Lycium Barbarum Polysaccharides Alleviates Oxidative Damage Induced by H2O2 Through Down-Regulating MicroRNA-194 in PC-12 and SH-SY5Y Cells

Background/Aims: Currently, scientists attempt to improve outcome of spinal cord injury (SCI) via reducing secondary injury during SCI. Oxidative stress is critical for pathophysiology of secondary damage, thus we mainly focused on the anti-oxidant effects of Lycium barbarum polysaccharides (LBPs) on PC-12 and SH-SY5Y cells as well as the underlying mechanisms. Methods: Oxidative stress was induced by H2O2 stimulation. Effects of LBPs on cell viability, apoptosis, and expression of proteins associated with apoptosis and autophagy in H2O2-induced cells were assessed by CCK-8 assay, flow cytometry assay and Western blot analysis, respectively. Then, expression of miR-194 was determined by qRT-PCR. Expression of miR-194 was dysregulated, and whether LBPs affected H2O2-treated cells through modulating miR-194 was verified. The expression of key kinases in the PI3K/AKT pathway and the intracellular levels of ROS and NO were testified by Western blot analysis and flow cytometry with fluorescent probes. Results: H2O2-induced decrease of cell viability and increases of apoptosis and autophagy in PC-12 cells were mitigated by LBPs treatment. Next, we found that miR-194 expression was both down-regulated by LBPs treatment in PC-12 and SH-SY5Y cells. More experiments consolidated that influence of LBPs on H2O2-treated cells was reversed by miR-194 overexpression while was augmented by miR-194 inhibition. LBPs elevated the phosphorylated levels of PI3K and AKT and reduced levels of ROS and NO through miR-194. Conclusion: LBPs alleviated H2O2-induced decrease of cell viability, and increase of apoptosis and autophagy through down-regulating miR-194. Moreover, LBPs activated the PI3K/AKT pathway and reduced oxidative stress through miR-194.

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Rhamnazin Ameliorates Traumatic Brain Injury in Mice via Reduction in Apoptosis, Oxidative Stress, and Inflammation

NeuroImmunoModulation ◽

10.1159/000516927 ◽

2021 ◽

pp. 1-8

Author(s):

Boxiao Yang ◽

Rui Zhang ◽

Qire Sa ◽

Yanli Du

Keyword(s):

Oxidative Stress ◽

Traumatic Brain Injury ◽

Flow Cytometry ◽

Brain Injury ◽

Western Blot ◽

Protective Effect ◽

Blot Analysis ◽

Western Blot Analysis ◽

Neuronal Degeneration ◽

Flow Cytometry Analysis

Background: Traumatic brain injury (TBI) is posing serious health challenges for people across the globe due to high morbidity and mortality. However, none of the agents prevents or limits the damage caused by TBI because of its multifactorial etiology. Thus, the discovery of novel agents which can act via several pathways could serve the purpose and afford favorable consequence against TBI. Therefore, in the present article, we intended to investigate the protective effect of rhamnazin (RMZ), a dimethoxyflavone against experimentally induced TBI in mice. Methods: The effect of RMZ was investigated on cerebral edema and grip test score after induction of experimental brain injury in rats. The effect of RMZ was also investigated on neuronal degeneration in brain tissues of the experimental mice via Nissl staining and flow cytometry analysis. The expression of Bax and Bcl-2 was also quantified using Western blot analysis. The level of inflammatory cytokines (TNF-α and IL-1β) and oxidative stress markers (malondialdehyde, superoxide dismutase, and glutathione peroxidase) was also determined using enzyme-linked immunosorbent assay. Results: RMZ showed a significant reduction in edema and improved grip strength. It also prevented neuronal degeneration via inhibition of neuronal apoptosis as shown by flow cytometry analysis. RMZ showed an antiapoptotic effect via reduction of Bax and increased the expression of Bcl-2 in Western blot analysis. It also showed to inhibit oxidative stress and inflammation compared to the TBI group. Conclusion: Collectively, our study is first to demonstrate the protective effect of RMZ against experimentally induced TBI in rats.

