C107-11 Rnase6 Attenuates Lung Injury From Staphylococcus Aureus Pneumonia By Enhancing Bacterial Clearance

Abstract Rationale Staphylococcus aureus (SA) pneumonia is a significant cause of severe respiratory failure in the ICU, frequently progressing to acute respiratory distress syndrome (ARDS). Due to its potent cytotoxins, SA infection leads to intense host inflammatory response and propensity for lung tissue destruction. Novel treatment strategies to lessen tissue damage are needed to combat the growing number of antibiotic-resistant SA strains, including Methicillin-resistant SA (MRSA) and Vancomycin-resistant SA (VRSA). During pneumonia, several ribonuclease (RNase) A superfamily enzymes direct host responses by regulating extracellular RNAs and modulating inflammation. One family member, RNase6 is highly expressed in myeloid cells and processes bacterial RNA for recognition by the endosomal pattern recognition receptor, TLR8. RNase6 has also been suggested to have anti-microbial properties in mouse models of urinary tract infection. We hypothesized that RNase6 plays an important role in host defense from SA pneumonia. Methods For in vitro studies, we differentiated THP-1 myeloid cells to macrophages using phorbol 12-myristate 13-acetate (PMA). For in vivo studies, we obtained RNase6EGFP/EGFP mice, which we will refer to as RNase6 KO mice. Results By secondary analysis of a published proteomics dataset, we observed RNase6 protein is increased in patient plasma during critical illness compared to controls. To characterize RNase6 secretion in vitro, we exposed PMA-differentiated THP-1 macrophages to Pam3CSK4, a synthetic lipopeptide and TLR 1/2 agonist, and assayed protein abundance by immunoblotting. We observed increased RNase6 abundance in supernatants but not cell lysates in a dose- and time-dependent manner. RNase6 expression, measured by qPCR, decreased in response to Pam3CSK4. To investigate RNase6 function during SA-induced lung infections, MRSA or vehicle control (saline) was delivered intranasally to RNase6 KO and WT mice. With SA infection, RNase6 mice demonstrated increased lung injury by histology, increased BAL cell count, increased cytokine secretion, and increased bacterial load, compared to controls. Conclusion RNase6, an RNase A superfamily member, is crucial for bacterial clearance during SA experimental pneumonia. Monocytes and macrophages secrete RNase6 in response to infection, which processes bacterial RNA into a recognizable pattern by TLR8. RNase6 augments early bacterial surveillance and innate immune responses to lessen disease burden, lung injury, and development of ARDS. This abstract is funded by: NHLBI