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ABSTRACT: To investigate whether atorvastatin can promote formation of neurites in cultured cortical neurons and the signaling mechanisms responsible for this effect.
Cultured rat cerebral cortical neurons were incubated with atorvastatin (0.05-10 μmol/L) for various lengths of time. For pharmacological experiments, inhibitors were added 30 min prior to addition of atorvastatin. Control cultures received a similar amount of DMSO. Following the treatment period, phase-contrast digital images were taken. Digital images of neurons were analyzed for total neurite branch length (TNBL), neurite number, terminal branch number, and soma area by SPOT Advanced Imaging software. After incubation with atorvastatin for 48 h, the levels of phosphorylated 3-phosphoinoside-dependent protein kinase-1 (PDK1), phospho-Akt, phosphorylated mammalian target of rapamycin (mTOR), phosphorylated 4E-binding protein 1 (4E-BP1), p70S6 kinase (p70S6K), and glycogen synthase kinase-3β (GSK-3β) in the cortical neurons were evaluated using Western blotting analyses.
Atorvastatin (0.05-10 μmol/L) resulted in dose-dependent increase in neurite number and length in these neurons. Pretreatment of the cortical neurons with phosphatidylinositol 3-kinase (PI3K) inhibitors LY294002 (30 μmol/L) and wortmannin (5 μmol/L), Akt inhibitor tricribine (1 μmol/L) or mTOR inhibitor rapamycin (100 nmol/L) blocked the atorvastatin-induced increase in neurite outgrowth, suggesting that atorvastatin promoted neurite outgrowth via activating the PI3K/Akt/mTOR signaling pathway. Atorvastatin (10 μmol/L) significantly increased the levels of phosphorylated PDK1, Akt and mTOR in the cortical neurons, which were prevented by LY294002 (30 μmol/L). Moreover, atorvastatin (10 μmol/L) stimulated the phosphorylation of 4E-BP1 and p70S6K, the substrates of mTOR, in the cortical neurons. In addition, atorvastatin (10 μmol/L) significantly increased the phosphorylated GSK-3β level in the cortical neurons, which was prevented by both LY294002 and tricribine.
These results suggest that activation of both the PI3K/Akt/mTOR and Akt/GSK-3β signaling pathways is responsible for the atorvastatin-induced neurite outgrowth in cultured cortical neurons.
Acta Pharmacologica Sinica 06/2012; 33(7):861-72. · 1.95 Impact Factor
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ABSTRACT: To investigate whether the age-related increase in interleukin-1beta (IL-1beta) and c-Jun N-terminal kinases (JNK) pathway was coupled with a decrease in cell survival signaling pathways and whether sodium ferulate (SF) treatment was effective in preventing these age-associated changes.
Groups of young and aged rats were fed for 4 weeks on a diet enriched in SF (100 mg/kg and 200 mg/kg per day). At the end of the period of dietary manipulation, Western blotting analysis was used to determine the expressions of IL-1beta, phosphorylated mitogen-activated protein kinase kinase (MKK)4, phospho-JNK, phospho-c-Jun, phosphorylated extracellular signal-regulated kinase (ERK1/2), phospho-MEK, phospho-Akt, phosphorylated ribosomal protein S6 protein kinase (p70S6K), and activated caspase-3 and caspase-7. Nissl staining was used to observe the morphological change in hippocampal CA1 regions. Immunohistochemical techniques for glial fibrillary acidic protein (GFAP) and integrin alphaM (OX-42) were used to determine the astrocyte and microglia activation.
IL-1beta protein levels, and phospho-MKK4, phospho-JNK1/2, and phospho-c-Jun were significantly enhanced in hippocampus prepared from age-matched control rats. Increased IL-1beta production and JNK1/2 activation was accompanied by downregulation of MEK/ERK1/2 pathway and Akt/p70S6K pathway, leading to cell apoptosis assessed by activation of caspase-3. Significantly, treatment of aged rats with SF (100 mg/kg and 200 mg/kg per day) for 4 weeks prevented the agerelated increase in IL-1beta and IL-1beta-induced JNK signaling pathway and also the age-related changes in ERK and Akt kinase.
SF plays neuroprotective roles through suppression of IL-1beta and IL-1beta-induced JNK signaling and upregulation of MEK/ERK1/2 and Akt/p70S6K survival pathways.
