Summary: In order to explore the role of activator protein-1 (AP-1) in the transcription of interleukin-5 (IL-5) gene regulated by protein kinase C (PKC) signal in peripheral blood T lymphocytes from asthmatic patient...Summary: In order to explore the role of activator protein-1 (AP-1) in the transcription of interleukin-5 (IL-5) gene regulated by protein kinase C (PKC) signal in peripheral blood T lymphocytes from asthmatic patient, T lymphocytes were isolated and purified from peripheral blood of each asthmatic patient. The T lymphocytes were randomly divided into 4 groups: group A (blank control), group B (treated with PKC agonist phorbol 12-myristate 13-acetate (PMA)), Group C (treated with PMA and AP-1 cis-element decoy oligodeoxynucleotides (decoy ODNs)), and group D (treated with PMA and AP-1 mutant decoy ODNs). The ODNs were transfected into the T cells of group C and D by cation liposome respectively. Reverse transcription-polymerase chain reaction (RT-PCR) was employed to assess IL-5 mRNA expression, and electrophoretic mobility shift assays (EMSA) for the activation of AP-1. The results showed that the activation of AP-1 (88 003.58±1 626.57) and the expression of IL-5 mRNA (0.8300±0.0294) in T lymphocytes stimulated with PMA were significantly higher than these in blank control (20 888.47±1103.56 and 0.3050±0.0208, respectively, P< 0.01), while the indexes (23 219.83±1 024.86 and 0.3425±0.0171 respectively) of T lymphocytes stimulated with PMA and AP-1 decoy ODNs were significantly inhibited, as compared with group B (P< 0.01). The indexes (87 107.41±1 342.92 and 0.8225±0.0222, respectively) in T lymphocytes stimulated with PMA and AP-1 mutant decoy ODNs did not exhibit significant changes, as compared with group B (P>0.05). The significant positive correlation was found between the activation of AP-1 and the expression of IL-5 mRNA (P< 0.01). It was concluded that AP-1 might participate in the signal transduction of PKC-triggered transcription of IL-5 gene in asthmatic T lymphocytes. This suggests the activation of PKC/AP-1 signal transduction cascade of T lymphocytes may play an important role in the pathogenesis of asthma.展开更多
AIM: To investigate the inhibitory effect of heparin-derived oligosaccharides (Oligs) on secretion of interleukin-4 (IL-4) and interleukin-5 (IL-5) from human peripheral blood T lymphocytes (PBTLs).METHODS: Oligs were...AIM: To investigate the inhibitory effect of heparin-derived oligosaccharides (Oligs) on secretion of interleukin-4 (IL-4) and interleukin-5 (IL-5) from human peripheral blood T lymphocytes (PBTLs).METHODS: Oligs were prepared by three different heparin depolymerization methods and separated by gel filtration chromatography. PBTLs from ten adult patients with allergic eosinophilic gastroenteritis were treated with phytahematoagglutinin (PHA) and Oligs. The supernatants from the cell culture of PBTLs were harvested and subjected to the determination of IL-4 and IL-5 contents by ELISA method.RESULTS: At the concentration of 5μg/mL, Oligs with different Mr had different effects on the secretion of IL-4 and IL-5. The tetrasaccharide with Mr of 1 142, produced by depolymerizing heparin with hydrogen peroxide, had the strongest inhibitory effect on the secretion of IL-4. It decreased the IL-4 content from 375.6±39.2 ng/L (PHA group) to 12.5±5.7 ng/L (P<0.01). The hexasaccharide with Mr of I 806, produced by depolymerizing heparin with βelimination method, had the strongest inhibitory effect on the secretion of IL-5. It decreased the IL-5 content from 289.2±33.4 ng/L (PHA group) to 22.0±5.2 ng/L (P<0.01).CONCLUSION: The inhibitory activity of Oligs on the secretion of IL-4 and IL-5 from human PBTLs closely depends on their molecular structure, and there may be an essential structure to act as an inhibitor. The most effective inhibitors of IL-4 and IL-5 secretion are tetrasaccharides and hexasaccharides, respectively.展开更多
