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.

Remodelling the inguinal adipose sensory system to switch on the furnace: Electroacupuncture stimulation induces brown adipose thermogenesis.

Brown and white adipose tissue mediate thermogenesis through the thermogenetic centre of the brain, but safe methods for activating thermogensis and knowledge of the associated molecular mechanisms are lacking. We investigated body surface electroacupuncture stimulation (ES) at ST25 (targeted at the abdomen) induction of brown adipose thermogenesis and the neural mechanism of this process. Inguinal white adipose tissue (iWAT) and interscapular brown adipose tissue (iBAT) were collected and the thermogenic protein expression levels were measured to evaluate iBAT thermogenesis capacity. The thermogenic centre activating region and sympathetic outflow were evaluated based on neural electrical activity and c-fos expression levels. iWAT sensory axon plasticity was analysed with whole-mount adipose tissue imaging. ES activated the sympathetic nerves in iBAT and the c-fos-positive cells induced sympathetic outflow activation to the iBAT from the medial preoptic area (MPA), the dorsomedial hypothalamus (DM) and the raphe pallidus nucleus (RPA). iWAT denervation mice exhibited decreased c-fos-positive cells in the DM and RPA, and lower recombinant uncoupling orotein 1 peroxisome proliferator-activated receptor, ß3-adrenergic receptor, and tyrosine hydroxylase expression. Remodelling the iWAT sensory axons recovered the signal from the MPA to the RPA and induced iBAT thermogenesis. The sympathetic denervation attenuated sensory nerve density. ES induced sympathetic outflow from the thermogenetic centres to iBAT, which mediated thermogenesis. iWAT sensory axon remodelling induced the MPA-DM-RPA-iBAT thermogenesis pathway.

Electroacupuncture stimulation ameliorates cognitive impairment induced by long-term high-fat diet by regulating microglial BDNF.

Long-term high-fat diet (HFD) in adolescents leads to impaired hippocampal function and increases the risk of cognitive impairment. Studies have shown that HFD activates hippocampal microglia and induces hippocampal inflammation, which is an important factor for cognitive impairment. Electroacupuncture stimulation (ES), a nerve stimulation therapy, is anti-inflammatory. This study explored its therapeutic potential and mechanism of action in obesity-related cognitive impairment. 4-week-old C57 mice were given either normal or HFD for 22 weeks. At 19 weeks, some of the HFD mice were treated with ES and nigericin sodium salt. The cognitive behavior was assessed through Morris water maze test at 23 weeks. Western blotting was used to detect the expression levels of pro-inflammatory molecules IL-1ß and IL-1R, synaptic plasticity related proteins synaptophysin and Postsynaptic Density-95 (PSD-95), and apoptotic molecules (Caspase-3 and Bcl-2), in the hippocampus. The number, morphology, and status of microglia, along with the brain-derived neurotrophic factor（BDNF） content, were analyzed using immunofluorescence. ES treatment improved cognitive deficits in HFD model mice, and decreased the expressions of microglial activation marker, CD68, and microglial BDNF. Inhibition of proinflammatory cytokine, IL-1ß, and IL-1R promoted PSD-95 and synaptophysin expressions. Peripheral NLRP3 inflammasome agonist injections exacerbated the cognitive deficits in HFD mice and promoted the expressions of IL-1ß and IL-1R in the hippocampus. The microglia showed obvious morphological damage and apoptosis. Collectively, our findings suggest that ES inhibits inflammation, regulates microglial BDNF, and causes remodeling of hippocampal function in mice to counteract obesity-like induced cognitive impairment. Overexcitation of peripheral inflammasome complexes induces hippocampal microglia apoptosis, which hinders the effects of ES.

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.

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.

STAT3 but Not STAT5 Contributes to the Protective Effect of Electroacupuncture Against Myocardial Ischemia/Reperfusion Injury in Mice.

