Effect of ursolic acid on proliferation and apoptosis of hepatic carcinoma HepG2 cell line and its partial mechanism / 中国现代普通外科进展

Objective:To investigate the effects of ursolic acid on proliferation and apoptosis of hepatic carcinoma cell line HepG2 and its partial mechanism.Methods:The human HepG2 cells were cultured by different concentrations of ursolic acid.The inhibitions of ursolic acid on cell proliferation were determined by using MTT.The effects of ursolic acid on on cell apoptosis and cell cycle were detected by using flow cytometry.The expressions of pERK1/2 and Cyclin D1 proteins after culturing by different concentrations of ursolic acid were tested by using Western blotting.Results:The human hepatoma HepG2 cell proliferations were inhibited by different concentrations of ursolic acid,and the effects showed dose and time-dependent manner(P＜0.05).The cell apoptosis rate for human hepatoma cell line HepG2 was achieved to the maximum under 60 μ mol/L ursolic acid for 72 h,which was (78.723 ± 3.623)％.Ursolic acid could significantly increase the Go/G1 phase cells proportion,and induce HepG2 apoptosis.Ursolic acid could inhibit the expressions of pERK1/2 and Cyclin D1 proteins,and inhibitory effects showed more apparent along with the concentration and time gradually increasing.Conclusion:Ursolic acid could inhibit the proliferation of human hepatic carcinoma HepG2 cell line,block of cells in G0/G1 phase,and promote the cell apoptosis.This might be related with down-regulation of the expressions of pERK1/2 and Cyclin D1 proteins.

Palmitoyl Serotonin Inhibits L-dopa-induced Abnormal Involuntary Movements in the Mouse Parkinson Model

L-3,4-dihydroxyphenylalanine (L-DOPA) is the most common treatment for patients with Parkinson's disease (PD). However, long term use of L-DOPA for PD therapy lead to abnormal involuntary movements (AIMs) known as dyskinesia. Fatty acid amide hydrolase (FAAH) is enriched protein in basal ganglia, and inhibition of the protein reduces dyskinetic behavior of mice. Palmitoyl serotonin (PA-5HT) is a hybrid molecule patterned after arachidonoyl serotonin, antagonist of FAAH. However, the effect of PA-5HT on L-DOPA-induced dyskinesia (LID) in PD have not yet been elucidated. To investigate whether PA-5HT relieve LID in PD and decrease hyperactivation of dopamine D1 receptors, we used the 6-hydroxydopomine (6-OHDA)-lesioned mouse model of PD and treated the L-DOPA (20 mg/kg) for 10 days with PA-5HT (0.3 mg/kg/day). The number of wall contacts with the forelimb in the cylinder test was significantly decreased by 6-OHDA lesion in mice and the pharmacotherapeutic effect of L-DOPA was also revealed in PA-5HT-treated mice. Moreover, in AIMs test, PA-5HT-treated mice showed significant reduction of locomotive, axial, limb, and orofacial AIMs score compared to the vehicle-treated mice. LID-induced hyper-phosphorylation of ERK1/2 and overexpression of FosB/ΔFosB was markedly decreased in 6-OHDA-lesioned striatum of PA-5HT-treated mice, indicating that PA-5HT decreased the dopamine D1 receptor-hyperactivation induced by chronic treatment of L-DOPA in dopamine-denervated striatum. These results suggest that PA-5HT effectively attenuates the development of LID and enhance of ERK1/2 phosphorylation and FosB/ΔFosB expression in the hemi-parkinsonian mouse model. PA-5HT may have beneficial effect on the LID in PD.

Effects of sertraline on the cell viability and expression of tyrosine hydroxylase and phosphorylated ERK1/2 in NGF-induced PC12 cells / 中华行为医学与脑科学杂志

Objective To investigate the effect of sertraline on the viability and the expression of tyrosine hydroxylase (TH) and phosphorylated ERK1/2 in NGF-induced rat pheochromocytoma (PC12) cells.Methods NGF-induced PC12 cells were pretreated or directly treated with different concentrations of sertraline for 24 or 48 hours and the pretreated groups were then subjected to serum withdrawal condition. Then cell viability was determined by the cell counting Kit-8 (CCK-8). The expression of Tyrosine hydroxylase (TH) and pERK1/2in NGF-induced PC12 cells was determined by immunohistochemistry and western blot respectively. Results The viability of NGF-induced PC12 was improved after administration with sertra]ine. After 24h sertraline administration, the cells activity of PC 12 cells at 20μM ( 1.32 ± 0. 11 ) , 10μM ( 1. 17 ± 0.05 ) of direct effect, and 20μM ( 1.15 ±0.11 ) of protect effect increased dramatically as compared with control group. But high dose ( 50μM )sertraline express high toxic effect to PC12 cells. The expression of TH was increased by sertraline 20 μM at both 24h(ratio of TH/β-actin = 1.27 ±0.05) and 48h(ratio of TH/β-actin = 1. 23 ±0.08) compare with control group,and the expression of pERK1/2 also increased dramatically by sertraline 20 μM at both 24h (ratio of (pERK1/2)/β-actin = 1.41±0.05) and 48h( ratio of (pERK1/2)/β-actin = 1.40 ±0.06) compare with control group(P＜0. 01, P ＜ 0. 05). Immunohistochemistry showed similar results. Conclusion These data suggest that the neuroprotective effect of sertraline may play an important role in depression therapy, and this effect might be mediated by TH and pERK1/2 up-regulation.

