| Objectives: Alzheimer’s disease(AD) is a degenerative disease of the central nervous system,which is characterized with progressive memory loss, cognitive decline,emotional changes,and major pathological hallmarks include intracellular neurofibrillary tangles(NFTs),extensive loss of neuronal, and extracellular senile β-amyloid(Aβ) plaques accumulation(senile plaques,SP). Plenty of studies indicated that the Aβ plays a very important role in the onset and development of AD,and the toxicity of Aβ is related with the oxygen free radicals.Aβ can not only damage the cell membrane,but also induce the production of oxidative stress. Recent studies have found that oxidative stress is another important pathological feature in addition to the three major pathological features. Oxidative stress is found early in AD patients.Given the crucial rule of oxidative stress in the pathogenesis of AD, the therapy of antioxidant stress have gain much of our attention. Although some trials about the antioxidants have been done,no exact answer is yet available as to whether or not antioxidants are truly protective in AD.Nrf2,as a nuclear transcription factor,is of great significance to the endogenous oxidative stress system.Under physiological conditions,Nrf2 is isloated in the cytoplasm by its repressor Kelch-like ECH-associated protein 1(Keap-1).When the body is under pathological conditions,Nrf2 is separated from Keap-1 and move into the nucleus to associate with other transcription factors,then recognize and bind to the anti-oxidant response element(ARE).Consequently,Nrf2/ARE signal pathway can upregulate the expression of numerous cytoprotective genes that work synergistically to against oxidantive stress.HO-1,induced by Nrf2/ARE signal pathway,has been proved to be protective to the central nervous system.The heme decomposition products,which is catalyzed by HO-1,are powerful scavengers of free radical in vivo. A recent investigation has shown that the expression of Nrf2 is significantly reduced in hippocampal neurons of AD patients. P38 MAPK belongs to the mitogen-activated protein kinase,which is widely present in mammalian cells, can be activated by a wide range of cellular stresses,such as oxygen free radicals, ischemia and hypoxia and other stimuli.Plenty of evidences showed that p38 MAPK activity is improtant for not only normal immune but also stress response such as inflammatory.Studies have indicated that p38 MAPK is activated in rats of AD model. In the present report,it is shown that in transgenic mice lacking of the gene of p38 MAPK,induce the expression of HO-1 protein via the transcription factor Nrf2.Paeonol(Pae) is a monomer component of Chinese medicine,which is extracted from the root bark of Moutan cortex,owns various pharmacologic activities,include anti-angiogenic,antioxidant and antiapoptosis.It is also sedative,vascular protective and analgesic.It has been reported to be neuroprotective in rats model of AD.Studies showed that paeonol can not only ameliorate the decline of learning and memory of rats model of AD,but also reduce the inflammatory response induced by Aβ. This study aims to explore weather paeonol can induce the expression of HO-1via activate Nrf2 /ARE pathway by downregulate the p38 MAPK.In this study,the AD rats model are builded by bilateral hippocampus injection of Aβ1-42,then we observe the behavioral changes,detect the expression of p38 MAPK,Nrf2、HO-1,explore the of pathogenesis AD and the antioxidant properties of paeonol.Methods:According to the random figure chart,sixty-eight male SD rats were assigned to four groups: the Sham group, the Sham+Paeonol(Paeonol 100mg/kg),group the AD group, and AD+Paeonol(Paeonol 100mg/kg)group.Every group had 17 rats.The rats were adaptive feeded for one week prior to the present experiment.The Sham+Paeonol group and the AD+Paeonol group were treated with paeonol once a day for 3 weeks,the Sham group and AD group were treated with saline of equal volume. After 3 weeks,the AD group and the AD+Paeonol group were injected with 10 ug Aβ1-42 into the rats’ bilateral hippocampus,and the Sham group and Sham+Pae group were injected with equal volume of saline.Seven days after modeling,we use morris water maze to test escape latent periods and the numbers of cross platform,and detect the SOD activity and MDA content of the hippocampus of four group rats’.We use HE staining to observe the morphological changes of neurons in the rats’ hippocampal CA1 area.Western Blot and immunostaining were used to detect the expression of p38 MAPK, Nrf2 and HO-1.Results:1 The result of Morris water maze test indicated that the latent period of escape of each group was not same(F=91.36, P<0.05) and neither does the frequency of crossing platform(F=26.30, P<0.05). Compared with the other three group, the latent period of escape in AD group was significantly longer(P <0.05). In the space probe test, the mean number of crossing platform of the Sham、Sham+Pae、AD+Pae group showed significant higher than the AD group(P <0.05).2 HE staining: The pyramidal cells of the hippocampal CA1 area of rats of Sham、Sham+Pae group were regularly arranged, normal morphology, closely packed. Compared with the Sham group, the pyramidal cells of AD model group were obviously less and sparsely arranged. By contrast, in the AD+Pae group, The number, arrangement and morphology of the neurons were all recovered to some extent.3 Immunostaining: Nrf2 was expressed both in cytoplasm and nucleus.Compared to the other three groups, the number of positive cells of Nrf2 in AD group was much less.HO-1 was expressed in cytoplasm, Compared to the other three groups, the number of positive cells of HO-1,in AD group was far less. Pp38 MAPK was expressed in nucleus.Compared to the other three groups, the number of positive cells of pp38 MAPK in AD group was more.4 The SOD activity and MDA contentFour groups have different SOD activity(F = 238.25, P < 0.05) and compared with AD model group,sham group and AD+Pae group had the higher SOD actixity in the rats’ hippocampals(both P < 0.05)Four groups have different MDA content(F = 68.89, P < 0.05) and compared with AD group,sham group and AD+pae group had the lower MDA content in the rats’ hippocampals(both P < 0.05).5 The results of Weston blot:Compared with the Sham group, the expression of pp38 MAPK was significantly higher(P<0.05),the expression of Nrf2、HO-1 was reduced in the AD group(P <0.05).The expression of pp38 MAPK was significantly reduced(P <0.05),and the expression of Nrf2、HO-1 was obviously increased in the Sham+Pae group compared to AD group(P <0.05).Conclusion: 1 The AD model,which was made by the injection of Aβ1-42 into the rat’s bilateral hippocampus, was successful and effective. 2 The reduced activity of SOD and the increased content of MDA in the hippocampus of rats of AD model,indicated the presence of oxidative stress in the brain.3 Paeonol could improve the ability of learning and memory,and reduced the response of oxidative stress in AD rats, indicating that the paeonol was neuroprotective.4 Paeonol could induced the expression of Nrf2 and HO-1 via down-regulated the expression of p38 MAPK.Through the down-regulating of p38 MAPK to activate intrinsic oxidative stress system may be one of the neuroprotive mechanisms of paeonol. |
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