Hibiscus Flower and Olive Leaf Extracts Activate Apoptosis in SH-SY5Y Cells.
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ABSTRACT:
SUBMITTER: Chiaino E
PROVIDER: S-EPMC8750347 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Accumulating evidences suggest that p53 is a key coordinator of cellular events triggered by oxidative stress often associated with the impairment in thiamine metabolism and its functions. However, there are limited data regarding the pursuant feedback between p53 transactivation and thiamine homeostasis. Impairment in thiamine metabolism can be induced experimentally via interference with the thiamine uptake and/or inhibition of the thiamin pyrophosphate-dependent enzymes using thiamine antagonists - amprolium (AM), oxythiamine (OT) or pyrithiamine (PT). We found that exposure of neuronally differentiated SH-SY5Y cells to AM, OT and PT triggered upregulation of p53 gene expression, post-translational modification of p53 via phosphorylation and activation of p53 DNA-binding activity. Phosphorylation of p53 at Ser20 was equally efficient in upregulation of thiamine transporter 1 (THTR1) by all antagonists. However, induction of the expressions of the pyruvate dehydrogenase E1 component subunit beta (PDHB) and oxoglutarate dehydrogenase (OGDH) required dual phosphorylation of p53 at Ser9 and Ser20, seen in cells treated with PT and OT. Moreover, pretreatment of the cells with a decoy oligonucleotide carrying wild-type p53-response element markedly attenuated OT-induced THTR1, PDHB and OGDH gene expression suggesting an important role of p53 in transactivation of these genes. Finally, analysis of gene and metabolic networks showed that OT triggers cell apoptosis through the p53-dependent intrinsic pathway.
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Project description: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.
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Project description:BackgroundTwo active compounds, baicalein and its glycoside baicalin were found in the dried root of Scutellaria baicalensis Georgi, and reported to be neuroprotective in vitro and in vivo. This study aims to evaluate the protective effects of baicalein on the rotenone-induced apoptosis in dopaminergic SH-SY5Y cells related to parkinsonism.MethodsCell viability and cytotoxicity were determined by MTT assay. The degree of nuclear apoptosis was evaluated with a fluorescent DNA-binding probe Hoechst 33258. The production of reactive oxidative species (ROS) and loss of mitochondrial membrane potential (??m) were determined by fluorescent staining with DCFH-DA and Rhodanmine 123, respectively. The expression of Bax, Bcl-2, cleaved caspase-3 and phosphorylated ERK1/2 was determined by the Western blots.ResultsBaicalein significantly increased viability and decreased rotenone-induced death of SH-SY5Y cells in a dose-dependent manner. Pre- and subsequent co-treatment with baicalein preserved the cell morphology and attenuated the nuclear apoptotic characteristics triggered by rotenone. Baicalein antagonized rotenone-induced overproduction of ROS, loss of ??m, the increased expression of Bax, cleaved caspase-3 and phosphorylated ERK1/2 and the decreased expression of Bcl-2.ConclusionThe antioxidative effect, mitochondrial protection and modulation of anti-and pro-apoptotic proteins are related to the neuroprotective effects of baicalein against rotenone induced cell death in SH-SY5Y cells.
| S-EPMC3275529 | biostudies-literature
Project description:Oleoylethanolamide (OEA) and palmitoylethanolamide (PEA) are endogenous lipids that act as agonists of the peroxisome proliferator-activated receptor α (PPARα). Recently, an interest in the role of these lipids in malignant tumors has emerged. Nevertheless, the effects of OEA and PEA on human neuroblastoma cells are still not documented. Type I interferons (IFNs) are immunomodulatory cytokines endowed with antiviral and anti-proliferative actions and are used in the treatment of various pathologies such as different cancer forms (i.e., non-Hodgkin's lymphoma, melanoma, leukemia), hepatitis B, hepatitis C, multiple sclerosis, and many others. In this study, we investigated the effect of OEA and PEA on human neuroblastoma SH-SY5Y cells treated with IFNβ. We focused on evaluating cell viability, cell proliferation, and cell signaling. Co-exposure to either OEA or PEA along with IFNβ leads to increased apoptotic cell death marked by the cleavage of caspase 3 and poly-(ADP ribose) polymerase (PARP) alongside a decrease in survivin and IKBα levels. Moreover, we found that OEA and PEA did not affect IFNβ signaling through the JAK-STAT pathway and the STAT1-inducible protein kinase R (PKR). OEA and PEA also increased the phosphorylation of p38 MAP kinase and programmed death-ligand 1 (PD-L1) expression both in full cell lysate and surface membranes. Furthermore, GW6471, a PPARα inhibitor, and the genetic silencing of the receptor were shown to lower PD-L1 and cleaved PARP levels. These results reveal the presence of a novel mechanism, independent of the IFNβ-prompted pathway, by which OEA and PEA can directly impair cell survival, proliferation, and clonogenicity through modulating and potentiating the intrinsic apoptotic pathway in human SH-SY5Y cells.
