Comparison of ECV-304 cells injured by the sera from patients with hypertension and type 2 diabetes mellitus

AIM: To compare the cellular injury model of ECV-304 induced by the sera from patients with the same blood stasis syndrome (BSS) in hypertension and type 2 diabetes mellitus (T2DM). METHODS: Studies were conducted in ECV-304 cell line, which was treated with different sera during growing, named as sera from patients with T2DM (with the sera from patients with BSS associated with T2DM), sera from patients with hypertension (with the sera from patients with BSS associated with hypertension) and control groups. The cell viability was measured by MTT colorimetry, and the morphological changes were identified by light microscopy and electron microscopy. The level of nitric oxide (NO) and endothelin (ET) in the cells was detected by nitric acid deoxidizing enzyme and non-balance method,respectively. Enzyme-linked immunosorbent assay (ELISA) was applied to assay von Willabrand factor (vWF), thrombomodulin (sTM) and endothelial cell protein C (EPCR) content in the cell culture supernatants. Intracellular free calcium ( _i) and cytoskeleton of the model cells were measured by laser scanning confocal microscopy (LSCM). RESULTS: The cells were damaged when treated with 100 mL/L sera for 24h. Compared with the control group, the level of NO, vWF, sTM, EPCR and intracellular calcium concentration in the damaged cells markedly increase while cell viability and the level of ET significantly decreased (P<0.05). Compared with the sera from patients with T2DM group, the excretion of NO and intracellular calcium concentration markedly increased while ET level, EPCR and cytoskeleton significantly decreased (P<0.05) in the cells induced by the sera from patients with hypertension. CONCLUSION: These results suggest that the sera of patients with BSS associated with T2DM or hypertension reduce cell viability and the level of ET, increase the level of NO, vWF, sTM, EPCR and intracellular free calcium ( _i) and damage cell shape and cytoskeleton. The results also clearly show that the changes of ET, NO, EPCR, _i and cytoskeleton in the cells between the sera from patients with the same BSS syndrome associate with T2DM and hypertension, which may partially explain the pathophysiology mechanism of "different disease and identical syndrome" in traditional Chinese medicine.