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

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

<b><i>Background:</i></b> 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. <b><i>Methods:</i></b> 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. <b><i>Results:</i></b> 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. <b><i>Conclusion:</i></b> Collectively, our study is first to demonstrate the protective effect of RMZ against experimentally induced TBI in rats.

scholarly journals Attenuation of astrogliosis and modulation of endothelial growth factor receptor in lipid rafts by simvastatin after traumatic brain injury

2010 ◽  
Vol 113 (3) ◽  
pp. 591-597 ◽  
Author(s):  
Hongtao Wu ◽  
Asim Mahmood ◽  
Dunyue Lu ◽  
Hao Jiang ◽  
Ye Xiong ◽  
...  
Keyword(s):  
Traumatic Brain Injury ◽  
Brain Injury ◽  
Growth Factor ◽  
Western Blot ◽  
Lipid Rafts ◽  
Blot Analysis ◽  
Western Blot Analysis ◽  
Growth Factor Receptor ◽  
Astrocyte Activation ◽  
Caveolin 1

Object The authors' previous studies have demonstrated that simvastatin treatment promotes neuronal survival and reduces inflammatory cytokine release from astrocytes after traumatic brain injury (TBI) in rats. Since reactive astrocytes produce inflammation mediators, in the current study the authors investigated the effect of simvastatin on astrocyte activation after TBI and its underlying signaling mechanisms. Methods Saline or simvastatin (1 mg/kg) was orally administered to rats starting at Day 1 after TBI and then daily for 14 days. Rats were killed at 1, 3, 7, and 14 days after treatment. Brain sections and tissues were prepared for immunohistochemical staining and Western blot analysis, respectively. Cultured astrocytes were subjected to oxygen-glucose deprivation (OGD) and followed by immunocytochemical staining with glial fibrillary acidic protein/caveolin-1 and Western blot analysis. Lipid rafts were isolated from the cell lysate and Western blotting was carried out to detect the changes in epidermal growth factor receptor (EGFR) expression and phosphorylation in the lipid rafts. Results Simvastatin significantly promoted neuronal survival after TBI and attenuated activation of astrocytes. Simvastatin modified the caveolin-1 expression in lipid rafts in astrocyte cell membrane, suppressed the phosphorylation of EGFR in lipid rafts of astrocytes after OGD, and inhibited the OGD-induced interleukin-1 production. Conclusions These data suggest that simvastatin reduces reactive astrogliosis and rescues neuronal cells after TBI. These beneficial effects of simvastatin may be mediated by inhibiting astrocyte activation after TBI through modifying the caveolin-1 expression in lipid rafts and the subsequent modulation of EGFR phosphorylation in lipid rafts.

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

2018 ◽  
Vol 50 (2) ◽  
pp. 460-472 ◽  
Author(s):  
Tong Niu ◽  
Liuzhong Jin ◽  
Shizhen Niu ◽  
Cunqi Gong ◽  
Hui Wang
Keyword(s):  
Oxidative Stress ◽  
Flow Cytometry ◽  
Western Blot ◽  
Cell Viability ◽  
Blot Analysis ◽  
Western Blot Analysis ◽  
Akt Pathway ◽  
Lycium Barbarum ◽  
Cord Injury ◽  
Lycium Barbarum Polysaccharides

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.

scholarly journals Protective Effect of Diphenhydramine against Traumatic Brain Injury in Rats via Modulation of Oxidative Stress and Inflammation

Pharmacology ◽  
2019 ◽  
Vol 105 (1-2) ◽  
pp. 47-53 ◽  
Author(s):  
Wenyong Pan ◽  
Zhigang Cao ◽  
Dongyang Liu ◽  
Yingbin Jiao
Keyword(s):  
Oxidative Stress ◽  
Traumatic Brain Injury ◽  
Brain Injury ◽  
Protective Effect ◽  
Neuronal Degeneration ◽  
Neuroprotective Effect ◽  
B Cell Lymphoma ◽  
Treated Group ◽  
Dependent Manner ◽  
And Oxidative Stress

Background: Traumatic brain injury (TBI) is considered a major burden across the globe affecting both individuals and their families. Therefore, the present study was conducted to determine the protective effect of diphenhydramine (DPM) against TBI in experimental rats. Methods: The effect of DPM was evaluated on the cerebral edema (CE) and neuronal degeneration after the induction of experimental brain injury in rats. The effect of DPM was also investigated on the inflammatory cytokines, for example, tumor necrosis factor-α and interleukin 1β and oxidative stress markers, such as malondialdehyde, superoxide dismutase, and glutathione peroxidase. Western blot analysis was used to investigate the effect of DPM on B-cell lymphoma 2 (Bcl-2), Bcl-2-associated X protein (Bax) and cleaved caspase-3. Results: Results of the study suggest that DPM causes reduction in CE and prevents neuronal degeneration. It also causes reduction in inflammation and oxidative stress in a dose-dependent manner. The level of Bax was found to be elevated, together with reduction in the Bcl-2 level in the DPM-treated group. Conclusion: DPM exerts a neuroprotective effect after TBI via the attenuation of oxidative stress, inflammation, and mitochondrial apoptosis pathways.

Study on the extract of Huangqi and Ezhu for the crosstalk of exosomes mediating tamoxifen resistance on LCC9 cells.

