Craniotomies are performed to treat a variety of intracranial pathology. Surgical site infection remains a complication of craniotomy despite the use of prophylactic antibiotics and universal sterile precautions. Infections occur in 1–3% of procedures, with approximately half caused by Staphylococcus aureus that forms a biofilm on the bone flap and is recalcitrant to systemic antibiotic therapy. We used an S. aureus-dsRed construct to compare the phagocytic capacity of leukocytes and microglia in vitro and in vivo using a mouse model of craniotomy infection. In addition, single-cell RNA sequencing (scRNA-seq) was applied to determine whether a transcriptional signature could be identified for phagocytic versus nonphagocytic cells in vivo. S. aureus was phagocytosed to equivalent extents in microglia, macrophages, neutrophils, and granulocytic myeloid-derived suppressor cells in vitro; however, microglial uptake of S. aureus was limited in vivo, whereas the other leukocyte populations exhibited phagocytic activity. scRNA-seq comparing the transcriptional signatures of phagocytic (S. aureus-dsRed+) versus nonphagocytic (S. aureus-dsRed−) leukocytes identified classical pathways enriched in phagocytic cells (i.e., reactive oxygen species [ROS]/reactive nitrogen species, lysosome, iron uptake, and transport), whereas nonphagocytic populations had increased ribosomal, IFN, and hypoxia signatures. scRNA-seq also revealed a robust ROS profile, which led to the exploration of craniotomy infection in NADPH oxidase 2 knockout mice. S. aureus burden, leukocyte recruitment, and intracellular bacterial load were significantly increased in NADPH oxidase 2 KO compared with wild-type animals. Collectively, these results highlight the importance of ROS generation in phagocytes for S. aureus biofilm containment, but not clearance, during craniotomy infection.

Epigenetics involves the study of various modes of adaptable transcriptional regulation, contributing to cell identity, characteristics, and function. During central nervous system (CNS) infection, epigenetic mechanisms can exert pronounced control over the maturation and antimicrobial properties of nearly every immune cell type.Epigenetics is a relatively new field, with the first mention of these marks proposed only a half-century ago and a substantial body of immunological epigenetic research emerging only in the last few decades. Here, we review the best-characterized epigenetic marks and their functions as well as illustrate how various immune cell populations responding to CNS infection utilize these marks to organize their activation state and inflammatory processes. We also discuss the metabolic and clinical implications of epigenetic marks and the rapidly expanding set of tools available to researchers that are enabling elucidation of increasingly detailed genetic regulatory pathways.These considerations paint an intricate picture of inflammatory regulation, where epigenetic marks influence genetic, signaling, and environmental elements to orchestrate a tailored immunological response to the threat at hand, cementing epigenetics as an important player in immunity.

Background Treatment of brain tumors, epilepsy, or hemodynamic abnormalities requires a craniotomy to access the brain. Nearly 1 million craniotomies are performed in the US annually, which increase to ~ 14 million worldwide and despite prophylaxis, infectious complications after craniotomy range from 1 to 3%. Approximately half are caused by Staphylococcus aureus (S. aureus), which forms a biofilm on the bone flap that is recalcitrant to antibiotics and immune-mediated clearance. However, the mechanisms responsible for the persistence of craniotomy infection remain largely unknown. The current study examined the role of IL-10 in promoting bacterial survival. Methods A mouse model of S. aureus craniotomy infection was used with wild type (WT), IL-10 knockout (KO), and IL-10 conditional KO mice where IL-10 was absent in microglia and monocytes/macrophages (CX3CR1CreIL-10 fl/fl) or neutrophils and granulocytic myeloid-derived suppressor cells (G-MDSCs; Mrp8CreIL-10 fl/fl), the major immune cell populations in the infected brain vs. subcutaneous galea, respectively. Mice were examined at various intervals post-infection to quantify bacterial burden, leukocyte recruitment, and inflammatory mediator production in the brain and galea to assess the role of IL-10 in craniotomy persistence. In addition, the role of G-MDSC-derived IL-10 on neutrophil activity was examined. Results Granulocytes (neutrophils and G-MDSCs) were the major producers of IL-10 during craniotomy infection. Bacterial burden was significantly reduced in IL-10 KO mice in the brain and galea at day 14 post-infection compared to WT animals, concomitant with increased CD4+ and γδ T cell recruitment and cytokine/chemokine production, indicative of a heightened proinflammatory response. S. aureus burden was reduced in Mrp8CreIL-10 fl/fl but not CX3CR1CreIL-10 fl/fl mice that was reversed following treatment with exogenous IL-10, suggesting that granulocyte-derived IL-10 was important for promoting S. aureus craniotomy infection. This was likely due, in part, to IL-10 production by G-MDSCs that inhibited neutrophil bactericidal activity and TNF production. Conclusion Collectively, these findings reveal a novel role for granulocyte-derived IL-10 in suppressing S. aureus clearance during craniotomy infection, which is one mechanism to account for biofilm persistence.

Staphylococcus aureus is a common cause of surgical-site infections, including those arising after craniotomy, which is performed to access the brain for the treatment of tumors, epilepsy, or hemorrhage. Craniotomy infection is characterized by complex spatial and temporal dynamics of leukocyte recruitment and microglial activation. We recently identified unique transcriptional profiles of these immune populations during S. aureus craniotomy infection. Epigenetic processes allow rapid and reversible control over gene transcription; however, little is known about how epigenetic pathways influence immunity to live S. aureus. An epigenetic compound library screen identified bromodomain and extraterminal domain–containing (BET) proteins and histone deacetylases (HDACs) as critical for regulating TNF, IL-6, IL-10, and CCL2 production by primary mouse microglia, macrophages, neutrophils, and granulocytic myeloid-derived suppressor cells in response to live S. aureus. Class I HDACs (c1HDACs) were increased in these cell types in vitro and in vivo during acute disease in a mouse model of S. aureus craniotomy infection. However, substantial reductions in c1HDACs were observed during chronic infection, highlighting temporal regulation and the importance of the tissue microenvironment for dictating c1HDAC expression. Microparticle delivery of HDAC and BET inhibitors in vivo caused widespread decreases in inflammatory mediator production, which significantly increased bacterial burden in the brain, galea, and bone flap. These findings identify histone acetylation as an important mechanism for regulating cytokine and chemokine production across diverse immune cell lineages that is critical for bacterial containment. Accordingly, aberrant epigenetic regulation may be important for promoting S. aureus persistence during craniotomy infection.

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