scholarly journals Translational control of Bcl-2 promotes apoptosis of gastric carcinoma cells

2019 ◽  
Author(s):  
Shuangfen Tao ◽  
Jianchun Gu ◽  
Qing Wang ◽  
Leizhen Zheng
Keyword(s):  
Gastric Carcinoma ◽  
Cell Survival ◽  
Translational Control ◽  
Annexin V ◽  
Luciferase Reporter ◽  
Bioinformatics Analyses ◽  
Apoptotic Protein ◽  
Detection Assay ◽  
Apoptosis Detection ◽  
Tumor Region

AbstractAberrantly expressed microRNAs (miRNAs) are essential for the tumorigenesis of gastric carcinoma (GC). Nevertheless, an effect of miR-383 on GC cells apoptosis has not been acknowledged previously. Here, we investigated the miR-383 level and anti-apoptotic protein Bcl-2 level in specimens of GC. Bioinformatics analyses was performed, and assay of luciferase-reporter was used for analyzing the relationship between Bcl-2 and miR-383. We analyzed survival of GC cells upon Fluorouracil treatment in an assay of CCK, and measured apoptosis of cells using flow cytometric FITC Annexin V apoptosis detection assay. The level of miR-383 was found extremely lower while the level of Bcl-2 levels was found extremely higher in GC specimens, in comparison with tissue from the adjacent non-tumor region. Additionally, the miR-383 level and Bcl-2 level inversely correlated in specimens of GC. In comparison with miR-383-high subjects, MiR-383-low subjects showed overall lower survival. Bioinformatics analyses revealed that miR-383 targeted the 3’-UTR of Bcl-2 mRNA to restrain its translation, demonstrated in a luciferase reporter assay. MiR-383 overexpression inhibited the Bcl-2-mediated survival of cell against apoptosis induced by Fluorouracil, while miR-383 depletion enhanced the cell survival. Together, these data indicate that suppression of miR-383 in GC improves the Bcl-2-mediated cell survival of GC against the chemotherapy-induced cell death. MiR-383 re-expression in cells of GC might augment apoptosis of GC cells during chemotherapy.

scholarly journals Translational control of Bcl-2 promotes apoptosis of gastric carcinoma cells

BMC Cancer ◽  
2021 ◽  
Vol 21 (1) ◽  
Author(s):  
Shuangfen Tao ◽  
Jianchun Gu ◽  
Qing Wang ◽  
Leizhen Zheng
Keyword(s):  
Gastric Carcinoma ◽  
Translational Control ◽  
Annexin V ◽  
Protein Translation ◽  
Luciferase Reporter ◽  
Bioinformatics Analyses ◽  
Detection Assay ◽  
Associated Proteins ◽  
Apoptosis Detection ◽  
Tumor Region

Abstract Background Anti-apoptotic protein Bcl-2 plays a substantial role in the carcinogenesis, whereas the regulation for Bcl-2 in gastric carcinoma (GC) is poorly understood. Specifically, a role of microRNA (miR)-383 in the control of Bcl-2 has not been shown in GC and thus addressed in the current study. Methods We investigated the levels of miR-383 and Bcl-2 in 50 GC specimens, and compared them with patients’ clinical characteristics. Bioinformatics analyses and luciferase-reporter assay were applied for analyzing the relationship between Bcl-2 and miR-383. An CCK assay was used to determine the survival of Fluorouracil-treated GC cells, and apoptosis of GC cells was assessed by flow cytometric FITC Annexin V apoptosis detection assay and expression of apoptosis-associated proteins. Results The levels of miR-383 were lower while the levels of Bcl-2 levels were higher in GC specimens, compared to tissue from the adjacent non-tumor region. Low miR-383 and high Bcl-2 seemed to be associated with high malignancy and metastasis. In GC specimens, the levels of Bcl-2 and miR-383 inversely correlated. The overall survival of miR-383-low cases was poorer. Mechanistically, miR-383 targeted the 3′-UTR of Bcl-2 mRNA to inhibit its protein translation. Overexpression of miR-383 downregulated Bcl-2, resulting in reduced survival of Fluorouracil-treated GC cells. Similar conclusion was drawn through analysis of published database. Conclusion MiR-383 reduces survival of Fluorouracil-treated GC cells through downregulating of Bcl-2.

