Signalling pathways involved in the stimulation of glycogen synthesis by insulin in rat hepatocytes

Peak, Matthew; Rochford, Justin J.; Borthwick, A C; Yeaman, Stephen J.; Agius, Loranne

doi:10.1007/s001250050861

Cited by 57 publications

(56 citation statements)

References 32 publications

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“…ERK2, P70 S6k , and PKB were assayed after immunoprecipition with their respective antibodies bound to protein A-sepharose CL-4B (ERK2 and P70 S6k ) or pansorbin (PKB), as described previously. 35 Kinase activities were expressed as milliunits per milligram of protein, in which 1 unit corresponds to the incorporation of 1 nmol of phosphate per minute. JNK1 and p38 MAPK activities were also assayed as previously described, 36 but with some modifications.…”

Section: Methodsmentioning

confidence: 99%

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Inhibition of rat hepatocyte proliferation by transforming growth factor ? and glucagon is associated with inhibition of ERK2 and p70 S6 kinase

Dixon¹,

Agius²,

Yeaman

et al. 1999

Hepatology

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Stimulation of hepatocyte proliferation by epidermal growth factor (EGF) and insulin is inhibited by transforming growth factor ␤ (TGF-␤) and by glucagon. It is also suppressed by inhibitors of various protein kinases, including rapamycin, which blocks activation of p70 S6 kinase (p70 S6k ), PD98059, which inhibits the activation of extracellular-regulated kinase (ERK), and SB 203580, an inhibitor of the p38 mitogen-activated protein kinase (p38 MAPK). In this study, we investigated whether the inhibition of proliferation by TGF-␤ involves these protein kinase cascades. Culture of hepatocytes with TGF-␤ for 16 hours decreased the stimulation by EGF of ERK2 and p70 S6k (by 50% and 35%, respectively), but did not affect the stimulation of either p38 MAPK, c-jun NH 2 -terminal kinase (JNK), or protein kinase B (PKB). Culture of hepatocytes with glucagon for 16 hours also inhibited the stimulation by EGF of activation of ERK2 and p70 S6k (by E50%). The inhibitory effects of glucagon were observed when the hormone was added either 10 minutes or 60 minutes before EGF addition, whereas no effects of TGF-␤ were observed after 10-minute or 60-minute incubation. These results suggest that the inhibition of hepatocyte proliferation by TGF-␤ may be in part mediated by inhibition of ERK2 and p70 S6k , but does not involve PKB, JNK, or p38 MAPK. Unlike glucagon, the effects of TGF-␤ are not elicited in response to short-term treatment. (HEPATOLOGY 1999;29:1418-1424 Hepatocyte proliferation following injury or partial hepatectomy is regulated by the action of both stimulatory (mitogenic) and inhibitory growth factors and hormones (reviewed in Michalopoulos 1 and Fausto 2 ). Studies in primary hepatocyte cultures have identified several complete mitogens capable of inducing DNA synthesis in serum-free medium,

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Section: Methodsmentioning

confidence: 99%

“…ERK1 and JNK2 were not assayed in view of our previous studies demonstrating very low levels of both kinase activities in rat hepatocytes. 35 Statistical Analysis. All values given are means Ϯ SEM for the number of hepatocyte preparations indicated, and statistical analysis was by the paired Student t test.…”

Section: Methodsmentioning

confidence: 99%

Inhibition of rat hepatocyte proliferation by transforming growth factor ? and glucagon is associated with inhibition of ERK2 and p70 S6 kinase

Dixon¹,

Agius²,

Yeaman

et al. 1999

Hepatology

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“…The immobilised immune complexes were recovered by centrifugation at 13,000×g, 4°C for 2 min and washed three times with lysis buffer. The pellet was incubated at 30°C for 30 min with 20 μl of assay mixture containing 25 mmol/l β-glycerophosphate, pH 7.4, 100 mmol/l sodium chloride, 10 mmol/l magnesium chloride, 2.5 μmol/l protein kinase A inhibitor, 100 nmol/l okadaic acid, 100 μmol/l crosstide, 50 μmol/l [γ 32 P]ATP as in [3].…”

