The anxiolytic drug opipramol inhibits insulin-induced lipogenesis in fat cells and insulin secretion in pancreatic islets

The antidepressant drug opipramol has been reported to exert antilipolytic effect in human adipocytes, suggesting that alongside its neuropharmacological properties, this agent might modulate lipid utilization by peripheral tissues. However, patients treated for depression or anxiety disorders by this tricyclic compound do not exhibit the body weight gain or the glucose tolerance alterations observed with various other antidepressant or antipsychotic agents such as amitriptyline and olanzapine, respectively. To examine whether opipramol reproduces or impairs other actions of insulin, its direct effects on glucose transport, lipogenesis and lipolysis were investigated in adipocytes while its influence on insulin secretion was studied in pancreatic islets. In mouse and rat adipocytes, opipramol did not activate triglyceride breakdown, but partially inhibited the lipolytic action of isoprenaline or forskolin, especially in the 10–100 μM range. At 100 μM, opipramol also inhibited the glucose incorporation into lipids without limiting the glucose transport in mouse adipocytes. In pancreatic islets, opipramol acutely impaired the stimulation of insulin secretion by various activators (high glucose, high potassium, forskolin...). Similar inhibitory effects were observed in mouse and rat pancreatic islets and were reproduced with 100 μM haloperidol, in a manner that was independent from alpha2-adrenoceptor activation but sensitive to Ca2+ release. All these results indicated that the anxiolytic drug opipramol is not only active in central nervous system but also in multiple peripheral tissues and endocrine organs. Due to its capacity to modulate the lipid and carbohydrate metabolisms, opipramol deserves further studies in order to explore its therapeutic potential for the treatment of obese and diabetic states. (AU)

The anxiolytic drug opipramol inhibits insulin-induced lipogenesis in fat cells and insulin secretion in pancreatic islets.

The antidepressant drug opipramol has been reported to exert antilipolytic effect in human adipocytes, suggesting that alongside its neuropharmacological properties, this agent might modulate lipid utilization by peripheral tissues. However, patients treated for depression or anxiety disorders by this tricyclic compound do not exhibit the body weight gain or the glucose tolerance alterations observed with various other antidepressant or antipsychotic agents such as amitriptyline and olanzapine, respectively. To examine whether opipramol reproduces or impairs other actions of insulin, its direct effects on glucose transport, lipogenesis and lipolysis were investigated in adipocytes while its influence on insulin secretion was studied in pancreatic islets. In mouse and rat adipocytes, opipramol did not activate triglyceride breakdown, but partially inhibited the lipolytic action of isoprenaline or forskolin, especially in the 10-100 µM range. At 100 µM, opipramol also inhibited the glucose incorporation into lipids without limiting the glucose transport in mouse adipocytes. In pancreatic islets, opipramol acutely impaired the stimulation of insulin secretion by various activators (high glucose, high potassium, forskolin...). Similar inhibitory effects were observed in mouse and rat pancreatic islets and were reproduced with 100 µM haloperidol, in a manner that was independent from alpha2-adrenoceptor activation but sensitive to Ca2+ release. All these results indicated that the anxiolytic drug opipramol is not only active in central nervous system but also in multiple peripheral tissues and endocrine organs. Due to its capacity to modulate the lipid and carbohydrate metabolisms, opipramol deserves further studies in order to explore its therapeutic potential for the treatment of obese and diabetic states.

The lack of Irs2 induces changes in the immunocytochemical expression of aromatase in the mouse retina.

