去甲肾上腺素通过PI3K/Akt信号通路上调Sox4表达促进肝癌细胞增殖

Norepinephrine promotes proliferation of hepatocellular carcinoma cells through enhancing Sox4 expression via PI3K/Akt signal pathway

目的探讨去甲肾上腺素对肝癌细胞自我更新能力的影响及其分子机制。方法实时荧光定量PCR和Western blot检测去甲肾上腺素对肝癌细胞(Huh7、PLC)中CD133表达水平的影响;肿瘤细胞球形成实验以及克隆形成实验检测去甲肾上腺素和哌唑嗪对肝癌细胞自我更新能力的作用;实时荧光定量PCR和Western blot检测去甲肾上腺素(10μmol/L)和哌唑嗪(5μmol/L)对肝癌细胞中干性相关基因表达的影响,并检测相关的信号通路。结果去甲肾上腺素使Huh7和PLC肝癌细胞中的CD133表达升高;去甲肾上腺素处理Huh7和PLC肝癌细胞后的成球率较对照组明显上升,哌唑嗪处理后明显降低(P<0.05);在克隆形成实验中,去甲肾上腺素处理Huh7和PLC肝癌细胞的后,克隆形成率高于哌唑嗪组和对照组(P<0.05);与对照组和哌唑嗪组相比,去甲肾上腺素组中Sox4基因表达明显升高,同时p-Akt表达相较于哌唑嗪和对照组相比也明显上调(P<0.05)。结论去甲肾上腺素激动肝癌细胞中α肾上腺素能受体,通过激活PI3K/Akt信号通路上调肝癌细胞中Sox4基因表达,从而提高肝癌细胞自我更新的能力。

Objective To determine the effect of norepinephrine on self-renewal capacity in hepatocellular carcinoma （HCC） cells and investigate the molecular mechanism. Methods Quantitative RT- PCR and Western blotting were used to detect the expression level of CD133 in the HCC cell lines Huh7 and PLC after the treatment of 10 Ixmol/L norepinephrine. Sphere and clone formation assays were used to investigate the impact of norepinephrine on self-renewal capacity of HCC cells. The effects of norepinephrine （10 μmol/L） and prazosin （5 μmol/L） on the expression of stem related genes and signal pathways were detected by RT-PCR and Western blotting. Results Norepinephrine promoted the expression of CD133 in Huh7 and PLC cells. Norepinephrine facilitated sphere and clone formation in the 2 cell lines, and prazosin blocked the effect of norepinephrine （ P 〈 0.05 ）. There were more sphere and clone formation in the cells after norepinephrine treatment than control cells and prazosin group （ P 〈 0.05 ）. Higher level of Sox4 and upregulation of p-Akt were observed in the norepinephrine-treated cells than prazosin-treated and control HCC cells （ P 〈 0. 05 ）. Conclusion Norepinephrine activates α-adrenergie receptor and up-regulates Sox4 expression via activating PI3K/Akt signal pathway, and thus improves self-renewal capacity in HCC cells.