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Paeonol Induces Protective Autophagy in Retinal Photoreceptor Cells

Frontiers in Pharmacology ◽

10.3389/fphar.2021.667959 ◽

2021 ◽

Vol 12 ◽

Author(s):

Daowei Zhang ◽

Jiawen Wu ◽

Jihong Wu ◽

Shenghai Zhang

Keyword(s):

Oxidative Stress ◽

Western Blot ◽

Cell Viability ◽

Blot Analysis ◽

Western Blot Analysis ◽

Critical Role ◽

Cell Counting ◽

Terminal Deoxynucleotidyl Transferase ◽

Retinal Photoreceptor

Background: Retinal photoreceptor (RP) cells are widely involved in retina-related diseases, and oxidative stress plays a critical role in retinal secondary damage. Herein, we investigated the effectiveness and potential mechanisms of autophagy of paeonol (Pae) in terms of oxidation resistance.Methods: The animal model was induced by light damage (LD) in vivo, whereas the in vitro model was established by H2O2 stimulation. The effectiveness of Pae was evaluated by hematoxylin and eosin, terminal deoxynucleotidyl transferase dUTP nick end labeling assay, immunofluorescence, transmission electron microscopy, electroretinogram, and Western blot analysis in vivo, and the underlying mechanisms of Pae were assessed by Cell Counting Kit-8 assay, reactive oxygen species (ROS) assay, and Western blot analysis in 661W cells. We mainly evaluated the effects of Pae on apoptosis and autophagy.Results: Increased apoptosis of the LD-induced and decreased autophagy of RPs were mitigated by Pae treatment. Pea, which increased the expression of mitochondrial functional protein cytochrome c, reversed the decreased cell viability and autophagy induced by oxidative stress in 661W cells. Experiments showed that autophagy was downregulated in PINK1/Parkin dependent and the BNIP3L/Nix dependent pathways under H2O2 stimulation and was upregulated by Pae treatment. Pae increased the cell viability and reduced ROS levels through autophagy.Conclusion: Pretreatment with Pae preserved RP cells by enhancing autophagy, which protected retinal function.

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Neogenin-1 Promotes Cell Proliferation, Motility, and Adhesion by Up-Regulation of Zinc Finger E-Box Binding Homeobox 1 Via Activating the Rac1/PI3K/AKT Pathway in Gastric Cancer Cells

Cellular Physiology and Biochemistry ◽

10.1159/000492255 ◽

2018 ◽

Vol 48 (4) ◽

pp. 1457-1467 ◽

Cited By ~ 6

Author(s):

Hengyi Qu ◽

Huabo Sun ◽

Xueping Wang

Keyword(s):

Gastric Cancer ◽

Cell Proliferation ◽

Western Blot ◽

Cell Viability ◽

Cell Lines ◽

Zinc Finger ◽

Blot Analysis ◽

Western Blot Analysis ◽

Akt Pathway ◽

E Box

Background/Aims: Neogenin-1 (Neo1) has been reported to be involved in diverse physiology and pathology functions, including cell proliferation, differentiation and migration. The present study aimed to explore the functional role of neogenin-1 (Neo1) in gastric cancer (GC), together with underlying mechanisms. Methods: Neo1 expression was analyzed by qRT-PCR and Western blot analysis in both human GC cell lines and normal gastric epithelial cell line. Neo1 was respectively overexpressed or silenced by transfection with pcDNA3.1 or siRNA, and then the cells were incubated with or without different concentrations of cisplatin, transforming growth factor (TGF)-β1, and/or inhibitors of Rac-1 and PI3K. Thereafter, cell viability, invasion, and adhesion were measured by CCK-8, wound healing and adhesion assays, respectively. The expression levels of key factors involved in epithelial mesenchymal transition (EMT) and the PI3K/AKT pathway were analyzed by Western blot analysis. Results: The results showed that the Neo1 level was significantly increased in GC cell lines, with the highest level in SGC-7901 cells. Overexpression of Neo1 significantly reduced the GC cell sensitivity to cisplatin and increased the cell viability, motility and adhesion ability, and while silencing of Neo1 showed contrary results. Moreover, overexpression of Neo1 dramatically downregulated the E-Cadherin level and upregulated the levels of N-Cadherin and Vimentin. In addition, the data revealed that Neo1 positively regulated the expression of Zinc finger E-box-binding homeobox 1 (ZEB1) by activating the Rac1/PI3K/AKT pathway. Conclusions: Neo1 could promote cell proliferation, motility, and adhesion by up-regulation of ZEB1 via activating the Rac1/PI3K/AKT pathway in GC cells.

