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Ischemia and hypoxia cause functional damage to brain tissues during stroke， and when blood supply is restored to brain tissues after ischemia， a large number of free radicals and calcium overload cause cerebral ischemia-reperfusion injury， which further aggravates the condition. Autophagy is a self-protection mechanism that maintains the homeostasis of the intracellular environment， but excessive autophagy causes brain tissue damage. MiRNA is a small endogenous non-coding RNA molecule that regulate various physiological activities at the gene level by binding to complementary sequences in the 3 '- UTR of its target gene mRNA， leading to translation inhibition or mRNA degradation. MiRNA not only directly acts on autophagy related proteins， but also participates in autophagy regulation induced by ischemia/reperfusion through various signaling pathways. However， there is still a lack of systematic induction and analysis of miRNA regulation of autophagy signaling pathways induced by cerebral ischemia/reperfusion. This article reviews the regulation of cellular autophagy during cerebral ischemia/ reperfusion by miRNA-124， miRNA-298， miRNA-202-5p， miRNA-142， miRNA-26b and so on through different signaling pathways， providing a systematic and theoretical approach for the study of autophagy in stroke.
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Cerebral ischemia/reperfusion injury （CIRI） is a complex cascade reaction process in which the blood flow and oxygen supply of brain tissue in the infarcted area recover after cerebral ischemia， resulting in secondary injury of ischemic brain tissue. At present， thrombolysis as soon as possible and restoration of cerebral blood supply are still the only strategies for the treatment of stroke， but a considerable number of patients' symptoms will be more serious after reperfusion， making patients face adverse outcomes such as neurological function injury and even death and seriously affecting the quality of life and safety of patients. Therefore， an in-depth exploration of the mechanism and treatment strategy of CIRI has important clinical significance. The phosphatidylinositol 3- kinase （PI3K）/protein kinase B （Akt） signaling pathway is one of the classic anti-apoptosis/reproductive-promoting signal transduction pathways， which is responsible for cell proliferation， growth， and differentiation. It is the key cascade signaling pathway of CIRI， located at the core site in many mechanisms such as mitochondrial disorder， apoptosis， autophagy， oxidative stress， and inflammation. It is closely related to the occurrence and development of CIRI. Traditional Chinese medicine has been used in the clinical treatment of stroke and its complications for thousands of years， and the clinical effect of traditional Chinese medicine in the prevention and treatment of CIRI has been affirmed by a large number of research results in recent years. It is further clarified that the monomers， active components， and their compound prescriptions of traditional Chinese medicine can directly or indirectly regulate the PI3K/Akt signaling pathway by virtue of the biological advantages of multi-targets， multi-components， and multi-pathways and play an overall protective role in CIRI. By analyzing the related research progress of traditional Chinese medicine in China and abroad in recent years， the authors summarized the role and mechanism of regulating the PI3K/Akt signaling pathway in the prevention and treatment of CIRI， so as to provide further theoretical basis for the study of the mechanism of clinical prevention and treatment of CIRI.
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Objective To investigate the potential effect and mechanism of curcumin in inhibiting synaptic injury in the cortex of rats with cerebral ischemia-reperfusion. Methods Sprague-Dawley rats were divided into sham-operated group, model group, low-dose curcumin (50 mg/kg) group, and high-dose curcumin (100 mg/kg) group. A model of middle cerebral artery occlusion for 2 hours and reperfusion for 24 hours was constructed, and curcumin was administered. Based on the neurological function score, the effects of curcumin on cerebral infarct volume, synaptic ultrastructure changes, inflammatory cell infiltration, and the expression of NLRP3, Caspase-1, Synapsin1, and CAMKⅡ were observed after the end of the animal treatment. Results The neurological function scores were 0, 3.25±0.43, 2.50±0.50, and 1.50±0.50 for the sham-operated group, model group, low-dose curcumin group, and high-dose curcumin group, respectively. The percentage of cerebral infarct volume was 0, (38.89±2.21)%, (33.48±1.77)%, and (23.69±2.19)%, respectively. Compared with the sham operation group, the model group had severe synaptic ultrastructure damage, extensive inflammatory cell infiltration, significantly increased expression of Caspase-1 and NLRP3 (P < 0.5), and significantly decreased expression of Synapsin1 and CAMKⅡ (P < 0.5). Curcumin treatment significantly inhibited synaptic damage, reduced inflammatory cell infiltration, decreased the expression of Caspase-1 and NLRP3 (P < 0.5), and increased the expression of Synapsin1 and CAMKII (P < 0.5), when compared with the model group. Conclusion Ischemia-reperfusion-mediated synaptic injury in rat brain triggers an inflammatory response in cortical nerve cells, and curcumin alleviates synaptic damage and reduces brain injury by inhibiting inflammatory factor levels.
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Aim To investigate the regulatory effect of geraniol on Nrf2/HO-1 signaling pathway after cerebral ischemia-reperfusion(I/R)in rats. Methods In this experiment,all the male SD rats were randomly divided into nine groups receiving the following treatments:sham operation(sham); sham operation+200 mg·kg-1 geraniol; I/R; I/R+50 mg·kg-1 geraniol; I/R+100 mg/kg geraniol; I/R+200 mg·kg-1 geraniol; edaravone; I/R+ brusatol(Nrf2 inhibitor); I/R+200mg·kg-1 geraniol+brusatol. All rats received intraperitoneal injection of geraniol for five consecutive days before MCAO and again after MCAO. During the construction of cerebral I/R injury models,the blood vessels were isolated without any suture in the sham operation and the sham operation +200 mg·kg-1 geraniol groups while the blood vessels with suture in other groups. The damage of neurological function was evaluated by the modified rating scale for neurological function. The TTC,HE and Tunel staining methods were used to determine the cerebral infarction volume,the damage of the ischemic cortex and the apoptosis of cortical cells,respectively. The oxidative stress-related parameters then were measured. The protein expressions of Nrf2 and HO-1 were detected by immunohistochemical staining and the target protein expressions of the injured cortex were detected by Western blot. Results Compared with the model group,it was found that the geraniol treatment significantly repaired neural injury,reduced cerebral infarction volume,cerebral cortex damage and cell apoptosis. Meanwhile,geraniol intervention could significantly increase the expression of Nrf2/HO-1 protein in the right-sided cortex and reduce oxidative stress level. Conclusion Geraniol can attenuate cerebral injury induced by ischemia-reperfusion in rats via activating Nrf2/HO-1 signaling pathway.
