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Treatment of glioblastoma multiforme (GBM) remains challenging. Unraveling the orchestration of glutamine metabolism may provide a novel viewpoint on GBM therapy. The study presented a full and comprehensive comprehending of the glutamine metabolism atlas and heterogeneity in GBM for facilitating the development of a more effective therapeutic choice. Transcriptome data from large GBM cohorts were integrated in this study. A glutamine metabolism-based classification was established through consensus clustering approach, and a classifier by LASSO analysis was defined for differentiating the classification. Prognosis, signaling pathway activity, tumor microenvironment, and responses to immune checkpoint blockade (ICB) and small molecular drugs were characterized in each cluster. A combinational therapy of glutaminase inhibitor CB839 with dihydroartemisinin (DHA) was proposed, and the influence on glutamine metabolism, apoptosis, reactive oxygen species (ROS), and migration was measured in U251 and U373 cells. We discovered that GBM presented heterogeneous glutamine metabolism-based clusters, with unique survival outcomes, activity of signaling pathways, tumor microenvironment, and responses to ICB and small molecular compounds. In addition, the classifier could accurately differentiate the two clusters. Strikingly, the combinational therapy of CB839 with DHA synergistically attenuated glutamine metabolism, triggered apoptosis and ROS accumulation, and impaired migrative capacity in GBM cells, demonstrating the excellent preclinical efficacy. Altogether, our findings unveil the glutamine metabolism heterogeneity in GBM and propose an innovative combination therapy of CB839 with DHA for this malignant disease.
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Artémisinines , Tumeurs du cerveau , Glioblastome , Glutamine , Glioblastome/métabolisme , Glioblastome/traitement médicamenteux , Humains , Glutamine/métabolisme , Lignée cellulaire tumorale , Tumeurs du cerveau/métabolisme , Tumeurs du cerveau/traitement médicamenteux , Artémisinines/usage thérapeutique , Artémisinines/pharmacologie , Espèces réactives de l'oxygène/métabolisme , Glutaminase/métabolisme , Glutaminase/antagonistes et inhibiteurs , Microenvironnement tumoral , Apoptose , Thiadiazoles/pharmacologie , Thiadiazoles/usage thérapeutique , Mouvement cellulaire , Benzèneacétamides/pharmacologie , Benzèneacétamides/usage thérapeutique , Antinéoplasiques/usage thérapeutique , Antinéoplasiques/pharmacologie , Protocoles de polychimiothérapie antinéoplasique/usage thérapeutique , Protocoles de polychimiothérapie antinéoplasique/pharmacologieRÉSUMÉ
Allyl sulfones are commonly present in bioactive compounds and organic building blocks. This work introduces a photocatalytic radical addition-elimination reaction involving readily accessible sulfonyl chlorides and allyl bromides. It delivers structurally diverse allylic sulfones in moderate to excellent yields, showcasing a high tolerance to functional groups. Notably, this method operates under mild reaction conditions without the need for oxidants, stoichiometric reducing metals, or additives.
