Phoenixin 20 promotes neuronal mitochondrial biogenesis via CREB-PGC-1α pathway.

Neurodegenerative disorders are dreadful diseases that affect millions of people worldwide. Mitochondrial dysfunction is closely associated with the development of neurodegenerative disorders. Phoenixin 20 is a newly discovered neuropeptide with a pleiotropic effect. This study showed that the presence of Phoenixin 20 promoted neuronal mitochondrial biogenesis in vitro. In cultured neuronal M17 cells, Phoenixin 20 increased the expression of mitochondrial regulators PGC-1α, NRF-1, and TFAM at both mRNA and protein levels. The treatment of Phoenixin 20 increased the ratio of mitochondrial vs nuclear DNA (mtDNA/nDNA) and the multiple mitochondrial gene expression as revealed by increasing mRNA expression of Tomm22, Timm50, Atp5d, Ndufs3, and protein expression of NDUFB8. At a cellular level, Phoenixin 20 promoted mitochondrial respiratory rate and cellular ATP production. Mechanistically, we found that Phoenixin 20 induced the phosphorylation of CREB, which suggests that Phoenixin 20 promoted the activation of the CREB pathway. The blockage of CREB by its selective inhibitor H89 prevented the effect of Phoenixin 20 on mitochondrial regulators and biogenesis. Moreover, the study showed that Phoenixin 20 induced the expression of its tentative receptor GPR173 at the mRNA and protein level, and the silence of GPR173 in neuronal cells ablated all its effect on mitochondrial regulation. Collectively, we showed that Phoenixin 20 promoted neuronal mitochondrial biogenesis via the regulation of CREB-PGC-1α pathway. This study revealed a new role and underlying mechanism of Phoenixin 20 in neuronal cells, suggesting it influences the therapeutic implication of neurodegenerative diseases.

Renal tubular cell death and inflammation response are regulated by the MAPK-ERK-CREB signaling pathway under hypoxia-reoxygenation injury.

Context: Cell death and inflammation response have been found to the primary features of acute kidney injury.Objective: The aim of our study is to figure out the molecular mechanism by which hypoxia-reoxygenation injury affects the viability of tubular cell death.Materials and methods: HK2 cells were treated with hypoxia-reoxygenation injury in vitro. Pathway agonist was added into the medium of HK2 cell to activate MAPK-EEK-CREB axis.Results: Hypoxia-reoxygenation injury reduced HK2 cell viability and increased cell apoptosis rate in vitro. Besides, inflammation response has been found to be induced by hypoxia-reoxygenation injury in HK2 cells in vitro. In addition, MAPK-ERK-CREB pathway was deactivated during hypoxia-reoxygenation injury. Interestingly, activation of MAPK-ERK-CREB pathway could attenuate hypoxia-reoxygenation injury-mediated HK2 cell apoptosis and inflammation. Mechanistically, MAPK-ERK-CREB pathway activation upregulated the transcription of anti-apoptotic genes and reduced the levels of pro-apoptotic factors under hypoxia-reoxygenation injury.Conclusions: Our results report a novel signaling pathway responsible for acute kidney injury-related tubular cell death. Activation of MAPK-ERK-CREB signaling could protect tubular cell against hypoxia-reoxygenation-related cell apoptosis and inflammation response.

The Three Ds of Transcription Activation by Glucagon: Direct, Delayed, and Dynamic.

Upon lowered blood glucose occurring during fasting, glucagon is secreted from pancreatic islets, exerting various metabolic effects to normalize glucose levels. A considerable portion of these effects is mediated by glucagon-activated transcription factors (TFs) in liver. Glucagon directly activates several TFs via immediate cyclic adenosine monophosphate (cAMP)- and calcium-dependent signaling events. Among these TFs, cAMP response element-binding protein (CREB) is a major factor. CREB recruits histone-modifying enzymes and cooperates with other TFs on the chromatin template to increase the rate of gene transcription. In addition to direct signal transduction, the transcriptional effects of glucagon are also influenced by dynamic TF cross talk. Specifically, assisted loading of one TF by a companion TF leads to increased binding and activity. Lastly, transcriptional regulation by glucagon is also exerted by TF cascades by which a primary TF induces the gene expression of secondary TFs that bring about their activity a few hours after the initial glucagon signal. This mechanism of a delayed response may be instrumental in establishing the temporal organization of the fasting response by which distinct metabolic events separate early from prolonged fasting. In this mini-review, we summarize recent advances and critical discoveries in glucagon-dependent gene regulation with a focus on direct TF activation, dynamic TF cross talk, and TF cascades.

