scholarly journals The Critical Role of Redox Homeostasis in Shikonin-Induced HL-60 Cell Differentiation via Unique Modulation of the Nrf2/ARE Pathway

2012 ◽  
Vol 2012 ◽  
pp. 1-12 ◽  
Author(s):  
Bo Zhang ◽  
Na Chen ◽  
Hongmei Chen ◽  
Zhenhua Wang ◽  
Qiusheng Zheng
Keyword(s):  
Drug Exposure ◽  
Target Genes ◽  
Redox Homeostasis ◽  
Critical Role ◽  
Leukemia Cell ◽  
Leukemia Cell Line ◽  
Downstream Target ◽  
Dose Dependent Increase

Among various cancer cell lines, the leukemia cell line HL-60 was most sensitive to Shikonin, with evidence showing both the prooxidative activities and proapoptotic effects of micromolar concentrations of Shikonin. However, the mechanism involved in the cytotoxicity of Shikonin in the submicromolar range has not been fully characterized. Using biochemical and free radical biological experimentsin vitro, we identified the prodifferentiated profiles of Shikonin and evaluated the redox homeostasis during HL-60 differentiation. The data showed a strong dose-response relationship between Shikonin exposure and the characteristics of HL-60 differentiation in terms of morphology changes, nitroblue tetrazolium (NBT) reductive activity, and the expression level of surface antigens CD11b/CD14. During drug exposure, intercellular redox homeostasis changes towards oxidation are necessary to support Shikonin-induced differentiation, which was proven by additional enzymatic and non-enzymatic redox modulators. A statistically significant and dose-dependent increase (P<0.05) was recorded with regard to the unique expression levels of the Nrf2/ARE downstream target genes in HL-60 cells undergoing late differentiation, which were restored with further antioxidants employed with the Shikonin treatment. Our research demonstrated that Shikonin is a differentiation-inducing agent, and its mechanisms involve the Nrf2/ARE pathway to modulate the intercellular redox homeostasis, thus facilitating differentiation.

scholarly journals NUSAP1 Promotes Gastric Cancer Progression Through Regulation of Yap Stability

2020 ◽  
Author(s):  
Hui Guo ◽  
Jianping Zou ◽  
Ling Zhou ◽  
Yan He ◽  
Miao Feng ◽  
...  
Keyword(s):  
Gastric Cancer ◽  
Cancer Progression ◽  
Target Genes ◽  
Hippo Pathway ◽  
Critical Role ◽  
Xenograft Model ◽  
Migration And Invasion

Abstract Background:Nucleolar and spindle associated protein (NUSAP1) is involved in tumor initiation, progression and metastasis. However, there are limited studies regarding the role of NUSAP1 in gastric cancer (GC). Methods: The expression profile and clinical significance of NUSAP1 in GC were analysed in online database using GEPIA, Oncomine and KM plotter, which was further confirmed in clinical specimens.The functional role of NUSAP1 were detected utilizing in vitro and in vivo assays. Western blotting, qRT-PCR, the cycloheximide-chase, immunofluorescence staining and Co-immunoprecipitaion (Co-IP) assays were performed to explore the possible molecular mechanism by which NUSAP1 stabilizes YAP protein. Results:In this study, we found that the expression of NUSAP1 was upregulated in GC tissues and correlates closely with progression and prognosis. Additionally, abnormal NUSAP1 expression promoted malignant behaviors of GC cells in vitro and in a xenograft model. Mechanistically, we discovered that NUSAP1 physically interacts with YAP and furthermore stabilizes YAP protein expression, which induces the transcription of Hippo pathway downstream target genes. Furthermore, the effects of NUSAP1 on GC cell growth, migration and invasion were mainly mediated by YAP. Conclusions:Our data demonstrates that the novel NUSAP1-YAP axis exerts an critical role in GC tumorigenesis and progression, and therefore could provide a novel therapeutic target for GC treatment.