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BACKGROUND: Ischemic stroke (IS) is a serious cerebrovascular disease characterized by significantly elevated mortality and disability rates, and the treatments available for this disease are limited. Neuroinflammation and oxidative stress are deemed the major causes of cerebral ischemic injury. N-Cinnamoylpyrrole alkaloids form a small group of natural products from the genus Piper and have not been extensively analyzed pharmacologically. Thus, identifying the effect and mechanism of N-cinnamoylpyrrole-derived alkaloids on IS is worthwhile. PURPOSE: The present research aimed to explore the antineuroinflammatory and antioxidative stress effects of N-cinnamoylpyrrole-derived alkaloids isolated from the genus Piper and to explain the effects and mechanism on IS. METHODS: N-cinnamoylpyrrole-derived alkaloids were isolated from Piper boehmeriaefolium var. tonkinense and Piper sarmentosum and identified by various chromatographic methods. Lipopolysaccharide (LPS)-induced BV-2 microglia and a mouse model intracerebroventricularly injected with LPS were used to evaluate the antineuroinflammatory and antioxidative stress effects. Oxygenâglucose deprivation/reperfusion (OGD/R) and transient middle cerebral artery occlusion (tMCAO) models were used to evaluate the effect of PB-1 on IS. To elucidate the fundamental mechanism, the functional target of PB-1 was identified by affinity-based protein profiling (ABPP) strategy and verified by cellular thermal shift assay (CETSA), drug affinity responsive target stability (DARTS), and circular dichroism (CD) analyses. The effect of PB-1 on the NF-κB and NRF2 signaling pathways was subsequently evaluated via western blotting and immunofluorescence staining. RESULTS: The results showed that N-cinnamoylpyrrole-derived alkaloids significantly affected neuroinflammation and oxidative stress. The representative compound, PB-1 not only inhibited neuroinflammation and oxidative stress induced by LPS or OGD/R insult, but also alleviated cerebral ischemic injury induced by tMCAO. Further molecular mechanism research found that PB-1 promoted antineuroinflammatory and antioxidative stress activities via the NF-κB and NRF2 signaling pathways by targeting eEF1A1. CONCLUSION: Our research initially unveiled that the therapeutic impact of PB-1 on cerebral ischemic injury might rely on its ability to target eEF1A1, leading to antineuroinflammatory and antioxidative stress effects. The novel discovery highlights eEF1A1 as a potential target for IS treatment and shows that PB-1, as a lead compound that targets eEF1A1, may be a promising therapeutic agent for IS.
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Alcaloïdes , Accident vasculaire cérébral ischémique , Piper , Pyrroles , Animaux , Mâle , Souris , Alcaloïdes/pharmacologie , Alcaloïdes/composition chimique , Anti-inflammatoires/pharmacologie , Anti-inflammatoires/composition chimique , Antioxydants/pharmacologie , Antioxydants/composition chimique , Modèles animaux de maladie humaine , Accident vasculaire cérébral ischémique/traitement médicamenteux , Lipopolysaccharides , Souris de lignée C57BL , Microglie/effets des médicaments et des substances chimiques , Neuroprotecteurs/pharmacologie , Neuroprotecteurs/composition chimique , Facteur-2 apparenté à NF-E2/métabolisme , Facteur de transcription NF-kappa B/métabolisme , Stress oxydatif/effets des médicaments et des substances chimiques , Piper/composition chimique , Extraits de plantes/pharmacologie , Extraits de plantes/composition chimique , Pyrroles/pharmacologie , Pyrroles/composition chimique , Cinnamates/composition chimique , Cinnamates/pharmacologie , Facteur-1 d'élongation de la chaîne peptidique/antagonistes et inhibiteurs , Facteur-1 d'élongation de la chaîne peptidique/métabolismeRÉSUMÉ
Background: Neutrophil extracellular traps (NETs) play a key role in thrombus formation in patients with coronavirus disease 2019 (COVID-19). However, the existing detection and observation methods for NETs are limited in their ability to provide quantitative, convenient, and accurate descriptions of in situ NETs. Therefore, establishing a quantitative description of the relationship between NETs and thrombosis remains a challenge. Objective: We employed morphological observations of blood cells and statistical analyses to investigate the correlation between the NETs/neutrophilic segmented granulocyte ratio and mortality risk in patients with COVID-19. Methods: Peripheral blood samples were collected from 117 hospitalized patients with COVID-19 between November 2022 and February 2023, and various blood cell parameters were measured. Two types of smudge cells were observed in the blood and counted: lymphatic and neutral smudge cells. Statistical data analysis was used to establish COVID-19 mortality risk assessment indicators. Results: Morphological observations of neutrophilic smudge cells revealed swelling, eruption, and NETs formation in the neutrophil nuclei. Subsequently, the NETs/neutrophilic segmented granulocyte ratio (NNSR) was calculated. A high concentration of NETs poses a fatal risk for thrombus formation in patients. Statistical analysis indicated that a high NNSR was more suitable for evaluating the risk of death in patients with COVID-19 compared to elevated fibrinogen (FIB) and D-dimer (DD) levels. Conclusion: Observing blood cell morphology is an effective method for the detection of NETs, NNSR are important markers for revealing the mortality risk of patients with COVID-19.
