The role of neutrophil extracellular traps in nonalcoholic steatohepatitis-associated hepatocellular carcinoma

Abstract

Abstract Introduction Nonalcoholic steatohepatitis (NASH) is a progressive form of nonalcoholic fatty liver disease characterized by steatosis, hepatocellular injury and chronic inflammation. Our recent findings showed that neutrophil forming neutrophil extracellular traps(NETs) in the NASH liver provide a pro-tumorigenic micro-environment. However, the mechanism has not been investigated. Here we sought that NASH induced-NETs suppress the development of anti-tumor immunity by promoting an immune-suppressive through Programmed eath-ligand 1 (PD-L1) signaling for HCC survival. Methods NASH-HCC mice were created by exposing C57BL/6 mice to streptozotocinand(STZ) and high fat diet. NET blockade was achieved with injection of DNase1, or by using PAD4 KO mice. Results There is increased neutrophil infiltration in the liver in the NASH-HCC mice and a concomitant decline in CD4+ and CD8+ T cells. Cit-H3, a specific NET marker persists in the liver through 20 weeks when HCC growth was visible. NETs blockade suppressed tumor growth. PD-L1 expression and PD-1, CTLA-4, LAG-3, TIM-3, the hallmarks of T cell exhaustion are all remarkably increased in CD8+ T cells from NASH-HCC liver compared with control and blocking NETs reduced their expressions. Furthermore, PD-L1 is expressed in the structure of NETs and neutrophils by exposure to stimuli which induce NET formation. The viability of CD4+ and CD8+ T cells isolated from spleen with the stimulation of pure NETs decreased. Summary The present results indicate that NASH induced NETs are critical to the inflammation driven carcinogenesis of NASH-HCC by promoting PD-1/PD-L-1 inhibitory signals, and demonstrate the therapeutic potential of disrupting NETs for the prevention of NASH-associated HCC.