Residual HCV-RNA after therapeutically induced SVR is associated with a more severe liver disease and a faster progression to cirrhosis [4]. In our previous study the presence of residual HCV-RNA detected 12–15 months after EOT was associated with a transient rise of NLR during and shortly after therapy but not with any other parameter tested, including serum liver enzyme levels [7]. Here we show that the presence of HCV-RNA in PBMC correlates with an adverse outcome in a follow-up period of more than 6 years. Thus, our results highlight the clinical significance of OCI during long-term follow-up.

We also found differences in the levels of six immune mediators between patients with and without HCV-RNA in PBMC. Treated patients without OCI had lower plasma levels of IL-18 and MDC/CCL22 than the OCI group and patients with CHC (Table 2, Table S3). Interleukin-18 plays a critical role in T helper type 1 (Th1) response during host antiviral defense. Serum level of IL-18 reflects the severity of liver disease during HCV infection [8] and it declines rapidly in patients with SVR after DAA therapy [9]. MDC/CCL22 recruits T helper type 2 (Th2) and T regulatory cells creating an immunosuppressive microenvironment and plays an important role in the development of liver cancer. Hepatitis C protein NS3/4A induces an increase in expression of intrahepatic CCL22 which is accompanied by diminished production of IFNγ, an immunomodulatory and antiviral cytokine which defines Th1 cells. This results in a switch of the Th1/Th2 ratio towards an anti-inflammatory state leading to viral persistence [9]. It was shown that plasma IFNγ level is lower in patients with CHC in comparison to healthy controls and it does not increase significantly even 12 weeks after successful DAA therapy [9–11]. This is in line with our results where the level of IFNγ was the lowest in patients with OCI and individuals with CHC.

Soluble fractalkine/CX3CL1 exhibits an anti-apoptotic effect across various cell types and has chemoattractive properties for immune cells. Its interaction with the receptor CX3CR1 can trigger either inflammatory or anti-inflammatory responses, depending on the local environmental conditions, such as tissue type or disease state. The fractalkine/CX3CR1 axis is upregulated during liver damage such as chronic hepatitis C and this signaling results in a reduction of hepatic inflammation and fibrosis [12]. Lymphotoxin alpha (LTα) is a pro-inflammatory cytokine crucial for lymphoid organ development and maintenance [13]. It also controls hepatic stellate cell function, lipoprotein homeostasis, and liver regeneration [14]. Although considered mainly as a Th2, immunosuppressive cytokine IL-13 regulates the balance of several other subtypes of T helper cells including Th1, Th17, and Treg cells and plays an important role in immunity, inflammation, and fibrosis [15, 16].

The results from this brief report suggest that in patients with OCI, the plasma cytokine/chemokine profile could be linked to viral persistence (higher MDC/CCL22, lower IFNγ) and point to a continued liver disease (IL-18) in comparison to patients without OCI where mediators with immunoregulatory (LTα, IL-13) and cytoprotective (fractalkine/CX3CL1) properties are more abundant. Elevated LTα, IL-13, and fractalkine/CX3CL1 in patients without OCI could also indicate a more pronounced restoration of immune balance after immune exhaustion caused by HCV infection. In line with this, the cytokine/chemokine profile of patients with OCI was more similar to that of the CHC group than to that of patients without residual HCV-RNA. However, as a result of the small size of groups analyzed in our study, we were not able to the determine cytokine/chemokine footprint which would reliably distinguish between patients with or without OCI. This limitation represents a major drawback of our study, and these findings need to be validated in larger cohorts. Additionally, repeated cytokine and HCV-RNA measurements over a longer observation period would provide more insight into the association between OCI and plasma cytokine/chemokine levels.

We found that PLR is associated with death in treated patients with CHC (Fig. 2a, b). High PLR reflects systemic inflammation and platelet activation, and it was previously found to predict inflammation and mortality in cardiovascular disease, liver cirrhosis, HCC, and in the general population [17,18,19]. On the other hand higher PLR was associated with a better response to the IFN-based therapy in CHC [20].