Objective: To explore the influence of inhibition of hypoxia-inducible factor-1 alpha (HIF-1 alpha) by RNA interference (RNAi) on tumorigenesis of human myeloma cell line (HMCL) RPMI8226 cells in nude mice.
Method: RNAi vector of HIF-1 alpha was constructed with commercial shRNA expression vector pSilencer 2. 1-U6 hygro. RT-PCR and western blot were used to detect HIF-1 alpha mRNA and protein expression respectively. Vascular endothelial growth factor (VEGF) secretion and cell cycle changes were detected by ELISA and flow cytometry respectively. Expression of target gene of HIF-1 alpha, VEGF and Glut-1 were tested under hypoxia condition. Tumorigenesis was observed after transfected cells were injected subcutaneously in nude mice.
Results: After interference, expression of HIF-1 alpha decreased significantly at both mRNA and protein level. Under normoxia condition, VEGF concentrations in HIF-la inhibited cells (RPMI8226-il and RPMI8226-i2) and non-inhibited cells (RPMI8226-c and RPMI8226) showed no differences. While under hypoxia condition, VEGF concentration in the above four cells was (506.0 +/- 53.2), (494.7 +/- 63.1), (984.4 +/- 61.9) and (938.2 +/- 62.2) pg/ml, respectively, being significantly lower in RPMI8226-il and RPMI8226-i2 cells than in RPMI8226-c and RPMI8226 cells (P <0.05). HIF-1 alpha interference was found to suppress the cells shift from S-phase to G1 induced by hypoxia. VEGF and Glut-1 expressions were markedly attenuated (P <0.05). The growth rate of HIF-1 alpha inhibition tumors in subcutaneous xenograft model decreased drastically.
Conclusions: RNAi inhibits HIF-1 alpha expression. Reduced tumor growth by HIF-1 alpha inhibition may partly through inhibiton of angiogenesis and glycolysis metabolism.