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OBJECTIVE To explore the protective effect and mechanism of Longshengzhi capsules on cerebral ischemia- reperfusion injury in rats. METHODS The model of middle cerebral artery occlusion （MCAO） was established by using the improved thread occlusion method. The experiment was divided into six groups： sham surgery group （only separating blood vessels without inserting thread plugs， given the same volume of normal saline）， model group （modeling， given the same volume of normal saline）， nimodipine group （positive control， modeling， dose of 20 mg/kg）， and low-dose， medium-dose， and high-dose groups of Longshengzhi capsules （modeling， doses of 0.72， 1.44 and 2.88 g/kg， respectively）， with 10 mice in each group. Each group was given corresponding medication solution/normal saline by gavage， once a day， for 7 consecutive days. One hour after the last administration， the Zea Longa scoring method was used to score the neurological deficits in each group of rats， and the ABC enzyme-linked immunosorbent assay was used to detect the serum levels of tumor necrosis factor-α （TNF-α） and interleukin-6 （IL-6） in rats； TTC staining was used to observe the volume of cerebral infarction in rats and calculate the cerebral infarction volume ratio. Hematoxylin eosin staining was used to observe the pathological changes in the brain tissue of rats. Immunohistochemical staining was used to detect the positive expression of NLRP3 protein in the brain tissue of rats. Real-time fluorescence quantitative PCR was used to detect mRNA relative expressions of Toll-like receptor 4 （TLR4） and nuclear factor-κB （NF-κB） in the brain tissue of rats. Western blot assay was adopted to detect the relative expressions of TLR4， NLRP3 and phosphorylated NF-κB （p-NF-κB） protein in the brain tissue of rats and its intracellular NF-κB protein. RESULTS Compared with the sham surgery group， the neural dysfunction score， serum levels of TNF-α and IL-6， cerebral infarction volume ratio， relative expression levels of NF-κB and TLR4 mRNA， as well as protein relative expressions of TLR4， NLRP3 and p-NF-κB in the brain tissue， and relative protein expression of intracellular NF-κB were increased significantly in the model group （P＜0.01）； the enlarged gap and significant edema were observed in cortical nerve cells of brain tissue in rats， with a large amount of inflammatory cell infiltration； the positive expression of NLRP3 protein in brain tissue of rats obviously increased. Compared with the model group， the levels of the above indicators in the medium-dose and high-dose groups of Longshengzhi capsules， as well as the Nimodipine group， were reversed to varying degrees， and most differences were statistically significant （P＜0.05 or P＜0.01）； the pathological morphology observation showed a significant improvement， and the positive expression of NLRP3 protein in the brain tissue of rats was obviously reduced. CONCLUSIONS Longshengzhi capsules may inhibit TLR4/NF-κB/NLRP3 signaling pathway and neuroinflammatory response， thereby achieving a protective effect against cerebral ischemia-reperfusion injury in rats.
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This study aims to investigate the mechanism of acacetin in protecting rats from cerebral ischemia-reperfusion injury via the Toll-like receptor 4(TLR4)/NOD-like receptor protein 3(NLRP3) signaling pathway. Wistar rats were randomized into sham, model, low-and high-dose acacetin, and nimodipine groups, with 10 rats in each group. The rat model of middle cerebral artery occlusion(MCAO) was established with the improved suture method in other groups except the sham group. The neurological deficit score and cerebral infarction volume of each group were evaluated 24 h after modeling. Enzyme-linked immunosorbent assay(ELISA) was employed to measure the levels of interleukin-1β(IL-1β), IL-6, tumor necrosis factor-α(TNF-α), malondialdehyde(MDA), supe-roxide dismutase(SOD), and glutathione(GSH). Western blot was employed to determine the expression levels of B-cell lymphonoma-2(Bcl-2), Bcl-2-associated X protein(Bax), and TLR4/NLRP3 signaling pathway-related proteins(TLR4, p-NF-κB/NF-κB, NLRP3, pro-caspase-1, cleaved caspase-1, pro-IL-1β, and cleaved IL-1β) in the rat brain tissue. Hematoxylin-eosin(HE) staining was employed to reveal the histopathological changes in the ischemic area. Compared with the sham group, the modeling of MCAO increased the neurological deficit score and cerebral infarction volume, elevated the IL-1β, IL-6, TNF-α, and MDA levels and lowered the SOD and GSH levels in the brain tissue(P<0.05). Compared with the MCAO model group, low-and high-dose acacetin and nimodipine decreased the neurological deficit score and cerebral infarction volume, lowered the IL-1β, IL-6, TNF-α, and MDA levels and elevated the SOD and GSH levels in the brain tissue(P<0.05). Compared with the sham group, the model group showed up-regulated protein levels of Bax, TLR4, p-NF-κB/NF-κB, NLRP3, pro-caspase-1, cleaved caspase-1, pro-IL-1β, and cleaved IL-1β and down-regulated protein level of Bcl-2 in the brain tissue(P<0.05). Compared with the MCAO model group, the acacetin and nimodipine groups showed down-regulated protein levels of Bax, TLR4, p-NF-κB/NF-κB, NLRP3, pro-caspase-1, cleaved caspase-1, pro-IL-1β, and cleaved IL-1β and up-regulated protein level of Bcl-2 in the brain tissue(P<0.05). In conclusion, acacetin regulates the TLR4/NLRP3 signaling pathway to inhibit neuroinflammatory response and oxidative stress, thus exerting the protective effect on cerebral ischemia-reperfusion injury in rats.
Subject(s)
Rats , Animals , NF-kappa B/metabolism , bcl-2-Associated X Protein , NLR Family, Pyrin Domain-Containing 3 Protein/metabolism , Tumor Necrosis Factor-alpha/metabolism , Rats, Sprague-Dawley , Caspase 1/metabolism , Toll-Like Receptor 4/metabolism , Nimodipine/pharmacology , Interleukin-6 , Rats, Wistar , Signal Transduction , Infarction, Middle Cerebral Artery , Reperfusion Injury/prevention & control , Superoxide Dismutase/metabolismABSTRACT
This study investigated the effect of Xiaoxuming Decoction(XXMD) on the activation of astrocytes after cerebral ischemia/reperfusion(I/R) injury. The model of cerebral IR injury was established using the middle cerebral artery occlusion method. Fluorocitrate(FC), an inhibitor of astrocyte activation, was applied to inhibit astrocyte activation. Rats were randomly divided into a sham group, a model group, a XXMD group, a XXMD+FC group, and a XXMD+Vehicle group. Neurobehavioral changes at 24 hours after cerebral IR injury, cerebral infarction, histopathological changes observed through HE staining, submicroscopic structure of astrocytes observed through transmission electron microscopy, fluorescence intensity of glial fibrillary acidic protein(GFAP) and thrombospondin 1(TSP1) measured through immunofluorescence, and expression of GFAP and TSP1 in brain tissue measured through Western blot were evaluated in rats from each group. The experimental results showed that neurobehavioral scores and cerebral infarct area significantly increased in the model group. The XXMD group, the XXMD+FC group, and the XXMD+Vehicle group all alleviated neurobehavioral changes in rats. The pathological changes in the brain were evident in the model group, while the XXMD group, the XXMD+FC group, and the XXMD+Vehicle group exhibited milder cerebral IR injury in rats. The submicroscopic structure of astrocytes in the model group showed significant swelling, whereas the XXMD group, the XXMD+FC group, and XXMD+Vehicle group protected the submicroscopic structure of astrocytes. The fluorescence intensity and protein expression of GFAP and TSP1 increased in the model group compared with those in the sham group. However, the XXMD group, the XXMD+FC group, and XXMD+Vehicle group all down-regulated the expression of GFAP and TSP1. The combination of XXMD and FC showed a more pronounced effect. These results indicate that XXMD can improve cerebral IR injury, possibly by inhibiting astrocyte activation and down-regulating the expression of GFAP and TSP1.
