The Recruitment of Innate Immune Cells in Liver Diseases: Uncovering the Role of NK Cells and pDC in HCC and NASH

Abstract

Background/ Objective Hepatocellular carcinoma (HCC) is the most common primary malignancy of the liver, with over half a million new cases diagnosed annually worldwide. The fatality of HCC is high despite administration of traditional chemotherapies. In recent years, immunotherapy has become a promising strategy to treat cancer, such as PD‐1/PDL‐1 immune checkpoint blockade. However, the interactions between immune cells and HCC and how immunotherapies work, in vivo, in HCC are still unclear. Non‐alcoholic steatohepatitis (NASH) is an extremely severe form of fatty liver disease and often associated with heavy infiltration of immune cells. Natural Killer (NK) cells and plasmacytoid dendritic cells (pDC) are critical in the innate immune response, and are known for their cytotoxic and interferon producing abilities in response to cancer cells and viral pathogens, respectively. To learn more about the role of innate immunity in liver diseases, we evaluated and characterized pDC and NK cells in HCC and NASH compared to their background hepatic tissue. Methods NK cells and pDC were isolated from HCC, NASH and hepatic tissue, stained with CD3, CD56, and CXCR6 or BDCA2 and CD123, respectively. The flow cytometry data were acquired and analyzed using FlowJo. E2‐2 immunostain for pDC was performed on FFPE tissue of HCC and NASH. Results The presence of pDC was identified in the lymphoid aggregates of HCC and background cirrhotic areas (6/9 cases). In contrast, in 3 HCC cases, no pDCs were appreciated either in tumors or their backgrounds, including two patients who were smokers and with a history of coronary artery disease. In addition, using flow cytometry we demonstrated that there were more pDC in tumor infiltrating lymphocytes (TILs) of HCC compared to adjacent and distant hepatic tissues (Figure 1). We also found that NK cells isolated from NASH livers do not express CXCR6, a marker for liver‐resident NK cells, in contrast to NK cells obtained from HCC that express high levels of CXCR6 in both tumor and non‐tumor areas. The NK cells from NASH appear similar to those isolated and expanded from peripheral blood as both show low levels of CXCR6 (Figure 2). This suggests that NK cells in NASH originated from the blood and completely outnumber those in the liver, while the NK cells in HCC are residents of the liver. Conclusions We have shown that both pDC and NK cells are recruited to the liver in patients with NASH or HCC. More pDC were found in HCC tumors compared to surrounding hepatic tissue. NK cells isolated from NASH do not express CXCR6, a marker for liver‐resident NK cells. This study will further uncover the roles of innate immunity in liver disease and assist in the establishment of novel immunotherapies. A. pDC were present in the lymphoid aggregates of HCC and infiltrating tumor tissue. B. The numbers of pDC in TILs of HCC, adjacent and distant hepatic tissue. Reduced expression of CXCR6 in NASH liver