Characterization of NK cell immunity in chronic HCV infection

Abstract

Natural killer (NK) cell responses are crucial for anti-viral and anti-tumor immunity. However, the immune responses of NK cells are impaired in chronically HCV-infected patients. While direct-acting antivirals (DAAs) treatment allows a complete viral clearance in HCV patients, the recovery of NK cell immunity is only fragmentary explored and underlying molecular mechanisms are fundamentally uncharacterized. The first part of this thesis aims to characterize the molecular phenotype of NK cells in chronic HCV patients before and after DAA treatment by using quantitative proteomics. In total, more than 4867 proteins were identified and quantified by mass spectrometry, covering both the surface markers and intracellular components. Principal component analysis and heatmap of correlation factors revealed that NK cell proteomes of the same patients one year after treatment did normalize their inter-individual variance to the level of healthy individuals but, notably, were found still different from the healthy controls. The dysregulated proteins before and after treatment complement our knowledge of impaired NK cell responses in chronically HCV infected patients and indicated both trained and impaired NK cell functions in cured patients. Among protein regulations that indicated trained NK cell immunity, the adhesion and degranulation-promoting adaptor protein (ADAP) was selected for functional studies. Thus, the second part of this thesis investigated the importance of ADAP in human NK cells. In vitro experiments showed that the Src kinases-associated phosphoprotein 1 and 2 are interaction partners of ADAP. Moreover, ADAP regulates cytokine production and degranulation upon the stimulation of K562 cells, as well as migration and adhesion upon stimulation of CXCR4. Interestingly, ADAP is also required for controlling the number of serially killed target cells for each NK cell and the ratio of cytotoxic NK cells. To sum up, these data show that ADAP promotes serial killing through regulating adhesion and migration in human NK cells, confirming that the up-regulation of ADAP is a part of trained immunity in cured HCV patients.