Electroacupuncture (EA) can help reduce infarct size and injury resulting from myocardial ischemia/reperfusion (I/R); however, the underlying molecular mechanism remains unknown. We previously reported that STAT5 plays a critical role in the cardioprotective effect of remote ischemic preconditioning (RIPC). Here, we assessed the effects of electroacupuncture pretreatment (EAP) on myocardial I/R injury in the presence and/or absence of Stat5 in mice and investigated whether EAP exerts its cardioprotective effects in a STAT5-dependent manner. Adult Stat5 fl/fl and Stat5-cKO mice were exposed to EAP at Neiguan (PC6) for 7 days before the induction of I/R injury by left anterior descending (LAD) coronary artery ligation. The myocardial infarct size (IS), area at risk, and apoptotic rate of cardiomyocytes were detected. RT-qPCR and western blotting were used to measure gene and protein expression, respectively, in homogenized heart tissues. RNA-seq was used to identify candidate genes and pathways. Our results showed that EAP decreased IS and the rate of cardiomyocyte apoptosis. We further found that STAT5 was activated by EAP in Stat5 fl/fl mice but not in Stat5-cKO mice, whereas the opposite was observed for STAT3. Following EAP, the levels of the antiapoptotic proteins Bcl-xL, Bcl-2, and p-AKT were increased in the presence of Stat5, while that of interleukin 10 (IL-10) was increased in both Stat5 fl/fl and Stat5-cKO. The gene expression profile in heart tissues was different between Stat5 fl/fl and the Stat5-cKO mice with EAP. Importantly, the top 30 DEGs under EAP in the Stat5-cKO mice were enriched in the IL-6/STAT3 signaling pathway. Our results revealed for the first time that the protective effect of EAP following myocardial I/R injury was attributable to, but not dependent on, STAT5. Additionally, we found that EAP could activate STAT3 signaling in the absence of the Stat5 gene, and could also activate antiapoptotic, survival, and anti-inflammatory signaling pathways.

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.

Epigenetic Landscape Changes Due to Acupuncture Treatment: From Clinical to Basic Research.

Acupuncture has been widely used for treating diseases since the ancient days in China, but the mechanism by which acupuncture exerts such powerful roles is unclear. Epigenetics, including DNA methylation, histone modification, and post-transcriptional regulation of miRNAs, is the study of heritable changes in gene expression that do not include DNA sequence alterations. Epigenetics has become a new strategy for the basic and clinical research of acupuncture in the last decade. Some investigators have been trying to illustrate the mechanism of acupuncture from an epigenetics perspective, which has shed new lights on the mechanisms and applications of acupuncture. Moreover, the introduction of epigenetics into the regulatory mechanism in acupuncture treatment has provided more objective and scientific support for acupuncture theories and brought new opportunities for the improvement of acupuncture studies. In this paper, we reviewed the literatures that has demonstrated that acupuncture could directly or indirectly affect epigenetics, in order to highlight the progress of acupuncture studies correlated to epigenetic regulations. We do have to disclose that the current evidence in this review is not enough to cover all the complex interactions between multiple epigenetic modifications and their regulations. However, the up-to-date results can help us to better understand acupuncture's clinical applications and laboratory research.

[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.

Electroacupuncture Regulates Inguinal White Adipose Tissue Browning by Promoting Sirtuin-1-Dependent PPARγ Deacetylation and Mitochondrial Biogenesis.

Background: Previous studies had suggested that electroacupuncture (EA) can promote white adipose tissue (WAT) browning to counter obesity. But the mechanism was still not very clear. Aim: In this study, we aim to study the effect of EA on promoting inguinal WAT (iWAT) browning and its possible mechanism. Method: Three-week-old rats were randomly divided into a normal diet (ND) group and a high-fat diet (HFD) group. After 10 weeks, the HFD rats were grouped into HFD + EA group and HFD control group. Rats in the EA group were electro-acupunctured for 4 weeks on Tianshu (ST25) acupoint under gas anesthesia with isoflurane, while the rats in HFD group were under gas anesthesia only. Body weight and cumulative food intake were monitored, and H&E staining was performed to assess adipocyte area. The effect of EA on WAT was assessed by qPCR, immunoblotting, immunoprecipitation and Co-immunoprecipitation. Mitochondria were isolated from IWAT to observe the expression of mitochondrial transcription factor A (TFAM). Results: The body weight, WAT/body weight ratio and cumulative food consumption obviously decreased (P < 0.05) in the EA group. The expressions of brown adipose tissue (BAT) markers were increased in the iWAT of EA rats. Nevertheless, the mRNA expressions of WAT genes were suppressed by 4-week EA treatment. Moreover, EA increased the protein expressions of SIRT-1, PPARγ, PGC-1α, UCP1 and PRDM16 which trigger the molecular conversion of iWAT browning. The decrease of PPARγ acetylation was also found in EA group, indicating EA could advance WAT-browning through SIRT-1 dependent PPARγ deacetylation pathway. Besides, we found that EA could activate AMPK to further regulate PGC-1α-TFAM-UCP1 pathway to induce mitochondrial biogenesis. Conclusion: In conclusion, EA can remodel WAT to BAT through inducing SIRT-1 dependent PPARγ deacetylation, and regulating PGC-1α-TFAM-UCP1 pathway to induce mitochondrial biogenesis. This may be one of the mechanisms by which EA affects weight loss.

[Effect of Electroacupuncture on Adenosine Receptor Expression in White Adipose Tissue of Diet-induced Obese Mice].

