丹参酮ⅡA对高糖诱导人脐静脉内皮细胞凋亡的作用

目的:观察丹参酮ⅡA对高糖刺激人脐静脉内皮细胞凋亡的影响,并探讨相关的作用机制。方法:四甲基偶氮唑蓝(MTT)法检测细胞存活率;流式细胞术Annexin V-FITC/PI双染法定量检测内皮细胞凋亡情况;免疫印迹法检测抗凋亡分子Bcl-2、促凋亡分子Bax以及细胞色素C(cytochrome C,Cyt C)的蛋白表达水平。结果:丹参酮ⅡA可抑制高糖诱导的人脐静脉内皮细胞存活率的减少以及凋亡率的增加。此外,丹参酮ⅡA处理后,Bcl-2蛋白表达显著增加,Bax蛋白表达显著减少,并且线粒体Cyt C释放受到明显抑制。结论:丹参酮ⅡA可以通过调节线粒体凋亡信号通路相关蛋白表达而对抗高糖诱导内皮细胞的凋亡。     

丹参酮ⅡA激活Nrf2/Nox4通路减轻脂多糖诱导的小鼠肺炎和纤维化

目的研究丹参酮ⅡA(Tan ⅡA)对脂多糖诱导小鼠肺炎和纤维化影响,探讨其潜在的作用机制。方法30只8周龄雄性C57BL/6小鼠,随机分成对照组、脂多糖(LPS)组和LPS+Tan ⅡA组。采用尾静脉注射LPS制作小鼠肺炎和纤维化动物模型,术后给予Tan ⅡA干预,每日一次,连续2周,实验第4周处死所有小鼠,收集血清和肺组织标本。采用HE染色检测肺组织炎症改变,Masson染色观察肺组织纤维化程度,ELISA检测血清IL-1β、TNF-α和IL-10蛋白表达,采用Western blot技术检测肺组织Nrf2、Nox4、Nqo1和Ho-1蛋白表达。结果与对照组相比,LPS组小鼠肺脏重量、肺脏/体重比显著增加,肺组织水肿、炎细胞浸润和纤维化明显加重,血清促炎蛋白IL-1β、TNF-α显著升高,抑炎蛋白IL-10表达下降(P<0.05)。Tan ⅡA处理显著减轻LPS导致的小鼠肺脏重量、肺脏/体重比,减轻肺水肿、炎细胞浸润和纤维化,减低血清IL-1β、TNF-α表达水平,升高IL-10表达水平。此外,Nrf2和Nox4蛋白表达水平LPS组高于对照组,Nqo1和Ho-1蛋白表达水平低于对照组,而Tan ⅡA处理可以上调Nrf2、Nqo1和Ho-1蛋白,降低Nox4蛋白表达水平(P<0.05)。结论Tan ⅡA减轻LPS诱发的小鼠炎症和纤维化,与激活Nrf2/Nox4通路相关。     

Nox4在同型半胱氨酸介导的动脉粥样硬化中作用的研究

同型半胱氨酸（Hcy）是心血管疾病的独立危险因子之一。Hcy能够诱导细胞活性氧（ROS）的生成增多引起内皮损伤从而导致动脉粥样硬化（AS）发生发展。内皮细胞中ROS主要来源于Nox4（NADPH oxidases 4）。NLRP3在冠状动脉粥样硬化及主动脉硬化中高表达。NLRP3是AS中IL-1家族细胞因子产生的关键介质,特别是当细胞内产生的大量ROS,可导致caspase-1前体大量剪切成具有生物活性的caspase-1 P10片段,推动下游IL-1β和IL-18等炎症因子的成熟及分泌,使内皮细胞功能受损,引起细胞炎症损伤及凋亡,正反馈加重血管损伤。本文主要探讨Nox4在Hcy介导的内皮细胞炎症及凋亡的作用,Hcy导致的AS提供可能的机制。     

