R. B. Hawkins1, J. C. Rincon1, S. L. Raymond1, S. D. Larson1 1University Of Florida,Division Of Pediatric Surgery,Gainesville, FL, USA
Introduction: Neonatal sepsis is a leading cause of morbidity and mortality, with over 1 million annual deaths worldwide. Neonates are at higher risk of sepsis and poor outcomes due to quantitative and functional impairments in immunity. Previous work from our laboratory has demonstrated that neonates rely on innate immunity during sepsis. However, functional aspects of innate immunity including neutrophil motility, neutrophil extracellular trap production, cytokine production, and antigen presentation are impaired in neonates. The purpose of this study was to better understand the quantitative and functional differences in neonatal dendritic cells at baseline and in response to sepsis compared to adults. We hypothesized that neonates have decreased dendritic cell populations at baseline and that antigen-presenting capabilities in response to sepsis are impaired.
Methods: Neonatal and adult C57BL/6 mice received an intraperitoneal injection of cecal slurry (CS) at 1.1 mg/g body weight to induce sepsis (LD40-70). Mice were euthanized 18 hours following CS. Splenocytes were harvested and myeloid (GR1+CD11b+) and dendritic cell activation (MHCII, CD80, CD86) markers were analyzed with flow cytometry. For in vitro studies, bone marrow-derived cells (BMDCs) were isolated from naïve neonatal and adult B6 mice and differentiated for 7 days with 20 ng/mL GM-CSF. Cells were stimulated with 100 ng/mL lipopolysaccharide (LPS) for 24 hours, and dendritic cell activation markers were analyzed using flow cytometry.
Results: Splenic GR1+CD11b+ populations were similar at baseline between neonatal and adult mice. However, 18 hours following CS, neonates had significant depletion of GR1+CD11+ cells (p<0.01) while adults had no significant change (Figure 1). Similarly, following in vitro stimulation with LPS, neonatal BMDCs experienced no expansion of the GR1+CD11b+ lineage, while adult BMDCs had significant myeloid expansion (p<0.001). In vitro LPS stimulation induced increased expression of MHCII with concurrent diminishment of CD86 and CD80 in adult but not neonatal BMDCs (p<0.001).
Conclusion: The adult response to sepsis involves early expansion of myeloid cells that differentiate into macrophages or neutrophils to fight infection. Neonatal mice have aberrant downregulation of splenic and bone marrow dendritic cells in response to infectious stimuli in both in vivo and in vitro experiments. Taken together, these results suggest that neonatal myeloid cells have reduced antigen presentation function, which may partially explain the diminished neonatal ability to mount an early, effective immune response during sepsis.
