Aparecida Emiko Hirata

Modulation of IR/PTP1B interaction and downstream signaling in insulin sensitive tissues of MSG-rats.

PTP1B has been shown to be a negative regulator of the insulin signal transduction in insulin resistant states. Herein we investigated IR/PTP1B interaction and downstream signaling in insulin sensitive tissues of 10 and 28-week-old MSG-insulin resistant rats which represent different stages of insulin resistance. Our results demonstrated that the increase in PTP1B expression and/or association with IR in MSG animals may contribute to the impaired insulin signaling mainly in liver and muscle. Although, adipose tissue of 10-week-old MSG rats showed higher PTP1B expression and IR/PTP1B interaction, they were not sufficient to impair all insulin signaling since IRS-2 phosphorylation and association with PI3-kinase and Akt serine phosphorylation were increased, which may contribute for the increased adiposity of these animals. In 28-week-old-MSG rats there was an increase in IR/PTP1B interaction and reduced insulin signaling in liver, muscle and adipocytes, and a more pronounced insulin resistance.

Regulation of insulin receptor substrate-2 tyrosine phosphorylation in animal models of insulin resistance.

Insulin induces a wide variety of growth and metabolic responses in many cell types. These actions are initiated by insulin binding to its receptor and involve a series of alternative and complementary pathways created by the multiple substrates of the insulin receptor (insulin receptor substrates [IRSs]). We investigated IRS-1 and IRS-2 tyrosine phosphorylation; their association with phosphatidylinositol-3-OH kinase (PI3-K); and the phosphorylation of Akt, a serine-threonine kinase situated downstream of PI3-K, in liver and muscle of two animal models of insulin resistance: epinephrine- or dexamethasone-treated rats. We used in vivo insulin infusion followed by tissue extraction, immunoprecipitation, and immunoblotting. IRS-1 and IRS-2 protein expression did not change in liver and muscle of the epinephrine-treated rats, but in dexamethasone-treated rats IRS-1 presented an increase in liver and a decrease in muscle tissue. PI3-K and Akt protein expression did not change in liver or muscle of the two animal models of insulin resistance. There was a downregulation in insulin-induced IRS-1 and IRS-2 tyrosine phosphorylation and association with PI3-K in both models of insulin resistance. In parallel, insulin-induced Akt phosphorylation was reduced in both tissues of epinephrine-treated rats, and in liver but not in muscle of dexamethasonetreated rats. The reduction in insulin-induced Akt phosphorylation may help to explain the insulin resistance in liver and muscle of epinephrine-treated rats and in the liver of dexamethasone-treated rats.

Regulation of IRS-2 tyrosine phosphorylation in fasting and diabetes.

Intracellular insulin signaling involves a series of alternative and complementary pathways created by the multiple substrates of the insulin receptor (IRS) and the various isoforms of the SH2 domain signaling molecules that can interact with substrate. In this study we investigated IRS-1 and IRS-2 tyrosine phosphorylation, their association with PI3-kinase and the phosphorylation of Akt, a serine-threonine kinase situated downstream to PI 3-kinase, in liver and muscle of two animal models of insulin resistance: 72 h of fasting and STZ-diabetic rats. There was an upregulation in insulin-induced IRS-1 and IRS-2 tyrosine phosphorylation and association with PI3-kinase in liver and muscle of both animal models of insulin resistance. However, Akt phosphorylation showed different regulation, increasing in fasting and decreasing in STZ-diabetic rats. Since an important difference between these two animal models of insulin resistance is the plasma glucose levels, we can suggest that in STZ diabetic rats, the reduction in Akt phosphorylation is probably related to hyperglycemia and may certainly contribute to the molecular mechanism of insulin resistance observed in these animals.

β3-Adrenergic-dependent and -independent mechanisms participate in cold-induced modulation of insulin signal transduction in brown adipose tissue of rats

During cold exposure, homeothermic animals mobilize glucose with higher efficiency than at thermoneutrality. An interaction between the insulin signal transduction machinery and high sympathetic tonus is thought to play an important role in this phenomenon. In the present study, rats were exposed to cold during 8 days and treated, or not, with a β3-adrenergic agonist, BRL37344 sodium 4-2-2-(3-chlorophenyl)-2-hydroxyethyl amino propyl phenoxy-acetic acid sodium (BRL37344), or antagonist, SR59230A 3-(2-ethylphenoxy)-[(1S)-1,2,3,4-tetrahydronaphth-1-ylamino]-(2S)-2-propanol oxalate (SR59230A), to evaluate the cross-talk between insulin and β3-adrenergic intracellular signaling in brown adipose tissue. The drugs did not modify food ingestion, body temperature, and body weight in control and cold-exposed rats. Treatment of control rats with BRL37344 led to higher insulin-induced tyrosine phosphorylation of the insulin receptors, insulin receptor substrate (IRS)-1 and ERK, higher insulin-induced IRS-1/PI3-kinase association, and higher [Ser473] phosphorylation of Akt. Cold exposure alone promoted higher insulin-induced tyrosine phosphorylation of the insulin receptors, IRS-1, IRS-2, and ERK, and higher insulin-induced IRS-1 and IRS-2/PI3-kinase association. Except for the regulation of ERK, SR59230A abolished all the cold-induced effects upon the insulin signal transduction pathway. However, this antagonist only partially inhibited the cold-induced increase of glucose uptake. Thus, the sympathetic tonus generated during cold-exposure acts, in brown adipose tissue, through the β3-adrenergic receptor and modulates insulin signal transduction, with the exception of ERK. However, insulin-independent mechanisms other than β3-adrenergic activation participate in cold-induced glucose uptake in brown adipose tissue of rats.