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COVID-19 , Pièges extracellulaires , Thrombose , Humains , Pièges extracellulaires/métabolisme , COVID-19/métabolisme , Granulocytes neutrophiles/métabolisme , Thrombose/métabolismeRÉSUMÉ
Nine undescribed sesquiterpene lactones, including two pseudoguaianolide dimers (1 and 2), a pseudoguaiac dilactone (3), and six pseudoguaianolides (4-9), along with 13 known analogues (10-22) were isolated from Parthenium hysterophorus. Among them, hysterolide A (1) possesses an unusual carbon skeleton with a unique cyclobutane ring connecting two pseudoguaianolides. Hysterolide C (3) is a sesquiterpene dilactone incorporating a bicyclo[5.1.0]octane core. Spectroscopic analyses, 13C NMR and ECD calculations, and X-ray diffraction elucidated their structures and absolute configurations. Moreover, all the isolates were assayed for their anti-inflammatory activities by inhibiting LPS-induced nitric oxide production in BV-2 microglia cells, wherein, nine compounds displayed significant inhibitory activities with IC50 of 0.52-6.32 µM. Furthermore, the preliminary structure-activity relationship was also established.
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Asteraceae , Sesquiterpènes , Structure moléculaire , Parthenium Hysterophorus , Lactones/pharmacologie , Lactones/composition chimique , Spectroscopie par résonance magnétique , Sesquiterpènes/pharmacologie , Sesquiterpènes/composition chimique , Asteraceae/composition chimiqueRÉSUMÉ
OBJECTIVE: To study the impact of the nursing model based on the interactive model of health behavior (IMCHB) on the self-efficacy and negative emotions of prostate cancer patients. METHODS: 70 cases clinically admitted to our hospital from January 1, 2021 to January 31, 2023 Patients who received radiotherapy for prostate cancer were taken as research subjects and divided into a control group (n=35) and an observation group (n=35) according to the random drawing method. The control group was given routine care, and the observation group was given a nursing model based on IMCHB on the basis of the control group. Compare the incidence of complications between the two groups, and use the Frankl Compliance Scale (FCS) to compare the compliance behaviors of the two groups, negative emotions on the day of admission and 1 day before discharge were statistically compared between the two groups Hamilton Depression Rating Scale (HAMD), Hamilton Anxiety Rating Scale (HAMA), and self-management efficacy General Self-Efficacy Scale (GSES)and quality of life World Health Organization Quality of Life Scale Short Form (WHOQOL-BREF). RESULTS: The incidence of radiation proctitis and radiation cystitis in observation group was lower than that in control group, the difference was statistically significant (P<0.05).The total compliance rate of medical compliance was 97.83%, which was higher than that of the control group, 84.78%, and the difference was statistically significant (P<0.05); 1 day before discharge, the HAMD and HAMA scores of the observation group were lower than those of the control group, GSES, and WHOQOL-BREF The score was higher than that of the control group, and the difference was statistically significant (P<0.05). CONCLUSION: The IMCHB nursing model can improve self-management efficiency and alleviate negative emotions when applied to patients undergoing radiotherapy for prostate cancer.
