转录因子KLF4介导纤溶酶原驱动的骨髓来源单核/巨噬细胞向疤痕相关巨噬细胞的转化

Transcription Factor KLF4 Mediates the Transformation of Scar-Associated Macrophages Driven by Plasminogen from Bone Marrow-Derived Monocytes/Macrophages

疤痕相关巨噬细胞(scar-associated macrophage,SAM)是肝纤维化的重要参与者,纤溶酶原(plasminogen,PLG)可驱动巨噬细胞获得SAM表型,但其分子机制尚不明确。该研究旨在探讨PLG驱动SAM表型转化的机制。分析已发表的正常、肝纤维化小鼠肝非实质细胞的单细胞测序数据,发现成熟SAM高表达转录因子Krüppel样因子4(Krüppel-like factor 4,KLF4)。分离小鼠原代骨髓来源单核/巨噬细胞,使用PLG处理以促进SAM转化,转染特异性siRNA/过表达质粒以敲减/过表达巨噬细胞中的KLF4,使用流式细胞术和Western blot检测蛋白表达情况,免疫荧光检测KLF4的亚细胞定位,实时荧光定量聚合酶链反应(real time quantitative polymerase chain reaction,RT-qPCR)检测mRNA表达情况。结果表明在PLG驱动SAM转化过程中,KLF4蛋白水平升高,且其定位在细胞核中。特异性敲减小鼠巨噬细胞中的KLF4表达能够抑制PLG驱动的SAM转化,SAM标志基因、功能基因的表达,SAM比例及诱导HSC产生细胞外基质的能力均下降;KLF4过表达则上调了SAM标志基因和功能基因的表达。综上所述,KLF4参与了PLG诱导的SAM表型转化。

SAM(scar-associated macrophage)plays an important role in liver fibrogenesis.PLG(plasminogen)drives the transformation of macrophage to SAM,but the underlying mechanism is still unclear.The aim of this study is to explore the mechanism underlying PLG-driven SAM transformation.scRNA-seq(single-cell RNA sequencing)data were analyzed.The results showed that transcriptional factor KLF4(Krüppel-like factor 4)was highly expressed by SAM.Isolated mouse primary BMDMs(bone marrow-derived monocyte/macrophages)were treated with PLG to induce SAM transformation.Specific siRNA was used to knock down KLF4 expression in BMDMs.The protein expression of genes was studied using flow cytometry and Western blot.Immunofluorescence was employed to study the localization of KLF4.RT-qPCR was used to detect the mRNA expression of genes.The results showed that PLG triggered the up-regulation and nuclear localization of KLF4.The knockdown of KLF4 expression blocked PLG-driven SAM transformation.The expressions of SAM feature/functional genes,the ratio of SAM,and the ability to induce extracellular matrix prodection by hepatic stellate cells were decreased.The overexpression of KLF4 increased SAM feature/functional gene expressions.In conclusion,KLF4 plays an important role in PLG-driven SAM transformation.