Ara-C induces apoptosis in monkey fibroblast cells.

The effect of cytosine arabinoside (Ara-C) on cell viability has been studied in African green monkey kidney fibroblasts (CV1-P). It has been shown previously that Ara-C- induced cell death in neurons is mediated by apoptosis. We investigated whether Ara-C can induce apoptosis also in CV1-P cells, and if the apoptosis is p53-associated. For comparison, human neuroblastoma cells (SH-SY5Y) were studied as a model of human neuronal cells. SYTO13/propidium iodide staining revealed condensed and fragmented nuclei in both cell lines. Ara-C treatment for 48 h induced approximately 24% apoptosis in CV1-P cells whereas approximately 55% of SH-SY5Y cells were apoptotic. Ara-C increased the level of p53 in both CV1-P and SH-SY5Y cells compared to control. The maximum level of p53 in SH-SY5Y cells was reached at 12 h and this then rapidly faded whereas CV1-P cells p53 levels remained elevated after reaching their maximum. Caspase-3 activity was 5-fold higher in human neuroblastoma cells than in monkey fibroblasts, this reflected the decreased cell viability. Our results prove that Ara-C- induced apoptosis in CV1-P cells is associated with an increase of p53 and activation of caspase-3. Ara-C-induced toxicity in CV1-P cells is modest compared to that seen in neuronal cells.     

Novel squamosamide derivative （compound FLZ） attenuates Aβ25-35^- induced toxicity in SH-SY5Y cells

AIM: The aim of the present study was to investigate the protective effect of compound N-[2-(4-hydroxy-phenyl)-ethyl]-2-(2,5-dimethoxy-phenyl)-3-(3-methoxy-4-hydroxy-phenyl)-acrylamide (compound FLZ), a novel synthetic analogue of nature squamosamide, on Abeta25-35-induced toxicity and its active mechanism in human neuroblastoma SH-SY5Y cells. METHODS: SH-SY5Y cells were pre-incubated with various concentrations of compound FLZ for 30 min and then cultivated with Abeta25-35 (25 micromol/L) for 48 h to induce neurotoxicity. Cell viability, lactate dehydrogenase (LDH) release, and the glutathione (GSH) level were determined by a biochemical analysis. The cell apoptotic ratio and intracellular reactive oxygen species (ROS) level were measured by a flow cytometry analysis. The expression of apoptosis protein (Bcl-2 and Bax) and cytochrome c release were assayed by the Western blot method. RESULTS: The pretreatment of SH-SY5Y cells with FLZ (1 and 10 micromol/L) markedly increased cell viability and decreased LDH release and morphological injury. Also, FLZ attenuated the Abeta25-35-induced apoptotic cell ratio, regulated the apoptosis protein (Bcl-2 and Bax) expression, and decreased the cytochrome c release from mitochondria. FLZ also significantly inhibited the generation of ROS and the depletion of GSH induced by Abeta25-35 in SH-SY5Y cells. CONCLUSION: FLZ has protective action against Abeta25-35-induced toxicity in SH-SY5Y cells, which might be mediated through its antioxidant property.     

Diclofenac, a non-steroidal anti-inflammatory drug, suppresses apoptosis induced by endoplasmic reticulum stresses by inhibiting caspase signaling

Non-steroidal anti-inflammatory drugs (NSAIDs) are frequently used in the treatment of inflammation and pain. In many reports, NSAIDs have induced apoptosis in a variety of cell lines such as colon cancer cells. On the other hand, more recently a few reports have found that NSAIDs protect against apoptosis. Here we investigate endoplasmic reticulum (ER)-stress-induced apoptosis of neuronal cells. The aim of this study is to examine the involvement of NSAIDs, in particular diclofenac, on ER-stress-induced apoptosis of human neuroblastoma SH-SY5Y cells. Diclofenac significantly suppressed SH-SY5Y cell death induced by two types of ER-stress-inducing agents: thapsigargin, an inhibitor of Ca2+-ATPase on the endoplasmic reticulum membrane, and tunicamycin, a glycosylation blocker. Other NSAIDs, such as indomethacin, ibuprofen, aspirin, and ketoprofen, also suppressed ER-stress-induced SH-SY5Y cell death. The dose-dependent anti-apoptotic effect of diclofenac did not correlate with the reduction of prostaglandin release. Administration of prostaglandin E2, which was a primary product of arachidonic metabolism, showed no effects against anti-apoptotic effects produced by diclofenac. Thapsigargin and tunicamycin each significantly activated caspase-3, -9, and -2 in the intrinsic apoptotic pathway in SH-SY5Y cells. Diclofenac suppressed the activation of caspases induced by both ER stresses. Thapsigargin and tunicamycin decreased the mitochondrial membrane potential in SH-SY5Y cells. Diclofenac suppressed the mitochondrial depolarization induced by both ER stresses. Diclofenac inhibited ER-stress-induced apoptosis of SH-SY5Y cells by suppressing the activation of caspases in the intrinsic apoptotic pathway. This is the first report to find that diclofenac has protective effects against ER-stress-induced apoptosis.     

