Purinergic signaling in myocardial ischemia-reperfusion injury.

Purines and their derivatives, extensively distributed in the body, act as a class of extracellular signaling molecules via a rich array of receptors, also known as purinoceptors (P1, P2X, and P2Y). They mediate multiple intracellular signal transduction pathways and participate in various physiological and pathological cell behaviors. Since the function in myocardial ischemia-reperfusion injury (MIRI), this review summarized the involvement of purinergic signal transduction in diversified pathological processes, including energy metabolism disorder, oxidative stress injury, calcium overload, inflammatory immune response, platelet aggregation, coronary vascular dysfunction, and cell necrosis and apoptosis. Moreover, increasing evidence suggests that purinergic signaling also mediates the prevention and treatment of MIRI, such as ischemic conditioning, pharmacological intervention, and some other therapies. In conclusion, this review exhibited that purinergic signaling mediates the complex processes of MIRI which shows its promising application and prospecting in the future.

Animal model of coronary microembolization under transthoracic echocardiographic guidance in rats.

The aim of the study was to develop a model of coronary microembolization (CME) in rats at a lower cost. We developed a novel rat model without thoracotomy and ventilation under the guidance of echocardiography. Rats were sacrificed at 3 h, 24 h and 1 month postoperatively in both the Echo-CME and Open-chest CME groups for the comparison of the modeling accuracy, mortality, cardiopulmonary circulation, pleural adhesion and ventilation-induced lung injury (VILI). Results showed that the coronary microthrombus formed at 3 h and reached its peak at 24 h postoperatively, which included platelet aggregation and fibrin web. The Echo-group increases success rates, decreased mortality, postoperative complications including pleural adhesion, cardiopulmonary dysfunction and VILI postoperatively than the Open-chest group at 1month postoperatively. The ejection fraction of the CME group decreased to 50% and obvious cardiac fibrosis formed at 3 months postoperatively. Our unique surgical method provided a platform to study molecular mechanisms and potential new pathways for CME treatment.

Electroacupuncture interventions alleviates myocardial ischemia reperfusion injury through regulating gut microbiota in rats.

Electroacupuncture (EA) intervention has a remarkable cardioprotection against myocardial ischemia reperfusion injury (MIRI). Recently, it has been suggested that the gut microbiota plays an important role in regulating the progression and prognosis of MIRI. The purpose of this study was to illustrate the relationship between gut microbiota and cardioprotection of EA on MIRI. We conducted a MIRI model by ligating the left anterior descending coronary artery for 30 min followed by reperfusion in male Sprague Dawley rats, which then received 7 days of EA intervention. Echocardiography was employed to evaluate left ventricular function. Fecal samples were collected for microbial analysis by 16S rDNA high-throughput sequencing. Blood samples and myocardium were collected for inflammatory cytokine detection by enzyme linked immunosorbent assay (ELISA) and Western blot. Hematoxylin & eosin (HE) staining and immunofluorescence of ileum tissue were performed for intestinal damage evaluation. After 7 days of EA intervention, the left ventricular function was improved with significantly increased ejection fraction and fractional shortening. Furthermore, we found that EA intervention reversed the changed gut microbiota induced by MIRI, including Clostridiales, RF39, S24-7, Desulfovibrio, and Allobaculum, improved the impaired gut barrier, reduced the production and circulation of lipopolysaccharide (LPS), inhibited the level of interleukin 6 (IL-6) and interleukin 12 (IL-12) in periphery and decreased the expression of Toll like receptor 4 (TLR4) and IL-6 in myocardium. EA intervention could improve the impaired gut mucosal barrier and reduce the production and circulation of LPS after MIRI through regulating gut microbiota, thus inhibiting the circulation and myocardium inflammation and finally exerted the cardioprotective effect.

Probiotics protect against RSV infection by modulating the microbiota-alveolar-macrophage axis.

