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. 2014 Sep 24;289(46):32178–32185. doi: 10.1074/jbc.M114.601096

FIGURE 2.

FIGURE 2.

ADRB3/HSL signaling up-regulates SphK1 in cultured adipocytes. 3T3-L1 adipocytes were treated with or without isoproterenol, in the presence and absence of HSL inhibitor BAY. Levels of SphK1 (A) and SphK2 (B) were measured by qPCR or Western blot analysis (D and E). Note that isoproterenol (ISO) treatment significantly up-regulated SphK1 expression, and the isoproterenol-increased SphK1 expression was completely abrogated in the presence of BAY. D, SphK1 and SphK2 protein levels were quantitated by NIH ImageJ software and normalized to actin loading control (*, p < 0.05, n = 4, t test). C, 3T3-L1 adipocytes were pretreated with or without HSL inhibitor Novo 13f (Novo, 10 μm, 0.5 h) followed by stimulation in the presence or absence of isoproterenol. Levels of SphK1 were measured by qPCR. F, quantitation of S1P in adipocyte culture media by LC-MS/MS method. Note that S1P production was significantly increased in isoproterenol-treated cells. Data in A–C are means ± S.D. of triplicate determinations. Panels A-F were repeated at least twice with similar results. **, p < 0.01, t test. Ctrl, control.