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Kaempferol and Kaempferide Attenuate Oleic Acid-Induced Lipid Accumulation and Oxidative Stress in HepG2 Cells

International Journal of Molecular Sciences ◽

10.3390/ijms22168847 ◽

2021 ◽

Vol 22 (16) ◽

pp. 8847

Author(s):

Fangfang Tie ◽

Jin Ding ◽

Na Hu ◽

Qi Dong ◽

Zhi Chen ◽

...

Keyword(s):

Oxidative Stress ◽

Oleic Acid ◽

Lipid Metabolism ◽

Western Blot ◽

Blot Analysis ◽

Lipid Accumulation ◽

Hepg2 Cells ◽

Western Blot Analysis ◽

Heme Oxygenase 1 ◽

And Oxidative Stress

Nonalcoholic fatty liver disease (NAFLD) is one of the most common liver diseases which lacks ideal treatment options. Kaempferol and kaempferide, two natural flavonol compounds isolated from Hippophae rhamnoides L., were reported to exhibit a strong regulatory effect on lipid metabolism, for which the mechanism is largely unknown. In the present study, we investigated the effects of kaempferol and kaempferide on oleic acid (OA)-treated HepG2 cells, a widely used in vitro model of NAFLD. The results indicated an increased accumulation of lipid droplets and triacylglycerol (TG) by OA, which was attenuated by kaempferol and kaempferide (5, 10 and 20 μM). Western blot analysis demonstrated that kaempferol and kaempferide reduced expression of lipogenesis-related proteins, including sterol regulatory element-binding protein 1 (SREBP1), fatty acid synthase (FAS) and stearoyl-CoA desaturase 1 (SCD-1). Expression of peroxisome proliferator-activated receptor γ (PPARγ) and CCAAT enhancer binding proteins β (C/EBPβ), two adipogenic transcription factors, was also decreased by kaempferol and kaempferide treatment. In addition, western blot analysis also demonstrated that kaempferol and kaempferide reduced expression of heme oxygenase-1 (HO-1) and nuclear transcription factor-erythroid 2-related factor 2 (Nrf2). Molecular docking was performed to identify the direct molecular targets of kaempferol and kaempferide, and their binding to SCD-1, a critical regulator in lipid metabolism, was revealed. Taken together, our findings demonstrate that kaempferol and kaempferide could attenuate OA-induced lipid accumulation and oxidative stress in HepG2 cells, which might benefit the treatment of NAFLD.

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MicroRNA-145 attenuates IL-6-induced enhancements of sensitivity to UVB irradiation by suppressing MyD88 in HaCaT cells

International Journal of Immunopathology and Pharmacology ◽

10.1177/2058738418795940 ◽

2018 ◽

Vol 32 ◽

pp. 205873841879594 ◽

Cited By ~ 3

Author(s):

Hui Dong ◽

Wei Jiang ◽

Hongquan Chen ◽

Shui Jiang ◽

Yunshu Zang ◽

...

Keyword(s):