| S-EPMC11013881 | biostudies-literature
Project description:Inflammation has long been implicated as a contributor to pathogenesis in many CNS illnesses, including Lyme neuroborreliosis. Borrelia burgdorferi is the spirochete that causes Lyme disease and it is known to potently induce the production of inflammatory mediators in a variety of cells. In experiments where B. burgdorferi was co-cultured in vitro with primary microglia, we observed robust expression and release of IL-6 and IL-8, CCL2 (MCP-1), CCL3 (MIP-1alpha), CCL4 (MIP-1beta) and CCL5 (RANTES), but we detected no induction of microglial apoptosis. In contrast, SH-SY5Y (SY) neuroblastoma cells co-cultured with B. burgdorferi expressed negligible amounts of inflammatory mediators and also remained resistant to apoptosis. When SY cells were co-cultured with microglia and B. burgdorferi, significant neuronal apoptosis consistently occurred. Confocal microscopy imaging of these cell cultures stained for apoptosis and with cell type-specific markers confirmed that it was predominantly the SY cells that were dying. Microarray analysis demonstrated an intense microglia-mediated inflammatory response to B. burgdorferi including up-regulation in gene transcripts for TLR-2 and NFkappabeta. Surprisingly, a pathway that exhibited profound changes in regard to inflammatory signaling was triggering receptor expressed on myeloid cells-1 (TREM1). Significant transcript alterations in essential p53 pathway genes also occurred in SY cells cultured in the presence of microglia and B. burgdorferi, which indicated a shift from cell survival to preparation for apoptosis when compared to SY cells cultured in the presence of B. burgdorferi alone. Taken together, these findings indicate that B. burgdorferi is not directly toxic to SY cells; rather, these cells become distressed and die in the inflammatory surroundings generated by microglia through a bystander effect. If, as we hypothesized, neuronal apoptosis is the key pathogenic event in Lyme neuroborreliosis, then targeting microglial responses may be a significant therapeutic approach for the treatment of this form of Lyme disease.
| S-EPMC2771360 | biostudies-literature
Project description:Acetamiprid (ACE), a commonly used neonicotinoid insecticide, is correlated with neurological symptoms, immunotoxicity and hepatotoxicity. Cellular stress and damage could play an important role in ACE-induced neurotoxicity; however, its mechanism has not been fully understood. We evaluated the effects of ACE on oxidative stress, endoplasmic reticulum (ER) stress, cellular death, mRNA expression levels of related genes and protein expressions of related molecular mechanisms in SH-SY5Y human neuroblastoma cells. The half maximal inhibition of enzyme activity (IC50) value of ACE was determined as 4.26 mM after 24 h of treatment by MTT assay. We revealed an increase in reactive oxygen species (ROS) production and calcium release. Significant increases were measured in inositol-requiring enzyme 1-alpha (IRE1-α) and binding immunoglobulin protein 90 (GRP90) levels as well as mRNA expression levels of caspase 3, 4 and 9 genes indicating enhanced ER stress. Apoptosis and ER stress-related genes were significantly upregulated at ≥2 mM. Indeed, ACE caused apoptosis and necroptosis while necrosis was not observed. There was a significant increase in the protein level of mitogen-activated protein kinase-8 (MAPK8) at 4 mM of ACE while no change was seen for nuclear factor kappa-B (NF-κB) and tumor necrosis factor-alpha (TNF-α). In conclusion, increased cellular stress markers could be proposed as an underlying mechanism of ACE-induced cell death in neural cells.
| S-EPMC8467785 | biostudies-literature
Project description:Alzheimer's disease (AD) is a progressive neurodegenerative disorder with an unclear cause. It appears that multiple factors participate in the process of neuronal damage including oxidative stress and accumulation of the protein amyloid β (Aβ) in the brain. The search for a treatment for this disorder is essential as current medications are limited to alleviating symptoms and palliative effects. The aim of this study is to investigate the effects of mint extracts on selected mechanisms implicated in the development of AD. To enable a thorough investigation of mechanisms, including effects on β-secretase (the enzyme that leads to the formation of Aβ), on Aβ aggregation, and on oxidative stress and apoptosis pathways, a neuronal cell model, SH-SY5Y cells, was selected. Six Mentha taxa were investigated for their in vitro β-secretase (BACE) and Aβ-aggregation inhibition activities. Moreover, their neuroprotective effects on H2O2-induced oxidative stress and apoptosis in SH-SY5Y cells were evaluated through caspase activity. Real-time PCR and Western blot analysis were carried out for the two most promising extracts to determine their effects on signalling pathways in SH-SY5Y cells. All mint extracts had strong BACE inhibition activity. M. requienii extracts showed excellent inhibition of Aβ-aggregation, while other extracts showed moderate inhibition. M. diemenica and M. requienii extracts lowered caspase activity. Exposure of SH-SY5Y cells to M. diemenica extracts resulted in a decrease in the expression of pro-apoptotic protein, Bax, and an elevation in the anti-apoptotic protein, Bcl-xL, potentially mediated by down-regulation of the ASK1-JNK pathway. These results indicate that mint extracts could prevent the formation of Aβ and also could prevent their aggregation if they had already formed. M. diemenica and M. requienii extracts have potential to suppress apoptosis at the cellular level. Hence, mint extracts could provide a source of efficacious compounds for a therapeutic approach for AD.