Subject(s)
Rats , Animals , Astrocytes , Brain Ischemia/metabolism , Brain , Reperfusion Injury/metabolism , Infarction, Middle Cerebral ArteryABSTRACT
OBJECTIVE@#To observe the effects of Xingnao Kaiqiao (regaining consciousness and opening orifices) acupuncture therapy on the expression of hypoxia-inducible factor 1α (HIF-1α) and Nod-like receptor protein 3 (NLRP3) in cerebral ischemia-reperfusion rats, and to explore the mechanism of acupuncture against cerebral ischemia-reperfusion injury.@*METHODS@#Seventy-two male SD rats were randomly divided into a sham-operation group, a model group, an acupuncture group and a non-point acupuncture group, with 18 rats in each one. Using modified Longa thread embolization method, the rat model of acute focal cerebral ischemia was prepared; and after 2 h ischemia, the reperfusion was performed to prepared the model of cerebral ischemia-reperfusion. Immediately after reperfusion, Xingnao Kaiqiao acupuncture method was applied to bilateral "Neiguan" (PC 6) and "Shuigou" (GV 26) in the acupuncture group, while in the non-point acupuncture group, acupuncture was delivered at non-points and all of the needles were retained for 30 min in these two groups. The samples were collected 24 h after reperfusion in the rats of each group. Zea-Longa neurological deficit score was used to evaluate the degree of cerebral neurological impairment, TTC staining was adopted to observe the volume percentage of cerebral infarction, HE staining was provided to observe the morphological changes of brain, and Western blot was applied for detecting the expression of HIF-1α and NLRP3 proteins in the cerebral cortex on the right side.@*RESULTS@#Compared with the sham-operation group, neurological deficit score and volume percentage of cerebral infarction were increased in the model group (P<0.01), and HIF-1α and NLRP3 protein expression was elevated (P<0.01). Compared with the model group, neurological deficit score and volume percentage of cerebral infarction were decreased (P<0.01), and HIF-1α and NLRP3 protein expression was lower (P<0.01) in the acupuncture group. There was no significant difference in above indexes in the non-point acupuncture group compared with the model group (P>0.05). Compared with the sham-operation group, the brain tissue of the rats in the model group and the non-point acupuncture group was loose and edema, and the nuclei were shriveled. The brain tissue morphology in the acupuncture group was similar to that of the sham-operation group.@*CONCLUSION@#Acupuncture can alleviate cerebral ischemia-reperfusion injury, and its mechanism may be related to the regulation of HIF-1α/NLRP3 signaling pathway to attenuate inflammatory response.
Subject(s)
Male , Animals , Rats , Rats, Sprague-Dawley , NLR Family, Pyrin Domain-Containing 3 Protein/genetics , Acupuncture Therapy , Reperfusion Injury/therapy , Brain Ischemia/therapy , Cerebral Infarction/therapy , NLR ProteinsABSTRACT
Cerebral ischemia-reperfusion is a serious cerebrovascular disease with high morbidity and mortality. In recent years， reperfusion therapy based on thrombolysis and thrombectomy has been the main treatment method for patients with ischemic stroke. Numerous studies have shown that Chinese medicine saponins can effectively interfere with cerebral ischemia-reperfusion in a multi-target and multi-way manner， which have great potential on the treatment and prevention of cerebral ischemia-reperfusion. Taking China National Knowledge Infrastructure （CNKI） literature database and Wanfang literature database as the analysis sources， this paper used SPSS statistics to summarize the number of papers on the treatment of cerebral ischemia-reperfusion with Chinese medicine saponins and CiteSpace to conduct cluster analysis on the high-frequency keywords of the research， thereby expounding the research hotspots and research status of Chinese medicine saponins in the treatment of cerebral ischemia-reperfusion. Based on literature analysis and summary of animal experiments on the treatment of cerebral ischemia-reperfusion with Chinese medicine saponins in the past two decades， Chinese medicine saponins exerted effects by anti-inflammation， inhibition of oxidative stress， immune regulation， protection of nerve cells， inhibition of thrombosis， promotion of thrombolysis， protection of mitochondria， blood-brain barrier repairing， and other ways. The specific mechanism， therapeutic effect， and signaling pathway of each Chinese medicine saponin have been summarized in this study， which provide a theoretical basis for the in-depth research， new drug development， and clinical application of Chinese medicine saponins for the treatment of cerebral ischemia- reperfusion.
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The purpose of this study is to investigate whether chrysin reduces cerebral ischemia-reperfusion injury(CIRI) by inhi-biting ferroptosis in rats. Male SD rats were randomly divided into a sham group, a model group, high-, medium-, and low-dose chrysin groups(200, 100, and 50 mg·kg~(-1)), and a positive drug group(Ginaton, 21.6 mg·kg~(-1)). The CIRI model was induced in rats by transient middle cerebral artery occlusion(tMCAO). The indexes were evaluated and the samples were taken 24 h after the operation. The neurological deficit score was used to detect neurological function. The 2,3,5-triphenyl tetrazolium chloride(TTC) staining was used to detect the cerebral infarction area. Hematoxylin-eosin(HE) staining and Nissl staining were used to observe the morphological structure of brain tissues. Prussian blue staining was used to observe the iron accumulation in the brain. Total iron, lipid pero-xide, and malondialdehyde in serum and brain tissues were detected by biochemical reagents. Real-time quantitative polymerase chain reaction(RT-qPCR), immunohistochemistry, and Western blot were used to detect mRNA and protein expression of solute carrier fa-mily 7 member 11(SLC7A11), transferrin receptor 1(TFR1), glutathione peroxidase 4(GPX4), acyl-CoA synthetase long chain family member 4(ACSL4), and prostaglandin-endoperoxide synthase 2(PTGS2) in brain tissues. Compared with the model group, the groups with drug intervention showed restored neurological function, decreased cerebral infarction rate, and alleviated pathological changes. The low-dose chrysin group was selected as the optimal dosing group. Compared with the model group, the chrysin groups showed reduced content of total iron, lipid peroxide, and malondialdehyde in brain tissues and serum, increased mRNA and protein expression levels of SLC7A11 and GPX4, and decreased mRNA and protein expression levels of TFR1, PTGS2, and ACSL4. Chrysin may regulate iron metabolism via regulating the related targets of ferroptosis and inhibit neuronal ferroptosis induced by CIRI.
