Elevated serum concentrations of the vasoactive protein ET-1 occur in the setting of systemic inflammatory response syndrome and contribute to distal organ hypoperfusion and pulmonary hypertension. Thus, understanding the cellular source and transcriptional regulation of systemic inflammatory stress-induced ET-1 expression may reveal therapeutic targets. Using a murine model of LPS-induced septic shock, we demonstrate that the hepatic macrophage is the primary source of elevated circulating ET-1, rather than the endothelium as previously proposed. Using pharmacologic inhibitors, ET-1 promoter luciferase assays, and by silencing and overexpressing NFκB inhibitory protein IκB expression, we demonstrate that LPS-induced ET-1 expression occurs via an NFκB dependent pathway. Finally, the specific role of the cRel/p65 inhibitory protein IκBβ was evaluated. Although cytoplasmic IκBβ inhibits activity of cRel containing NFκB dimers, nuclear IκBβ stabilizes NFκB/DNA binding and enhances gene expression. Using targeted pharmacologic therapies to specifically prevent IκBβ/NFκB signaling, as well as mice genetically modified to overexpress IκBβ, we show that nuclear IκBβ is both necessary and sufficient to drive LPS-induced ET-1 expression. Together, these results mechanistically link the innate immune response mediated by IκBβ/NFκB to ET-1 expression and potentially reveals therapeutic targets for patients with gram-negative septic shock.

Design: Cell viability, ACTH secretion (enzyme-linked immunosorbent assay), apoptosis, and gene expression profile were investigated on AtT-20 cells. In vivo efficacy was examined in an athymic nude mouse AtT-20 xenograft model. SAHA efficacy against human-derived corticotroph tumor (hCtT) (n = 8) was tested in vitro.Setting: National Institutes of Health.Intervention: SAHA (0.5 to 8 mM). Main Outcome Measures:AtT-20 and hCtT cell survival, in vitro/in vivo ACTH measurements.Results: SAHA (1 mM) reduced AtT-20 viability to 75% at 24 hours, 43% at 48 hours (analysis of variance; P = 0.002). Apoptosis was confirmed with elevated BAX/Bcl2 ratio and FACS. Intriguingly, early (3-hour) significant decline (70%; P , 0.0001) of secreted ACTH and diminished POMC transcription was observed with SAHA (1 mM). Microarray analysis revealed a direct association between liver X receptor alpha (LXRa) and POMC expression. Accordingly, SAHA reduced LXRa in AtT-20 cells but not in normal murine corticotrophs. Xenografted nude-mice tumor involution (126 6 33/160 6 35 vs 337 6 49 mm 3 ; P = 0.0005) was observed with 5-day intraperitoneal SAHA, with reversal of elevated ACTH (P , 0.0001). SAHA did not affect serum ACTH in nontumor mice. Lastly, we confirmed that SAHA (1 mM/24 h) decreased hCtT survival (78.92%; P = 0.0007) and ACTH secretion (83.64%; P = 0.03).Conclusion: Our findings demonstrate SAHA's efficacy in reducing survival and ACTH secretion in AtT-20 and hCtT cells, providing a potential intervention for recurrent/unremitting CD. (J Clin

OBJECTIVEVon Hippel-Lindau disease (VHL) is a tumor predisposition syndrome characterized by CNS hemangioblastomas (HBs) and clear cell renal cell carcinomas (RCCs) due to hypoxia-inducible factor activation (pseudohypoxia). Because of the lack of effective medical therapies for VHL, HBs and RCCs account for significant morbidity and mortality, ultimately necessitating numerous neurological and renal surgeries. Propranolol is an FDA-approved pan-beta adrenergic antagonist with antitumor effects against infantile hemangiomas (IHs) and possibly VHL HBs. Here, the authors investigated the antitumor efficacy of propranolol against pseudohypoxia-driven VHL-HBs and VHL-RCCs.METHODSPatient-derived VHL-associated HBs (VHL-HBs) or 786-O-VHL−/− RCC cells were treated with clinically relevant concentrations of propranolol in vitro and assessed with viability assays, flow cytometry, quantitative real-time polymerase chain reaction, and western blotting. In vivo confirmation of propranolol antitumor activity was confirmed in athymic nude mice bearing 786-O xenograft tumors. Lastly, patients enrolled in a VHL natural history study (NCT00005902) were analyzed for incidental propranolol intake. Propranolol activity against VHL-HBs was assessed retrospectively with volumetric HB growth kinetic analysis.RESULTSPropranolol decreased HB and RCC viability in vitro with IC50 (half maximal inhibitory concentration) values of 50 µM and 200 µM, respectively. Similar to prior reports in infantile hemangiomas, propranolol induced apoptosis and paradoxically increased VEGF-A mRNA expression in patient-derived VHL-HBs and 786-O cells. While intracellular VEGF protein levels were not affected by propranolol treatment, propranolol decreased HIF expression in 786-O cells (7.6-fold reduction, p < 0.005). Propranolol attenuated tumor progression compared with control (33% volume reduction at 7 days, p < 0.005) in 786-O xenografted tumor-bearing mice. Three patients (harboring 25 growing CNS HBs) started propranolol therapy during the longitudinal VHL-HB study. HBs in these patients tended to grow slower (median growth rate 27.1 mm3/year vs 13.3 mm3/year) during propranolol treatment (p < 0.0004).CONCLUSIONSPropranolol decreases VHL-HB and VHL-related RCC viability in vitro likely by modulation of VEGF expression and by inducing apoptosis. Propranolol abrogates 786-O xenograft tumor progression in vivo, and retrospective clinical data suggest that propranolol curtails HB growth. These results suggest that propranolol may play a role in the treatment of VHL-related tumors.

Increased and delayed CRH-mediated glucose uptake differentially occurs in adenomatous corticotrophs. Delayed secretagogue-stimulated F-FDG PET could improve microadenoma detection.