Expression, regulation and function of the serum and glucocorticoid-regulated kinase 1 in pituitary corticotrophs

Serum and glucocorticoid-regulated kinase 1 (Sgk1) is a glucocorticoid early response gene; its function however has mainly been elucidated in the context of mineralocorticoid signaling. Here we investigated the expression and function of Sgk1 in the pituitary gland, which is one of the primary glucocorticoid targets. We found that Sgk1 is highly expressed in the human pituitary gland and co-localises to ACTH. In the AtT-20 murine corticotroph cell line, glucocorticoids up-regulated Sgk1 mRNA and protein levels, whilst decreasing POMC transcription and ACTH release. The dexamethasone-induced increase in Sgk1 protein was abolished by the steroid receptor antagonist RU 486 and reduced by the PI3K inhibitor LY-294002. Sgk1 overexpression increased CREB- and AP-1-dependent transcription, POMC transcription and ACTH secretion, but did not influence intracellular cAMP levels, suggesting a regulation of POMC transcription and ACTH release by Sgk1 via the PKA and MAPK pathways. Sgk1 overexpression and CRH had additive effects on POMC promoter activity, but not on ACTH secretion. Sgk1 knockdown by siRNA decreased POMC promoter activity, demonstrating the significance of Sgk1-dependent signalling for basal POMC expression. In summary, Sgk1 is strongly stimulated by glucocorticoids in pituitary corticotrophs; however it enhances AP-1 and CRE activity downstream of PKA, thereby increasing POMC transcription under basal conditions as well as ACTH secretion following glucocorticoid stimulation. These data represent the first direct evidence that Sgk1 effects are antagonistic to the classical glucocorticoid properties. We suggest that Sgk1 integrates different pathways such as the glucocorticoid pathway, the PI3K pathway and the downstream part of the PKA pathway, and acts as a cell-regulated counter regulatory mechanism aiming to weaken the potentially detrimental glucocorticoid effects. The therapeutical targeting of the Sgk1 pathway might open new possibilities for the treatment of Cushing's disease.Die Serum- und Glukokortikoid-regulierte Kinase 1 (Sgk1) ist ein direktes frühes Zielgen von Glukokortikoiden; ihre Funktion wurde jedoch bislang hauptsächlich im Kontext von Mineralokortikoid-Signalwegen studiert. Wir untersuchten hier die Expression und Funktion von Sgk1 in der Hypophyse, die eines der primären Ziele der Glukokortikoidwirkung darstellt. Wir fanden dass Sgk1 in der humanen Hypophyse hoch exprimiert wird und dort mit ACTH co-lokalisiert. In der murinen corticotrophen Zelllinie AtT-20 wurden Sgk1 mRNA und Protein durch Glukokortikoide up-reguliert, während POMC Transkription und ACTH Ausschüttung gleichzeitig vermindert wurden. Der Dexamethason-stimulierte Anstieg von Sgk1 Protein wurde durch den Steroidrezeptorantagonisten RU 486 vollständig gehemmt und durch den PI3K-Inhibitor LY-29402 leicht vermindert. Sgk1 overexpression bewirkte eine Steigerung der CREB- und AP-1-abhängigen Transkription, der POMC Transkription und der ACTH Ausschüttung aber beeinflusste die intrazellulären cAMP Konzentrationen nicht; das weist auf eine Regulation der POMC Transkription und ACTH Ausschüttung durch Sgk1 über die PKA und MAPK Signalwege hin. Sgk1 overexpression und CRH zeigten additive Effekte in Bezug auf POMC Promoteraktivität, jedoch nicht auf ACTH Ausschüttung. Sgk1 knockdown durch siRNA erniedrigte POMC Promoteraktivität, was die Bedeutung der Sgk1-abhängigen Signalaktivität für die basale POMC Expression aufzeigt. Zusammenfassend wird Sgk1 in hypophysären Corticotrophen durch Glukukortikoide stark stimuliert; es verstärkt jedoch AP-1- und CRE-Aktivität downstream der PKA und steigert dadurch die basale POMC Transkription sowie die Glukokortikoid-stimulierte ACTH Ausschüttung. Diese Daten repräsentieren den ersten direkten Hinweis darauf, dass Sgk1 antagonistisch gegenüber den klassischen Glukokortikoid-Effekten wirkt. Dies könnte bedeuten, dass Sgk1 verschiedene Signalwege, wie zum Beispiel den Glukokortikoid-Signalweg, den PI3K Signalweg und das Endstück des PKA Signalwegs, integriert, und dadurch als ein zellgesteuerter Gegenregulationsmechanismus zur Abschwächung von potentiell schädlichen Glukokortikoidwirkungen agiert. Die therapeutische Beeinflussung des Sgk1 Signalwegs könnte neue Perspektiven für die Behandlung von Morbus Cushing eröffnen.submitted by Marie Helene ReiterAbweichender Titel laut Übersetzung der Verfasserin/des VerfassersZsfassung in dt. SpracheWien, Med. Univ., Diss., 2011OeBB(VLID)171651

