epidermal-growth-factor and Adrenal-Cortex-Neoplasms

Adrenal incidentalomas (AI) are very common and mostly they are non-functioning adenomas (NFA). NFAs are often associated with insulin resistance and metabolic syndrome. Several biomarkers, including certain growth factors, may participatein the pathogenesis ofmetabolic changes in patients with adrenal adenomas.Patients with NFA and age-matched control subjects were enrolled in the study. Data on age, gender, presence of metabolic syndrome or its components were obtained for each subject. Blood samples were obtained and glycemia, insulinemia, lipid profile, and selected growth factor levels were measured. Forty-three patients with NFA and 40 controls were included in the study. Differences were not found in the metabolic syndrome and its components prevalence or in the biochemical profile between patients and the control group. Significant differences were noticed in the levels of IGF1, IGF2, and IGFBP3 (p=0.016, p=0.005, p=0.004, respectively), but there were no differences in VEGF or EGF concentrations. In NFA patients, an association between glycemia and EGF levels was present (p=0.026). No significant correlations between tumor size and insulin or growth factor concentrations were present in AI patients. Significantly higher serum IGF1, IGF2, and IGFBP3 concentrations in NFA patients may support the role of the IGF axis in the pathogenesis of adrenocortical lesions.No correlation between IGFs or IGFBP3 and parameters of glucose or lipid metabolism was found. Present results may support the role of the growth hormone axis rather than hyperinsulinemia and insulin resistance in the pathogenesis of adrenocortical adenomas.

Topics: Adrenal Cortex Neoplasms; Adrenocortical Adenoma; Adult; Aged; Aged, 80 and over; Biomarkers, Tumor; Case-Control Studies; Epidermal Growth Factor; Female; Humans; Insulin-Like Growth Factor Binding Protein 3; Insulin-Like Growth Factor I; Insulin-Like Growth Factor II; Male; Middle Aged; Prognosis

Hypercortisolism is a common endocrine disorder in dogs, caused by a cortisol-secreting adrenocortical tumor (AT) in approximately 15% of cases. In adrenocortical carcinomas of humans, activation of the phosphatidylinositol 3 kinase (PI3K) signaling pathway by insulin-like growth factor (IGF) signaling represents a promising therapeutic target.. To investigate the involvement of PI3K signaling in the pathogenesis of ATs in dogs and to identify pathway components that may hold promise as future therapeutic targets or as prognostic markers.. Analyses were performed on 36 canine cortisol-secreting ATs (11 adenomas and 25 carcinomas) and 15 normal adrenal glands of dogs.. mRNA expression analysis was performed for PI3K target genes, PI3K inhibitor phosphatase and tensin homolog (PTEN), IGFs, IGF receptors, IGF binding proteins and epidermal growth factor receptors. Mutation analysis was performed on genes encoding PTEN and PI3K catalytic subunit (PIK3CA).. Target gene expression indicated PI3K activation in carcinomas, but not in adenomas. No amino acid-changing mutations were detected in PTEN or PIK3CA and no significant alterations in IGF-II or IGFR1 expression were detected. In carcinomas, ERBB2 expression tended to be higher than in normal adrenal glands, and higher expression of inhibitor of differentiation 1 and 2 (ID1 and ID2) was detected in carcinomas with recurrence within 2.5 years after adrenalectomy.. Based on these results, ERBB2 might be a promising therapeutic target in ATs in dogs, whereas ID1 and 2 might be valuable as prognostic markers and therapeutic targets.