scholarly journals CircRNA-DOPEY2 enhances the chemosensitivity of esophageal cancer cells by inhibiting CPEB4-mediated Mcl-1 translation

2021 ◽  
Vol 40 (1) ◽  
Author(s):  
Zhenchuan Liu ◽  
Shaorui Gu ◽  
Kaiqin Wu ◽  
Lei Li ◽  
Chenglai Dong ◽  
...  
Keyword(s):  
Cancer Progression ◽  
Cisplatin Resistance ◽  
Critical Role ◽  
Circular Rnas ◽  
Major Barrier ◽  
Downstream Target ◽  
Cisplatin Sensitivity

Abstract Background Cisplatin-based chemotherapy is a mainstay systematic therapy for advanced esophageal squamous cell carcinoma (ESCC), and cisplatin resistance, which is not uncommon, is the major barrier to improving patient outcomes. Circular RNAs (circRNAs) are novel noncoding RNAs that are implicated in cancer progression, but their involvement in modulating cisplatin responsiveness in ESCC remains unknown. Methods Bioinformatics analysis was used to profile and identify the circRNAs involved in cisplatin responsiveness in ESCC. The chemosensitive role of cDOPEY2 was confirmed both in vitro and in vivo. The molecular mechanism of cDOPEY2 was investigated by mass spectrometry, immunoprecipitation, and ubiquitination analyses. Results We report that a novel circRNA (cDOPYE2, hsa_circ_0008078) was markedly downregulated in cisplatin-resistant ESCC cells (ESCC-CR) compared with parental chemosensitive cells. Re-expression of cDOPEY2 substantially enhanced the cell-killing ability of cisplatin by augmenting the apoptotic process in ESCC-CR cells, which was achieved by decreasing the abundance of the antiapoptotic protein Mcl-1. Mechanistically, we showed that cDOPEY2 acted as a protein scaffold to enhance the interaction between the cytoplasmic polyadenylation element binding protein (CPEB4) and the E3 ligase TRIM25, which in turn facilitated the ubiquitination and degradation of CPEB4. The increased Mcl-1 expression in ESCC-CR cells was dependent on the binding of CPEB4 to its untranslated mRNA, and depletion of CPEB4 mediated by cDOPEY2 reversed this effect. Rescue experiments confirmed that the critical role of cDOPEY2 in maintaining cisplatin sensitivity was dependent on the depletion of CEPB4 and its downstream target Mcl-1. Clinical and in vivo data further corroborated the significant relevance of cDOPEY2 to cisplatin responsiveness in ESCC. Conclusions We provide evidence that cDOPEY2 inhibits CPEB4-mediated Mcl-1 translation by promoting the ubiquitination and degradation of CPEB4 to alleviate cisplatin resistance, indicating that cDOPEY2 may serve as a valuable biomarker and potential therapeutic target in ESCC.

MLL-AF4 Down-Regulates CDKN1B (P27) Independent of Cell Cycle Progression.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 2563-2563
Author(s):  
Zhenbiao Xia ◽  
Relja Popovic ◽  
Tara Lorenz ◽  
Donna Santillan ◽  
Frank Erfurth ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Cell Growth ◽  
Cell Line ◽  
Cell Cycle Progression ◽  
Target Genes ◽  
Leukemia Cell ◽  
Leukemia Cell Line ◽  
Luciferase Reporter ◽  
Cycle Progression ◽  
Downstream Target