Diabetes in the older patient : heterogeneity requires individualisation of therapeutic strategies

Owing to the worldwide increase in life expectancy, the high incidence of diabetes in older individuals and the improved survival of people with diabetes, about one-third of all individuals with diabetes are now older than 65 years. Evidence is accumulating that type 2 diabetes is associated with cognitive impairment, dementia and frailty. Older people with diabetes have significantly more comorbidities, such as myocardial infarction, stroke, peripheral arterial disease and renal impairment, compared with those without diabetes. However, as a consequence of the increased use of multifactorial risk factor intervention, a considerable number of older individuals can now survive for many years without any vascular complications. Given the heterogeneity of older individuals with type 2 diabetes, an individualised approach is warranted, which must take into account the health status, presence or absence of complications, and life expectancy. In doing so, undertreatment of otherwise healthy older individuals and overtreatment of those who are frail may be avoided. Specifically, overtreatment of hyperglycaemia in older patients is potentially harmful; in particular, insulin and sulfonylureas should be avoided or, if necessary, used with caution. Instead, glucose-dependent drugs that do not induce hypoglycaemia are preferable since older patients with diabetes and impaired kidney function are especially vulnerable to this adverse event.(VLID)358448

CPEB Interacts with an Ovary-specific eIF4E and 4E-T in Early Xenopus Oocytes

International audienceCPEB (cytoplasmic polyadenylation element-binding protein) is an important regulator of translation in oocytes and neurons. Although previous studies of CPEB in late Xenopus oocytes involve the eIF4E-binding protein maskin as the key factor for the repression of maternal mRNA, a second mechanism must exist, since maskin is absent earlier in oogenesis. Using co-immunoprecipitation and gel filtration assays, we show that CPEB specifically interacts, via protein/protein interactions, with the RNA helicase Xp54, the RNA-binding proteins P100(Pat1) and RAP55, the eIF4E-binding protein 4E-T, and an eIF4E protein. Remarkably, these CPEB complex proteins have been characterized, in one or more organism, as P-body, maternal, or neuronal granule components. We do not detect interactions with eIF4E1a, the canonical cap-binding factor, eIF4G, or eIF4A or with proteins expressed late in oogenesis, including maskin, PARN, and 4E-BP1. The eIF4E protein was identified as eIF4E1b, a close homolog of eIF4E1a, whose expression is restricted to oocytes and early embryos. Although eIF4E1b possesses all residues required for cap and eIF4G binding, it binds m(7)GTP weakly, and in pull-down assays, rather than binding eIF4G, it binds 4E-T, in a manner independent of the consensus eIF4E-binding site, YSKEELL. Wild type and Y-A mutant 4E-T (which binds eIF4E1b but not eIF4E1a), when tethered to a reporter mRNA, represses its translation in a cap-dependent manner, and injection of eIF4E1b antibody accelerates meiotic maturation. Altogether, our data suggest that CPEB, partnered with several highly conserved RNA-binding partners, inhibits protein synthesis in oocytes using a novel pairing of 4E-T and eIF4E1b

Circulating angiopoietin-2 and soluble Tie-2 in type 2 diabetes mellitus: a cross-sectional study

