Electroacupuncture regulates histone acetylation of Bcl-2 and Caspase-3 genes to improve ischemic stroke injury.

Background: Imbalances between Bcl-2 and caspase-3 are significant evidence of apoptosis, which is considered an influential factor in rapidly occurring neuronal cell death and the decline of neurological function after stroke. Studies have shown that acupuncture can reduce poststroke brain cell damage via either an increase in Bcl-2 or a reduction in caspase-3 exposure. The current study aimed to investigate whether acupuncture could modulate Bcl-2 and caspase-3 expression through histone acetylation modifications, which could potentially serve as a neuroprotective mechanism. Methods: This study used TTC staining, Nissl staining, Clark neurological system score, and Evans Blue (EB) extravasation to evaluate neurological damage following stroke. The expression of Bcl-2/caspase-3 mRNA was detected by real-time fluorescence quantification of PCR (real-time PCR), whereas the protein expression levels of Bcl-2, Bax, caspase-3, and cleaved caspase-3 were assessed using western blotting. TUNEL staining of the ischemic cortical neurons determined apoptosis in the ischemic cortex. Histone acetyltransferase (HAT) and histone deacetylase (HDAC) activities, along with the protein performance of AceH3, H3K9ace, and H3K27ace, were detected to evaluate the degree of histone acetylation. The acetylation enrichment levels of H3K9 and K3K27 in the Bcl-2/caspase-3 gene were assessed using Chromatin Immunoprecipitation (ChIP) assay. Results: Our data demonstrated that electroacupuncture (EA) exerts a significant neuroprotective effect in middle cerebral artery occlusion (MCAO) rats, as evidenced by a reduction in infarct volume, neuronal damage, Blood-Brain Barrier (BBB) disruption, and decreased apoptosis of ischemic cortical neurons. EA treatment can promote the mRNA and protein expression of the Bcl-2 gene in the ischemic brain while reducing the mRNA and protein expression levels of caspase-3 and effectively decreasing the protein expression levels of Bax and cleaved caspase-3. More importantly, EA treatment enhanced the level of histone acetylation, including Ace-H3, H3K9ace, and H3K27ace, significantly enhanced the occupancy of H3K9ace/H3K27ace at the Bcl-2 promoter, and reduced the enrichment of H3K9ace and H3K27ace at the caspase-3 promoter. However, the Histone Acetyltransferase inhibitor (HATi) treatment reversed these effects. Conclusions: Our data demonstrated that EA mediated the expression levels of Bcl-2 and caspase-3 in MCAO rats by regulating the occupancy of acetylated H3K9/H3K27 at the promoters of these two genes, thus exerting a cerebral protective effect in ischemic reperfusion (I/R) injury.

Characteristics of Electroacupuncture Preconditioning and Treatment on Regulation of the Brain Transcriptome in Rats with Ischemic Stroke.

BACKGROUND: Accumulating evidence suggests that acupuncture may serve as a potent strategy to mitigate the deleterious effects of ischemic stroke on neural tissue. The present investigation delineated the neuroprotective potential of electroacupuncture (EA) administered pre-and post-stroke, with a focus on determining the commonalities and disparities between these two therapeutic approaches in ameliorating ischemic stroke-induced brain injury. The ultimate objective is to inform optimal timing for acupuncture intervention in the clinical management and prevention of stroke. METHODS: The extent of cerebral infarction was quantified with 2,3,5-triphenyltetrazolium chloride staining. The integrity of the blood-brain barrier was assessed by evaluating the extravasation of Evans blue (EB) dye, while neurological function was appraised using the Longa neurological scoring system. RNA sequencing was employed to examine the transcriptomic landscape of ischemic brain tissue, with subsequent bioinformatics annotation of the sequencing data facilitated by Metascape. RESULTS: (1) A notable decrease in the ischemic infarct volume was observed in both the EA-preconditioned plus middle cerebral artery occlusion (MCAO), EA-preconditioned plus middle cerebral artery occlusion (EAM) and MCAO plus EA-treated (MEA) groups, compared to the MCAO group. Furthermore, the decreased leakage of EB and reduction in neurological function impairment scores were evident in the EAM and MEA groups compared with the MCAO group. (2) Relative to the Sham group, the MCAO group exhibited a total of 4798 differentially expressed genes (DEGs), with 67.84% demonstrating an expression fold change (FC) greater than 1.5, and 34.16% exceeding a FC of 2. The EAM and MEA groups displayed 4020 and 1956 DEGs, respectively, compared to the MCAO group. In both groups, more than 55% of DEGs showed an expression FC surpassing 1.5, whereas only approximately 10% exhibited a change greater than 2-fold. Remarkably, EA preconditioning and EA treatment resulted in the reversal of 18.72% and 28.91% of DEGs, respectively, in the MCAO group. (3) The DEGs upregulated in response to ischemic stroke were predominantly implicated in immune inflammatory processes and cellular apoptosis, whereas the downregulated DEGs were associated with neurogenesis and neuronal signal transduction. The MEA-induced upregulated DEGs were primarily involved in neural transmission and metabolic processes, whereas the downregulated DEGs were linked to excessive inflammatory responses to physical and chemical stimuli, as well as cell matrix adhesion chemotaxis. In the context of EAM, the upregulated DEGs were chiefly related to protein biosynthesis, and energy and metabolic processes, whereas the downregulated genes were connected to gene transcriptional activity, synaptic function, and neuronal architecture. CONCLUSIONS: Both preconditioning and post-event treatment with acupuncture demonstrated efficacy in mitigating pathological damage to brain tissue in a rat model of ischemic stroke, albeit with some divergences in their gene targets. The integration of EA preconditioning and treatment may potentially confer enhanced neuroprotection in the clinical management of stroke patients.

