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dc.contributor.author廖昆
dc.contributor.author夏斌
dc.contributor.author张玉静
dc.contributor.author殷花
dc.contributor.author何承勇
dc.contributor.author林育纯
dc.contributor.author林忠宁
dc.date.accessioned2016-05-17T02:11:25Z
dc.date.available2016-05-17T02:11:25Z
dc.date.issued2014-10-28
dc.identifier.citation热带医学杂志,2014,(10):11-15
dc.identifier.issn1672-3619
dc.identifier.otherRDYZ201410002
dc.identifier.urihttps://dspace.xmu.edu.cn/handle/2288/102256
dc.description.abstract目的初步探讨孕烷X受体(PXr)对黄曲霉毒素b1(Afb1)诱导肝细胞dnA损伤和坏死性凋亡的影响。方法采用已构建的PXr高表达l02-PXr和空白载体对照l02-P b细胞;实时荧光定量PCr(Q rT-PCr)检测细胞nr1I2和CyP3A4 M rnA水平改变;蛋白免疫印迹(WESTErn blOTTIng)检测细胞内PXr和坏死性凋亡下游效应自噬分子lC3-Ⅰ和lC3-Ⅱ蛋白相对表达含量;双核微核试验(CbMn)检测细胞遗传损伤情况;采用噻唑蓝(MTT)法测定Afb1对细胞活性抑制影响;利用坏死性凋亡抑制剂nEC-1构建坏死性凋亡抑制的细胞模型,验证Afb1诱导的坏死性凋亡的效应。结果与l02-P b细胞相比,l02-PXr细胞nr1I2 M rnA和PXr蛋白显著上调(均P<0.001)。Afb1显著地诱导l02-P b和l02-PXr细胞中CyP3A4 M rnA上调(均P<0.05),在l02-PXr细胞中的效应更为明显。与对照组相比,Afb1在5~30μMOl/l呈剂量反应关系诱导l02-P b和l02-PXr细胞的微核率增高(均P<0.05),l02-PXr细胞更为明显;同时,Afb1明显地诱导两株细胞的核芽率和核桥率,但随Afb1剂量增高都有下降趋势。细胞活性随Afb1浓度(1.875~120μMOl/l)增加呈剂量反应关系抑制(均P<0.05);且相对于l02-P b细胞,l02-PXr细胞对Afb1处理48 H诱导的细胞活性抑制作用更为敏感(P<0.05)。nEC-1可显著性抑制Afb1诱导的l02-PXr细胞活性抑制率(P<0.05),然而却不能降低Afb1诱导l02-P b细胞活性抑制率。此外,Afb1显著性诱导l02-P b和l02-PXr坏死性凋亡下游lC3-Ⅱ的上调(均P<0.05);且与l02-P b细胞相比,nEC-1对Afb1诱导的l02-PXr细胞活性抑制和lC3-Ⅱ上调的抑制效果更为明显(P<0.05)。结论 PXr参与Afb1诱导人肝细胞dnA损伤介导的坏死性凋亡,与PXr促进Afb1诱导CyP3A4基因上调有关。
dc.description.abstractObjective To investigate the effects of pregnane X receptor(PXR) over expression on aflatoxin B1(AFB1)-induced DNA damage and necroptosis in human normal liver L02 cells.Methods The established cells models of stable transfection of over expression PXR(L02-PXR) and null vector p Babe-puro(L02-p B) were used.The background levels of NR1I2 m RNA and PXR protein, and the expression of AFB1-induced CYP3A4 m RNA and LC3-I / LC-3II protein were determined by the real time PCR(q RT-PCR) and Western blotting, respectively.The cytokinesis-block micronucleus(CBMN)assay was adopted to evaluate the genotoxicity.The cell viability inhibition rate was determined by MTT assay, after treatment with different doses of AFB1.The inhibition models of necroptosis were established by treatment with necroptosis inhibitor Nec-1.Results The expression of NR1I2 m RNA and PXR protein in L02-PXR cells were higher than that in L02-p B cells(all P<0.001).The level of CYP3A4 m RNA was significantly up regulated in L02-p B and L02-PXR cells by treatment with AFB1(all P<0.05).Compared with control group(Ctrl), MN frequencies in L02-p B and L02-PXR cells were significantly increased by treatment with AFB1 in a dose-dependent manner(all P <0.05), especially, in L02-PXR cells.Meanwhile, NBD and NBP frequencies were significantly increased by treatment with AFB1.However, AFB1 with a higher dose induced downward trends in frequencies of NBD and NBP.Moreover, the inhibition rate of cell viability was increased after treatment with AFB1(1.875~120 μmol / L) in a dose-dependent manner(all P <0.05); specifically, the inhibitory effects of AFB1-treatment after 48 h were significantly stronger in L02-PXR cells than in L02-p B cells(P <0.05).Interestingly, necroptosis inhibitor Nec-1 could inhibit AFB1-induced cell death in L02-PXR cells(P<0.05).On the contrary, Nec-1 could not prevent L02-p B cells from death by treatment with AFB1.In addition, the expression of necroptotic LC3-II, a classical marker of autophagy, was significantly increased by treatment with AFB1 in two cell lines(all P <0.05).Notably, pre-treatment with Nec-1 was able to block the inducement of necroptotic LC3-II in a more efficiently way in L02-PXR cells than in L02-p B cells(P <0.05).Conclusion PXR involved in the effect of AFB1 on necroptosis by DNA damage mediation in human liver cells;specifically, the up regulation of CYP3A4 gene may relate to the AFB1-induced DNA damage.
dc.description.sponsorship国家自然科学基金(81172705;81072334); 广东省自然科学基金(S2011020002769); 福建省自然科学基金(2014J01372)
dc.language.isozh_CN
dc.subject孕烷X受体
dc.subject坏死性凋亡
dc.subject黄曲霉毒素B1
dc.subject肝细胞毒性
dc.subjectpregnane X receptor
dc.subjectnecroptosis
dc.subjectaflatoxin B1
dc.subjecthepatotoxicity
dc.title孕烷X受体参与AFB1诱导的人肝L02细胞坏死性凋亡
dc.title.alternativePregnane X receptor involves in the effect of aflatoxin B1 on necroptosis in human normal L02 liver cells
dc.typeArticle


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