Expression profiles of CD11b, galectin-1, beclin-1, and caspase-3 in nasal polyposis*

Abstract The acidic leucine-rich nuclear phosphoprotein 32B (ANP32B, also called APRIL) is a member of a conserved superfamily of nuclear proteins that includes ANP32A/pp32, a factor that binds histones and inhibits their acetylation and regulates cell growth and differentiation in a tissue-specific manner. Recently, ANP32B was identified as a novel histone chaperone, and it can interact with the transcription factor KLF5, leading to transcriptional repression of a KLF5-downstream gene through stimulation of promoter region-specific histone incorporation and inhibition of histone acetylation. Additionally, ANP32B and/or ANP32A also serve as adaptor molecules linking the HuR nucleocytoplasmic shuttle protein and the nuclear export receptor CRM1 to regulate the cytoplasmic accumulation of some transcripts such as c-fos and CD83. However, its biological activity is still poorly understood. By the two-dimensional electrophoresis plus MALDI-TOF/TOF tandem mass spectrometry-based analysis of subcellular protein expression profiles, we identified ANP32B protein to become a small fragment in the cytosols of apoptotic leukemic cell line induced by NSC606985, a camptothecin analog. The ongoing immunoblot analyses confirmed that ANP32B protein was cleaved during cellular context-independent and caspase-3 activation-dependent apoptosis induced by etoposide, doxorubin and arsenic trioxide besides NSC606985. Further in vitro proteolytic experiments supported that ANP32B is a direct substrate of caspase-3, and the site-directed mutagenesis analysis identified the unclassical aspartate (AEVD163) of ANP32B sequence to be the caspase-3 targeted sites. Thus, we investigated the potential role of ANP32B in apoptosis induction. Our results showed that the suppression of ANP32B expression by siRNA in acute myeloid leukemic cell line U937 cells strongly enhances NSC606985 and etoposide-induced apoptosis. Based on these findings, this work also analyzed molecular mechanism of anti-apoptotic effect of ANP32B, and some interesting findings were confirmed.

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Coupling Activation of Pro-Apoptotic Caspases With Autophagy in the Meckel´s Cartilage

Physiological Research ◽

10.33549/physiolres.933947 ◽

2018 ◽

pp. 135-140 ◽

Cited By ~ 1

Author(s):

P. Bíliková ◽

E. Švandová ◽

B. Veselá ◽

J. Doubek ◽

A. Poliard ◽

...

Keyword(s):

Caspase 3 ◽

Beclin 1 ◽

Hypertrophic Chondrocytes ◽

Caspase 8 ◽

Major Mechanism ◽

Histological Sections ◽

Temporary Structure ◽

Caspase 2 ◽

First Time

Mammalian Meckel´s cartilage is a temporary structure associated with mandible development. Notably, its elimination is not executed by apoptosis, and autophagy was suggested as the major mechanism. Simultaneous reports point to pro-apoptotic caspases as novel participants in autophagic pathways in general. The aim of this research was to find out whether activation of pro-apoptotic caspases (-2, -3, -6, -7, -8 and -9) was associated with autophagy of the Meckel´s cartilage chondrocytes. Active caspases were examined in serial histological sections of mouse mandible using immunodetection and were correlated with incidence of autophagy based on Beclin-1 expression. Caspase-2 and caspase-8 were found in Beclin-1 positive regions, whereas caspase-3, -6, -7 and -9 were not present. Caspase-8 was further correlated with Fas/FasL and HIF-1alpha, potential triggers for its activation. Some Fas and FasL positivity was observed in the chondrocytes but caspase-8 activation was found also in FasL deficient cartilage. HIF-1alpha was abundantly present in the hypertrophic chondrocytes. Taken together, caspase-8 activation in the Meckel´s cartilage was demonstrated for the first time. Caspase-8 and caspase-2 were the only pro-apoptotic caspases detected in the Beclin-1 positive segment of the cartilage. Activation of caspase-8 appears FasL/Fas independent but may be switched on by HIF-1alpha.

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Induction of apoptosis and autophagy in T-lymphocytes of patients with Systemic Lupus Erythematosus

Kazan medical journal ◽

10.17816/kmj2020-347 ◽

2020 ◽

Vol 101 (3) ◽

pp. 347-355

Author(s):

Y V Skibo ◽

A R Fathullina ◽

B R Ibragimov ◽

S N Abramov ◽

R R Ismagilova ◽

...

