sirolimus and ursodoxicoltaurine

Vascular calcification increases the risk of cardiovascular disease and death in patients with chronic kidney disease (CKD). Increased activity of mammalian target of rapamycin complex 1 (mTORC1) and endoplasmic reticulum (ER) stress-unfolded protein response (UPR) are independently reported to partake in the pathogenesis of vascular calcification in CKD. However, the association between mTORC1 activity and ER stress-UPR remains unknown. We report here that components of the uremic state [activation of the receptor for advanced glycation end products (RAGE) and hyperphosphatemia] potentiate vascular smooth muscle cell (VSMC) calcification by inducing persistent and exaggerated activity of mTORC1. This gives rise to prolonged and excessive ER stress-UPR as well as attenuated levels of sestrin 1 ( Sesn1) and Sesn3 feeding back to inhibit mTORC1 activity. Activating transcription factor 4 arising from the UPR mediates cell death via expression of CCAAT/enhancer-binding protein (c/EBP) homologous protein (CHOP), impairs the generation of pyrophosphate, a potent inhibitor of mineralization, and potentiates VSMC transdifferentiation to the osteochondrocytic phenotype. Short-term treatment of CKD mice with rapamycin, an inhibitor of mTORC1, or tauroursodeoxycholic acid, a bile acid that restores ER homeostasis, normalized mTORC1 activity, molecular markers of UPR, and calcium content of aortas. Collectively, these data highlight that increased and/or protracted mTORC1 activity arising from the uremic state leads to dysregulated ER stress-UPR and VSMC calcification. Manipulation of the mTORC1-ER stress-UPR pathway opens up new therapeutic strategies for the prevention and treatment of vascular calcification in CKD.

Topics: Activating Transcription Factor 4; Animals; Aorta; Aortic Diseases; Cell Death; Cell Proliferation; Cell Transdifferentiation; Disease Models, Animal; Endoplasmic Reticulum Stress; Extracellular Signal-Regulated MAP Kinases; HEK293 Cells; Humans; Mechanistic Target of Rapamycin Complex 1; Mice, Mutant Strains; Muscle, Smooth, Vascular; Osteogenesis; Phosphorylation; Receptor for Advanced Glycation End Products; S100 Proteins; Signal Transduction; Sirolimus; Taurochenodeoxycholic Acid; Unfolded Protein Response; Uremia; Vascular Calcification

Tauroursodeoxycholic acid (TUDCA) is a cytoprotective bile acid frequently prescribed to patients with cholestatic diseases. Several mechanisms of action have been investigated, but the possibility that cyclic adenosine monophosphate responsive element binding protein (CREB), a transcription factor promoting cell survival, mediates TUDCA's protective effects has not been considered. We examined whether TUDCA activates CREB and whether this activation can protect biliary epithelial cells. Cholangiocytes were stressed by exposure to CCI-779, which inhibits signaling though the kinase mTOR (mammalian target of rapamycin), resulting in cell cycle arrest and apoptosis. Incubation of normal rat cholangiocytes (NRC) cells, with TUDCA resulted in phosphorylation of CREB (Western blotting analysis) and activation of CREB transcription activity (luciferase reporter assay). Inhibition of calcium signals and inhibition of protein kinase C prevented the TUDCA-induced activation of CREB. CCI-779 decreased the viability of rat cholangiocytes in a dose-dependent manner (MTT [3-(4,5-dimethythiazol-2-yl)-2,5-diphenyl tetrazolium bromide] assay). TUDCA protected against CCI-779 cytotoxicity. A dominant negative form of CREB was stably transduced in NRC cells (NRC-M1). TUDCA protection was decreased in NRC-M1. While CCI-779 induced apoptosis in NRC cells as determined by caspase 3 activity, TUDCA attenuated CCI-779-induced apoptosis, an effect absent in NRC-M1. Finally, CCI-779 blocked proliferation of both NRC and NRC-M1 (thymidine incorporation) and this was unaffected by TUDCA. In conclusion, TUDCA activates CREB in cholangiocytes, reducing the apoptotic effect of CCI-779. These findings suggest a novel cytoprotective mechanism for this bile acid.

Topics: Animals; Apoptosis; Bile Ducts; Blotting, Western; Cell Line, Tumor; Cell Proliferation; Cell Survival; Cyclic AMP Response Element-Binding Protein; Cytoprotection; Phosphorylation; Protein Kinases; Rats; Sirolimus; Taurochenodeoxycholic Acid; TOR Serine-Threonine Kinases; Transcription, Genetic