Electro-acupuncture treatment ameliorates intestinal inflammatory injury in cerebral ischemia-reperfusion rats via regulating the balance of Treg / γδ T cells.

Electro-acupuncture (EA) has an anti-inflammatory role in ischemic stroke, but whether the protective effect of EA involves the regulation of the intestine barrier and Treg/ γδ T cells is unclear. Cerebral ischemia-reperfusion (I/R) injury was induced by middle cerebral artery occlusion(MCAO) for 2 h followed by reperfusion for 24 h. The rats have treated with EA at the "Baihui" acupoint(GV20). Triphenyl tetrazolium chloride (TTC) staining and Longa neurologic score were performed to evaluate the outcomes after ischemic stroke. Inflammatory factor expression levels in the serum, ischemic hemisphere brain, and small intestine were detected by ELISA or RT-qPCR. Additionally, the morphology change of the small intestine was evaluated by analyzing villus height and smooth muscle thickness. Meanwhile, the expression of tight-junction proteins, including Zonula Occludens-1 (ZO-1), Occludin, and Claudin-1, were detected to evaluate the impact of EA on mucosal permeability in the small intestine. The percentages of regulatory T cells (Tregs) (CD45+CD4+Foxp3+) and γδ T cells (CD45+CD4-γδ T+) were measured to assess the effect of EA on intestinal T cells. EA decreased the brain infarction volume and intestine barrier injury in ischemic stroke rats. At the same time, it effectively suppressed the post-stroke inflammation in the brain and small intestine. More importantly, EA treatment increased the percentage of Tregs in the small intestine while reducing the rate of γδ T cells, and ultimately increased the ratio of Treg/ γδ T cells. These results demonstrated that EA ameliorated intestinal inflammation damage by regulating the Treg/ γδ T cell polarity shift and improving the intestine barrier integrity in rats with I/R injury. This may be one of the mechanisms underlying the anti-ischemic injury effects of acupuncture on stroke.

Electro-acupuncture treatment inhibits the inflammatory response by regulating γδ T and Treg cells in ischemic stroke.

BACKGROUND: Electro-acupuncture (EA) is an effective and safe treatment for ischemic stroke. It is not only capable of reducing cerebral damage but also alleviating intestinal inflammation. However, its mechanism has not been fully elucidated. METHODS: All rats were randomly divided into three experimental groups: the SHAM group, the MCAO group, and the MEA (MCAO+EA) group. Ischemic-reperfusion (I/R) injury was induced by MCAO surgery. Rats in the MEA group were treated with EA stimulation in the "Baihui" acupoint (1 mA, 2/15 Hz, 20 min for each time). The Real-time (RT)-qPCR was used to evaluate the mRNA expression of inflammation factors in the ischemic brain and the small intestine after I/R injury. In addition, our research evaluated the effects of EA on regulatory T cells (Tregs) and γδ T cells in the small intestine and brain via Flow cytometry analysis. Finally, we applied CM-Dil and CFSE injection and explored the potential connections of T cells between the ischemic hemisphere and the small intestine. RESULTS: Our results suggested that EA treatment could significantly reduce the inflammation response in the ischemic brain and small intestine 3 days after I/R injury in rats. To be specific, EA increased the percentage of Tregs in the brain and the small intestine and decreased intestinal and cerebral γδ T cells. Concomitantly, after EA treatment, the percentage of cerebral CD3+TCRγδ+CFSE+ cells dropped from 12.06% to 6.52% compared with the MCAO group. CONCLUSIONS: These findings revealed that EA could regulate the Tregs and γδ T cells in the ischemic brain and the small intestine, which indicated its effect on inhibiting inflammation. And, EA could inhibit the mobilization of intestinal T cells, which may contribute to the protection of EA after ischemic stroke.

Network pharmacology and experimental evidence reveal the protective mechanism of Yi-Qi Cong-Ming decoction on age-related hearing loss.