Topics: Adenoma; Adrenal Cortex Neoplasms; Animals; Carcinoma; Dog Diseases; Dogs; Epidermal Growth Factor; Female; Gene Expression Regulation, Neoplastic; Hydrocortisone; Male; Phosphatidylinositol 3-Kinase; PTEN Phosphohydrolase; RNA, Messenger; Signal Transduction; Somatomedins

Aldosterone-producing adenomas (APA) are a frequent cause of secondary hypertension characterized by autonomous hypersecretion of aldosterone. However, the molecular mechanisms involved in adrenal tumorigenesis and deregulated aldosterone secretion are currently unknown. To identify putative functional genes, a transcriptional screening was performed on 8 APA and 3 normal adrenals (NA) using oligonucleotide microarrays. Data were next validated on an expanded set of samples by real-time PCR (APA, n=19; NA, n=10). The epidermal growth factor-like teratocarcinoma-derived growth factor-1 (TDGF-1) was upregulated in APA compared with NA (14.7-fold and 21.4-fold by microarray and real-time PCR, respectively). In vitro studies and Western blot analysis using the NCI H295R adrenocortical cell line showed that TDGF-1 increased Akt phosphorylation on Thr308 and Ser473, consistent with activation of phosphatidylinositol 3-kinase/Akt signaling, and also demonstrated a concomitant inactivation of the Akt substrate glycogen synthesis kinase-3beta via Ser9 phosphorylation. Furthermore, TDGF-1 mediated a 3.8+/-0.4-fold increase in aldosterone secretion (n=4) that was specifically blocked by the phosphatidylinositol 3-kinase inhibitors wortmannin (50 nmol/L) and LY294002 (20 micromol/L). Finally, TDGF-1 protected H295R cells from apoptosis induced by staurosporine, causing a decrease in caspase-3 activity, a reduction in the inactivation of poly(ADP-ribose) polymerase, and an inhibition of DNA fragmentation, detected by the TUNEL reaction and fluorescence microscopy that was blocked by LY294002. Taken together, our data suggest that TDGF-1, which is significantly upregulated in APA and mediates aldosterone hypersecretion and deregulated growth in adrenocortical cells in vitro, may represent a key player in the development and pathophysiology of primary aldosteronism.

Topics: Adenoma; Adrenal Cortex Neoplasms; Adrenalectomy; Adult; Aged; Aldosterone; Apoptosis; Biomarkers, Tumor; Biopsy, Needle; Blotting, Western; Caspase 3; Cell Proliferation; Epidermal Growth Factor; Female; GPI-Linked Proteins; Humans; Immunohistochemistry; In Situ Nick-End Labeling; Intercellular Signaling Peptides and Proteins; Male; Membrane Glycoproteins; Middle Aged; Neoplasm Proteins; Oligonucleotide Array Sequence Analysis; Reverse Transcriptase Polymerase Chain Reaction; RNA, Neoplasm; Signal Transduction; Tissue Culture Techniques; Tumor Cells, Cultured; Up-Regulation

We investigated the effects of epidermal growth factor (EGF) and prostaglandins (PG) on the expression of aromatase (CYP19) in human adrenocortical carcinoma cell line NCI-H295R cells. EGF significantly increased aromatase activity and CYP19 gene transcript in NCI-H295R cells. Exon PII was selected from among several tissue-specific exon I regions. Promoter II that abuts on exon PII was activated by EGF. PGE(2) also significantly increased aromatase activity, CYP19 gene transcript, and promoter II activity. The results of experiments using protein kinase (PK) inhibitors suggest that the cAMP-PKA signaling pathway is involved in the up-regulation of aromatase expression by EGF. PGE(2) activated promoter II activity in 4 h, while 12 h was required for its activation by EGF. In addition, PGE(2) was secreted from NCI-H295R cells in response to EGF. Selective agonists for prostaglandin receptors EP(1) and EP(2) significantly increased aromatase activity, which was decreased by the corresponding antagonists. Finally, antagonists for EP(1) and EP(2) inhibited the up-regulation of aromatase expression following EGF. These results suggest that PGE(2) secondarily acts as an autocrine signal in the up-regulation of aromatase expression by EGF in NCI-H295R cells.