Subject(s)
Rats , Male , Animals , Rats, Sprague-Dawley , Ferroptosis , Signal Transduction , Brain Ischemia/metabolism , Cyclooxygenase 2/metabolism , RNA, Messenger , Cerebral Infarction , Reperfusion Injury/metabolism , Malondialdehyde , Infarction, Middle Cerebral ArteryABSTRACT
Based on GC-MS and network pharmacology, the active constituents, potential targets, and mechanism of essential oil from Gleditsiae Fructus Abnormalis(EOGFA) against cerebral ischemia/reperfusion(I/R) injury were explored, and the effective constituents were verified by experiment. To be specific, GC-MS was used identify the constituents of the volatile oil. Secondly, the targets of the constituents and disease were predicted by network pharmacology, and the drug-constituent-target network was constructed, followed by Gene Ontology(GO) term enrichment and Kyoto Encyclopedia of Genes and Genomes(KEGG) pathway enrichment of the core targets. Molecular docking was performed to investigate the binding affinity between the active constituents and the targets. Finally, SD rats were used for experimental verification. The I/R injury model was established, and the neurological behavior score, infarct volume, and pathological morphology of brain tissue were measured in each group. The content of interleukin-1β(IL-1β), interleukin-6(IL-6), and tumor necrosis factor-alpha(TNF-α) was determined by enzyme-linked immunosorbent assay(ELISA), and the protein expression of vascular endothelial growth factor(VEGF) by Western blot. A total of 22 active constituents and 17 core targets were screened out. The core targets were involved in 56 GO terms and the major KEGG pathways of TNF signaling pathway, VEGF signaling pathway, and sphingolipid signaling pathway. Molecular docking showed that the active constituents had high affinity to the targets. The results of animal experiment suggested that EOGFA can alleviate the neurological impairment, decrease the cerebral infarct volume and the content of IL-1β, IL-6 and TNF-α, and down-regulate the expression of VEGF. The experiment verified the part results of network pharmacology. This study reflects the multi-component, multi-target, and multi-pathway characteristics of EOGFA. The mechanism of its active constituents is related to TNF and VEGF pathways, which provides a new direction for in-depth research on and secondary development of Gleditsiae Fructus Abnormalis.
Subject(s)
Animals , Rats , Rats, Sprague-Dawley , Network Pharmacology , Oils, Volatile , Gas Chromatography-Mass Spectrometry , Interleukin-6 , Molecular Docking Simulation , Tumor Necrosis Factor-alpha , Vascular Endothelial Growth Factor A , Reperfusion Injury , Cerebral InfarctionABSTRACT
This study explores the effect of total flavonoids of Rhododendra simsii(TFR) on middle cerebral artery occlusion(MCAO)-induced cerebral injury in rats and oxygen-glucose deprivation/reoxygenation(OGD/R) injury in PC12 cells and the underlying mechanism. The MCAO method was used to induce focal ischemic cerebral injury in rats. Male SD rats were randomized into sham group, model group, and TFR group. After MCAO, TFR(60 mg·kg~(-1)) was administered for 3 days. The content of tumor necrosis factor-α(TNF-α), interleukin-1(IL-1), and interleukin-6(IL-6) in serum was detected by enzyme-linked immunosorbent assay(ELISA). The pathological changes of brain tissue and cerebral infarction were observed based on hematoxylin and eosin(HE) staining and 2,3,5-triphenyltetrazolium chloride(TTC) staining. RT-qPCR and Western blot were used to detect the mRNA and protein levels of calcium release-activated calcium channel modulator 1(ORAI1), stromal interaction molecule 1(STIM1), stromal intera-ction molecule 2(STIM2), protein kinase B(PKB), and cysteinyl aspartate specific proteinase 3(caspase-3) in brain tissues. The OGD/R method was employed to induce injury in PC12 cells. Cells were randomized into the normal group, model group, gene silencing group, TFR(30 μg·mL~(-1)) group, and TFR(30 μg·mL~(-1))+gene overexpression plasmid group. Intracellular Ca~(2+) concentration and apoptosis rate of PC12 cells were measured by laser scanning confocal microscopy and flow cytometry. The effect of STIM-ORAI-regulated store-operated calcium entry(SOCE) pathway on TFR was explored based on gene silencing and gene overexpression techniques. The results showed that TFR significantly alleviated the histopathological damage of brains in MCAO rats after 3 days of admini-stration, reduced the contents of TNF-α, IL-1, and IL-6 in the serum, down-regulated the expression of ORAI1, STIM1, STIM2, and caspase-3 genes, and up-regulated the expression of PKB gene in brain tissues of MCAO rats. TFR significantly decreased OGD/R induced Ca~(2+) overload and apoptosis in PC12 cells. However, it induced TFR-like effect by ORAI1, STIM1 and STIM2 genes silencing. However, overexpression of these genes significantly blocked the effect of TFR in reducing Ca~(2+) overload and apoptosis in PC12 cells. In summary, in the early stage of focal cerebral ischemia-reperfusion injury and OGD/R-induced injury in PC12 cells TFR attenuates ischemic brain injury by inhibiting the STIM-ORAI-regulated SOCE pathway and reducing Ca~(2+) overload and inflammatory factor expression, and apoptosis.
Subject(s)
Animals , Male , Rats , Apoptosis , Brain Ischemia/metabolism , Caspase 3 , Interleukin-1 , Interleukin-6 , Rats, Sprague-Dawley , Reperfusion Injury/metabolism , Tumor Necrosis Factor-alpha/genetics , Flavonoids/pharmacology , Rhododendron/chemistryABSTRACT
This study aims to explore the effect of Xiaoxuming Decoction on synaptic plasticity in rats with acute cerebral ischemia-reperfusion. A rat model of cerebral ischemia-reperfusion injury was established by middle cerebral artery occlusion(MCAO). Rats were randomly assigned into a sham group, a MCAO group, and a Xiaoxuming Decoction(60 g·kg~(-1)·d~(-1)) group. The Longa score was rated to assess the neurological function of rats with cerebral ischemia for 1.5 h and reperfusion for 24 h. The 2,3,5-triphenyltetrazolium chloride(TTC) staining and hematoxylin-eosin(HE) staining were employed to observe the cerebral infarction and the pathological changes of brain tissue after cerebral ischemia, respectively. Transmission electron microscopy was employed to detect the structural changes of neurons and synapses in the ischemic penumbra, and immunofluorescence, Western blot to determine the expression of synaptophysin(SYN), neuronal nuclei(NEUN), and postsynaptic density 95(PSD95) in the ischemic penumbra. The experimental results showed that the modeling increased the Longa score and led to cerebral infarction after 24 h of ischemia-reperfusion. Compared with the model group, Xiaoxuming Decoction intervention significantly decreased the Longa score and reduced the formation of cerebral infarction area. The modeling led to the shrinking and vacuolar changes of nuclei in the brain tissue, disordered cell arrangement, and severe cortical ischemia-reperfusion injury, while the pathological damage in the Xiaoxuming Decoction group was mild. The modeling blurred the synaptic boundaries and broadened the synaptic gap, while such changes were recovered in the Xiaoxuming Decoction group. The modeling decreased the fluorescence intensity of NEUN and SYN, while the intensity in Xiaoxuming Decoction group was significantly higher than that in the model group. The expression of SYN and PSD95 in the ischemic penumbra was down-regulated in the model group, while such down-regulation can be alleviated by Xiaoxuming Decoction. In summary, Xiaoxuming Decoction may improve the synaptic plasticity of ischemic penumbra during acute cerebral ischemia-reperfusion by up-regulating the expression of SYN and PSD95.
