Background: Tumor endothelial cells (TECs) perform tumor angiogenesis, which is essential for tumor growth and metastasis. Tumor cells produce large amounts of lactic acid from glycolysis; however, the mechanism underlying the survival of TECs to enable tumor angiogenesis under high lactic acid conditions in tumors remains poorly understood.
Methodology: The metabolomes of TECs and normal endothelial cells (NECs) were analyzed by capillary electrophoresis time-of-flight mass spectrometry. The expressions of pH regulators in TECs and NECs were determined by quantitative reverse transcription-PCR. Cell proliferation was measured by the MTS assay. Western blotting and ELISA were used to validate monocarboxylate transporter 1 and carbonic anhydrase 2 (CAII) protein expression within the cells, respectively. Human tumor xenograft models were used to access the effect of CA inhibition on tumor angiogenesis. Immunohistochemical staining was used to observe CAII expression, quantify tumor microvasculature, microvessel pericyte coverage, and hypoxia.
Results: The present study shows that, unlike NECs, TECs proliferate in lactic acidic. TECs showed an upregulated CAII expression both in vitro and in vivo. CAII knockdown decreased TEC survival under lactic acidosis and nutrient-replete conditions. Vascular endothelial growth factor A and vascular endothelial growth factor receptor signaling induced CAII expression in NECs. CAII inhibition with acetazolamide minimally reduced tumor angiogenesis in vivo. However, matured blood vessel number increased after acetazolamide treatment, similar to bevacizumab treatment. Additionally, acetazolamide-treated mice showed decreased lung metastasis.
Conclusion: These findings suggest that due to their effect on blood vessel maturity, pH regulators like CAII are promising targets of antiangiogenic therapy. Video Abstract.
Keywords: Angiogenesis; Carbonic anhydrase 2 (CAII); Lactic acidosis; Tumor endothelial cells; pH regulation.