2021 ◽  
Vol 39 (15_suppl) ◽  
pp. e12517-e12517
Author(s):  
Shasha Cui ◽  
Dongni Zhang ◽  
Wenping Lu
Keyword(s):  
Breast Cancer ◽  
Stem Cells ◽  
Flow Cytometry ◽  
Cancer Stem Cells ◽  
Western Blot ◽  
Blot Analysis ◽  
Western Blot Analysis ◽  
Tamoxifen Resistance ◽  
Negative Control ◽  
Flow Cytometry Analysis

e12517 Background: Estrogen receptor (ER)-positive breast cancer is the most frequent subtype accounting for about 70% of all cases. Tamoxifen (TAM) is still the most effective drug for treating ER+/PR+ breast cancer. However, recurrence and metastasis still occur in 30-50% of patients due to TAM resistance. Furthermore, studies have shown that breast cancer stem cells (BCSCs) play an important role in TAM resistance, and exosomes regulate dynamic changes of BCSCs in tumor microenvironment. Additionally, the extract of Huangqi and Ezhu (EHE) has been shown to inhibit multiple TAM-associated resistance signaling pathways. In the current study, we investigated the effect of EHE to reverse TAM-resistance in LCC9 cells via crosstalk of distinct types of exosomes. Methods: Exosomes were extracted from LCC9 cells by ultracentrifugation and characterized by particle size detection, transmission electron microscopy and western blot analysis. Subsequently, isolated LCC9 exosomes were used to treat bone marrow stem cells (BMSCs) with EHE only, TAM only, EHE and TAM (EHETAM), and no treatment (negative control, NC). After treatment, exosomes of BMSCs were isolated and incubated further with LCC9 cells. CCK-8 assay and flow cytometry analysis were performed, and protein expression in treated LCC9 cells was tested by western blot analysis. Results: Data from CCK-8 assay showed that compared with the NC group, proliferation of LCC9 cells was significantly reduced in the presence of exosomes of BMSCs treated with TAM (82.9%, 77.1% , 75.4%), EHE (73.2 %, 69.5% , 68.2 %) and EHETAM (68.9 %, 52.3%，54.1% ) after 24h, 48h, and 72 h, respectively (p < 0.001). EHE and EHETAM groups have more effective inhibition on cell proliferation than TAM only group on the different time separately(p < 0.001).Flow cytometry analysis demonstrated that compared with the NC group(6.1%), increased apoptosis was observed for groups of TAM (8.1%), EHE (12.5%) and EHETAM (12.2%) (p < 0.001). Compared with TAM only group, EHE and EHETAM groups also have significantly higher cell death rate (p < 0.001). Lower CD44 and HER2 expression and higher ERα expression were observed in EHE, TAM, and EHETAM than those of NC group by western blot analysis of treated LCC9 cells. Conclusions: EHE treatment can reverse TAM-resistance by interfering with the crosstalk between BMSCs and breast cancer cells exosomes to induce more active apoptosis and reduce TAM-resistant CD44+ cancer stem cells.

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

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.

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

Blood ◽  
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.

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

Blood ◽  
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.

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

2013 ◽  
Vol 2013 ◽  
pp. 1-8 ◽  
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.

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

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.

scholarly journals Dimethyl Fumarate Ameliorates Nucleus Pulposus Cell Dysfunction through Activating the Nrf2/HO-1 Pathway in Intervertebral Disc Degeneration

2021 ◽  
Vol 2021 ◽  
pp. 1-13
Author(s):  
Ruihong Wang ◽  
Dawei Luo ◽  
Zhiwei Li ◽  
Huimin Han
Keyword(s):  
Oxidative Stress ◽  
Intervertebral Disc ◽  
Western Blot ◽  
Er Stress ◽  
Nucleus Pulposus ◽  
Blot Analysis ◽  
Western Blot Analysis ◽  
Dimethyl Fumarate ◽  
Matrix Composition ◽  
Type I

Background. Oxidative stress, inflammation, and nucleus pulposus cells (NPCs) apoptosis are involved in pathogenesis of intervertebral disc (IVD) degeneration (IVDD). Dimethyl fumarate (DMF) has been found to effectively depress oxidative stress and inflammation via the Nrf2 pathway. Hence, this project was designed to explore the underlying mechanisms of how DMF protects NPCs from damage by LPS challenge. Methods and Results. CCK8 assay and flow cytometry of apoptosis indicated that DMF treatment attenuated LPS-induced NPC damage. Western blot analysis demonstrated that DMF enhanced the expressions of nuclear factor-erythroid 2-related factor 2 (Nrf2) and heme oxygenase-1 (HO-1) in LPS-challenged NPCs. DMF treatment significantly decreased the accumulation of ROS, downregulated inflammatory cytokines (p-NF-κB, IL-1β, and TNF-α), and ER stress-associated apoptosis proteins (Bip, calpain-1, caspase-12, caspase-3, and Bax) in LPS-challenged NPCs. The level of antiapoptotic protein Bcl-2 was promoted by DMF treatment in LPS-challenged NPCs. Glutathione (GSH) assay showed that DMF treatment improved reduced to oxidized glutathione ratio in LPS-challenged NPCs. Furthermore, the results of western blot analysis indicated that in LPS-challenged NPCs, DMF treatment ameliorated the elevated levels of matrix degradation enzymes (MMP-13, aggrecanase 1) and type I collagen and the reduced levels of matrix composition (type II collagen and ACAN). However, Nrf2 knockdown abolished these protective effects of DMF. Conclusion. Our data suggested that treatment with DMF mitigated LPS-induced oxidative stress, inflammation, and ER stress-associated apoptosis in NPCs via the Nrf2/HO-1 signaling pathway, thus reliving LPS-induced dysfunction of NPCs, which offered a novel potential pharmacological treatment strategy for IVDD.

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