ABSTRACT
OBJECTIVE@#Our previous research showed that Naotaifang (a compound traditional Chinese herbal medicine) extract (NTE) has clinically beneficial effects on neurological improvement of patients with acute cerebral ischemia. In this study, we investigated whether NTE protected acute brain injury in rats and whether its effects on ferroptosis could be linked to the dysfunction of glutathione peroxidase 4 (GPX4) and iron metabolism.@*METHODS@#We established an acute brain injury model of middle cerebral artery occlusion (MCAO) in rats, in which we could observe the accumulation of iron in neurons, as detected by Perl's staining. Using assay kits, we measured expression levels of ferroptosis biomarkers, such as iron, glutathione (GSH), reactive oxygen species (ROS) and malonaldehyde (MDA); further the expression levels of transferrin receptor 1 (TFR1), divalent metal transporter 1 (DMT1), solute carrier family 7 member 11 (SLC7A11) and GPX4 were determined using immunohistochemical analysis, real-time quantitative polymerase chain reaction and Western blot assays.@*RESULTS@#We found that treatment with NTE reduced the expression levels of TFR1 and DMT1, reduced ROS, MDA and iron accumulation and reduced neurobehavioral scores, relative to untreated MCAO rats. Treatment with NTE increased the expression levels of SLC7A11, GPX4 and GSH, and the number of Nissl bodies in the MCAO rats.@*CONCLUSION@#Taken together, our data suggest that acute cerebral ischemia induces neuronal ferroptosis and the effects of treating MCAO rats with NTE involved inhibition of ferroptosis through the TFR1/DMT1 and SCL7A11/GPX4 pathways.
ABSTRACT
Objective: To explore the molecular mechanism of the supplemented Naotaifang (sNTF) in the treatment of vascular dementia (VD), and the mRNA expression profiles of hippocampal tissue of VD model rats before and after the intervention of modified sNTF were investigated by microarray analysis. Methods: VD model was established by bilateral common carotid artery ligation. VD rats were treated with sNTF for 30 days. HE staining and Morris water maze were used to evaluate the therapeutic effect of sNTF. The mRNA expression profiles data of VD model rats before and after intervention of sNTF were obtained by Agilent mRNA expression chip. The significantly differentially expressed genes were screened by microarray analysis, and the protein-protein interaction (PPI) network was constructed. The biological processes and signaling pathways in which differentially expressed genes were mainly involved and analyzed by GO and pathway enrichment. Immunohistochemistry and qRT-PCR were used to verify the chip analysis results. Results: HE staining and Morris water maze experiments showed that VD rats showed cerebral ischemia, hippocampal neuron damage, and decreased spatial learning and memory function, but sNTF can partially reverse this trend. 469 differential expression genes were screened by microarray analysis, including 180 up-regulated genes and 289 down-regulated genes. IL6, FGF2, TNF, and IL1b may be the main pharmacodynamic targets of sNTF in the treatment of VD rats, and the results were verified by immunohistochemistry and qRT-PCR. GO and pathway enrichment analysis showed that these genes were closely related to biological processes such as inflammation and apoptosis, and these genes were mainly involved in the regulation of TNF signaling pathway, toll-like receptor signaling pathway and the apoptosis pathway. Conclusion: The results suggested that the therapeutic effect of DSS on AD involves multiple genes and pathways, and and inhibition of hippocampal neuroinflammation may be one of the important mechanisms of its anti-VD.
ABSTRACT
Aim To investigate the effects of Jiawei Naotaifang on cerebral infarction area, pathological changes of brain tissue and estrogen level of focal cere-bral Iischemia in female ovariectomized rats, and cor-relation between estrogen levels and cerebral infarction area. Methods SD rats were randomly divided into sham operation group, ovariectomized group, cerebral ischemia group,model group,and drug groups(estro-gen group, Jiawei Naotaifang high dose group, Jiawei Naotaifang middle dose group, Jiawei Naotaifang low dose group). The rats in the ovariectomized group, model group, drug groups were ovariectomized, elev-enth days after the ovariectomy. The rats in the drug groups were given intragastric administration for three days. The rats in the model group, cerebral ischemia group and drug groups were prepared for cerebral is-chemia models. Neurological function scores were scored 24 hours after the success of the model, serum levels of estrogen were detected, and the brain was stained with 2, 3, 5-triphenyltetrazolium chloride (TTC) and hematoxylin-eosin staining(HE), TTC staining was used to measure the area of cerebral in-farction, and HE staining was used to observe the pathological changes of brain tissues. Results Com-pared with cerebral ischemia group,cerebral infarction area of rats in the model group increased significantly, the estrogen level was lower and the necrosis and py-knosis of cortical and hippocampus cells of rats in the model group were more obvious. Compared with model group,the cerebral infarction area of rats in the drug groups was reduced,the estrogen levels were elevated, especially in Jiawei Naotaifang high dose group and es-trogen group. The cell morphology of rats,in the estro-gen group,Jiawei Naotaifang high dose group and mid-dle dose group, was improved obviously. Cerebral in-farction area was negatively correlated with the level of estrogen. Conclusions The cerebral infarction area of cerebral ischemia in female ovariectomized rat is signif-icantly correlated with the level of estrogen. Jiawei Naotaifang can reduce the damage and alleviate brain injury of cerebral ischemia in female ovariectomized rats,which may be related to the improvement of estro-gen level.
