Fernando Díaz

Role of 17β-estradiol administration on insulin sensitivity in the rat: implications for the insulin receptor

The role of 17beta-estradiol in the early steps of insulin action is only partially known, although its effect on glucose homeostasis has been reported. In this paper, we attempt to prove the influence of 17beta-estradiol on the insulin receptor of ovariectomized rats treated with different hormonal doses. Our results show that high doses of estradiol impair insulin sensitivity while low doses improve it. We think that these results are the consequence of changes at a molecular level, because high doses of estradiol produced lower expression of the insulin receptor gene, lower content of this receptor in target tissues, and lower phosphorylation of insulin receptor in these tissues. However, low doses of estradiol seem to produce just the opposite. The possible existence of consensus response elements in the insulin receptor gene promoter to estradiol could be controlling the expression of this gene, this control being dose and timing dependent. Moreover, we cannot discard a possible effect of estradiol on the activity of protein tyrosine phosphatases, and therefore, on the activity of the insulin receptor. These new findings improve knowledge about the possible risk for insulin resistance in women taking oral contraceptives or receiving hormonal replacement therapy around the menopause, but could also open the door towards the possible utilization of 17beta-estradiol in some diabetes cases.

Sex-related differences in spatial learning during the early postnatal development of the rat

Some authors have reported that male rats younger than 21 days old are unable to perform spatial learning correctly because they have still not developed the ability to use extra-maze cues. In experiment 1, we analyzed spatial learning in 14-, 21-, 30- and 42-day-old rats using the Morris water maze (MWM). According to our results, a good performance was observed in 30-day-old male rats whereas this was not observed in female rats until they were 42 days old. In experiment 2 we studied the role of sex hormones in this kind of learning using the MWM and 30-day-old rats (castrated male rats and female rats treated with testosterone propionate (TP) after birth). The latter group, the male control group and the castrated males all solved the task correctly. The objective of experiment 3 was to determine possible differences between the sexes in the use of taxon strategies in the T water maze. To summarize, sexual dimorphism was only observed in spatial learning during development.

Effects of pregnancy on insulin receptor in liver, skeletal muscle and adipose tissue of rats

The mechanism responsible for insulin resistance during pregnancy remains unclear. Considerable evidence indicates that the insulin receptor plays an important role in insulin sensitivity. It seems possible that the hormonal milieu during gestation could have an effect on the insulin receptor. In the present study, measurements of tyrosine phosphorylation and protein content of the insulin receptor and expression of its gene in liver, skeletal muscle and adipose tissue indicate that during pregnancy significant changes occur in these parameters. We found that at the end of early gestation (day 10), muscle and adipose tissue are very sensitive to insulin action because the amount, phosphorylation and gene expression of insulin receptor is higher than in late gestation (days 15–20), while the tissue which is most sensitive to insulin action in late gestation is the liver. Our hypothesis is that these results are connected with changes in the concentrations of estradiol and progesterone observed during pregnancy. In conclusion, our previous and present findings seem to demonstrate that the different concentrations of gestational hormones play an important role in insulin sensitivity in this period and that each tissue responds in the most appropriate manner to guarantee the gestation in its entirety.

Soy isoflavones, diet and physical exercise modify serum cytokines in healthy obese postmenopausal women

OBJECTIVES Evaluate the effect of diet, physical exercise, and a daily oral intake of a soy isoflavones extract (Fisiogen(®)) contained 200 mg of Glycine max, which corresponded to 80 mg of isoflavone (60.8 mg of genistein, 16 mg of daidzein and 3.2 mg of glicitein) on leptin and other adipokines plasma levels in healthy obese postmenopausal women. METHODS A multicentric randomized longitudinal prospective cohort study was conducted in a sample of 87 healthy obese postmenopausal women. Patients were randomly assigned to a 1200 kcal diet and exercise group (control group) or a group of 1200 kcal diet, exercise, and daily oral intake of daily oral intake of a soy isoflavones extract (Fisiogen(®)) contained 200 mg of Glycine max, which corresponded to 80 mg of isoflavone (60.8 mg of genistein, 16 mg of daidzein and 3.2 mg of glicitein) (soy isoflavones group) along 6 months. Main outcome measures were: anthropometric measures, body composition, leptin, adiponectin, TNF-alpha, homocysteine, C-reactive protein, glucose, insulin, lipid profile and oestradiol serum levels, Kupperman index and Cervantes Scale. RESULTS Mean serum leptin and TNF-alpha levels declined after 6 months in both groups of the study, but only women in the soy isoflavones group showed a significant increase of mean serum levels of adiponectin. CONCLUSIONS Diet, physical exercise and daily oral intake of a soy isoflavones extract (Fisiogen(®)) contained 200 mg of Glycine max, which corresponded to 80 mg of isoflavone (60.8 mg of genistein, 16 mg of daidzein and 3.2 mg of glicitein) have a beneficial effect on serum leptin, adiponectin and TNF-α in healthy obese postmenopausal women after 6 months of treatment.