Respiratory syncytial virus (RSV) is leading cause of respiratory tract infections in early childhood. Gut microbiota is closely related with the pulmonary antiviral immunity. Recent evidence shows that gut dysbiosis is involved in the pathogenesis of RSV infection. Therefore; pharmacological and therapeutic strategies aiming to readjust the gut dysbiosis are increasingly important for the treatment of RSV infection. In this study, we evaluated the therapeutic effects of a probiotic mixture on RSV-infected mice. This probiotic mixture consisted of Lactobacillus rhamnosus GG, Escherichia coli Nissle 1917 and VSL#3 was orally administered to neonatal mice on a daily basis either for 1 week in advance or for 3 days starting from the day of RSV infection. We showed that administration of the probiotics protected against RSV-induced lung pathology by suppressing RSV infection and exerting an antiviral response via alveolar macrophage (AM)-derived IFN-ß. Furthermore, administration of the probiotics reversed gut dysbiosis and significantly increased the abundance of short-chain fatty acid (SCFA)-producing bacteria in RSV-infected mice, which consequently led to elevated serum SCFA levels. Moreover, administration of the probiotics restored lung microbiota in RSV-infected mice. We demonstrated that the increased production of IFN-ß in AMs was attributed to the increased acetate in circulation and the levels of Corynebacterium and Lactobacillus in lungs. In conclusion, we reveal that probiotics protect against RSV infection in neonatal mice through a microbiota-AM axis, suggesting that the probiotics may be a promising candidate to prevent and treat RSV infection, and deserve more research and development in future.

A novel oleanolic acid derivative HA-19 ameliorates muscle atrophy via promoting protein synthesis and preventing protein degradation.

Muscle atrophy refers to a decrease in the size of muscles in the body, occurs in certain muscles with inactivity in many diseases and lacks effective therapies up to date. Natural products still play an important role in drug discovery. In the present study, derivatives of a natural product, oleanolic acid, were screened with myoblast differentiation and myotube atrophy assays, respectively. Results revealed that one of the derivatives, HA-19 showed the most potent anti-muscle atrophy activity, and was used for further studies. We demonstrated that HA-19 led to the increase of the protein synthesis by activating mechanistic target of rapamycin complex 1 (mTORC1)/p70 S6K pathways, and also enhanced myoblast proliferation and terminal differentiation via up-regulating of the myogenic transcription factors Pax7, MyoD and Myogenin. The interesting thing was that HA-19 also suppressed protein degradation to prevent myotube atrophy by down-regulating negative growth factors, FoxO1, MuRF1 and Atrogin-1. The results were also supported by puromycin labelling and protein ubiquitination assays. These data revealed that HA-19 possessed a "dual effect" on inhibition of muscle atrophy. In disuse-induced muscle atrophy mice model, HA-19 treatment significantly increased the weights of bilateral tibialis anterior (TA), gastrocnemius (Gastroc.), quadriceps (Quad.), suggesting the effectiveness of HA-19 to remit disuse-induced muscle atrophy. Our finding demonstrated that HA-19 has a great potential as an inhibitor or lead compound for the anti-muscle atrophy drug discovery.

[Research progress of histone 3 methyltransferase MLL4].

Mixed linked leukemia 4 (MLL4) is a specific methyltransferase of histone 3 position lysine 4 (H3K4). It is also one of the important members of COMPASS/Set1-like protein complex. Both MLL4 protein itself and its mediated H3K4 methylation modification can cause changes in chromatin structure and function, thus regulating gene transcription and expression. With the studies of MLL4 protein in recent years, the roles of MLL4 gene, MLL4 protein and protein complex in the development of tissues and organs, tumor diseases and other physiological and pathophysiological processes have been gradually revealed. In this paper, the research progress of MLL4 gene, MLL4 protein characteristics, biological function and its effect on disease were reviewed, in order to further understand the effect of histone methyltransferase on gene expression regulation, as well as its non-enzyme dependent function. This paper may provide new ideas for the prevention, diagnosis and treatment of related diseases.

A systematic review and meta-analysis of acupuncture for improving learning and memory ability in animals.