松果菊苷对TNFα诱导的SH-SY5Y细胞凋亡的保护作用

目的　探讨肉苁蓉提取物松果菊苷对TNFα诱导的SH SY5Y细胞凋亡的保护作用。方法　用MTT法检测细胞存活率,DNA琼脂糖凝胶电泳和流式细胞仪检测细胞凋亡的发生,以激光共聚焦显微镜荧光染色法检测细胞内活性氧的产生和线粒体膜电位的变化,并用荧光酶标仪测定caspase 3的活性。结果　100μg·L-1 TNFα处理细胞 36h显著降低细胞的存活率;诱导细胞发生凋亡,凋亡率达37%;细胞内活性氧水平及caspase 3的活性升高;而线粒体膜电位却明显降低,红 /绿荧光强度的比值由正常的 5 97降低为 0 35左右。而预先给予 1, 10或者 100mg·L-1浓度的松果菊苷处理细胞 2h,可提高细胞存活率;并可有效抑制DNAladder的发生;流式细胞仪检测凋亡率分别降低到25 9%, 18 3% 和 8 2%;激光共聚焦显微镜结果显示松果菊苷可明显抑制细胞内活性氧产生;并可逐渐恢复线粒体的高能量状态;caspase 3的活性不断降低,并呈现了一定的剂量依赖性。结论　松果菊苷能抑制TNFα诱导的SH SY5Y细胞凋亡,其神经细胞保护作用可能与降低细胞内活性氧水平,抑制caspase 3的活性和维持线粒体膜电位的高能状态有关。     

Involvement of Wnt/beta-catenin signaling in tripchlorolide protecting against oligomeric beta-amyloid-(1-42)-induced neuronal apoptosis

This study is to explore whether the Wnt/beta-catenin signaling pathway is involved in the process of tripchlorolide (T4) protecting against oligomeric Abeta(1-42)-induced neuronal apoptosis. Primary cultured cortical neurons were used for the experiments on day 6 or 7. The oligomeric Abeta(1-42) (5 micromol x L(-1) for 24 h) was applied to induce neuronal apoptosis. Prior to treatment with Abeta(1-42) for 24 h, the cultured neurons were pre-incubated with T4 (2.5, 10, and 40 nmol x L(-1)), Wnt3a (Wnt signaling agonists) and Dkk1 (inhibitors) for indicated time. Then the cell viability, neuronal apoptosis, and protein levels of Wnt, glycogen synthase kinase 3beta (GSK3beta), beta-catenin and phospho-beta-catenin were measured by MTT assay, TUNEL staining and Western blotting, respectively. The result demonstrated that oligomeric Abeta(1-42) induced apoptotic neuronal cell death in a time- and dose-dependent manner. Pretreatment with T4 significantly increased the neuronal cell survival and attenuated neuronal apoptosis. Moreover, oligomeric Abeta(1-42)-induced phosphorylation of beta-catenin and GSK3beta was markedly inhibited by T4. Additionally, T4 stabilized cytoplasmic beta-catenin. These results indicate that tripchlorolide protects against the neurotoxicity of Abeta by regulating Wnt/beta-catenin signaling pathway. This may provide insight into the clinical application of tripchlorolide to Alzheimer's disease.     