Insulin receptor substrate (Irs) belongs to a family of proteins that mediate the intracellular signaling of insulin and IGF-1. Insulin receptor substrate 2 (Irs2) is necessary for retinal function, since its failure in Irs2-deficient mice in hyperglycemic situation promotes photoreceptor degeneration and visual dysfunction, like in diabetic retinopathy. The expression of P450 aromatase, which catalyzes androgen aromatization to form 17ß-estradiol, increases in some neurodegenerative diseases thus promoting the local synthesis of neuroestrogens that exert relevant neuroprotective functions. Aromatase is also expressed in neurons and glial cells of the central nervous system (CNS), including the retina. To further understand the role of Irs2 at the retinal level, we performed an immunocytochemical study in adult normoglycemic Irs2-deficient mice. For this aim, the retinal immunoexpression of neuromodulators, such as aromatase, glutamine synthetase (GS), and tyrosine hydroxylase (TH) was analyzed, joint to a morphometric and planimetric study of the retinal layers. Comparing with wild-type (WT) control mice, the Irs2-knockout (Irs2-KO) animals showed a significant increase in the immunopositivity to aromatase in almost all of the retinal layers. Besides, Irs2-KO mice exhibited a decreased immunopositive reaction for GS and TH, in Müller and amacrine cells, respectively; morphological variations were also found in these retinal cell types. Furthermore, the retina of Irs2-KO mice displayed alterations in the structural organization, and a generalized decrease in the retinal thickness was observed in each of the layers, except for the inner nuclear layer. Our findings suggest that the absence of Irs2 induces retinal neurodegenerative changes in Müller and amacrine cells that are unrelated to hyperglycemia. Accordingly, in the Irs2-KO mice, the increased retinal immunocytochemical reactivity of aromatase could be associated with an attempt to repair such neural retina injuries by promoting local neuroprotective mediators.

Role of Flavonoids in The Interactions among Obesity, Inflammation, and Autophagy.

Nowadays, obesity is considered as one of the main concerns for public health worldwide, since it encompasses up to 39% of overweight and 13% obese (WHO) adults. It develops because of the imbalance in the energy intake/expenditure ratio, which leads to excess nutrients and results in dysfunction of adipose tissue. The hypertrophy of adipocytes and the nutrients excess trigger the induction of inflammatory signaling through various pathways, among others, an increase in the expression of pro-inflammatory adipocytokines, and stress of the endoplasmic reticulum (ER). A better understanding of obesity and preventing its complications are beneficial for obese patients on two facets: treating obesity, and treating and preventing the pathologies associated with it. Hitherto, therapeutic itineraries in most cases are based on lifestyle modifications, bariatric surgery, and pharmacotherapy despite none of them have achieved optimal results. Therefore, diet can play an important role in the prevention of adiposity, as well as the associated disorders. Recent results have shown that flavonoids intake have an essential role in protecting against oxidative damage phenomena, and presents biochemical and pharmacological functions beneficial to human health. This review summarizes the current knowledge of the anti-inflammatory actions and autophagic flux of natural flavonoids, and their molecular mechanisms for preventing and/or treating obesity.

Evidences for Expression and Location of ANGPTL8 in Human Adipose Tissue.

The metabolism of triglycerides (TGs) is regulated, among others, by the lipoprotein lipase (LPL) that hydrolyses the TGs on endothelial cells. In turn, LPL is inhibited by the ANGPTLs family of proteins, such as ANGPTL3, 4, and, 8; the latter is the least known. In this work, we have tried to establish the expression and localisation of the Angiopoietin-like 8 (ANGPTL8) protein in the visceral adipose tissue (VAT) of morbid-obese and non-obese patients. 109 subjects (66 women and 43 men) undergoing laparoscopic surgery participated in this study. A blood sample and a portion of the VAT were obtained, and the patients were classified according to their Body Mass Index (BMI) as non-obese (19.5-30 kg/m2) and morbid-obese (40-50 kg/m2). No significant changes in ANGPTL8 plasma levels were determined by EIA in obese patients. The immunocytochemistry and Western blotting showed the presence of increased ANGPTL8 in morbid-obese patients (p < 0.05). In-situ hybridisation and a real time polymerase chain reaction (RT-PCR) confirmed that the mRNA that encodes ANGPTL8 was present in adipocytes, without differences in their nutritional state (p = 0.89), and even in the endothelial cells. Our data suggests that ANGPT8 plasmatic levels do not change significantly in patients with morbid obesity, although there is a modest difference related to gender. Besides, we demonstrate that in visceral adipose tissue, ANGPTL8 is well defined in the cytoplasm of adipocytes coexisting with perilipin-1 and its mRNA, also is present in endothelial cells. These findings suggest the possibility that among other functions, ANGPTL8 could perform either a paracrine and/or an endocrine role in the adipose tissue.