CONTEXT: Yi-Qi Cong-Ming (YQCM) decoction has been widely used to prevent age-related hearing loss (ARHL), the most prevalent neurodegenerative disease in the elderly. OBJECTIVE: To explore the mechanism of YQCM decoction in the treatment of ARHL. MATERIALS AND METHODS: The chemical constituents of YQCM were screened from the Traditional Chinese Medicine Systems Pharmacology Database. Potential targets of YQCM against ARHL were predicted by DrugBank, GeneCards, and OMIM database. Protein-protein network and enrichment analysis were used for exploring possible molecular mechanisms. Molecular docking and an in vitro model of ARHL by exposing auditory cells with 100 µM H2O2 for 3 h were applied. Cell viability and mitochondrial membrane potential (ΔΨM) were detected by CCK-8 and high-content analysis. γH2AX and cleaved caspase-3 were detected by Western blot. RESULTS: The main compounds have good affinities with hub targets, especially AKT1, PTGS2, and CASP3. GO and KEGG analysis showed that the main biological process and key targets were related to negative regulation of the apoptotic process. H2O2 treatment could reduce the cell viability by 68% and impaired ΔΨM, while 90 µg/mL YQCM pre-treatment could restore the cell viability by 97.45% and increase ΔΨM (2-fold higher). YQCM pre-treatment also reduced γH2AX and cleaved caspase-3 protein levels. CONCLUSIONS: Our study suggested that YQCM prevents ARHL by modulating the apoptosis process in auditory hair cells. Moreover, this study proved that bioinformatics analysis combined with molecular docking and cell model is a promising method to explore other possible pharmacological interventions of ARHL.

Genome-Wide Transcription Analysis of Electroacupuncture Precondition-Induced Ischemic Tolerance on SD Rat With Ischemia-Reperfusion Injury.

Acupuncture promotes the recovery of neurological function by the overall improvement of ischemic brain injury. It is not only regarded as a rehabilitative treatment but also a pretreatment method for stroke. However, its mechanism has not been fully elucidated. In this study, rats were treated with electroacupuncture (EA) at Baihui (GV20) for 30 min/day for 6 days, ahead of conducting cerebral ischemia-reperfusion (I/R) injury. Infarction volume, Evans blue leakage, and neurological deficits were evaluated at 24 h after I/R injury. Then, the ipsilateral ischemic brain was isolated for RNA sequencing (RNA-Seq) to identify molecular consequences. The results showed that EA pretreatment decreased blood-brain barrier (BBB) permeability, reduced brain infarction volume, and improved neurological outcomes. EA pretreatment could upregulate expression of antivirus and immunity activity-associated genes (such as Ifit1, Ifit3, Irf7, and Oasla) and downregulate expression of matrix disruption-associated genes (Col24a1, Col11a1, Col27a1, etc.) in healthy rats. In addition, it could partially reverse or ameliorate genome-wide transcription changes of the ipsilateral ischemic brain. For the first time, this study provides insight into genomic network modulation of a healthy rat with EA treatment and a EA-preconditioned rat under subsequent I/R injury, which is helpful in explaining acupuncture precondition-induced ischemic tolerance of stroke. It also provides new strategies and targets for the prevention of ischemic stroke.

Electroacupuncture improves cardiac function and reduces infarct size by modulating cardiac autonomic remodeling in a mouse model of myocardial ischemia.

BACKGROUND: Sympathetic and parasympathetic nerve remodeling play an important role in cardiac function after myocardial ischemia (MI) injury. Increasing evidence indicates that electroacupuncture (EA) can regulate cardiac function by modulating the autonomic nervous system (ANS), but little is known about its effectiveness on neural remodeling post-MI. OBJECTIVES: To investigate the role of EA in ANS remodeling post-MI. METHODS: Adult male C57/BL6 mice were equally divided into the Control (Ctrl), MI and EA groups after generating the MI model by ligating the left anterior descending (LAD) coronary artery. Echocardiography and 2,3,5-triphenyltetrazolium (TTC) staining were employed to evaluate cardiac function and infarct size after EA treatment for five consecutive days. Serum norepinephrine (NE) levels were measured by ELISA to quantify sympathetic activation. Then, ANS remodeling was detected by immunohistochemistry (IHC), RT-qPCR, and Western blotting. RESULTS: Our preliminary findings showed that EA increased ejection fraction and fractional shortening and reduced infarct area after MI injury. Serum NE levels in the EA group were significantly decreased compared with those in the MI group. IHC staining results demonstrated that the density of growth associated protein (GAP)43 and tyrosine hydroxylase (TH) positive nerve fibers in the EA group were decreased with increased choline acetyltransferase (CHAT) and vesicular acetylcholine transporter (VACHT). Meanwhile, the results verified that mRNA and protein expression of GAP43 and TH were significantly inhibited by EA treatment in the MI mice, accompanied by elevated CHAT and VACHT. CONCLUSIONS: EA treatment could improve cardiac function and reduce infarct size by modulating sympathetic and parasympathetic nerve remodeling post-MI, thus helping the cardiac ANS reach a new balance to try to protect the heart from further possible injury.

Astragali Radix Isoflavones Synergistically Alleviate Cerebral Ischemia and Reperfusion Injury Via Activating Estrogen Receptor-PI3K-Akt Signaling Pathway.

