Department of Cell and Molecular Biology, Institute of Acute Abdominal Diseases of Integrated Traditional Chinese and Western Medicine, Tianjin Nankai Hospital, Tianjin 300100, China; Nankai Clinical College, Tianjin Medical University, Tianjin 300107, China.
Center for Reproductive Medicine, Tianjin Central Hospital of Obstetrics and Gynecology, Tianjin 300100, China.
Life Sci. 2019 Jan 15;217:243-250. doi: 10.1016/j.lfs.2018.12.019. Epub 2018 Dec 11.
Pancreatic stellate cells (PSCs) play a critical role in the development of pancreatic fibrosis. Any agents that can affect PSC activation could become potential candidates for treating pancreatic fibrosis. FTY720 can attenuate chronic pancreatic fibrosis by suppressing T-cell infiltration, but its effect on PSCs remains unknown. This study was conducted to investigate the effects of FTY720 on PSC activation in cultured rat PSCs.
The viability of PSCs after FTY720 treatment was detected by MTT. Cell proliferation and migration analysis was performed using the iCELLigence System and a Transwell assay. Cell apoptosis was assessed by flow cytometry, western blot and an activity assay. The mitochondrial membrane potential (MMP) was assessed by JC-1 staining. The expression of α-SMA, collagen I, fibronectin, Beclin-1, Atg5, P62 and LC3B were analysed by immunofluorescence, quantitative real-time PCR and western blot. Rapamycin and phenformin hydrochloride were used to determine whether FTY720 inhibits PSC autophagy by the AMPK/mTOR pathway.
FTY720 supressed PSC viability, proliferation and migration. FTY720 inhibited PSC activation, induced PSC apoptosis and supressed PSC autophagy. We also confirmed that FTY720 inhibited PSC autophagy via the AMPK/mTOR pathway.
Our results indicated that FTY720 inhibited PSC activation by promoting cell apoptosis and inhibiting PSC autophagy by suppressing AMPK and activating the mTOR pathway. These findings may explain the therapeutic mechanisms of FTY720 in treating pancreatic fibrosis and further suggest that targeting autophagy and the related signalling pathways may provide new strategies for the treatment of pancreatic fibrosis.
胰腺星状细胞（PSCs）在胰腺纤维化的发展中起着关键作用。任何能够影响 PSC 激活的药物都可能成为治疗胰腺纤维化的潜在候选药物。FTY720 通过抑制 T 细胞浸润来减轻慢性胰腺纤维化，但它对 PSCs 的影响尚不清楚。本研究旨在探讨 FTY720 对培养的大鼠 PSCs 激活的影响。
用 MTT 检测 FTY720 处理后 PSCs 的活力。使用 iCELLigence 系统和 Transwell 测定法进行细胞增殖和迁移分析。通过流式细胞术、western blot 和活性测定评估细胞凋亡。通过 JC-1 染色评估线粒体膜电位（MMP）。通过免疫荧光、实时定量 PCR 和 western blot 分析 α-SMA、胶原 I、纤维连接蛋白、Beclin-1、Atg5、P62 和 LC3B 的表达。使用雷帕霉素和盐酸苯乙双胍来确定 FTY720 是否通过 AMPK/mTOR 通路抑制 PSC 自噬。
FTY720 抑制 PSC 活力、增殖和迁移。FTY720 抑制 PSC 激活，诱导 PSC 凋亡，抑制 PSC 自噬。我们还证实 FTY720 通过 AMPK/mTOR 通路抑制 PSC 自噬。
我们的结果表明，FTY720 通过促进细胞凋亡和抑制 PSC 自噬来抑制 PSC 激活，抑制 AMPK 并激活 mTOR 通路。这些发现可能解释了 FTY720 在治疗胰腺纤维化中的治疗机制，并进一步表明靶向自噬和相关信号通路可能为胰腺纤维化的治疗提供新策略。