The influence of the lack of insulin receptor substrate 2 (IRS2) on the thyroid gland.

Involvement of IRS2 in the proliferative effects of IGF-I of follicular thyroid cells has been described, but there are no evidences for in vivo participation of IRS2. This study aimed to analyse the in vivo relevance of IRS2 in the proliferation and apoptosis of thyroid cells by immunocytochemical studies for PCNA, Ki67, and active-caspase-3 in thyroid cells of IRS2 knockout (IRS2-KO) mice, jointly to TUNEL assay. Thyroid hormones were lower in IRS2-KO mice than in their wild-type (WT) counterparts. Increases in the area, perimeter and diameter of thyroid follicles of IRS2-KO mice were observed, which also showed increased proliferation rate of follicular cells and decreased percentage of apoptotic cells that was more evident in the central than in the marginal region of the gland. Sex-related differences were also found, since the follicular epithelium height was higher in male than in female mice. The percentage of proliferating cells showed significant changes in male but not in female mice, and apoptotic cells were more abundant in female than in male IRS2-KO animals, without significant differences between WT-animals. Therefore, our results suggest that IRS2 could be involved in the maintenance of thyroid cells population and in the normal physiology of the thyroid gland.

Prolactin system in the hippocampus.

Among the more than 300 biological actions described for prolactin, its role in the neurogenic capacity of the hippocampus, which increases synaptogenesis and neuronal plasticity, consolidates memory and acts as a neuronal protector against excitotoxicity-effects mediated through its receptors are more recently known. The detection of prolactin in the hippocampus and its receptors, specifically in the Ammon's horn and dentate gyrus, opened up a new field of study on the possible neuroprotective effect of hormones in a structure involved in learning and memory, as well as in emotional and behavioral processes. It is currently known, although controversial, that prolactin may be related to sex and age and that the hormone could be synthesized in the hippocampus itself. However, the regulatory mechanisms of changes in prolactin or in its hippocampal receptors still remain unknown. This review introduces the reader to general aspects concerning prolactin and its receptors and to what is currently known about the role prolactin plays in the brain and, in particular, in the hippocampus.

Sequential testicular atrophy involves changes in cellular proliferation and apoptosis associated with variations in aromatase P450 expression levels in Irs-2-deficient mice.

Insulin receptor substrate 2 (Irs-2) is an intracellular protein susceptible to phosphorylation after activation of the insulin receptor. Its suppression affects testis development and its absence induces peripheral resistance to insulin. The aim of this study was to identify changes induced by the deletion of Irs-2 in the testicular structure and by the altered expression of cytochrome P450 aromatase, a protein necessary for the development and maturation of germ cells. Adult knockout (KO) mice (Irs-2-/- , 6 and 12 weeks old) and age-matched wild-type (WT) mice were used in this study. Immunohistochemistry and Western blot analyses were performed to study proliferation (PCNA), apoptosis (active caspase-3) and P450 aromatase expression in testicular histological sections. Deletion of Irs-2 decreased the number of epithelial cells in the seminiferous tubule and rete testis. Aberrant cells were frequently detected in the epithelia of Irs-2-/- mice, accompanied by variations in spermatogonia, which were shown to exhibit small hyperchromatic nuclei as well as polynuclear and anuclear structures. The amount of cell proliferation was significantly lower in Irs-2-/- mice than in WT mice, whereas apoptotic processes were more common in Irs-2-/- mice. Aromatase P450 reactivity was higher in 6-week-old KO mice than in WT mice of the same age and was even higher at 12 weeks. Our results suggest that Irs-2 is a key element in spermatogenesis because silencing Irs-2 induces the sequential development of testicular atrophy. The effects are observed mainly in germ cells present in the seminiferous tubule, which may be due to changes in cytochrome P450 aromatase expression.

