Modulation of nuclear steroid receptors by ursodeoxycholic acid inhibits TGF-beta1-induced E2F-1/p53-mediated apoptosis of rat hepatocytes.
We have recently shown that both ursodeoxycholic acid (UDCA) and tauroursodeoxycholic acid (TUDCA) prevent transforming growth factor beta1 (TGF-beta1)-induced hepatocyte apoptosis by modulating the E2F-1/p53/Bax pathway. In addition, activation of glucocorticoid (GR) and mineralocorticoid receptors (MR) inhibits apoptosis in various systems. UDCA induces a ligand-independent activation of the GR, thus potentially regulating a number of targets. In this study, we investigated the role of GR and MR during TGF-beta1-induced hepatocyte apoptosis, and identified additional antiapoptotic targets for UDCA. Our results showed that in primary hepatocytes, TGF-beta1 induced 40-50% decreases in gr and mr mRNA expression (p < 0.01), together with up to 10-fold reductions in their protein levels (p < 0.01). Notably, pretreatment with UDCA resulted in a significant upregulation of nuclear steroid receptors (p < 0.05), which coincided with 2- and 3-fold increases in the level of GR and MR nuclear translocation, respectively, when compared with that of TGF-beta1 alone (p < 0.05). Similarly, TUDCA induced GR and MR nuclear translocations (p < 0.05) and markedly prevented MR protein changes associated with TGF-beta1 (p < 0.05) without affecting GR protein levels. Moreover, when interference RNA was used to inhibit GR and MR, UDCA no longer protected hepatocytes against TGF-beta1-induced apoptosis. In fact, the protective effect of UDCA in TGF-beta1-associated caspase activation decreased from 65 to <10% when GR or MR function was blocked. Finally, the TGF-beta1-induced E2F-1/Mdm-2/p53 apoptotic pathway, normally inhibited by UDCA, was not regulated by the bile acid after GR or MR silencing. These results demonstrate that UDCA protects against apoptosis through an additional pathway that involves nuclear receptors GR and MR as key factors. Further, the E2F-1/Mdm-2/p53 apoptotic pathway appears to be a prime target for UDCA-induced steroid receptor activation.
Solá
Susana
S
Centro de Patogénese Molecular, Faculty of Pharmacy, University of Lisbon, Lisbon 1600-083, Portugal.
Castro
Rui E
RE
Kren
Betsy T
BT
Steer
Clifford J
CJ
Rodrigues
Cecília M P
CM
D021581:Active Transport, Cell Nucleus; D000818:Animals; D017209:Apoptosis; D020169:Caspases / Q000378:metabolism; D018797:Cell Cycle Proteins; D002467:Cell Nucleus / Q000378:metabolism*; D003600:Cytosol / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism*; D003720:Densitometry; D050684:E2F Transcription Factors; D050687:E2F1 Transcription Factor; D004789:Enzyme Activation; D022781:Hepatocytes / Q000378:metabolism*; D015151:Immunoblotting; D008297:Male; D012313:RNA / Q000378:metabolism; D034622:RNA Interference; D012333:RNA, Messenger / Q000378:metabolism; D034741:RNA, Small Interfering / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D011987:Receptors, Steroid / Q000737:chemistry*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013997:Time Factors; D014157:Transcription Factors / Q000378:metabolism*; D014162:Transfection; D016212:Transforming Growth Factor beta / Q000378:metabolism*; D053773:Transforming Growth Factor beta1; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism*; D014580:Ursodeoxycholic Acid / Q000494:pharmacology*
D018797:Cell Cycle Proteins; D004268:DNA-Binding Proteins; D050684:E2F Transcription Factors; D050687:E2F1 Transcription Factor; C493755:E2f1 protein, rat; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D011987:Receptors, Steroid; C508962:Tgfb1 protein, rat; D014157:Transcription Factors; D016212:Transforming Growth Factor beta; D053773:Transforming Growth Factor beta1; D016159:Tumor Suppressor Protein p53; D012313:RNA; D014580:Ursodeoxycholic Acid; D020169:Caspases