Effects of epidermal growth factor on gluconeogenesis and cellular redox state do not require Na+/H+ exchange or Na+/K+-ATPase activities

Most reports on the effects of epidermal growth factor (EGF) on gluconeogenesis have indicated that such effects depend on the substrate used and are only observable after a lag time of 30-40 min. Recently, an immediate and transient effect of EGF on glucose synthesis was described in a perfused liver system. Here we extend the study of the effect of EGF on gluconeogenesis to isolated hepatocytes from fasted rats. The delayed effect of EGF on gluconeogenesis was studied by adding the substrate 40 min after the peptide. Under these conditions EGF increased glucose synthesis from pyruvate, decreased it when the substrate was lactate or glycerol and did not modify gluconeogensis from fructose or dihydroxyacetone. EGF did not affect the metabolic flux through glycolysis, determined as the production of lactate+pyruvate from 30 mM glucose. Furthermore, EGF did not modify the metabolic flux through pyruvate kinase, determined as the production of lactate+pyruvate from 1 mM dihydroxyacetone. The differing effects of EGF on gluconeogenesis depending on the substrate used can be explained by the effects of EGF on the cytosolic redox state (measured as the lactate/pyruvate ratio). About 20 min after the addition of EGF, the mitochondrial redox state (measured as the 3-hydroxybutyrate/acetoacetate ratio) decreased. This effect of EGF was blocked by ammonium, which also abolished the effect of the peptide on gluconeogenesis. Thus the effect of EGF at the mitochondrial level appears to be necessary for its effects on gluconeogenesis. Taken together, our results indicate that the delayed effects of EGF on gluconeogenesis are secondary to the effects of the peptide at both the mitochondrial and cytosolic levels. In addition to these delayed effects, we observed that EGF rapidly and transiently stimulated glucose synthesis from lactate, decreased the cytosolic redox state and increased oxygen consumption. All of these rapid effects required the presence of extracellular calcium and disappeared in the presence of rotenone, suggesting that this rapid effect of EGF on gluconeogenesis is secondary to the stimulation of mitochondrial respiration.

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Extracellular cysteine/cystine redox regulates the p44/p42 MAPK pathway by metalloproteinase-dependent epidermal growth factor receptor signaling

AJP Gastrointestinal and Liver Physiology ◽

10.1152/ajpgi.00280.2004 ◽

2005 ◽

Vol 289 (1) ◽

pp. G70-G78 ◽

Cited By ~ 46

Author(s):

Yvonne S. Nkabyo ◽

Young-Mi Go ◽

Thomas R. Ziegler ◽

Dean P. Jones

Keyword(s):

Epidermal Growth Factor Receptor ◽

Growth Factor ◽

Epidermal Growth Factor ◽

Redox State ◽

Mapk Pathway ◽

Growth Factor Receptor ◽

Mitogen Activated Protein Kinase ◽

Growth Factor Signaling ◽

Epidermal Growth ◽

Mapk Inhibitors

Previous research shows that stimulation of proliferation of colon carcinoma (Caco-2) cells by a more reduced extracellular cysteine/cystine (Cys/CySS) redox state occurs with no apparent effect on intracellular glutathione and that this stimulation is lost on addition of epidermal growth factor. The purpose of the present study was to determine whether a more reduced extracellular Cys/CySS redox state activates the mitogenic p44/p42 mitogen-activated protein kinase (MAPK) pathway and whether this is signaled through the epidermal growth factor receptor (EGFR). Caco-2 cells were exposed to a range of physiological extracellular redox conditions from −150 to 0 mV. In the absence of added growth factors, the most reduced (−150 mV) redox state induced an 80% increase in EGFR phosphorylation, and this was followed by a marked increase in phosphorylation of p44/p42 MAPK. Inhibitors of EGFR (AG1478) and p44/p42 MAPK (U0126) phosphorylation blocked redox-dependent p44/p42 phosphorylation, indicating that signaling occurred by EGFR. These effects were inhibited by pretreatment with a nonpermeant alkylating agent, showing that signaling involved thiols accessible to the extracellular space. The EGFR ligand TGF-α was increased in culture medium at more reduced redox states. Redox-dependent phosphorylation of EGFR was completely prevented by a metalloproteinase inhibitor (GM6001), and an antibody to TGF-α partially inhibited the phosphorylation of p44/p42 MAPK by redox. Thus the data show that a redox-dependent activation of metalloproteinase can stimulate the mitogenic p44/p42 MAPK pathway by a TGF-α-dependent mechanism. Because Cys availability and Cys/CySS redox are dependent on nutrition, disease, and environmental exposures, the results suggest that cell proliferation could be influenced physiologically by Cys-dependent redox effects on growth factor signaling pathways.

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