Variations in adrenal gland medulla and dopamine effects induced by the lack of Irs2

The adrenomedullary chromaffin cells' hormonal pathway has been related to the pathophysiology of diabetes mellitus. In mice, the deletion of insulin receptor substrate type 2 (Irs2) causes peripheral insulin resistance and reduction in Beta-cell mass, leading to overt diabetes, with gender differences on adrenergic signaling. To further unravel the relevance of Irs2 on glycemic control, we analyzed in adult Irs2 deficient (Irs2-/-) mice, of both sexes but still normoglycemic, dopamine effects on insulin secretion and glycerol release, as well as their adrenal medulla by an immunohistochemical and morphologic approach. In isolated islets, 10 μM dopamine significantly inhibited insulin release in wild-type (WT) and female Irs2−/− mice; however, male Irs2−/− islets were insensitive to that catecholamine. Similarly, on isolated adipocytes, gender differences were observed between WT and Irs2-/- mice in basal and evoked glycerol release with crescent concentrations of dopamine. By immunohistochemistry, reactivity to tyrosine hydroxylase (TH) in female mice was significantly higher in the adrenal medulla of Irs2-/- compared to WT; although no differences for TH-immunopositivity were observed between the male groups of mice. However, compared to their corresponding WT animals, adrenomedullary chromaffin cells of Irs2-/- mice showed a significant decrease in the cellular and nuclear areas, and even in their percentage of apoptosis. Therefore, our observations suggest that, together with gender differences on dopamine responses in Irs2-/- mice, disturbances in adrenomedullary chromaffin cells could be related to deficiency of Irs2. Accordingly, Irs2 could be necessary for adequate glucose homeostasis and maintenance of the population of the adrenomedullary chromaffin cells

Variations in adrenal gland medulla and dopamine effects induced by the lack of Irs2.

The adrenomedullary chromaffin cells' hormonal pathway has been related to the pathophysiology of diabetes mellitus. In mice, the deletion of insulin receptor substrate type 2 (Irs2) causes peripheral insulin resistance and reduction in ß-cell mass, leading to overt diabetes, with gender differences on adrenergic signaling. To further unravel the relevance of Irs2 on glycemic control, we analyzed in adult Irs2 deficient (Irs2-/-) mice, of both sexes but still normoglycemic, dopamine effects on insulin secretion and glycerol release, as well as their adrenal medulla by an immunohistochemical and morphologic approach. In isolated islets, 10 µM dopamine significantly inhibited insulin release in wild-type (WT) and female Irs2-/- mice; however, male Irs2-/- islets were insensitive to that catecholamine. Similarly, on isolated adipocytes, gender differences were observed between WT and Irs2-/- mice in basal and evoked glycerol release with crescent concentrations of dopamine. By immunohistochemistry, reactivity to tyrosine hydroxylase (TH) in female mice was significantly higher in the adrenal medulla of Irs2-/- compared to WT; although no differences for TH-immunopositivity were observed between the male groups of mice. However, compared to their corresponding WT animals, adrenomedullary chromaffin cells of Irs2-/- mice showed a significant decrease in the cellular and nuclear areas, and even in their percentage of apoptosis. Therefore, our observations suggest that, together with gender differences on dopamine responses in Irs2-/- mice, disturbances in adrenomedullary chromaffin cells could be related to deficiency of Irs2. Accordingly, Irs2 could be necessary for adequate glucose homeostasis and maintenance of the population of the adrenomedullary chromaffin cells.

Relation among Aromatase P450 and Tumoral Growth in Human Prolactinomas.