Western Blot ◽

Cell Viability ◽

Lupus Erythematosus ◽

Blot Analysis ◽

Cell Apoptosis ◽

Western Blot Analysis ◽

Ultraviolet B ◽

Hacat Cells ◽

Related Factors ◽

Uvb Irradiation

MicroRNAs (miRNAs/miRs) play vital roles in various immune diseases including systemic lupus erythematosus (SLE). The current study aimed to assess the role of miR-145 in interleukin-6 (IL-6)-treated HaCaT cells under ultraviolet B (UVB) irradiation and further explore the potential regulatory mechanism. HaCaT cells were pretreated with IL-6 and then exposed to UVB to assess the effect of IL-6 on sensitivity of HaCaT cells to UVB irradiation. The levels of miR-145 and MyD88 were altered by transfection and the transfected efficiency was verified by quantitative reverse transcription polymerase chain reaction (qRT-PCR)/western blot analysis. Cell viability, percentage of apoptotic cells and expression levels of apoptosis-related factors were measured by trypan blue assay, flow cytometry assay, and western blot analysis, respectively. In addition, the levels of c-Jun N-terminal kinases (JNK) and nuclear factor-κB (NF-κB) signaling pathway-related factors were assessed by western blot analysis. IL-6 treatments significantly aggravated the reduction of cell viability and promotion of cell apoptosis caused by UVB irradiation in HaCaT cells. Interestingly, miR-145 level was augmented by UVB exposure and miR-145 mimic alleviated IL-6-induced increase of sensitivity to UVB irradiation in HaCaT cells, as dramatically increased cell viability and reduced cell apoptosis. Opposite effects were observed in miR-145 inhibitor-transfected cells. Meanwhile, MyD88 was negatively regulated by miR-145 and MyD88 mediated the regulatory effect of miR-145 on IL-6- and UVB-treated cells. In addition, miR-145 mimic inhibited the JNK and NF-κB pathways by down-regulating MyD88. In conclusion, the present study demonstrated that miR-145 alleviated IL-6-induced increase of sensitivity to UVB irradiation by down-regulating MyD88 in HaCaT cells.

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Role of EGFR/ErbB2 and PI3K/AKT/e-NOS in Lycium barbarum polysaccharides Ameliorating Endothelial Dysfunction Induced by Oxidative Stress

The American Journal of Chinese Medicine ◽

10.1142/s0192415x19500782 ◽

2019 ◽

Vol 47 (07) ◽

pp. 1523-1539 ◽

Cited By ~ 2

Author(s):

Wenjuan Zhang ◽

Huifang Yang ◽

Lingqin Zhu ◽

Yan Luo ◽

Lihong Nie ◽

...

Keyword(s):

Oxidative Stress ◽

Endothelial Dysfunction ◽

Cell Viability ◽

Therapeutic Option ◽

Critical Role ◽

Ang Ii ◽

Lycium Barbarum ◽

Underlying Mechanisms ◽

Lycium Barbarum Polysaccharides

Lycium barbarum polysaccharides (LBP) are the major ingredients of wolfberry. In this study, we investigated the role of LBP in endothelial dysfunction induced by oxidative stress and the underlying mechanisms using thoracic aortic endothelial cells of rat (RAECs) as a model. We found that Ang II inhibits cell viability of RAECs with 10[Formula: see text][Formula: see text]mol/L of Ang II treatment for 24[Formula: see text]h most potential ([Formula: see text]), the level of reactive oxygen species (ROS) is increased by Ang II treatment ([Formula: see text]), and the expression of Occludin and Zonula occludens-1 (ZO-1) is decreased by Ang II treatment ([Formula: see text]). However, preincubation of cells with LBP could inhibit the changes caused by Ang II, LBP increased cell viability ([Formula: see text]), decreased the level of ROS ([Formula: see text]), and up-regulated the expression of Occludin ([Formula: see text]) and ZO-1. In addition, Ang II treatment increased the expression of EGFR and p-EGFR (Try1172) and which can be inhibited by LBP. On the contrary, expression of ErbB2, p-ErbB2 (Try1248), PI3K, p-e-NOS (Ser1177) ([Formula: see text]), and p-AKT (Ser473) ([Formula: see text]) was inhibited by Ang II treatment and which can be increased by LBP. Treatment of the cells with inhibitors showed that the regulation of p-e-NOS and p-AKT expression by Ang II and LBP can be blocked by PI3K inhibitor wortmannin but not EGFR and ErbB2 inhibitor AC480. Taken together, our results suggested that LBP plays a critical role in maintaining the integrality of blood vessel endothelium through reduced production of ROS via regulating the activity of EGFR, ErbB2, PI3K/AKT/e-NOS, and which may offer a novel therapeutic option in the management of endothelial dysfunction.

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Nutlin-3A, an MDM2 Antagonist, Induces p53-Dependent Cell Cycle Arrest and Apoptosis in Hodgkin Lymphoma (HL).