Keyword(s):

Systemic Lupus Erythematosus ◽

T Lymphocytes ◽

Lupus Erythematosus ◽

Peripheral Blood ◽

Caspase 3 ◽

Immunomagnetic Separation ◽

Beclin 1 ◽

Systemic Lupus ◽

Software Environment ◽

Average Value

Aim. To analyze the expression of key apoptosis (Bcl-2, caspase-3) and autophagy (Beclin 1, Vps34, p62 and LC3) proteins regulators in peripheral blood T-lymphocytes of patients with systemic lupus erythematosus. Methods. The object of the study was peripheral blood T-lymphocytes of healthy donors and patients with systemic lupus erythematosus. To obtain T cells, we used the immunomagnetic separation method. Protein expression was analyzed using the Western blot method. Statistically analyzing the results was performed using the R software environment. The data was represented using boxplots. Groups were compared using the MannWhitney test. Results. According to the results of the study of the apoptotic proteins, we found an increased content of caspase-3 and the absence of significant changes in the content of the anti-apoptotic protein Bcl-2 in patients with lupus, which indicates active apoptotic activity. A comparative analysis of Beclin 1 and Vps34 showed their increased content in the cells of patients, which indicates the activation of autophagy. The analysis of two isoforms of LC3 protein revealed their low content in the group of patients. Since the scatter of indicators was very different from the average value, we analyzed these indicators depending on the severity of the disease. In the acute course group, high content of protein LC3-I was detected, the content of form II was lower. In the group with the subacute course, the number of both isoforms is lower than in the other groups. In the group with a chronic course, significant increases of protein LC3-II and a decrease in the ratio of LC3-I/LC3-II were found. Conclusion. The study showed that depending on the severity of systemic lupus erythematosus, the content of protein LC3 isoforms changes, which can be used for differential diagnosis of disease forms.

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Ameliorative Effects of Nano-Selenium Against Fluoride Stress Induced Hepatocytes Autophagy and Apoptosis in Mice

10.21203/rs.3.rs-44205/v1 ◽

2020 ◽

Author(s):

Yajing Wang ◽

Bingxian Liu ◽

Qingyue Han ◽

Khalid Mehmood ◽

Fazul Nabi ◽

...

Keyword(s):

Caspase 3 ◽

Beclin 1 ◽

Protective Effects ◽

Human Beings ◽

Caspase 9 ◽

Nano Material ◽

Cyt C ◽

Hepatocyte Damage

Abstract Background: Fluorine is widespread in the environment, and the injurious impacts of fluoride underscore its significance for public health. The long-term presence of fluorine in environment could be a risk in hepatotoxicity for both human beings and animals. Important role of selenium in mitigation of heavy metal toxicity via regulating autophagy and apoptosis is well-known. Further, nano-Se is a common artificial nano material, with higher biological activity and lower toxicity. The aim of the current study was to examine whether nano-Se supplementation can reduce the effects of fluoride-induced hepatocytes autophagy and apoptosis. Results: Here, we report that fluoride exposure induces apoptosis and autophagy with nucleus broken, dissolved and disappeared of hepatocyte, contributing to its hepatotoxicity. More importantly, Cyt-C and Beclin-1/Bcl-2 pathways are involved in the regulation of autophagy and apoptosis via targeting Caspase-9, Caspase-3, P53, Bax, LC3, ATG-5, P62 and mTOR expressions. Conclusion: Nano-Se is capable to alleviate fluoride-induced hepatocyte damage, that selenium can be prefer to prevent chronic fluorosis-induced autophagy and apoptosis by regulating Cyt-C and Beclin-1/Bcl-2 signaling pathway. In precisely, NaF-induced the liver injury by activating autophagy and apoptosis, which indicate that fluorine exposure, pose an ecological risk to human beings and animals. Nano-Se has protective effects against fluoride-induced hepatocytes.

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FAK and S6K1 Inhibitor, Neferine, Dually Induces Autophagy and Apoptosis in Human Neuroblastoma Cells

Molecules ◽

10.3390/molecules23123110 ◽

2018 ◽

Vol 23 (12) ◽

pp. 3110 ◽

Cited By ~ 12

Author(s):

Dinh-Chuong Pham ◽

Yu-Chuan Chang ◽

Shian-Ren Lin ◽

Yuh-Ming Fuh ◽

May-Jywan Tsai ◽

...