<p>Abstract</p> <p>Background</p> <p>Type 2 diabetes is associated with increased levels of Angiopoietin-2 (Ang-2) and soluble Tie-2 (sTie-2), but its impact on vascular disease is still unknown. This study aimed to further explore the associations of Ang-2 and sTie-2 with metabolic control and diabetic complications.</p> <p>Methods</p> <p>In a cross-sectional designed study, levels of Ang-2 and sTie-2 as well as their relationships to cardiometabolic parameters were determined in 80 type 2 diabetic subjects (age 65 ± 7 years, female 47.4%).</p> <p>Results</p> <p>After controlling for age and BMI, Ang-2 levels were associated with levels of sTie-2, diastolic blood pressure, plasma insulin, homeostasis model assessment of insulin resistance (HOMA-IR), creatinine, glomerular filtration rate (GFR), and gamma-glutamyl transferase (GGT) (all p < 0.02). Presence of diabetic macrovascular complications, polyneuropathy and insulin therapy were associated with higher Ang-2 levels (p < 0.05). Conversely, sTie-2 levels were associated with glycated hemoglobin (HbA_1c), fasting plasma glucose and insulin, HOMA-IR, triglyceride, and liver function parameters (all p < 0.03). Multiple linear regression analysis showed that Ang-2 remained significantly associated only with levels of GGT (p < 0.04), whereas sTie-2 remained significantly associated with HbA<b>_1c</b>, insulin levels, and HOMA-IR (p < 0.03). No differences in Ang-2 and sTie-2 levels were observed with regard to gender of participants.</p> <p>Conclusions</p> <p>Ang-2 is independently associated with levels of GGT while sTie-2 is independently associated with levels of HbA<b>_1c</b>, plasma insulin and HOMA-IR in type 2 diabetic subjects. Therefore we suggest that the associations of Ang-2 and sTie-2 with type 2 diabetes are based on different patho-physiological mechanisms.</p

Factors predicting long-term comorbidities in patients with Cushings syndrome in remission

Purpose In Cushings syndrome, comorbidities often persist after remission of glucocorticoid excess. Here, we aim to identify factors predicting long-term comorbidities in patients with Cushings syndrome in remission. Methods In a retrospective cross-sectional study, 118 patients with Cushings syndrome in remission (52 pituitary, 58 adrenal, 8 ectopic) were followed for a median of 7.9 years (range 238) after the last surgery. Associations between baseline anthropometric, metabolic, hormonal parameters at diagnosis, and comorbidities (obesity, diabetes, hyperlipidemia, hypertension, osteoporosis, depression) at last follow-up, were tested by uni- and multivariate regression analysis. Results In patients with manifest comorbidities at diagnosis, remission of Cushings syndrome resolved diabetes in 56% of cases, hypertension in 36% of cases, hyperlipidaemia in 23%, and depression in 52% of cases. In a multivariate regression analysis, age, fasting glucose, BMI, and the number of comorbidities at diagnosis were positive predictors of the number of long-term comorbidities, while baseline 24-h urinary free cortisol (UFC) negatively correlated with the persistence of long-term comorbidities. The negative relationship between baseline UFC and long-term comorbidities was also found when pituitary and adrenal Cushings cases were analyzed separately. Baseline UFC was negatively related to the time of exposure to excess glucocorticoids. Conclusions Long-term comorbidities after remission of Cushings syndrome depend not only on the presence of classic cardiovascular risk factors (age, hyperglycemia, BMI), but also on the extent of glucocorticoid excess. Lower baseline UFC is associated with a higher number of long-term comorbidities, possibly due to the longer exposure to excess glucocorticoids in milder Cushings syndrome.(VLID)363261

Characterization of GPR101 transcripts structure and expression patterns

We recently showed that Xq26.3 microduplications cause X-linked acrogigantism (X-LAG). X-LAG patients mainly present with growth hormone and prolactin-secreting adenomas and share a minimal duplicated region containing at least four genes. GPR101 was the only gene highly expressed in their pituitary lesions, but little is known about its expression patterns. GPR101 transcripts were characterized in human tissues by 5’-RACE and RNAseq, while the putative promoter was bioinformatically predicted. We investigated GPR101 mRNA and protein expression by RT-qPCR, whole-mount in situ hybridization, and immunostaining, in human, rhesus monkey, rat, and zebrafish. We identified four GPR101 isoforms characterized by different 5’ untranslated regions (UTRs) and a common 6.1 kb-long 3’UTR. GPR101 expression was very low or absent in almost all adult human tissues examined, except for specific brain regions. Strong GPR101 staining was observed in human fetal pituitary and during adolescence, whereas very weak/absent expression was detected during childhood and adult life. In contrast to humans, adult pituitaries of monkey and rat expressed GPR101, but in different cell types. Gpr101 is expressed in the brain and pituitary during rat and zebrafish development; in rat pituitary Gpr101 is expressed only after birth and showed sexual dimorphism. This study shows that different GPR101 transcripts exist and that the brain is the major site of GPR101 expression across different species, although divergent species- and temporal-specific expression patterns are evident. These findings suggest an important role for GPR101 in brain and pituitary development and likely reflect the very different growth, development and maturation patterns among species