丹参酮IIA对低氧条件下人肝癌HepG2细胞增殖、凋亡的影响及与HIF-1α、VEGF和野生型P53蛋白表达的关系

 目的:探讨低氧条件下丹参酮IIA（Tan IIA）对人肝癌HepG2细胞增殖和凋亡的影响及其分子机制。方法:用氯化钴（CoCl2）创建低氧模型,实验分为常氧对照组、低氧对照组和低氧+Tan IIA处理组。不同浓度的Tan IIA 分别作用于低氧下人肝癌HepG2细胞24 h、48 h和72 h,采用MTT法测定Tan IIA 对低氧下HepG2细胞增殖的抑制作用。不同浓度的Tan IIA 分别作用于低氧条件下HepG2细胞24 h和48 h后,Hoechst 33258染色法检测细胞核的形态学变化并计算凋亡率。不同浓度的Tan IIA 作用于低氧条件下人肝癌HepG2细胞48 h后,Western blotting检测低氧诱导因子1α (HIF-1α)、血管内皮生长因子（VEGF）和野生型P53的蛋白表达情况。结果:低氧条件下Tan IIA以时间和剂量依赖方式抑制HepG2细胞的生长和增殖。Tan IIA 作用于低氧下的HepG2细胞后,可见典型的凋亡细胞形态学特征,细胞凋亡率呈时间、剂量依赖性的增加。Western blotting免疫印迹法显示常氧对照组的HIF-1α和VEGF表达较低,而低氧对照组的HIF-1α、VEGF蛋白表达较常氧组升高,低氧下随着Tan IIA 浓度的升高,HIF-1α和VEGF蛋白的表达明显降低,野生型P53蛋白的表达随着Tan IIA 浓度的升高而升高。结论: 低氧条件下,Tan IIA能抑制肝癌HepG2细胞增殖并诱导其凋亡,其机制可能与抑制HIF-1α和VEGF蛋白表达,上调P53蛋白表达有关。     

Tanshinone IIA Attenuates Bleomycin-Induced Pulmonary Fibrosis via Modulating Angiotensin-Converting Enzyme 2/ Angiotensin-(1-7) Axis in Rats

Pulmonary fibrosis (PF) is a common complication in those interstitial lung diseases patients, which will result in poor prognosis and short survival. Traditional therapeutic methods such as glucocorticoid and cytotoxic drugs are insufficient for treating PF and may cause severe side effects. Recent studies showed that traditional Chinese herbal abstraction such as Tanshinone IIA (TIIA) was displayed significant anti-PF effects in animal models. However, the exact mechanisms underlying the protective effects of TIIA were not fully understood. Here we further investigated the protective effects of TIIA and its mechanisms underlying. PF models of rat were induced by bleomycin (BLM); TIIA was administered subsequently. The PF changes were identified by histopathological analyses. The results showed that BLM resulted in severe PF and alveolar inflammation; together with significant elevation of transforming growth factor-β 1 (TGF-β1). Angiotensin-converting enzyme 2 (ACE-2) together with angiotensin-(1-7) [ANG-(1-7)] were both greatly reduced after BLM administration. TIIA treatment notably attenuated BLM induced PF and inflammation, decreased expression of TGF-β1 and reversed ACE-2 and ANG-(1-7) production in rat lungs. Thus we may draw the conclusion that TIIA may exert protective effects on BLM induced PF in rats, and the ACE-2/ANG-(1-7) axis may ascribe to those protective effects.     

Changes in phosphate metabolism in thymoma cells suggest mechanisms for resistance to dexamethasone-induced apoptosis. A 31P NMR spectroscopic study of cell extracts