Abstract The MLL gene, involved in many chromosomal translocations associated with acute myeloid and lymphoid leukemia, has more than forty known partner genes with which it is able to form in- frame fusions. MLL fusion genes transform hematopoietic cells in vitro, and cause leukemia in mouse models. However, the mechanism is still not clear. Characterizing important downstream target genes may provide rational therapeutic strategies for the treatment of MLL-associated leukemia. We explored potential downstream target genes of the most prevalent MLL fusion protein, MLL-AF4, which is primarily associated with pro-B ALL and is involved in the majority of infant leukemia. To this end, we developed an inducible MLL-AF4 fusion cell line. Overexpression of MLL-AF4 does not lead to increased proliferation in this cell line, but rather, cell growth is slowed compared to similar cell lines inducibly expressing truncated MLL. To try to understand the reason for slower cell growth, we assayed for expression of several CDK inhibitors. We found that in the MLL-AF4 induced cell line, the amount of CDKN1B (cyclin-dependent kinase inhibitor P27) was dramatically decreased both at the RNA and protein levels, in contrast, the levels of CDKN1A (P21) and CDKN2A (P16) were unchanged. Interestingly, we did not observe an increased percentage of cells in S phase of the cell cycle. To explore whether CDKN1B might be a direct target of MLL-AF4, we employed chromatin immunoprecipitation (ChIP) assays and luciferase reporter gene assays. We observed that MLL-AF4 binds to the CDKN1B promoter in vivo and represses CDKN1B promoter activity. Further, we confirmed CDKN1B promoter binding by ChIP assays in the MLL-AF4 leukemia cell line MV4-11. Our results suggest that the CDKN1B may be a downstream target of MLL-AF4, and that MLL-AF4 inhibits CDKN1B expression independent of cell cycle progression.

Abstract 448: Etv2-Mir130a-Jarid2 Cascade Regulates Vascular Patterning During Embryogenesis

2017 ◽  
Vol 37 (suppl_1) ◽  
Author(s):  
Bhairab N Singh ◽  
Naoyuki Tahara ◽  
Yasuhiko Kawakami ◽  
Naoko Koyano-Nakagawa ◽  
Wuming Gong ◽  
...  
Keyword(s):  
In Situ Hybridization ◽  
Critical Role ◽  
Yolk Sac ◽  
Zebrafish Embryos ◽  
Vascular Patterning ◽  
Downstream Target

Remodeling of the pre-existing primitive vasculature is necessary for the formation of a complex branched vascular architecture. However, the factors that modulate these processes are incompletely defined. Previously, we defined the role of microRNAs (miRNAs) in endothelial specification. In the present study, we further examined the Etv2-Cre mediated ablation of Dicer L/L and characterized the perturbed vascular patterning in the embryo proper and yolk-sac. We mechanistically defined an important role for miR-130a , an Etv2 downstream target, in the mediation of vascular patterning and angiogenesis in vitro and in vivo . Inducible overexpression of miR-130a resulted in robust induction of vascular sprouts and angiogenesis with increased uptake of acetylated-LDL. Mechanistically, miR-130a directly regulates Jarid2 expression by binding to its 3’-UTR region. CRISPR/Cas9 mediated knockout of miR-130a showed increased levels of Jarid2 in the ES/EB system. Further, the levels of Jarid2 transcripts were increased in the Etv2-null embryos at E8.5. In the in vivo settings, injection of miR-130a specific morpholinos in zebrafish embryos resulted in perturbed vascular patterning with reduced levels of endothelial transcripts in the miR-130a morphants. qPCR and in situ hybridization techniques demonstrated increased expression of jarid2a in the miR-130a morphants in vivo . These findings demonstrate a critical role for Etv2-miR-130a-Jarid2 in vascular patterning both in vitro and in vivo .

scholarly journals GATA3 interacts with and stabilizes HIF-1α to enhance cancer cell invasiveness

Oncogene ◽  
2017 ◽  
Vol 36 (30) ◽  
pp. 4243-4252 ◽  
Author(s):  
M-C Lin ◽  
J-J Lin ◽  
C-L Hsu ◽  
H-F Juan ◽  
P-J Lou ◽  
...  
Keyword(s):  
Target Genes ◽  
Critical Role ◽  
Hypoxia Inducible Factor ◽  
Disease Free Survival ◽  
Underlying Mechanism ◽  
Immunodeficient Mice ◽  
Mediated Effects ◽  
Cell Invasiveness