BACKGROUND: Memory loss is the most prominent symptoms of brain aging, but there is currently no evidence-based treatment strategy. Acupuncture has been widely used in China and the effectiveness for improving learning and memory has been mentioned in previous studies. We conducted this systematic review and meta-analysis to evaluate the effectiveness of acupuncture for improving learning and memory in animal experiments. METHODS: We searched Pubmed, Embase, Ovid Medline(R), the China National Knowledge Infrastructure (CNKI), Chinese Science and Technology Periodical Database (VIP) and Wanfang data Information Site to collect studies published up to December 2015. Study quality for each included article was evaluated according to the CAMARADES 10-item checklist. Outcome measure is Morris water maze. A meta-analysis was conducted according to the Cochrane systematic review method by using RevMan 5.3 software. RESULTS: Forty-two studies involving 944 animals were included. The quality score of the studies ranged from 2 to 8, with a mean of 5.3. Meta-analysis results showed that 24 studies reported significant effect of acupuncture for decreasing escape latency (-3.00, 95 % CI: -3.78 ~ -2.23, P < 0.00001), 14 studies reported significant effect of acupuncture for increasing frequency of cross platform (2.57, 95 % CI: 1.92 ~ 3.22, P < 0.00001), and 7 studies reported significant effect of acupuncture for increasing time in target quadrant (2.00, 95 % CI: 1.10 ~ 2.91, P < 0.00001) compared with the control group. CONCLUSIONS: These findings show acupuncture has a potential role in improving learning and memory ability in animal models, suggesting it as a candidate therapy for memory loss of aged brain.

[Bibliometric Analysis of Applying Proteomics Technology in Acupuncture and Moxibustion Re- search].

OBJECTIVE: To analyze document features and current research status of applying proteomics technology in acupuncture and moxibustion research, thereby further providing reference for further research and development in this area. METHODS: CNKI, Wangfang Data, CBM, PubMed, and Medline were included. Retrieved were literatures involving applying proteomics technology in acupunc- ture and moxibustion from January 2004 to December 2015. Document features and current research features were statistically analyzed using bibliometrics. RESULTS: Totally 91 articles were included, 60 from journals, 8 conference theses, and 23 dissertations. The 60 articles were published in 47 kinds of journals, and 56 of them were published in core journals. Theories of acupuncture and moxibustion (correlated with meridians, zang-fu organs) and mechanism research were hotspots, involving 19 kinds of disease models (dominated as gastric mucosal lesion, cerebral ischemia, and spinal cord injury). Two- dimensional gel based electrophoresis technology was used in 51 articles (69. 86%). CONCLUSIONS: Proteomics technology has been widely applied in the field of acupuncture and moxibustion research. It could promote researches on meridians and zang-fu organs correlated theories and mechanisms. The depth and breadth of the study is worth further widening.

A protocol of histone modification-based mechanistic study of acupuncture in patients with stable angina pectoris.

BACKGROUND: Angina pectoris (Angina) is a medical condition related to myocardial ischemia. Although acupuncture has been widely accepted as a clinical approach for angina, there is no sufficient evidence of its effectiveness against this syndrome, and its mechanisms have not yet been well elucidated. We develop this protocol to confirm the clinical efficacy of electro-acupuncture on stable angina pectoris by needling on acupoint Neiguan (PC6). Furthermore, we employ high-throughput sequencing technology to investigate the gene expression profiling and determine involvement of histone modifications in the regulation of genes after electro-acupuncture treatment. METHODS/DESIGN: A randomized, controlled, double-blinded (assessor and patients) trial will be carried out. Sixty participants will be randomly assigned to two acupuncture treatment groups and one control group in a 1:1:1 ratio. Participants in acupuncture groups will receive 12 sessions of electro-acupuncture treatment across 4 weeks, followed by a 12-week randomization period. The acupuncture groups are divided into Neiguan (PC6) on Pericardium Meridian of Hand-jueyin or a non-acupoint. The primary clinical measure of effect is the frequency of angina attacks between these groups for four weeks after randomization. RNAs are extracted from peripheral neutrophils collected from all participants on day 0, day 30, and week 16, and are processed to RNA-Seq. We then investigate profiles of histone modifications by ChIP-Seq, for H3 Lysine 4 (H3K4me) and acetylation of H3 Lysine 27 (H3K27ac), in the presence or absence of acupuncture treatment. DISCUSSION: This study determines the efficacy and mechanisms of electro-acupuncture on stable angina pectoris. We focus on effectiveness of acupuncture on alleviating symptoms of myocardial ischemia and the gene regulation and the chromatin remodeling marks, including H3K4me1, H3K4me2, and H3K27ac, which could be key factors for regulating gene expressions caused by electro-acupuncture treatment at Neiguan. This is the first genome-wide study of electro-acupuncture treatment in angina patients, and will provide valuable information for future studies in the fields of acupuncture and its underlying mechanisms. Fourteen patients have been recruited since recruitment opened in November of 2012. This study is scheduled to end in November of 2014. TRIALS REGISTRATION: ChiCTR-TRC-12002668.