Isoflavones are major neuroprotective components of a medicinal herb Astragali Radix, against cerebral ischemia-reperfusion injury but the mechanisms of neuroprotection remain unclear. Calycosin and formononetin are two major AR isoflavones while daidzein is the metabolite of formononetin after absorption. Herein, we aim to investigate the synergistic neuroprotective effects of those isoflavones of Astragali Radix against cerebral ischemia-reperfusion injury. Calycosin, formononetin and daidzein were organized with different combinations whose effects observed in both in vitro and in vivo experimental models. In the in vitro study, primary cultured neurons were subjected to oxygen-glucose deprivation plus reoxygenation (OGD/RO) or l-glutamate treatment. In the in vivo study, rats were subjected to middle cerebral artery occlusion to induce cerebral ischemia and reperfusion. All three isoflavones pre-treatment alone decreased brain infarct volume and improved neurological deficits in rats, and dose-dependently attenuated neural death induced by l-glutamate treatment and OGD/RO in cultured neurons. Interestingly, the combined formulas of those isoflavones revealed synergistically activated estrogen receptor (estrogen receptors)-PI3K-Akt signaling pathway. Using ER antagonist and phosphatidylinositol 3-kinase (PI3K) inhibitor blocked the neuroprotective effects of those isoflavones. In conclusion, isoflavones could synergistically alleviate cerebral ischemia-reperfusion injury via activating ER-PI3K-Akt pathway.

Acupuncture for Cancer-Induced Bone Pain in Animal Models: A Systemic Review and Meta-Analysis.

BACKGROUND: Cancer-induced bone pain (CIBP) is a highly prevalent symptom, which afflicts vast majority of patients who suffer from cancer. The current treatment options failed to achieve satisfactory effect and the side effects were prominent. Recent randomized controlled trials (RCTs) of animal demonstrate the benefit of acupuncture for CIBP. We sought to determine if the pooled data from available RCTs supports the use of acupuncture for CIBP. METHODS: A literature search for randomized controlled trials was conducted in six electronic databases from inception to May 31, 2019. Meta-analysis was performed with Review Manager 5.3 software; the publication bias was assessed by Stata 12.0 software. We used random effects model for pooling data because heterogeneity is absolute among studies to some extent. RESULTS: Twenty-four trials were included in the review, of which 12 trials provided detailed data for meta-analyses. Preliminary evidence indicates that compared to wait list/sham group, acupuncture was effective on increasing paw withdrawal threshold (PWT) and paw withdrawal latency (PWL). Compared to medicine, acupuncture was less effective on PWT, but as effective as medicine on PWL. Acupuncture can reinforce medicine's effect on PWT and PWL. Compared to the control group, acupuncture was superior to increase body weight (BW), decrease spinal cord glial fibrillary acidic protein (GFAP), and interleukin-1ß (IL-1ß). Furthermore, some studies showed acupuncture delay or partially reverse morphine tolerance. Three studies found acupuncture has no effect on PWT, but 2 of them found acupuncture could enhance small dose of Celebrex's effect on CIBP. CONCLUSIONS: Acupuncture was superior to wait list/sham acupuncture on increasing PWT and has no less effect on increasing PWL compared to medicine; acupuncture improved the efficacy of drugs, increased the CIBP animals' body weight, and decreased their spinal cord GFAP and IL-1ß. High-quality studies are necessary to confirm the results.

Proteomics-Guided Study on Buyang Huanwu Decoction for Its Neuroprotective and Neurogenic Mechanisms for Transient Ischemic Stroke: Involvements of EGFR/PI3K/Akt/Bad/14-3-3 and Jak2/Stat3/Cyclin D1 Signaling Cascades.

Buyang Huanwu Decoction (BHD), a classic traditional Chinese medicine (TCM) formula, has been used for recovering neurological dysfunctions and treating post-stroke disability in China for 200 years. In the present study, we investigated the effects of BHD on inhibiting neuronal apoptosis, promoting proliferation and differentiation of neural stem cells (NSCs) and neurite formation and enhancing learning and memory functional recovery in an experimental rat ischemic stroke model. BHD significantly reduced infarct volume and decreased cell apoptosis in the ischemic brain. BHD enhanced neuronal cell viability in vitro. BHD dose-dependently promoted the proliferation of NSCs in ischemic rat brains in vivo. Moreover, BHD promoted neuronal and astrocyte differentiation in primary cultured NSCs in vitro. Water maze test revealed that BHD promoted the recovery of learning function but not memory functions in the transient ischemic rats. We then investigated the changes of the cellular signaling molecules by using two-dimension (2D) gel electrophoresis and focused on the PI3K/Akt/Bad and Jak2/Stat3/cyclin D1signaling pathway to uncover its underlying mechanisms for its neuroprotective and neurogenetic effects. BHD significantly upregulated the expression of p-PI3K, p-Akt, and p-Bad as well as the expression of p-Jak, p-Stat3, and cyclin D1 in vitro and in vivo. In addition, BHD upregulated Hes1 and downregulated cav-1 in vitro and in vivo. Taken together, these results suggest that BHD has neuroprotective effects and neurogenesis-promoting effects via activating PI3K/Akt/Bad and Jak2/Stat3/Cyclin D1 signaling pathways. Graphical Abstract Buyang Huanwu Decoction (BHD) activates the PI3K-AKT-BAD pathway in the ischemic brain for neuroprotection. BHD also activates JAK2/STAT3/Cyclin D1 signaling cascades for promoting neurogenesis in the hippocampus of post-ischemic brains. Moreover, BHD inhibits the expression of caveolin-1 and increases the expression of HES1 for promoting neuronal differentiation. The neuroprotective and neurogenesis-promoting effects in the hippocampus of post-ischemic brains promote learning ability.