Subject(s)
Rats , Animals , Rats, Sprague-Dawley , Brain Ischemia/drug therapy , Reperfusion Injury/metabolism , Infarction, Middle Cerebral Artery , Neuronal Plasticity , ReperfusionABSTRACT
Based on the O-GlcNAc transferase(OGT)-PTEN-induced putative kinase 1(PINK1) pathway, the mechanism of 3,4-dihydroxybenzaldehyde(DBD) on mitochondrial quality control was investigated. Middle cerebral artery occlusion/reperfusion(MCAO/R) rats were established. SD rats were randomized into sham operation group(sham), model group(MCAO/R), DBD-L group(5 mg·kg~(-1)), and DBD-H group(10 mg·kg~(-1)). After 7 days of administration(ig), MCAO/R was induced in rats except the sham group with the suture method. Twenty-four h after reperfusion, the neurological function and the percentage of cerebral infarct area were measured. Based on hematoxylin and eosin(HE) staining and Nissl staining, the pathological damage of cerebral neurons was examined. Then the ultrastructure of mitochondria was observed under the electron microscope, and the co-localization of light chain-3(LC3), sequestosome-1(SQSTM1/P62), and Beclin1 was further detected by immunofluorescence staining. It has been reported that the quality of mitochondria can be ensured by inducing mitochondrial autophagy through the OGT-PINK1 pathway. Therefore, Western blot was employed to detect the expression of OGT, mitophagy-related proteins PINK1 and E3 ubiquitin ligase(Parkin), and mitochondrial kinetic proteins dynamin-like protein 1(Drp1) and optic atrophy 1(Opa1). The results showed that MCAO/R group had neurological dysfunction, large cerebral infarct area(P<0.01), damaged morphological structure of neurons, decreased number of Nissl bodies, mitochondrial swelling, disappearance of mitochondrial cristae, decrease of cells with LC3 and Beclin1, rise of cells with P62(P<0.01), inhibited expression of OGT, PINK1, and Parkin, up-regulated expression of Drp1, and down-regulated expression of Opa1 compared with the sham group(P<0.01). However, DBD improved the behavioral deficits and mitochondrial health of MCAO/R rats, as manifested by the improved morphology and structure of neurons and mitochondria and the increased Nissl bodies. Moreover, DBD increased cells with LC3 and Beclin1 and decreased cells with P62(P<0.01). In addition, DBD promoted the expression of OGT, PINK1, Parkin, and Opa1 and inhibited the expression of Drp1, enhancing mitophagy(P<0.05, P<0.01). In conclusion, DBD can trigger PINK1/Parkin-mediated brain mitophagy through the OGT-PINK1 pathway, which plays a positive role in maintaining the health of the mitochondrial network. This may be a mitochondrial therapeutic mechanism to promote nerve cell survival and improve cerebral ischemia/reperfusion injury.
Subject(s)
Animals , Rats , Rats, Sprague-Dawley , Beclin-1 , Mitochondria , Cerebral Infarction , Protein KinasesABSTRACT
OBJECTIVE@#To improve the classical 4-vessel occlusion (4VO) model established by Pulsinelli and Brierley.@*METHODS@#Thirty-two male SD rats were randomized into sham operation group, I4VO-Con10 group, I4VO-Int10 group and I4VO-Int15 group. The sham surgery group underwent exposure of the bilateral vertebral arteries and carotid arteries without occlusion to block blood flow. The I4VO-Con10 group experienced continuous ischemia by occluding the bilateral vertebral arteries and carotid arteries for 10 minutes followed by reperfusion for 24 hours. The I4VO-Int10 and I4VO-Int15 groups were subjected to intermittent ischemia. The I4VO- Int10 group underwent 5 minutes of ischemia, followed by 5 minutes of reperfusion and another 5 minutes of ischemia, and then reperfusion for 24 hours. The I4VO-Int15 group experienced 5 minutes of ischemia followed by two cycles of 5 minutes of reperfusion and 5 minutes of ischemia, and then reperfusion for 24 hours. The regional cerebral blood flow (rCBF) was monitored with laser Doppler scanning, and survival of the rats was observed. HE staining was used to observe hippocampal pathologies to determine the optimal method for modeling. Another 48 rats were randomized into 6 groups, including a sham operation group and 5 model groups established using the optimal method. The 5 I4VO model groups were further divided based on the reperfusion time points (1, 3, 7, 14, and 28 days) into I4VO-D1, I4VO-D3, I4VO-D7, I4VO- D14, and I4VO- D28 groups. Body weight changes and survival of the rats were recorded. HE staining was used to observe morphological changes in the hippocampal, retinal and optic tract tissues. The Y-maze test and light/dark box test were used to evaluate cognitive and visual functions of the rats in I4VO-D28 group.@*RESULTS@#Occlusion for 5 min for 3 times at the interval of 5 min was the optimal method for 4VO modeling. In the latter 48 rats, the body weight was significantly lower than that of the sham-operated rats at 1, 3, 7, 14 and 28 days after modeling without significant difference in survival rate among the groups. The rats with intermittent vessel occlusion exhibited progressive deterioration of hippocampal neuronal injury and neuronal loss. Cognitive impairment was observed in the rats in I4VO-D28 group, but no obvious ischemic injury of the retina or the optic tract was detected.@*CONCLUSION@#The improved 4VO model can successfully mimic the main pathological processes of global cerebral ischemia-reperfusion injury without causing visual impairment in rats.
