Hypoxia-Mediated Regulation of TGFβ in Hepatocellular Carcinoma Angiogenesis: A Novel HIF1α-miR145-Smad2/3-TSP1 Axis

Background and Aims: The role of transforming growth factor β (TGFβ) in regulating angiogenesis is still under debate. While TGFβ has been found to have an inhibitory effect on angiogenesis, abundant TGFβ and active angiogenesis seem to coexist in tumor microenvironments. The aim of this study was to investigate how tumors overcome TGFβ's anti-angiogenesis effect.

Methods: We used mouse xenograft models, transgenic zebrafish models, clinical samples, and cell models.

Results: Our experiments showed that TGFβ suppressed physiological angiogenesis in zebrafish but did not affect angiogenesis in mouse hepatoma. Interestingly, the anti-angiogenesis effect of TGFβ was weakened in zebrafish treated with HIF1α stabilizer, while it was restored in hepatoma xenografts from mice administered with HIF1α inhibitor. This indicates that the hypoxic microenvironment may attenuate TGFβ's anti-angiogenesis role. In vitro analyses showed that TGFβ inhibited the migration and tube formation of endothelial cells (EC) in normoxia but not in hypoxia conditions by upregulating TSP1, a critical anti-angiogenic factor. Moreover, HIF1α induced miR145 expression upon hypoxia, which abrogated TGFβ's role in upregulating TSP1 and repressing angiogenesis by binding to Smad2/3 and inhibiting their expression in EC. Further examination of primary EC isolated from human hepatocellular carcinoma (HCC) tissues and their adjacent non-tumor liver revealed that Smad2, Smad3, and TSP1 were dramatically decreased, while miR145 was significantly elevated in TEC. The decreases of Smad3 and TSP1 were negatively correlated with tumor angiogenesis in hepatocellular carcinoma tissues.

Conclusions: Our findings suggest that a novel HIF1α-miR145-Smad2/3-TSP1 axis in endothelial cells abrogates the anti-angiogenesis role of TGFβ, highlighting the hypoxic microenvironment as a key factor for tumors to overcome TGFβ's anti-angiogenesis control.

Lay summary: Our study reveals that the role of TGFβ in physiological and HCC angiogenesis can be switched by the hypoxic microenvironment. We identified a novel mechanism by which tumors overcome TGFβ's anti-angiogenesis effect and provided insight into the complex regulatory network of TGFβ in HCC angiogenesis, which may have important implications for precision medicine.