[Effects of electroacupuncture pretreatment on cardiac function and immune inflammatory response in mice with acute myocardial ischemia].

OBJECTIVE: To observe the effects of electroacupuncture (EA) pretreatment on the cardiac ejection fraction (EF), the number of macrophages in spleen and heart, and the expression of nucleotide-binding oligomerization domain-like receptor protein 3 (NLRP3) and interleukin-1ß (IL-1ß) in myocardium in mice with acute myocardial ischemia, and to explore the possible mechanism of EA pretreatment on promoting myocardial protection. METHODS: A total of 30 male C57BL/6J mice were randomly divided into a control group, a model group and an EA pretreatment group, 10 rats in each group. The acute myocardial ischemia model was established by ligating the left anterior descending branch of the coronary artery in the model group and EA pretreatment group, while threading but no ligating at left anterior descending branch of the coronary artery was applied in the control group. In the EA pretreatment group, mice were intervented with EA at bilateral "Neiguan" (PC 6), disperse-dense wave, frequency of 2 Hz/15 Hz, intensity of 2 mA; each EA treatment last for 20 min, once a day, and 3-day treatment was given before model establishment. The EF value was evaluated by ultrasonic cardiogram; the number of macrophages in spleen and heart was measured by flow cytometry; the expression level of NLRP3 and IL-1ß in myocardium was measured by Western blot. RESULTS: Compared with the control group, the EF value was decreased in the model group (P<0.001), the number of macrophages in the heart and spleen was increased (P<0.001), and the expression level of NLRP3 and IL-1ß in the myocardium was increased (P<0.001, P<0.01). Compared with the model group, the EF value was increased in the EA pretreatment group (P<0.01), the number of macrophages in the heart and spleen was decreased (P<0.01), and the expression level of NLRP3 and IL-1ß in the myocardium was decreased (P<0.01, P<0.05). CONCLUSION: EA pretreatment could reduce the number of macrophages in spleen and heart, down-regulate the expression of NLRP3 and IL-1ß in myocardial tissue in mice with acute myocardial ischemia, which could relieve the local inflammatory response and achieve the myocardial protective effect.

Electroacupuncture Relieves Pain and Attenuates Inflammation Progression Through Inducing IL-10 Production in CFA-Induced Mice.

The therapeutic effect of electroacupuncture (EA) on inflammatory pain has been well recognized clinically, but the mechanism is unclear. Interleukin-10 (IL-10), which is produced by regulatory T (Treg) cell, is a key anti-inflammatory cytokine for relieving inflammatory pain. Therefore, the aim of this study is to investigate whether EA could inhibit CFA-induced pain and attenuate inflammation progression by regulating the activation of immunocyte and inducing the expression of IL-10. In this study, mice were treated with EA (2/100 Hz, 2 mA) for five consecutive days after 1 day of CFA injection. The behavioral tests were measured and analyzed after the daily EA treatment; then, hind paw, spinal cord, and spleen tissues were prepared for assessment. The results showed that EA treatment significantly increased the mechanical threshold and thermal latency after CFA injection and boosted the expression of IL-10 in paw and spinal cord tissues. EA treatment promoted Treg cells; suppressed macrophage and neutrophils cells; reduced the expression of IL-1ß, NLRP3, and TNF-α; and ultimately relieved inflammatory pain. The findings suggested that the analgesic and anti-inflammatory effect of EA treatment could be partially associated with suppression of pro-inflammatory cytokines mediated by induction of IL-10.

Electroacupuncture preconditioning attenuates acute myocardial ischemia injury through inhibiting NLRP3 inflammasome activation in mice.

AIMS: Electro-acupuncture pretreatment (EAP) plays a protective role in myocardial ischemia (MI) injury. However, the underlying mechanism remains unclear. A growing body of evidence suggests postinfarction inflammatory response directly affects the remodeling of ventricular function. The purpose of this study was to investigate whether EAP alleviates MI through NLRP3 inflammasome inhibition. MATERIALS AND METHODS: We constructed an AMI model by ligating the left anterior descending (LAD) coronary artery after 3 days of EAP with C57BL/6 mice. Echocardiography and TTC staining were employed to evaluate cardiac function and infarct size after 24 h of ischemia. HE staining and immunohistochemistry were employed to determine inflammatory level. Then, inflammasome activation was detected by western blotting, and macrophage polarization and neutrophil infiltration were observed by flow cytometry. KEY FINDINGS: Our preliminary findings showed that EAP reduced the infarct area and increased fractional shortening (FS) and ejection fraction (EF) and decreased the degree of inflammation after AMI injury. Meanwhile, EAP inhibited the expression of NLRP3, cleaved caspase-1 and IL-1ß in ischemia myocardial tissue, companied by inhibiting the expression of F4/80+, CD11b+, CD206low macrophages and activated M2 macrophage, and decreasing Ly-6G+CD11b+ neutrophils in ischemia myocardial and spleen tissue. SIGNIFICANCE: EAP inhibits the activation of NLRP3 inflammasome, promotes M2 polarization of macrophages and reduces the recruitment of neutrophils in damaged myocardium, thereby decreases the infarct size and improves the cardiac function.

