Project description:Background: Electroacupuncture (EA) treatment in ischemic stroke has been highlighted recently; however, the specific mechanism is still elusive. Autophagy is considered a new target for cerebral ischemia/reperfusion (I/R), but whether it plays a role of protecting or causing rapid cell apoptosis remains unclear. Studies have reported that the reduction in lysine 16 of histone H4 acetylation coheres with autophagy induction. The primary purpose of the study was to explore whether EA could alleviate I/R via autophagy-mediated histone H4 lysine 16 acetylation in the middle cerebral artery occlusion (MCAO) rat model. Methods: One hundred and twenty male Sprague-Dawley rats were divided into five groups: control group, MCAO group, MCAO+EA group, MCAO+EA+hMOF siRNA group, and MCAO+EA+Sirt1 inhibitor group. EA was applied to "Baihui" (Du20) and "Renzhong" (Du26) at 5 min after modeling and 16 h after the first EA intervention. The structure and molecular markers of the rat brain were evaluated. Results: EA significantly alleviated I/R injury by upregulating the expressions of Sirt1, Beclin1, and LC3-II and downregulating the expressions of hMOF and H4K16ac. In contrast, the Sirt1 inhibitor lowered the increase in Sirt1, Beclin1, and LC3-II and enhanced the level of hMOF and H4K16ac expressions associated with EA treatment. Besides, ChIP assay revealed that the binding of H4K16ac in the Beclin1 promoter region of the autophagy target gene was significantly raised in the MCAO+EA group and MCAO+EA+hMOF siRNA group. Conclusions: EA treatment inhibited the H4K16ac process, facilitated autophagy, and alleviated I/R injury. These findings suggested that regulating histone H4 lysine 16 acetylation-mediated autophagy may be a key mechanism of EA at Du20 and Du26 to treat I/R.
Project description:BackgroundIschemic stroke is a deadly disease that poses a serious threat to human life. Superoxide dismutase 3 (SOD3, ECSOD) is the main antioxidant enzyme that removes superoxide anions from cells. This study aimed to investigate the effect of SOD3 overexpression on cerebral ischemia-reperfusion injury in rats.MethodsGV230-EGFP-ECSOD, the recombinant SOD3-overexpressed vector, was constructed by genetic engineering technology, and mesenchymal stem cells (MSCs) were infected with lentiviral packaging. In animal experiment, cerebral ischemia-reperfusion injury model rats were successfully established. ECSOD-MSCs are the MSCs that successfully transfected with SOD3 overexpression vector. The animals were injected with ECSOD-MSCs (ECSOD-MSC group), normal MSCs (MSCs group), PBS (PBS group), and not do any processing (Model group) via the tail vein. Then MRI was used to detect the infarct volume of rats, modified Neurological Severity Scores (mNSS), and immunohistochemistry were used to evaluate the expression of neurological function and apoptosis-related genes in rats.ResultsWestern blot analysis revealed that the SOD3 was highly expressed in MSCs. Animal experiments showed that the transplantation of ECSOD-MSCs significantly reduced the infarct volume of ischemic stroke rats (p < 0.05), significantly improved neurological function in rats (p < 0.05), and found proapoptotic gene, Bax, expression was significantly decreased (p < 0.05), the expression of anti-apoptotic gene, Bcl-2, was significantly increased (p < 0.05). The highly expressed SOD3 has no correction with brain infarct volume, and the highly expressed SOD3 has a positive correlation with cell apoptosis. It is speculated that overexpression of SOD3 affects the expression of Bax and Bcl-2, and improves apoptosis to alleviate ischemic stroke.ConclusionOur results indicated that MSCs transfected with SOD3 can effectively alleviate cerebral ischemia-reperfusion injury in rats.