目的探讨甲基转移酶5(methyltransferase-like 5,METTL5)在三阴乳腺癌(triple-negative breast cancer,TNBC)中的作用和潜在机制。方法采用免疫组织化学方法和Western blot检测TNBC肿瘤组织和细胞系中METTL5的表达情况。用靶向METTL5的s...目的探讨甲基转移酶5(methyltransferase-like 5,METTL5)在三阴乳腺癌(triple-negative breast cancer,TNBC)中的作用和潜在机制。方法采用免疫组织化学方法和Western blot检测TNBC肿瘤组织和细胞系中METTL5的表达情况。用靶向METTL5的shRNA(shRNA-METTL5)转染TNBC细胞后,用CCK-8、集落形成、伤口愈合以及Transwell实验分别检测细胞增殖活性、迁移与侵袭,Western blot检测Wnt/β-catenin信号关键蛋白的表达。构建异种移植瘤模型,验证敲降METTL5对TNBC细胞在体内生长以及Wnt/β-catenin信号活性的影响。结果METTL5在TNBC肿瘤组织和细胞系中表达上调(P<0.01)。敲降METTL5可抑制TNBC细胞的增殖、迁移和侵袭并降低了Wnt/β-catenin信号分子β-catenin、细胞周期蛋白(Cyclin)D1、基质金属蛋白酶(MMP)-2和MMP-7的表达(均P<0.01)。体内实验显示,敲降METTL5减缓了移植瘤生长和Wnt/β-catenin信号活性。结论敲降METTL5能抑制TNBC细胞的增殖、迁移与侵袭,其作用可能与抑制Wnt/β-catenin信号通路有关。展开更多
文摘Summary: In order to explore the role of activator protein-1 (AP-1) in the transcription of interleukin-5 (IL-5) gene regulated by protein kinase C (PKC) signal in peripheral blood T lymphocytes from asthmatic patient, T lymphocytes were isolated and purified from peripheral blood of each asthmatic patient. The T lymphocytes were randomly divided into 4 groups: group A (blank control), group B (treated with PKC agonist phorbol 12-myristate 13-acetate (PMA)), Group C (treated with PMA and AP-1 cis-element decoy oligodeoxynucleotides (decoy ODNs)), and group D (treated with PMA and AP-1 mutant decoy ODNs). The ODNs were transfected into the T cells of group C and D by cation liposome respectively. Reverse transcription-polymerase chain reaction (RT-PCR) was employed to assess IL-5 mRNA expression, and electrophoretic mobility shift assays (EMSA) for the activation of AP-1. The results showed that the activation of AP-1 (88 003.58±1 626.57) and the expression of IL-5 mRNA (0.8300±0.0294) in T lymphocytes stimulated with PMA were significantly higher than these in blank control (20 888.47±1103.56 and 0.3050±0.0208, respectively, P< 0.01), while the indexes (23 219.83±1 024.86 and 0.3425±0.0171 respectively) of T lymphocytes stimulated with PMA and AP-1 decoy ODNs were significantly inhibited, as compared with group B (P< 0.01). The indexes (87 107.41±1 342.92 and 0.8225±0.0222, respectively) in T lymphocytes stimulated with PMA and AP-1 mutant decoy ODNs did not exhibit significant changes, as compared with group B (P>0.05). The significant positive correlation was found between the activation of AP-1 and the expression of IL-5 mRNA (P< 0.01). It was concluded that AP-1 might participate in the signal transduction of PKC-triggered transcription of IL-5 gene in asthmatic T lymphocytes. This suggests the activation of PKC/AP-1 signal transduction cascade of T lymphocytes may play an important role in the pathogenesis of asthma.
基金Supported by the National Natural Science Foundation of China,No.30370613 and the Major state Basic Research Development Program of China,2001CCA01600
文摘AIM: To investigate the inhibitory effect of heparin-derived oligosaccharides (Oligs) on secretion of interleukin-4 (IL-4) and interleukin-5 (IL-5) from human peripheral blood T lymphocytes (PBTLs).METHODS: Oligs were prepared by three different heparin depolymerization methods and separated by gel filtration chromatography. PBTLs from ten adult patients with allergic eosinophilic gastroenteritis were treated with phytahematoagglutinin (PHA) and Oligs. The supernatants from the cell culture of PBTLs were harvested and subjected to the determination of IL-4 and IL-5 contents by ELISA method.RESULTS: At the concentration of 5μg/mL, Oligs with different Mr had different effects on the secretion of IL-4 and IL-5. The tetrasaccharide with Mr of 1 142, produced by depolymerizing heparin with hydrogen peroxide, had the strongest inhibitory effect on the secretion of IL-4. It decreased the IL-4 content from 375.6±39.2 ng/L (PHA group) to 12.5±5.7 ng/L (P<0.01). The hexasaccharide with Mr of I 806, produced by depolymerizing heparin with βelimination method, had the strongest inhibitory effect on the secretion of IL-5. It decreased the IL-5 content from 289.2±33.4 ng/L (PHA group) to 22.0±5.2 ng/L (P<0.01).CONCLUSION: The inhibitory activity of Oligs on the secretion of IL-4 and IL-5 from human PBTLs closely depends on their molecular structure, and there may be an essential structure to act as an inhibitor. The most effective inhibitors of IL-4 and IL-5 secretion are tetrasaccharides and hexasaccharides, respectively.