Peng Xianglin, Pu Feifei, Zhou Fangzheng, Dai Xiyong, Xu Feng, Wang Junwen, Feng Jing, Xia Ping
Department of Orthopedics, Traditional Chinese and Western Medicine Hospital of Wuhan, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, 430022, China.
Department of Orthopedics, Wuhan No.1 Hospital, Wuhan, 430022, China.
Sci Rep. 2025 Mar 25;15(1):10240. doi: 10.1038/s41598-025-94452-w.
Autophagy is a widespread physiological process in the body, which also protects the host by degrading invading pathogens and harmful substances during pathological conditions. Nevertheless, Mycobacterium tuberculosis (MTB), the causative agent of tuberculosis, has evolved strategies to subvert autophagy by modulating microRNA (miRNA) expression, enabling its escape from host defenses. In this study, we established an in vitro model using the human macrophage cell line infected with the highly virulent MTB strain H37Rv. Through RNA sequencing and bioinformatic analysis post H37Rv infection, we screened 14 differentially expressed miRNAs. We predicted and demonstrated that miR-30c-1-3p inhibits autophagy and promotes MTB survival by targeting ATG4B and ATG9B during the infection process. The results showed that miR-30c-1-3p expression was gradually increased before 12 h of H37Rv infection, followed by a decrease. Overexpression of miR-30c-1-3p suppressed autophagic activity. We also identified the targeting of miR-30c-1-3p to ATG4B and ATG9B for the first time, and overexpression of both ATG4B and ATG9B, alone or together, on the basis with upregulation of miR-30c-1-3p reversed the inhibition of autophagy. Autophagy levels were analyzed at different levels by western blot, immunofluorescence, and transmission electron microscopy, all of which showed that upregulation of miR-30c-1-3p inhibited autophagy during H37Rv infection. Additionally, the intervention of miR-30c-1-3p mimics resulted in an increased bacterial load in macrophages, suggesting that MTB achieves immune evasion by upregulating miR-30c-1-3p during infection. In conclusion, our study provides a valuable target for the development of host-directed anti-tuberculosis therapy as well as a new diagnostic marker.
自噬是机体中一种广泛存在的生理过程，在病理状态下，它还可通过降解入侵的病原体和有害物质来保护宿主。然而，结核病的病原体结核分枝杆菌（MTB）已进化出通过调节微小RNA（miRNA）表达来破坏自噬的策略，从而使其能够逃避宿主防御。在本研究中，我们利用感染高毒力MTB菌株H37Rv的人巨噬细胞系建立了体外模型。通过对H37Rv感染后的RNA测序和生物信息学分析，我们筛选出14种差异表达的miRNA。我们预测并证明，在感染过程中，miR-30c-1-3p通过靶向ATG4B和ATG9B抑制自噬并促进MTB存活。结果显示，在H37Rv感染12小时前，miR-30c-1-3p表达逐渐增加，随后下降。miR-30c-1-3p的过表达抑制了自噬活性。我们还首次鉴定了miR-30c-1-3p对ATG4B和ATG9B的靶向作用，并且在miR-30c-1-3p上调的基础上单独或共同过表达ATG4B和ATG9B可逆转对自噬的抑制作用。通过蛋白质免疫印迹、免疫荧光和透射电子显微镜在不同水平分析自噬水平，所有结果均表明，miR-30c-1-3p的上调在H37Rv感染期间抑制了自噬。此外，miR-30c-1-3p模拟物的干预导致巨噬细胞中细菌载量增加，这表明MTB在感染期间通过上调miR-3