Project description:BackgroundThis study aimed to investigate dynamic urinary proteome changes of electroacupuncture (EP) on cerebral ischemia-reperfusion (CI/R) injured rats and to explore the therapeutic biological mechanisms of EP.MethodsFirst, changed urinary proteins were found in EP stimulation in healthy rats. Then, we used a CI/R injury rat model induced by Pulsinelli's four-vessel occlusion (4-VO) method to explore the function of EP on urinary proteome in CI/R injury. Urine samples were collected for proteome analysis by liquid chromatography-tandem mass spectrometry (LC-MS/MS) and bioinformatics analysis.ResultsIn total, 384 proteins were identified, among which 47 proteins (23 upregulated, 24 downregulated) were differentially expressed with 0.6-log FC and p < .05. Gene ontology analysis revealed that the cell redox homeostasis, acute-phase response, response to lipopolysaccharide, and cellular response to glucocorticoid stimulus were significantly enriched. The partially biologically connected differential proteins were found by the Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis in the EP group. With the CI/R rat model, 80 proteins (27 upregulated, 53 downregulated) were significantly changed in the CI/R rats compared to the controls. Among these differentially expressed proteins (DEPs), 23 proteins (17 upregulated, six downregulated) showed significant changes after EP treatment (0.6-log FC change, p < .05). The main related biological processes were aging, immune response, acute-phase response, liver regeneration, protein catabolic process, and response to oxidative stress. Many metabolic pathways were enriched by KEGG analysis.ConclusionOur results indicate that the EP could alleviate cerebral damage induced by ischemia-reperfusion through an anti-inflammatory and metabolism regulation mechanism. The urinary proteome might reflect the pathophysiological changes in EP pretreatment in the treatment and prevention of CI/R injury.
Project description:Electroacupuncture (EA) therapy via alternating current stimulation on the scalp over the motor cortex is used for the treatment of brain disorders. Perinatal hypoxia-ischemia (HI), a brain injury in newborns, leads to long-term neurologic complications. Here, we investigated whether EA could promote functional improvements and neurogenesis in a neonatal HI rat model. A neonatal HI rat model was induced by permanent ligation of the left carotid artery in postnatal day 7 pups. EA for neonatal HI rats was performed at 2 Hz (1, 3, or 5 mA; 20 min) from 4-6 weeks after birth. HI rats undergoing EA had improved motor and memory function, with the greatest improvement after 3 mA EA. The corpus callosum was significantly thicker and showed a significant increase in proliferating astrocytes in the 3 mA EA group. We observed proliferating cells and a greater number of newly developed neurons and astrocytes in the subventricular zone and dentate gyrus of the 3 mA EA group than in those of the HI group. These results suggest that EA promotes functional improvements following neonatal HI assault via the proliferation and differentiation of neural stem cells. This effect was the strongest after 3 mA EA, suggesting that this is the optimal treatment dose.
Project description:BackgroundIschemic stroke is a severe disorder with high incidence, disability rate and mortality. Multiple pathogenesis mechanisms are involved in ischemic stroke, such as inflammation and neuronal cell apoptosis. Protein inhibitor of activated signal transducer and activators of transcription 1 (PIAS1) plays a crucial role in various biological processes, including inflammation. PIAS1 is also downregulated in ischemia-reperfusion injury and involved in the disease processes. However, the role of PIAS1 in cerebral ischemia is unclear.MethodsSprague-Dawley (SD) rats were induced with middle cerebral artery occlusion (MCAO). The role and mechanisms of PIAS1 in ischemic cerebral infarction were explored by Longa test, 2,3,5-triphenyltetrazolium chloride (TTC) staining, Morris water maze (MWM) test, hematoxylin-eosin (HE) staining, quantification of brain water content, reverse transcription-quantitative polymerase chain reaction (RT-qPCR), enzyme-linked immunosorbent assay (ELISA), terminal deoxynucleotidyl transferase deoxyuridine triphosphate (dUTP) nick end labeling (TUNEL), Western blot and immunofluorescence assays.ResultsThe expression of PIAS1 in MCAO-induced rat was declined compared to sham rats. Overexpression of PIAS1 reduced the Longa neurological scores, the percent of infarction area, the pathological abnormality, the escape latency of swimming and the percent of brain water content, and increased the number of platform crossings and time in the target quadrant in the MCAO-induced rats. Besides, overexpression of PIAS1 decreased the MCAO-induced the contents of IL-1β, IL-6 and TNF-α, but further elevated the concentrations of IL-10 in both sera and brain tissues. Moreover, overexpression of PIAS1 reversed the MCAO-induced apoptosis rate and the relative protein level of Bax, cleaved caspase3 and Bcl-2. Overexpression of PIAS1 also reversed the level of proteins involved in NF-κB pathway.ConclusionPIAS1 reduced inflammation and apoptosis, thereby alleviating ischemic cerebral infarction in MCAO-induced rats through regulation NF-κB pathway.