Treatment of the mouse thymoma-derived WEHI7.2 cell line with dexamethasone, a synthetic glucocorticoid, causes the cells to undergo apoptosis. Previous studies have shown that WEHI7.2 cell variants with an increased antioxidant defense exhibit increased resistance to dexamethasone-induced apoptosis, suggesting that oxidative stress may play a role in glucocorticoid-induced apoptosis. In this work we compared metabolic profiles of WEHI7.2 parental cells with those of WEHI7.2 variants with an increased antioxidant defense or overexpressing bcl-2, to determine whether bolstering the antioxidant defense results in altered metabolic parameters that could translate into increased resistance to dexamethasone-induced apoptosis. WEHI7.2 parental cells and cells overexpressing catalase, thioredoxin or bcl-2, or selected for resistance to 200 micro M H(2)O(2) were cultured in low-glucose DMEM medium supplemented with 10% calf serum, and extracted using chloroform-methanol-water (1:1:1). Metabolites contained in the aqueous and organic phases of the extracts were processed separately and subjected to high-resolution (31)P NMR spectroscopy. In most of the steroid-resistant variants, ATP levels and energetic status were decreased compared with the steroid-sensitive parental cell line, while the concentrations of hexose and triose phosphates were increased. Furthermore, the ratio of choline-containing phospholipids to ethanolamine-containing phospholipids was generally reduced in steroid-resistant cells. Phosphatidylethanolamine and its derivatives contain a higher amount of polyunsaturated fatty acids (PUFA) than the choline-containing analogs, and PUFA are readily oxidized by reactive oxygen species. Therefore, an increased initial amount of phosphatidylethanolamine may increase the 'buffering capacity' of this antioxidant and may thus contribute to the steroid resistance of WEHI7.2 variants.     

HIV-1 triggers apoptosis in primary osteoblasts and HOBIT cells through TNFalpha activation

Several HIV-1 infected patients show bone loss and osteopenia/osteoporosis during the course of disease. The mechanisms underlying this degenerative process are largely unsettled and it has not been determined yet whether bone dysfunction is linked to HIV-1-mediated direct and/or indirect effects on osteoblasts/osteoclasts cross-talk regulation. This study investigated the effects of HIV-1(IIIb) and HIV-1(ADA) strains on osteoblasts using the osteoblast-derived cell line (HOBIT) and primary human osteoblasts as cellular models. The challenge of these cell cultures by both HIV-1 strains triggered a significant apoptosis activation unrelated to viral infection, since proviral HIV-1 DNA and supernatant HIV-1 RNA were not detected by real time PCR or b-DNA assays respectively. Under the experimental conditions, even heat-inactivated HIV-1 or cross-linked recombinant gp120 treatment of HOBIT and osteoblasts induced programmed cell death, suggesting that apoptosis is regulated by the interaction between HIV-1 gp120 and cell membrane. The analysis of cell culture supernatants showed a significant up-regulation of TNFalpha, a pleiotropic protein considered an apoptosis inducer in the osteoblast model. In fact, pretreatment of HOBIT and osteoblast cell cultures with anti-TNFalpha polyclonal antibody tackled effectively HIV-1 related induction of cell apoptosis. As a whole, these results indicate that HIV-1 may impair bone mass structure homeostasis by TNFalpha regulated osteoblast apoptosis.     

Tetrahydroxystilbene glucoside attenuates MPP-induced apoptosis in PC12 cells by inhibiting ROS generation and modulating JNK activation

It is known that oxidative stress plays a major role in the progression of Parkinson's disease (PD). Previous studies have suggested that 2,3,5,4′-tetrahydroxystilbene-2-O-β-d-glucoside (TSG), an active component extracted from a traditional Chinese herb Polygonum multiflorum Thunb., has significant antioxidant and free radical-scavenging activities. This is the first study that investigated the protective effects of TSG against MPP⁺-induced apoptosis in PC12 cells and determined the underlying mechanism. The results showed that incubation of PC12 cells with TSG before exposing them to MPP⁺ could significantly decrease cell viability loss and reverse cell apoptosis in a dose-dependent manner. The anti-apoptotic effects of TSG were probably mediated via the inhibition of ROS generation and modulation of JNK activation because TSG blocked ROS increase and JNK phosphorylation induced by MPP⁺. Taken together, these results indicated that TSG may provide a useful therapeutic strategy for the treatment of neurodegenerative diseases such as PD.     

