Triptolide suppresses growth and hormone secretion in murine pituitary corticotroph tumor cells via NF-kappaB signaling pathway.

Triptolide is a principal diterpene triepoxide from the Chinese medical plant Tripterygium wilfordii Hook. f., whose extracts have been utilized in dealing with diverse diseases in traditional Chinese medicine for centuries. Recently, the antitumor effect of triptolide has been found in several pre-clinical neoplasm models, but its effect on pituitary corticotroph adenomas has not been investigated so far. In this study, we are aiming to figure out the antitumor effect of triptolide and address the underlying molecular mechanism in AtT20 murine corticotroph cell line. Our results demonstrated that triptolide inhibited cell viability and colony number of AtT20 cells in a dose- and time-dependent pattern. Triptolide also suppressed proopiomelanocortin (Pomc) mRNA expression and extracellular adrenocorticotropic hormone (ACTH) secretion in AtT20 cells. Flow cytometry prompted that triptolide leaded to G2/M phase arrest, apoptosis program and mitochondrial membrane depolarization in AtT20 cells. Moreover, dose-dependent activation of caspase-3 and decreased Bcl2/Bax proportion were observed after triptolide treatment. By western blot analysis we found that triptolide impeded phosphorylation of NF-κB p65 subunit and extracellular signal-regulated kinase (ERK), along with reduction of cyclin D1, without any impact on other NF-κB related protein expression like total p65, p50, IκB-α, p-IκB-α. Furthermore, the mouse xenograft model revealed the inhibition of tumor growth and hormone secretion after triptolide administration. Altogether this compound might be a potential pharmaceutical choice in managing Cushing's disease.

Tanshinone IIA induces apoptosis via inhibition of Wnt/ß­catenin/MGMT signaling in AtT­20 cells.

A strategy to suppress the expression of the DNA repair enzyme O6­methylguanine­DNA methyltransferase (MGMT) by inhibition of Wnt/ß­catenin signaling may be useful as a novel treatment for pituitary adenoma. Previous studies have reported that Tanshinone IIA (TSA), a major quinone compound isolated from Salvia miltiorrhiza, had antitumor effects. However, whether TSA has antitumor effects against pituitary adenoma and whether the mechanisms are associated with the Wnt/ß­catenin/MGMT pathway remains to be clarified. In the present study, TSA treatment caused apoptosis in AtT­20 cells in a concentration­dependent manner, as demonstrated by cell viability reduction, phophatidylserine externalization detected by Annexin V staining and mitochondrial membrane potential disruption detected by JC­1 staining, which were associated with activation of caspase­3 and DNA fragmentation detected by TUNEL in AtT­20 cells. T­cell factor (TCF)­lymphoid­enhancing factor (LEF) reporter activity was determined by dual luciferase reporter assay and the interaction between ß­catenin and TCF­4 were detected using a co­immunoprecipitation kit. The results indicated TSA treatment increased ß­catenin phosphorylation, inhibited ß­catenin nuclear translocation, reduced ß­catenin/TCF­4 complex formation and TCF­LEF luciferase reporter activity, and subsequently reduced the expression of cyclin D1 and MGMT. Notably, overexpression of MGMT in ß­catenin knock down AtT­20 cells abrogated the TSA­mediated effects in AtT­20 cells. In conclusion, TSA induced apoptosis via inhibition of Wnt/ß­catenin­dependent MGMT expression, which may provide novel insights into the understanding of the mechanism of the antitumor effects of Salvia miltiorrhiza.

Postnatal ablation of POMC neurons induces an obese phenotype characterized by decreased food intake and enhanced anxiety-like behavior.

Proopiomelanocortin (POMC) neurons in the arcuate nucleus of the hypothalamus are central components of systems regulating appetite and energy homeostasis. Here we report on the establishment of a mouse model in which the ribonuclease III ribonuclease Dicer-1 has been specifically deleted from POMC-expressing neurons (POMC(ΔDCR)), leading to postnatal cell death. Mice are born phenotypically normal, at the expected genetic ratio and with normal hypothalamic POMC-mRNA levels. At 6 weeks of age, no POMC neurons/cells could be detected either in the arcuate nucleus or in the pituitary of POMC(ΔDCR) mice. POMC(ΔDCR) develop progressive obesity secondary to decreased energy expenditure but unrelated to food intake, which was surprisingly lower than in control mice. Reduced expression of AgRP and ghrelin receptor in the hypothalamus and reduced uncoupling protein 1 expression in brown adipose tissue can potentially explain the decreased food intake and decreased heat production, respectively, in these mice. Fasting glucose levels were dramatically elevated in POMC(ΔDCR) mice and the glucose tolerance test revealed marked glucose intolerance in these mice. Secondary to corticotrope ablation, basal and stress-induced corticosterone levels were undetectable in POMC(ΔDCR) mice. Despite this lack of activation of the neuroendocrine stress response, POMC(ΔDCR) mice exhibited an anxiogenic phenotype, which was accompanied with elevated levels of hypothalamic corticotropin-releasing factor and arginine-vasopressin transcripts. In conclusion, postnatal ablation of POMC neurons leads to enhanced anxiety and the development of obesity despite decreased food intake and glucocorticoid deficiency.