scholarly journals MiR-429 Induces Gastric Carcinoma Cell Apoptosis Through Bcl-2

2015 ◽  
Vol 37 (4) ◽  
pp. 1572-1580 ◽  
Author(s):  
Ping Zhu ◽  
Jingping Zhang ◽  
Jianfei Zhu ◽  
Jun Shi ◽  
Qiuwei Zhu ◽  
...  
Keyword(s):  
Gastric Carcinoma ◽  
Cell Survival ◽  
Cell Apoptosis ◽  
Annexin V ◽  
Luciferase Reporter Assay ◽  
Luciferase Reporter ◽  
Reporter Assay ◽  
Bioinformatics Analyses ◽  
Gastric Carcinoma Cell ◽  
Underlying Mechanisms

Background/Aims: MicroRNAs (miRNAs) play an essential role in the tumorigenesis of gastric carcinoma (GC). MiR-429 has been recently reported to inhibit GC growth, but the underlying mechanisms are not clear. Methods: Here, we studied the levels of miR-429 and anti-apoptotic protein Bcl-2 in GC specimens. We performed bioinformatics analyses and used luciferase-reporter assay to analyze the relationship between miR-429 and Bcl-2 in GC cells. Cell survival upon Fluorouracil treatment was analyzed in a CCK assay. Cell apoptosis was measured by flow cytometry based FITC Annexin V apoptosis detection assay. Results: MiR-429 levels were significantly decreased and Bcl-2 levels were significantly increased in GC specimens, compared to the paired adjacent non-tumor gastric tissue. Moreover, the levels of miR-429 and Bcl-2 inversely correlated in GC specimens. MiR-429-low subjects had an overall inferior survival, compared to miR-429-high subjects. Bioinformatics analyses showed that miR-429 targeted the 3'-UTR of Bcl-2 mRNA to inhibit its translation, which was confirmed by luciferase-reporter assay. Overexpression of miR-429 inhibited Bcl-2-mediated cell survival against apoptosis induced by Fluorouracil, while depletion of miR-429 augmented it. Conclusion: Our data suggest that miR-429 suppression in GC promotes Bcl-2-mediated cancer cell survival against chemotherapy-induced cell death. Re-expression of miR-429 levels in GC cells may enhance cancer apoptosis during chemotherapy.

Targeting apoptosis and autophagy by a novel bcl-2 inhibitor, GX15-070, in neuroblastoma.

2013 ◽  
Vol 31 (15_suppl) ◽  
pp. 10048-10048
Author(s):  
Herve Sartelet ◽  
Sonia Cournoyer ◽  
Anissa Addioui ◽  
Assila Belounis ◽  
Mona Beaunoyer ◽  
...  
Keyword(s):  
Cell Death ◽  
Cell Survival ◽  
Annexin V ◽  
High Sensitivity ◽  
Western Blots ◽  
Pediatric Tumor ◽  
Nsg Mice ◽  
Apoptotic Protein

10048 Background: Neuroblastoma (NB) is a frequent pediatric tumor with poor prognosis. The disregulation of the anti-apoptotic protein Bcl-2 is crucial for the tumoral development and chemoresistance. Autophagy is also implicated in tumor cell survival and chemoresistance. The aim of our study was to demonstrate the in vitro and in vivo therapeutic efficiency of GX 15-070, a Bcl-2 inhibitor, used alone and in combination with conventional drugs used in the treatment of NB and hydroxychloroquine (HCQ), a known autophagy inhibitor. Methods: Using 6 NB cell lines, cell viability (MTT) assays were done at progressively increased concentrations of GX 15-070 alone or in combination with cisplatin or with Z-VAD-FMK, a broad-spectrum caspase inhibitor. Apoptosis was tested by evaluating the cleavage of caspase 3 by western blots (WB) and the Annexin V/7-AAD staining studied by FACS. To assess if autophagy was modified by GX 15-070, the cleavage of LC3 protein was tested by WB and cell survival were tested with combination of GX 15-070 and HCQ. To verify the anti-tumor activity in vivo of GX 15-070, orthotopic injections were made on NSG mice treated with GX 15-070 alone and in combination with HCQ. Results: It was observed a high sensitivity of the NB cells to GX 15-070 with increase of cell death and a potential synergistic of this molecule when it’s combined with cisplatin or HCQ. This cell death was due to apoptosis and may also be inhibited by Z-VAD-FMK. GX 15-070 alone or associated to cisplatin increased the autophagy. The in vivo study showed that GX 15-070 treatment used alone or in combination with HCQ significantly decreased the size of the tumor. Conclusions: Our results support the interest of GX 15-070 in the treatment of NB alone or in combination with classical drugs. Our studies also support that activation of apoptosis associated with inhibition of autophagy have a synergistic potential against tumoral progression and must have to be considered in further mechanistic studies for the optimization of more efficient combined therapies in the treatment of NB.