Acupuncture promotes white adipose tissue browning by inducing UCP1 expression on DIO mice.

BACKGROUND: To study the influence of acupuncture and its possible mechanism on white adipose tissue of high fat diet-induced obese. METHODS: Four-week-old C57BL/6 J mice were randomly divided into a normal diet group and a high-fat diet (HFD) group. After 8 weeks, the HFD mice were randomly divided into Electro-acupuncture (EA) group and control group. Mice in the EA group were electro-acupunctured, under physical restraint, on Zusanli (ST36) and Neiting (ST44) acupoints, while the mice in the control group were under physical restraint only. Body weight and food intake were monitored, and serum leptin, cholesterol and triglyceride levels were measured by using biochemistrical methods. The effect of EA on white adipose tissues (WAT) was assessed by qPCR, immunoblotting, immunohistochemistry (IHC), immunoprecipitation and cold endurance experiment. RESULTS: The WAT/body weight ratio decreased (P < 0.05) in the EA group, albeit no significant difference on food consumption between EA and control groups. The difference in the darkness of Epi-WAT between EA and control groups could be distinguished visually. HE staining indicated that the EA mice had an increased number of UCP1-immunoreactive paucilocular adipocytes in their WAT. The expressions of brown adipose tissue (BAT) markers, including UCP1, COX4il and Nrtf1 were increased in the WAT of EA mice, acetylation of Pparγ was decreased by electro-acupuncture. CONCLUSION: EA can remodel WAT to BAT through inducing UCP1 expression, and this may be one of the mechanisms by which acupuncture affects weight loss.

Electro-acupuncture at Neiguan pretreatment alters genome-wide gene expressions and protects rat myocardium against ischemia-reperfusion.

This study investigated genome-wide gene expressions and the cardioprotective effects of electro-acupuncture pretreatment at the PC6 Neiguan acupoint on myocardial ischemia reperfusion (I/R) injury. Male SD rats were randomly divided into four groups: sham operation (SO), I/R, electro-acupuncture at the PC6 Neiguan acupoint pretreatment (EA) and electro-acupuncture at non-acupoint pretreatment (NA). Compared with the I/R group, the survival rate of the EA group was significantly increased, the arrhythmia score, infarction area, serum concentrations of CK, LDH and CK-Mb and plasma level of cTnT were significantly decreased. RNA-seq results showed that 725 genes were up-regulated and 861 genes were down-regulated under I/R conditions compared to the SO group; both EA and NA reversed some of these gene expression levels (592 in EA and 238 in NA group). KEGG pathway analysis indicated that these genes were involved in multiple pathways, including ECM, MAPK signaling, apoptosis, cytokine and leukocyte pathways. In addition, some pathways were uniquely regulated by EA, but not NA pretreatment, such as oxidative stress, cardiac muscle contraction, gap junction, vascular smooth muscle contraction, hypertrophic, NOD-like receptor, and P53 and B-cell receptor pathways. This study was first to reveal the gene expression signatures of acute myocardial I/R injury and electro-acupuncture pretreatment in rats.

Effect of electroacupuncture at "Neiguan" (PC6) on pain and brain orexin 1 receptor in mice with inflammatory pain. / 电针"å† å ³"å¯¹ç‚Žæ€§ç—›å°é¼ è„‘å† é£Ÿæ¬²ç´ 1受体的影响.