Silica nanoparticles induced intrinsic apoptosis in neuroblastoma SH-SY5Y cells via CytC/Apaf-1 pathway

The present study was to investigate effects of Silica nanoparticles (SiNPs) on nervous system and explore potential mechanisms in human neuroblastoma cells (SH-SY5Y). Cytotoxicity was detected by cell viability and Lactate dehydrogenase (LDH) release. Flow cytometry analysis was applied to assess mitochondrial membrane potential (MMP) loss, intracellular Ca²⁺ and apoptosis. To clarify the mechanism of SiNPs-induced apoptosis, intrinsic apoptosis-related proteins were detected. Our results showed that SiNPs caused cytotoxicity, cell membrane damage and Ca²⁺ increase in a dose-dependent manner in SH-SY5Y cells. Both the mitochondrial membrane potential (MMP) loss and potential mitochondria damage resulted in Cyt C release to the cytoplasm. The elevated Cyt C and Apaf1 further triggered intrinsic apoptosis via executive molecular caspase-9 and caspase-3. The present study confirmed that SiNPs induced intrinsic apoptosis in neuroblastoma SH-SY5Y cells via CytC/Apaf-1 pathway and provided a better understanding of the potential toxicity induced by SiNPs on human neurocyte.     

Potential autophagy enhancers protect against fipronil-induced apoptosis in SH-SY5Y cells

Oxidative stress created by environmental toxicants activates several signaling pathways. Autophagy is one of the first lines of defense against oxidative stress damage. The autophagy pathway can be induced and up-regulated in response to intracellular reactive oxygen species (ROS). Recently, we reported that fipronil (FPN)-induced mitochondria-dependent apoptosis is mediated through ROS in human neuroblastoma SH-SY5Y cells. In this study, we explored the role of autophagy to prevent FPN neurotoxicity. We investigated the modulation of FPN-induced apoptosis according to autophagy regulation. FPN activated caspase-9 and caspase-3, and induced nuclear fragmentation and condensation, all of which indicate that FPN-induced cell death was due to apoptosis. In addition, we observed FPN-induced autophagic cell death by monitoring the expression of LC3-II and Beclin-1. Exposure to FPN in SH-SY5Y cells led to the production of ROS. Treatment with N-acetyl-cysteine (NAC) effectively blocked both apoptosis and autophagy. Interestingly, pretreatment with rapamycin, an autophagy inducer, significantly enhanced the viability of FPN-exposed cells; the enhancement of cell viability was partially due to alleviation of FPN-induced apoptosis via a decrease in levels of cleaved caspase-3. However, pretreatment with 3-methyladenine (3MA) a specific inhibitor for autophagy, remarkably strengthened FPN toxicity and further induced activation of caspase-3 in these cells. Our studies suggest that FPN-induced cytotoxicity is modified by autophagy regulation and that rapamycin is neuroprotective against FPN-induced apoptosis through enhancing autophagy.     

Fumonisin B1-induced apoptosis in neuroblastoma, glioblastoma and hypothalamic cell lines

Fumonisin B(1) (FB(1)) is a mycotoxin produced by Fusarium verticilliodes, which commonly infects corn across the world. Fusarium fungi may also be found in moisture-damaged buildings. In this study, we investigated the role of apoptosis in the toxicity of FB(1) in four different cell lines. Activation of caspase-3-like protease, DNA fragmentation and expression of p53 and Bcl-2 family proteins were studied in mouse GT1-7 hypothalamic, rat C6 glioblastoma, human U-118MG glioblastoma, and human SH-SY5Y neuroblastoma cells exposed to 0.1-100microM FB(1) for 0-144h. Caspase-3-like protease activity increased in all cell lines, except SH-SY5Y, at 48-144h, and internucleosomal DNA fragmentation occurred in all of the cell lines, pointing to a role for apoptosis in the toxicity of FB(1). However, the expressions of p53 or pro- or antiapoptotic Bcl-2 family proteins (Bax, Bcl-2, Bcl-X(L) and Mcl-1) were not affected in any of the cell lines even after prolonged exposure to FB(1) at high doses. The results of this study, together with the results of our previous studies, provide evidence that FB(1) is a potential neurotoxin, but that the toxicity of FB(1) varies between different cell lines. The sensitivity of these cell lines towards FB(1) is as follows: U-118MG>GT1-7>C6>SH-SY5Y cells. These results are consistent with the assumption that cells of glial origin may be more sensitive towards FB(1) than cells of neural origin.     

