目的探讨METTL14通过调控巨噬细胞分化抑制宫颈癌病理性发展及相关机制。方法检测宫颈癌病变样本METTL14 m RNA和蛋白,以及IL-6、iNOS、Arg-1和CD206表达变化。PMA诱导THP-1细胞转化为巨噬细胞,慢病毒过表达或抑制METTL14表达,检测IL-6...目的探讨METTL14通过调控巨噬细胞分化抑制宫颈癌病理性发展及相关机制。方法检测宫颈癌病变样本METTL14 m RNA和蛋白,以及IL-6、iNOS、Arg-1和CD206表达变化。PMA诱导THP-1细胞转化为巨噬细胞,慢病毒过表达或抑制METTL14表达,检测IL-6、iNO、Arg-1和CD206表达变化以及PI3K/AKT/GSK3β/β-catenin信号通路相关蛋白表达情况。随后加入PI3K/AKT/GSK3β/β-catenin信号通路激动剂和抑制剂,检测过表达或抑制METTL14后,巨噬细胞IL-6、iNO、Arg-1和CD206表达变化,并取其上清制成条件培养基,孵育Hela细胞,检测细胞凋亡和增殖情况。结果1)宫颈癌病变组织中METTL14 mRNA和蛋白表达降低(P<0.05),巨噬细胞M1型标志物IL-6和iNOS表达明显降低(P<0.05),而M2型标志物Arg-1和CD206表达明显升高(P<0.05)。2)巨噬细胞过表达METTL14后,IL-6和iNOS表达明显升高(P<0.05),而Arg-1和CD206表达明显降低(P<0.05),M1/M2比例升高;抑制METTL14表达后,M1型标志物降低(P<0.05),M2型标志物升高(P<0.05),M1/M2比例降低。3)巨噬细胞中转染OE-METTL14慢病毒组PI3K/AKT/GSK3β/β-catenin信号通路被抑制(P<0.05);加入PI3K/AKT激动剂后,M1型标志物降低而M2型标记物升高(P<0.05),M1/M2比例降低;OE-METTL14可逆转此趋势。Sh-METTL14慢病毒组PI3K/AKT/GSK3β/β-catenin信号通路被激活(P<0.05),加入PI3K/AKT抑制剂后,M1型标志物升高而M2型标记物降低(P<0.05),M1/M2比例升高;Sh-METTL14可逆转此趋势。4)取转染OE-METTL14慢病毒后的巨噬细胞上清培养Hela细胞,可见细胞凋亡明显增多(P<0.05),增殖明显减少(P<0.05)。Sh-METTL14组的Hela细胞则表现出细胞凋亡减少(P<0.05),增殖增多(P<0.05)。结论METTL14通过PI3K/AKT/GSK3β/β-catenin信号通路调控巨噬细胞分化可能有促进宫颈癌细胞凋亡,抑制增殖的作用。展开更多
AIM: To develop a safe and effective agent for cholangiocarcinoma(CCA) chemotherapy. METHODS: A drug combination experiment was conducted to determine the effects of β-escin in c o m b i n a t i o n w i t h c h e m o...AIM: To develop a safe and effective agent for cholangiocarcinoma(CCA) chemotherapy. METHODS: A drug combination experiment was conducted to determine the effects of β-escin in c o m b i n a t i o n w i t h c h e m o t h e ra p y o n C C A c e l l s. 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide assay was performed to determine the effects of β-escin and common chemotherapeutics on the proliferation of human CCA cells(QBC939, Sk-Ch A-1, and MZ-Ch A-1). Immunocytochemistry was used to detect the expression of P-glycoprotein(P-gp) protein. Luciferase reporter assay was used to detect the activation of the Wnt/β-catenin pathway. The protein levels of P-gp, p S9-GSK3β, p T216-GSK3β, GSK3β, β-catenin, and p-β-catenin were further confirmed by western blotting.RESULTS: The drug sensitivity of QBC939 and QBC939/5-fluorouracil(5-FU) cells to 5-FU, vincristine sulfate(VCR), or mitomycin C was significantly enhanced by β-escin compared with either agent alone(P < 0.05). In addition, the combination of β-escin(20 μmol/L) with 5-FU and VCR was synergic with a combination index < 1. Further investigation found that the m RNA and protein expression of P-gp was downregulated by β-escin. Moreover, β-escin induced GSK3β phosphorylation at Tyr-216 and dephosphorylation at Ser-9, resulting in phosphorylation and degradation of β-catenin. Interestingly, activation of the GSK3β/β-catenin pathway induced by Wnt3 a resulted in upregulation of P-gp, which was effectively abolished by β-escin, indicating that β-escin down-regulated P-gp expression in a GSK3β-dependent manner.CONCLUSION: β-escin was a potent reverser of P-gpdependent multidrug resistance, with said effect likely being achieved via inhibition of the GSK3β/β-catenin pathway and thus suggesting a promising strategy of developing combination drugs for CCA.展开更多