Acta Pharmacologica Sinica 01/2009; 29(12):1399-408. · 1.95 Impact Factor
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ABSTRACT: Aim: To investigate whether the age-related increase in interleukin-1β (IL-1β) and c-Jun N-terminal kinases (JNK) pathway was coupled with a decrease in cell survival signaling pathways and whether sodium ferulate (SF) treatment was effective in preventing these age-associated changes. Methods: Groups of young and aged rats were fed for 4 weeks on a diet enriched in SF (100 mg/kg and 200 mg/kg per day). At the end of the period of dietary manipulation, Western blotting analysis was used to determine the expressions of IL-1β, phosphorylated mitogen-activated protein kinase kinase (MKK)4, phospho-JNK, phospho-c-Jun, phosphorylated extracellular signal-regulated kinase (ERK1/2), phospho-MEK, phospho-Akt, phosphorylated ribosomal protein S6 protein kinase (p70S6K), and activated caspase-3 and caspase-7. Nissl staining was used to observe the morphological change in hippocampal CA1 regions. Immunohistochemical techniques for glial fibrillary acidic protein (GFAP) and integrin αM (OX-42) were used to determine the astrocyte and microglia activation. Results: IL-1β protein levels, and phospho-MKK4, phospho-JNK1/2, and phospho-c-Jun were significantly enhanced in hippocampus prepared from age-matched control rats. Increased IL-1β production and JNK1/2 activation was accompanied by down-regulation of MEK/ERK1/2 pathway and Akt/p70S6K pathway, leading to cell apoptosis assessed by activation of caspase-3. Significantly, treatment of aged rats with SF (100 mg/kg and 200 mg/kg per day) for 4 weeks prevented the age-related increase in IL-1β and IL-1β-induced JNK signaling pathway and also the age-related changes in ERK and Akt kinase. Conclusion: SF plays neuroprotective roles through suppression of IL-1β and IL-1β-induced JNK signaling and upregulation of MEK/ERK1/2 and Akt/p70S6K survival pathways.
Acta Pharmacologica Sinica 12/2008; 29(12):1399 - 1408. · 1.95 Impact Factor
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ABSTRACT: Aim: To investigate whether the age-related increase in interleukin-1β (IL-1β) and c-Jun N-terminal kinases (JNK) pathway was coupled with a decrease in cell survival signaling pathways and whether sodium ferulate (SF) treatment was effective in preventing these age-associated changes.
Acta Pharmacologica Sinica 11/2008; 29(12):1399-1408. · 1.95 Impact Factor
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ABSTRACT: To investigate whether sodium ferulate (SF) can protect cortical neurons from glutamate-induced neurotoxicity and the mechanisms responsible for this protection.
Cultured cortical neurons were incubated with 50 micromol/L glutamate for either 30 min or 24 h, with or without pre-incubation with SF (100, 200, and 500 micromol/L, respectively). LY294002, wortmannin, PD98059, and U0126 were added respectively to the cells 1 h prior to SF treatment. After incubation with glutamate for 24 h, neuronal apoptosis was quantified by scoring the percentage of cells with apoptotic nuclear morphology after Hoechst 33258 staining. After incubation with glutamate for either 30 min or 24 h, cellular extracts were prepared for Western blotting of active caspase-3, poly (ADP-ribose) polymerase (PARP), mu-calpain, Bcl-2, phospho-Akt, phosphorylated ribosomal protein S6 protein kinase (p70S6K), phospho-mitogen-activated protein kinase kinase (MEK1/2) and phosphorylated extracellular signal-regulated kinase (ERK) 1/2.
SF reduced glutamate-evoked apoptotic morphology, active caspase-3 protein expression, and PARP cleavage and inhibited the glutamate-induced upregulation of the mu-calpain protein level. The inhibition of the phosphatidylinositol 3-kinase (PI3K) and the MEK/ERK1/2 pathways partly abrogated the protective effect of SF against glutamate-induced neuronal apoptosis. SF prevented the glutamate-induced decrease in the activity of the PI3K/Akt/p70S6K and the MEK/ERK1/2 pathways. Moreover, incubation of cortical neurons with SF for 30 min inhibited the reduction of the Bcl-2 expression induced by glutamate.
