Growth differentiation factor 11 modulates metabolism, mitigating the pro-tumoral behavior provided by M2-like macrophages in hepatocellular carcinoma-derived cells

Abstract

BACKGROUND

Hepatocellular carcinoma (HCC) is one of the most aggressive tumors worldwide. Chronic inflammation contributes to tumor evolution, and the infiltration of tumor-associated macrophages (TAMs), also known as M2-like macrophages, is associated with the most aggressive behavior. Therefore, these macrophages provide the primary growth and migratory factors to the tumor cells, including those of HCC. Current therapies are not well optimized for eliminating transformed cells or neutralizing the tumor immune microenvironment leukocytes, such as TAMs. Growth differentiation factor 11 (GDF11) may represent a promising dual therapeutic target due to its reported anti-tumorigenic and immunomodulatory properties.

AIM

To characterize the effects of GDF11 in M2-like macrophages and the HCC cell interaction using a functional in vitro model.

METHODS

This research used THP-1 and Huh7 cell lines. We applied recombinant GDF11 (50 ng/mL) every 24 hours on THP-1 differentiated macrophages with M2-like polarization using interleukin-4 and interleukin-13. Firstly, the GDF11 effects on signaling, viability, proliferation, metabolism, and redox state in macrophages were characterized. Subsequently, we extracted conditioned media (CM) from macrophages and performed indirect co-cultures with Huh7 cells. The functional parameters were proliferation and migration assays. Finally, we characterized secretion in the CM using the cytokine array membrane assay.

RESULTS

The present study demonstrated that GDF11 activates the canonical pathway Smad2/3 without cytotoxic or proliferative effects. We provide evidence that GDF11 also diminishes the pro-tumoral properties of M2-like macrophages. GDF11 promoted the reduction of the M2-like macrophage marker, cluster of differentiation 206, indicating a loss of pro-tumoral properties in these leukocytes. Furthermore, this molecule induced changes in metabolism and an increase in reactive oxygen species. Using CM derived from GDF11-treated M2-like macrophages, we observed a reduction in the proliferation and migratory capacity of liver cancer cells. Moreover, the cytokine profile was affected by GDF11 stimulus, demonstrating that this molecule alters the pro-tumoral properties of TAMs, which in turn impact the behavior of HCC-derived cells.

CONCLUSION

This in vitro study suggests that mitigating tumor-promoting or M2-like macrophages with GDF11 may be an effective strategy for controlling the aggressiveness of HCC.

Keywords: Hepatocellular carcinoma; Growth differentiation factor 11; Tumor-associated macrophages; M2-like macrophages; Tumor immune microenvironment

Core Tip: Hepatocellular carcinoma (HCC) is an aggressive tumor, and the infiltration of tumor-associated macrophages or M2-like macrophages increases aggressiveness. We need therapies to mitigate this pathology in the immune context. In the present study, our in vitro model demonstrates that growth differentiation factor 11 (GDF11) counteracts the pro-tumorigenic properties of M2-like macrophages. This was shown in the reduction of cluster of differentiation 206; furthermore, it induces changes in metabolism and the redox state. Conditioned media from GDF11-treated M2-like macrophages possessed changes in their cytokine profile that neutralized the proliferation and migration of HCC cells, demonstrating that GDF11 alters the pro-tumoral properties of macrophages. This study suggests that mitigating tumor-associated macrophages may be an effective strategy for controlling HCC.