香椿果抗补体活性多酚对补体损伤神经细胞的保护作用研究

目的评价香椿果抗补体活性多酚XCG-7对补体损伤神经细胞的保护作用。方法采用眼镜蛇毒因子特异激活血清补体,诱导人神经母细胞瘤细胞SH-SY5Y损伤。通过测定乳酸脱氢酶释放量、细胞活力和Caspase-3/7活性等指标,评价XCG-7对补体损伤神经细胞的保护作用。结果 XCG-7对SH-SY5Y细胞的生长具有促进作用,明显抑制补体损伤导致的细胞活力下降,在一定程度上减轻乳酸脱氢酶的释放,抑制细胞凋亡的发生。结论香椿果抗补体活性多酚XCG-7能促进SH-SY5Y细胞的生长,对补体损伤神经细胞具有一定的保护作用。     

Protective effect of tubuloside B on TNFalpha-induced apoptosis in neuronal cells.

AIM: To investigate the neuroprotective effect of tubuloside B, one of the phenylethanoids isolated from the stems of Cistanche salsa, on tumor necrosis factor-alpha (TNFalpha)-induced apoptosis in SH-SY5Y neuronal cells. METHODS: Cell viability was analyzed using MTT assay. Apoptotic cells were detected using Hoechst33342 staining, and confirmed by DNA fragmentation and flow cytometric analysis. The activity of caspase-3 was measured with special assay kit. The concentration of free intracellular calcium was determined with the probe Indo-1 by spectrometer. The level of intracellular reactive oxygen species and the potential of mitochondrial membrane were determined by laser scanning confocal microscopy (LSCM) combined with fluorescence probe H2DCFDA or JC-1 respectively. RESULTS: SH-SY5Y cells treated with TNFalpha 100 microg/L for 36 h showed typical morphological changes of apoptosis. DNA ladder could be observed by agarose gel electrophoresis. The highest percentage of apoptotic cells accumulated to 37.5 %. Following 36 h treatment with TNFalpha, accumulation of intracellular ROS and [Ca2+]i and decrease in mitochondrial membrane potential were observed, and caspase-3 activity increased by about five-fold compared with controls. However, pretreatment with tubuloside B (1, 10, or 100 mg/L) for 2 h attenuated the TNFalpha-mediated apoptosis. The antiapoptotic action of tubuloside B was partially dependent on an anti-oxidative stress effects, maintain of mitochondria function, decrease of concentration of free intracellular calcium and inhibition of caspase-3 activity. CONCLUSION: Tubuloside B has the neuroprotective capacity to antagonize TNFalpha-induced apoptosis in SH-SY5Y cells and may be useful in treating some neurodegenerative diseases.     

Identification, design, synthesis, and pharmacological activity of (4-ethyl-piperazin-1-yl)-phenylmethanone derivatives with neuroprotective properties against beta-amyloid-induced toxicity

In search of novel therapeutic approaches for Alzheimer's disease (AD), we report herein the identification, design, synthesis, and pharmacological activity of (4-ethyl-piperaz-1-yl)-phenylmethanone derivatives with neuroprotective properties against beta-amyloid-induced toxicity. (4-ethyl-piperaz-1-yl)-phenylmethanone is a common substructure shared by molecules isolated from plants of the Asteraceae genus, traditionally used as restorative of lost or declining mental functions. (4-Ethyl-piperaz-1-yl)-phenylmethanone displayed strong neuroprotective properties against Abeta1-42 and reversed Abeta1-42-induced ATP depletion on neuronal cells, suggesting a mitochondrial site of action. Abeta1-42 has been described to induce a hyperactivity of the glutamate network in neuronal cells. (4-Ethyl-piperaz-1-yl)-phenylmethanone also inhibited the neurotoxic effect that glutamate displayed on PC12 cells, suggesting that the reduction of glutamate-induced neurotoxicity may be one of the mechanisms by which this compound exerts its neuroprotective properties against the deleterious effects of the Abeta1-42. These data suggest that the identified (4-ethyl-piperaz-1-yl)-phenylmethanone chemical entity exerts neuroprotective properties and may serve as a lead compound for the development of novel therapies for AD.     

Curcumin inhibits appoptosin-induced apoptosis via upregulating heme oxygenase-1 expression in SH-SY5Y cells

Aim:

Appoptosin (SLC25A38) is a pro-apoptotic protein, which is upregulated in Alzheimer''s disease (AD) brains and plays an important role in promoting the pathological progress of AD. The aim of this study was to investigate the effects of curcumin from the rhizome of Curcuma longa on appoptosin-induced apoptosis in SH-SY5Y cells.