Blood ◽

10.1182/blood.v108.11.2506.2506 ◽

2006 ◽

Vol 108 (11) ◽

pp. 2506-2506

Author(s):

Elias Drakos ◽

Athanasios Thomaides ◽

Jiang Li ◽

Marina Konopleva ◽

L. Jeffrey Medeiros ◽

...

Keyword(s):

Cell Cycle ◽

Flow Cytometry ◽

Cell Death ◽

Western Blot ◽

Cell Cycle Arrest ◽

Small Molecule ◽

Blot Analysis ◽

Western Blot Analysis ◽

P53 Gene ◽

Cycle Arrest

Abstract p53 is the most frequently mutated tumor suppressor gene in human cancer. However, in Hodgkin lymphoma (HL) p53 is mutated only in a small subset of cases suggesting that modulation of wild-type-p53 (wt-p53) levels in Hodgkin and Reed-Sternberg (HRS) cells may have therapeutic implications in these patients. MDM2 (HDM2 in humans) is a physiologic negative regulator of p53 levels through a well-established auto-regulatory feedback loop. Nutlin-3A is a recently developed small molecule, which antagonizes mdm2 through disruption of p53-MDM2 interaction resulting in p53 stabilization. We hypothesized that nutlin 3A may stabilize p53 in HRS cells carrying wt-p53 gene, thus leading to p53-dependent apoptosis and G1-S cell cycle arrest. We used two novel classical HL cell lines recently established in our Institution, MDA-V and MDA-E, which have been shown to carry wt-p53 gene. As a control, we used a HL cell line L-428 harboring a mutant p53 (mt-p53) gene product (deletion at exon 4). We investigated effects on apoptosis and cell cycle arrest after treatment of cultured HRS cells with nutlin-3A or a 150-fold less active enantiomere, nutlin-3B. Treatment with nutlin-3A resulted in substantial cell death (up to 65%) in a concentration-dependent manner associated with increased apoptosis as shown by apoptotic morphology (DAPI immunofluorescence), annexin V binding (flow cytometry) and caspase activation (Western blot analysis) in MDA-V and MDA-E cells, but not in L-428 cells. Nutlin-3A-induced apoptotic cell death was accompanied by stabilization of p53 protein as detected by western blot analysis and immunofluorescence and up-regulation of pro-apoptotic Bax, a known target of p53. Inhibition of nuclear export by leptomycin B stabilized p53 at a similar level as compared to nutlin-3A treatment in these cells, suggesting that nutlin-3A stabilized p53 through inhibition of MDM2-mediated degradation of the protein. By contrast, no changes in cell viability, growth or apoptosis were seen after treatment with the inactive nutlin-3B small molecule. Treatment with nutlin-3A also resulted in a significant decrease (up to 85%) of cells in S-phase and a dose-dependent increase of cells in G1 phase of cell cycle as detected by flow cytometry, in MDA-V and MDA-E cells, but not in L-428 cells. Cell cycle arrest was associated with up-regulation of the cyclin-dependent kinase inhibitor p21, a transcriptional target of p53. In contrast, treatment of HRS cells with nutlin-3B had no effects on the cell cycle irrespective of p53 mutation status. Furthermore, combined treatment with nutlin-3A and doxorubicin revealed synergistic effects and enhanced cytotoxicity in HRS cells with wt-p53 gene. Targeting MDM2 with the specific antagonist nutlin-3A that leads to non-genotoxic p53 activation, apoptosis induction and cell cycle inhibition may provide a new therapeutic approach for patients with HL.

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Monoclonal Antibody Against ROR1 in Chronic Lymphocytic Leukemia Cells Induced Apoptosis Via PI3-Kinase/AKT/CREB Pathway

Blood ◽

10.1182/blood.v120.21.1769.1769 ◽

2012 ◽

Vol 120 (21) ◽

pp. 1769-1769

Author(s):

Amir Hossein Daneshmanesh ◽

Mohammad Hojjat-Farsangi ◽

Asa Sandin ◽

Abdul Salam Khan ◽

Ali Moshfegh ◽

...

