Stevioside enhances apoptosis induced by serum deprivation in PC12 cells

In the laboratory, using a PC12 cell system, studies have been conducted on the effects of various chemicals on apoptosis, as it is considered to be an essential part of normal development, maintenance, and defense in organisms. Stevioside is a natural sweetener extracted from the leaves of Stevia rebaudiana. Since it is widely used as a sugar replacement, it was decided to evaluate the toxicological effects of low concentrations of stevioside on apoptosis induced by serum deprivation using the PC12 cell system. It was found that based on data from DNA electrophoresis and TUNEL signal assays stevioside enhanced apoptosis induced by serum deprivation. This enhancement was caused by increased expression of Bax and of cytochrome c released into the cytosol. These findings suggest that stevioside affects the regulation of the normal apoptotic condition. Further investigation will be needed to clarify the detailed mechanism of the enhancement due to the treatment with stevioside.     

Stevioside enhances apoptosis induced by serum deprivation in PC12 cells

In the laboratory, using a PC12 cell system, studies have been conducted on the effects of various chemicals on apoptosis, as it is considered to be an essential part of normal development, maintenance, and defense in organisms. Stevioside is a natural sweetener extracted from the leaves of Stevia rebaudiana. Since it is widely used as a sugar replacement, it was decided to evaluate the toxicological effects of low concentrations of stevioside on apoptosis induced by serum deprivation using the PC12 cell system. It was found that based on data from DNA electrophoresis and TUNEL signal assays stevioside enhanced apoptosis induced by serum deprivation. This enhancement was caused by increased expression of Bax and of cytochrome c released into the cytosol. These findings suggest that stevioside affects the regulation of the normal apoptotic condition. Further investigation will be needed to clarify the detailed mechanism of the enhancement due to the treatment with stevioside.     

Diethyl phthalate enhances apoptosis induced by serum deprivation in PC12 cells

In this study, to examine the mechanism of diethyl phthalate toxicity to cells, the effects of diethyl phthalate on apoptosis in a PC12 cell system were investigated by assaying apoptotic factors such as caspase-3, Bax, cytochrome c and DNA damage. Diethyl phthalate was shown to enhance the apoptosis induced by serum deprivation according to the results of DNA electrophoresis and TUNEL signal assays, although it could not induce apoptosis itself in the cells. This enhancement was thought to be because of an increase in caspase-3-like activity. In addition, the expression of bax and contents of cytochrome c in the cytosol showed a tendency to increase the cells exposed to diethyl phthalate. These results indicated that diethyl phthalate, a potential endocrine disrupter, affects the apoptotic system in PC12 cells. Diethyl phthalate may enhance oxidative stress such as that induced by reactive oxygen species in PC12 cells.     

Minimal systems analysis of mitochondria-dependent apoptosis induced by cisplatin

Recently, it was reported that the role of mitochondria-reactive oxygen species (ROS) generating pathway in cisplatin-induced apoptosis is remarkable. Since a variety of molecules are involved in the pathway, a comprehensive approach to delineate the biological interactions of the molecules is required. However, quantitative modeling of the mitochondria-ROS generating pathway based on experiment and systemic analysis using the model have not been attempted so far. Thus, we conducted experiments to measure the concentration changes of critical molecules associated with mitochondrial apoptosis in both human mesothelioma H2052 and their ρ⁰ cells lacking mitochondrial DNA (mtDNA). Based on the experiments, a novel mathematical model that can represent the essential dynamics of the mitochondrial apoptotic pathway induced by cisplatin was developed. The kinetic parameter values of the mathematical model were estimated from the experimental data. Then, we have investigated the dynamical properties of this model and predicted the apoptosis levels for various concentrations of cisplatin beyond the range of experiments. From parametric perturbation analysis, we further found that apoptosis will reach its saturation level beyond a certain critical cisplatin concentration.     

Efflux of potassium ion is an important reason of HL-60 cells apoptosis induced by tachyplesin

AIM: To investigate the role of intercellular potassium in tachyplesin-induced HL-60 cells apoptosis. METHODS: The concentration of intercellular potassium, cell volume and mitochondrial membrane potential were examined by flow cytometry. RESULTS: The concentration of intercellular potassium reduced in a time-dependent manner in tachyplesin-treated HL-60 cells. In addition, the loss of mitochondrial membrane potential was tightly coupled with the shrinkage of cell volume. Different caspase inhibitors protected against DNA degradation but did not prevent the loss of HL-60 cell viability induced by tachyplesin. Ba2+, which was a kind of blocker of volume-regulatory K+ channels, increased the viability of tachyplesin-treated HL-60 cells and maintained mitochondrial membrane potential and cell volume. CONCLUSION: Efflux of K+ was an important reason for apoptosis in tachyplesin-treated HL-60 cells. Efflux of K+ affected the viability of tachyplesin-treated HL-60 cells independent of the process of caspase activation.     