Following the publication,concerns have been raised about a number of figures in this article.An unexpected area of similarity was identified in terms of the cellular data,where the results from differently performed ...Following the publication,concerns have been raised about a number of figures in this article.An unexpected area of similarity was identified in terms of the cellular data,where the results from differently performed experiments were intended to have been shown,although the areas immediately surrounding this area featured comparatively different distributions of cells.In addition,the western blots in this article were presented with atypical,unusually shaped and possibly anomalous protein bands in many cases.展开更多
目的探讨驱动蛋白家族成员11(kinesin family member 11,KIF11)、β-连环蛋白(β-catenin)、糖原合成酶激酶-3β(glycogen synthase kinase-3β,GSK-3β)在宫颈癌中的表达情况及临床意义。方法采用免疫组化法检测KIF11、β-catenin、GS...目的探讨驱动蛋白家族成员11(kinesin family member 11,KIF11)、β-连环蛋白(β-catenin)、糖原合成酶激酶-3β(glycogen synthase kinase-3β,GSK-3β)在宫颈癌中的表达情况及临床意义。方法采用免疫组化法检测KIF11、β-catenin、GSK-3β在102例宫颈癌、52例高级别鳞状上皮内病变(high-grade squamous intraepithelial lesion,HSIL)、46例低级别鳞状上皮内病变(low-grade squamous intraepithelial lesion,LSIL)及40例慢性宫颈炎组织中的表达情况,分析三者的表达与宫颈癌患者临床病理特征的关系,分析三者之间的相关性,COX比例风险模型分析影响宫颈癌患者预后的影响因素。结果随着宫颈病变进展,KIF11、β-catenin阳性率逐渐升高,GSK-3β阳性率逐渐减低(P<0.05)。KIF11、β-catenin、GSK-3β在宫颈癌组织中的阳性表达在国际妇产科联盟(International Federation of Gynecology and Obstetrics,FIGO)分期、分化程度、淋巴结转移方面比较,差异有统计学意义(P<0.05),但在不同年龄及病理类型间比较,差异无统计学意义(P>0.05)。宫颈癌患者组织中KIF11与β-catenin的表达呈正相关(r=0.461,P<0.05),β-catenin与GSK-3β的表达呈负相关(r=-0.692,P<0.05),KIF11与GSK-3β的表达呈负相关(r=-0.336,P<0.05)。KIF11、β-catenin阳性表达患者的平均生存时间短于阴性表达患者,GSK-3β阳性表达患者的平均生存时间长于阴性表达患者。COX回归分析显示,FIGO分期、淋巴结转移、KIF11、β-catenin为宫颈癌患者预后的独立危险因素,GSK-3β为独立保护因素。结论KIF11、β-catenin、GSK-3β在宫颈癌患者组织中异常表达,KIF11可能参与宫颈癌发生、发展中Wnt/β-catenin通路的调节,三者联合检测可为宫颈癌的诊断及预后提供新的参考。展开更多
文摘目的探讨METTL14通过调控巨噬细胞分化抑制宫颈癌病理性发展及相关机制。方法检测宫颈癌病变样本METTL14 m RNA和蛋白,以及IL-6、iNOS、Arg-1和CD206表达变化。PMA诱导THP-1细胞转化为巨噬细胞,慢病毒过表达或抑制METTL14表达,检测IL-6、iNO、Arg-1和CD206表达变化以及PI3K/AKT/GSK3β/β-catenin信号通路相关蛋白表达情况。随后加入PI3K/AKT/GSK3β/β-catenin信号通路激动剂和抑制剂,检测过表达或抑制METTL14后,巨噬细胞IL-6、iNO、Arg-1和CD206表达变化,并取其上清制成条件培养基,孵育Hela细胞,检测细胞凋亡和增殖情况。结果1)宫颈癌病变组织中METTL14 mRNA和蛋白表达降低(P<0.05),巨噬细胞M1型标志物IL-6和iNOS表达明显降低(P<0.05),而M2型标志物Arg-1和CD206表达明显升高(P<0.05)。2)巨噬细胞过表达METTL14后,IL-6和iNOS表达明显升高(P<0.05),而Arg-1和CD206表达明显降低(P<0.05),M1/M2比例升高;抑制METTL14表达后,M1型标志物降低(P<0.05),M2型标志物升高(P<0.05),M1/M2比例降低。3)巨噬细胞中转染OE-METTL14慢病毒组PI3K/AKT/GSK3β/β-catenin信号通路被抑制(P<0.05);加入PI3K/AKT激动剂后,M1型标志物降低而M2型标记物升高(P<0.05),M1/M2比例降低;OE-METTL14可逆转此趋势。Sh-METTL14慢病毒组PI3K/AKT/GSK3β/β-catenin信号通路被激活(P<0.05),加入PI3K/AKT抑制剂后,M1型标志物升高而M2型标记物降低(P<0.05),M1/M2比例升高;Sh-METTL14可逆转此趋势。4)取转染OE-METTL14慢病毒后的巨噬细胞上清培养Hela细胞,可见细胞凋亡明显增多(P<0.05),增殖明显减少(P<0.05)。Sh-METTL14组的Hela细胞则表现出细胞凋亡减少(P<0.05),增殖增多(P<0.05)。结论METTL14通过PI3K/AKT/GSK3β/β-catenin信号通路调控巨噬细胞分化可能有促进宫颈癌细胞凋亡,抑制增殖的作用。
基金Supported by National Nature Science Foundation of China,No.81101502the National Science Foundation for Fostering Talents in Basic Research of the National Natural Science Foundation of China,No.J1310027