| S-EPMC7285141 | biostudies-literature
Project description:We have recently reported that, in SH-SY5Y cells, mu-opioid receptor occupancy activates phospholipase C via a pertussis toxin-sensitive G-protein. In the present study we have further characterized the mechanisms involved in this process. Fentanyl (0.1 microM) caused a monophasic increase in inositol 1,4,5-trisphosphate mass formation, with a peak (20.5 +/- 3.6 pmol/mg of protein) at 15 s. Incubation in Ca(2+)-free buffer abolished this response, while Ca2+ replacement 1 min later restored the stimulation of inositol 1,4,5-trisphosphate formation (20.1 +/- 0.6 pmol/mg of protein). In addition, nifedipine (1 nM-0.1 mM), an L-type Ca(2+)-channel antagonist, caused a dose-dependent inhibition of inositol 1,4,5-trisphosphate formation, with an IC50 of 60.3 +/- 1.1 nM. Elevation of endogenous beta/gamma subunits by selective activation of delta-opioid and alpha 2 adrenoceptors failed to stimulate phospholipase C. Fentanyl also caused a dose-dependent (EC50 of 16.2 +/- 1.0 nM), additive enhancement of carbachol-induced inositol 1,4,5-trisphosphate formation. In summary, we have demonstrated that in SH-SY5Y cells activation of the mu-opioid receptor allows Ca2+ influx to activate phospholipase C. However, the possible role of this mechanism in the process of analgesia remains to be elucidated.
| S-EPMC1136401 | biostudies-other
Project description:Cultured SH-SY5Y human neuroblastoma cells are used in neurotoxicity assays. These cells express markers of the cholinergic and dopaminergic systems. Acetylcholinesterase (AChE) activity has been reported in these cells. Neurotoxic organophosphate compounds that inhibit AChE, also inhibit butyrylcholinesterase (BChE). We confirmed the presence of AChE in the cell lysate by activity assays, Western blot, and liquid chromatography-tandem mass spectrometry (LC-MS/MS) of immunopurified AChE. A nondenaturing gel stained for AChE activity identified the catalytically active AChE in SH-SY5Y cells as the unstable monomer. We also identified immature BChE in the cell lysate. The concentration of active BChE protein was similar to that of active AChE protein. The rate of substrate hydrolysis by AChE was 10-fold higher than substrate hydrolysis by BChE. The higher rate was due to the 10-fold higher specific activity of AChE over BChE (5000 units/mg for AChE; 500 units/mg for BChE). Neither cholinesterase was secreted. Tryptic peptides of immunopurified AChE and BChE were identified by LC-MS/MS on an Orbitrap Lumos Fusion mass spectrometer. The unfolded protein chaperone, binding immunoglobulin protein BiP/GRP78, was identified in the mass spectral data from all cholinesterase samples, suggesting that BiP was co-extracted with cholinesterase. This suggests that the cytoplasmic cholinesterases are immature forms of AChE and BChE that bind to BiP. It was concluded that SH-SY5Y cells express active AChE and active BChE, but the proteins do not mature to glycosylated tetramers.
| S-EPMC9124689 | biostudies-literature
Project description:Background In childhood, metastatic neuroblastoma (NB) is the most common extracranial solid tumor, but there are no appropriate drugs for its treatment. Dihydroartemisinin (DHA), a drug for malaria treatment, has therapeutic potential in several cancers; however, its mechanisms remain unclear. This study aimed to investigate the anti-proliferation effect of DHA on SH-SY5Y cells and to explore its mechanism in vitro. Methods We used 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay to measure the half-maximal inhibitory concentration (IC50) of DHA; western blot was used to determine protein levels; propidium iodide (PI) staining was used to determine apoptotic cells; JC-1 staining to measure mitochondrial membrane potential; and dichloro-dihydro-fluorescein diacetate (DCFH-DA) staining was used to determine reactive oxygen species (ROS). Metabonomic analysis was performed by using ultra-high performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS)-based untargeted metabolomics. Multivariate statistical analysis was performed to screen potential metabolites associated with DHA treatment in SH-SY5Y cells. Results It was shown that DHA inhibited SH-SY5Y cell proliferation and increased poly (ADP-ribose) polymerase (PARP-1) and caspase 3 in a dose-dependent manner. In Further, DHA promoted ROS generation and γH2AX expression. In addition, a total of 125 proposed metabolites in SH-SY5Y cells and 45 vital metabolic pathways were identified through UHPLC-MS/MS-based untargeted metabolomic analysis. Conclusions These data suggest that DHA could regulate taurine, linoleic acid, phenylalanine metabolism, and tryptophan metabolism, which are involved in the anti-proliferation effect of DHA in SH-SY5Y cells.
| S-EPMC9442203 | biostudies-literature