Inhibition of apoptosis facilitates necrosis induced by cisplatin in gastric cancer cells

Although cisplatin has been shown to induce both apoptosis and necrosis in cancer cells, the potential interconnections between these modes of cell death induced by the drug remain unknown. We studied this phenomenon in gastric cancer cell lines and identified one cell line (SGC-7901) that underwent apoptosis, and another cell line (BGC-823) that primarily underwent nonapoptotic cell death, in response to cisplatin. Apoptosis in cisplatin-treated SGC-7901 cells seemed to be caspase dependent and was mediated, at least in part, by the BH3-only protein, Noxa. This was evidenced by the rapid upregulation of Noxa and inhibition of apoptosis by small interfering RNA knockdown of Noxa. Nonapoptotic cell death induced by cisplatin in BGC-823 cells was characterized by lack of DNA fragmentation, delayed externalization of phosphatidylserine, caspase independence, plasma membrane disruption, and intracellular vacuole formation, indicative of necrosis. Surprisingly, blockage of apoptosis induction by a general caspase inhibitor or by Noxa small interfering RNA in SGC-7901 failed to protect against cisplatin-induced cell death. Under such conditions, SGC-7901 cells displayed cellular features associated with necrosis. Cisplatin-induced apoptosis, thus, seems to precede necrosis when the apoptotic machinery is operative. When the apoptosis program is defective, necrotic cell death takes place as an alternative pathway leading to cell demise. Induction of different modes of cell death that are interrelated in the same cells by cisplatin has the potential to be exploited in formulating new adjuvant cancer therapies.     

hTERT反义寡核苷酸增强DDP介导细胞凋亡作用

目的研究端粒酶催化亚基(hTERT)反义寡核苷酸(ASODN)对HeLa细胞端粒酶活性的抑制及其对顺铂(DDP)诱导细胞凋亡的影响。方法用逆转录聚合酶链反应技术(RT-PCR)定量端粒酶重复扩增法(TRAP)检测细胞的端粒酶活性。观察细胞凋亡的形态学变化,流式细胞仪对细胞凋亡进行定量分析。结果ASODN作用后,细胞的端粒酶活性明显降低,且这种作用具有明显的时间和剂量依赖性。0.05、0.1μmol.L-1和0.2μmol.L-1的硫代ASODN治疗后,HeLa细胞的端粒酶活性分别下降了21.8%、52.4%和71.1%。0.2μmol.L-1的硫代ASODN作用HeLa24、48h和72h后,细胞的端粒酶活性分别下降了12.48%、38.27%和71.10%。细胞转染0.2μmol.L-1浓度的ASODN24h后再与1.5、3.0mg.L-1浓度的顺铂联合作用,吖啶橙染色可见典型的凋亡形态,并且凋亡百分率(50.35%、29.67%)分别与RSODN联合顺铂组(19.33%、12.13%)、单用顺铂组(19.67%、11.38%)比较,差异有显著性(P<0.05)。结论hTERT反义寡核苷酸能有效抑制其端粒酶活性,并且促进DDP诱导HeLa细胞凋亡。     

大黄素对抗顺铂引起的WI-38细胞凋亡

目的　从细胞水平研究大黄素对顺铂引起WI 38细胞凋亡的影响。方法　采用MTT法检测细胞毒性 ,用形态学观察、DNA凝胶电泳及流式细胞仪检测细胞凋亡。结果　WI 38细胞经大黄素和顺铂同时处理 2 2h后 ,30mg·L- 1大黄素可明显减轻顺铂引起的细胞毒性 ,其IC50 值由 (16± 3)mg·L- 1增加至 (34± 6 )mg·L- 1;可明显抑制顺铂引起的细胞形态学改变、核异染色质边集和DNA片段化 ,使顺铂 10和 30mg·L- 1导致的细胞凋亡率由 35 .5 6 %和33.99%降至 9.2 1%和 10 .2 5 % ,S期细胞百分数由6 2 .6 6 %和 4 8.4 6 %降至 4 8.6 7%和 36 .18%。结论　大黄素可对抗顺铂所致WI 38细胞凋亡 ,可能与其对细胞周期的影响有关     