Propranolol Inhibits the Growth of Cervical Cancer Cells by Inhibiting Cyclic Guanosine Monophosphate/Protein Kinase G (cGMP/PKG) Pathway

2022 ◽  
Vol 12 (2) ◽  
pp. 422-426
Author(s):  
Mi Li ◽  
Yanqin Ji
Keyword(s):  
Cervical Cancer ◽  
Annexin V ◽  
Cyclic Guanosine Monophosphate ◽  
Siha Cell ◽  
Guanosine Monophosphate ◽  
Dependent Manner ◽  
Cell Vitality ◽  
Detection Assay ◽  
Cervical Cancer Cells ◽  
Apoptosis Detection

This study assesses the therapeutic effect of propranolol on cervical cancer and its mechanism. Propranolol’s effect on cervical cancer was evaluated by MTT, Western blotting, flow cytometry and colony formation. By searching Drug Bank and String, cGMP/PKG signaling might be downstream targets of propranolol for subsequent analysis. Our results found that propranolol could significantly inhibit Hela and SiHA cell vitality and clone formation in a dose dependent manner. Further, Annexin V-PE/7-AAD Apoptosis Detection assay showed that propranolol could increase Hela and SiHA cell apoptosis. Finally, propranolol attenuated the phosphorylation level of VASP at Ser239 which is critical for PKG activation. In conclusion, propranolol suppressed cervical cancer cell proliferation via inhibition of cGMP/PKG signaling, which provides an affordable and effective method for cervical cancer remedy.

Low Affinity Interaction with CLL B-Cell Receptors Induces Apoptosis by Down-Regulating Mcl-1 and Up-Regulating BimEL.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 1115-1115
Author(s):  
Santanu Paul ◽  
Rasmi Abu-Helu ◽  
Patricia Mongini ◽  
Christine N. Metz ◽  
Charles C. Chu ◽  
...  
Keyword(s):  
B Cell ◽  
Cell Survival ◽  
Binding Sites ◽  
Annexin V ◽  
Parp Cleavage ◽  
Antigen Binding ◽  
Cellular Interactions ◽  
Apoptotic Protein ◽  
Survival And Growth

Abstract The B-cell receptor (BCR) engages antigens and triggers divergent signaling pathways, inducing survival, proliferation, and differentiation or anergy and apoptosis. The decision as to which pathway is activated and which functional outcome ensues is regulated by a series of receptor and cellular interactions. In particular, signal transduction via the BCR is influenced by the affinity of receptor-ligand binding and by the number of antigen-binding sites engaged. To address the role of BCR-antigen-binding affinity and valency in CLL, we used a series of anti-μ chain mAbs that differ in their affinities for the ligand as surrogates of antigen binding to the BCR (HB57: Ka = 5x108, “high”; Mu53: Ka = 2x107, “intermediate”‘ and P24: Ka = ∼2x106 M−1, “low”). We also mimicked the valency of the surrogate antigens (and hence the number of binding sites occupied) by preparing these mAbs in either their native dimeric state or in multimeric forms by conjugating them to dextran and to DYNA beads M-450. The native mAbs and the mAbs bound to dextran are soluble; the mAbs bound to beads are insoluble. The functional effects on CLL B cells of each mAb, in all three forms (i.e., soluble dimeric, soluble multimeric on dextran, and insoluble multimeric on beads), were then assessed in vitro. We noted striking and consistent differences in CLL cell responses when the mAb of different affinities for the BCR were used in the soluble, native dimeric state. Whereas the higher affinity anti-μ mAbs (HB57 and Mu53) induced minimal to modest proliferation, the low affinity mAb P24 drastically reduced proliferation. Similarly the soluble, dimeric forms of the same three mAbs had striking and consistently different effects on CLL cell survival. In particular, the low affinity mAb induced significant apoptosis as measured by Annexin V/propidium iodide immunofluorescent analyses (mean 61.2%; range 32–99.2%) as well as by PARP cleavage. Western blotting studies of cell lysates from CLL cells stimulated with the low affinity mAb vs. the higher affinity mAbs or isotype matched control mAbs revealed downregulation of the anti-apoptotic protein Mcl-1 and upregulation of the pro-apoptotic protein BimEL. The multimeric forms of the three anti-μ mAbs, both soluble and insoluble, increased CLL cell survival and proliferation, irrespective of affinities, by maintaining Mcl-1 levels and failing to upregulate Bim. This study illustrates the importance of the “quality” of BCR-antigen interaction for CLL cell survival and growth. Soluble low affinity interactions with antigens of reduced valency lead to CLL cell death, whereas soluble low affinity interactions with multivalent antigens lead to cell survival and growth. The use of soluble, monomeric, low affinity ligands reactive with shared stereotypic BCRs, therefore, should be considered as an immunotherapeutic approach in CLL.

