自然杀伤细胞来源的外泌体 miR-223 通过抑制自噬抑制肝星状细胞激活。
Exosomal miR-223 derived from natural killer cells inhibits hepatic stellate cell activation by suppressing autophagy.
机构信息
Department of Infectious Diseases, Xiangya Hospital of Central South University, No. 87 Xiangya Road, Changsha, 410008, Hunan, China.
Department of Ophthalmology, The Second Xiangya Hospital of Central South University, Changsha, 410011, Hunan, China.
出版信息
Mol Med. 2020 Sep 1;26(1):81. doi: 10.1186/s10020-020-00207-w.
BACKGROUND
Activation of hepatic stellate cells (HSCs) is a prominent driver of liver fibrosis. We previously demonstrated that exosomes derived from natural killer (NK) cells (NK-Exo) attenuated TGF-β1-induced HSC activation. Herein, this study was designed to investigate the mechanism underlying the action of NK-Exo.
METHODS
NK-Exo was isolated from NK-92MI cells and then administered into TGF-β1-treated LX-2 (human HSC line) cells. MiR-223 expression in NK-Exo was downregulated by transfecting NK-92MI cells with miR-223 inhibitor followed by exosome isolation. The HSC activation was evaluated by determining cell proliferation using CCK-8 assay and measuring the protein levels of α-SMA and CoL1A1 using western blot in LX-2 cells. The expression of miR-223 was detected by qRT-PCR. The interaction between miR-223 and ATG7 was analyzed by a dual-luciferase activity assay. The autophagy was evaluated by measuring the autophagy-related proteins using western blot.
RESULTS
miR-223 was highly expressed in NK-Exo and inhibition of miR-223 expression in NK-Exo abrogated the inhibitory effect of NK-Exo on TGF-β-induced HSC activation. ATG7 was confirmed as a direct target of miR-223. Furthermore, treatment with the autophagy activator rapamycin and ATG7 overexpression in LX-2 cells abolished the HSC activation-suppressive effect of NK-Exo.
CONCLUSION
NK-Exo attenuated TGF-β-induced HSC activation by transferring miR-223 that inhibited autophagy via targeting ATG7.
背景
肝星状细胞（HSCs）的激活是肝纤维化的主要驱动因素。我们之前的研究表明，自然杀伤（NK）细胞衍生的外泌体（NK-Exo）可减轻 TGF-β1 诱导的 HSC 激活。在此，本研究旨在探讨 NK-Exo 作用的机制。
方法
从 NK-92MI 细胞中分离 NK-Exo，然后将其给予 TGF-β1 处理的 LX-2（人 HSC 系）细胞。通过用 miR-223 抑制剂转染 NK-92MI 细胞，然后进行外泌体分离，下调 NK-Exo 中的 miR-223 表达。通过 CCK-8 测定法测定细胞增殖，通过 Western blot 测定 LX-2 细胞中 α-SMA 和 CoL1A1 的蛋白水平来评估 HSC 激活。通过 qRT-PCR 检测 miR-223 的表达。通过双荧光素酶活性测定分析 miR-223 与 ATG7 之间的相互作用。通过 Western blot 测定评估自噬。
结果
miR-223 在 NK-Exo 中高表达，并且抑制 NK-Exo 中的 miR-223 表达可消除 NK-Exo 对 TGF-β 诱导的 HSC 激活的抑制作用。ATG7 被确认为 miR-223 的直接靶标。此外，在 LX-2 细胞中用自噬激活剂雷帕霉素处理和过表达 ATG7 可消除 NK-Exo 对 HSC 激活抑制作用。
结论
NK-Exo 通过转移 miR-223 减轻 TGF-β 诱导的 HSC 激活，miR-223 通过靶向 ATG7 抑制自噬。
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