Abstract GATA binding protein 3 (GATA3) is indispensable in development of human organs. However, the role of GATA3 in cancers remains elusive. Hypoxia inducible factor (HIF)-1 plays an important role in pathogenesis of human cancers. Regulation of HIF-1α degradation is orchestrated through collaboration of its interacting proteins. In this study, we discover that GATA3 is upregulated in head and neck squamous cell carcinoma (HNSCC) and is an independent predictor for poor disease-free survival. GATA3 promotes invasive behaviours of HNSCC and melanoma cells in vitro and in immunodeficient mice. Mechanistically, GATA3 physically associates with HIF-1α under hypoxia to inhibit ubiquitination and proteasomal degradation of HIF-1α, which is independent of HIF-1α prolyl hydroxylation. Chromatin immunoprecipitation assays show that the GATA3/HIF-1α complex binds to and regulates HIF-1 target genes, which is also supported by the microarray analysis. Notably, the GATA3-mediated invasiveness can be significantly reversed by HIF-1α knockdown, suggesting a critical role of HIF-1α in the underlying mechanism of GATA3-mediated effects. Our findings suggest that GATA3 stabilizes HIF-1α to enhance cancer invasiveness under hypoxia and support the GATA3/HIF-1α axis as a potential therapeutic target for cancer treatment.

scholarly journals In Vitro Modulation of P-Glycoprotein Activity by Euphorbia intisy Essential Oil on Acute Myeloid Leukemia Cell Line HL-60R

2021 ◽  
Vol 14 (2) ◽  
pp. 111
Author(s):  
Paola Poma ◽  
Manuela Labbozzetta ◽  
Aro Vonjy Ramarosandratana ◽  
Sergio Rosselli ◽  
Marco Tutone ◽  
...  
Keyword(s):  
Essential Oil ◽  
Target Genes ◽  
Biological Activities ◽  
Mrna Level ◽  
Leukemia Cell ◽  
Constitutive Expression ◽  
Leukemia Cell Line ◽  
Medicinal Uses ◽  
P Glycoprotein

Euphorbia species have a large spectrum of traditional medicinal uses. We tested the biological activities of the essential oil (EO) of Euphorbia intisy Drake in an acquired multidrug resistance leukemia model to assess whether the EO obtained by hydrodistillation of stems was able to reverse the resistant phenotype. HL-60R cell lines are characterized by the overexpression of P-glycoprotein (P-gp), inhibitors of apoptosis proteins (IAPs) and constitutive expression of NF-κB. EO chemical composition was determined by GC/MS analysis; cytotoxic activity of EO by MTS assay alone or in combination with doxorubicin; pro-apoptotic effect and doxorubicin accumulation were analyzed by flow cytometry; P-gp ATPase activity was measured by P-gp-Glo™ assay systems kit. The ability to inhibit NF-κB and its target genes was also assessed. E. intisy EO exhibited a comparable cytotoxic effect and ability to block P-gp in both the HL-60 and its MDR variant HL-60R. In addition, EO suppressed P-gp protein expression and significantly downregulated MDR1 mRNA level, as well as some IAPs proteins, probably through the inhibition of NF-κB. Our results suggest that E. intisy EO could reverse P-gp-mediated drug resistance in tumor cells acting as a chemosensitizing agent.

scholarly journals Role of a versatile peptide motif controlling Hox nuclear export and autophagy in the Drosophila fat body

2020 ◽  
Vol 133 (18) ◽  
pp. jcs241943
Author(s):  
Marilyne Duffraisse ◽  
Rachel Paul ◽  
Julie Carnesecchi ◽  
Bruno Hudry ◽  
Agnes Banreti ◽  
...  
Keyword(s):  
Nuclear Export ◽  
Target Genes ◽  
Fat Body ◽  
Nuclear Export Signal ◽  
Peptide Motif ◽  
Hox Proteins ◽  
Downstream Target