Limonin, a Component of Dictamni Radicis Cortex, Inhibits Eugenol-Induced Calcium and cAMP Levels and PKA/CREB Signaling Pathway in Non-Neuronal 3T3-L1 Cells.

Limonin, one of the major components in dictamni radicis cortex (DRC), has been shown to play various biological roles in cancer, inflammation, and obesity in many different cell types and tissues. Recently, the odorant-induced signal transduction pathway (OST) has gained attention not only because of its function in the perception of smell but also because of its numerous physiological functions in non-neuronal cells. However, little is known about the effects of limonin and DRC on the OST pathway in non-neuronal cells. We investigated odorant-stimulated increases in Ca(2+) and cAMP, major second messengers in the OST pathway, in non-neuronal 3T3-L1 cells pretreated with limonin and ethanol extracts of DRC. Limonin and the extracts significantly decreased eugenol-induced Ca(2+) and cAMP levels and upregulated phosphorylation of CREB and PKA. Our results demonstrated that limonin and DRC extract inhibit the OST pathway in non-neuronal cells by modulating Ca(2+) and cAMP levels and phosphorylation of CREB.

Beneficial Effect of Flavone Derivatives on Aß-Induced Memory Deficit Is Mediated by Peroxisome Proliferator-Activated Receptor γ Coactivator 1α: A Comparative Study.

In the present study, the neuroprotective effect of 5-hydroxy-6,7,4'-trimethoxyflavone (flavone 1), a natural flavone, was investigated in comparison with another flavone, 5,7,4'-trihydroxyflavone (flavone 2) on the hippocampus of amyloid beta (Aß)-injected rats. Rats were treated with the 2 flavones (1 mg/kg/d) for 1 week before Aß injection. Seven days after Aß administration, memory function of rats was assessed in a passive avoidance test (PAT). Changes in the levels of mitochondrial transcription factor A (TFAM), peroxisome proliferator-activated receptor γ coactivator 1 α (PGC-1α), phospho-adenosine monophosphate (AMP)-activated protein kinase (pAMPK), AMPK, phospho-cAMP-responsive element-binding protein (CREB), CREB, and nuclear respiratory factor 1 (NRF-1) proteins were determined by Western blot analysis. Our results showed an improvement in memory in rats pretreated with flavonoids. At the molecular level, phosphorylation of CREB, known as the master modulator of memory processes, increased. On the other hand, the level of mitochondrial biogenesis factors, PGC-1α and its downstream molecules NRF-1 and TFAM significantly increased by dietary administration of 2 flavones. In addition, flavone 1 and flavone 2 prevented mitochondrial swelling and mitochondrial membrane potential reduction. Our results provided evidence that flavone 1 is more effective than flavone 2 presumably due to its O-methylated groups. In conclusion, it seems that in addition to classical antioxidant effect, flavones exert part of their protective effects through mitochondrial biogenesis.

Activation of hepatic CREBH and Insig signaling in the anti-hypertriglyceridemic mechanism of R-α-lipoic acid.