OBJECTIVES: To observe the effect of electroacupuncture (EA) at "Neiguan" (PC6) on pain response in mice injected with complete Freund's adjuvant (CFA) in the hind paw, so as to investigate the mechanism of orexin 1 receptor (OX1R) -endogenous cannabinoid 1 receptor (CB1R) pathway in acupuncture analgesia. METHODS: A total of 48 male C57BL/6 mice were used in the present study. In the first part of this study, 18 mice were randomized into control, model and EA groups, with 6 mice in each group. In the second part of this study, 30 mice were randomized into control, model, EA, EA+Naloxone, EA+OX1R antagonist (SB33486) groups, with 6 mice in each group. Inflammatory pain model was established by subcutaneous injection of 20 µL CFA solution in the left hind paw. EA (2 Hz, 2 mA ) was applied to bilateral PC6 for 20 min, once a day for 5 consecutive days. The mice in the EA+Naloxone and EA+SB33486 groups were intraperitoneally injected with naloxone (10 mg/kg) or SB33486 (15 mg/kg) 15 min before EA intervention on day 5, respectively. Tail-flick method and Von Frey method were used to detect the thermal pain threshold and mechanical pain threshold of mice. Quantitative real-time PCR was used to detect the expression level of ß-endorphin mRNA in periaqueductal gray (PAG) of mice. The expression of OX1R positive cells in the lateral hypothalamic area (LH) and CB1R positive cells in the ventrolateral periaqueductal gray (vlPAG) were detected by immunofluorescence. RESULTS: Compared with the control group, the thermal pain threshold and mechanical pain threshold of the model group were decreased (P<0.001), the expression level of ß-endorphin mRNA in PAG was decreased (P<0.001), and the numbers of OX1R positive cells in LH and CB1R positive cells in vlPAG were decreased (P<0.05, P<0.001). Compared with the model group, the thermal pain threshold and mechanical pain threshold of the EA group were significantly increased (P<0.001), and the numbers of OX1R positive cells in LH and CB1R positive cells in vlPAG were increased (P<0.01, P<0.001). Compared with the EA group, the mechanical pain threshold in the EA+SB33486 group was significantly decreased (P<0.01), but there was no significant difference in the mechanical pain threshold between the EA+Naloxone group and EA group, and the numbers of OX1R positive neurons in LH and CB1R positive neurons in vlPAG were decreased in the EA+SB33486 group (P<0.001). CONCLUSIONS: EA at PC6 can achieve analgesic effect on CFA mice by activating the OX1R-CB1R pathway in the brain, and this effect is opioid-independent.

Effect of electroacupuncture on inflammatory response and immune cells in mice with chronic inflammatory pain. / ç”µé’ˆå¯¹æ ¢æ€§ç‚Žæ€§ç—›å°é¼ ç‚Žæ€§ååº”å’Œå ç–«ç»†èƒžçš„å½±å“.

OBJECTIVES: To observe the effects of electroacupuncture(EA) on local inflammatory mediators and macrophage polarization, and immune cells in the spleen of mice with chronic inflammatory pain induced by complete Freund's adjuvant (CFA) in the hind paw, so as to investigate the immunoinflammatory regulatory mechanisms of EA in relieving pain and swelling in mice with chronic inflammatory pain. METHODS: Thirty C57BL/6 mice were randomly divided into control, model, and EA groups, with 10 mice in each group. Chronic inflammatory pain model were established by subcutaneous injection of 20 µL CFA solution in the left hind paw for 7 consecutive days. After modeling, mice in the EA group received EA at bilateral "Zusanli"(ST36) for 20 min (2 Hz/100 Hz, 1 mA) once a day for 18 consecutive days. Mechanical pain threshold, heat pain thresholds, and paw thickness were measured before and after mode-ling, and after interventions. Western blot was used to detect the expression of tumor necrosis factor-alpha (TNF-α), interleukin (IL)-1ß, and NOD-like receptor protein 3 (NLRP3) in the paw tissue. Immunohistochemistry was used to detect the positive expression of M1-type macrophage marker inducible nitric oride synthase (iNOS) and M2-type marker CD206 in the paw, and flow cytometry was used to detect the proportion of F4/80+ CD11b+ macrophages, Ly6G+ CD11b+ neutrophils, and CD25+ Foxp3+ regulatory T cells (Treg) in the spleen. RESULTS: Compared with the control group, mechanical pain and heat pain thresholds were significantly reduced(P<0.000 1), while paw thickness, expressions of IL-1ß, TNF-α, and NLRP3 in the paw, and positive expression of M1 macrophage marker iNOS in the paw, the proportions of macrophages and neutrophils in the spleen were significantly increased (P<0.000 1, P<0.001) in the model group. Compared with the model group, mechanical pain threshold and heat pain thresholds, CD206 positive expression in the paw, and Treg cell proportion in spleen were significantly increased (P<0.01), while paw thickness, the expressions of IL-1ß, TNF-α and NLRP3 in the paw, as well as the positive expression of M1 macrophage marker iNOS in the paw, the proportions of macrophages and neutrophils in the spleen were significantly reduced (P<0.001, P<0.01, P<0.05)in mice of the EA group after intervention. CONCLUSIONS: EA may alleviate pain and swelling in mice with chronic inflammatory pain by regulating the numbers of macrophages, neutrophils, and Treg cells, as well as promoting M2 polarization of local macrophages and inhibiting the release of pro-inflammatory cytokines.