OBJECTIVE: To investigate the effects of electroacupuncture (EA) on the expression of adenosine receptor (AR) in the white adipose tissue (WAT) of diet-induced obese (DIO) mice, so as to reveal a peripheral mechanism of EA underlying improvement of body weight. METHODS: Forty three-week-old C 57 BL/6 male mice were divided into normal diet group (n=12) and high fat diet group (n=28) randomly, and fed by normal diet and high fat diet for 12 weeks, respectively. In the high fat diet group, mice with body weight over 20% heavier than that of the normal diet group were considered as obese mice. The normal diet mice and the obese mice were divided into normal group (CD, n=5) and normal plus EA group (CD+EA, n=7), or obese group (HFD, n=6) and obese plus EA group (HFD+EA, n=12). The CD+EA group and the HFD+EA group were treated with EA at "Zusanli"(ST 36) and "Neiting"(ST 44, 2 Hz/15 Hz, 0.6-1.0 mA) for 20 min, 6 times a week for 4 weeks. Body weight, ratio of WAT/body weight were calculated, qPCR and Western blot were applied to detect mRNA and protein levels of adenosine receptors in the epididymal adipose tissue (Epi-WAT), respectively. RESULTS: Compared with the normal diet group, high fat diet significantly increased body weight in C 57 BL/6 mice after feeding for 12 weeks (P<0.01); 18 out of 28 mice in the high fat diet group were classified as obesity. After treatment, the body weight and the ratio of Epi-WAT/body weight of the HFD group were increased than those in the CD group (P<0.05), the change of body weight in the HFD group was bigger than that in the CD group (P<0.01). Compared with the HFD group, the body weight and the ratio of Epi-WAT/body weight of the HFD+EA group were decreased after EA (P<0.05), the change of body weight was also significantly increased (P<0.01). No significant differences were found among the four groups in the expression level of A1R mRNA (P>0.05). The expression of A3R mRNA in the HFD group was lower than that in the CD group (P<0.01), while the expressions of A2A R and A2BR proteins were decreased in the HFD group than in the CD group (P<0.01). In comparison with the HFD group, the expression levels of A2AR and A2BR mRNAs and proteins were significantly up-regulated in the HFD+EA group, respectively (P<0.05, P<0.01). CONCLUSIONS: EA intervention is able to reduce the body weight of DIO mice, which Feb be associated with its effects in regulating the expression of A2AR and A2BR in WAT, suggesting a new mechanism of EA in accelerating peripheral WAT metabolism.

Cardioprotective Effect of Electroacupuncture Pretreatment on Myocardial Ischemia/Reperfusion Injury via Antiapoptotic Signaling.

Objectives. Our previous study has used RNA-seq technology to show that apoptotic molecules were involved in the myocardial protection of electroacupuncture pretreatment (EAP) on the ischemia/reperfusion (I/R) animal model. Therefore, this study was designed to investigate how EAP protects myocardium against myocardial I/R injury through antiapoptotic mechanism. Methods. By using rats with myocardial I/R, we ligated the left anterior descending artery (LAD) for 30 minutes followed by 4 hr of reperfusion after EAP at the Neiguan (PC6) acupoint for 12 days; we employed arrhythmia scores, serum myocardial enzymes, and cardiac troponin T (cTnT) to evaluate the cardioprotective effect. Heart tissues were harvested for western blot analyses for the expressions of pro- and antiapoptotic signaling molecules. Results. Our preliminary findings showed that EAP increased the survival of the animals along with declined arrhythmia scores and decreased CK, LDH, CK-Mb, and cTnT levels. Further analyses with the heart tissues detected reduced myocardial fiber damage, decreased number of apoptotic cells and the protein expressions of Cyt c and cleaved caspase 3, and the elevated level of Endo G and AIF after EAP intervention. At the same time, the protein expressions of antiapoptotic molecules, including Xiap, BclxL, and Bcl2, were obviously increased. Conclusions. The present study suggested that EAP protected the myocardium from I/R injury at least partially through the activation of endogenous antiapoptotic signaling.

Electroacupuncture Reduces Weight Gain Induced by Rosiglitazone through PPARγ and Leptin Receptor in CNS.

We investigate the effect of electroacupuncture (EA) on protecting the weight gain side effect of rosiglitazone (RSG) in type 2 diabetes mellitus (T2DM) rats and its possible mechanism in central nervous system (CNS). Our study showed that RSG (5 mg/kg) significantly increased the body weight and food intake of the T2DM rats. After six-week treatment with RSG combined with EA, body weight, food intake, and the ratio of IWAT to body weight decreased significantly, whereas the ratio of BAT to body weight increased markedly. HE staining indicated that the T2DM-RSG rats had increased size of adipocytes in their IWAT, but EA treatment reduced the size of adipocytes. EA effectively reduced the lipid contents without affecting the antidiabetic effect of RSG. Furthermore, we noticed that the expression of PPARγ gene in hypothalamus was reduced by EA, while the expressions of leptin receptor and signal transducer and activator of transcription 3 (STAT3) were increased. Our results suggest that EA is an effective approach for inhibiting weight gain in T2DM rats treated by RSG. The possible mechanism might be through increased levels of leptin receptor and STAT3 and decreased PPARγ expression, by which food intake of the rats was reduced and RSG-induced weight gain was inhibited.

