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MGMT is down-regulated independently of promoter DNA methylation in rats with all-trans retinoic acidinduced spina bifida aperta

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Neural Regeneration Research 2019年第2期14卷 361-368页

作者： He-Nan Zhang Yi Guo Wei Ma Jia Xue Wei-Lin Wang Zheng-Wei Yuan Key Laboratory of Health Ministry for Congenital Malformation Shengjing Hospital China Medical University Department of Pediatric Surgery Shengjing Hospital China Medical University

O6-methylguanine DNA methyltransferase(MGMT), a DNA repair enzyme, has been reported in some congenital malformations, but it is less frequently reported in neural tube defects. This study investigated MGMT mRNA expression and methylation levels in the early embryo and in different embryonic stages, as well as the relationship between MGMT and neural tube defects. Spina bifida aperta was induced in rats by a single intragastric administration of all-trans retinoic acid on embryonic day(E) 10, whereas normal control rats received the same amount of olive oil on the same embryonic day. DNA damage was assessed by detecting γ-H2 A.X in spina bifida aperta rats. Real time-polymerase chain reaction was used to examine mRNA expression of MGMT in normal control and spina bifida aperta rats. In normal controls, the MGMT mRNA expression decreased with increasing embryonic days, and was remarkably reduced from E11 to E14, reaching a minimum at E18. In the spina bifida aperta model, γ-H2 A.X protein expression was increased, and mRNA expression of MGMT was markedly decreased on E14, E16, and E18. Bisulfite sequencing polymerase chain reaction for MGMT promoter methylation demonstrated that almost all CpG sites in the MGMT promoter remained unmethylated in both spina bifida aperta rats and normal controls, and there was no significant difference in methylation level between the two groups on either E14 or E18. Our results show that DNA damage occurs in spina bifida aperta rats. The mRNA expression of MGMT is downregulated, and this downregulation is independent of promoter DNA methylation.

关键词： nerve regeneration neural tube defects spina bifida aperta spinal cord all-trans retinoic acid O6-methylguanine DNA methyltransferase gene expression DNA methylation promoter bisulfite sequencing polymerase chain reaction neural regeneration

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Identification of target genes of transcription factor CEBPB in acute promyelocytic leukemia cells induced by all-trans retinoic acid

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Asian Pacific Journal of Tropical Medicine 2013年第6期6卷 473-480页

作者： Lei Yu Yang-De Zhang Jun Zhou De-Ming Yao Xiang Li Department of Hepatic Surgery National Hepatobiliary and Enteric Surgery Research CenterMinistry of HealthCentral South UniversityChina

Objective:To indentify target genes of transcription factor CCA AT enhancer-binding protein P(CEBPB) in acute proinyelocytie leukemia cells induced by all-tram retinoie ***: A new strategy for high—throughput identification of direct target genes was established by combining chromatin immunoprecipitation(ChIP) with in vitro ***,106 potential CKBPB binding fragments from the genome of the all-trans retinoie acid(ATRA)-treated NB4 cells were ***:Of them,82 were mapped in proximity to known or previously predicted genes;7 were randomly picked up for further confirmation by ChlP-PCR and 3 genes (CALM,1TPR2 and 0RM2) were found to be specificaUy up-regulated in the ATRA-treated NB4 cells,indicating that they might lie the down-stream target genes of ***:Our results provided new insight into the mechanisms of ATRA-induced granulocytic differentiation.

关键词： Chromatin immunoprecipitation all-trans retinoic acid CEBP Granulopoiesis Neutrophils

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Effects of prostaglandin E_2 and all-trans retinoic acid combination on induced differentiation of human acute promyelocytic leukemia NB_4 cells

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Chinese Medical Journal 1997年第10期 48-53页

作者：陈子兴

Objective A human promyelocytic leukemia (APL) cell line NB 4 was used to demonstrate the synergistic effects between all trans retinoic acid (ATRA) and prostaglandin E 2 (PGE 2) on growth inhibition and cytodifferentiation induction. Methods NB 4 cells were cultured in the presence of either ATRA or PGE 2 as a single agent or in combinations at various ratio. Cell growth was measured and myeloid differentiation was tested on consecutive days over the whole course of culture. Results PGE 2 and ATRA synergistically induced the myeloid differentiation of NB 4 cells. In comparison with ATRA alone, the combination of PGE 2 with ATRA caused an almost 20 fold decrease of effective concentration of ATRA. Conclusion The combination of ATRA and PGE 2 at an appropriate ratio may provide a convenient oral regimen of combined differentiation therapy for a better clinical outcome in APL patients.