ABSTRACT
Objective To investigate the effect of Jiawei Naotaifang on neuronal apoptosis and the mechanism in ovariectomized rats with cerebral ischemia. Methods Female Sprague-Dawley rats(n=40)were randomly divided into sham group(n=10),model group(n=10),es-trogen group(n=10)and Jiawei Naotaifang group(n=10).The model group,estrogen group and Jiawei Naotai-fang group were ovariectomized.Eleven days after ovariectomy,the estrogen group and Jiawei Naotaifang group were given estrogen and Jiawei Naotaifang respectively intragastrically for three days.14 days after ovariecto-my,the model group,estrogen group and Jiawei Naotaifang group were modeled cerebral ischemia with Langa's method.24 hours after modeling,the apoptosis rate of neurons was detected with TUNEL,and the activation of extracellular signal-regulated kinase 1/2(ERK1/2)and c-Jun N-terminal kinase(p-JNK)in hippocampus were de-tected with Western blotting. Results Compared with the model group, the apoptosis rates decreased in Jiawei Naotaifang group and the estrogen group(P<0.001),with more activation of ERK1/2(P<0.01)and less activation of JNK(P<0.01). Conclusion Jiawei Naotaifang can protect neuron from apoptosis by promoting the activation of ERK1/2 and inhibiting the activation of p-JNK.
ABSTRACT
Through comparative study on Naotaifang containing serum and plasma proteomics (peptide), this article revealed differential proteins (peptides) in the Naotaifang. The characteristics of differential proteins were identified with mass spectrometry. It provides scientific evidences for the pharmacodynamic material basis and Chinese herbal medicine plasma pharmacological method development in the exploration of Naotaifang. A total of 20 healthy adult SD rats were randomly divided into the control group, Naotaifang treatment group according to their weights. Ten rats in each group. Intragastric administration of medication was given for seven consecutive days. Before surgery, rats were fed with water but without food. One hour after the last drug administration, 10% chloral hydrate was injected for intraperitoneal anesthesia. Blood was taken through the common carotid artery. Serum and plasma samples were made after blood was taken from each rat. Serum and plasma samples of five rats were randomly selected from each group. And the two-dimensional electrophoresis (2-DE) technique was used in the comparative study of serum pro-teomics (peptide). The 300 DPI scanning and PDQuest 7.3.0 were used in the analysis. The ESI-MS/MS was used to identify important differences in proteins and screen characteristic serum and plasma protein. The results showed that 20 differential proteins of 5 plasma samples were identified. There were 15 types of proteins expressing up-regulation and 5 types expressing down-regulation. Comparative analysis on the 2-DE gel pictures of Naotaifang containing serum, 19 differential proteins of 5 plasma samples were identified, among which 15 types of proteins express up-regulation and 4 down-regulation. Comparative analysis on the 2-DE gel pictures of Naotaifang containing serum and Naotaifang containing plasma showed that 24 differential proteins of 5 plasma samples were identified, among which 9 types of proteins express up-regulation and 15 down-regulation. The highly expressed proteins were selected to MALDI-TOF-MS between Naotaifang containing serum and Naotaifang containing plasma. There were six successful-ly identified proteins, which were inter-alpha trypsin inhibitor, heavy chain 3, group-specific component, comple-ment factor B, Receptor Complexed with A Heterodimeric Fc, isoform CRA-d, Transferrin. It was concluded that protein with obvious changes in the Naotaifang containing serum and plasma may be related with fibrinolysis and an-ticoagulant. These proteins are involved in angiogenesis, inflammation and other pathological regulations of physiolog-ical processes. They are of great significance in the study of effective target and its signal transduction pathway of Naotaifang.
ABSTRACT
Objective To observe effects of Naotaifang on MMP-9, NF-κB and TIMP-1 after focal cerebral ischemia in rats. Methods The rats were randomly divided into sham operation group, model group, and Naotaifang low- (3 g/kg), medium- (9 g/kg), high- dose (27 g/kg) group. After 3 days of corresponding therapy by intragastric administration once a day, the regional cerebral ischemia model was made by middle cerebral artery occlusion (MCAO) with suture method. Following 3 days, the rats were treated with previous method. On the third day, hippocampal C2 region of ischemic tissue was detected by HE dyeing. And the contents of MMP-9, NF-κB and TIMP1 proteins in hippocampal C2 region were measured by immunohistochemical method. Results The number of normal brain cells in high dose group of Naotaifang was more than that of the model group, and only a few cells appeared nucleus pycnosis. The MMP-9 expression of all dose groups of Naotaifang were significantly decreased than model group (P<0.05). The NF-κB expression of high and medium dose groups of Naotaifang were significantly decreased (P<0.05). The TIMP1 expression of all dose groups of Naotaifang were significantly increased compared with sham operation group (P<0.05). Conclusion The mechanism of Naotaifang protecting blood brain barrier against injury of cerebral ischemia may be involved in ameliorating MMP, NF-κB and increasing TIMP1 expression.
ABSTRACT
Objective:To approach the effect of Naotangfang extracts on expression of vascular endothelial growth factor(VEGF) after focal cerebral infarct in rats. Methods:30 Sprague-Dawley(SD) rats were randomly divided into 6 groups:sham operation group,model group,Naotaifang extract groups(low,middle,and high dosage) ,and Naofukang group. Some indexes were detected such as neurological scores,VEGF mRNA expression by RT-PCR,and the number of VEGF positive cells by immunohistochemical methods. Results:The symptoms in nervous function were improved in the Admin. groups,especially in the Naotaifang middle dosage group. The number of VEGF positive cells and VEGF PCR production in Naotaifng middle dosage group was higher than that in the model group. Conclusion:Naotaifang has neuroprotective effect against focal cerebral ischemic injury by improving the expression of VEGF.