Chronic 17β-estradiol treatment improves skeletal muscle insulin signaling pathway components in insulin resistance associated with aging

Insulin resistance is a common feature of aging in both humans and rats. In the case of females, it seems to be related to loss of gonadal function, due mainly due to a decrease in plasma estrogen levels. Several causes have been postulated for this insulin resistance, among them changes in several steps of the insulin pathway. In view of these findings, the purpose of the present study was to examine the role of chronic 17β-estradiol treatment on insulin sensitivity during the aging process, and its effects on levels of the insulin-sensitive glucose transporter Glut4 (both total and plasma membrane localized), the interaction between p85α subunit of PI3-k and IRS-1, Tyr- and Ser-612 phosphorylation of IRS-1 levels, and Ser-473 phosphorylation of Akt. The present findings indicate that 17β-estradiol treatment is able to minimize the deleterious effect of aging on insulin sensitivity, at least at the level of plasma membrane localized Glut4. Nevertheless further research is needed to determine this conclusively.

Acute effects of 17β-estradiol and genistein on insulin sensitivity and spatial memory in aged ovariectomized female rats

Aging is characterized by decline in metabolic function and insulin resistance, and both seem to be in the basis of neurodegenerative diseases and cognitive dysfunction. Estrogens prevent age-related changes, and phytoestrogens influence learning and memory. Our hypothesis was that estradiol and genistein, using rapid-action mechanisms, are able to modify insulin sensitivity, process of learning, and spatial memory. Young and aged ovariectomized rats received acute treatment with estradiol or genistein. Aged animals were more insulin-resistant than young. In each age, estradiol and genistein-treated animals were less insulin-resistant than the others, except in the case of young animals treated with high doses of genistein. In aged rats, no differences between groups were found in spatial memory test, showing a poor performance in the water maze task. However, young females treated with estradiol or high doses of genistein performed well in spatial memory task like the control group. Only rats treated with high doses of genistein showed an optimal spatial memory similar to the control group. Conversely, acute treatment with high doses of phytoestrogens improved spatial memory consolidation only in young rats, supporting the critical period hypothesis for the beneficial effects of estrogens on memory. Therefore, genistein treatment seems to be suitable treatment in aged rats in order to prevent insulin resistance but not memory decline associated with aging. Acute genistein treatment is not effective to restore insulin resistance associated to the early loss of ovarian function, although it can be useful to improve memory deficits in this condition.

Influence of gonadal steroids on the glial fibrillary acidic protein‐immunoreactive astrocyte population in young rat hippocampus

It is known that expression of glial fibrillary acidic protein (GFAP) as an astrocyte‐specific marker can be regulated by levels of circulating gonadal steroids during postnatal development. In addition, astrocytes play an important role in the physiology of the hippocampus, a brain region considered sexually dimorphic at the neuronal level in rodents. To evaluate the contribution of glial cells to gender‐related differences in the hippocampus, we estimated the number of GFAP‐immunoreactive (GFAP‐IR) astrocytes in the hippocampus (CA1 and CA3 areas, dorsal and ventral regions) of male and female rats aged 30 days. Groups of 30‐day‐old masculinized females (TP‐females; injected with testosterone propionate at birth) and feminized males (FLU‐males, castrated and treated with flutamide, an androgen receptor antagonist) were included to assess the effects of gonadal hormones on these hippocampal astrocytes. Using the optical fractionator method, the total number of GFAP‐IR cells found in CA1 and CA3 areas was significantly higher in males compared to that in age‐matched females. This numerical pattern was reversed in TP‐females and FLU‐males in both hippocampal areas. In addition, more GFAP‐IR cells were found in dorsal hippocampus than in the ventral region in the CA1 area from all experimental groups, whereas this result was found in the CA3 area from males and TP‐females. Our results suggest an essential contribution of gonadal hormones to gender differences found in the astrocyte population of the rat hippocampus during development.

