M. Thorn1, R. A. Burga1, G. Point1, N. J. Espat1,2, S. C. Katz1,2 1Roger Williams Medical Center,Department Of Surgery,Providence, RI, USA 2Boston University,Department Of Surgery,Boston, MA, USA
Introduction: Obstructive jaundice (ObJ) is a life-threatening clinical condition that results in liver inflammation and impaired immunity. The immunological mechanisms for liver injury due to ObJ are incompletely understood. Programmed death-1 (PD-1, CD279) is an immunoinhibitory molecule expressed by T cells and classically results in T cell exhaustion. Recent reports suggest that PD-1 may have surprising pro-inflammatory effects when activated on myeloid cells (MC) by programmed death ligand-1 (PDL-1). We hypothesized that acute inflammation due to ObJ is driven by PD-1 activation on intrahepatic polymorphonuclear cells (PMN) and Kupffer cells (KC).
Methods: Bile duct ligation (BDL) or sham operations were performed in C57Bl/6 mice and liver leukocytes isolated at 3, 7, or 10 days. In vivo anti-PD-1 injections (200 µg/mouse) were given intraperitoneally. Liver lysate cytokines were evaluated by ELISA and cell phenotype by flow cytometry. KC were defined as CD11b+CD11c-F4/80+, PMN as CD11b+Gr1+, and MC as CD11b+.
Results: Intrahepatic PD-1 expression was not limited to liver T cells, but also expressed at significant levels by MC, including KC and PMN as early as 3 days following BDL. At baseline, 6.9% of liver PMN and 4.1% of KC expressed PD-1. Following 7 days of ObJ, PD-1 expression increased to 18.5% and 24.3% among liver PMN (p=0.014) and KC (p=0.004). PD-1 expression levels declined slightly on day 10 following BDL, but remained elevated on PMN (12.5%, p<0.001) and KC (14.3%, p=0.003) compared to baseline. Both liver PMN (14.3-32.2%+) and KC (31.2-65.3%+) expressed high levels of PDL-1 in jaundiced mice. Following BDL, we observed PD-1-dependent elevations of CXCL2, a chemokine that attracts MC. Intrahepatic CXCL2 levels were 465.9 pg/ml in jaundiced mice, but decreased to 225.9 pg/ml with anti-PD-1 treatment (p=0.045). Accordingly, anti-PD-1 treatment led to a contraction of the liver PMN population as a percentage of liver leukocytes on day 3 following BDL (19.4% vs 9.5%, p=0.03). We also examined the effects of PD-1 blockade on liver HIF1, VEGF, and IL6 levels to measure the broad anti-inflammatory effects of this treatment. While VEGF and HIF1 levels were not affected by anti-PD-1 injections, IL6 levels decreased from 376.1 pg/ml to 255.3 pg/ml in jaundiced animals treated with PD-1 blockade (p=0.009).
Conclusion: We have demonstrated increased expression of PD-1 on liver PMN and KC in response to ObJ. As these cell types also expressed PDL-1, they are likely able to activate PD-1, which we demonstrate to mediate inflammation via increased CXCL2 and IL6 production in the liver. PD-1 blockade may reduce inflammation and restore normal T cell function in the inflamed liver.