Unusual electro-optical behavior in a wide-temperature BPIII cell.

A low driving voltage and fast response blue phase III (BPIII) liquid-crystal device with very low dielectric anisotropy is demonstrated. To stabilize BPIII in a wide temperature range (> 15°C), a chiral molecule with good solubility was chosen. By studying field-dependent polarization state of the transmitting light, it was found that the field-induced birefringence becomes saturated in the high field. However, the transmitting intensity exhibits a tendency to increase as the electric field increases. This indicates that the electro-optical behavior in BPIII device may be from the flexoelectric effect, which induces tilted optical axis and then induces birefringence. Because the phase transition from BPIII to chiral nematic phase does not happen, the device shows no hysteresis effect and no residual birefringence, exhibits fast response, and can be a candidate for fast photonic application.

[Anti-depressive effect of acupuncture on selective serotonin reuptake inhibitors].

OBJECTIVE: To evaluate anti-depressive effects of acupuncture on selective serotonin reuptake inhibitors (SSRIs). METHODS: Totally 60 patients with depression were randomly assigned to the control group (30 cases) and the treatment group (30 cases). All patients took one kind of SSRIs. Those in the treatment group were additionally treated by acupuncture. All were treated for 6 weeks. Patients' efficacies were evaluated with Hamilton Depression Scale (HAMD), Self-rating Depression Scale (SDS), and Eisenberg antidepressant side effects scale (Asberg). RESULTS: Compared with the control group after 6 weeks of treatment, the cured-markedly effective rate was improved by 33.4% in the treatment group (P < 0.05). The HAMD was lower in the treatment group. The tendency of interaction of sleep disorder factor and anxiety/somatization factor was different between at the end of 1-week treatment and at the end of 6-week treatment in the treatment group (P < 0.05). The SDS score decreased at the end of 6-week treatment in the treatment group. The reduction rate was elevated by 19.23% (P < 0.05). By the end of 6-week treatment, the average score of Asberg decreased by 3.77 score in average in the treatment group, while it decreased by 0.07 score in average in the control group (P < 0.05). CONCLUSION: Acupuncture could effectively improve anti-depressive effects of SSRIs and reduce their adverse reactions.

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.

Electroacupuncture preconditioning protects against myocardial ischemia-reperfusion injury by modulating dynamic inflammatory response.

Background: The protective effects of electroacupuncture (EA) preconditioning against myocardial ischemia-reperfusion injury (MIRI) have been reported. However, the underlying mechanism remains unclear. Recent research has indicated that the dynamic inflammatory response following MIRI plays an essential role in the progression of myocardial injury. This study aimed to investigate the myocardial protective effects of EA preconditioning on MIRI in rats and to explore the relevant mechanism from the perspective of dynamic inflammatory response. Methods: A MIRI model was employed, and the rats were subjected to EA on Neiguan for four days prior to modeling. The myocardial protective effect of EA preconditioning was evaluated by echocardiography, Evans blue and triphenyltetrazolium chloride staining. Real-time polymerase chain reaction, Western blot, hematoxylin & eosin staining, and immunohistochemistry were utilized to detect the content of mitochondrial DNA, NOD receptor family protein 3 (NLRP3) inflammasome activation, neutrophil recruitment and macrophage infiltration in blood samples and myocardium below the ligation. Results: We found that EA preconditioning could accelerate the recovery of left ventricle function after MIRI and reduce the myocardial infarction area, thereby protecting the myocardium against MIRI. Furthermore, EA preconditioning was observed to ameliorate mitochondrial impairment, reduce the level of plasma mitochondrial DNA, modulate NLRP3 inflammasome activation, attenuate neutrophil infiltration, and promote the polarization of M1 macrophages towards M2 macrophages in the myocardium after MIRI. Conclusion: EA preconditioning could reduce plasma mtDNA, suppress overactivation of the NLRP3 inflammasome, facilitate the transition from the acute pro-inflammatory phase to the anti-inflammatory reparative phase after MIRI, and ultimately confer cardioprotective benefits.