Keyword(s):

Flow Cytometry ◽

Western Blot ◽

Signaling Pathways ◽

Blot Analysis ◽

Western Blot Analysis ◽

Pi3 Kinase ◽

Signaling Proteins ◽

Downstream Signaling ◽

Induced Apoptosis ◽

Creb Pathway

Abstract Abstract 1769 Background: Phosphoinositide 3-kinase (PI3K)/AKT cascade regulates cell survival, proliferation and differentiation in a variety of cells. In CLL cells PI3K pathway is constitutively activated leading to AKT activation and phosphorylation of cAMP response element-binding protein (CREB). CREB is a transcription factor overexpressed and constitutively phosphorylated in a variety of cancers and seems to have a role in tumor pathobiology. There is a great need to develop novel strategies for targeted therapy in CLL. Monoclonal antibodies (mAbs) specifically targeting leukemic cells might be a rewarding approach. ROR1 is a type I transmembrane receptor tyrosine kinase belonging to one of the twenty families of receptor tyrosine kinases (RTKs). ROR1 is overexpressed on CLL cells but not in white blood cells of healthy donors. ROR1 is constitutively phosphorylated in CLL and siRNA transfection induced apoptosis. We have developed a unique anti-ROR1 mAb directed against CRD (cysteine-rich domain) of the extracellular region of ROR1 capable of inducing direct apoptosis of primary CLL cells. Our anti-CRD mAb induced dephosphorylation of the ROR1 molecule. Aims: To study the apoptotic effect of an anti-ROR1 CRD mAb and effects on downstream signaling pathways involved in CLL, specially the PI3-kinase/AKT/CREB pathway using primary CLL cells. Methods: Using a peptide-based mouse mAb generation method we produced several mAbs against the three extracellular domains of ROR1. In the current study we used one of the best anti-ROR1 antibodies, an anti-CRD mAb raised against the CRD region of ROR1 (Daneshmanesh et al., Leukemia. 2012 Jun;26(6):1348-55). Flow cytometry was used for surface staining of ROR1. Primary CLL cells were incubated with the anti-ROR1 CRD mAb and apoptosis was detected by the MTT assay and Annexin V/propidium iodide (flow cytometry) methods in a 24 h assay. Antibody untreated and treated cell lysates were prepared and subjected to Western blot analysis for identification of signaling molecules involved in apoptosis induced by the anti-ROR1 CRD mAb. We analysed total and phosphorylated levels of the following signaling proteins: AKT, p-AKT, PI3K, p-PI3K, CREB, p-CREB, ERK, p-ERK, PKC and p-PKC. Phosphoproteins were measured before incubation with the mAb and after 20 min-2 h. Results: ROR1 surface expression was detected on 80–85% of the CLL cells. The frequency of apoptotic cells induced by the anti-CRD mAb was in the range of 45–50% which is in accordance with our previous reports (see above). Time kinetics experiments using anti-ROR1 CRD mAb incubated with primary CLL cells revealed dephosphorylation of ROR1 downstream signaling molecules. We analysed the following molecules known to be involved in CLL: PKC, PI3-kinase and ERK1/2. After co-culturing CLL cells with the anti-ROR1 CRD mAb, Western blot analysis showed decreased level of phosphorylated AKT in treated compared to untreated samples. No changes in the phosphorylation levels of ERK1/2 and PKC proteins were seen. Furthermore, we analysed the PI3-kinase protein which is upstream of AKT, and noticed that in CLL cells treated with the anti-ROR1 CRD mAb, the phosphorylation intensity of PI3-kinase p85 isoform has decreased but not p55 isoforrn. Moreover, we also studied the CREB phosphorylation in treated and untreated CLL samples and detected dephosphorylation of CREB in treated as compared to untreated samples. Conclusion: Incubation of CLL cells with an anti-ROR1 CRD mAb induced apoptosis of primary CLL cells. Apoptosis was preceded by dephosphorylation within 2 h of PI3-kinase, AKT and CREB proteins indicating deactivation of these signaling proteins by the anti-ROR1 mab. In untreated CLL cells no effect on phosphorylation of these proteins was noted. Furthermore our ROR1 mAb did not dephosphorylate PKC or ERK. Our data may suggest that activation of CREB molecule might occur via the PI3K/AKT pathway and may be a survival signal in CLL cells associated with the aberrant expression of ROR1. The constitutive phosphorylation of PKC and ERK1/2 seen in CLL might not be related to the overexpression of ROR1. Further studies are warranted for a better understanding of signaling pathways associated with ROR1 and the downstream signaling effects of ROR1 targeting drugs. Disclosures: No relevant conflicts of interest to declare.