Methods:

SH-SY5Y cells were pretreated with curcumin, then transfected with appoptosin or vector. The apoptotic cells were detected with Annexin V staining analysis by flow cytometry. The expression of cleaved caspase-3, appoptosin, heme oxygenase-1 (HO-1) was examined using Western blotting. Intracellular level of ROS was measured with DCFH-DA staining by flow cytometry analysis. Mitochondrial membrane potential (ΔΨm) was detected with JC-1 staining under a fluorescence microscope and quantified by fluorescence ratio detection.Overexpression of appoptosin in SH-SY5Y cells markedly increased cell apoptosis accompanied by reduced HO-1 expression, increased intracellular heme level, ROS overproduction and ΔΨm impairment. Treatment of SH-SY5Y cells with curcumin (2.5–20 μmol/L) for 24 h did not significantly affect their viability. However, pretreatment with curcumin (2.5–20 μmol/L) dose-dependently attenuated all above-mentioned pathological changes in appoptosin-transfected SH-SY5Y cells.

Results:

Overexpression of appoptosin in SH-SY5Y cells markedly increased cell apoptosis accompanied by reduced HO-1 expression, increased intracellular heme level, ROS overproduction and ΔΨm impairment. Treatment of SH-SY5Y cells with curcumin (2.5–20 μmol/L) for 24 h did not significantly affect their viability. However, pretreatment with curcumin (2.5–20 μmol/L) dose-dependently attenuated all above-mentioned pathological changes in appoptosin-transfected SH-SY5Y cells.

Conclusion:

Curcumin inhibits appoptosin-induced apoptosis in SH-SY5Y cells by upregulating the expression of HO-1, reducing the production of intracellular heme and ROS, and preventing the ΔΨm loss.     

Identification of molecular signatures predicting the carcinogenicity of polycyclic aromatic hydrocarbons (PAHs)

There are multiple lines of evidence showing that environmental toxicants including pesticides may contribute to neuronal cell death. Fipronil (FPN) is a phenylpyrazole insecticide that acts on insect GABA receptors. Although the action of FPN is restricted to insect neuronal or muscular transmitter systems, a few studies have assessed the effects of this neurotoxicant on neuronal cell death distinct from an insect. To determine the mechanisms underlying FPN-induced neuronal cell death, we evaluated the ability of this chemical to induce oxidative stress and studied the involvement of mitogen activated protein kinases (MAPKs) in FPN-induced apoptosis stress in human neuroblastoma SH-SY5Y (SH-SY5Y) cells. Exposure of SH-SY5Y cells to FPN led to the production of reactive oxygen species (ROS) and apoptotic cell death via activation of caspase-9 and caspase-3. Interestingly, the antioxidant, N-acetyl-cysteine (NAC) attenuated apoptotic cell death and ROS production induced by FPN. These results indicated that oxidative stress plays a central role in FPN-induced cytotoxicity. Mitochondrial complex I activity was also inhibited by FPN treatment. These finding indicate that FPN triggers intrinsic apoptosis via the mitochondrial signaling pathway that is initiated by the generation of ROS. Furthermore, FPN treatment induced phosphorylation of MAPK members. Activation of these protein kinases by FPN was involved in the onset of apoptosis as inhibitors specific to these kinases protect against FPN-induced cell death as well as ROS generation. Our data indicate that FPN-induced apoptosis is mediated primarily by the generation of ROS and activation of MAPK members followed by activation of the intrinsic apoptotic pathway.     

左旋黄皮酰胺对冈田酸和β淀粉样肽25-35神经毒性的保护作用

探讨左旋黄皮酰胺对冈田酸(okadaic acid，OA)诱导的人神经瘤细胞(SH-SY5Y)和去卵巢(ovariectomy，OVX)及单侧侧脑室注射Aβ_25-35所致神经元损伤的保护作用。通过MTT试验、LDH释放测定试验、Hoechst 33258荧光染色试验以及SH-SY5Y细胞检测，考察左旋黄皮酰胺拮抗冈田酸诱导的细胞毒作用。通过避暗试验、电镜检测、Nissl体染色及HE染色，考察左旋黄皮酰胺对去卵巢及侧脑室注射Aβ_25-35大鼠神经元的保护作用。左旋黄皮酰胺可明显拮抗冈田酸诱导的细胞毒作用，提高去卵巢及侧脑室注射Aβ_25-35大鼠的学习记忆能力，保护海马及皮层神经元。左旋黄皮酰胺可拮抗冈田酸及Aβ_25-35诱导的神经毒性，具有神经保护作用。     