scholarly journals Endothelial Cell Autophagy in Atherosclerosis is Regulated by miR-30-Mediated Translational Control of ATG6

2015 ◽  
Vol 37 (4) ◽  
pp. 1369-1378 ◽  
Author(s):  
Tao Zhang ◽  
Feng Tian ◽  
Jing Wang ◽  
Jing Jing ◽  
Shan-Shan Zhou ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Endothelial Cell ◽  
Translational Control ◽  
Cell Injury ◽  
Protein Translation ◽  
Luciferase Reporter ◽  
Protective Effects ◽  
Bioinformatics Analyses

Background/Aims: Endothelial cell injury and subsequent death play an essential role in the pathogenesis of atherosclerosis. Autophagy of endothelial cells has a protective role against development of atherosclerosis, whereas the molecular regulation of endothelial cell autophagy is unclear. MicroRNA-30 (miR-30) is a known autophagy suppressor in some biological processes, while it is unknown whether this regulatory axis may be similarly involved in the development of atherosclerosis. Here, we aimed to answer these questions in the current study. Methods: We examined the levels of endothelial cell autophagy in ApoE (-/-) mice suppled with high-fat diet (HFD), a mouse model for atherosclerosis (simplified as HFD mice). We analyzed the levels of autophagy-associated protein 6 (ATG6, or Beclin-1) and the levels of miR-30 in the purified CD31+ endothelial cells from mouse aorta. Prediction of the binding between miR-30 and 3'-UTR of ATG6 mRNA was performed by bioinformatics analyses and confirmed by a dual luciferase reporter assay. The effects of miR-30 were further analyzed in an in vitro model using oxidized low-density lipoprotein (ox-LDL)-treated human aortic endothelial cells (HAECs). Results: HFD mice developed atherosclerosis in 12 weeks, while the control ApoE (-/-) mice that had received normal diet (simplified as NOR mice) did not. Compared to NOR mice, HFD mice had significantly lower levels of endothelial cell autophagy, resulting from decreases in ATG6 protein, but not mRNA. The decreases in ATG6 in endothelial cells were due to HFD-induced increases in miR-30, which suppressed the translation of ATG6 mRNA via 3′-UTR binding. These in vivo findings were reproduced in vitro on ox-LDL-treated HAECs. Conclusion: Upregulation of miR-30 by HFD may impair the protective effects of endothelial cell autophagy against development of atherosclerosis through suppressing protein translation of ATG6.

scholarly journals miR-23b Negatively Regulates Sepsis-Induced Inflammatory Responses by Targeting ADAM10 in Human THP-1 Monocytes

