Hepatocellular carcinoma serum derived exosomal HIF-1α induces phosphoinositide-3 kinase/protein kinase B signaling to induce the angiogenesis and proliferation of HCC

Qingyu Xu, Ran You, Guowen Yin, Jianping Gu


Background: Tumor exosomes are nanovesicles mostly secreted by tumor cells that play roles of paracrine signaling during tumor progression, including tumor-stromal interactions, activation of proliferative pathways and inducing immunosuppression. The hypoxia-inducible factor (HIF) family serve role of the principal molecular mediators of hypoxia in physiological and pathophysiological course. HIF-1α excreted from tumor and stromal cells serves an important role in tumor angiogenesis, especially in hepatocellular carcinoma (HCC). However, exosomal HIF-1α from HCC serum has not been studied extensively. The present study aimed to determine role of HIF-1α derived from HCC serum exosomes in the development of HCC.
Methods: We extracted exosomes from the serum of 48 patients suffering HCC and 24 healthy individuals. The exosomes were examined by transmission electron microscopy and nanoparticle tracking analysis. The concentrations of exosomal HIF-1α were measured by ELISA. In vitro experiments were performed to testify the exosomal role in human umbilical endothelial cells (HUVECs) and Huh-7 cells by tube formation, cell viability and western blotting analyses. Nude mouse xenograft Huh-7 tumors were generated by injecting HCC patient serum exosomes, healthy individuals’ serum exosomes and PBS into nude mice. Tumors in vivo were isolated, weighed and subjected to immunohistochemical assays.
Results: HCC patients exhibited a greater capacity to convert HIF-1α in serum exosomes than normal individuals (P=0.0007), and high serum exosomal HIF-1α levels were correlated with Barcelona Clinic Liver Cancer (BCLC) staging (P=0.031) and the phosphoinositide-3 kinase (PI3K)/protein kinase B (AKT) signaling pathway. The results of flow cytometry analysis demonstrated that the HUVECs cocultured with HCC exosomes turned down the percentage of HUVECs in the G1 phase but turned up the percentage of HUVECs in the S phase (P<0.05). HUVECs and Huh-7 cells cultured with HCC serum exosomes enhanced the proliferation of HUVECs and Huh-7 cells, and the tube formation of HUVECs, while exosomes from healthy individuals and the controls did not. The results demonstrated that serum exosomes from patients with HCC appeared to accelerate tumor growth compared to the control group (P<0.05) and immunohistochemical analysis revealed that the nude mice injected with exosomes from HCC patient serum exhibited higher microvessel density (MVD) (P<0.05).
Conclusions: HCC serum derived exosomes could induce angiogenesis and the proliferation of HCC potentially via the HIF-1α-PI3K/AKT signaling pathway, it making the prevention and treatment of HCC angiogenesis and proliferation available.