Expression of Phosphorylated ERK1/2 and cFos Proteins in Vestibular Nuclei by Selective Stimulation of Horizontal Semicircular Canal

BACKGROUND: There is a little information about the effect of selective vestibular stimulation on the expression of activity-dependent metabolic markers in the vestibular nuclei. The purpose of this study was to evaluate effect of afferent excitation of the horizontal semicircular canal on expression of phosphorylated ERK1/2 (pERK1/2) and cFos proteins in the vestibular nuclei. METHODS: The horizontal semicircular canal of Sprague-Dawley rats was selectively stimulated by using the sinusoidal horizontal stimulator with 10-minute duration of stimulation. Conventional immunohistochemical method was used to visualize pERK1/2 or cFos immunoreactive neurons in the vestibular nuclei following rotation. RESULTS: Five minutes after stimulation of the horizontal semicircular canal there was a high expression of pERK1/2 protein in the medial vestibular nucleus among 4 major subnuclei of the central vestibular nuclear complex. On the contrary, immunoreactivity of cFos protein was observed in the medial and inferior vestibular nucleus 2 hours after horizontal sinusoidal rotation. The lateral vestibular nucleus was free from the expression of pERK1/2 and cFos proteins in response to excitation of the horizontal semicircular canal. However, in the vestibular nuclei of unilaterally labyrinthectomized rats expression of pERK and cFos proteins was markedly suppressed in ipsi-lesional side as well as contra-lesional side following stimulation of the horizontal semicircular canal. Furthermore no expression of pERK1/2 and cFos protein in the bilateral vestibular nuclei of bilaterally labyrinthectomized rats was noted after stimulation of the horizontal semicircular canal. CONCLUSIONS: Therefore these results of present study suggest that excitatory afferent signals from the peripheral vestibular receptors are essential for protein translation for pERK1/2 and cFos in response to stimulation of the semicircular canal.

Activation of Vestibular Neurons Projecting to Autonomic Brain Stem Nuclei Following Acute Hypotension in Rats

Extracellular regulated protein kinase1/2 (pERK1/2) is one of the major regulatory factors for transcription of the c-fos oncogene in neurons. The purpose of this study was to evaluate the expression of phosphorylated ERK1/2 within the vestibular nuclei (VN) of rats following acute arterial hypotension. Following the acute arterial hypotension induced by rapid hemorrhage, a significant number of pERK1/2-immunoreactive neurons appeared bilaterally in the caudal aspect of the medial and inferior VN. No labeling of pERK1/2 was observed in the lateral VN. The peak expression of pERK1/2 in these nuclei occurred within 5 min after hemorrhage. However, in bilaterally labyrinthectomized rats, the appearance of pERK1/2-immunoreactive neurons was eliminated in the VN. Western blot confirmed the effect of bilateral labyrinthectomy on pERK1/2 protein expression in the medial vestibular nucleus 5 min after hemorrhage. These results suggest that, following acute hypotension, afferent signals from the peripheral vestibular receptors are required for activation of ERK 1/2 in the VN.

Expressions of the pERK1/2 and the cFos Proteins at an Early Stage of Transient Global Ischemia-reperfusion Injury in the Hippocampus of Rats

PURPOSE: This study was to evaluate temporal changes in the expressions of the phosphorylated extracellular-regulated kinase1/2 (pERK1/2), the phosphorylated MAPK/ERK kinase1/2 (pMEK1/2) and the cFos proteins in the hippocampus of rats following transient global ischemia. METHODS: Transient global ischemia was induced in the forebrains of Sprague-Dawley rats by using a 4-vessel occlusion for 20 min under anesthetic condition. Hematoxyline-eosin staining showed typical microscopic findings that represented neuronal cell death in hippocampal CA1 regions 5 days after transient global ischemia. Four-vessel occlusion-reperfusion produced ischemic injury in major forebrain structures, such as the striatum, the cortex and the hippocampus, in the finding of triphenyltetrazolium chloride (TTC) staining. RESULTS: A high density of pERK1/2 immunoreactivity existed in the pyramidal-cell layers of the CA2-3 regions and in the granular-cell layers of the dentate gyrus 5 min after ischemia. Following ischemia, expression of the pMEK1/2 protein showed temporal changes similar to that of the pERK1/2 protein. A significant expression of the cFos protein was noted in the pyramidal-cell layers of the CA2-3 regions and in the granular-cell layers of the dentate gyrus 2 hours after global ischemia. CONCLUSION: Intracellular signaling cascades of the ERK or the cFos protein take part in early cellular events in the hippocampus of rats in response to ischemic insult.

Immunohistochemical Identification of Phosphorylated Extracellular Signal-Regulated Kinase1/2 in Rat Vestibular Nuclei by Unilateral Labyrinthectomy

This study evaluated the expression of phosphorylated signal-regulated kinase1/2 (pERK1/2), which is one of the main factors regulating transcription of the cfos oncogene in neurons, in the vestibular nuclei of Sprague-Dawley rats following unilateral labyrinthectomy (UL). Surgical UL was performed to eliminate afferent signals from the peripheral vestibular receptors in the inner ear, under a surgical microscope, 2 hours after anesthesia. Significant numbers of pERK1/2 immunoreactive neurons were seen in the superior, medial, and inferior vestibular nuclei. There were more pERK1/2 immunoreactive cells in the vestibular nuclei contralateral than in the vestibular nuclei ipsilateral to the injured labyrinth, which resulted in significant asymmetric expression of pERK1/2 immunoreactive cells. Subsequently, the pERK1/2 immunoreactivity decreased rapidly, disappearing 90 min after labyrinthectomy. No pERK1/2 labeling was seen in the lateral vestibular nucleus. These results suggest that intracellular signal pathways for the activation of extracellular signal-regulated kinase in the vestibular nuclei are involved in lesion-neural plasticity in the vestibular system