Keyword(s):

Cell Migration ◽

Cancer Cells ◽

Molecular Mechanisms ◽

Caspase 3 ◽

Neuroblastoma Cells ◽

Beclin 1 ◽

Potential Candidate ◽

Human Neuroblastoma Cells ◽

Human Neuroblastoma ◽

Anti Cancer

Human neuroblastoma cancer is the most typical extracranial solid tumor. Yet, new remedial treatment therapies are demanded to overcome its sluggish survival rate. Neferine, isolated from the lotus embryos, inhibits the proliferation of various cancer cells. This study aimed to evaluate the anti-cancer activity of neferine in IMR32 human neuroblastoma cells and to expose the concealable molecular mechanisms. IMR32 cells were treated with different concentrations of neferine, followed by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay to assess cell viability. In an effort to determine the molecular mechanisms in neferine-incubated IMR32 cells, cell cycle arrest, cell migration, and focal adhesion kinase (FAK), the 70-kDa ribosomal S6 kinase 1 (S6K1), poly (ADP-ribose) polymerase (PARP), caspase-3, Beclin-1, and microtubule-associated protein 1A/1B-light chain 3 (LC3) protein expressions were investigated. Neferine strongly disrupted the neuroblastoma cell growth via induction of G2/M phase arrest. Furthermore, neferine provoked autophagy and apoptosis in IMR32 cells, confirmed by p-FAK, and p-S6K1 reduction, LC3-II accumulation, Beclin-1 overexpression, and cleaved caspase-3/PARP improvement. Finally, neferine markedly retarded cell migration of neuroblastoma cancer cells. As a result, our findings for the first time showed an explicit anti-cancer effect of neferine in IMR32 cells, suggesting that neferine might be a potential candidate against human neuroblastoma cells to improve clinical outcomes with further in vivo investigation.

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Abstract 43: SIRT1 Enhances Oxidized Low-density Lipoprotein Induced Autophagy in Human Valve Interstitial Cells

Circulation Research ◽

10.1161/res.119.suppl_1.43 ◽

2016 ◽

Vol 119 (suppl_1) ◽

Author(s):

Huadong Li ◽

Jiawei Shi ◽

Weihua Qiao ◽

Nianguo Dong ◽

Gangjian Qin

Keyword(s):

Aortic Valve ◽

Heart Valve ◽

Western Blotting ◽

Caspase 3 ◽

Beclin 1 ◽

Calcium Deposition ◽

Control Group ◽

Ros Generation ◽

Aortic Valves ◽

Over Expression

Objectives: Sirt1, an NAD + -dependent class III deacetylase, has been shown to regulate autophagy during cardiac remodeling. Its role in calcific aortic valve disease has not been studied. Methods: Aortic valve samples were collected from patients with calcific aortic valve diseases complicated with hyper-lipidemia (at the time of heart valve replacement) (CAVD group), and from patients with dilated cardiomyopathy with non-calcified valves and a normal blood lipid profile (at the time of heart transplantation) (control group); the expression and distribution of calcification and Sirt1 in the aortic valves were analyzed by immuno-histochemical staining and Western blotting. In addition, primary human valvular intersititial cells (VICs) were isolated from normal valves in control group, and their response to DiI-ox-LDL treatments was analyzed in the presence of adenovirus-mediated Sirt1 over-expression (Ad-SIRT1) and Sirt1 inhibitor EX527, followed by assessments of the levels of reactive oxygen species (ROS), autophagy, apoptosis and calcification with biochemical methods. Results: The H&E, Masson, Von Kossa and α-SMA immuno-fluorescence staining showed that the aortic valves in CAVD group were thickened by excessive elastin, collagen expression and calcium deposition, which was associated with an elevated expression of osteocalcin and a reduced expression of Sirt1, as compared to control group. In vitro, the primary VICs were incubated with different concentrations (25, 50 and 100μg/mL) of ox-LDL for 48 h. ROS levels increased in an ox-LDL dose dependent manner. In addition, ox-LDL treatment led to a slight increase in the level of LC3 and Beclin-1, autophagic death-related hMOF and acetyl-histone H4, and a significant decrease in the level of Sirt1 through Western blotting. Interestingly, co-treatment of VICs with EX527 further increased ROS generation and expression of Caspase 3, Bax, and β-Catenin, while Sirt1 over-expression by Ad-SIRT1 resulted in increased expression of LC3 and Beclin-1, decreased expression of Caspase 3, Bax, Wnt3a and β-Catenin and reduced ROS generation. Conclusion: Our data suggest that Sirt1 may play a protective role in ox-LDL-induced VIC autophagy and heart valve calcification.