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Is Oxidative Stress in Mice Brain Regions Diminished by 2-[(2,6-Dichlorobenzylidene)amino]-5,6-dihydro-4H-cyclopenta[b]thiophene-3-carbonitrile?

Oxidative Medicine and Cellular Longevity ◽

10.1155/2013/194192 ◽

2013 ◽

Vol 2013 ◽

pp. 1-8 ◽

Cited By ~ 2

Author(s):

A. C. Fortes ◽

A. A. C. Almeida ◽

G. A. L. Oliveira ◽

P. S. Santos ◽

W. De Lucca Junior ◽

...

Keyword(s):

Oxidative Stress ◽

Lipid Peroxidation ◽

Superoxide Dismutase ◽

Western Blot ◽

Blot Analysis ◽

Western Blot Analysis ◽

Brain Regions ◽

Saline Control ◽

Control Group ◽

Nitrite Content

2-[(2,6-Dichlorobenzylidene)amino]-5,6-dihydro-4H-cyclopenta[b]thiophene-3-carbonitrile, 5TIO1, is a new 2-aminothiophene derivative with promising pharmacological activities. The aim of this study was to evaluate its antioxidant activity in different areas of mice central nervous system. Male Swiss adult mice were intraperitoneally treated with Tween 80 dissolved in 0.9% saline (control group) and 5TIO1 (0.1, 1, and 10 mg kg−1). Brain homogenates—hippocampus, striatum, frontal cortex, and cerebellum—were obtained after 24 h of observation. Superoxide dismutase and catalase activities, lipid peroxidation and nitrite content were measured using spectrophotometrical methods. To clarify the 5TIO1’s mechanism on oxidative stress, western blot analysis of superoxide dismutase and catalase was also performed. 5TIO1 decreased lipid peroxidation and nitrite content in all brain areas and increased the antioxidant enzymatic activities, specially, in cerebellum. The data of Western blot analysis did not demonstrate evidence of the upregulation of these enzymes after the administration of this compound. Our findings strongly support that 5TIO1 can protect the brain against neuronal damages regularly observed during neuropathologies.

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Acetylshikonin Suppresses Diffuse Large B-Cell Lymphoma Cell Growth By Targeting The TOPK Signaling Pathway

10.21203/rs.3.rs-146077/v1 ◽

2021 ◽

Author(s):

Jieke Cui ◽

Rong Guo ◽

Yingjun Wang ◽

Yue Song ◽

Xuewen Song ◽

...

Keyword(s):

Flow Cytometry ◽

Cell Growth ◽

Western Blot ◽

B Cell ◽

Blot Analysis ◽

Western Blot Analysis ◽

Cell Lymphoma ◽

B Cell Lymphoma ◽

Large B Cell Lymphoma ◽

Large B Cell

Abstract Background: Diffuse large B-cell lymphoma (DLBCL) is one of the most common causes of cancer death worldwide, and responds badly to the existing treatment. Thus, identifying the novel therapeutic targets of DLBCL are urgent. Methods and results: In this study, we found that the T-lymphokine-activated killer cell-originated protein kinase (TOPK) was highly expressed in DLBCL cells and tissues. The TOPK expression were analyzed by bioinformatics analysis, immunohistochemistry (IHC) and western blot analysis. TOPK knockdown inhibited cell growth and induced apoptosis of DLBCL cells with MTS and flow cytometry. Further experiments demonstrated that acetylshikonin, the targeted compound of TOPK, could attenuate the cell growth and aggravate the cell apoptosis through TOPK/extra cellular signal-regulated kinase (ERK)-1/2 signaling using MTS, flow cytometry and western blot analysis. In addition, we demonstrated that TOPK overexpression signiﬁcantly reduced the acetylshikonin effect on cell proliferation and apoptosis in U2932 and OCI-LY8 cells using MTS, flow cytometry and western blot analysis. Conclusions: Taken together, the present study suggests that the targeted inhibition of TOPK by acetylshikonin may be a promising approach to the treatment of DLBCL.

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