The pituitary gland is part of hypothalamic-pituitary-gonadal axis, which controls development, reproduction, and aging in humans and animals. In addition, the pituitary gland is regulated mainly by hormones and neurotransmitters released from the hypothalamus and by systemic hormones secreted by target glands. Aromatase P450, the enzyme responsible for the catabolization of aromatizable androgens to estrogens, is expressed in different parts of body, including the pituitary gland. Moreover, aromatase P450 is involved in sexual dimorphism where alteration in the level of aromatase can initiate a number of diseases in both genders. On the other hand, the direct actions of estrogens, mainly estradiol, are well known for stimulating prolactin release. Numerous studies have shown that changes in the levels of estrogens, among other factors, have been implicated in the genesis and development of prolactinoma. The pituitary gland can produce estradiol locally in several types of endocrine cells, and it is possible that aromatase could be responsible for the maintenance of the population of lactotroph cells and the modulation of the action of central or peripheral regulators. Aromatase overexpression due to inappropriate gene regulation has clinical effects such as the pathogenesis of prolactinomas. The present study reports on the synthesis of pituitary aromatase, its regulation by gonadal steroids, and the physiological roles of aromatase on pituitary endocrine cells. The involvement of aromatase in the pathogenesis of pituitary tumors, mainly prolactinomas, through the auto-paracrine production of estradiol is reviewed.

Endothelial immunocytochemical expression of pituitary IL-1ß and its relation to ACTH-positive cells is regulated by corticosterone in the male rat.

Interleukin-1 beta (IL-1ß) is a cytokine linking the neuroendocrine system and metabolic homeostasis. We have previously demonstrated the relevance of IL-1ß for maintaining the pituitary ACTH-producing cells by immuno-blocking its effects in pituitary cultures. However, the morphological characteristics and the intimate relationship of the pituitary cells expressing IL-1ß and ACTH remain unknown. For determining pituitary variations of immunoreactivity for IL-1ß and its relation with ACTH-positive cells under stress situations, we performed an immunohistochemical analysis of the expression of IL-1ß and ACTH in the pituitary gland of adult rats, in the absence or presence of corticosterone, by establishing different groups: untreated, sham-operated, and bilaterally adrenalectomized animals. In the rats subjected to surgery, the glucocorticoid was administered on the same day of the intervention and on the third day post-surgery. Interestingly, it was observed that IL-1ß was located in the pituitary endothelial cells at the hypophyseal portal vessels, regardless of the treatment schedule. When comparing the pituitary immunoreactive surface to IL-1ß expression without corticosterone, adrenalectomized animals displayed a significantly greater area than the sham-operated animals. Corticosterone significantly inhibited the effect of adrenalectomy depending on the time interval it was administered. By in situ hybridization, IL-1ß mRNA expression was also correlated with immnunocytochemical expression of pituitary IL-1ß. Our results demonstrate that IL-1ß is a constitutive element in endothelial portal pituitary vessels and under stress experimental conditions IL-1ß increases its expression and its relation with ACTH-positive cells, suggesting that IL-1ß could participate in an autocrine-paracrine fashion thereby modulating the pituitary population of ACTH-positive cells.

Pituitary Aromatase P450 May Be Involved in Maintenance of the Population of Luteinizing Hormone-Positive Pituitary Cells in Mice.

As aromatase P450 is located in several pituitary cells, testosterone can be transformed into 17ß-estradiol in the gland by the enzyme. The possible role of this transformation in pituitary function remains to be elucidated, but some evidence suggests a physiological and pathophysiological role for pituitary aromatase. To determine its relevance in the modulation of pituitary function, mainly associated with reproduction, luteinizing hormone (LH)-positive cells in the hypophysis of female and male aromatase knockout (ArKO) mice were studied. In all LH-positive cells, significant increases in the cellular (p < 0.01) and nuclear (p < 0.05) areas were found in the ArKO mice compared to the wild-type mice. In the ArKO mice, LH-positive cells were more abundant (p < 0.01); they were characterized by a stronger cytoplasmic reaction and the cells were more polygonal and exhibited more short, thick cytoplasmic prolongations than those in the wild-type mice. Moreover, LH-positive cells showed a greater proliferation rate in the ArKO mice compared to the wild-type mice (p < 0.01). These findings suggest that the local production of estradiol mediated by pituitary aromatase is necessary for the regulation of LH-positive gonadotropic cells, exerting an autoparacrine inhibitory regulation. These results could underlie the higher pituitary aromatase expression observed in male versus female mice. Similar effects were found in ArKO male and female mice, suggesting that in both sexes the effects of estrogens on maintenance of the LH-positive pituitary cell population could be related to the local aromatization of testosterone to estradiol inside the hypophysis. Therefore, aromatase could modulate pituitary LH-positive cells in males through local estradiol synthesis.