[Effects of electroacupuncture on food intake and expression of lipid receptors of taste buds in the tongue and hippocampus in obese rats].

OBJECTIVE: To observe the effect of electroacupuncture (EA) on food intake, body weight, number of taste bud cells and the expression of lipid taste bud receptor (CD36), Gα-gustducin, post-synaptic density protein 95 (PSD95) and neurofilament light chain (NFL) proteins in the tongue or hippocampus in obese rats, so as to explore its mechanism underlying reducing body weight. METHODS: A total of 30 male SD rats were randomly divided into control, model and EA groups (n=10 in each group, 5 rats for H.E. staining and immunohistochemistry, and 5 for Western blot). The obesity model was established by feeding the rats with high fat diet for 11 weeks. Following successful modeling, EA (2 Hz/15 Hz, 1.0-1.2 mA) was applied to "Tianshu" (ST25) for 30 min, once a day, 5 times/week for 5 weeks. The body length, body weight and maximum daily food consumption were recorded every day, followed by calculating the lee's index. Histopathological changes of the circumvallate papillae (CVP) and number of taste bud cells and CD36 were detected by HE staining and immunohistochemistry (IHC), separately. The expression levels of CD36, PSD95 and NFL proteins in the hippocampus were detected by Western blot. RESULTS: The body weight, Lee's index and daily food consumption were significantly higher in the model group than in the control group (P<0.01), and were significantly lowered after EA intervention in comparison with the model group (P<0.01), suggesting an improvement of obesity. H.E. staining displayed that the CVP area and the number of taste bud cells were obviously decreased in the model group in contrast to the control group (P<0.01), and were notably increased in the EA group in contrast to the model group (P<0.05, P<0.01). IHC and Western blot showed that the expression levels of CD36 in the tongue and hippocampus were obviously up-regulated in the model group relevant to the control group (P<0.01, P<0.05), and considerably down-regulated in the EA group relevant to the model group (P<0.05, P<0.01). The expression levels of Gα-gustducin in the tongue, and PSD95 and NFL in the hippocampus were remarkably decreased in the model group relevant to the control group (P<0.01, P<0.05), and significantly increased in the EA group relevant to the model group (P<0.01, P<0.05). CONCLUSION: EA can reduce daily food consumption and body weight in obese rats, which is associated with its effects in down-regulating the expression of CD36 in taste buds and hippocampus, and up-regulating the expression of Gα-gustducin in the tongue, and PSD95 and NFL proteins in the hippocampus. It suggests that EA may regulate the feeding behavior of obese rats by influencing the cognitive memory mechanism involved in CD36 in hippocampus.

Effectiveness of Acupuncture in Treatment of Simple Obesity in Animal Models: A Systematic Review and Meta-Analysis.

BACKGROUND: Simple obesity has become a global risk to health of human beings. Acupuncture, as one of traditional Chinese medicine therapies, has been widely used in obesity treatment in recent years. However, the individual heterogeneity which makes acupuncture's efficiency unstable leads to some controversy. So more evidence-based results are necessary to judge the effectiveness of acupuncture in treatment of simple obesity. Compared with clinical trials, animal experiments are controllable, and the underlying mechanism is more likely to be explored. Last but not the least, more and more experimental studies on acupuncture for animal obesity have been published. Therefore, we conducted the systematic review and meta-analysis to evaluate the effectiveness of acupuncture in treating simple obesity in animal experiments. METHODS: Randomized Controlled Trials (RCTs) on acupuncture for simple obesity animal models were searched from six databases: PubMed, MEDLINE, CNKI, VIP, WanFang Date, and CMB from inception to February 2017 and updated on April 12, 2019. RevMan 5.3 software was used for meta-analysis. Treatment effects were summarized as relative risk (RR) and Standard mean difference (SMD) with 95% of confidence interval (CI). RESULTS: A total of 108 trials involving 5731 rats were included. Meta-analysis showed that acupuncture had better effect on reducing weight (SMD -2.60, 95%CI: -2.93 to -2.26, p<0.00001) and Lee's index (SMD -2.62, 95%CI:-3.18 to -2.06, p<0.00001) compared with control group. However, the methodological quality of included studies was generally poor. Details of blinding were not reported in most studies. In spite of high heterogeneity being observed on the merged data, sensitivity analysis using the leave-one-out approach, subgroup analysis based on different acupuncture techniques, and rat strains and meta-regression all failed to find the sources of heterogeneity. The asymmetric funnel plot suggested publication bias. Besides, adverse events were not reported in any reports. CONCLUSIONS: Our review provided positive evidence of acupuncture for simple obesity. Unfortunately, none of the firm conclusions can be drawn due to methodological flaws, high heterogeneity, and publication bias. More high-quality trials are needed in future to get objective conclusions.

Electroacupuncture Reduces Body Weight by Regulating Fat Browning-Related Proteins of Adipose Tissue in HFD-Induced Obese Mice.