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Mechanism of Action of MK-0457 Efficacy in Monotherapy and in Combination with Vorinostat in Wild Type and T315I Mutant BCR/ABL Positive Leukemia Cells.

Blood ◽

10.1182/blood.v110.11.1051.1051 ◽

2007 ◽

Vol 110 (11) ◽

pp. 1051-1051 ◽

Cited By ~ 6

Author(s):

Seiichi Okabe ◽

Tetsuzo Tauchi ◽

Junko H. Ohyashiki ◽

Kazuma Ohyashiki

Keyword(s):

Gene Expression ◽

Intracellular Signaling ◽

Caspase 3 ◽

Expression Profiles ◽

Gene Expression Profiles ◽

Leukemia Cells ◽

Wild Type ◽

Imatinib Resistance ◽

T315i Mutation ◽

Induced Apoptosis

Abstract Mutation in the ABL kinase domain is the principal mechanism of imatinib resistance in patients with BCR/ABL-positive leukemia. MK-0457 is a small molecule inhibitor of the Aurora kinase family with potent cross-reactivity towards wild type and T315I BCR/ABL and FLT-3, and the compound has shown activity in preclinical models of solid tumors and FLT-3 driven leukemia. MK-0457 is presently being evaluated in a clinical trial in chronic myelogenous leukemia (CML) patients with imatinib resistance such as T315I mutation, but the mechanism of MK-0457 is not fully evaluated. In this study, the gene expression profiles of CML cell line, K562, exposed to imatinib or MK-0457 were analyzed and compared. When their gene expression profiles were compared, 937 genes in imatinib and 895 genes in MK-0457 were increased and 625 genes were overlapped. In contrast, 597 genes in imatinib and 537 genes in MK-0457 were decreased and 396 genes were overlapped. These down regulated genes included heat shock proteins (HSPs) suggesting these results may relate to the BCR/ABL stability. Next, we examined the intracellular signaling of MK-0457 in BCR/ABL positive cells with T315I mutation by using BAF-3 p185 BCR/ABL (p185) and BAF-3 BCR/ABL T315I (T315I) cell lines. MK-0457 potently induced apoptosis of p185 cells and T315I cells in 72 hours treatment. IC50 of MK-0457 was 100 nM (p185) and 250 nM (T315I). We found that caspase 3, and poly (ADP-ribose) polymerase (PARP) were activated and BCR/ABL phosphorylation was reduced after MK-0457 treatment in a dose dependent manner. It has been reported that histone deacetylase inhibitor (e.g., vorinostat) also depleted BCR/ABL, as well as induced apoptosis and sensitized BCR/ABL-expressing leukemia cells to apoptosis induced by imatinib. These data suggest that MK-0457 may be synergistic with histone deacetylase inhibitors in light of recent publications that have shown that vorinostat induces acetylation of the heat shock protein (HSP), depletes the HPS90 client proteins, including BCR/ABL, and enhances the imatinib-induced apoptosis in BCR/ABL-expressing cells. We found that combination of MK-0457 and vorinostat synergistically increased the apoptosis of p185 cells and T315I cells in 72 hours treatment. Caspase 3 and PARP activation were also synergistically increased after vorinostat and MK-0457 treatment. We examined the intracellular signaling by using these cell lines. Phosphorylation of BCR/ABL, signal-transducing activators of transcription 5 (STAT5), one of the src kinase, Lyn, Crk-L were reduced after treatment with MK-0457 + vorinostat. We also found that cyclin D1 was decreased after MK-0457 or vorinostat and MK-0457 treatment. Histone H4 is also acetylated after vorinostat treatment, but phosphorylation of BCR/ABL was not reduced directly. We evaluated the activity of MK-0457 and vorinostat in primary Ph positive acute lymphoblastic leukemia cells with the T315I mutation. We found that MK-0457 potently induced cell growth inhibition of primary T315I cells in 72 hours treatment. Moreover, combination of vorinostat and MK-0457 synergistically increased the extent of apoptosis in primary T315I cells. This study provides further insight into the mechanism of action of MK-0457 in wild type and T315I mutant BCR-ABL cells and provides mechanistic rational for the synergistic induction of apoptosis observed for the combination of MK-0457 + vorinostat.

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