Role of Phosphatidylinositol-3-Kinase in Insulin Receptor Signaling: Studies with Inhibitor, LY294002

Sánchez‐Margalet, Víctor; Goldfine, I D; Vlahos, Chris J.; Sung, Chin K.

doi:10.1006/bbrc.1994.2480

Cited by 123 publications

(73 citation statements)

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“…PI3K plays a central role in regulating glucose transport and glycogen synthesis (Sánchez-Margalet et al 1994, Sánchez-Margalet 2000. In this study, we show that short-term incubation with homocysteine thiolactone inhibits insulin-receptor signaling and insulinstimulated glycogen synthesis.…”

Section: Introductionmentioning

confidence: 56%

Homocysteine thiolactone inhibits insulin signaling, and glutathione has a protective effect

Najib¹,

Sanchez-Margalet²

2001

Journal of Molecular Endocrinology

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Hyperhomocysteinemia and insulin resistance are independent factors for cardiovascular disease. Most of the angiotoxic effects of homocysteine are related to the formation of homocysteine thiolactone and the consequent increase in oxidative stress. The oxidative stress has also been shown to impair insulin action, therefore leading to insulin resistance. In order to study a putative direct effect of homocysteine on insulin signaling, we have characterized the molecular counter-regulation of the early events in the signal transduction of the insulin receptor, and the metabolic end-point of glycogen synthesis. We employed HTC rat hepatoma cells transfected with the human insulin receptor. A 10 min exposure to homocysteine thiolactone (50 µM) resulted in a significant inhibition of insulin-stimulated tyrosine phosphorylation of the insulin receptor -subunit and its substrates IRS-1 and p60-70, as well as their association with the p85 regulatory subunit of phosphatidylinositol 3-kinase.These effects led to impairment of the insulinstimulated phosphatidylinositol 3-kinase activity, which plays a central role in regulating insulin action. Thus, insulin-stimulated glycogen synthesis was also inhibited by homocysteine thiolactone. To investigate whether oxidative stress was mediating the counter-regulatory effect of homocysteine thiolactone on insulin signaling, we preincubated the cells (5 min) with 250 µM glutathione prior to the incubation with homocysteine (10 min) and subsequent insulin challenge. Glutathione completely abolished the effects of homocysteine thiolactone on insulin-receptor signaling and restored the insulin-stimulated glycogen synthesis. In conclusion, these data suggest that homocysteine thiolactone impairs insulin signaling by a mechanism involving oxidative stress, leading to a defect in insulin action.

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Section: Introductionmentioning

confidence: 56%

Homocysteine thiolactone inhibits insulin signaling, and glutathione has a protective effect

Najib¹,

Sanchez-Margalet²

2001

Journal of Molecular Endocrinology

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“…Insulin increases glucose uptake into cells, partly through the translocation of GLUT4 from intracellular compartments to the plasma membrane in muscle and adipose tissues [37]. Distinct experimental approaches have led to the conclusion that PI 3-kinase is necessary for insulin-stimulated GLUT4 translocation [38][39][40][41][42][43][44]. Evidence from other sources has also demonstrated a correlation between PI 3-kinase activity and glycogen metabolism [45,46].…”