Subject(s)
Rats , Male , Animals , Rats, Sprague-Dawley , Brain Ischemia , Cerebral Infarction , Reperfusion Injury , Body WeightABSTRACT
ObjectiveTo investigate the mechanism of Huazhuo Jiedu Huoxue Tongluo prescription in alleviating cerebral ischemia-reperfusion injury via regulating nerve cell autophagy based on c-Jun N-terminal kinase（JNK）signaling pathway . MethodSixty SD rats were randomly divided into 6 groups： sham group， middle cerebral artery occlusion/reperfusion （MCAO/R） group （model group）， Huazhuo Jiedu Huoxue Tongluo prescription group ［traditional Chinese medicine （TCM） group（25.0 g·kg-1）］， JNK inhibitor SP600125 （SP） group（5 mg·kg-1）， TCM+SP group and JNK agonist Anisomycin （Ani） group（15 mg·kg-1）. After 24 h of modeling， TCM group and TCM+SP group were given TCM decoction （ig） for 3 consecutive days， and the other groups were given equal volume of normal saline （ig）. Neurological deficit was evaluated by neurological function score and cerebral infarct volume was determined by 2，3，5-triphenyltetrazole chloride （TTC） staining. Hematoxylin-eosin （HE） staining and Nissl staining were used to observe the structural changes of brain tissue and the damage of neurons， respectively. Terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling assay （TUNEL） was performed to detect cell apoptosis. The ultrastructure of autophagosomes was observed by transmission electron microscope. Western blot was employed to detect the protein expressions of microtubule-associated protein 1 light chain 3A/B （LC3A/B）， autophagy related 5 （Atg5）， the ortholog of yeast Atg6 （Beclin1）， p62， B-cell lymphoma 2 （Bcl-2）， JNK， phosphorylated （p）-JNK and c-Jun in brain tissue. The mRNA expressions of LC3A/B， Beclin1， p62， Atg5， Bcl-2， JNK and c-Jun were detected by Real-time fluorescence quantitative polymerase chain reaction （Real-time PCR）. ResultCompared with the sham group， the model group had elevated neurological deficit score （P<0.05）， enlarged cerebral infarct volume （P<0.05）and typical infarction manifestations formed in hippocampal region and its surrounding brain tissue. In addition， there were a large number of neuronal cell degeneration， necrosis， liquefaction， nucleus pyknosis and deep staining， and inflammatory cell infiltration in the cortex in the model group， and severe swelling of mitochondria， lysosomes， autophagosomes and autophagolysosomes were clearly seen under electron microscope. TUNEL positive cells were increased （P<0.05）， and cell apoptosis was severe. The nuclear membrane and nucleolus of neurons in brain tissue were blurred with discontinuous processes， and Nissl bodies in cytoplasm were stained light with reduced number. Western blot revealed that the model group had up-regulated protein expressions of LC3A/B， Beclin1， Atg5， JNK， p-JNK and c-Jun in brain tissue （P<0.05）， while down-regulated protein expressions of p62 and Bcl-2 （P<0.05）as compared with the sham group. Real-time PCR indicated that the mRNA expressions of LC3A/B， Beclin1， Atg5， JNK and c-Jun in the model group were higher （P<0.05） while the mRNA expressions of p62 and Bcl-2 were lower （P<0.05） than those in the sham group. Compared with the conditions in model group， the neurological deficit scores of TCM， SP and TCM+SP groups were lowered （P<0.05）， and the cerebral infarct volume was reduced （P<0.05）， with improved pathological status of brain tissue， especially in the TCM group. Furthermore， there were abundant and basically normal mitochondrial cristae， slightly dilated endoplasmic reticulum， slightly swollen golgi apparatus， slightly fused nuclear membrane， and few visible lysosomes， autophagosomes and autophagolysosomes. TUNEL positive cells were decreased （P<0.05）， displaying reduced apoptosis， especially in the TCM group. The nucleolus and nuclear membrane of neurons in brain tissue were discernible， and Nissl bodies in cytoplasm was reduced to a certain degree as compared with those in the model group. Western blot showed a decrease in the protein expressions of LC3A/B， Beclin1， Atg5， JNK， p-JNK and c-Jun in the TCM group ，the SP group，and the TCM+SP group（P<0.05），while an increase in the protein expressions of p62 in the TCM group and SP group（P<0.05），and an increase in the protein expressions of Bcl-2 in the TCM group and TCM+SP group. By Real-time PCR， the mRNA expressions of LC3A， LC3B， Beclin1， Atg5， JNK and c-Jun had a down-regulation（P<0.05） while the mRNA expression of p62 a up-regulation in the TCM group ，the SP group，and the TCM+SP group（P<0.05），and the mRNA expression of Bcl-2 a up-regulation in the TCM group and the TCM+SP group（P<0.05）.Scores of the Ani group were raised （P<0.05）， and infarct volume was increased significantly， with aggravated neuronal cell necrosis and obvious inflammatory infiltration. Moreover， there were neuronal nuclear membrane fusion with abnormal protrusion， increased heterochromatin aggregation in edge， severe mitochondrial swelling， endoplasmic reticulum expansion， increased lysosomes， increased intracytoplasmic vesicles， and visible autophagosomes and autophagolysosomes. TUNEL positive cells were increased （P<0.05）， representing severe apoptosis. The number of Nissl bodies dropped with light staining， and the nucleolus and nuclear membrane were blurred. Real-time PCR found that the mNRA expressions of Atg5， c-Jun， JNK were up-regulated （P<0.05），while Beclin1， p62， Bcl-2 were were down-regulated in the Ani group （P<0.05）. Compared with the TCM group and SP group，the protein expressions of LC3A/B， Beclin1， Atg5， JNK， p-JNK， c-Jun were decreased，and p62， Bcl-2 were increased in the Ani group（P<0.05）. Compared with the TCM group，the mRNA expressions of JNK mRNA had a down-regulation in the SP group and TCM+SP group，while LC3A， LC3B， Atg5， c-Jun， JNK had an up-regulation（P<0.05） and Bcl-2 had a down-regulation in the Ani group（P<0.05）. Compared with the SP group，the mRNA expressions of Atg5， c-Jun， JNK had an up-regulation（P<0.05）， and Beclin1， p62， Bcl-2 had a down-regulation in the Ani group（P<0.05）. ConclusionHuazhuo Jiedu Huoxue Tongluo prescription significantly up-regulates the protein and mRNA expressions of LC3A/B， Beclin1， Atg5， JNK， p-JNK and c-Jun， and down-regulates the protein and mRNA expressions of p62 and Bcl-2， suggesting that the prescription can inhibit autophagy through JNK signaling pathway to reduce ischemia/reperfusion injury in rats.
