S. P. Walker1, N. Galbraith1, C. Bishop1, M. Reid1, S. Gardner1, H. Polk1 1University Of Louisville,Department Of Surgery,Louisville, KY, USA
Introduction:
Major infection and trauma lead to poorly understood pro-inflammatory and compensatory anti-inflammatory immunological responses. Some patients have resulting monocyte impairment which is predictive of nosocomial infection and death. Attempts to augment host defenses in surgical patients, such as interferon-gamma (IFN-γ), have thus far failed to improve mortality. An incomplete understanding exists of the underlying immunological mechanisms, including the role of programmed cell death-ligand 1 (PD-L1), a negative co-stimulatory molecule. The purpose of this study was to determine the effects of IFN-γ on monocyte function and PD-L1 expression.
Methods:
Venous blood was taken using EDTA tubes from healthy volunteers and stimulated ex-vivo with 100 ng/mL of lipopolysaccharide (LPS) for 18 hours in the presence or absence of recombinant IFN-γ (100 ng/mL). The monocyte inflammatory response was measured using quantitative measurements of surface protein expression of human leukocyte antigen-DR (HLA-DR) and PD-L1 on CD14 positive monocytes using flow cytometry. Cytokine responses including tumor necrosis factor-alpha (TNF-α) were measured by ELISA. Statistical analyses was determined using Paired T-test or Wilcoxon signed-rank test as appropriate, with significance set at 0.05. Six donors were used for each experiment.
Results:
Exposure to LPS led to an increase in HLA-DR expression and increased TNF-α production. IFN-γ alone offered minimal effects on HLA-DR or TNF-α levels. The addition of the immunoadjuvant IFN-γ augmented HLA-DR expression and TNF-α levels at 18 h (p<0.05). Monocyte PD-L1 expression was low at baseline (0 h) in healthy volunteers, but increased in the presence of either LPS or IFN-γ alone (p<0.05). IFN-γ treatment in combination with LPS led to a synergistic increase in PD-L1 expression at 6 h and 18 h after stimulation (p<0.05).
Conclusion:
In this study we found that IFN-γ acts to increase monocyte responsiveness as measured by increased HLA-DR expression and TNF-α production in the presence of LPS. Monocyte PD-L1, a negative co-stimulatory molecule, increased in the presence of LPS. Adjuvant IFN-γ treatment further increased PD-L1 expression. These results suggest that IFN-γ therapy stimulates a negative feedback loop through the PD-1-PD-L1 axis, which could have therapeutic implications in approaching immunoadjuvant therapy in the “high risk” patient.