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Tumeurs de la prostate , Qualité de vie , Mâle , Humains , Auto-efficacité , Comportement en matière de santé , Modèles de soins infirmiers , Tumeurs de la prostate/radiothérapie , ÉmotionsRÉSUMÉ
Objective To provide anatomical basis for clinical treatment of acromioclavicular joint dislocation by studying the morphology of coracoid process of human scapula. Methods A total of 500 patients with shoulder injury were selected from the Affiliated Hospital of Traditional Chinese Medicine of Southwest Medical University in Sichuan Province, and 300 patients were selected as subjects, including 159 cases of right shoulder and 141 cases of left shoulder. CT scan images and 3D reconstruction results of scapula of the subjects were collected. The basic morphological characteristics of coracoid process CT images of the subjects were observed, and the relevant parameters were measured, including the longest horizontal distance of the coracoid process tip and the thickness of the midpoint (cd, pp’), the distance from the upper part of the coracoid process scapula to the base and the thickness of the midpoint (mn, kk’). The distance from the apex of the coracoid process to the base of the coracoid process (ab), the longest horizontal distance of the recursion part of the coracoid process (ef), the distance of as (point s was the intersection of point a perpendicular to mn), the distance of hj (point h and j were the intersection of the base of the coracoid process and the recursion part respectively), and ik (point i was the intersection of point k perpendicular to mn and the coracoid process retraction). Results According to the morphological characteristics of coracoid process, they were divided into five types, including peanut 29. 7%; Short rod type accounted for 27. 4%; Melon seed type accounted for 12. 6%; Rod type accounted for 17. 0%; Wedge type accounted for 13. 3%. Through data comparison, it was found that the distance ef and distance hj on the left were larger than those on the right, P<0. 05. All types had statistical difference in comparison distance cd, P<0. 05. The melon seed type showed statistical differences with peanut type, wedge type, long stick type and short stick type in thickness pp’, distance ab and as of point p, P<0. 05. In the comparison of point K thickness kk’, there was statistical difference between melon seed type and other four types, P<0. 05. In the distance ab comparison, there was statistical difference between the short bar type and the other four types, P < 0. 05. Conclusion The study on the morphology of coracoid process can provide anatomical basis for clinical reconstruction of coracoid ligament to treat acromioclavicular joint dislocation.
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Acid sphingomyelinase deficiency (ASMD), also known as type A and B Niemann-Pick disease, is a group of intra-lysosomal lipid storage diseases caused by mutations in the SMPD1 gene that decrease acid sphingomyelinase activity or even cause deletion, resulting in abnormal deposition of sphingolipids. This disease can be diagnosed by bone marrow aspiration, pathological biopsy, acid sphingomyelinase activity measurement and SMPD1 gene testing. In recent years, with the rapid progress of molecular diagnostic techniques, new insights have been gained in the laboratory diagnosis of ASMD by means of molecular genetic tests, biomarkers and acid sphingomyelinase activity assay. This article will review the diagnostic progress of ASMD, aiming to reduce the misdiagnosis and leakage of the disease and improve the clinicians′ understanding of the disease.
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OBJECTIVE@#To investigate the toxicity management and efficacy evaluation of BCMA-chimeric antigen receptor T cells(CART) in the treatment of relapsed and refractory multiple myeloma (MM).@*METHODS@#The efficacy and adverse reactions of 21 patients with MM who received BCMA-CART treatment at the First Affiliated Hospital of Wenzhou Medical University from December 2017 to September 2020 were evaluated, and the efficacy assessment and survival analysis for high-risk patients and non-high-risk patients were evaluated.@*RESULTS@#After infusion of BCMA-CART cells in 21 MM patients, the number of effective cases was 17, of which the complete remission (sCR/CR) was 10, and the partial remission (VGPR/PR) was 7. The median OS time for all patients was 19.4 months, and the median PFS time was 7.9 months. The number of patients with extramedullary disease(EMD), high-risk genetics, and ISS stage Ⅲ were 5, 15 and 8, and the effective number was 3, 11 and 6, respectively. The treatment of 3 patients without high-risk factors was effective. The median OS and median PFS of patients with EMD were 14.2 and 2.5 months, respectively, which were shorter than those of patients without EMD (19.4 months and 8.9 months, respectively). The median OS and median PFS of patients with high-risk cytogenetic factors and ISS Ⅲ were not significantly different from those of non-high-risk patients. Cytokine release syndrane (CRS) occurred in 20 patients, of which 14 cases were Grade 1 CRS, while 6 were Grade 2, no CRS of Grade 3 or above occurred. IL-6 receptor inhibitors were used in 9 patients. All CRS were controlled effectively, and no patients had neurological toxicity.@*CONCLUSION@#BCMA-CART is a certain curative effect in the treatment of relapsed and refractory multiple myeloma, and the adverse reactions can be well controlled through close monitoring and timely treatment.
