ANG II-stimulated DNA synthesis is mediated by ANG II receptor-dependent Ca²⁺/PKC as well as EGF receptor-dependent PI3K/Akt/mTOR/p70S6K1 signal pathways in mouse embryonic stem cells

Effect of angiotensin II (ANG II) on mouse embryonic stem (ES) cell proliferation was examined. ANG II increased [³H] thymidine incorporation in a time- (>4 h) and dose- (>10^-9 M) dependent manner. The ANG II-induced increase in [³H] thymidine incorporation was blocked by inhibition of ANG II type 1 (AT₁) receptor but not by ANG II type 2 (AT₂) receptor, and AT₁ receptor was expressed. ANG II increased inositol phosphates formation and [Ca ²⁺]_i, and translocated PKC α, δ, and ζ to the membrane fraction. Consequently, the inhibition of PLC/PKC suppressed ANG II-induced increase in [³H] thymidine incorporation. The inhibition of EGF receptor kinase or tyrosine kinase prevented ANG II-induced increase in [³H] thymidine incorporation. ANG II phosphorylated EGF receptor and increased Akt, mTOR, and p70S6K1 phosphorylation blocked by AG 1478 (EGF receptor kinase blocker). ANG II-induced increase in [³H] thymidine incorporation was blocked by the inhibition of p44/42 MAPKs but not by p38 MAPK inhibition. Indeed, ANG II phosphorylated p44/42 MAPKs, which was prevented by the inhibition of the PKC and AT₁ receptor. ANG II increased c-fos, c-jun, and c-myc levels. ANG II also increased the protein levels of cyclin D1, cyclin E, cyclin-dependent kinase (CDK) 2, and CDK4 but decreased the p21 ^cip1/waf1 and p27^kip1, CDK inhibitory proteins. These proteins were blocked by the inhibition of AT₁ receptor, PLC/PKC, p44/42 MAPKs, EGF receptor, or tyrosine kinase. In conclusion, ANG II-stimulated DNA synthesis is mediated by ANG II receptor-dependent Ca²⁺/PKC and EGF receptor-dependent PI3K/Akt/mTOR/p70S6K1 signal pathways in mouse ES cells.