Host response and cell fates of human U937 monocytes upon infection with vancomycin-resistant E. faecium

Abstract

In recent years, Enterococcus faecium has become a high-priority human opportunistic pathogen, especially due to its antibiotic and environmental resistance, promoting hospital-acquired infections. However, its interaction with host cells remains poorly understood. In this study, we investigated the interaction of four recently collected, clinical vancomycin-resistant Enterococcus faecium (VREfm) isolates with the human U937 cell line. Early after infection, we observed close bacterial association with U937 monocytes and features suggestive of internalization. We thus investigated host cell fates after washing at 2 hours post infection (hpi) to concentrate on effects induced by internalized bacteria or those with direct host cell contact. Whereas viability was not impaired at 6 hpi, moderate cell death was induced to an isolate-specific extent at 24 hpi, which was abrogated when direct host-pathogen contact as prevented via boyden chamber cell culture inserts, but was enhanced by TNF-R1 deficiency. Host mRNA sequencing revealed a consistent pattern of differentially expressed genes across all VREfm isolates, suggesting a shared innate immune and stress-related response at 6 hpi, with functionally enriched biological processes related to leukocyte activation, cytokine signaling, programmed cell death, and RNA stability. Together, these data show that VREfm triggers an early pro-inflammatory transcriptional response in U937 monocytes without immediate cytotoxicity in U937 monocytes, while late-stage isolate-specific cell death induction depends on direct host cell contact or internalization and involves TNF-R1 mediated regulation. Our findings may assist future efforts to understand how VREfm triggers host cell damage that contributes to severe inflammation and systemic infection such as persistent bacteremia.