Hepatic Gi signaling regulates whole-body glucose homeostasis
Mario Rossi, … , Owen P. McGuinness, Jürgen Wess
Mario Rossi, … , Owen P. McGuinness, Jürgen Wess
Published January 16, 2018
Citation Information: J Clin Invest. 2018;128(2):746-759. https://doi.org/10.1172/JCI94505.
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Research Article Endocrinology

Hepatic Gi signaling regulates whole-body glucose homeostasis

Abstract

An increase in hepatic glucose production (HGP) is a key feature of type 2 diabetes. Excessive signaling through hepatic Gs–linked glucagon receptors critically contributes to pathologically elevated HGP. Here, we tested the hypothesis that this metabolic impairment can be counteracted by enhancing hepatic Gi signaling. Specifically, we used a chemogenetic approach to selectively activate Gi-type G proteins in mouse hepatocytes in vivo. Unexpectedly, activation of hepatic Gi signaling triggered a pronounced increase in HGP and severely impaired glucose homeostasis. Moreover, increased Gi signaling stimulated glucose release in human hepatocytes. A lack of functional Gi-type G proteins in hepatocytes reduced blood glucose levels and protected mice against the metabolic deficits caused by the consumption of a high-fat diet. Additionally, we delineated a signaling cascade that links hepatic Gi signaling to ROS production, JNK activation, and a subsequent increase in HGP. Taken together, our data support the concept that drugs able to block hepatic Gi–coupled GPCRs may prove beneficial as antidiabetic drugs.

Authors

Mario Rossi, Lu Zhu, Sara M. McMillin, Sai Prasad Pydi, Shanu Jain, Lei Wang, Yinghong Cui, Regina J. Lee, Amanda H. Cohen, Hideaki Kaneto, Morris J. Birnbaum, Yanling Ma, Yaron Rotman, Jie Liu, Travis J. Cyphert, Toren Finkel, Owen P. McGuinness, Jürgen Wess

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Figure 4

Hepatic Di signaling increases ROS production, JNK phosphorylation, and OCR.

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Hepatic Di signaling increases ROS production, JNK phosphorylation, and ...
(A–C and E) Studies using Hep-Di hepatocytes. (D) Studies using control hepatocytes. (A) Enhanced ROS production caused by hepatocyte Di signaling was PTX sensitive. CNO (10 μM) treatment of Hep-Di hepatocytes led to time-dependent increases in ROS production. These responses were completely abolished in the presence of PTX (300 ng/ml). Data represent the mean ± SEM from 3 independent experiments. (B) Increased p-JNK caused by hepatocyte Di signaling was NAC sensitive. Primary hepatocytes from Hep-Di mice were incubated with CNO (10 μM) for the indicated durations at 37°C, followed by Western blot analysis of cell lysates. Incubations were done with or without the ROS scavenger NAC (5 mM). Representative immunoblots are shown. Quantitative data represent the mean ± SEM from 7 independent experiments (CNO incubation time: 15 min). p-JNK expression levels were normalized to total JNK expression. (C and D) Measurement of the OCR of primary mouse hepatocytes. CNO (10 μM) treatment of Hep-Di hepatocytes (C) caused a marked increase in the OCR. These effects were almost abolished in the presence of PTX (300 ng/ml). The mitochondrial inhibitors antimycin A and rotenone demonstrated that the increased OCR derived from mitochondrial metabolism. CNO had no significant effect on the OCR in control hepatocytes (D). The curves shown are representative of 3 independent experiments. The OCR was measured using Seahorse technology. (E) Glucose output assays were performed with primary Hep-Di hepatocytes. The ability of CNO (10 μM) and glucagon (100 nM) to stimulate glucose release from primary Hep-Di hepatocytes was examined. Experiments were done with or without NAC (5 mM). Data represent the mean ± SEM from 7 (B) or 3 (A and E) independent experiments. *P < 0.05, **P < 0.01, and ***P < 0.001, compared with the corresponding control group. Statistical significance was determined by (B and E) 2-way ANOVA followed by Bonferroni’s post-hoc test and (A, C, and D) 2-tailed Student’s t test. See complete unedited blots in the supplemental material.