白花丹醌对人肝星状细胞Nox4/ROS及α-SMA生成的影响

 目的: 研究白花丹醌(plumbagin)对转化生长因子β1(TGF-β1)刺激的体外培养人肝星状细胞(HSC-LX2)NADPH氧化酶4(Nox4 )的mRNA和蛋白表达、活性氧簇(ROS)水平及α-平滑肌肌动蛋白(α-SMA)蛋白表达的影响。方法: 体外培养HSC-LX2,随机设立空白组、TGF-β1刺激的模型组、TGF-β1+ plumbagin (2 μmol/L、1.5 μmol/L及1 μmol/L)组;细胞与各药物共同孵育72 h后,采用RT-PCR检测细胞Nox4 mRNA的表达;原位装载探针法测定细胞内ROS的水平;Western blot检测细胞内Nox4和α-SMA的蛋白含量。结果: 与模型组比较,白花丹醌作用72 h后,2 μmol/L和1.5 μmol/L plumbagin能明显降低HSC-LX2细胞Nox4 mRNA和蛋白的表达(P<0.01),下调ROS水平,降低α-SMA的蛋白表达(P<0.01)。结论: Plumbagin可能是通过下调Nox4的表达进而降低ROS生成从而发挥其抑制HSC-LX2细胞活化的作用。     

RETRACTED: Resibufogenin inhibited colorectal cancer cell growth and tumorigenesis through triggering ferroptosis and ROS production mediated by GPX4 inactivation

Resibufogenin (RB) has been used for cancer treatment, but the underlying mechanisms are still unclear. This study aimed to investigate the effects of RB treatment on colorectal cancer (CRC) cells, and to determine the underlying mechanisms. The cell counting kit-8 assay was used to determine cell viability. Cell morphology was observed under light microscopy, and terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling assay was employed to detect cell apoptosis. Intracellular ferrous iron (Fe²⁺), malondialdehyde (MDA), glutathione (GSH), and reactive oxygen species levels were detected by using commercial iron assay kit, MDA assay kit, GSH assay kit, and 2,7-dichlorodihydrofluorescein diacetate probes, respectively. The protein expressions were determined by Western blot and immunohistochemistry. RB inhibited cell viability in the CRC cell lines (HT29 and SW480) in a dose- and time-dependent manner, and caused cytotoxicity to the normal colonic epithelial cell line (NCM460) at high dose. Similarly, RB induced morphological changes in CRC cells from normal to round shape, and promoted cell death. Of note, RB triggered oxidative stress and ferroptotic cell death in CRC cells, and only ferroptosis inhibitors (deferoxamine and ferrostatin-1), instead of inhibitors for other types of cell death (apoptosis, autophagy, and necroptosis), reversed the inhibitory effects of RB on CRC cell proliferation. Furthermore, glutathione peroxidase 4 (GPX4) was inactivated by RB treatment, and overexpression of GPX4 alleviated RB-induced oxidative cell death in CRC cells. Consistently, the in vivo experiments validated that RB also triggered oxidative stress, and inhibited CRC cells growth and tumorigenicity in mice models. RB can inhibit CRC cells growth and tumorigenesis by triggering ferroptotic cell death in a GPX4 inactivation-dependent manner.     

红景天苷对小鼠腹腔巨噬细胞体外增殖、凋亡、吞噬、ROS和NO产生的影响

目的:研究红景天苷(Sal)对小鼠腹腔巨噬细胞体外增殖、凋亡、吞噬、胞内活性氧簇(ROS)及分泌一氧化氮(NO)的影响,初步探讨其对小鼠腹腔巨噬细胞的免疫调节作用。方法:无菌分离小鼠腹腔巨噬细胞,并制备单细胞悬液,以不同终浓度(80μmol/L、160μmol/L及320μmol/L)的Sal和巨噬细胞共培养4 h,再以脂多糖(LPS)和γ-干扰素(IFN-γ)进行共刺激。利用MTT比色法检测Sal对巨噬细胞体外增殖的影响。用放线菌酮(CHX)诱导巨噬细胞凋亡,用Sytox G reen染色结合荧光酶标仪检测Sal对CHX诱导巨噬细胞凋亡的影响。用流式细胞术(FCM)检测Sal对巨噬细胞吞噬功能的影响。用2-7-二氯氢化荧光素乙二脂(H2DCFDA)染色法结合荧光酶标仪检测Sal对胞内ROS产生的影响;用G riess反应检测Sal对巨噬细胞分泌NO的影响。结果:MTT比色法检测显示,终浓度为80、160、320μmol/L的Sal均可显著促进LPS+IFN-γ刺激巨噬细胞增殖(P<0.05)。荧光酶标仪检测Syto xG reen染色法的结果显示,160μmol/L的Sal可抑制CHX诱导的巨噬细胞凋亡(P<0.01)。FCM结果显示,各浓度的Sal均能促进单纯药物组和实验药物组LPS+IFN-γ刺激巨噬细胞的吞噬功能(P<0.05)。用荧光酶标仪检测DH2DCFDA染色结果表明,各浓度的Sal对LPS+IFN-γ刺激的巨噬细胞胞内ROS的产生均具有显著的抑制作用(P<0.01)。Griess反应检测NO含量的结果显示,各浓度的Sal对LPS+IFN-γ刺激巨噬细胞产生NO均具有促进作用(P<0.05)。结论:Sal对LPS和IFN-γ刺激的巨噬细胞增殖具有显著的促进作用,对CHX诱导的巨噬细胞凋亡具有显著的抑制作用,对静息态和活化态的巨噬细胞的吞噬功能均有增强作用,并能减少LPS和IFN-γ活化的巨噬细胞胞内ROS的产生;但能促进LPS和IFN-γ活化的巨噬细胞NO的分泌。     