Acrylonitrile induced apoptosis via oxidative stress in neuroblastoma SH-SY5Y cell

Acrylonitrile (ACN) is a chemical that is widely used in the production of plastics, acrylic fibers, synthetic rubbers and resins. It has been reported that ACN can cause oxidative stress, a condition which is well recognized as an apoptotic initiator; however, information regarding ACN-induced apoptosis is limited. This present study investigated whether ACN induces apoptosis in human neuroblastoma SH-SY5Y cells, and whether its apoptotic induction involves oxidative stress. The results showed that ACN caused activation of caspase-3, a key enzyme involved in apoptosis, in a dose- and time-dependent manner. Detection of sub-G1 apoptotic cell death and apoptotic nuclear condensation revealed that ACN caused an increase in the number of apoptotic cells indicating ACN induces apoptosis in SH-SY5Y cells. ACN dose- and time-dependently increased the level of proapoptotic protein, Bax. Pretreatment with N-acetylcysteine (NAC), an antioxidant, attenuated caspase-3 activation by ACN, as evidenced by a reduction in proteolysis of PARP, a known caspase-3 substrate, as well as in the number of sub-G1 apoptotic cells. Moreover, induction of Bax by ACN was abolished by NAC. Taken together, the results indicate that ACN induces apoptosis in SH-SY5Y cells via a mechanism involving generation of oxidative stress-mediated Bax induction.     

Acidic oligosaccharide sugar chain, a marine-derived acidic oligosaccharide, inhibits the cytotoxicity and aggregation of amyloid beta protein

In this paper, we investigated interactions of the acidic oligosaccharide sugar chain (AOSC), derived from brown algae Echlonia kurome OKAM, with amyloid beta protein (Abeta). We observed that AOSC inhibited the toxicity induced by Abeta in both primarily cortical cells and the SH-SY5Y cell line. We also observed that AOSC inhibited the apoptosis induced by Abeta in SH-SY5Y by reducing the elevated level of intracellular calcium concentration ([Ca(2+)](i)) and suppressing the generation of reactive oxygen species. Surface plasmon resonance analysis demonstrated that AOSC had affinity for both freshly-dissolved Abeta and 48-h incubated Abeta. Furthermore, AOSC blocked the fibril formation of Abeta, which may be responsible for its anti-cytotoxic effects. Thus, our results indicate that AOSC might be a potentially therapeutic compound for Alzheimer's disease.     

beta-Adrenoceptor blockers protect against staurosporine-induced apoptosis in SH-SY5Y neuroblastoma cells

The beta-adrenoceptor blockers exhibit a well-characterized anti-apoptotic property in the heart and kidney while less is known about the effect of this class of drugs on neuronal apoptosis. We studied the effects of three beta-adrenoceptor blockers propranolol (1-(isoproplyamino)-3-(naphthalene-1-yloxy)propan-2-ol), atenolol (2-[4-[2-hydroxy-3-(1-methylethylamino)propoxyl]phenyl]ehanamide), and ICI 118551 (1-[2,3-(dihydro-7-methyl-1H-iden-4-yl)oxy]-3-[(1-methylethyl)amino]-2-butanol), against staurosporine-induced apoptosis in SH-SY5Y human neuroblastoma cells. Staurosporine increased caspase 3-like activity, DNA fragmentation, PARP cleavage, and the number of TUNEL positive cells consistent with the induction of apoptosis. Propranolol and ICI 118551, but not atenolol, demonstrated a concentration-dependent inhibition of caspase 3-like activity. Propranolol and ICI 118551 directly inhibited the enzymatic activity of recombinant caspase 9 while atenolol did not; however, none of the beta-adrenoceptor blockers that were examined directly blocked caspases 2 or 3 activity. In isolated mitochondria, propranolol and ICI 118551 inhibited staurosporine-induced cytochrome c release while atenolol did not. We conclude that propranolol and ICI 118551 protect SH-SY5Y cells against staurosporine-induced apoptosis through a dual action on the mitochondria and on caspase 9 in a cell type and an apoptotic paradigm where the conventional inhibitors of mitochondrial permeability transition such as cyclosporin A and bongkrekic acid demonstrate no protection.     