梓醇对顺铂诱导大鼠卵巢颗粒细胞损伤的保护作用

目的探讨梓醇对顺铂诱导大鼠卵巢颗粒细胞损伤的保护作用及作用机制。方法收集大鼠卵巢颗粒细胞,经原代培养后设为对照组、模型组(顺铂50 nmol/L)和梓醇50、100、200μmol/L组,各组细胞按相应方式进行处理。采用MTT法测定细胞活力;流式细胞术Annexin V/PI双染法考察细胞凋亡情况;免疫组化法观察细胞增殖能力,WB法定量细胞凋亡蛋白的分子水平。结果顺铂可显著抑制卵巢颗粒细胞活力、增殖,诱导细胞凋亡;在给予梓醇进行干预后,大鼠卵巢颗粒细胞活力增强、增殖上调,细胞凋亡减弱;经顺铂进行刺激损伤,卵巢颗粒细胞中caspase-3、caspase-9的活化蛋白表达量显著上升,而不同浓度的梓醇可显著下调细胞中cleaved caspase-3、cleaved caspase-9的蛋白表达量。结论梓醇可以改善大鼠细胞活力、提高细胞增殖和抗凋亡能力,对顺铂诱导的卵巢颗粒细胞损伤产生明显的保护作用。     

乙酰葛根素对氧糖剥离诱导海马神经元凋亡的影响

目的研究新型异黄酮类化合物乙酰葛根素对氧糖剥离神经元的保护作用及其机制。方法采用氧糖剥离建立细胞模型。DAPI法观察神经元凋亡率,RT-PCR法检测NF-κBp65、HIF-1α及p53mRNA的表达,Western blot法测定Hsp70蛋白的含量及乙酰葛根素对其的影响。结果与模型组相比较,乙酰葛根素可以减少凋亡细胞数;降低NF-κBp65、HIF-1α及p53mRNA表达;乙酰葛根素(低,中和高浓度组)增加Hsp70蛋白的表达。结论乙酰葛根素降低NF-κBp65、HIF-1α、p53的表达和升高Hsp70蛋白表达。     

Cytotoxicity, apoptosis, and in vitro DNA damage induced by potassium chromate

Cr(6+) is a known human cytotoxic and carcinogenic agent that requires intracellular reduction for activation. We have analyzed the cytotoxic and DNA binding properties of K(2)CrO(4) (Cr(6+)) in comparison with those of Cl(3)Cr (Cr(3+)). The results indicate that K(2)CrO(4) exhibits higher cytotoxicity than Cl(3)Cr in several human and murine cell lines. The cytotoxic activity of K(2)CrO(4) is also indicated by the fact that is able to produce cell killing through apoptosis in cisplatin-resistant cells transformed by H-ras oncogene. Moreover, in vitro DNA binding experiments show that, in the presence of ascorbate (the major intracellular reductant of Cr(6+)), K(2)CrO(4) induces both interstrand cross-links and strand breaks. Because the chromate anion is by itself unreactive toward DNA, these data suggest that the cytotoxicity of K(2)CrO(4) may be associated with the DNA binding of reactive intermediate chromium species resulting from reduction of Cr(6+).     

Effect of manoalide on apoptosis induced by deprivation of growth factors in vascular endothelial cells

研究ｍａｎｏａｌｉｄｅ对除去生长因子（ａＦＧＦ和血清）而诱导的血管内皮细胞凋亡的影响．方法：通过细胞形态观察,ＤＮＡ凝胶电泳及荧光显微术等方法确定ｍａｎｏａｌｉｄｅ对细胞凋亡的抑制或促进作用．结果：向去除ａＦＧＦ和血清的培养液中加低浓度的ｍａｎｏａｌｉｄｅ（１－４μｍｏｌ·Ｌ－１）,培养细胞４８ｈ,细胞的脱壁和ＤＮＡ片断化受到抑制;ｍａｎｏａｌｉｄｅ浓度为７μｍｏｌ·Ｌ－１时,促进细胞脱壁和ＤＮＡ片断化．结论：低浓度的ｍａｎｏａｌｉｄｅ（２μｍｏｌ·Ｌ－１）抑制血管内皮细胞凋亡,而较高浓度的ｍａｎｏａｌｉｄｅ（７μｍｏｌ·Ｌ－１）促进该细胞凋亡．     

