Department of Hematology, The First Affiliated Hospital of Kunming Medical University, Hematology Research Center of Yunnan Province, Kunming, 650032, China.
Fujian Medical University, Fujian Institute of Hematology, Fujian Provincial Key Laboratory on Hematology, Fujian Medical University Union Hospital, Fuzhou, 350001, China.
Biochem Biophys Res Commun. 2020 Jan 8;521(2):434-440. doi: 10.1016/j.bbrc.2019.09.129. Epub 2019 Oct 29.
In this study, we aimed to explore the role of liver kinase b1 (Lkb1) in the biological characteristics and immune regulation of amniotic mesenchymal stem cells (AMSCs). AMSCs were identified via the cell surface markers using flow cytometry. We knocked down the expression of Lkb1 in AMSCs using lentivirus-mediated Lkb1-specific shRNA. The efficiency of the knockdown was detected by flow cytometry, RT-qPCR, and western blot. The AMSC-related phenotype was determined by flow cytometric analysis via staining surface markers. Fibroblast colony-forming cells (CFU-F) assay and Ki-67 intracellular staining assay were used to determine the proliferative capacity. The differentiated and immunosuppressive capabilities were determined by conditional induction of differentiation and co-culture experiments. We observed that AMSCs along with Lkb1 knockdown (AMSCs-Lkb1) displayed similar cellular morphology and surface antigen expression patterns as those observed in AMSCs. However, AMSCs-Lkb1 exhibited an enhanced differentiation capacity towards osteogenesis and chondrogenesis while it showed defective proliferation and increased apoptosis. Furthermore, AMSCs-Lkb1 showed an enhanced immunosuppressive capacity by directly inhibiting conventional T cells and indirectly inducing production of regulatory T cells (Treg). Interestingly, Treg produced by AMSCs-Lkb1 displayed stronger proliferative capacity as compared to those produced by AMSCs. Our results indicate that Lkb1 plays a vital role in maintaining self-renewal of AMSCs and regulating immune equivalence, and may hold potential for the clinical management of diseases such as GVHD.
在这项研究中，我们旨在探讨肝激酶 b1（Lkb1）在羊膜间充质干细胞（AMSCs）的生物学特性和免疫调节中的作用。通过流式细胞术使用细胞表面标志物鉴定 AMSCs。我们使用慢病毒介导的 Lkb1 特异性 shRNA 敲低 AMSCs 中的 Lkb1 表达。通过流式细胞术、RT-qPCR 和 Western blot 检测敲低效率。通过表面标志物染色的流式细胞术分析确定 AMSC 相关表型。成纤维细胞集落形成细胞（CFU-F）测定和 Ki-67 细胞内染色测定用于确定增殖能力。通过条件诱导分化和共培养实验确定分化和免疫抑制能力。我们观察到，与 Lkb1 敲低的 AMSCs（AMSCs-Lkb1）相比，AMSCs-Lkb1 显示出与 AMSCs 相似的细胞形态和表面抗原表达模式。然而，AMSCs-Lkb1 表现出增强的成骨和成软骨分化能力，同时表现出增殖缺陷和凋亡增加。此外，AMSCs-Lkb1 通过直接抑制常规 T 细胞和间接诱导调节性 T 细胞（Treg）的产生来增强免疫抑制能力。有趣的是，与 AMSCs 产生的 Treg 相比，AMSCs-Lkb1 产生的 Treg 显示出更强的增殖能力。我们的结果表明，Lkb1 在维持 AMSCs 的自我更新和调节免疫平衡方面发挥着重要作用，并且可能为 GVHD 等疾病的临床管理提供潜在的治疗靶点。
