Table_1_Transcriptome Architecture of Osteoblastic Cells Infected With Staphylococcus aureus Reveals Strong Inflammatory Responses and Signatures of Metabolic and Epigenetic Dysregulation.xlsx

Staphylococcus aureus is an opportunistic pathogen that causes a range of devastating diseases including chronic osteomyelitis, which partially relies on the internalization and persistence of S. aureus in osteoblasts. The identification of the mechanisms of the osteoblast response to intracellular S. aureus is thus crucial to improve the knowledge of this infectious pathology. Since the signal from specifically infected bacteria-bearing cells is diluted and the results are confounded by bystander effects of uninfected cells, we developed a novel model of long-term infection. Using a flow cytometric approach we isolated only S. aureus-bearing cells from mixed populations that allows to identify signals specific to intracellular infection. Here we present an in-depth analysis of the effect of long-term S. aureus infection on the transcriptional program of human osteoblast-like cells. After RNA-seq and KEGG and Reactome pathway enrichment analysis, the remodeled transcriptomic profile of infected cells revealed exacerbated immune and inflammatory responses, as well as metabolic dysregulations that likely influence the intracellular life of bacteria. Numerous genes encoding epigenetic regulators were downregulated. The later included genes coding for components of chromatin-repressive complexes (e.g., NuRD, BAHD1 and PRC1) and epifactors involved in DNA methylation. Sets of genes encoding proteins of cell adhesion or neurotransmission were also deregulated. Our results suggest that intracellular S. aureus infection has a long-term impact on the genome and epigenome of host cells, which may exert patho-physiological dysfunctions additionally to the defense response during the infection process. Overall, these results not only improve our conceptual understanding of biological processes involved in the long-term S. aureus infections of osteoblast-like cells, but also provide an atlas of deregulated host genes and biological pathways and identify novel markers and potential candidates for prophylactic and therapeutic approaches.

金黄色葡萄球菌（Staphylococcus aureus）是一种机会致病菌，可引发多种毁灭性疾病，其中包括慢性骨髓炎（chronic osteomyelitis），而该疾病的发生一定程度上依赖于金黄色葡萄球菌在成骨细胞（osteoblasts）内的内化与定植。因此，阐明成骨细胞对胞内金黄色葡萄球菌的应答机制，对于深化我们对该感染性病理过程的认知至关重要。
由于携带特异性感染细菌的细胞所产生的信号会被稀释，且实验结果会受到未感染细胞的旁观者效应干扰，我们构建了一种新型长期感染模型。通过流式细胞术（flow cytometric）方法，我们从混合细胞群中仅分离出携带金黄色葡萄球菌的细胞，该方法可精准识别胞内感染相关的特异性信号。
本研究深入分析了长期金黄色葡萄球菌感染对人成骨样细胞转录程序的影响。在完成RNA测序（RNA-seq）、京都基因与基因组百科全书（KEGG）及Reactome通路富集分析后，我们发现感染细胞的转录组谱发生重塑，表现为免疫与炎症反应的加剧，同时伴随可能影响细菌胞内生存状态的代谢失调。
大量编码表观遗传调控因子的基因出现下调，其中包括编码染色质抑制复合物组分的基因（如NuRD、BAHD1和PRC1）以及参与DNA甲基化的表观调控因子。编码细胞黏附蛋白或神经传递相关蛋白的基因集也出现表达失调。
我们的研究结果表明，胞内金黄色葡萄球菌感染会对宿主细胞的基因组与表观基因组产生长期影响，这除了会在感染过程中引发防御应答外，还可能导致病理生理功能异常。
总体而言，这些研究结果不仅深化了我们对成骨样细胞长期金黄色葡萄球菌感染相关生物学过程的概念性认知，还提供了一份失调宿主基因与生物学通路的图谱，并鉴定出可用于预防与治疗策略的新型标志物及潜在候选靶点。