用非细胞体系研究HeLa细胞细胞质在顺铂诱导凋亡中的作用

在构建Bcl-2高表达及其对照细胞株基础上, 用非细胞体系研究了顺铂诱导的HeLa细胞凋亡中细胞质的作用. 实验结果显示凋亡细胞胞质提取物可引起正常细胞核的固缩及染色质边缘化, 类似完整细胞凋亡的改变, 而Bcl-2高表达株的胞质提取物具有抗凋亡能力. 说明引起凋亡的物质及对抗凋亡的物质在细胞质中都存在, 细胞质在顺铂诱导的HeLa细胞凋亡中起着较重要的作用.     

Basic fibroblast growth factor sensitizes NIH 3T3 cells to apoptosis induced by cisplatin

One mechanism by which chemotherapeutic agents kill tumor cells is by induction of apoptosis. Basic fibroblast growth factor (bFGF/FGF-2) has been reported to inhibit apoptosis in NIH 3T3 cells treated with chemotherapy drugs. We have investigated how bFGF modulates apoptosis induced by cisplatin in NIH 3T3 cells. Treatment with 10 microgram/ml cisplatin for 12 h induced apoptosis in 2 to 13% of the cells at 24 h post-treatment. Preincubation with 10 ng/ml bFGF for 24 h led to cisplatin-induced apoptosis in 20% to 50% of the cells. Preincubation with lower concentrations of bFGF (0.1-1 ng/ml) or simultaneous addition of bFGF and cisplatin had no effect on the amount of apoptosis. Pretreatment with bFGF also significantly decreased the dose-dependent survival of NIH 3T3 cells exposed to cisplatin, as determined by colony formation. Cells treated with 10 ng/ml bFGF showed a distinct morphology, appearing smaller and more refractile, before cisplatin exposure. The enhancement of cisplatin-induced apoptosis and the morphology shift demonstrated the same dose response to bFGF, and both effects were reversible if bFGF was removed from the medium for 24 h before cisplatin treatment. Mitogenic response to bFGF by NIH 3T3 cells saturated at 0.5 ng/ml, as measured by (3)H-thymidine uptake, and this response was blocked by coaddition of suramin, an inhibitor of FGF ligand-receptor interactions. Suramin did not reverse the enhancement of cisplatin-induced apoptosis by bFGF. Therefore, bFGF sensitized NIH 3T3 cells to cisplatin, and this effect might be mediated through a pathway separate from that used for mitogenic signaling.     

Function of HeLa cell cytoplasm in apoptosis induced by cisplatin in a cell-free system

在构建Ｂｃｌ－２高表达及其对照细胞株基础上，用非细胞体系研究了顺铂诱导的ＨｅＬａ细胞凋亡中细胞质的作用．实验结果显示凋亡细胞胞质提取物可引起正常细胞核的固缩及染色质边缘化，类似完整细胞凋亡的改变，而Ｂｃｌ－２高表达株的胞质提取物具有抗凋亡能力．说明引起凋亡的物质及对抗凋亡的物质在细胞质中都存在，细胞质在顺铂诱导的ＨｅＬａ细胞凋亡中起着较重要的作用．     

An ent-kaurene diterpene enhances apoptosis induced by tumor necrosis factor in human leukemia cells

Some antitumor agents, including tumor necrosis factor-alpha (TNF-alpha) and camptothecin (CPT), often cause resistance of tumor cells to antitumor agents through activation of the nuclear factor-kappa B (NF-kappa B) pathway that leads to up-regulation of anti-apoptotic proteins. Therefore, co-treatment of an inhibitor of the NF-kappa B pathway with antitumor agents is a useful strategy for chemotherapy. Here we report that ent-11 alpha-hydroxy-16-kauren-15-one (KD) selectively inhibits NF-kappa B-dependent gene expression due to treatment with TNF-alpha. KD in combination with TNF-alpha caused a dramatic increase in apoptosis in human leukemia cells accompanied by activation of caspases. A broad-spectrum inhibitor of caspases decreased the apoptosis induced by treatment with KD and TNF-alpha. KD in combination with CPT also caused an increase in apoptosis. These results suggest that the apoptotic potency of co-treatment of KD with TNF-alpha or CPT is elicited through selective inhibition of NF-kappa B-dependent anti-apoptotic proteins and thus may provide a basis for the development of useful approaches to the treatment of leukemia.     

