Fig. 2

GCGR overexpression partially sensitizes SNU398 to glucagon signaling. (a) qPCR mRNA levels of GCGR in SNU398, following lentiviral CMV-driven mammalian expression, compared to Primary Human Hepatocytes (PHH). Data represent a single experiment with 3 biological replicates (3 separate RNA samples). eGFP used as a control. b Protein assessment of GCGR and eGFP overexpression in SNU398. Lysate number denotes independent protein sample. c Quantification of cAMP in SNU398 expressing either eGFP or GCGR and treated with 100 nM glucagon (100 G). Data points represent a single experiment of 2 technical replicates. veh: vehicle (0.05 M acetic acid), 20F: 20uM forskolin (positive control). d Protein analysis of downstream effectors of cAMP signaling in SNU398 cells expressing either eGFP or GCGR and treated with 100 nM glucagon (+). (−): vehicle treated. e Protein localization of p-CREB following glucagon treatment of SNU398 cells expressing either eGFP or GCGR. f qPCR mRNA expression of G6PC in SNU398 cells expressing either eGFP or GCGR and treated with 100nM glucagon (100 G). Data represent a single experiment with 3 biological replicates (3 separate RNA samples). Error bars: ±SEM. veh: vehicle (0.05M acetic acid). ns: not significant, p > 0.05, ordinary one-way ANOVA with Tukey’s multiple comparisons test. g Glucose quantification in SNU398 expressing either eGFP or GCGR and cultured in no glucose media with 1 mM pyruvate for 1 day. 0mM and 25mM glucose media was used as control