Objective: This study investigated the influence of electroacupuncture (EA) and its potential underlying mechanisms on adipose tissue in obese mice. Methods: Three-week-old male C56BL/6 mice were randomly divided to feed or not to feed high-fat diet (HFD), named HFD group and chow diet (CD) group, respectively. After 12 weeks, CD and HFD mice were randomly divided into two groups, respectively, to receive or not receive EA for 4 weeks. Body weight (BW) was monitored. Intraperitoneal glucose tolerance test and metabolic chamber recordings were performed. Blood samples and adipose tissue were collected for the analysis of leptin, triglyceride levels, and fat browning-related proteins. Results: EA significantly reduced food intake, BW, and white adipose tissue (WAT)/BW ratio; decreased the adipocyte size and serum concentrations of triglyceride (TG) and cholesterol; and increased oxygen consumption in HFD mice. Compared with the CD mice, the HFD mice had elevated fasting serum glucose level and impaired glucose tolerance; however, these parameters were decreased by EA treatment. Meanwhile, EA promoted the protein and mRNA expressions of UCP1, PRDM16, and PGC-1α in adipose tissue, and activated sympathetic nerves via p-TH, A2AR, and ß3AR in white adipose tissue. Conclusions: EA reduced food intake, BW, TG, and cholesterol, and improved glucose tolerance in HFD mice. This ameliorative effect of EA on obesity-related symptoms associated with its promoted adipose tissue plasticity via activating sympathetic nerves.

[Effect of electroacupuncture on myocardial infarct size and expression of inflammatory cytokines and sympathetic-active substances in the myocardial ischemic tissue of rats].

OBJECTIVE: To investigate the effect of electroacupuncture (EA) on the expression of interleukin-8 (IL-8), interleukin-10 (IL-10), tyrosine hydroxylase (TH), ß3-adrenergic receptor (ß3AR), and endothelial nitric oxide synthase (eNOS) in myocardial tissue in ischemic myocardial injury rats, so as to reveal its underlying mechanisms in myocardial protection via anti-inflammation and sympathetic nerve remodeling. METHODS: A total of 48 male Sprague-Dawley rats were randomly divided into sham-operation (sham, n＝9), sham ＋EA (n＝9), model (n＝15) and EA (n＝15) groups. The myocardial ischemia (MI) model was established by ligation of the left anterior descending branch of the left coronary artery. EA (2 Hz/15 Hz，1.5－2 mA) was applied to bilateral "Neiguan" (PC6) for 30 min, once daily for 4 days. The myocardial infarct size was detected by 2, 3, 5 triphenyltetrazolium chloride (TTC) staining, myocardial histopathological changes and inflammatory infiltration were assessed by H．E. staining, and the expression of IL-8, IL-10, TH, ß3AR, and eNOS in the myocardium was determined by using Western blot. RESULTS: Compared with the sham group, a marked myocardial infarction was found in the left ventricle tissue, accompanied with disordered arrangement of myocardial fibers and higher degree of inflammatory cell infiltration, and increased expression of IL-8, TH, ß3AR and eNOS in the myocardium in the model group (P<0.01), but without significant change in the expression of IL-10 (P>0.05). After EA intervention and in comparison with the model group, the myocardial infarct size was significantly reduced (P<0.01), the severity of inflammatory cell infiltration and disordered arrangement of myocardial fibers were relieved, and the expression of IL-10 and eNOS proteins were significantly up-regulated (P<0.05), and the markedly up-regulated expression of IL-8, TH, and ß3AR were significantly suppressed in the EA group (P<0.01)．. CONCLUSION: EA intervention can reduce the myocardial infarct size (protective effect) in MI rats possibly by reducing inflammatory reaction and sympathetic nerve remodeling.

[Effects of electroacupuncture on sympathetic nerve-related substance in myocardial tissue in mice with myocardial ischemia].

OBJECTIVE: To observe the effects of electroacupuncture (EA) on sympathetic nerve-related substance in myocardial tissue in mice with myocardial ischemia (MI), and to explore its possible mechanism. METHODS: Thirty adult male C57BL/6 mice were randomly divided into a sham operation group, a model group and an EA group, 10 mice in each one. The model of MI was established in the model group and EA group by ligating the left anterior descending branch of coronary artery. The mice in the sham operation group were not treated with ligating at left anterior descending branch of coronary artery, but the remaining procedure was similar with the model group. The mice in the EA group were treated with EA at "Neiguan" (PC 6) with 2 Hz/100 Hz of frequency and 2 mA of intensity, 20 min per treatment, once a day for totally 5 days. No EA was given for model group and sham operation group. The electrocardiogram was recorded and △ST value was calculated to evaluate the model. TTC staining was applied to evaluate the infarct size. Immunohistochemical (IHC) method was applied to evaluate the positive nerve fiber density in myocardial tissue. Western blot method was applied to test the protein expression levels of neuregulin-1 (NRG-1), tyrosine hydroxylase (TH), growth associated protein-43 (GAP-43). RESULTS: The electrocardiogram (lead II) results indicated compared with the sham operation group, the S-T segments in the model group and EA group were increased obviously (both P<0.01), indicating the MI model was established successfully. The TTC staining results indicated compared with sham operation group, the infarction size was significantly increased in the model group (P<0.01); compared with the model group, the infarction size in the EA group was significantly reduced (P<0.01). The IHC results indicated compared with the sham operation group, the positive nerve fiber density in myocardial was increased in the model group (P<0.01); compared with the model group, the positive nerve fiber density in myocardial was reduced in the EA group (P<0.05). The Western blot results indicated compared with the sham operation group, the expression levels of TH, NRG-1 and GAP-43 were significantly increased in the model group (P<0.01); compared with the model group, the expression level of TH and GAP-43 were significantly reduced (P<0.01) and that of NRG-1 was increased in the EA group (P<0.05). CONCLUSION: EA could increase the expression of NRG-1 and reduce the expression of TH and GAP-43 in myocardial tissues in MI mice, which could suppress sympathetic nerve hyperexcitability after infarction to achieve myocardial protection effect.