ABSTRACTHox proteins are major regulators of embryonic development, acting in the nucleus to regulate the expression of their numerous downstream target genes. By analyzing deletion forms of the Drosophila Hox protein Ultrabithorax (Ubx), we identified the presence of an unconventional nuclear export signal (NES) that overlaps with a highly conserved motif originally described as mediating the interaction with the PBC proteins, a generic and crucial class of Hox transcriptional cofactors that act in development and cancer. We show that this unconventional NES is involved in the interaction with the major exportin protein CRM1 (also known as Embargoed in flies) in vivo and in vitro. We find that this interaction is tightly regulated in the Drosophila fat body to control the autophagy-repressive activity of Ubx during larval development. The role of the PBC interaction motif as part of an unconventional NES was also uncovered in other Drosophila and human Hox proteins, highlighting the evolutionary conservation of this novel function. Together, our results reveal the extreme molecular versatility of a unique short peptide motif for controlling the context-dependent activity of Hox proteins both at transcriptional and non-transcriptional levels.

MDM2 Inhibitor Nutlin-3a Induces p53-Dependent Apoptosis Via Transcription-Dependent and -Independent Pathways and Overcomes Fludarabine-Resistance in CLL.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 445-445
Author(s):  
Kensuke Kojima ◽  
Marina Konopleva ◽  
Ismael J. Samudio ◽  
Teresa McQueen ◽  
Twee Tsao ◽  
...  
Keyword(s):  
Transcriptional Activation ◽  
Target Genes ◽  
Critical Role ◽  
Leukemia Cell ◽  
Annexin V ◽  
Current Standard ◽  
Induced Apoptosis ◽  
Mdm2 Overexpression ◽  
Mdm2 Inhibitor

Abstract Fludarabine containing combination therapies are the current standard for CLL therapy, in which p53-mediated induction of apoptosis contributes to leukemia cell killing. Although TP53 mutations occur in only 5–10% of patients with CLL, inactivation of the p53 pathway also occurs through Mdm2 overexpression or Atm deficiency. We investigate if recently developed potent and selective small-molecule antagonists of Mdm2, Nutlins, can overcome functional p53 inactivation associated with Mdm2 overexpression or Atm deficiency in CLL. Nutlin-3a caused a dose- and time-dependent increase in the percentage of Annexin V-positive cells in 20 primary CLL samples, irrespective of Mdm2 or Atm status. Samples with low Atm levels (n=3) were resistant to fludarabine. In addition to the transcriptional activation of target genes in the nucleus, cytoplasmic p53 can trigger transcription-independent apoptosis at the mitochondria. Samples with a cytoplasmic p53 localization pattern (n=7) showed a higher percentage of Nutlin-induced apoptosis than those with a nuclear pattern (n=8; P < .05). Furthermore, the inhibitory effect of cycloheximide pretreatment showed a negative correlation with the degree of Nutlin-induced apoptosis (r = - 0.617, P < .05). These findings suggest that the transcription-independent pathway may have stronger apoptogenic activity than the transcription-dependent pathway. The degree of Nutlin-3a-induced apoptosis directly correlated with apoptosis induced by the same concentrations (1–10 μM) of fludarabine in the early (24 hr) time period, and the Nutlin-3a/fludarabine combination induced synergistic apoptosis [averaged combination index (CI): 0.68]. Interestingly, this synergism was not affected by Mdm2 overexpression (CI: 0.59, n=12) or Atm deficiency (CI: 0.79, n=3). We conclude that (1) Mdm2 inhibitor Nutlin-3a efficiently induces p53-dependent apoptosis in wild-type p53 CLL cells, independent of Mdm2 overexpression or Atm deficiency, (2) transactivation-dependent apoptosis does not always play a major role in p53-dependent apoptosis and the exclusive activation of transactivation-independent pathway can fully induce apoptosis, (3) p53 activation plays a critical role in the early phase of fludarabine-induced apoptosis in vitro, and (4) Mdm2 inhibition and fludarabine synergistically induce apoptosis, which may overcome fludarabine-resistance in CLL.