Project description:Diabetic peripheral neuropathy (DPN) is one of the most common complications of diabetes mellitus (DM) and affects over one-third of all patients. Neuropathic pain and nerve dysfunction induced by DM is related to the increase of advanced glycation end products (AGEs) produced by reactive dicarbonyl compounds in a hyperglycemia environment. AGEs induce the expression of pro-inflammatory cytokines via the main receptor (RAGE), which has been documented to play a crucial role in the pathogenesis of diabetic peripheral neuropathy. Electroacupuncture (EA) has been reported to have a positive effect on paralgesia caused by various diseases, but the mechanism is unclear. In this study, we used high-fat-fed low-dose streptozotocin-induced rats as a model of type 2 diabetes (T2DM). Persistent metabolic disorder led to mechanical and thermal hyperalgesia, as well as intraepidermal nerve fiber density reduction and nerve demyelination. EA improved neurological hyperalgesia, decreased the pro-inflammatory cytokines, reduced the generation of AGEs and RAGE, and regulated the glyoxalase system in the EA group. Taken together, our study suggested that EA plays a role in the treatment of T2DM-induced DPN, and is probably related to the regulation of metabolism and the secondary influence on the GLO/AGE/RAGE axis.
Project description:Receptor for advanced glycation end products (RAGE) plays a role in inflammatory reactions. The soluble form of RAGE (sRAGE) acts as a decoy to inhibit interactions of RAGE with advanced glycation end products such as High mobility group box 1 (HMGB1). We have demonstrated that HMGB1 directs Th17 skewing by regulating dendritic cell (DC) functions in a previous study. However, the protective effects of HMGB1 blockade with sRAGE in the development of neutrophilic asthma remain unclear. Here, we showed that allergen challenge decreased expression of sRAGE in a murine model of neutrophilic asthma, correlating well with neutrophil counts and interleukin (IL)-17 production. When HMGB1 signalling was blocked by intratracheal administration of sRAGE before sensitisation, HMGB1 expression, neutrophilic inflammation, and Th17-type responses were reduced significantly. Anti-asthma effects of sRAGE were achieved by inhibition of RAGE and IL-23 expression in airway CD11c+ antigen-presenting cells. Finally, we showed that sRAGE inhibited Th17 polarisation induced by recombinant HMGB1 (rHMGB1)-activated dendritic cells (DCs) in vitro. Adoptive transfer of rHMGB1-activated DCs was sufficient to restore airway inflammation, whereas transfer of rHMGB1 plus sRAGE-activated DCs significantly reduced neutrophilic inflammation. Thus, sRAGE prevents Th17-mediated airway inflammation in neutrophilic asthma at least partly by blocking HMGB1/RAGE signalling in DCs.
Project description:Objective To explore the mechanism of electroacupuncture stimulation of the hand-taiyin meridian in regulating the molecular network of rats with cerebral ischemia-reperfusion injury based on transcriptomics. Methods Male SD rats were randomly divided into sham operation group, model group, and electroacupuncture (EA) group. Middle cerebral artery embolization/reperfusion injury (MCAO/R) was used to establish the model group and EA group. The sham operation group only performed sham operation without modeling and any intervention, and the model group was bound daily. The EA group received electroacupuncture to stimulate the acupoints of hand-taiyin meridian for 14 days. Then, neurological scores, pathomorphological observations, and Tunel staining were performed. Finally, the affected hippocampus of the rat was used for transcriptome sequencing and RT-PCR detection. Results After electroacupuncture intervention in rats, neurological function scores were improved, and neuronal apoptosis was reduced. The results of transcriptomics showed that a total of 1097 differentially expressed genes were obtained, of which 422 were upregulated and 675 were downregulated. The bioinformatics analysis showed that those differentially expressed genes were related to axon development, neuron projection development, neuron projection morphogenesis, plasma membrane cell projection morphogenesis, cell part morphogenesis, notch signaling pathway, long-term potentiation, MAPK signaling pathway, Hedgehog signaling pathway, and so on. The results of RT-PCR showed that Caspase 9 mRNA increased and BDNF, Grin2a, and PlexinD1 mRNA decreased after electroacupuncture intervention (P < 0.05). Conclusion Electroacupuncture intervention on hand-taiyin meridian may reduce neurological function scores, inhibit neuron apoptosis, and enhance neuronal repair neuroreparation in MCAO/R rats, which may be related to the regulation of genes such as Caspase 9, BDNF, Grin2a, and PlexinD1.