关键词： prostaglandin all-trans E2 Effects

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The Effects of all-trans Retinoic Acid on Vasculogenic Mimicry Formation Ability in CD133+ Glioma Stem Cells and Its Mechanisms

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Journal of Biosciences and Medicines 2017年第4期5卷 42-54页

作者： Ronghua Tang Chen Liang Jian Shangguan Shiwen Guo Department of Neurosurgery First Affiliated Hospital of Xi’an Jiaotong University Xi’an China

Objective: to investigate the effects of all-trans retinoic acid (ATRA) on vasculogenic mimicry formation in glioma stem cells. Methods: U87 stem cells were harvested through a suspension culture assay from the U87 cells, identified by CD133 and nestin, and counted by a flow cytometry. To investigate the VM formation ability of U87 stem cells with the treatment of various concentrations of ATRA, a Matrigel-based tube formation assay was used in the present study in vitro and tube-like structure (typical tube, TT;atypical tube AT) was observed and counted. Then the expressions of VEGF, VEGFR-2 and CD133 were measured throughout real time q-PCR, western blotting and immunofluorescence techniques. The data, presented as the mean ± standard deviation, were analyzed using SPSS software. One-way analysis of variance was used to compare groups and Fisher’s least significant difference tests were performed for subsequent comparisons between groups. P Results: Most of the harvested spheroid cells were positive for nestin and 88.4% were positive forCD133. The CD133+ U87 cells were cultured into tube like structure loaded on the top of Matrigel and the quantity of tubes was decreased under the treatment of ATRA. In addition, the expressions of VEGF, VEGFR-2 and CD133 were significantly reduced under the treatment of ATRA, particularly in the higher concentration groups (20 and 40 μmol, P Conclusions: ATRA may inhibit the establishment of VM differing from stem cells in glioma, and these effects may attribute to the effects of ATRA’s promotion of the differentiation of stem cells and/or down regulation of the expressions of proangiogenic factors VEGF and its receptor VEGFR-2. Thus, the results of the present study indicated a novel idea for the treatment of GBM and enriched the anti-glioma mechanisms of ARTA.

关键词： all-trans Retinoic Acid Vasculogenic Mimicry Glioma Stem Cells

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COMPARISON OF CLINICAL OBSERVATIONS BETWEEN PATIENTS WITH ACUTE PROMYELOCYTIC LEUKEMIA TREATED WITH all-trans RETINOIC ACID AND CHEMOTHERAPY

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Medical Bulletin of Shanghai Jiaotong University 1992年第2期15卷 83-87页

作者：张芬琴吴立德李秀松孙关林蔡敬仁王振义 Department of Internal Medicine Ruijin Hosptial and Shanghai Institute of Hematology SSMU Shanghai

Clinical observations were retrospectively compared between 2 matched groups of patients with acute promyelocytic leukemia (APL) each 20. The first group were treated with chemotherapy, the other with all-tram retinoic acid (ATRA) alone at a dose of 45-60mg/M^2/d. The complete remission (CR) rate of ATRA group was significantly higher than that of chemotherapy (90% vs 55%). The time for obtaining CR as well as the duration of fever and hospitalization were shorter and the amount of blood transfused was less in the former than in the latter group. Seven cases were complicated by DIC and 4 died in the group of chemotherapy, while no case was by of DIC or death in the ATRA group. The mechanism was discussed. ATRA is an alternative effective drug for remission induction therapy in APL with high rate of CR.

关键词： acute promyelocytic leukemia all-trans retinoic acid complete remission rate

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The E3 ubiquitin protein ligase MDM2 dictates ATRA-induced osteoblastic differentiation of osteosarcoma cells by

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中国药理学通报 2015年第B11期31卷 64-65页

作者： ZHOUQian SHAOXue-jing ZhejiangProvinceKeyLaboratoryofAnti-CancerDrugResearch CollegeofPharmaceuticalSciencesZhejiangUniversityHangzhou310058

Aim Retinoic acid receptor alpha （RARα） plays a critical role in the differentiation process of osteo- sarcoma cells induced by all-trans retinoic acid （ATRA）. However, degradation of RARoL through ubiquitin pro- teasome pathway weakens the differentiation efficiency of osteosarcoma cells. Therefore, it is necessary to explore the regulatory mechanisms involved in RARoL degradation. Methods U2OS cells were mainly used as our research models. （1） Protein levels were detected by Western blot; （2） The interactions between MDM2 and RARalpha were determined by immunofluorescence and immunoprecipitation; （3） Related proteins and genes were investiga- ted via transfection; （4） The expression levels of MDM2 and OPN in patient biopsies were detected by immunohis- tochemistry; （5） Alkaline phosphatase activity was assessed by colorimetric assays using the BCIP/NBT Alkaline Phosphatase Color Development kit. Results In this report, it demonstrated that interaction with MDM2 leaded to strong stimulation of RARoL polyubiquitination and degradation by proteasomes. MDM2 appeared to function as an ubiquitin E3 ligase in this process, since the MDM2 RING domain mutant inhibited the ubiquitination of RARoL. Furthermore, MDM2 was capable of stimulating RARoL polyubiquitination under cell-free conditions. Moreover, it also provided evidence that silencing or inhibiting MDM2 promotes the differentiation of U2OS cells as induced by ATRA. Conclusions MDM2 serves as an E3 ubiquitin ligase to regulate the degradation of RARoL and becomes a novel therapeutic target for ATRA-based differentiation therapeutic approaches in osteosarcoma.