The results indicate that PI3K/Akt/p70S6K and the MEK/ERK signaling pathways play important roles in the protective effect of SF against glutamate toxicity in cortical neurons.
Acta Pharmacologica Sinica 01/2008; 28(12):1881-90. · 1.95 Impact Factor
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ABSTRACT: To observe the effects of sodium ferulate (SF) on amyloid beta (Abeta)1-40-induced p38 mitogen-activated protein kinase (MAPK) signal transduction pathway and the neuroprotective effects of SF.
Rats were injected intracerebroventricularly with Abeta1-40. Six hours after injection, Western blotting was used to determine the expressions of phosphorylated mitogen-activated protein kinase kinase (MKK) 3/MKK6, phospho-p38 MAPK, interleukin (IL)-1beta, phospho-MAPK activating protein kinase 2 (MAPKAPK-2), the 27 kDa heat shock protein (Hsp27), procaspase-9, -3, and -7 cleavage, and poly (ADP-ribose) polymerase (PARP) cleavage. Seven days after injection, Nissl staining was used to observe the morphological change in hippocampal CA1 regions.
Intracerebroventricular injection of Abeta1-40 induced an increase in phosphorylated MKK3/MKK6 and p38 MAPK expressions in hippocampal tissue. These increases, in combination with enhanced interleukin (IL)-1beta protein expression and reduced phospho-MAPKAPK2 and phospho-Hsp27 expression, mediate the Abeta-induced activation of cell death events as assessed by cleavage of procaspase-9, -3, and -7 and caspase-3 substrate PARP cleavage. Pretreatment with SF (100 mg/kg and 200 mg/kg daily, 3 weeks) significantly prevented Abeta1-40-induced increases in phosphorylated MKK3/MKK6 and p38 MAPK expression. The Abeta1-40-induced increase in IL-1beta protein level was attenuated by pretreatment with SF. In addition, Abeta1-40-induced decreases in phosphorylated MAPKAPK2 and Hsp27 expression were abrogated by administration of SF. In parallel with these findings, Abeta1-40-induced changes in activation of caspase-9, caspase-7, and caspase-3 were inhibited by pretreatment with SF.
SF prevents Abeta1-40-induced neurotoxicity through suppression of MKK3/MKK6-p38 MAPK activity and IL-1beta expression and upregulation of phospho-Hsp27 expression.
Acta Pharmacologica Sinica 11/2006; 27(10):1309-16. · 1.95 Impact Factor
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ABSTRACT: To observe whether an amyloid beta (Abeta)-induced increase in interleukin (IL)-1beta was accompanied by an increase in the p38 mitogen-activated protein kinase (MAPK) pathway and a decrease in the cell survival pathway, and whether sodium ferulate (SF) treatment was effective in preventing these Abeta-induced changes.
Rats were injected intracerebroventricularly with Abeta25-35. Seven days after injection, immunohistochemical techniques for glial fibrillary acidic protein (GFAP) were used to determine the astrocyte infiltration and activation in hippocampal CA1 areas. The expression of IL-1beta, extracellular signal-regulated kinase (ERK), p38 MAPK, Akt/protein kinase B (PKB), Fas ligand and caspase-3 were determined by Western blotting. The caspase-3 activity was measured by cleavage of the caspase-3 substrate (Ac-DEVD-pNA). Reverse transcription-polymerase chain reaction was used to analyze the changes in IL-1beta mRNA levels.
Intracerebroventricular injection of Abeta25-35 elicited astrocyte activation and infiltration and caused a strong inflammatory reaction characterized by increased IL-1beta production and elevated levels of IL-1beta mRNA. Increased IL-1beta synthesis was accompanied by increased activation of p38 MAPK and downregulation of phospho-ERK and phospho-Akt/PKB in hippocampal CA regions prepared from Abeta-treated rats, leading to cell death as assessed by activation of caspase-3. SF significantly prevented Abeta-induced increases in IL-1beta and p38 MAPK activation and also Abeta-induced changes in phospho-ERK and phospho-Akt/PKB expression levels.
SF prevents Abeta-induced neurotoxicity through suppression of p38 MAPK activation and upregulation of phospho-ERK and phospho-Akt/PKB expression.
Acta Pharmacologica Sinica 09/2005; 26(8):943-51. · 1.95 Impact Factor
