TLR9-mediated immune sensing of bacterial DNA in decompensated cirrhosis is stage-dependent

Background:

Altered local and systemic levels of type-I interferons (IFN) have been linked to impaired antibacterial capacity in animal models of cirrhosis and to a higher risk of death in patients with alcoholic cirrhosis. DNA sensing by immune competent cells is known to elicit strong type-I IFN responses, which may play a role in the context of bacterial translocation in cirrhosis.

Aims:

To characterize immune responses towards CpG-rich single stranded DNA oligonucleotides from E. coli (ssE.coli CpG) in monocytes from patients with decompensated cirrhosis and from healthy controls.

Methods:

CD14-positive monocytes were isolated by density gradient separation and immunomagnetic sorting. Cells were transfected with ssE.coli CpG and bioactive type-I IFN in supernatants as well as in serum was measured using a IFN reporter cell assay. IL-6, TNF, and IL-10 were quantified using ELISA and mRNA expression using qRT-PCR.

Results:

Serum type-I IFN bioactivity was elevated above 30 pg/ml IFN-β equivalent in 32% of patients with decompensated cirrhosis and ascites. As compared to healthy individuals, purified monocytes from patients with decompensated cirrhosis secreted higher levels of IL-6 (p = 0.007) and type-I IFN (p= 0.001) after isolation indicating recent pro-inflammatory activation. In contrast to LPS, ss E.coli CpG was a potent inducer of type-I IFN release and IFNA, IFNB, and IFN-response gene expression. Overall, the treatment of monocytes from patients with cirrhosis with ss E.coli CpG in vitro resulted in a significantly increased release of IL-6 (p = 0.0001) and bioactive IFNα/β (p = 0.0002) compared to healthy individuals, although some patients had diminished type-I IFN secretion. In patients with decompensated cirrhosis (n = 43), advanced disease states were associated with reduced ssE.coli CpG-induced type-I IFN release from monocytes, which correlated with serum bilirubin (r =-0.41; p= 0.008), international normalized ratio (r =-0.33; p= 0.04), and serum albumin (r = 0.41; p= 0.01). Prevention of endosomal acidification with chloroquine abrogated type-I IFN release, indicating a primary role of endosomal but not cytoplasmatic DNA sensing in this process.

Conclusions:

Monocytes from patients with decompensated cirrhosis are potent producers of type-I IFN in response to bacterial DNA. Consistent with a concept of cirrhosis-associated immune dysfunction in cirrhosis, both, TLR9-mediated hyperinflammatory as well as blunted immune responses are observed in different disease states.