Experimental and Translational Research Center, Beijing Friendship Hospital, Capital Medical University, Beijing, P.R. China.
Beijing Clinical Medicine Institute, Beijing, P.R. China.
FASEB J. 2021 Oct;35(10):e21918. doi: 10.1096/fj.202100374R.
The role of LOXL1 in fibrosis via mediating ECM crosslinking and stabilization is well established; however, the role of hepatic stellate cells (HSCs)-specific LOXL1 in the development of fibrosis remains unknown. We generated HSCs-specific Loxl1-depleted mice (Loxl1 mice) to investigate the HSCs-specific contribution of LOXL1 in the pathogenesis of fibrosis. Loxl1 mice were used as the control. Furthermore, we used RNA sequencing to explore the underlying changes in the transcriptome. Results of the sirius red staining, type I collagen immunolabeling, and hydroxyproline content analysis, coupled with the reduced expression of profibrogenic genes revealed that Loxl1 mice with CCl -induced fibrosis exhibited decreased hepatic fibrosis. In addition, Loxl1 mice exhibited reduced macrophage tissue infiltration by CD68-positive cells and decreased expression of inflammatory genes compared with the controls. RNA sequencing identified integrin α8 (ITGA8) as a key modulator of LOXL1-mediated liver fibrosis. Functional analyses showed that siRNA silencing of Itga8 in cultured fibroblasts led to a decline in the LOXL1 expression and inhibition of fibroblast activation. Mechanistic analyses indicated that LOXL1 activated the FAK/PI3K/AKT/HIF1a signaling pathway, and the addition of inhibitors of FAK or PI3K reversed these results via downregulation of LOXL1. Furthermore, HIF1a directly interacted with LOXL1 and upregulated its expression, indicating that LOXL1 can positively self-regulate by forming a positive feedback loop with the FAK/PI3K/AKT/HIF1a pathway. We demonstrated that HSCs-specific Loxl1 deficiency prevented fibrosis, inflammation and that ITGA8/FAK/PI3K/AKT/HIF1a was essential for the function and expression of LOXL1. Knowledge of this approach can provide novel mechanisms and targets to treat fibrosis in the future.
LOXL1 在通过介导细胞外基质交联和稳定化来调节纤维化方面的作用已得到充分证实；然而，肝星状细胞（HSCs）特异性 LOXL1 在纤维化发展中的作用仍不清楚。我们构建了 HSCs 特异性 Loxl1 敲除小鼠（Loxl1 小鼠），以研究 LOXL1 在纤维化发病机制中的 HSCs 特异性作用。将 Loxl1 小鼠作为对照。此外，我们还使用 RNA 测序来探索转录组的潜在变化。天狼星红染色、I 型胶原免疫标记和羟脯氨酸含量分析的结果，以及纤维化基因表达的下调表明，CCl4 诱导纤维化的 Loxl1 小鼠表现出肝纤维化减少。此外，与对照组相比，Loxl1 小鼠的巨噬细胞组织浸润减少，炎症基因的表达降低。RNA 测序确定整合素 α8（ITGA8）是 LOXL1 介导的肝纤维化的关键调节剂。功能分析表明，在培养的成纤维细胞中沉默 Itga8 会导致 LOXL1 表达下降和成纤维细胞激活抑制。机制分析表明，LOXL1 激活了 FAK/PI3K/AKT/HIF1a 信号通路，而 FAK 或 PI3K 的抑制剂可通过下调 LOXL1 逆转这些结果。此外，HIF1a 直接与 LOXL1 相互作用并上调其表达，表明 LOXL1 可以通过与 FAK/PI3K/AKT/HIF1a 通路形成正反馈环来自我正向调节。我们证明 HSCs 特异性 Loxl1 缺失可预防纤维化、炎症，并且 ITGA8/FAK/PI3K/AKT/HIF1a 对于 LOXL1 的功能和表达是必需的。了解这种方法可以为未来治疗纤维化提供新的机制和靶点。
