Discussion
IL-35 has been reported to have a role in liver disease, including viral
hepatitis, hepatic fibrosis, liver cirrhosis, chemical-induced liver
injury, as well as HCC [10, 14-18]. Additionally, a large amount of
HCC is associated with impairment of the proliferation, cytokine
production, and cytotoxic effector functions accompanied by a chronic
hepatitis virus-specific T cell immune reaction [19]. In this study
we found large amounts of IL-35 in HCC patient serum and tumor tissues,
which indicated that IL-35 might be involved in HCC initiation and
progression. Thus, a better understanding of the relationship between
the level of IL-35 expression and the anti-tumor immune changes seemed
necessary.
The immune cells and secreted cytokines present in the tumor
microenvironment contribute to tumor progression or active repression of
tumor development. A total of 1270 DEGs were detected in the tumor and
non-tumor tissues. Through online IPA pathway analysis we found that
most of the overlapped DEGs were involved in T cell immune activities.
Based on our previous work in which over-expressed IL-35 was shown to
exert strong inhibition on CD4+T cells through
Tregs-derived IL-10, we determined the levels of Foxp3 and IL-10
expression to explore the changes in Tregs [20]. We found that there
were higher Foxp3 and IL-10 expression in IL-35 high group cases, which
suggested that IL-35 was positively correlated with Tregs frequency and
function.
Peripheral and tissue resident CD8+T cells have roles
in the anti-tumor effect and IL-35 suppressed cytotoxicity and
pro-inflammatory cytokine secretion [21]. Hacohen et al. (17)
developed cytolytic activity as a quantitative measure of the activity
of tumor-infiltrating lymphocytes, especially CD8+T
cells, based on two key effector genes (GZMA and PRF1[22]. In this
study we calculated the index “CYT” and detected the level of IFN-γ
expression to reflect the cytotoxic status and anti-tumor cytokine
secretion capability between the two compared groups. Our results showed
that both the cytolytic activity and IFN-γ secretion were significantly
decreased when there was a large amount of IL-35 present in the liver
microenvironment.
In the past few years, immune inhibitory checkpoints have been
increasingly recognized as important elements in the immunosuppression
of chronic inflammation and tumor immunity. There are studies that have
shown Tregs-derived IL-35 induce CD8+T cell exhaustion that limits
effective anti-tumor immunity [13]. We checked the expression of T
cell inhibitory receptor to reflect the intratumoral T cell exhaustion
status. Our results revealed that both PD1 and LAG3 are highly expressed
in the IL-35 high group samples, which indicated that IL-35 might
promote inhibitory receptor induction and exhaustion.