Down-regulation of nectin-4 inhibits apoptosis in systemic lupus erythematous (SLE) through targeting Bcl-2/Bax pathway

Purpose: The purpose of this study was to investigate the potential role of nectin-4 in systemic lupus erythematous (SLE) cell apoptosis during the disease development and its potential mechanism. Methods: Human peripheral blood mononuclear cells (PBMCs) were obtained for the isolation of monocytes and T lymphocytes. siRNA-nectin-4 plasma was constructed for the transfection into T cells using Lipofectamine 2000 reagent. siRNA with no nectin-4 sequence was transfected into T cells for the control group. mRNA expression of nectin-4 in cells was analyzed using RT-PCR method. Effect of netin-4 expression on T cell apoptosis was analyzed with Annexin V-FITC cell apoptosis kit. Moreover, effects of nectin-4 expression on cell apoptotic-related proteins expressions were detected using western blotting analysis. Results: Nectin-4 was significantly overexpressed in cells from SLE group compared with healthy control (HC) group (P<0.05). When T cells were transfected with sinectin-4, nectin-4 slicing increased cell apoptosis in HC group but significantly decreased apoptosis in SLE group (P<0.05). Nectin-4 slicing significantly decreased CD40L and CD17 expressions in SLE (P<0.05), but performed no effect on CD11a expression. Moreover, nectin-4 down-regulation could significantly decrease Bcl-2, Bcl-XL, and caspase-6 expressions but increase Bax level in SLE group. Conclusion: The data presented in this study suggested that nectin-4 may be a therapeutic target for SLE through affecting the cell apoptosis.     

Exacerbation of apoptosis of cortical neurons following traumatic brain injury in par-4 transgenic mice

Traumatic brain injury (TBI) is a significant clinical problem, yet few effective strategies for treating it have emerged. People that sustain and survive a TBI are left with significant cognitive, behavioral, and communicative disabilities. Apoptotic neuronal death occurs following TBI. Prostate apoptosis response-4 (Par-4) is a death domain-containing protein initially characterized as a critical regulator of apoptosis in prostate cancer cells. We have recently generated and characterized Par-4 transgenic mice in which the expression of the par-4 transgene was limited to cells of neuronal lineage. We now provide evidence that, in cortical neurons from these mice, Par-4 drastically increases apoptotic neuronal death in both in vitro and in vivo models of TBI. In vitro experiments were performed in 7-day-old primary cultures of cortical neurons using a previously published, scratch-induced mechanical trauma model. Neurons that overexpress Par-4 showed not only a significant decrease in overall neuron survival after TBI compared to wild-type cells, but also exhibited a sharper decrease in mitochondrial transmembrane potential, a higher degree of free radical accumulation, and earlier activation of caspase-3 than wild-type cells did. In vivo experiments were performed utilizing a weight drop TBI model. A significantly increased volume of cortical injury and exacerbated activation of caspase-3 were observed in Par-4 transgenic mice when compared to those in wild-type mice. These data suggests that aberrant Par-4 expression exacerbates neuronal cell death following TBI by altering mitochondrial function, enhancing oxidative damage, and execution of apoptosis via caspase activation.     

