Fig. 4.

Intracellular Ca²⁺ measurements, VEGF-A release, and proliferation in response to ANG II in angiomyolipoma cells. A: representative tracings of intracellular Ca²⁺ concentration ([Ca²⁺]_i) responses to ANG II (1 μM) in TRI102 and TRI103 cells. B: measurements of changes in [Ca²⁺]_i (calculated as the maximum fura-2 ratio value after treatment minus the average baseline ratio value) in the presence or absence of the ARB valsartan (Val; 1 μM). Values are expressed as means ± SE. ***P < 0.001. C: measurements of changes in [Ca²⁺]_i in response to thapsigargin (TG; 1 μM). Inhibition of endoplasmic reticulum (ER) Ca²⁺-ATPase produced equivalent Ca²⁺ responses in TRI102 and TRI103 cells that were not affected by valsartan, demonstrating the specificity of the ANG II effect. D: VEGF-A levels were measured by ELISA in conditioned media from TRI102 and TRI103 cells that were seeded at equal density, cultured to near confluence, serum deprived, and treated for 24 h with or without the mammalian target of rapamycin (mTOR) complex 1 (mTORC1) inhibitor RAD-001 (20 nM) or ANG II (10 or 100 nM). Values are expressed as means ± SEM. **P < 0.01. E: cell number quantitation was performed by crystal violet assay on TRI102 and TRI103 cells in the presence or absence of ANG II (100 nM), valsartan (1 μM), or the combination for 48 h. Bars represent means ± SEM. *P < 0.05; **P < 0.01.