[Effect of different electrical current intensities of electroacupuncture preconditioning on cardiac function and macrophage polarization in mice with acute myocardial ischemia].

OBJECTIVE: To observe the effect of different intensities of electroacupuncture (EA) preconditioning on car-diac function and polarization state of macrophages in mice with acute myocardial ischemia (AMI), so as to explore its possible mechanism underlying improvement of AMI. METHODS: A total of 50 male C57BL/6J mice were randomly divided into sham ope-ration, AMI model, and EA pretreatment groups (0.5 mA, 1 mA, 3 mA subgroups), with 10 mice in each group/subgroup. The mice in the EA pretreatment groups were subjected to EA stimulation of bilateral "Neiguan"(PC6) with 0.5, 1.0 and 3 mA respectively and frequency of 2 Hz/15 Hz for 20 min, once a day, for 3 days. The acute myocardial ischemia model was established by ligating the anterior descending branch (ADB) of the left coronary artery, while the sham operation only had a surgical suture trans-passed below the ADB but without ligation. The myocardial infarction area was measured after TTC staining, and the cardiac function ï¼»left ventricular ejection fraction (EF), short-axis contraction rate (FS)ï¼½ was detected by using echocardiography. The M1 macrophages were labeled with CD11b+F480+CD206low, M2 macrophages were labeled with CD11b+F480+CD206high and detected by using flow cytometry, and the expression levels of myocardial interleukin-1ß (IL-1ß), tumor necrosis factor-α (TNF-α), Toll-like receptor-4 (TLR4) proteins were detected by using Western blot. RESULTS: Compared with the sham operation group, the model group had a significant increase in the infarction area (P<0.000 1), number of cardiac macrophages and percentage of M1 type macrophages (P<0.000 1), and the expression levels of myocardial IL-1ß, TNF-α, TLR4 proteins (P<0.001, P<0.01), and a remarkable decrease in the levels of EF, FS and the percentage of M2 type macrophages (P<0.000 1). In contrast to those of the model group, the area of myocardial infarction (P<0.000 1, P<0.01), expression levels of myocardial IL-1ß, TNF-α, TLR4 proteins (P<0.01, P<0.05, P<0.001) in the 0.5 mA, 1 mA and 3 mA groups, number of macrophages and percentage of M1 macrophages (P<0.05) in the 1 mA group were significantly decreased, while the levels of EF and FS (P<0.000 1, P<0.05, P<0.001) in the 3 EA groups, and percentage of M2 macrophage (P<0.05) in the 1 mA group were significantly increased. Comparison among the 3 EA groups displayed that the effects of 1 mA group were significantly superior to those of 0.5 and 3 mA groups in up-regulating EF and FS (P<0.01, P<0.001), and in down-regulating the area of infarct myocardium (P<0.01, P<0.000 1), and the expression of TLR4 protein (P<0.01), and 0.5 mA group in the expression of IL-1ß and TNF-α proteins (P<0.05). CONCLUSION: EA preconditioning with electrical current intensities of 0.5 mA, 1 mA and 3 mA can effectively reduce myocardial infarction size, improve cardiac function in mice with AMI, which may be related with its effects in reducing the number of cardiac macrophages and down-regulating the expression of myocardial IL-1ß, TNF-α and TLR4 proteins. The therapeutic effect of 1 mA is better than that of 0.5 and 3 mA.

Multi-material basis and multi-mechanisms of the Dahuang Zhechong pill for regulating Treg/Th1 balance in hepatocellular carcinoma.