Topics: Adrenal Cortex Neoplasms; Alprostadil; Aromatase; Benzylamines; Cell Line, Tumor; Dinoprost; Dinoprostone; Epidermal Growth Factor; Exons; Flavonoids; Humans; Isoquinolines; Promoter Regions, Genetic; Prostaglandins; Prostaglandins A; Prostaglandins E; Protein Kinase Inhibitors; Receptors, Prostaglandin E; Receptors, Prostaglandin E, EP1 Subtype; Receptors, Prostaglandin E, EP2 Subtype; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Stimulation, Chemical; Sulfonamides

We tested the ability of epidermal growth factor (EGF) to regulate a key enzyme in the adrenal synthesis of glucocorticoids: human type II 3beta-hydroxysteroid dehydrogenase/Delta(5)-Delta(4)-isomerase (3 beta HSD). EGF treatment (25 ng/ml) of human adrenocortical carcinoma cells (H295R) resulted in a 5-fold increase in cortisol production and a corresponding 2-fold increase in 3 beta HSD mRNA. Experiments were performed to determine whether EGF is acting through a previously identified signal transducer and activator of transcription 5 (Stat5)-responsive element located from -110 to -118 in the human type II 3 beta HSD promoter. A Stat5 expression construct was cotransfected with a 3 beta HSD-chloramphenol acetyltransferase (CAT) reporter construct comprised of nucleotides -301-->+45 of the human type II 3 beta HSD promoter linked to the CAT reporter gene sequence. The addition of EGF at doses as low as 10 ng/ml resulted in an 11- to 15-fold increase in CAT activity. The introduction of 3-bp point mutations into critical nucleotides in the Stat5 response element obviated the EGF response. Either Stat5a or Stat5b isoforms induced CAT reporter expression upon treatment with EGF. These results demonstrate the ability of EGF to regulate the expression of a critical enzyme (3 beta HSD) in the production of cortisol and suggest a molecular mechanism by which this regulation occurs.

Topics: Adrenal Cortex Neoplasms; Carcinoma; DNA-Binding Proteins; Dose-Response Relationship, Drug; Epidermal Growth Factor; HeLa Cells; Humans; Hydrocortisone; Isoenzymes; Milk Proteins; Multienzyme Complexes; Progesterone Reductase; Response Elements; RNA, Messenger; STAT5 Transcription Factor; Steroid Isomerases; Trans-Activators; Transcription, Genetic; Transduction, Genetic; Tumor Cells, Cultured; Tumor Suppressor Proteins

Epidermal growth factor (EGF) and basic fibroblast growth factor (bFGF) are pluripotent growth factors that stimulate both the proliferation and steroidogenesis of adrenocortical cells. Here we demonstrate that EGF and bFGF specifically induce mRNA of 3beta-hydroxysteroid dehydrogenase type II (3betaHSD II) and suppress that of 17alpha-hydroxylase/lyase P450 (CYP17) in human adrenocortical H295R cells. The induction of 3betaHSD II mRNA did not occur until 6 h after the growth factor treatment and was completely abolished in the presence of a protein synthesis inhibitor, cycloheximide (CHX), suggesting that the induction required de novo protein synthesis. The CYP17 mRNA suppression began at almost the same time as the induction of the 3betaHSD II mRNA. Interestingly, the CYP17 mRNA level was increased by the CHX treatment. Both the 3betaHSD II and CYP17 mRNAs were repressed by treatment with a calmodulin kinase II (CaMK II) inhibitor, KN-93, and were enhanced by a mitogen-activated protein kinase (MAPK) inhibitor, PD98059. The PD98059-mediated induction of the 3betaHSD II mRNA was completely blocked by the CHX treatment. Interestingly, treatment with EGF in the presence of both PD98059 and CHX produced a greater increase in the CYP17 mRNA than did treatment in the presence of PD98059 alone. These results suggest that CHX-sensitive factor(s) and CaMK II- and MAPK-signaling pathways may have important roles in both induction of 3betaHSD II and suppression of CYP17 by EGF or bFGF in H295R cells.

Topics: 3-Hydroxysteroid Dehydrogenases; Adrenal Cortex Neoplasms; Blotting, Northern; Blotting, Southern; Epidermal Growth Factor; Fibroblast Growth Factor 2; Gene Expression; Humans; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Steroid 17-alpha-Hydroxylase; Tumor Cells, Cultured