The activation of sterol regulatory element binding proteins (SREBPs) is regulated by insulin-induced genes 1 and 2 (Insig-1 and Insig-2) and SCAP. We previously reported that feeding R-α-lipoic acid (LA) to Zucker diabetic fatty (ZDF) rats improves severe hypertriglyceridemia. In this study, we investigated the role of cyclic AMP-responsive element binding protein H (CREBH) in the lipid-lowering mechanism of LA and its involvement in the SREBP-1c and Insig pathway. Incubation of McA cells with LA (0.2 mM) or glucose (6 mM) stimulated activation of CREBH. LA treatment further induced mRNA expression of Insig-1 and Insig-2a, but not Insig-2b, in glucose-treated cells. In vivo, feeding LA to obesity-induced hyperlipidemic ZDF rats activated hepatic CREBH and stimulated transcription and translation of Insig-1 and Insig-2a. Activation of CREBH and Insigs induced by LA suppressed processing of SREBP-1c precursor into nuclear SREBP-1c, which subsequently inhibited expression of genes involved in fatty acid synthesis, including FASN, ACC and SCD-1, and reduced triglyceride (TG) contents in both glucose-treated cells and ZDF rat livers. Additionally, LA treatment also decreased abundances of very low density lipoprotein (VLDL)-associated apolipoproteins, apoB100 and apoE, in glucose-treated cells and livers of ZDF rats, leading to decreased secretion of VLDL and improvement of hypertriglyceridemia. This study unveils a novel molecular mechanism whereby LA lowers TG via activation of hepatic CREBH and increased expression of Insig-1 and Insig-2a to inhibit de novo lipogenesis and VLDL secretion. These findings provide novel insight into the therapeutic potential of LA as an anti-hypertriglyceridemia dietary molecule.

Activation of Hippocampal CREB by Rolipram Partially Recovers Balance Between TNF-α and IL-10 Levels and Improves Cognitive Deficits in Diabetic Rats.

Diabetes damages the central nervous system, inducing cognitive dysfunction and structural changes, known as diabetic encephalopathy (DE). Some research suggests that the pathogenesis of DE may involve an inflammatory imbalance in the nervous system, along with ß-amyloid deposition, similar to Alzheimer's disease. Less data have been yet provided to prove that mechanism. The aim of this study was to evaluate the influence of diabetes on the expression of TNF-α, IL-10, and cAMP response element-binding protein (CREB)/phosphorylated CREB (pCREB). Moreover, we investigated whether rolipram can improve memory, suppress the inflammatory response, and improve balance of CREB/pCREB in the hippocampus of diabetic rats. We used a 4-week high-fat diet and a low dose of streptozocin (30 mg/kg) to induce diabetes with hyperinsulinemia and hyperglycemia. Cognitive impairment was induced over a period of 4 months, and rolipram treatment was concomitantly given. Cognitive impairment was evaluated with the Morris water maze test. We also assessed expression of the pro-inflammatory cytokine TNF-α and the anti-inflammatory cytokine IL-10. We found that memory in rats with long-term diabetes was impaired. Treatment with rolipram increased expression of CREB and pCREB, reduced the inflammatory reaction (decreased TNF-α levels and increased IL-10 levels), and prevented cognitive impairment in these diabetic animals. This present study suggests that rolipram improves cognitive function by activating the CREB signaling pathway and alleviating neuroinflammation in type 2 diabetic rats. Rolipram may have therapeutic potential in DE.

Inhibition of phosphatidylinositol 3-kinase/Akt signaling attenuates hypoxia-induced pulmonary artery remodeling and suppresses CREB depletion in arterial smooth muscle cells.

Hypoxia-induced pulmonary hypertension is characterized by progressive remodeling of the pulmonary artery (PA) system and loss of the transcription factor, cAMP response element binding protein (CREB) in PA smooth muscle cells (SMCs). Previous in vitro studies suggested that platelet-derived growth factor, a mitogen produced in the hypoxic arterial wall, elicits loss of CREB in medial SMCs via the PI3K/Akt pathway. These events trigger switching of SMCs from a quiescent, contractile phenotype to a proliferative, migratory, dedifferentiated, and synthetic phenotype, which contributes to PA thickening. Here, we investigated whether inhibition of PI3K or Akt could attenuate arterial remodeling in the lung and prevent CREB loss in PA medial SMCs in rats subjected to chronic hypoxia. Inhibition of either enzyme-blunted hypoxia-induced PA remodeling and SMC CREB depletion and diminished SMC proliferation and collagen deposition. Inhibition of Akt, but not PI3K, suppressed muscularization of distal arterioles and blunted right ventricular hypertrophy. Interestingly, mean PA pressure was elevated equally by hypoxia in untreated and inhibitor-treated groups but was normalized acutely by the Rho kinase inhibitor, Fasudil. We conclude that PI3K and Akt inhibitors can attenuate hypoxia-induced PA remodeling and SMC CREB depletion but fail to block the development of pulmonary hypertension because of their inability to repress Rho kinase-mediated vasoconstriction.