Relevance of pituitary aromatase and estradiol on the maintenance of the population of prolactin-positive cells in male mice.

In previous studies we demonstrated the expression of aromatase in pituitary cells. This expression is gender related, and is also associated with the presence of prolactinomas. To ascertain the relevance of aromatase in modulating the populations of prolactin-positive pituitary cells an immunocytochemical and morphometric study of prolactin-positive pituitary cells was carried out using the pituitary glands of adult male and female aromatase-knockout (ArKO) mice. Additionally has been determined if pituitary aromatase is involved in a gender-linked differentiated regulation of the prolactin-producing pituitary cells. Compared to wild-type mice, the knockout animals of both genders showed a significant decrease (p<0.01) in the cellular and nuclear areas of their prolactin cells, as well as in the percentages of the prolactin-positive cells and the proliferating prolactin cells. Our results suggest that estradiol is responsible for the maintenance of the population of prolactin cell in males and, so as not to disturb the endocrine reproductive environment, estradiol is synthesized inside the pituitary by circulating testosterone via means of aromatase P450, which acts in paracrine way. This new role for pituitary aromatase may well explain the previous findings establishing that the pituitary expression of aromatase is higher in males than in females, and the association between the development of prolactinomas and the increased expression of aromatase in tumours.

Dopamine modulates insulin release and is involved in the survival of rat pancreatic beta cells.

The local synthesis of dopamine and its effects on insulin release have been described in isolated islets. Thus, it may be accepted that dopamine exerts an auto-paracrine regulation of insulin secretion from pancreatic beta cells. The aim of the present study is to analyze whether dopamine is a regulator of the proliferation and apoptosis of rat pancreatic beta cells after glucose-stimulated insulin secretion. Glucose stimulated pancreatic islets obtained from male Wistar rats were cultured with 1 or 10 µM dopamine from 1 to 12 h. Insulin secretion was analyzed by RIA. The cellular proliferation rate of pancreatic islets and beta cells was studied with immunocytochemical double labelling for both insulin and PCNA (proliferating cell nuclear antigen), and active caspase-3 was detected to evaluate apoptosis. The secretion of insulin from isolated islets was significantly inhibited (p<0.01), by treatment with 1 and 10 µM dopamine, with no differences between either dose as early as 1 h after treatment. The percentage of insulin-positive cells in the islets decreased significantly (p<0.01) after 1 h of treatment up to 12 h. The proliferation rate of insulin-positive cells in the islets decreased significantly (p<0.01) following treatment with dopamine. Apoptosis in pancreatic islets and beta cells was increased by treatment with 1 and 10 µM dopamine along 12 h. In conclusion, these results suggest that dopamine could modulate the proliferation and apoptosis of pancreatic beta cells and that dopamine may be involved in the maintenance of pancreatic islets.

Local transformations of androgens into estradiol by aromatase P450 is involved in the regulation of prolactin and the proliferation of pituitary prolactin-positive cells.