Section: Discussionmentioning

confidence: 99%

Defects in insulin signal transduction in liver and muscle of pregnant rats

et al. 1997

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Insulin resistance during pregnancy has been reported both in women and in experimental animals [1][2][3][4][5][6][7], but the mechanism of this resistance remains unclear. In vivo studies have shown that pregnant rats become progressively resistant to insulin after day 16 of gestation [5]. In these animals, the insulin resistance of skeletal muscle has been clearly demonstrated in vitro, while the insulin resistance in vivo in peripheral tissues and liver has been substantiated in studies using the euglycaemic-hyperinsulinaemic clamp technique [6,7].Insulin initiates its metabolic and growth-promoting effects by binding to the a subunit of its tetrameric receptor, thereby activating the kinase in the b subunit [8]. This interaction catalyses the intramolecular autophosphorylation of specific tyrosine residues of the b subunit which further enhances the tyrosine kinase activity of the receptor toward other protein substrates [8]. In most cells, this primary event leads to the subsequent tyrosyl phosphorylation of a cytoplasmic protein with an apparent molecular weight of 160-185 kDa, called insulin receptor substrate 1 (IRS-1) [9][10][11]. Considerable evidence indicates that insulin receptor tyrosine Diabetologia (1997) 40: 179-186 Defects in insulin signal transduction in liver and muscle of pregnant rats Summary Pregnancy is known to induce insulin resistance, but the exact molecular mechanism involved is unknown. In the present study, we have examined the levels and phosphorylation state of the insulin receptor and of insulin receptor substrate 1(IRS-1), as well as the association between IRS-1 and phosphatidylinositol 3-kinase (PI 3-kinase) in the liver and muscle of pregnant rats (day 20 of gestation) by immunoprecipitation and immunoblotting with anti-insulin receptor, anti-IRS-1, anti-PI 3-kinase and antiphosphotyrosine antibodies. There were no changes in the insulin receptor concentration in the liver and muscle of pregnant rats. However, insulin stimulation of receptor autophosphorylation, as determined by immunoblotting with antiphosphotyrosine antibody, was reduced by 30 ± 6 % (p < 0.02) in muscle and 36 ± 5 % (p < 0.01) in liver at day 20 of gestation. IRS-1 protein levels decreased by 45 ± 6 % (p < 0.002) in liver and by 56 ± 9 % (p < 0.002) in muscle of pregnant rats. In samples previously immunoprecipitated with anti-IRS-1 antibody and blotted with antiphosphotyrosine antibody, the insulin-stimulated IRS-1 phosphorylation levels in the muscle and liver of pregnant rats decreased by 70 ± 9 % (p < 0.01) and 75 ± 8 % (p < 0.01), respectively. The insulin-stimulated IRS-1 association with PI 3-kinase decreased by 81 ± 6 % in muscle (p < 0.01) and 79 ± 11 % (p < 0.01) in the liver during pregnancy. These data suggest that changes in the early steps of insulin signal transduction may have a role in the insulin resistance observed in pregnancy. [Diabetologia (1997) 40: 179-186]

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“…In preliminary experiments (our unpublished data), we verified that E 2 promoted cell survival/growth under low serum conditions and further established that the PI-3K inhibitor wortmannin inhibited this effect of E 2 . To confirm the requirement of PI-3K activity for the cell survival induced by E 2 , MCF7 cells were treated with E 2 in the presence or absence of LY294002, a specific PI-3K inhibitor (Sanchez-Margalet et al, 1994) in serum-free medium. In the absence of E 2 , there was significant cell loss reflected in MTT uptake over a 5-d period (p Ͻ 0.05).…”

Section: Inhibition Of Apoptosis In Mcf7mentioning

confidence: 99%

Estradiol Abrogates Apoptosis in Breast Cancer Cells through Inactivation of BAD: Ras-dependent Nongenomic Pathways Requiring Signaling through ERK and Akt

Fernando

Wimalasena

2004

MBoC

113

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Estrogens such as 17-β estradiol (E2) play a critical role in sporadic breast cancer progression and decrease apoptosis in breast cancer cells. Our studies using estrogen receptor-positive MCF7 cells show that E2 abrogates apoptosis possibly through phosphorylation/inactivation of the proapoptotic protein BAD, which was rapidly phosphorylated at S112 and S136. Inhibition of BAD protein expression with specific antisense oligonucleotides reduced the effectiveness of tumor necrosis factor-α, H2O2, and serum starvation in causing apoptosis. Furthermore, the ability of E2 to prevent tumor necrosis factor-α-induced apoptosis was blocked by overexpression of the BAD S112A/S136A mutant but not the wild-type BAD. BAD S112A/S136A, which lacks phosphorylation sites for p90RSK1 and Akt, was not phosphorylated in response to E2 in vitro. E2 treatment rapidly activated phosphatidylinositol 3-kinase (PI-3K)/Akt and p90RSK1 to an extent similar to insulin-like growth factor-1 treatment. In agreement with p90RSK1 activation, E2 also rapidly activated extracellular signal-regulated kinase, and this activity was down-regulated by chemical and biological inhibition of PI-3K suggestive of cross talk between signaling pathways responding to E2. Dominant negative Ras blocked E2-induced BAD phosphorylation and the Raf-activator RasV12T35S induced BAD phosphorylation as well as enhanced E2-induced phosphorylation at S112. Chemical inhibition of PI-3K and mitogen-activated protein kinase kinase 1 inhibited E2-induced BAD phosphorylation at S112 and S136 and expression of dominant negative Ras-induced apoptosis in proliferating cells. Together, these data demonstrate a new nongenomic mechanism by which E2 prevents apoptosis.

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Role of Phosphatidylinositol-3-Kinase in Insulin Receptor Signaling: Studies with Inhibitor, LY294002

Cited by 123 publications

References 0 publications

Homocysteine thiolactone inhibits insulin signaling, and glutathione has a protective effect

Homocysteine thiolactone inhibits insulin signaling, and glutathione has a protective effect

Defects in insulin signal transduction in liver and muscle of pregnant rats

Estradiol Abrogates Apoptosis in Breast Cancer Cells through Inactivation of BAD: Ras-dependent Nongenomic Pathways Requiring Signaling through ERK and Akt

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