BACKGROUND: Dahuang Zhechong pill (DHZCP) improves the inhibitory immune status of mice with hepatocellular carcinoma (HCC) by regulating Treg/Th1 balance. HYPOTHESIS/PURPOSE: To study the multi-material basis and multi-mechanisms of DHZCP against HCC by regulating Treg/Th1 balance in vitro and in vivo. METHODS: UPLC-MS/MS was used to detect the dynamic changes in 29 characteristic components of different polar parts of DHZCP. H&E and TUNEL were used to check pathological condition in HCC mice. The number of CD4+T, CD8+T, Treg, Th1, and Th1-like Treg cells was counted by flow cytometry. TGF-ß, IL-10, IFN-γ, and TNF-α content were detected by ELISA. α-Ketoglutarate and glutamine levels were detected by Trace1310/TSQ8000 GC-MS/MS. p-Smad2, and p-Smad3 protein levels were detected by WB, mRNA expression of Smad2, alanine-serine-cysteine transporter-2, glutaminase, and glutamate dehydrogenase were detected by RT-PCR. Simca-p multivariate data analysis software was used to evaluate the relationship between the different polar parts of DHZCP and the proportion of Treg cells. RESULTS: Water-soluble (PW) and ethyl acetate (PE) polar parts of DHZCP affected the HCC immune system by inhibiting the differentiation of Tregs, reversing the balance of Treg/Th1, and significantly reduced the tumor volume and weight. However, petroleum ether and n-butanol polar parts had no above actions. The changes in emodin, chrysophanol, aloe vera emodin, emodin-8-O-ß-D-glycoside, gallic acid, naringenin, baicalein, wogonin, norwogonin, apigenin, chrysin, glycyrrhizin, formononetin, and palmitic acid were closely related to the changes of Treg cells, which is the main material basis of DHZCP inhibition of Treg differentiation. Additionally, PW mainly inhibit the differentiation of Treg cells by affecting the metabolism of hepatoma cells, improving tumor microenvironment acidity, and glutamine depletion. However, PE inhibited the differentiation of Treg cells mainly by regulating the TGF-ß/Smad pathway. CONCLUSION: In this study, accurate analysis of multi-component was combined with pharmacodynamic evaluations to identify the pharmacodynamic substances of DHZCP in regulating Treg/Th1 balance, and clarified the multi-target mechanism of DHZCP to improve tumor immunity. The study style offers a novel approach for pharmacological research on TCM.

[Effect of electroacupuncture preconditioning on expression of Gasdermin D, Caspase-1 and IL-1ß in rats with myocardial ischemia reperfusion injury].

OBJECTIVE: To observe the effect of electroacupuncture(EA) preconditioning on expression of Caspase-1, Gasdermin D(GSDMD) and interleukin-1ß(IL-1ß) in myocardial tissue of myocardial ischemia reperfusion injury (MIRI) rats in order to explore its underlying mechanisms in resisting MIRI. METHODS: Forty male rats were randomly divided into 4 groups: normal control (normal), sham operation (sham), MIRI model and EA groups. The MIRI model was established by ligation of the left anterior descending branch of the left coronary artery for 30 min and perfusion. EA (2 Hz/100 Hz, 1 mA) was applied to bilateral "Neiguan" (PC6) for 20 min, once a day for 3 consecutive days. The echocardiography was used to analyze the left ventricular end-diastolic dimension (LVEDD), left ventricular end-systolic dimension (LVESD) and left ventricular ejection fraction (LVEF, by using Teichholz formula) 4 h after modeling. The myocardial TTC staining was used to observe the proportion of the infarct area, and Western blot was used to detect the expression levels of GSDMD, Caspase-1, IL-1ß proteins in the myocardium. RESULTS: Compared with the normal group, the immunoactivity of GSDMD was increased in the sham group (P<0.05). Compared with the sham group, the LVEF was significantly decreased (P<0.000 1), while the myocardial infarction area, immunoactivity of GSDMD, and the expression levels of Caspase-1, GSDMD and IL-1ß proteins were considerably increased in the model group (P<0.000 1, P<0.001). In comparison with the model group, the decreased ejection fraction and the increased myocardial infarction area, and Caspase-1, GSDMD and IL-1ß expression were reversed in the EA group (P<0.001, P<0.000 1, P<0.01). CONCLUSION: EA preconditioning may ameliorate myocardial injury in MIRI rats which may be associated with its function in down-regulating the expression of myocardial Caspase-1 protein to reduce cardiomyocyte pyroptosis.