4 Discussion
Despite recent progress in CRC treatment, patients’ quality of life has
not significantly improved. Nearly 50% of patients experience tumor
recurrence and drug resistance20. Therefore, it is
necessary to explore CRC pathogenesis and develop new treatment
alternatives. IFN-λ is a new type of interferon, recently discovered and
initially recognized for its antiviral activity21, 22.
Later, its anti-tumor and immunomodulatory activities have attracted
researchers’ attention6-9, 23-25. Many groups have
reported the anti-tumor effects of IFN-λ in tumor models—these include
liver cancer14, lung cancer15,
esophageal cancer26, breast
cancer27, prostate cancer28, and
fibrosarcoma29. We investigated the effects and
mechanism of subcutaneous IL-28B injection on colon tumors. We confirmed
that IL-28B inhibited MC38 mouse colon cancer models and regulated
immune cell behavior. Furthermore, we found that M2 macrophages play an
important role in IL-28B anti-tumor activity.
We found that IL-28B effectively inhibited colon cancer growth. Tumor
volume and quality were significantly reduced compared with the control
group. The necrosis of tumor tissue after IL-28B expression was
significantly increased. IFN-III exerts direct anti-tumor effects by
inhibiting tumor cell proliferation and promoting tumor cell apoptosis
and cell cycle stagnation30. However, we found that
even IL-28B concentrations of 1000 ng/mL did not affect MC38 apoptosis
and cell viability in vitro . Interestingly, we confirmed the same
result with IL-28B-treated RM-1 cells (mouse prostate cancer cell line)in vitro . We speculated that the role of IL-28B in colon cancer
might be related to its immunomodulatory effects.
IFN-λ exerts indirect anti-tumor activity by regulating host immune
cells such as natural killer cells31 and T
cells32. Therefore, we detected immune cells in the
peripheral blood, spleen, and tumor tissues. We found that IL-28B
regulates immune cells, consistent with previous reports. Interestingly,
the difference in M2 macrophages in the tumor tissue was the most
significant. After IL-28B treatment, M2 macrophages in tumor tissues
were significantly reduced. To the best of our knowledge, macrophages
have two types of activation, M1 and M2. M2 macrophages exert a powerful
effect on tumor growth and are involved in the proliferation,
angiogenesis, tumor invasion, and metastasis of tumor
cells33. However,
the regulatory mechanism of IL-28B in M2 macrophages is not well
understood.
Therefore, we cultivated BMDMs in vitro to explore the regulatory
mechanism of IL-28B on M2 macrophages. CD11b, F4/80, and CD206 were
labeled M2 in vitro . In the pre-experiment, we found IL-28B
addition before differentiation into M2 led to no significant effects.
Li et al.34 found that IFN-λ time-dependently
influences cell behavior and biology. Therefore, we tuned the timing of
IL-28B addition. Compared with that in the control group, IL-28B
inhibited the polarization of BMDMs to M2 in the IL-28B-treated groups.
Consistent with in vivo results, the relative abundance of M2
macrophages was significantly reduced. Furthermore, we analyzed TGF-β
and Arg-1 expression and cytokine secretion in M2 macrophages. We found
that TGF-β and Arg-1 expression was significantly reduced. These results
indicated that IL-28B directly regulated M2 macrophages. Clinical
studies show that tumor treatment and prognosis are associated with M2
macrophages in tumor tissue30. Therefore, we believe
that IL-28B exerted anti-tumor effects by adjusting M2 macrophage
abundance and behavior in the tumor microenvironment.
Signal transducer and activator of
transcription (STAT) and MAPK pathways are closely related to the
polarization of M2 macrophages35. Yu
et al. found that in the tumor
microenvironment, M2 polarization is related to STAT3 overexpression,
and STAT3 activation can increase blood vessel production, transfer, and
promote the expression of the proliferation-related
genes36. JNK affects the polarization of M2
macrophages, and in turn, the induction of M2-IL-4 activates
JNK37. Our results showed that IL-28B affected the
polarization of M2 macrophages by downregulating the JNK and STAT3
pathways.