| 丁美玲 冯文强 张 松 曹海超 聂勇战.RNA 特异腺苷脱氨酶 1（ p150 亚型）抑制肝细胞脂质合成[J].,2015,15(5):829-833 |
| RNA 特异腺苷脱氨酶 1（ p150 亚型）抑制肝细胞脂质合成 |
| RNA-specific Adenosine Deaminase 1 p150 Isoform Inhibits Lipid Synthesisin Hepatocytes |
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| DOI： |
| 中文关键词: ADAR1 -p150 脂质合成 高内 涵筛选系统 |
| 英文关键词: ADAR1 -p150 Lipid synthesis High content screening system |
| 基金项目:科技部重大国际合作项目（ 811 10200） |
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| 中文摘要: |
| 目 的： 在油酸诱导的肝细胞脂肪变模型中，检测 RNA 特异腺苷脱氨酶 1 p1 50亚型（ ADAR1-p1 50）高表达细胞系中脂肪合成
的变化。方法： 利用 本课题组前期摸索的油酸刺激人胚胎肝细胞 L-02 细胞系脂肪变的条件， qRT-PCR 和 Western-Blot 检测油酸刺
激组和对照 组 ADAR1 -p150表达变 化； 将构 建成功 的 ADAR1 -p150过表达慢病毒载体 GV166-ADAR1 -p150及空 载体病毒
GV166-control 感染 L-02 细胞， 检测感染细胞中 ADAR1 -p150的 mRNA 和蛋白 表达水平； 通过油红 O 染色和 BODIPY 染色观察
L-02 ADAR1-p150和 L-02 control 细胞中脂滴形成， 并进一步利用 高内 涵系统检测其荧光强度，对脂滴合成作定量分析。 结果：
L-02 细胞在 油酸刺 激 后 ADAR1 -p150 的 mRNA 和蛋白 水平 降低；成 功 构建 ADAR1 -p150 过表达 慢 病毒载体 GV166-
ADAR1 -p150及空 载体病毒 GV1 66-control， qRT-PCR 及 Western-Blot 检测 显示病毒转染 GV1 66- ADAR1 -p1 50后 ADAR1 -p150
在细胞中的表达水平显著升高；油红 O 染色和 BODIPY 染色发现 L-02 ADAR1 -p150较 L-02 control 细胞胞质中脂滴数量减少。 高
内 涵筛选系统检测提示 L-02 ADAR1-p1 50组中脂滴的荧光强度明显较 L-02 control 组低。 结论： 成功构建 ADAR1-p1 50过表达稳
定转染 L-02 细胞系， 并证实高表达 ADAR1 -p150能够抑制脂肪合成。 |
| 英文摘要: |
| Objective:To investigate the regulatory effect of RNA-specific adenosine deaminase 1 p150 isoform (ADAR1 -p150)
on fat synthesis in oleic acid-induced hepatic steatosis cell model.Methods:Oleic acid stimulated steatosis model in human embryonic
liver cell line L-02 cells was constructed according to our previously reported protocols. ADAR1 -p150 expression was analyzed by
qRT-PCR and Western-Blot in both control group and oleic acid (OA) group. ADAR1-p1 50 overexpression lentiviral vector
GV166-ADAR1 -p150 and control lentiviral vector GV166-control were constructed to infect L-02 cell line, the mRNA and protein expression levels of ADAR1 -p150 were detected by qRT-PCR and Western-Blot. The lipid droplets formation of L-02 ADAR1 -p150 and
L-02 control cell lines was observed by oil red O staining and BODIPY staining. High Content Screening system was performed to detect
the average fluorescence intensity of lipid droplets.Results:The mRNA and protein expression levels of ADAR1-p1 50 were down-regulated in OA group as compared with that in control group. ADAR1-p1 50 overexpression lentiviral vector GV1 66- ADAR1 -p150 and
empty vector virus GV166-control were successfully constructed. qRT-PCR and Western-Blot analysis showed that ADAR1 -p150 expression significantly increased in GV166- ADAR1 -p150 treated cells. Oil Red O staining and BODIPY staining revealed that the number of lipid droplets in L-02 ADAR1 -p150 cells was less than that in L-02 control cells. High content screening system demonstrated that
the lipid droplets fluorescence intensity of L-02 ADAR1-p1 50 cells was significantly lower than that of L-02 control cells.Conclusion:Stable ADAR1 -p150 overexpressing cell line was successfully established. More importantly, high expression of ADAR1-p1 50 was capable of inhibiting fat synthesis in L-02 cells. |
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