文摘AIM: To develop a safe and effective agent for cholangiocarcinoma(CCA) chemotherapy. METHODS: A drug combination experiment was conducted to determine the effects of β-escin in c o m b i n a t i o n w i t h c h e m o t h e ra p y o n C C A c e l l s. 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide assay was performed to determine the effects of β-escin and common chemotherapeutics on the proliferation of human CCA cells(QBC939, Sk-Ch A-1, and MZ-Ch A-1). Immunocytochemistry was used to detect the expression of P-glycoprotein(P-gp) protein. Luciferase reporter assay was used to detect the activation of the Wnt/β-catenin pathway. The protein levels of P-gp, p S9-GSK3β, p T216-GSK3β, GSK3β, β-catenin, and p-β-catenin were further confirmed by western blotting.RESULTS: The drug sensitivity of QBC939 and QBC939/5-fluorouracil(5-FU) cells to 5-FU, vincristine sulfate(VCR), or mitomycin C was significantly enhanced by β-escin compared with either agent alone(P < 0.05). In addition, the combination of β-escin(20 μmol/L) with 5-FU and VCR was synergic with a combination index < 1. Further investigation found that the m RNA and protein expression of P-gp was downregulated by β-escin. Moreover, β-escin induced GSK3β phosphorylation at Tyr-216 and dephosphorylation at Ser-9, resulting in phosphorylation and degradation of β-catenin. Interestingly, activation of the GSK3β/β-catenin pathway induced by Wnt3 a resulted in upregulation of P-gp, which was effectively abolished by β-escin, indicating that β-escin down-regulated P-gp expression in a GSK3β-dependent manner.CONCLUSION: β-escin was a potent reverser of P-gpdependent multidrug resistance, with said effect likely being achieved via inhibition of the GSK3β/β-catenin pathway and thus suggesting a promising strategy of developing combination drugs for CCA.
文摘Following the publication,concerns have been raised about a number of figures in this article.An unexpected area of similarity was identified in terms of the cellular data,where the results from differently performed experiments were intended to have been shown,although the areas immediately surrounding this area featured comparatively different distributions of cells.In addition,the western blots in this article were presented with atypical,unusually shaped and possibly anomalous protein bands in many cases.
文摘目的探讨驱动蛋白家族成员11(kinesin family member 11,KIF11)、β-连环蛋白(β-catenin)、糖原合成酶激酶-3β(glycogen synthase kinase-3β,GSK-3β)在宫颈癌中的表达情况及临床意义。方法采用免疫组化法检测KIF11、β-catenin、GSK-3β在102例宫颈癌、52例高级别鳞状上皮内病变(high-grade squamous intraepithelial lesion,HSIL)、46例低级别鳞状上皮内病变(low-grade squamous intraepithelial lesion,LSIL)及40例慢性宫颈炎组织中的表达情况,分析三者的表达与宫颈癌患者临床病理特征的关系,分析三者之间的相关性,COX比例风险模型分析影响宫颈癌患者预后的影响因素。结果随着宫颈病变进展,KIF11、β-catenin阳性率逐渐升高,GSK-3β阳性率逐渐减低(P<0.05)。KIF11、β-catenin、GSK-3β在宫颈癌组织中的阳性表达在国际妇产科联盟(International Federation of Gynecology and Obstetrics,FIGO)分期、分化程度、淋巴结转移方面比较,差异有统计学意义(P<0.05),但在不同年龄及病理类型间比较,差异无统计学意义(P>0.05)。宫颈癌患者组织中KIF11与β-catenin的表达呈正相关(r=0.461,P<0.05),β-catenin与GSK-3β的表达呈负相关(r=-0.692,P<0.05),KIF11与GSK-3β的表达呈负相关(r=-0.336,P<0.05)。KIF11、β-catenin阳性表达患者的平均生存时间短于阴性表达患者,GSK-3β阳性表达患者的平均生存时间长于阴性表达患者。COX回归分析显示,FIGO分期、淋巴结转移、KIF11、β-catenin为宫颈癌患者预后的独立危险因素,GSK-3β为独立保护因素。结论KIF11、β-catenin、GSK-3β在宫颈癌患者组织中异常表达,KIF11可能参与宫颈癌发生、发展中Wnt/β-catenin通路的调节,三者联合检测可为宫颈癌的诊断及预后提供新的参考。