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Humains , Antigène de maturation des cellules B , Immunothérapie adoptive/effets indésirables , Myélome multiple/thérapie , Récepteurs chimériques pour l'antigène , Induction de rémissionRÉSUMÉ
Objective To determine the potential sources of food safety risk and food safety related awareness of employees in the farmer markets in Yangpu District of Shanghai, and to provide evidence for food safety management. Methods The investigation on food safety and food-borne disease knowledge were conducted in 13 farmer markets in Yangpu District, and to assess the impact of employees′ behavior on food safety. Results A total of 368 employees in the 13 farmer markets were included in the study.Employees with high school education and above had better knowledge of food safety (P < 0.001).There was significant difference in the physical examination and habit of smoking in public places among the employees with different education levels.In addition, the employees in various positions in the markets differed significantly in the knowledge of food safety and behavior, such as taking physical examination, washing hands before meal and continuing working when feeling sick. Conclusion Food safety related risk in the farmer markets in Yangpu District mainly originates from the inadequate knowledge of the employees with low education levels, which further affects their behavior.Market management should strengthen the training of food safety related knowledge in the employees and advocate healthy behavior, such as taking physical examination and stop smoking, which contributes to creating a healthy work and shopping environment.
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Objective To assess the correlation between magnetic resonance imaging (MRI)and clinical staging of pelvic organ prolapse (POP)by using pubococcygeal line (PCL)and midpubic line (MPL)respectively.And to determine which(MRI)reference line for staging pelvic organ prolapse had the highest agreement with clinical staging.Methods A retrospective study of 32 women with pelvic organ prolapse who underwent dynamic pelvic MRI from Jan 2013 to May 2013 was conducted.One radiologist staged descent on MRI for each pelvic compartment (anterior,middle,posterior),using PCL and MPL lines.Agreement between MRI and clinical staging was estimated respectively.Results Thirty-two women were included.Agreement between clinical and MPL staging was morderate in the anterior (κ=0.44,SE=0.13)and middle compartment (κ=0.46,SE=0.12),but fair in the posterior com-partment (κ=0.23,SE=0.10).Agreement between clinical and PCL staging was poor in all compartments of pelvic.Conclusion MPL appeared more reliable and simple than the PCL for the evaluation of pelvic prolapse on MRI.
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Objective To demonstrate the carotid artery hemodynamic characteristics in patients with common carotid artery occlusion,and to investigate the value of color Doppler ultrasonography in evaluating it.Methods 30 patients with common carotid artery (CCA) occlusion or severe stenosis (stenosis rate>70%) were as the patients group,while 30 volunteers with the normal indicators of carotid artery ultrasonography as the control group.In both groups,the examinations of the CCA,the internal carotid artery (ICA),the external carotid artery (ECA) and the vertebral artery (VA) were taken by color Doppler ultrasonography under quiescent conditions.Bilateral carotid ultrasonography studies were performed,which included the peak systolic velocity (PSV),end-diastolic velocity (EDV),mean velocity (MV),and resistance index (RI).Results The peak systolic velocity (PSV),EDV and MV of ICA were obviously lower in case group at affected side than in the control group (P<0.05 or 0.01).The blood flow of ECA at the affected side was retrograde or bidirectional.PSV,EDV and MV of the external carotid artery (ECA) were obviously higher in the healthy side in patients than in the control group (all P<0.01).There was a significant difference in blood flow velocity of bilateral vertebral artery (VA) between the patients group and the control group,and the PSV,EDV and MV were obviously higher in the former than in the latter (all P<0.01).And the RI was different between the two groups (P<0.05).Conclusions The pressure of ICA is dropped after the common carotid artery occlusion,and then ECA reversely feeds ICA by collateral circulation.The blood flow velocities of bilateral vertebral artery and contralateral ECA are significantly increased to compensate for cerebral blood supply through collateral circulation channels.Color Doppler ultrasonography can reflect this phenomenon more fully and accurately,and provide clinically valuable diagnostic information.