[Effects of electroacupuncture on inflammatory response of cardiac muscle tissue in mice with acute myocardial ischemia].

OBJECTIVE: To observe the effects of electroacupuncture (EA) on inflammatory reaction of acute myocardial ischemia (MI) in mice, and to explore its action mechanism. METHODS: Forty adult male C57BL/6 mice were randomly divided into a control group, a sham operation group, a model group and an EA group, 10 mice in each one. The model was established in the model group and EA group by ligating the left anterior descending branch of coronary artery. The mice in the EA group were treated with EA at "Neiguan" (PC 6) with 2 mA of intensity and 2 Hz /100 Hz of frequency; EA was given 30 min per treatment, once a day for totally 5 days. The mice in the control group and model group were treated with immobilization and no EA was given. The mice in the sham operation group were not treated with ligating at the left anterior descending branch of coronary artery, but the remaining procedure was identical to the model group. The electrocardiogram was recorded and △ST was calculated to evaluate the model. TTC and HE staining methods were applied to evaluate the infarct size and pathologic change of myocardial tissue, respectively. Western blot method was applied to test the protein expression levels of tumor necrosis factor-α (TNF-α), nuclear factor-κB p65 (NF-κB p65), interleukin-1ß (IL-1ß) and interleukin-8 (IL-8). RESULTS: Compared with the sham operation group, the S-T segments in the model group and EA group were increased obviously after modeling (both P<0.01), indicating the MI model was established successfully. The TTC and HE staining results indicated, compared with the sham operation group, the model group had larger infarction size (P<0.01), more myocardial fibers injury and inflammatory infiltration; compared with the model group, the infarction size of the EA group was significantly reduced (P<0.01), and the myocardial fibers injury and inflammatory infiltration were improved. Compared with the control group, the protein expression levels in the sham operation group were similar (all P>0.05); compared with the sham operation group, the expression levels of TNF-α, NF-κB p65, IL-1ß and IL-8 were significantly increased in the model group (P<0.01, P<0.05); compared with the model group, the expression levels of TNF-α, NF-κB p65, IL-1ß and IL-8 were significantly reduced in the EA group (all P<0.05). CONCLUSION: EA might reduce the protein expression levels of TNF-α, NF-κB p65, IL-1ß and IL-8 in cardiac muscle tissue to inhibit inflammatory reaction and achieve myocardial protective effect in mice with acute myocardial ischemia.

[Effects of electroacupuncture on the expression of adenosine receptors in the heart tissue of myocardial ischemia rats].

OBJECTIVE: To explore the impact of electroacupuncture (EA) on the protein expression of adenosine receptors in the heart of the rats with myocardial ischemia (MI). METHODS: Thirty healthy male SD rats were divided randomly into a control group (n=6), a model group (n=12) and an EA group (n=12). We ligated the left anterior descending artery (LAD) for MI model in the model group and EA group, and exposed the heart after opening the chest without ligation in the control group. EA, 2 Hz /15 Hz and 1.5-2 mA, was applied at bilateral"Neiguan"(PC 6) in the EA group for 20 min, once a day for continuous 5 days. No intervention except grabbing and fixation was used in the control group and model group. We applied 2% TTC staining to observe the infarct size of myocardium, colorimetry to analyze serum lactic dehydrogenase (LDH), creatine kinase (CK), creatine kinase isoenzyme (CK-MB), radio-immunity assessment to detect cardiac troponin T (cTnT), Western blot to evaluate the adenosine A1 receptor (A1AR), A2aAR, A2bAR and A3AR. RESULTS: After treatment, myocardial infarction of (27.56±3.24)% was obvious in the model group; the myocardial infarction in the EA group was (21.04±3.61)%, with statistical significance (P<0.05). The expressions of serum LDH, CK, CK-MB and cTnT levels in the model group increased compared with those in the control group (all P<0.01), and the expressions of LDH, CK, CK-MB and cTnT levels in the EA group decreased compared with those in the model group (P<0.05，P<0.01). The A1AR expression in the model group was not different from that in the control group (P>0.05), and A2aAR、A2bAR、A3AR expressions decreased (P<0.05, P<0.01). A2aAR and A2bAR expressions in the EA group increased compared with those in the model group (both P<0.01), and there was no statistical significance between A1AR and A3AR expressions (both P>0.05). . CONCLUSION: EA may achieve cardioprotective effect by regulating the expressions of A2aAR and A2bAR in myocardial tissue, which induce the corresponding signal cascade for reducing myocardial infarction area.