Page 63 - 《中国药房》2021年24期

Effects of Sulforaphane on the Proliferation and Apoptosis of Human Renal Tubular Epithelial Cells
Induced by High Glucose and Its Mechanism
ZHOU Lei，AI Wang，CHEN Xiaojuan，WEN Chunyue（Dept. of Nephrology，Wuhan Hospital of Traditional
Chinese Medicine，Wuhan 430010，China）

ABSTRACT OBJECTIVE：To study the effects of sulforaphane on the proliferation and apoptosis of human renal tubular
epithelial cells HK-2 induced by high glucose，and to investigate its mechanism primarily. METHODS：HK-2 cells were divided
into normal group，high glucose group，irbesartan group （positive control，1 μ mol/L），sulforaphane low，medium and high
concentration groups（10，20，40 μmol/L）. The cells in normal group were cultured in DMEM medium for 96 hours. The cells in
other groups were cultured in high glucose DMEM medium （containing 40 mmol/L glucose） for 48 hours. After inducing cell
injury，the cells were added with corresponding drugs for 48 hours. Survival rate and apoptotic rate of cells were detected. mRNA
expression of cyclin D1，caspase-3，Bcl-2 and Bax as well as protein expression of p-mTOR，p-AMPK，p-Akt and p-PI3K were
also determined. In addition，HK-2 cells were divided into normal group，high glucose group，sulforaphane high concentration
group （40 μmol/L），acardicin group （AMPK agonist，1 mmol/L），sulforaphane high concentration+compound C group
（sulforaphane 40 μmol/L+AMPK inhibitor compound C 40 μmol/L），perifoxine group（Akt inhibitor，19.95 μmol/L）、sulforaphane
high concentration + SC79 group（sulforaphane 40 μ mol/L + Akt agonist SC79 4 μ mol/L）. After cultured with the same method，
protein expression of p-mTOR，p-AMPK，p-Akt and p-PI3K were detected in HK-2 cells. RESULTS：Compared with normal
group，survival rate of HK-2 cells，mRNA expression of cyclin D1 and Bcl-2 as well as protein expression of p-AMPK were
decreased significantly in high glucose group （P＜0.05）；apoptotic rate，mRNA expression of caspase-3 and Bax，protein
expression of p-mTOR，p-Akt and p-PI3K in HK-2 cells were increased significantly （P＜0.05）. Compared with high glucose
group， above indexes of sulforaphane low， medium and high concentration groups， irbesartan group were all improved
significantly （P＜0.05）； the improvement of above indexes in sulforaphane medium and high concentration groups were
significantly better those of sulforaphane low concentration group（P＜0.05）. There was no significant difference in above indexes
between sulforaphane high concentration group and irbesartan group（P＞0.05）. Compared with sulforaphane high concentration
group，there were no significant difference in the protein expression of p-AMPK，p-mTOR in acardicin group and p-mTOR，p-Akt
and p-PI3K in perifoxine group（P＞0.05）；the protein expression of p-AMPK in sulforaphane high concentration+compound C
group was decreased significantly（P＜0.05），while the protein expression of p-mTOR was increased significantly（P＜0.05）；the
protein expression of p-mTOR、p-Akt、p-PI3K in sulforaphane high concentration+SC79 group were increased significantly（P＜
0.05）. CONCLUSIONS：Sulforaphane can promote the proliferation of renal tubular epithelial cells and inhibit its apoptosis；its
mechanism may be associated with up-regulating the expression of p-AMPK and down-regulating the expression of p-mTOR，p-Akt
and p-PI3K.
KEYWORDS Sulforaphane；High glucose；Human renal tubular epithelial cells；PI3K/Akt signaling pathway；AMPK/mTOR
signaling pathway；Proliferation；Apoptosis

糖尿病肾病是糖尿病的常见并发症，是严重危害人 凋亡的途径之一 。此外，萝卜硫素还可通过激活 Akt
[7]
类身体健康的重大疾病之一。有研究表明，持续的高糖 来增强Nrf2介导的外源抗氧化防御，且Akt和腺苷一磷
环境可导致细胞凋亡，是肾小管上皮细胞损伤的重要原 酸活化蛋白激酶（AMPK）的磷酸化可以下调哺乳动物
因之一 。而肾小管上皮细胞损伤是糖尿病肾病的关键 雷帕霉素靶蛋白（mTOR）表达，进而诱导细胞自噬 。
[1]
[8]
诱发因素，亦是导致糖尿病肾病转向终末期的重要原 基于此，本研究拟通过建立高糖诱导的肾小管上皮细胞
因 [2－3] 。萝卜硫素是一种天然的单体物质，广泛分布于 损伤模型，考察萝卜硫素对其增殖、凋亡的影响，并基于
十字花科植物中。有研究指出，萝卜硫素在缓解2型糖 PI3K/Akt、AMPK/mTOR 双信号通路初步探讨其可能的
尿病患者胰岛素抵抗方面的效果较好，但其机制尚不明 作用机制，旨在为阐明萝卜硫素治疗糖尿病肾病的机制
[4]
确 。有研究报道，萝卜硫素可通过抑制核因子 E2 相 提供参考。
关因子 2（Nrf2）/血红素加氧酶 1 信号通路来缓解过氧 1 材料
化氢（H2O2）诱导的肾小管上皮细胞氧化应激损伤，而 1.1 主要仪器
氧化应激损伤是促进肾脏组织细胞凋亡和坏死的因素 本研究所用主要仪器包括：Multiskan SkyHigh型全
之一 [5－6] 。 波长酶标仪、VeritiPro 96-Well 型聚合酶链反应（PCR）
已有研究表明，磷脂酰肌醇3激酶（PI3K）/蛋白激酶 仪、Form Steri-Cycle 型 CO2 培养箱（美国 Thermo Fisher
B（Akt）信号通路是萝卜硫素及其衍生物促进肿瘤细胞 Scientific 公 司），Mini-Protean 型 小 垂 直 板 电 泳 槽 、

中国药房 2021年第32卷第24期 China Pharmacy 2021 Vol. 32 No. 24 ·3001 ·