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Objective To survey and analyze characteristics of brucellosis epidemic in Weinan city of Shaanxi province for the purpose of setting up prevention and control measures for the disease. Methods According to "The Executing Plan for the Work of Surveying Brucellosis Disease in Shaanxi Province", 35 villages(towns) of designated monitoring locations and 24 villages (towns) of randomized monitoring locations in five countries of Weinan were chosen to survey brucellosis disease. The five countries were Chengcheng, Dali, Heyang, Tongguan and Hancheng. High risk populations with a history of contacting livestock and livestock products aged between 7 and 60 underwent clinical and serology examination[rose bengal plate agglutination test(RBPT) and standard tube agglutination test(SAT)]. All manipulation methods and judging standards were in accord with the "Diagnostic Standard for Brucellosis" (WS 269-2007). Results In the designated monitoring location, a total of 8664 people at high risk were investigated, among whom 1407 people were tested by RBPT test and 27 people were positive,the positive rate was 1.92%(27/1407); 27 people were tested by SAT test and 27 people were positive, the positive rate was 100% (27/27); 25 people were diagnosed and the diagnosis rate was 92.59%(25/27). In the randomized monitoring location, a total of 3464 people at high risk were investigated, among whom 411 people were tested by RBPT test and 3 people were positive, the positive rate was 0.73%(3/411 ), 3 people were tested by SAT test which were all positive and made a definite diagnosis. Twenty-eight new cases were made a definite diagnosis and its incidence was 2.06 in a hundred thousand(28/1 361 618). Conclusions The infection of human brucellosis in Weinan city stays at higher level. The governments should increase input for the monitoring,investigating and disinfecting to prevent the disease from increasing and outspreading.
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<p><b>OBJECTIVE</b>To construct anti-CD20scFv/CD80/CD28/zeta recombinant gene modified T cells, test its effectiveness of eradicating CD20 positive primary chronic lymphocytic leukemia (CLL) cells and provide a promising tool for tumor adoptive immunotherapy.</p><p><b>METHODS</b>The recombinant vectors were transduced into PA 317 cells and high titer retroviruses were obtained to infect human peripheral blood T lymphocytes. Resistant T cells were obtained by G418 selection for one week. Then transduced T lymphocytes and primary CLL cells were co-cultured. The status of primary chronic lymphocytic leukemia cells were observed by microscope. The level of IL-2 and IFN-gamma in the culture medium were measured.</p><p><b>RESULTS</b>Primary T cells expressing anti-CD20scFv/IgGFc/CD80/CD28/zeta could be constructed successfully. These T cells were able to lyse CD20+ targets and secrete high levels of IL-2 (1301.00 pg/ml) and IFN-gamma (602.18 pg/ml) in vitro.</p><p><b>CONCLUSION</b>(1) Recombinant gene modified T cells can be constructed successfully. (2) Recombinant gene modified T cells can specially kill CD20 positive primary CLL cells in vitro.</p>
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Humains , Antigènes CD20 , Génétique , Antigène CD80 , Génétique , Antigène CD28 , Génétique , Vecteurs génétiques , Immunothérapie adoptive , Interféron gamma , Sécrétions corporelles , Interleukine-2 , Sécrétions corporelles , Leucémie chronique lymphocytaire à cellules B , Anatomopathologie , Retroviridae , Génétique , Lymphocytes T , Allergie et immunologie , Sécrétions corporelles , Transfection , Cellules cancéreuses en cultureRÉSUMÉ
<p><b>OBJECTIVE</b>To investigate the effects of hydroquinone (HQ) on expression of topoisomerase IIα (TOPOIIα) in human bone marrow mononuclear cells, and to explore the role and possible regulatory mechanism of TOPOIIα involved in toxicity of HQ to hematopoietic cells.</p><p><b>METHODS</b>After human bone marrow mononuclear cells were exposed to 50 µmol/L HQ (used the cells which were exposed to sterile distilled water as control); the activity of TOPOII was measured by TOPOII assay kit; the expression levels of TOPOIIα mRNA and protein were detected by RT-PCR technique and Western blotting method respectively; the chromatin immunoprecipitation (ChIP) assay was carried out to study the possible mechanism of TOPOIIα expression changes.