Project description:Cerebral ischemia is one of the leading causes of death. Reperfusion is a critical stage after thrombolysis or thrombectomy, accompanied by oxidative stress, excitotoxicity, neuroinflammation, and defects in synapse structure. The process is closely related to the dephosphorylation of actin-binding proteins (e.g., cofilin-1) by specific phosphatases. Although studies of the molecular mechanisms of the actin cytoskeleton have been ongoing for decades, limited studies have directly investigated reperfusion-induced reorganization of actin-binding protein, and little is known about the gene expression of actin-binding proteins. The exact mechanism is still uncertain. The motor cortex is very important to save nerve function; therefore, we chose the penumbra to study the relationship between cerebral ischemia-reperfusion and actin-binding protein. After transient middle cerebral artery occlusion (MCAO) and reperfusion, we confirmed reperfusion and motor function deficit by cerebral blood flow and gait analysis. PCR was used to screen the high expression mRNAs in penumbra of the motor cortex. The high expression of cofilin in this region was confirmed by immunohistochemistry (IHC) and Western blot (WB). The change in cofilin-1 expression appears at the same time as gait imbalance, especially maximum variation and left front swing. It is suggested that cofilin-1 may partially affect motor cortex function. This result provides a potential mechanism for understanding cerebral ischemia-reperfusion.
Project description:BackgroundElectroacupuncture (EA) has been shown to promote functional recovery after cerebral ischemia-reperfusion (I/R) injury. However, the contribution of mitochondrial dynamics to recovery remains unclear. The aim of this study was to investigate whether mitochondrial dynamics are involved in the effects of EA on cerebral I/R injury.MethodsThe rats with cerebral I/R injury were established by the middle cerebral artery occlusion/reperfusion. Subsequently, EA was applied to Baihui (GV20) and Dazhui (GV14) acupoints, with 2 Hz/5 Hz in frequency, 1.0 mA in intensity, 20 min each time, once a day for seven consecutive days. The therapeutic outcomes were assessed by modified neurological severity score (mNSS), 2,3,5-Triphenyte-trazolium chloride (TTC) staining, and hematoxylin-eosin (HE) staining. Mitochondrial morphology was observed under transmission electron microscopy. Adenosine triphosphate (ATP) content and ATP synthases (ATPases) activity were evaluated to measure mitochondrial function using ELISA. Finally, mitochondrial dynamics-related molecules, including dynamin-related protein 1 (Drp1), fission 1 (Fis1), mitofusin 1 (Mfn1), mitofusin 2 (Mfn2), and optic atrophy 1 (OPA1), were detected by Western blot and immunofluorescence staining.ResultsCerebral I/R injury induced neurological dysfunction, cerebral infarction and neuronal injury, all of which were ameliorated by EA. And EA improved mitochondrial morphology and function. Moreover, EA altered the balance of mitochondrial dynamics. Specifically, the data showed a significant decrease in the expression of Drp1 and Fis1, leading to the inhibition of mitochondrial fission. Additionally, Mfn1, Mfn2 and Opa1, which are related to mitochondrial fusion, were effectively promoted after EA treatment. However, sham EA did not show any neuroprotective effects in rats with cerebral I/R injury.ConclusionsIn summary, our study indicates that the balance of mitochondrial dynamics is crucial for EA therapy to treat cerebral I/R injury.