关键词： osteosarcoma RARα degradation MDM2 all-trans retinoic acid differentiation

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Identification and Characterization of Human Genomic Binding Sites for Retinoic Acid Receptor/Retinoid X Receptor Heterodimers

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Advances in Biological Chemistry 2015年第2期5卷 58-72页

作者： Kenta Hosoda Masashi Sato Kazuyuki Yanai Department of Biomolecular Science Faculty of Science Toho University Chiba Japan

all-trans retinoic acid (ATRA) triggers a wide range of effects on vertebrate development by regulating cell proliferation, differentiation, and apoptosis. ATRA activates retinoic acid receptors (RARs) which heterodimerize with retinoid X receptors (RXRs). RAR/RXR heterodimers function as ATRA-dependent transcriptional regulators by binding to retinoic acid response elements (RAREs). To identify RAR/RXR heterodimer-binding sites in the human genome, we performed a modified yeast one-hybrid assays and identified 193 RAR/RXR heterodimer-binding fragments in the human genome. The putative target genes included genes involved in development process and cell differentiation. Gel mobility shift assays indicated that 160 putative RAREs could directly interact with the RAR/RXR heterodimer. Moreover, 19 functional regulatory single nucleotide polymorphisms (rSNPs) on the RAR/RXR-binding sequences were identified by analyzing the difference in the DNA-binding affinities. These results provide insights into the molecular mechanisms underlying the physiological and pathological actions of RAR/RXR heterodimers.

关键词： all-trans Retinoic Acid Retinoic Acid Receptor Retinoid X Receptor Yeast One-Hybrid System Polymorphism Regulatory SNP

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EXPERIMENTAL STUDY ON DIMETHYLSULFOXIDEINDUCED DIFFERENTIATION OF HUMAN ESOPHAGEALEPITHELIAL CARCINOMA CELLS

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Medical Bulletin of Shanghai Jiaotong University 1998年第Z1期19卷 34-39页

作者：谭立军李慧易静汤雪明沈忠英

Objective The treatment of solid tumors by means of differentiation induction has not yet beengained such broken - through success as all-trans retinoic acid (ATRA) in acute promyelocytic leukemia (APL).To explore more effective inducers for solid tumor therapy, dimethyl sulfoxide (DMSO) is considered as acandidate. Methods In the present study, DMSO was used as inducer to human esophageal cancer cell lines invitro, in compared with classical inducer ATRA, in terms of morphology, cell cycle, growth inhibition, cytokeratin4 expression, gap junction-mediated dye transfer and tumorigenecity in nude mice. Results DMSO as well asATRA induced differentiation of human esophageal carcinoma cells. However, DMSO was confirmed to be moreeffective for induced differentiation of esophageal carcinoma cells than ATRA. Conclusion The results suggestedthat DMSO might have a good prospect in the treatment of solid tumors.

关键词： induced differentiation esophageal cancer tumor therapy dimethylsulfoxide all-trans retinoic acid

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Sweet syndrome and differentiation syndrome in a patient with acute promyelocytic leukemia

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World Journal of Clinical Cases 2015年第2期3卷 196-198页

作者： Guillermo Solano-López Mar Llamas-Velasco Maria José Concha-Garzón Esteban Daudén Department of Dermatology Hospital Universitario de la Princesa

The differentiation syndrome is an inflammatory reaction with increased capillary permeability that occurs in up to 25% of patients with acute promyelocytic leukemia treated with all-trans retinoic acid. A 50-year-old man with acute promyelocytic leukemia underwent chemotherapy with idarubicin and all-trans retinoic acid. On day +21 the patient developed pruritic prepatelar papules as well as several 10 mm subcutaneous nodules in both thighs accompanied by persistent fever. On the day +25 the patient presented with bilateral pulmonary crackles, infiltrates in the right lower lobe and severe hypotension which required dopamine infusion. Biopsy of one of the thighs nodules was performed. A Sweet syndrome associated to a differentiation syndrome was suspected. all-trans retinoic acid therapy was discontinued and dexamethasone was administered. In 48 h the patient showed remission of the fever and the infiltrates and the skin lesions acquired a residual aspect. It is debatable whether these two syndromes are distinct entities with common mechanisms or whether they are poles of the same spectrum. Dermatologists and hematologists must be aware of these two syndromes and its pathophysiologic association.

关键词： Differentiation syndrome Sweet syndrome Acute promyelocytic leukemia all-trans retinoic acid

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