过表达miR-138抑制TCF-4对甲状腺癌细胞CAL-62增殖，凋亡和运动的调节作用

目的　探究过表达miR-138抑制T细胞因子4（TCF-4）对甲状腺癌细胞CAL-62增殖,凋亡和运动的调节作用机制。 方法　转染miR-138 mimic于CAL-62细胞,使用RT-PCR检测miR-138的表达;使用TCF-4构建pcDNA过表达载体转染CAL-62细胞,使用Westen blot检测TCF-4、增殖核抗原67（Ki67）、增殖细胞核抗原（PCNA）、半胱天冬酶3（caspase-3）、半胱天冬酶9（caspase-9）、上皮型钙黏蛋白（E-cadherin）、波形蛋白（Vimentin）和靶基因cycD、c-Myc的表达;使用EDU染色检测细胞增殖;使用Hoechst染色检测细胞凋亡情况;transwell检测细胞侵袭能力。 结果　相比空白组,miR-138转染+TCF-4组caspase-3、caspase-9、Vimentin蛋白表达显著减少,且有显著性差异（P<0.01）;Ki67、PCNA、E-cadherin、cycD、c-Myc蛋白表达显著增加（P<0.01）;甲状腺肿瘤细胞凋亡受到显著抑制（P<0.01）,增殖、侵染受到显著的促进（P<0.01）。相比TCF-4组,miR-138转染+TCF-4组caspase-3、caspase-9、Vimentin蛋白表达显著增加、（P<0.01）;Ki67、PCNA、E-cadherin、cycD、c-Myc蛋白表达显著减少（P<0.01）,抑制了甲状腺肿瘤细胞增殖、侵染（P＜0.01）,凋亡受到了显著的促进（P<0.01）。 结论　表达miR-138抑制TCF-4使得甲状腺癌细胞CAL-62的增殖、侵染受到显著抑制,凋亡受到显著促进。     

过表达miR-138抑制TCF-4对甲状腺癌细胞CAL-62增殖，凋亡和运动的调节作用

目的　探究过表达miR-138抑制T细胞因子4（TCF-4）对甲状腺癌细胞CAL-62增殖,凋亡和运动的调节作用机制。 方法　转染miR-138 mimic于CAL-62细胞,使用RT-PCR检测miR-138的表达;使用TCF-4构建pcDNA过表达载体转染CAL-62细胞,使用Westen blot检测TCF-4、增殖核抗原67（Ki67）、增殖细胞核抗原（PCNA）、半胱天冬酶3（caspase-3）、半胱天冬酶9（caspase-9）、上皮型钙黏蛋白（E-cadherin）、波形蛋白（Vimentin）和靶基因cycD、c-Myc的表达;使用EDU染色检测细胞增殖;使用Hoechst染色检测细胞凋亡情况;transwell检测细胞侵袭能力。 结果　相比空白组,miR-138转染+TCF-4组caspase-3、caspase-9、Vimentin蛋白表达显著减少,且有显著性差异（P<0.01）;Ki67、PCNA、E-cadherin、cycD、c-Myc蛋白表达显著增加（P<0.01）;甲状腺肿瘤细胞凋亡受到显著抑制（P<0.01）,增殖、侵染受到显著的促进（P<0.01）。相比TCF-4组,miR-138转染+TCF-4组caspase-3、caspase-9、Vimentin蛋白表达显著增加、（P<0.01）;Ki67、PCNA、E-cadherin、cycD、c-Myc蛋白表达显著减少（P<0.01）,抑制了甲状腺肿瘤细胞增殖、侵染（P＜0.01）,凋亡受到了显著的促进（P<0.01）。 结论　表达miR-138抑制TCF-4使得甲状腺癌细胞CAL-62的增殖、侵染受到显著抑制,凋亡受到显著促进。     

Role of TM4SF1 in regulating breast cancer cell migration and apoptosis through PI3K/AKT/mTOR pathway