Amyloid precursor protein binding protein Fe65 is cleaved by caspases during DNA damage-induced apoptosis

Caspases cleave several cellular proteins to execute cell death by apoptosis. The identification of novel substrates of caspases could provide an important clue for elucidation of new apoptosis signaling pathways. In this study, we tested whether an amyloid precursor protein (APP) binding protein Fe65 is proteolytically degraded in neuronal cell death by apoptosis, using a neuron-like cell line, human neuroblastoma SH-SY5Y cells. When treated with DNA damaging agents, etoposide (ETP) and camptothecin (CPT), SH-SY5Y cells underwent apoptosis in a dose-dependent manner. Interestingly, Fe65 (97 kDa) was cleaved to a 65 kDa product during DNA damage-induced apoptosis. Furthermore, the cleavage of Fe65 was accompanied by activation of caspases-9 and -3. The restriction cleavage of Fe65 was completely suppressed by the treatment with a pan-caspase inhibitor N-benzyloxycarbonyl-Val-Ala-Asp(OMe) fluoromethylketone (z-VAD-fmk). These results reveal the restriction cleavage of Fe65 by caspases during DNA damage-induced apoptosis. Since Fe65 has been shown to suppress APP processing to amyloid β (Aβ) production, our findings may provide a new insight into the molecular mechanism by which DNA damage induces Aβ production and subsequent neuronal cell death in Alzheimer's disease (AD).     

PINK1/Parkin-mediated mitophagy play a protective role in manganese induced apoptosis in SH-SY5Y cells

Manganese (Mn) as an environmental risk factor of Parkinson's disease (PD) is considered to cause manganism. Mitophagy is thought to play a key role in elimination the injured mitochondria. The goal of this paper was to explore whether the PINK1/Parkin-mediated mitophagy is activated and its role in Mn-induced mitochondrial dysfunction and cell death in SH-SY5Y cells. Here, we investigated effects of MnCl₂ on ROS generation, mitochondrial membrane potential (MMP/ΔΨ_m) and apoptosis by FACS and examined PINK1/Parkin-mediated mitophagy by western-blotting and the co-localization of mitochondria and acidic lysosomes. Further, we explore the role of mitophagy in Mn-induced apoptosis by inhibition the mitophagy by knockdown Parkin level. Results show that MnCl₂ dose-dependently caused ΔΨ_m decrease, ROS generation and apoptosis of dopaminergic SH-SY5Y cells. Moreover, Mn could induce mitophagy and PINK1/Parkin-mediated pathway was activated in SH-SY5Y cells. Transient transfection of Parkin siRNA knockdown the expressing level of parkin inhibited Mn-induced mitophagy and aggravated apoptosis of SH-SY5Y cells. In conclusion, our study demonstrated that Mn may induce PINK1/Parkin-mediated mitophagy, which may exert significant neuro-protective effect against Mn-induced dopaminergic neuronal cells apoptosis.     

TEMPOL protects human neuroblastoma SH-SY5Y cells against β-amyloid-induced cell toxicity

Amyloid-β peptide (Aβ) has been implicated in the pathogenesis of Alzheimer's disease (AD). It can cause cell death in Alzheimer's disease by evoking a cascade of oxidative damage to neurons. Antioxidant compounds may help to elucidate and develop a treatment for Alzheimer's disease. In the present study, we investigated the protective effect of TEMPOL (4-hydroxy-2,2,6,6-tetramethyl-1-piperidinyloxy), a cyclic nitroxide which is particularly effective at reducing oxidative injury, on Aβ(1-42)-induced SH-SY5Y cell toxicity. Exposure of cells to 20 μM Aβ(1-42) for 48 h caused viability loss and apoptotic increase, and pre-treatment with TEMPOL for 24 h significantly reduced the viability loss and apoptotic rate. In addition, TEMPOL inhibited Aβ(1-42)-induced superoxide anion generation and hydroxyl radical generation to a striking degree. Based on these results, it is concluded that TEMPOL effectively protects SH-SY5Y cells against β-amyloid-induced damage by suppressing the generation of reactive oxygen species especially, superoxide anion.