2019 ◽  
Vol 2019 ◽  
pp. 1-13 ◽  
Author(s):  
Wenying Zhang ◽  
Furong Lu ◽  
Yuliu Xie ◽  
Yao Lin ◽  
Tian Zhao ◽  
...  
Keyword(s):  
Western Blot ◽  
Inflammatory Cytokines ◽  
Mononuclear Cells ◽  
Inflammatory Responses ◽  
Annexin V ◽  
Luciferase Reporter ◽  
Inflammatory Factor ◽  
Bioinformatics Analyses ◽  
Peripheral Blood Mononuclear ◽  
Adam10 Expression

Background. Previous studies have demonstrated pivotal roles of disintegrin and metalloproteinase 10 (ADAM10) in the pathogenesis of sepsis. MicroRNA- (miR-) 23b has emerged as an anti-inflammatory factor that prevents multiple autoimmune diseases. However, the underlying mechanisms of miR-23b in the regulation of ADAM10 and sepsis remain uncharacterized. Methods. The expression levels of ADAM10 and miR-23b were detected by quantitative RT-PCR and western blot analysis. Cytokine production and THP-1 cell apoptosis were measured by enzyme-linked immunosorbent and annexin V apoptosis assays. Bioinformatics analyses and qRT-PCR, western blot, and luciferase reporter assays were performed to identify ADAM10 as the target gene of miR-23b. Results. miR-23b expression was downregulated in the peripheral blood mononuclear cells of sepsis patients and LPS-induced THP-1 cells and was negatively correlated with the expression of ADAM10 and inflammatory cytokines. miR-23b regulated ADAM10 expression by directly binding to the 3′-UTR of ADAM10 mRNA. The overexpression of miR-23b alleviated the LPS-stimulated production of inflammatory cytokines (TNF-α, IL-1β, and IL-6) and apoptosis by targeting ADAM10 in THP-1 cells. The inhibitor or knockdown of ADAM10 elicited effects similar to those of miR-23b on THP-1 cells upon LPS stimulation. Conclusions. The present study demonstrated that miR-23b negatively regulated LPS-induced inflammatory responses by targeting ADAM10. The molecular regulatory mechanism of miR-23b in ADAM10 expression and sepsis-induced inflammatory consequences may provide potential therapeutic targets for sepsis.

scholarly journals Long non-coding RNA CASC7 Contributes to the Progression of Sepsis-Induced Liver Injury by Targeting miR-217/TLR4 Axis

2021 ◽  
Author(s):  
Xiaoqian Zhou ◽  
Tao Zhang ◽  
Yanhua Tang ◽  
Wenyong Jiang ◽  
Weiwen Yang ◽  
...  
Keyword(s):  
Liver Injury ◽  
Annexin V ◽  
Luciferase Reporter ◽  
Dependent Manner ◽  
Luciferase Reporter Gene ◽  
Non Coding Rna ◽  
Enhanced Expression ◽  
Apoptosis Detection

Abstract Background: Sepsis is a system inflammation disease that can lead to liver injury. Long non-coding RNAs as crucial regulators participate in the regulation of sepsis-induced liver injury. However, the role of lncRNA CASC7 (CASC7) in the modulation of sepsis-induced liver injury remains elusive. Here, we aimed to explore the effect of CASC7 on the sepsis-induced liver injury. Methods: The sepsis mouse model was established in BALB/c mice by the treatment of lipopolysaccharide (LPS). The effect of CASC7 on sepsis-induced liver injury was analyzed by Hematoxylin and Eosin (HE) staining, ELISA assays, TUNEL detection kit, CCK‐8 assays, and Annexin V-FITC Apoptosis Detection Kit in vivo or in vitro. The mechanism investigation was performed using RNA pull-down, luciferase reporter gene assays, qPCR assays, and Western blot analysis.Results: The expression of CASC7 was elevated in a time-dependent manner in the liver tissues of the sepsis mice and LPS-treated LO2 cells. The depletion of CASC7 decreased the LPS treatment-induced liver injury in the sepsis mice. The treatment of LPS enhanced the apoptosis in the sepsis mice, while the depletion of CASC7 blocked this enhancement in the system. The CASC7 knockdown inhibited the LPS-enhanced expression of TNF-α and IL-1β in the mice. CASC7 served as a sponge for the miR-217 in the liver cells. CASC7 promoted the progression of sepsis-induced liver injury by sponging miR-217. MiR-217 attenuated sepsis-induced liver injury by targeting TLR4. Conclusions: Thus, we conclude that CASC7 contributes to the progression of sepsis-induced liver injury by targeting miR-217/TLR4 axis.