Purpose: The purpose of this study was to investigate the effect of transmembrane-4-l-six-family-1 (TM4SF1) on breast cancer cell line MDA-MB-231 invasion and apoptosis and its mechanism through PI3K/AKT/mTOR pathway. Methods: siRNA-TM4SF1 and pcDNA-TM4SF1 plasma were constructed and then transfected into MDA-MB-231 cells respectively. Real time (RT)-PCR was used to measure the mRNA expression of TM4SF1 in each group. Also, matrigel method and Annexin V-FITC were used to detect the effect of TM4SF1 expression on MDA-MB-231 cell migration and apoptosis respectively. Besides, western blotting analyze was used to assay the effects of TM4SF1 expression on PI3K/AKT/mTOR pathway associated proteins expressions. Results: The results showed that after being transfected with siRNA-TM4SF1, TM4SF1 expression was significantly declined, while it was significantly increased after cells were transfected with pcDNA-TM4SF1 (P<0.05). Compared with the controls, TM4SF1 overexpression significantly contributed MDA-MB-231 cell migration but decreased apoptotic cells (P<0.05), which were opposite to the results when TM4SF1 was sliced in cells. Moreover, TM4SF1 slicing significantly decreased the expressions of phosphorylated (p)-AKT, p-mTOR, and p-P70 (P<0.05). Conclusion: Our study suggested that TM4SF1 may be a therapeutic target for breast cancer treatment and may loan insight into the mechanisms behind the development and metastasis of advanced breast cancer.     

ATF4重组慢病毒抑制C2C12细胞增殖并促凋亡

目的 构建人活性转录因子4(ATF4)慢病毒,探讨C2C12细胞成骨分化过程中ATF4基因慢病毒修饰对其增殖和凋亡的影响.方法 构建ATF4重组慢病毒载体质粒,然后与2个包装质粒共转入293T细胞中包装成ATF4慢病毒(LV-ATF4);用病毒感染C2C12细胞,并用流式细胞仪(FCM)检测BMP2诱导C2C12细胞分化时对其增殖凋亡的影响,免疫印迹法检测凋亡相关蛋白的表达,电子显微镜观察凋亡细胞的形态学结构变化.结果 成功构建和包装了ATF4重组慢病毒.FCM检测结果表明,BMP2+ LV-ATF4处理组S期细胞(14.89％)低于BMP2+ LV-GFP组(30.64％) (P ＜0.05);BMP2+ LV-ATF4处理组细胞凋亡率(31.06％)高于BMP2+ LV-GFP组(11.39％)(P＜0.05);凋亡相关蛋白的表达与FCM结果一致.结论 在BMP2诱导C2C12细胞成骨分化时,LV-ATT4慢病毒可促进C2C12细胞的凋亡,抑制其增殖.     

Methotrexate induces poly(ADP-ribose) polymerase-dependent, caspase 3-independent apoptosis in subsets of proliferating CD4+ T cells

The mechanism of action of methotrexate (MTX) in autoimmune diseases (AID) is unclear. A pro-apoptotic effect has been demonstrated in mitogen-stimulated peripheral blood mononuclear cells (PBMC), but studies employing conventional antigens have disputed a pro-apoptotic effect. CD4+ T helper (Th) cells play a significant role in most AID. We therefore examined directly, by flow cytometry, the uptake of MTX by the T helper (Th) cells stimulated for 6 days with Candida albicans (CA) or tetanus toxoid (TT), and its consequences with respect to induction of apoptosis. While none of the resting Th cells took up MTX, nearly all the dividing Th cells did, and this abrogated further cell division. Among dividing Th cells, MTX induced an approximately sixfold increase over baseline levels in the proportion of apoptotic cells. This proportion could be reverted to baseline by the addition of folic acid. Exposure of CA-stimulated PBMC to MTX significantly increased their level of cleaved poly(ADP-ribose) polymerase (PARP), and a similar tendency was observed in TT-stimulated cells. Unlike CA and TT, the mitogen phytohaemagglutinin (PHA) induced proliferation of both CD4- and CD4+ T cells, and induced apoptosis in both undivided and divided Th cells. PHA-induced apoptosis involved activation of caspase-3 and the anti-apoptotic protein Bcl-2 in addition to PARP cleavage, suggesting that PHA induces apoptosis via different pathways than CA and TT. We suggest that the latter are more representative of stimulation with self-antigens in AID, and that a pro-apoptotic effect of MTX on self-antigen-stimulated Th cells contributes to the effect of MTX in the treatment of AID.