【摘要】：目的探討苯代謝物氫醌(HQ)對人白血病細胞K562中解旋酶WRN基因mRNA轉(zhuǎn)錄及蛋白表達的影響。方法常規(guī)培養(yǎng)白血病細胞K562至生長對數(shù)期,低劑量HQ組、中劑量HQ組、高劑量HQ組分別以15、30、60μmol/L濃度HQ重復(fù)間隔染毒48及72 h,以等體積的PBS培養(yǎng)的細胞組為空白對照組。采用單細胞凝膠電泳法(SCGE)檢測細胞DNA損傷;采用Taqman探針實時熒光定量PCR法檢測WRN基因mRNA水平,采用免疫印跡分析WRN基因蛋白表達水平,計算mRNA及蛋白的相對表達量。結(jié)果 (1)HQ可導(dǎo)致細胞DNA損傷,且損傷效應(yīng)隨染毒濃度增加而增大,72 h比48 h的DNA損傷程度增加,呈時間-劑量依賴性;(2)HQ染毒48 h,WRN基因mRNA在各組差異均無統(tǒng)計學(xué)意義(P0.05);HQ染毒72 h,各劑量組與空白對照組比較差異有統(tǒng)計學(xué)意義(P0.05),低、中劑量組與高劑量組比較,差異有統(tǒng)計學(xué)意義(P0.05),低劑量組與中劑量組組間差異無統(tǒng)計學(xué)意義(P0.05);(3)HQ染毒48 h,WRN蛋白相對表達量在各組差異無統(tǒng)計學(xué)意義(P0.05);HQ染毒72 h,各劑量組與空白對照組比較,蛋白表達隨著染毒劑量增大而降低,差異有統(tǒng)計學(xué)意義(P0.05),HQ各劑量組之間兩兩比較差異無統(tǒng)計學(xué)意義(P0.05)。結(jié)論 HQ可導(dǎo)致K562細胞DNA損傷,且呈時間-劑量依賴性,其機制可能與HQ下調(diào)WRN基因mRNA轉(zhuǎn)錄及蛋白的表達有關(guān)。
[Abstract]:Objective to investigate the effect of benzene metabolite hydroquinone (HQ) on mRNA transcription and protein expression of helicase WRN gene in human leukemia cell line K562. Methods K562 leukemic cells were cultured from normal culture to logarithmic phase, low dose HQ group, middle dose HQ group and high dose HQ group were exposed to HQ at 1530 ~ 60 渭 mol/L concentration for 48 h and 72 h, respectively. The cells cultured with the same volume of PBS were used as blank control group. Single cell gel electrophoresis (SCGE) was used to detect DNA damage, Taqman probe real-time fluorescence quantitative PCR method was used to detect the mRNA level of WRN gene, and Western blotting was used to analyze the expression level of WRN gene protein, and to calculate the relative expression of mRNA and protein. Results (1) HQ induced DNA damage in cells, and the damage effect increased with the increase of the concentration of DNA. The degree of DNA damage was increased at 72 h than 48 h in a time-dose dependent manner. (2) there was no significant difference in mRNA of WRN gene in 48 h after HQ exposure (P0.05); 72 hours after exposure to HQ, the difference between each dose group and the blank control group was statistically significant (P0.05), and the difference between the middle dose group and the high dose group was statistically significant (P0.05), the difference between the middle dose group and the high dose group was significant (P0.05). There was no significant difference between the low dose group and the middle dose group (P0.05). (3) there was no significant difference in the relative expression of WRN protein in each group at 48 h after exposure to HQ (P0.05); After 72 hours of HQ exposure, the protein expression decreased with the increase of the dose of HQ compared with the blank control group (P0.05). There was no significant difference between the two groups (P0.05). Conclusion HQ can induce DNA damage in K562 cells in a time-dose dependent manner, and its mechanism may be related to the down-regulation of WRN gene mRNA transcription and protein expression by HQ.
【作者單位】： 貴州醫(yī)科大學(xué)醫(yī)學(xué)檢驗學(xué)院;貴陽中醫(yī)學(xué)院第二附屬醫(yī)院檢驗科;
【基金】：國家自然科學(xué)基金資助項目(編號:81360437)
【分類號】：R114

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