In previous studies we demonstrated the immunohistochemical expression of aromatase in pituitary cells. In order to determine whether pituitary aromatase is involved in the paracrine regulation of prolactin-producing pituitary cells and the physiological relevance of pituitary aromatase in the control of these cells, an in vivo and in vitro immunocytochemical and morphometric study of prolactin-positive pituitary cells was carried out on the pituitary glands of adult male rats treated with the aromatase antagonist fadrozole. Moreover, we analyzed the expression of mRNA for the enzyme in pituitary cells of male adult rats by in situ hybridization. The aromatase-mRNA was seen to be located in the cytoplasm of 41% of pituitary cells and was well correlated with the immunocytochemical staining. After in vivo treatment with fadrozole, the size (cellular and nuclear areas) of prolactin cells, as well as the percentage of prolactin-positive cells and the percentage of proliferating-prolactin cells, was significantly decreased. Moreover, fadrozole decreased serum prolactin levels. In vitro, treatment with fadrozole plus testosterone induced similar effects on prolactin-positive cells, inhibiting their cellular proliferation. Our results suggest that under physiological conditions aromatase P450 exerts a relevant control over male pituitary prolactin-cells, probably transforming testosterone to estradiol in the pituitary gland.

Differential sensitivity to adrenergic stimulation underlies the sexual dimorphism in the development of diabetes caused by Irs-2 deficiency.

The diabetic phenotype caused by the deletion of insulin receptor substrate-2 (Irs-2) in mice displays a sexual dimorphism. Whereas the majority of male Irs-2(-/-) mice are overtly diabetic by 12 weeks of age, female Irs-2(-/-) animals develop mild obesity and progress less rapidly to diabetes. Here we investigated ß-cell function and lipolysis as potential explanations for the gender-related differences in this model. Glucose-stimulated insulin secretion was enhanced in islets from male null mice as compared to male WT whereas this response in female Irs-2(-/-) islets was identical to that of female controls. The ability of α(2)-adrenoceptor (α(2)-AR) agonists to inhibit insulin secretion was attenuated in male Irs2 null mice. Consistent with this, the expression of the α(2A)-AR was reduced in male Irs-2(-/-) islets. The response of male Irs-2(-/-) islets to forskolin was enhanced, owing to increased production of cAMP. Basal lipolysis was increased in male Irs-2(-/-) but decreased in female Irs-2(-/-) mice, concordant with the observation that adipose tissue is sparse in males whereas female Irs2 null mice are mildly obese. Adipocytes from both male and female Irs-2(-/-) were resistant to the anti-lipolytic effects of insulin but female Irs-2(-/-) fat cells were additionally resistant to the catabolic effects of beta-adrenergic agonists. This catecholamine resistance was associated with impaired generation of cAMP. Consequently, targets of cAMP-dependent protein kinase (PKA) which mediate lipolysis were not phosphorylated in adipose tissue of female Irs-2(-/-) mice. Our findings suggest that IRS-2 deficiency in mice alters the expression and/or sensitivity of components of adrenergic signaling.

The imidazoline I2-site ligands BU 224 and 2-BFI inhibit MAO-A and MAO-B activities, hydrogen peroxide production, and lipolysis in rodent and human adipocytes.

Numerous imidazolinic agents exhibit antihyperglycaemic properties and have been described to promote insulin secretion, however their effects on adipose tissue development have been poorly investigated. Since white adipose tissue (WAT) plays an important role in glucose homeostasis and expresses imidazoline (I(2)) binding sites abundantly, this work aimed at studying extrapancreatic actions of two I(2)-site ligands, BU 224 and 2-BFI in adipocytes. Interaction with monoamine oxidase (MAO) was investigated by measuring the ability to modulate [(14)C]tyramine oxidation and hydrogen peroxide production. Direct influence on glucose uptake or on lipolytic activity was tested on mouse, rat, rabbit and human adipocytes. BU 224 and 2-BFI behaved as reversible inhibitors of both MAO-A and -B, as demonstrated by total inhibition of tyramine oxidation in human adipocytes and platelets or in liver from rats previously treated with selective MAO-inhibitors. Moreover, they weakly inhibited semicarbazide-sensitive amine oxidase. Like classical MAO-inhibitors, they were unable to produce hydrogen peroxide and to activate glucose uptake but prevented tyramine to do so in rodent or human adipocytes. BU 224 and 2-BFI also differed from MAO-inhibitors since they inhibited lipolysis at millimolar concentrations via a still undefined pathway independent of alpha(2)-adrenoceptor stimulation, beta-adrenergic antagonism and MAO activation. However, chronic treatment of obese Zucker rats with 2-BFI did not modify the maximal lipolytic capacity or the mild insulin resistance status of their adipocytes. Taken together, our observations demonstrate on WAT novel effects of BU 224 and 2-BFI different from their already reported actions on brain or endocrine pancreas.