Section: Enzyme Activity Determinationmentioning

confidence: 99%

“…The stimulation of glycogen synthesis [3,4], like insulin action in other cell types, is thought to occur downstream from the activation of the lipid kinase phosphoinositide-3 (PI-3)-kinase [5,6]. The lipid products of PI-3-kinase activate a member of the AGC class of protein kinases termed phosphoinositide-dependent protein kinase 1 (PDK-1), which itself can phosphorylate and activate other protein kinases of the AGC family.…”

Section: Introductionmentioning

confidence: 99%

The role of protein kinase B/Akt in insulin-induced inactivation of phosphorylase in rat hepatocytes

et al. 2005

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Aims/hypothesis: An insulin signalling pathway leading from activation of protein kinase B (PKB, also known as Akt) to phosphorylation (inactivation) of glycogen synthase kinase-3 (GSK-3) and activation of glycogen synthase is well characterised. However, in hepatocytes, inactivation of GSK-3 is not the main mechanism by which insulin stimulates glycogen synthesis. We therefore tested whether activation of PKB causes inactivation of glycogen phosphorylase. Materials and methods: We used a conditionally active form of PKB, produced using recombinant adenovirus, to test the role of acute PKB activation in the control of glycogen phosphorylase and glycogen synthesis in hepatocytes. Results: Conditional activation of PKB mimicked the inactivation of phosphorylase, the activation of glycogen synthase, and the stimulation of glycogen synthesis caused by insulin. In contrast, inhibition of GSK-3 caused activation of glycogen synthase but did not mimic the stimulation of glycogen synthesis by insulin. PKB activation and GSK-3 inhibition had additive effects on the activation of glycogen synthase, indicating convergent mechanisms downstream of PKB involving inactivation of either phosphorylase or GSK-3. Glycogen synthesis correlated inversely with the activity of phosphorylase-a, irrespective of whether this was modulated by insulin, by PKB activation or by a selective phosphorylase ligand, supporting an essential role for phosphorylase inactivation in the glycogenic action of insulin in hepatocytes. Conclusions/interpretation: In hepatocytes, the acute activation of PKB, but not the inhibition of GSK-3, mimics the stimulation of glycogen synthesis by insulin. This is explained by a pathway downstream of PKB leading to inactivation of phosphorylase, activation of glycogen synthase, and stimulation of glycogen synthesis, independent of the GSK-3 pathway.

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“…3,[36][37][38] To underline this in the present study, a concentration dependence was performed. Cells were treated with 0.1 nmol/L glucagon for 2 hours in the absence or presence of 100 ng/mL IL-6 or 10 nmol/L insulin with or without increasing concentrations of wortmannin.…”

Section: Attenuation Of Thementioning

confidence: 99%

Phosphatidylinositol 3-kinase and protein kinase C contribute to the inhibition by interleukin 6 of phosphoenolpyruvate carboxykinase gene expression in cultured rat hepatocytes

2000

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Signalling pathways involved in the stimulation of glycogen synthesis by insulin in rat hepatocytes

Cited by 57 publications

References 32 publications

Inhibition of rat hepatocyte proliferation by transforming growth factor ? and glucagon is associated with inhibition of ERK2 and p70 S6 kinase

Inhibition of rat hepatocyte proliferation by transforming growth factor ? and glucagon is associated with inhibition of ERK2 and p70 S6 kinase

The role of protein kinase B/Akt in insulin-induced inactivation of phosphorylase in rat hepatocytes

Phosphatidylinositol 3-kinase and protein kinase C contribute to the inhibition by interleukin 6 of phosphoenolpyruvate carboxykinase gene expression in cultured rat hepatocytes

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