Doxorubicin blocks proliferation of cancer cells through proteolytic activation of CREB3L1.

Doxorubicin is used extensively for chemotherapy of diverse types of cancer, yet the mechanism through which it inhibits proliferation of cancer cells remains unclear. Here we report that doxorubicin stimulates de novo synthesis of ceramide, which in turn activates CREB3L1, a transcription factor synthesized as a membrane-bound precursor. Doxorubicin stimulates proteolytic cleavage of CREB3L1 by Site-1 Protease and Site-2 Protease, allowing the NH(2)-terminal domain of CREB3L1 to enter the nucleus where it activates transcription of genes encoding inhibitors of the cell cycle, including p21. Knockdown of CREB3L1 mRNA in human hepatoma Huh7 cells and immortalized human fibroblast SV589 cells conferred increased resistance to doxorubicin, whereas overexpression of CREB3L1 in human breast cancer MCF-7 cells markedly enhanced the sensitivity of these cells to doxorubicin. These results suggest that measurement of CREB3L1 expression may be a useful biomarker in identifying cancer cells sensitive to doxorubicin.DOI:http://dx.doi.org/10.7554/eLife.00090.001.

Imipramine induces brain-derived neurotrophic factor mRNA expression in cultured astrocytes.

Depression is one of the most prevalent and livelihood-threatening forms of mental illnesses and the neural circuitry underlying depression remains incompletely understood. Recent studies suggest that the neuronal plasticity involved with brain-derived neurotrophic factor (BDNF) plays an important role in the recovery from depression. Some antidepressants are reported to induce BDNF expression in vivo; however, the mechanisms have been considered solely in neurons and not fully elucidated. In the present study, we evaluated the effects of imipramine, a classic tricyclic antidepressant drug, on BDNF expression in cultured rat brain astrocytes. Imipramine dose-dependently increased BDNF mRNA expression in astrocytes. The imipramine-induced BDNF increase was suppressed with inhibitors for protein kinase A (PKA) or MEK/ERK. Moreover, imipramine exposure activated transcription factor cAMP response element binding protein (CREB) in a dose-dependent manner. These results suggested that imipramine induced BDNF expression through CREB activation via PKA and/or ERK pathways. Imipramine treatment in depression might exert antidepressant action through BDNF production from astrocytes, and glial BDNF expression might be a target of developing novel antidepressants.

17 Beta-estradiol prevents focal cerebral ischemic damages via activation of Akt and CREB in association with reduced PTEN phosphorylation in rats.

This study aimed to assess the signaling pathway of the neuroprotective action of estrogen in the cerebral ischemic injury evoked by subjecting rats to 2-h occlusion of the middle cerebral artery (MCA) followed by 24-h reperfusion. Rats received 17 beta-estradiol (1, 4 and 10 mg/kg, i.p.) 24 h before and 5 min after the completion of 2-h MCA occlusion. The cerebral infarct area was consistently observed in the cortex and striatum of the left hemisphere. Increased terminal deoxynucleotidyl transferase-mediated deoxyuridine-biotin nick-end labeling (TUNEL)-positive cells and DNA fragmentation in the penumbral zone were significantly reduced by 17 beta-estradiol. In line with these results, 17 beta-estradiol significantly increased Akt and cyclic AMP response element binding protein (CREB) with increased Bcl-2 protein in the ischemic area, whereas the elevated the phosphatase and tensin homolog deleted from chromosome10 (PTEN) phosphorylation was significantly reduced with decreased Bax protein and cytochrome c release. Inhibition of DNA fragmentation, PTEN phosphorylation, and Akt activation by 17 beta-estradiol were antagonized by iberiotoxin, a maxi-K channel blocker. Taken together, it is suggested that suppression of cerebral ischemic injury by 17 beta-estradiol may be ascribed to the maxi-K channel opening-coupled downregulation of PTEN phosphorylation and upregulation of Akt and CREB phosphorylation with resultant increase in Bcl-2 protein and decrease in Bax protein and cytochrome c release.