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OBJECTIVE@#To observe the effect of Tongdu Tiaoshen (promoting the circulation of the governor vessel and regulating the spirit) electroacupuncture (EA) pretreatment on pyroptosis mediated by peroxisome proliferators-activated receptor γ (PPARγ) of the cerebral cortex in rats with cerebral ischemia reperfusion injury (CIRI) and explore the potential mechanism of EA for the prevention and treatment of CIRI.@*METHODS@#A total of 110 clean-grade male SD rats were randomly divided into a sham-operation group, a model group, an EA group, an EA + inhibitor group and an agonist group, 22 rats in each group. In the EA group, before modeling, EA was applied to "Baihui" (GV 20), "Fengfu" (GV 16) and "Dazhui" (GV 14), with disperse-dense wave, 2 Hz/5 Hz in frequency, 1 to 2 mA in intensity, lasting 20 min; once a day, consecutively for 7 days. On the base of the intervention as the EA group, on the day 7, the intraperitoneal injection with the PPARγ inhibitor, GW9662 (10 mg/kg) was delivered in the EA + inhibitor group. In the agonist group, on the day 7, the PPARγ agonist, pioglitazone hydrochloride (10 mg/kg) was injected intraperitoneally. At the end of intervention, except the sham-operation group, the modified thread embolization method was adopted to establish the right CIRI model in the rats of the other groups. Using the score of the modified neurological severity score (mNSS), the neurological defect condition of rats was evaluated. TTC staining was adopted to detect the relative cerebral infarction volume of rat, TUNEL staining was used to detect apoptosis of cerebral cortical nerve cells and the transmission electron microscope was used to observe pyroptosis of cerebral cortical neural cells. The positive expression of PPARγ and nucleotide-binding to oligomerization domain-like receptor protein 3 (NLRP3) in the cerebral cortex was detected with the immunofluorescence staining. The protein expression of PPARγ, NLRP3, cysteinyl aspartate specific protease-1 (caspase-1), gasdermin D (GSDMD) and GSDMD-N terminal (GSDMD-N) in the cerebral cortex was detected with Western blot. Using the quantitative real-time fluorescence-PCR, the mRNA expression of PPARγ, NLRP3, caspase-1 and GSDMD of the cerebral cortex was detected. The contents of interleukin (IL)-1β and IL-18 in the cerebral cortex of rats were determined by ELISA.@*RESULTS@#Compared with the sham-operation group, the mNSS, the relative cerebral infarction volume and the TUNEL positive cells rate were increased (P<0.01), pyroptosis was severe, the protein and mRNA expression levels of PPARγ, NLRP3, caspase-1 and GSDMD were elevated (P<0.01); and the protein expression of GSDMD-N and contents of IL-1β and IL-18 were increased (P<0.01) in the model group. When compared with the model group, the mNSS, the relative cerebral infarction volume and the TUNEL positive cells rate were decreased (P<0.01), pyroptosis was alleviated, the protein and mRNA expression levels of PPARγ were increased (P<0.01), the protein and mRNA expression levels of NLRP3, caspase-1 and GSDMD were decreased (P<0.01), the protein expression of GSDMD-N was reduced (P<0.01); and the contents of IL-1β and IL-18 were lower (P<0.01) in the EA group and the agonist group; while, in the EA + inhibitor group, the protein expression of PPARγ was increased (P<0.01), the protein and mRNA expression levels of NLRP3 and GSDMD were decreased (P<0.01, P<0.05), the mRNA expression of caspase-1 was reduced (P<0.01); and the contents of IL-1β and IL-18 were lower (P<0.01). When compared with the EA + inhibitor group, the mNSS, the relative cerebral infarction volume and the TUNEL positive cells rate were decreased (P<0.05, P<0.01), pyroptosis was alleviated, the protein and mRNA expression levels of PPARγ were increased (P<0.01), the protein and mRNA expression levels of NLRP3, caspase-1 and GSDMD were decreased (P<0.01), the protein expression of GSDMD-N was reduced (P<0.01); and the contents of IL-1β and IL-18 were declined (P<0.01) in the EA group. Compared with the agonist group, in the EA group, the relative cerebral infarction volume and the TUNEL positive cells rate were increased (P<0.05, P<0.01), the mRNA expression of PPARγ was decreased (P<0.01) and the protein expression of GSDMD-N was elevated (P<0.05); and the contents of IL-1β and IL-18 were higher (P<0.01).@*CONCLUSION@#Tongdu Tiaoshen EA pretreatment can attenuate the neurological impairment in the rats with CIRI, and the underlying mechanism is related to the up-regulation of PPARγ inducing the inhibition of NLRP3 in the cerebral cortex of rats so that pyroptosis is affected.
Subject(s)
Male , Animals , Rats , Rats, Sprague-Dawley , PPAR gamma/genetics , Pyroptosis , Interleukin-18 , Electroacupuncture , NLR Family, Pyrin Domain-Containing 3 Protein , Cerebral Cortex , Cerebral Infarction/therapy , Caspases , RNA, MessengerABSTRACT
OBJECTIVE@#To investigate the neuroprotective effect of electroacupuncture (EA) at "Quchi" (LI 11) and "Zusanli" (ST 36) in the rats with cerebral ischemic reperfusion and the potential mechanism of microglia pyroptosis.@*METHODS@#Sixty SD rats were randomly divided into a sham-operation group, a model group and an EA group, with 20 rats in each group. The Zea Longa method was employed to establish the rat model of the middle cerebral artery occlusion and reperfusion (MACO/R) in the left brain. In the EA group, since the 2nd day of modeling, EA was given at "Quchi" (LI 11) and "Zusanli" (ST 36) of right side with disperse-dense wave, 4 Hz/20 Hz in frequency and 0.2 mA in current intensity, 30 min each time, once a day for lasting 7 consecutive days. The reduction rate of cerebral blood flow was measured with laser Doppler flowmetry during operation. The neurological function of rats was observed using Zea Longa neurobehavioral score. The cerebral infarction volume was detected by TTC staining method. The microglia positive expression in the ischemic side of the cortex was detected with the immunofluorescence method. Under transmission electron microscope, the ultrastructure of cell in the ischemic cortex was observed. The mRNA expression levels of nucleotide-binding oligomerization domain-like receptor protein 3 (NLRP3), apoptosis-associated speck-like protein containing a caspase recruitment domain (ASC), cysteinyl aspartate specific proteinase-1 (Caspase-1) and gasdermin D (GSDMD) in the ischemic cortex were detected using real-time PCR.@*RESULTS@#Compared with the sham-operation group, in the model group, the reduction rate of cerebral blood flow was increased during operation (P<0.001); Zea Longa neurobehavional score and the percentage of cerebral infarction volume were increased (P<0.001), the numbers of M1-type microglia marked by CD68+ and M2-type microglia marked by TMEM119+ were elevated in the ischemic cortex (P<0.001), the mRNA expression of NLRP3, ASC, Caspase-1 and GSDMD was increased (P<0.001, P<0.01); the cytomembrane structure was destroyed, with more cell membrane pores formed in the ischemic cortex. Compared with the model group, after intervention, Zea Longa neurobehavioral score and the percentage of cerebral infarction volume were reduced (P<0.05), the number of M1-type microglia marked by CD68+ was reduced (P<0.05) and the number of M2-type microglia marked by TMEM119+ was increased (P<0.05); and the mRNA expression of NLRP3, ASC, Caspase-1 and GSDMD was decreased (P<0.01, P<0.05) in the EA group. Even though the cytomembrane structure was incomplete, there were less membrane pores presented in the ischemic cortex in the EA group after intervention.@*CONCLUSION@#The intervention with EA attenuates the neurological dysfunction and reduces the volume of cerebral infarction in the rats with cerebral ischemic reperfusion. The underlying mechanism is related to the inhibition of microglia pyroptosis through modulating NLRP3/Caspase-1/GSDMD axis.
