Biology Department, Middle Tennessee State University, Murfreesboro, TN, United States of America.
Microbiology, Immunology, and Molecular Genetics Department, University of Kentucky, Lexington, KY, United States of America.
PLoS One. 2020 Aug 28;15(8):e0233818. doi: 10.1371/journal.pone.0233818. eCollection 2020.
Macrophages serve as a first line of defense against infection with the facultative intracellular pathogen, Cryptococcus neoformans (Cn). However, the ability of these innate phagocytic cells to destroy ingested Cn is strongly influenced by polarization state with classically (M1) activated macrophages better able to control cryptococcal infections than alternatively (M2) activated cells. While earlier studies have demonstrated that intracellular Cn minimally affects the expression of M1 and M2 markers, the impact on the broader transcriptome associated with these states remains unclear. To investigate this, an in vitro cell culture model of intracellular infection together with RNA sequencing-based transcriptome profiling was used to measure the impact of Cn infection on gene expression in both polarization states. The gene expression profile of both M1 and M2 cells was extensively altered to become more like naive (M0) macrophages. Gene ontology analysis suggested that this involved changes in the activity of the Janus kinase-signal transducers and activators of transcription (JAK-STAT), p53, and nuclear factor-κB (NF-κB) pathways. Analyses of the principle polarization markers at the protein-level also revealed discrepancies between the RNA- and protein-level responses. In contrast to earlier studies, intracellular Cn was found to increase protein levels of the M1 marker iNos. In addition, common gene expression changes were identified that occurred post-Cn infection, independent of polarization state. This included upregulation of the transcriptional co-regulator Cited1, which was also apparent at the protein level in M1-polarized macrophages. These changes constitute a transcriptional signature of macrophage Cn infection and provide new insights into how Cn impacts gene expression and the phenotype of host phagocytes.
巨噬细胞作为防御机会性细胞内病原体新型隐球菌（Cn）感染的第一道防线。然而，这些先天吞噬细胞破坏吞噬的 Cn 的能力受到极化状态的强烈影响，经典（M1）激活的巨噬细胞比替代（M2）激活的细胞更能控制隐球菌感染。虽然早期的研究已经表明，细胞内 Cn 对 M1 和 M2 标志物的表达影响最小，但与这些状态相关的更广泛转录组的影响仍不清楚。为了研究这一点，使用体内细胞培养感染模型和基于 RNA 测序的转录组谱分析来测量 Cn 感染对两种极化状态下基因表达的影响。M1 和 M2 细胞的基因表达谱都发生了广泛改变，变得更像未极化（M0）巨噬细胞。基因本体分析表明，这涉及到 Janus 激酶信号转导和转录激活因子（JAK-STAT）、p53 和核因子-κB（NF-κB）途径活性的变化。对蛋白水平的主要极化标记物的分析也揭示了 RNA 和蛋白水平反应之间的差异。与早期的研究相反，细胞内 Cn 被发现增加了 M1 标记物 iNos 的蛋白水平。此外，还鉴定了发生在 Cn 感染后的常见基因表达变化，与极化状态无关。这包括转录共调节剂 Cited1 的上调，在 M1 极化的巨噬细胞中也可以在蛋白水平上观察到。这些变化构成了巨噬细胞 Cn 感染的转录特征，并为 Cn 如何影响基因表达和宿主吞噬细胞表型提供了新的见解。