Mecamylamine prevents neuronal apoptosis induced by glutamate and low potassium via differential anticholinergic-independent mechanisms

Neuronal loss via apoptosis caused by various stimuli may be the fundamental mechanism underlying chronic and acute neurodegenerative diseases. A drug inhibiting neuronal apoptosis may lead to a practical treatment for these diseases. In this study, treatment with mecamylamine, a classical antagonist of nicotinic acetylcholine receptors (nAChRs), prevented neuronal apoptosis induced by 75 microM glutamate and by low potassium (LK) in cerebellar granule neurons (CGNs) with EC(50)s of 35 and 293 microM, respectively. Two other antagonists of nAChRs, dihydro-beta-erythroidine and tubocurarine, failed to inhibit these two kinds of apoptosis. Mecamylamine inhibited the NMDA (30 microM)-evoked current and competed with [(3)H]MK-801. Furthermore, two inhibiters of the c-Jun N-terminal kinase (JNK) pathway prevented LK-induced apoptosis. Mecamylamine reversed the phosphorylation levels of JNK and c-Jun as well as the expression of c-Jun caused by LK in a Western blot assay. In addition, the JNK/c-Jun pathway was not involved in glutamate-induced cell death of CGNs. Our results suggest that mecamylamine prevents glutamate-induced apoptosis by blocking NMDA receptors at the MK-801 site and LK-induced apoptosis by inhibiting the activation of the JNK/c-Jun pathway.     

Effect of manoalide on apoptosis induced by deprivation of growth factors in vascular endothelial cells.

AIM: To study effect of manoalide on apoptosis induced by deprivation of acidic fibroblast growth factor (aFGF) and serum in vascular endothelial cells (VEC). METHODS: Morphologic changes were observed by light microscopy. Viability was determined by counting the cells that attached to dishes after treatments. DNA fragmentation was analyzed by agarose gel electrophoresis and fluorescence microscopy. RESULTS: The cells deprived of aFGF and serum were exposed to manoalide 1-4 mumol. L-1 for 48 h, detachment and DNA fragmentation of these cells were suppressed. At 7 mumol. L-1, manoalide promoted detachment and DNA fragmentation of VEC. CONCLUSION: manoalide 2 mumol.L-1 inhibited, but 7 mumol.L-1 promoted, apoptosis of VEC.     

Antiapoptotic effect of ras in the apoptosis induced by serum deprivation and exposure to actinomycin D

Serum deprivation or exposure of NIH 3T3 cells to actinomycin D (0.25–1.0 mg/ml ; 1 h) was associated with the accumulation of numerous apoptotic cells, as identified by their condensed nuclei and the decrease in cell size. In contrasts, v-H-ras-transformed NIH 3T3 cells were found to be resistant to this apoptosis induction. When v-H-ras-transformed cells were first pretreated for 24 h with 50 μM mevastatin, an agent which is known to be capable to deactivate the ras funcion, cell viability decreased and apoptotic cells became abundant (~60–80%) 72 h after serum deprivation or exposure to actinomycin D. During the serum deprivation of NIH 3T3 cells, appearance of the apoptotic cells was preceded by G₁ phase arrest. Accumulation of cells in the G₁ phase was also observed in v-H-ras-transformed cells 24 h after serum deprivation. At later times (48–72 h), v-H-ras-transformed cells seemed to be capable of breaking through the G₁ arrest and were then found to be distributed normally in the cell cycle. Received: 6 May 1996 / Accepted: 2 September 1996     

1-甲基-3-异丁基黄嘌呤延迟去除生长因子诱导的血管内皮细胞凋亡(英文)

目的:研究1-甲基-3-异丁基黄嘌呤(MIBX)对去除生长因子(aFGF和血清)诱导的血管内皮细胞凋亡的影响.方法:通过细胞存活率的分析,荧光显微技术和DNA凝胶电泳等方法,检测MIBX对细胞凋亡的影响.结果:用25-200μmol/L的MIBX处理培养在无aFGF和血清的培养液中的血管内皮细胞,50-200μmol/L的MIBX在处理6h明显抑制了凋亡小体的形成和DNA的片断化.但是同样浓度的MIBX处理细胞12h以后,处理组和对照组之间无明显差别.结论:MIBX延迟去除aFGF和血清诱导的血管内皮细胞凋亡.