</p><p><b>RESULTS</b>(1) The activity of TOPOII was inhibited obviously; the protein and mRNA expression of TOPOIIα were 0.017 ± 0.029 and 0.610 ± 0.128, significantly lower than that in the control with the significant difference (P < 0.01) after treated with HQ for 10 h; (2) The decreased content of TOPOIIα was associated with descended level of histone H4 acetylation than in the control, from 1.198 ± 0.056 to 0.324 ± 0.229, with the significant difference (P < 0.01), without accompanied descended level of histone H3 acetylation, from 1.253 ± 0.045 to 1.177 ± 0.025 (P > 0.05); (3) TOPOIIα mRNA expression decreased gradually after HQ processing, and the chemical modification (histone H4 acetylation) of TOPOIIα promoter happened prior to the mRNA expression.</p><p><b>CONCLUSION</b>HQ could repress the expression of TOPOIIα in human bone marrow mononuclear cells; the change of histone chemical modification plays an important role in the benzene's hematopoietic toxicity.</p>
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Adulte , Femelle , Humains , Mâle , Jeune adulte , Acétylation , Antigènes néoplasiques , Métabolisme , Cellules de la moelle osseuse , Métabolisme , Cellules cultivées , ADN topoisomérases de type II , Métabolisme , Protéines de liaison à l'ADN , Métabolisme , Histone , Métabolisme , Hydroquinones , ToxicitéRÉSUMÉ
<p><b>AIM</b>To construct recombinant retroviruses expressing anti-CD20 scFv/CD80/CD28/zeta gene and detect its expression in Jurkat cells.</p><p><b>METHOD</b>CD28-zetacDNA were amplified from plasmids pBULLET and inserted into pLNCX vector that contained anti-CD20 scFv/CD80 gene. The recombinant plasmids were transfected into PA 317 cells. Retroviruses were harvested from culture medium of PA 317 cells. Then NIH 3T3 were transfected with retroviruses. Objective gene expression was determined by PCR and FACS. Jurkat cells were transfected with high titer of retroviruses and resistant clones were obtained by G418 selection. Objective mRNA was determined by RT- PCR.</p><p><b>RESULTS</b>The recombinant eukaryotic vector was constructed successfully by PCR and enzyme digestion analysis and objective gene was amplified from NIH 3T3 cells transfected with retroviruses by PCR; FACS showed that objective protein could be expressed in NIH 3T3 cells. Objective gene was amplified from Jurkat cells transfected with retroviruses by RT-PCR.</p><p><b>CONCLUSION</b>Recombinant retrovirus expressing anti-CD20 scFv/CD80/CD28/zeta gene was successfully constructed and objective protein could be expressed in Jurkat cells.</p>
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Humains , Antigènes CD20 , Génétique , Métabolisme , Antigène CD80 , Génétique , Métabolisme , Antigène CD28 , Génétique , Métabolisme , Vecteurs génétiques , Fragments Fab d'immunoglobuline , Génétique , Métabolisme , Région variable d'immunoglobuline , Génétique , Métabolisme , Cellules Jurkat , Protéines de fusion recombinantes , Génétique , Métabolisme , Retroviridae , Génétique , Lymphocytes T , Métabolisme , TransfectionRÉSUMÉ
<p><b>OBJECTIVE</b>To construct anti-CD20scFv/CD80/CD28/zeta recombinant gene modified T cells, test its effectiveness of eradicating CD20+ lymphoma cells and provide a probably new approach to tumor adoptive immunotherapy.</p><p><b>METHODS</b>CD28-zeta cDNA were amplified from vector pBULLET and inserted into pLNCX vector that contained anti-CD20scFv/CD80 gene. The recombinant vectors were transduced into PA317 cells and high titer retroviruses were obtained to infect human peripheral blood T lymphocytes. Resistant T cells were obtained by G418 selection at one week. Then transduced T lymphocytes and lymphoma cell lines Daudi Raji were cocultured. The cytotoxicity and cytokine production of transduced T cells were determined by non-radio-activation cytotoxicity assay and ELISA respectively.</p><p><b>RESULTS</b>The recombinant eukaryotic vector was constructed successfully as proved by enzyme digestion analysis and sequencing. These T cells were able to lyse CD20+ target cells and secrete high levels of IL-2 and IFN-gamma in vitro.</p><p><b>CONCLUSION</b>Recombinant gene modified T cells can be constructed successfully. It can specially kill CD20 positive lymphoma cells in vitro.</p>