LncRNA HEIH promotes cell proliferation, migration and invasion by suppressing miR-214-3p in gastric carcinoma

2020 ◽  
Author(s):  
Lei Jiang ◽  
Luyao Zhang ◽  
Qian Chen ◽  
Shigang Qiao ◽  
Feng Zhou ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Gastric Carcinoma ◽  
Quantitative Polymerase Chain Reaction ◽  
Annexin V ◽  
Fluorescein Isothiocyanate ◽  
Cell Counting ◽  
Migration And Invasion ◽  
Gastric Carcinoma Cell ◽  
Normal Tissues ◽  
Apoptosis Detection

Abstract The present study aimed to investigate the function of long non-coding RNA HEIH in gastric carcinoma (GC). Adjacent normal tissues and GC tissues were obtained from 72 patients. Real-time quantitative polymerase chain reaction (RT-qPCR) was utilized to measure the expression of HEIH in cancer tissues and cells. Cell Counting Kit-8 and transwell assays were employed to evaluate cell proliferation, migration and invasion. An Annexin V-fluorescein-isothiocyanate (FITC)/propidium iodide (PI) Apoptosis Detection Kit was used to evaluate the apoptosis ratio. RT-qPCR was used to detect the expression level of miR-214-3p. The expression of HEIH in GC tissues was higher than in adjacent normal tissues. The expression of HEIH was upregulated in MKN-45, NCL-N87, KATO III cell lines compared within normal gastric epithelial cells. Knockdown of lncRNA HEIH significantly decreased the number of migrated and invaded cells. Additionally, downregulation of HEIH could increase GC cell apoptosis compared with the non-specific control (NC) group. We also proved that miR-214-3p was the direct target of lncRNA HEIH, and that overexpression of miR-214-3p could reverse the effects of HEIH. Silencing of HEIH could suppress Gastric Carcinoma cell proliferation, migration and invasion by inhibiting miR-214-3p. Thus, HEIH might represent a novel biomarker and therapeutic target.

scholarly journals EGFRvIII Promotes Cell Survival during Endoplasmic Reticulum Stress through a Reticulocalbin 1-Dependent Mechanism

Cancers ◽  
2021 ◽  
Vol 13 (6) ◽  
pp. 1198
Author(s):  
Juliana Gomez ◽  
Zammam Areeb ◽  
Sarah F. Stuart ◽  
Hong P. T. Nguyen ◽  
Lucia Paradiso ◽  
...  
Keyword(s):  
Endoplasmic Reticulum ◽  
Cell Viability ◽  
Er Stress ◽  
Cell Survival ◽  
Glioblastoma Cells ◽  
Over Expression ◽  
Stress Markers ◽  
Apoptotic Protein ◽  
Novel Association ◽  
Reduced Activity

Reticulocalbin 1 (RCN1) is an endoplasmic reticulum (ER)-residing protein, involved in promoting cell survival during pathophysiological conditions that lead to ER stress. However, the key upstream receptor tyrosine kinase that regulates RCN1 expression and its potential role in cell survival in the glioblastoma setting have not been determined. Here, we demonstrate that RCN1 expression significantly correlates with poor glioblastoma patient survival. We also demonstrate that glioblastoma cells with expression of EGFRvIII receptor also have high RCN1 expression. Over-expression of wildtype EGFR also correlated with high RCN1 expression, suggesting that EGFR and EGFRvIII regulate RCN1 expression. Importantly, cells that expressed EGFRvIII and subsequently showed high RCN1 expression displayed greater cell viability under ER stress compared to EGFRvIII negative glioblastoma cells. Consistently, we also demonstrated that RCN1 knockdown reduced cell viability and exogenous introduction of RCN1 enhanced cell viability following induction of ER stress. Mechanistically, we demonstrate that the EGFRvIII-RCN1-driven increase in cell survival is due to the inactivation of the ER stress markers ATF4 and ATF6, maintained expression of the anti-apoptotic protein Bcl-2 and reduced activity of caspase 3/7. Our current findings identify that EGFRvIII regulates RCN1 expression and that this novel association promotes cell survival in glioblastoma cells during ER stress.

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