patient with autoimmune thyroiditis

In the present study we have analyzed the changes in the expressed antibody repertoire and in temporal fluctuations of antibody levels in serum that followed infusion of normal IgG (IVIg) in a patient with autoimmune thyroiditis. Administration of IVIg resulted in the stimulation of IgM production, in alterations of expressed antibody activity in serum that could not merely be accounted for by the passive transfer of antibody specificities contained in IVIg, in transient down-regulation of B cells clones expressing a specific disease-related idiotype and in the increase in serum in recipient's autoantibodies specifically reactive with F(ab′)₂ fragments of IVIg. In addition, infusion of IVIg shifted the pattern of spontaneous fluctuations of autoantibody activities in the patient's serum from a pattern indicative of disconnected events in the immune network to a pattern similar to that which is consistently observed in healthy controls. These results suggest that normal IgG may modulate autoreactivity by selecting expressed antibody repertoire through V region-dependent interactions with antibodies.     

Effects for Sequential Treatment of siAkt and Paclitaxel on Gastric Cancer Cell Lines

Real-time screening of cellular response on the drugs could provide valuable insights for the early detection of therapeutic efficiency and the evaluation of disease progression. Cancer cells have the ability to vary widely in response to stress in a manner to adjust the signaling pathway to promote the survival or having a resistance to stimulation. Cell-based label-free technologies using electronic impedance sensor have strategies for constructing the signature profiles of each cells. To achieve exquisite sensitivity to substantially change of live-cell response have an important role that predict the potential of therapeutic effects. In this study, we use an impedance-based real-time cell analysis system to investigate dynamic phenotypes of cells described as a cellular index value. We show that gastric cancer cells generated characteristic kinetic patterns that corresponded to the treatment order of therapeutics. The kinetic feature of the cells offers insightful information that cannot be acquired from a conventional single end-point assay. Furthermore, we employ a ''sequential treatment strategy'' to increase cytotoxic effects with minimizing the use of chemotherapeutics. Specifically, treatment of paclitaxel (PTX) after down-regulating Akt gene expression using RNAi reduces the cell proliferation and increases apoptosis. We propose that the sequential treatment may exhibit more effective approach rather than traditional combination therapy. Moreover, the dynamic monitoring of cell-drug interaction enables us to obtain a better understanding of the temporal effects in vitro.     

RNA干扰Notch1基因对乳腺癌MCF-7细胞增殖及凋亡的影响

目的探讨RNA干扰技术沉默Notch1基因表达对人乳腺癌细胞增殖和凋亡的影响。方法设计并合成靶向Notch1基因的小分子干扰RNA质粒,在转染试剂Sofast介导下转染人乳腺癌细胞株MCF-7,用RT-PCR和Western blot法检测转染前、后Notch1基因的表达,挑选干扰效率最强的一组表达载体;cck8比色法检测分析各组细胞的存活率;流式细胞术检测细胞凋亡比例;Western blot法检测转染各组MCF-7细胞Notch1、NF-κB及Caspase-3蛋白表达。结果 Notch1-shRNA能有效封闭Notch1基因的表达,Notch1基因和蛋白表达水平明显降低(P<0.05);Notch1-shRNA能明显抑制细胞增殖(P<0.05);转染48h后细胞凋亡比例增加(P<0.01)。NF-κB蛋白水平表达降低,Caspase-3蛋白表达水平增高。结论利用RNA干扰技术沉默Notch1基因的表达可以明显抑制MCF-7细胞的增殖,促进MCF-7细胞凋亡,其机制可能通过NF-κB信号通路调节相关凋亡蛋白的表达,进而影响细胞的凋亡和增殖,靶向Notch1的RNA干扰技术在乳腺癌的基因治疗中具有一定的研究价值。     

强心苷类化合物对肺癌细胞A549和H1975的不同生长抑制作用

目的研究3种强心苷药物地高辛、吉妥辛和狄吉妥辛对体外培养的人肺腺癌细胞系A549(EGFR野生型/KRAS突变型)和H1975(EGFR突变型/KRAS野生型)的抑制作用。方法 3种强心苷药物处理A549和H1975后,用MTS法检测细胞药物敏感度,流式细胞术检测细胞凋亡率和增殖情况。结果 3种强心苷药物对A549半数抑制浓度均大于10μmol/L,而对H1975在0.1μmol/L左右。达到相同程度的凋亡率和增殖抑制,A549所需药物浓度是H1975的10倍以上。结论强心苷类药物对H1975比A549具有更显著的诱导凋亡和抑制增殖的作用。