[Analysis on the academic cultivation of young TCM scholars based on Professor LIU Zhicheng's academic track].

In view of the public academic papers of Professor LIU Zhicheng, his academic track is analyzed, including validating effect, exploring mechanism, improving efficacy, expanding disease variety, deepening mechanism, optimizing program and seeking breakthrough. On this basis, combined with the self-character of TCM, the potential path for the academic cultivation of young scholars is refined: focusing on clinical practice, confirming clinical efficacy, deepening mechanism exploration, improving clinical therapeutic effect, optimizing clinical program, extending clinical disease spectrum, inheriting, seeking, breaking through, innovating, expecting and topping over. It is believed that during the academic cultivation, the importance is to form the stable research direction and continuity, organize research team, absorb actively new knowledge and thoughts, and adopt multidisciplinary cross and coordination so as to achieve the greater success ultimately.

Effects of Ziziphus jujuba fruit extracts on cytochrome P450 (CYP1A2) activity in rats.

Drug-drug interactions have become a serious problem in the clinic, since plant-based medicines are extensively used. The present study investigated the effects of Ziziphus jujuba fruit (ZJ) extract on the pharmacokinetics of phenacetin, a typical substrate of a cytochrome P450 enzyme CYP 1A2, in rats. The rats were pretreated with the water extract (1.0 g · kg(-1)) or the ethanolic extract (3.6 g · kg(-1)) of ZJ for 10 days, and the pharmacokinetics of phenacetin was investigated after intravenous administration. In an in vitro assay, acetaminophen formation in the hepatic microsomes of ZJ-treated rats was investigated to assess CYP1A2 activity. Our results demonstrated that the treatment with the water and ethanolic extracts of ZJ decreased the plasma concentration of phenacetin and increased the plasma concentration of acetaminophen, resulting in a 43.2% and 15.5% reduction in the AUC0-120 of phenacetin, respectively, and a 53.2% and 64.9% increase in the AUC0-120 of acetaminophen, respectively after intravenous administration. The water or ethanolic extract of ZJ significantly increased the clearance of phenacetin and acetaminophen formation in hepatic microsomes. In conclusion, ZJ extracts displayed effects on the pharmacokinetics of phenacetin and increased the CYP1A2 activity in rats. Therefore, precaution on drug-drug interactions should be taken when ZJ is co-administered with drugs metabolized by CYP1A2, which may result in decreased concentrations of these drugs.

[Acupuncture Intervention Reduced Weight Gain Induced by Hypoglycemic Agents through Food Intake-related Targets in Central Nervous System].

Clinical practice shows that thiazolidinediones (TZDs) induce weight gain in patients with type-II diabetes mellitus during treatment, which restrains its application and generalization clinically. It has been demonstrated that acupuncture therapy is useful in easing obesity in clinical trials. In the present paper, we summarize the underlying mechanism of weight gain induced by TZDs through food intake-related targets in the central nervous system and analyze the possible effects of acupuncture therapy. Acupuncture therapy is expected to reduce weight gain side effect of TZDs through 1) lowering permeability of blood brain barrier to reduce TZDs concentration in the brain, 2) upregulating the expression of hypothalamic leptin and inhibiting hypothalamic neuropiptide Y expression, and 3) down-regulating activities of peroxisome proliferator-activated receptor to reduce energy intake and fat syntheses.

In vitro and in vivo assessment of CYP2C9-mediated herb-herb interaction of Euphorbiae Pekinensis Radix and Glycyrrhizae Radix.

According to traditional Chinese medicine theories, Euphorbiae Pekinensis Radix and Glycyrrhizae Radix should not be used together in one prescription, because their interaction leads to an unexpected consequence. However, the mechanism remains unclear. The purpose of this study was to find out whether CYP2C9 was involved in this herb-herb interaction by using tolbutamide as a probe substrate in vivo and in vitro. Both Euphorbiae Pekinensis Radix and Glycyrrhizae Radix showed induction activity toward CYP2C9, while the combination of them showed a more potent induction activity toward CYP2C9 in vivo. In vitro study revealed only the combination of the herbs could induce the activity of CYP2C9. Thus, both in vivo and in vitro study indicated combination of Glycyrrhizae Radix and Euphorbiae Pekinensis Radix could induce the activity of CYP2C9 to a high level, which may result in decreased plasma levels of major active ingredients of these two herbs, as well as other herbs in the prescriptions. Further research also appears to be necessary to identify the main enzymes involved in the metabolism of the active ingredients in Glycyrrhizae Radix and Euphorbiae Pekinensis Radix.