17β-Estradiol and/or progesterone protect from insulin resistance in STZ-induced diabetic rats

Recent clinical and experimental evidences suggest that sex steroids protect from insulin resistance associated with diabetes. Therefore, we have assessed the influence of E2 and/or P4 on insulin sensitivity by euglicaemic-hyperinsulinaemic clamp in ovariectomized streptozotocin-induced diabetic rats, focusing on key proteins of insulin signaling in skeletal muscle. Although low plasma levels of E2 (days 6 and 11) increased Glut-4 plasma membrane content and subsequent improved insulin sensitivity, they could not fully reverse hyperglycaemia negative effects on p85alpha-IRS-1 association and IRS-1 content during 11 days. However, high plasma levels of E2 (day 16) could reverse hyperglycaemia effects not only on Glut-4 plasma membrane content but also on p85alpha-IRS-1 association and IRS-1 protein content level. In contrast, P4 treatment only improved insulin sensitivity when its plasma concentration was low (days 6 and 11) and its effects were not associated with any proteins study in this paper. The combined therapy had a synergic effect on insulin sensitivity when their plasma levels were low (day 6) or high (day 16), that could be associated with Glut-4 plasma membrane content modulation, p85alpha-IRS-1 association and IRS-1 amount. These new findings improve our understanding of biochemical basis of insulin resistance due to hyperglycaemia and could open up new possibilities of treatment in uncontrolled type 1 DM.

Leptin and its receptor are controlled by 17β-estradiol in peripheral tissues of ovariectomized rats

It has been widely shown that there is a complex interaction between sex steroids and leptin effects on body weight. In this sense, the absence of female sex steroids is linked to a significant increase in body weight, which seems to be related to an impairment of the central actions of leptin. The present study was designed to elucidate the effects of two different treatments with 17β-estradiol on leptin receptor and serum leptin levels in ovariectomized rats, a model of postmenopausal condition. Our results have shown that plasma leptin levels in ovariectomized rats were lower than in estradiol-treated animals, thereby supporting a positive effect of this steroid. Recent information has extended leptin actions to peripheral tissues, mainly to insulin-dependent tissues, this effect being related to metabolic actions. To better understand the peripheral effects of leptin and their possible regulation by estradiol treatment, we have analyzed leptin receptor expression in the skeletal muscle and the adipose tissue. Our results showed a tissue-specific regulation of this protein: Ob-Rb expression in the adipose tissue decreased when the time of treatment or the dose of estradiol administered increased, suggesting less sensitivity to leptin in this tissue, whereas in the skeletal muscle the changes in this protein followed the same profile as the plasma leptin levels. We think that this specific regulation could ensure a different response of each tissue toward the same serum leptin level. Further studies to clarify this situation are ongoing.

Insulin sensitivity in streptozotocin‐induced diabetic rats treated with different doses of 17β‐oestradiol or progesterone

It has been reported that in streptozotocin (STZ)‐induced diabetes, hyperglycaemia leads to progressive insulin resistance of the peripheral tissues. In this study, we tried to elucidate the effects of hyperglycaemia on insulin sensitivity and insulin signalling in ovariectomized (STZ)‐induced diabetic rats. In addition, we attempted to demonstrate the role of 17β‐oestradiol and progesterone on insulin sensitivity, focusing on their effects on key proteins of skeletal muscle, insulin receptor (IR) and glucose transporter‐4 (Glut‐4). Our results show that hyperglycaemia could modulate insulin signalling, at the IR and Glut‐4 level, in different ways depending on exposure time. 17β‐Oestradiol and progesterone have different effects on insulin signalling. 17β‐Oestradiol treatment improves insulin sensitivity, but its action is dependent on the exposure time and its plasma level. During the early period of treatment (days 6–11), this hormone counteracts the effects of hyperglycaemia downstream of the IR, whereas during the later period of treatment (days 11–16), it may counteract the effects of hyperglycaemia by modulating IR relative tyrosine phosphorylation. By contrast, progesterone only improves insulin sensitivity during the early period of treatment (days 6–11), and this effect is not associated with changes in IR and Glut‐4 content. Both hormones have a protective role in skeletal muscle against the effects of glucose toxicity, but their effects begin at different stages of treatment. These new findings improve our understanding of insulin resistance in type 1 diabetes mellitus and of the risk/benefit ratio when 17β‐oestradiol and progesterone are used in oral contraceptives or hormone replacement therapy taken by menopausal women with controlled type 1 diabetes mellitus.