Methylamine but not mafenide mimics insulin-like activity of the semicarbazide-sensitive amine oxidase-substrate benzylamine on glucose tolerance and on human adipocyte metabolism.

It has been reported that benzylamine reduces blood glucose in rabbits, stimulates hexose uptake, and inhibits lipolysis in mouse, rabbit, and human adipocytes. In the presence of vanadate, benzylamine is also able to improve glucose disposal in normoglycaemic and diabetic rats. Such insulin-mimicking properties are the consequence of hydrogen peroxide production during benzylamine oxidation by semicarbazide-sensitive amine oxidase (SSAO). The aim of the study was to determine whether other SSAO-substrates could share such potential antidiabetic properties. Thus, mafenide, a synthetic antimicrobial sulfonamide structurally related to benzylamine, and which has been recently reported to interact with SSAO, was tested in the above mentioned models, in parallel with methylamine, a proposed endogenous SSAO-substrate. All tested amines stimulated glucose uptake and inhibited lipolysis in rat and mouse fat cells. Methylamine and benzylamine, but not mafenide, reduced the hyperglycaemic response during a glucose tolerance test in rabbits while the three amines tested were devoid of insulin-releasing activity under both in vivo and in vitro conditions. In human adipocytes, mafenide did not stimulate glucose transport since it was not a high-affinity substrate for SSAO and generated less hydrogen peroxide than benzylamine or methylamine. Therefore, mafenide could not be considered as an antidiabetic drug despite being oxidized and exhibiting insulin-mimicking effects in rat and mouse adipocytes. By contrast, the endogenous substrate methylamine improved glucose utilization in all in vitro and in vivo models, leading to consider novel SSAO substrates as drugs with potential anti-hyperglycaemic properties.

Benzylamine exhibits insulin-like effects on glucose disposal, glucose transport, and fat cell lipolysis in rabbits and diabetic mice.

Benzylamine, a substrate of semicarbazide-sensitive amine oxidase (SSAO), stimulates glucose transport in rat adipocytes and improves glucose disposal in diabetic rats only in the presence of vanadate. These effects have been described to result from a synergism between the hydrogen peroxide formed during amine oxidation and vanadate, via the generation of pervanadate, a powerful insulin mimicker. However, it has also been reported that benzylamine alone can stimulate glucose uptake and inhibit lipolysis in human fat cells. In this work, we therefore investigated whether benzylamine on its own was able to induce both in vivo and in vitro insulin-like responses in animal models other than rat. In rabbits, the i.v. infusion of 7 micromol/kg benzylamine before a glucose tolerance test resulted in a net reduction of the hyperglycemic response without a change in insulin secretion. Benzylamine also improved glucose tolerance and reduced lipid mobilization in hyperglycemic/obese mice. In vitro, 0.1 mM benzylamine stimulated glucose transport and inhibited lipolysis in mouse and rabbit adipocytes. These effects were blocked by previous treatments with semicarbazide, a SSAO inhibitor. Levels of benzylamine oxidation were more elevated in mouse than in rabbit adipose tissues, whereas the reverse was observed for skeletal muscles. Finally, benzylamine was unable to stimulate insulin secretion by isolated pancreatic islets from both species and SSAO activity was hardly detectable in pancreas. Together, our results bring evidence that benzylamine on its own can improve glucose tolerance in rabbit and mouse, likely by stimulating glucose uptake via amine oxidase activation in insulin-sensitive tissues.