Reversal of Methotrexate-Induced Folate Pool Depletion by Thymidine in a Human Leukemia Cell Line in Vitro

1993 ◽  
Vol 79 (6) ◽  
pp. 433-438 ◽  
Author(s):  
Pratima Sur ◽  
Yoshinobu Matsuo ◽  
Takeshi Otanl ◽  
Jun Minowada
Keyword(s):  
Cell Cycle ◽  
Cell Growth ◽  
Cell Line ◽  
Leukemia Cell ◽  
Leukemia Cell Line ◽  
Human Leukemia ◽  
Folate Level ◽  
Ifn Γ

In an In vitro study using a human monocytic leukemia cell line, U-937, the effects of interferon-γ (IFN-γ) in combination with the antifolate methotrexate and the role of thymidine introduced as a biochemical modulator were investigated. Methotrexate alone or in combination with INF-γ was found to enhance the induction of morphologic and functional monocytic differentiation in the U-937 cell line. Various cellular effects with the addition of thymidine to the medium with methotrexate and IFN-γ were studied. Enhanced inhibition of cell growth and perturbation of the cell cycle were noted when methotrexate and IFN-γ were used in combination, but not when methotrexate was used alone. The reduction of cellular folate by methotrexate was also enhanced in combination with IFN-γ. Cell cycle delay, resulting in cell growth inhibition of folate depletion, caused the induction of differentiation in U-937 cells, which was found to be greater with methotrexate + IFN-γ than with methotrexate alone. Cellular differentiation, as assessed by nitroblue tetrazolium reduction assay, indirect immunofluorescence and morphology, showed better effects towards the differentiation of U-937 cells when the agents were used in combination. However, addition of thymidine to the medium was found to cancel all the aforementioned effects. The addition of thymidine to the medium also caused reversal of the inhibitory effect of methotrexate and IFN-γ on cell growth and repletion of the endogenous folate level. Repletion of the folate level by exogenous thymidine is a new possibility for the role of the thymidine in cellular growth.

scholarly journals Inhibiting Apoptosis of CTLL-2 Cells to Enhance their GVL Effects via Anti-Fas Ribozyme

2004 ◽  
Vol 36 (8) ◽  
pp. 559-565
Author(s):  
Min Zhang ◽  
Fang Liu ◽  
Lin-Bo Liu ◽  
Yong You ◽  
Zhi-Chao Chen ◽  
...  
Keyword(s):  
Cell Line ◽  
Caspase 3 ◽  
Fas Ligand ◽  
Hammerhead Ribozyme ◽  
Leukemia Cell ◽  
Leukemia Cell Line ◽  
Killing Activity ◽  
Fas Mrna

Abstract To investigate the inhibition role of anti-Fas hammerhead ribozyme on fas expression and Fas-mediated apoptosis of CTL cell line CTLL-2 cells, the cDNA of an anti-Fas hammerhead ribozyme was synthesized, its expression plasmid was constructed and transfected into CTLL-2 cells by electroporation. fas expression of CTLL-2 cells was detected by RT-PCR and Western blot. CTLL-2 cell viability was measured using MTT assay when co-cultured with mouse T cell leukemia cell line EL4 cells that highly expressed Fas ligand (FasL). Meanwhile, caspase-3 proteolytic activity was detected, and cell apoptosis was measured by flow cytometry and Hochest-PI double staining. Killing activity of CTLL-2 cells was detected by lactate dehydrogenase (LDH) releasing assay in vitro. Results showed that the expression of both Fas mRNA and protein in CTLL-2 cells were decreased after transfection of anti-Fas ribozyme. Compared with mocktransfected group and mutant ribozyme-transfected group, viability of CTLL-2 cells co-cultured with EL4 cells was increased significantly and cells killing activity was enhanced after transfected with anti-Fas ribozyme, while the caspase-3 activity and apoptosis rate was significantly decreased. The results demonstrated anti-Fas ribozyme could efficiently cleave Fas and inhibit Fas-mediated apoptosis of CTLL-2 cells to improve their viability. Our study made a basis for enhancing CTLL-2 cells anti-leukemia effect in DLI.

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