Hepatocellular carcinoma (HCC) is a hypervascular
tumor, and accumulating evidence has indicated that stimulation of angiogenesis by hepatitis B virus (HBV) may contribute to HCC
malignancy. The small
protein of hepatitis B virus
surface antigen (
HBsAg),
SHBs, is the most abundant HBV
protein and has a close clinical association with HCC; however, whether
SHBs contributes to HCC angiogenesis remains unknown. This study reports that the forced expression of
SHBs in HCC cells promoted xenograft
tumor growth and increased the microvessel density (MVD) within the
tumors. Consistently,
HBsAg was also positively correlated with MVD counts in HCC patients' specimens. The
conditioned media from the
SHBs-transfected HCC cells increased the capillary tube formation and migration of human umbilical vein endothelial cells (HUVECs). Intriguingly, the overexpression of
SHBs increased
vascular endothelial growth factor A (VEGFA) expression at both the
mRNA and
protein levels. Higher VEGFA expression levels were also observed in xenograft
tumors transplanted with
SHBs-expressing HCC cells and in
HBsAg-positive HCC
tumor tissues than in their negative controls. As expected, in the culture supernatants, the secretion of VEGFA was also significantly enhanced from HCC cells expressing
SHBs, which promoted HUVEC migration and vessel formation. Furthermore, all three unfolded protein response (UPR) sensors,
inositol-requiring
enzyme 1α (IRE1α),
protein kinase RNA-like endoplasmic reticulum (ER)
kinase (PERK), and
activating transcription factor 6 (ATF6), associated with ER stress were found to be activated in
SHBs-expressing cells and correlated with VEGFA
protein expression and secretion. Taken together, these results suggest an important role of
SHBs in HCC angiogenesis and may highlight a potential target for preventive and therapeutic intervention for HBV-related HCC and its malignant progression. IMPORTANCE
Chronic hepatitis B virus infection is one of the important risk factors for the development and progression of
hepatocellular carcinoma (HCC). HCC is characteristic of hypervascularization even at early phases of the disease due to the overexpression of angiogenic factors like
vascular endothelial growth factor A (VEGFA). However, a detailed mechanism of HBV-induced angiogenesis remains to be established. In this study, we demonstrate for the first time that the most abundant HBV
protein, i.e., small
surface antigen (
SHBs), can enhance the angiogenic capacity of HCC cells by the upregulation of VEGFA expression both in vitro and in vivo. Mechanistically,
SHBs induced endoplasmic reticulum (ER) stress, which consequently activated unfolded protein response (UPR) signaling to increase VEGFA expression and secretion. This study suggests that
SHBs plays an important proangiogenic role in HBV-associated HCC and may represent a potential target for antiangiogenic
therapy in HCC.