Subject(s)
Animals , Rats , Rats, Sprague-Dawley , Caspase 1/genetics , NLR Family, Pyrin Domain-Containing 3 Protein/genetics , Electroacupuncture , Cerebral Infarction/therapy , RNA, MessengerABSTRACT
ObjectiveTo investigate the effects of Buyang Huanwutang （BYWHT） on reducing inflammatory injury and improving neurological function after cerebral ischemia-reperfusion in rats via activating the adiponectin （APN） pathway. MethodMale SD rats were randomized into sham， model， BYHWT （16 g·kg-1， twice a day）， and BYHWT + APN inhibitor （GW9662） groups. In the sham group， blood vessels were isolated. The rat model of middle cerebral artery occlusion （MCAO） was established. The rats in the BYHWT+GW9662 group was treated with subcutaneous injection of GW9662 at 4 m·kg-1 30 min before MCAO surgery and BYHWT at 16 g·kg-1 by gavage after MCAO surgery， once in the morning and once in the evening. The immunofluorescence （IF） assay was employed to observe the expression of adiponectin receptor 1 （AdipoR1） and the colocalization of AdipoR1 with neuronal nuclei （NeuN） and ionized calcium binding adaptor molecule 1 （Iba1） in the brain. Enzyme-linked immunosorbent assay （ELISA） was employed to detect the expression of APN in the serum and brain. The balance beam test was carried out to examine the balance ability， and the grasping test to assess the recovery of limb strength. The immunofluorescence assay was used to detect the expression of myeloperoxidase （MPO）. Western blot was employed to determine the expression of tumor necrosis factor-alpha （TNF-α）， interleukin （IL）-1β， IL-6， and IL-10 and in the brain. ResultCompared with the sham group， the modeling promoted the expression of AdipoR1 （P<0.01）， lowered the APN levels in the serum and brain （P<0.05， P<0.01）， increased the score in the balance beam test （P<0.01）， and decreased the grasping strength of forepaw （P<0.01）， which were accompanied with increased MPO， TNF-α， IL-1β， and IL-6 levels （P<0.01） and decreased IL-10 level （P<0.01）. Compared with the model group， BYHWT promoted the expression of AdipoR1 （P<0.01）， elevated APN levels in the serum and brain （P<0.05， P<0.01）， and increased the grasping strength of forepaw （P<0.01）， which were accompanied with lowered MPO， TNF-α， IL-1β， and IL-6 levels （P<0.01） and elevated IL-10 level （P<0.01）. All the above effects were partially blocked by GW9662. ConclusionBYHWT can reduce inflammatory injury and improve neurological function in the rat model of cerebral ischemia-reperfusion by activating the APN pathway.
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Objective:To explore the mechanism of gypenoside ⅩⅦ against cerebral ischemia/reperfusion (I/R) through nuclear factor erythroid 2-related factor 2/antioxidant responsive element (Nrf2/ARE) signaling pathway.Methods:Forty SPF Sprague Dawley (SD) rats were randomly divided into sham operated group, I/R model group, 25, 50 and 100 mg/kg gypenoside ⅩⅦ groups ( n = 8). Gypenoside ⅩⅦ groups were administered 25, 50 or 100 mg/kg (0.01 mL/g) gypenoside ⅩⅦ by intragastric administration for 14 days; the other two groups received the same dose of saline. Rat cerebral I/R model was established by modified line bolt method; rats in the sham operated group underwent the same procedure without producing substantial embolization. After 24 hours of reperfusion, the neurological deficit scores of the rats in each group were assessed. Rat abdominal aortic whole blood was collected and the serum reactive oxygen species (ROS), heme oxygenase-1 (HO-1), γ-glutamylcysteine synthase (γ-GCS), superoxide dismutase (SOD), quinone NADH oxidoreductase 1 (NQO1), and malondialdehyde (MDA) were detected. Then whole brain tissue was harvested and penumbra tissue was isolated from cerebral cortex, the general condition of rat brain tissue and the volume of cerebral infarction were evaluated, the histopathological changes in the brain were observed under light microscopy, the mRNA expressions of Nrf2 and Keap1 were measured by real-time fluorescent quantitative polymerase chain reaction (RT-qPCR), the protein expressions of Nrf2 and Keap1 were determined by Western blotting. Results:After 24 hours of reperfusion, compared with the sham operated group, the score of neurological deficit and infarct volume were significantly increased, the NQO1, SOD and γ-GCS levels in serum were significantly decreased, MDA, HO-1 and ROS levels in serum were significantly increased, the Nrf2 and Keap1 mRNA and protein expressions in the ischemic penumbra were significantly increased in rats from I/R model group. Compared with the I/R model group, the neurological deficit scores (1.50±0.53, 1.37±0.52 vs. 2.75±0.46) and brain infarct volume [(19.8±5.1)%, (21.4±6.4)% vs. (42.3±5.8)%] were significantly reduced, serum NQO1, SOD, HO-1 and γ-GCS were significantly increased [NQO1 (ng/L): 186.05±10.38, 220.75±16.22 vs. 131.36±5.95, SOD (kU/L): 63.23±5.30, 72.70±8.62 vs. 36.75±6.55, HO-1 (ng/L): 60.57±7.93, 60.35±4.72 vs. 42.72±4.95, γ-GCS (kU/L): 8.81±0.53, 8.72±0.69 vs. 6.80±0.56], serum MDA and ROS levels were significantly reduced [MDA (μmol/L): 5.94±0.66, 5.61±0.53 vs. 10.88±1.34, ROS (kU/L): 69.11±4.23, 67.12±4.52 vs. 104.43±7.54], the mRNA and protein expressions of Nrf2 and Keap1 in the ischemic penumbra were significantly increased in rats from 50 mg/kg and 100 mg/kg gypenoside ⅩⅦ groups [Nrf2 mRNA (2 -△△Ct): 1.90±0.13, 2.13±0.18 vs. 1.48±0.11, Keap1 mRNA (2 -△△Ct): 1.78±0.11, 1.85±0.10 vs. 1.43±0.10, Nrf2/β-actin: 0.73±0.04, 0.79±0.03 vs. 0.60±0.03, Keap1/β-actin: 0.71±0.01, 0.76±0.03 vs. 0.61±0.01], all the comparative differences were statistically significant (all P < 0.01); 25 mg/kg gypenoside ⅩⅦ had no significant effect. Conclusion:Gypenoside ⅩⅦ (50 mg/kg and 100 mg/kg) may play a role in anti-cerebral I/R injury by regulating NQO1, SOD, HO-1, γ-GCS, ROS and MDA through Nrf2/ARE signaling pathway.