Effect of electroacupuncture on Nrf2/NLRP3/Caspase-1 pathway mediated-pyroptosis in mice with Parkinson's disease. / ç”µé’ˆå¯¹å¸•é‡‘æ£®ç— å°é¼ Nrf2/NLRP3/Caspase-1通路介导的细胞焦亡的影响.

OBJECTIVES: To observe the effect of electroacupuncture (EA) on nuclear transcription factor E2 related factor 2 (Nrf2)/NOD-like receptor family pyrin domain-containing protein 3 (NLRP3)/cysteine aspartic acid specific protease-1 (Caspase-1) pathway in the substantia nigra (SN) of mice with Parkinson's disease (PD), so as to explore the neuroprotective mechanism of EA. METHODS: Forty C57BL/6 male mice were randomly divided into 4 groups, namely, control, PD model, EA and sham-EA groups, with 10 mice in each group. The PD mouse model was established by gavage of rotenone for 4 weeks. Mice in the EA group were given EA stimulation (1 mA, 2 Hz) at "Fengfu" (GV36), bilateral "Taichong" (LR3) and "Zusanli" (ST36) for 30 min, once daily for 2 consecutive weeks. And mice in the sham-EA group were given acupuncture at the subcutaneous areas of the same acupoints without EA stimulation. The open-field test was used for assessment of mouse behavior. The levels of interleukin-1ß (IL-1ß) and interleukin-18 (IL-18) in the serum were detected by enzyme-linked immunosorbent assay . The positive expression of tyrosine hydroxylase (TH) in SN was determined by immunohistochemistry. The mRNA expression levels of Nrf2, NLRP3, Caspase-1, gasdermin D(GSDMD), IL-1ß, IL-18 and the protein expression levels of Nrf2, NLRP3, Caspase-1 and GSDMD in the SN were detected by quantitative real-time PCR and Western blot, separately. RESULTS: After modeling, compared with the control group, the behavioral score was increased (P<0.01), the total exercise time, the total distance and the average speed were decreased (P<0.01), and the positive expression of TH and the mRNA and protein expression levels of Nrf2 in the SN were decreased (P<0.01), while the contents of IL-1ß and IL-18 in serum, the mRNA and protein expression levels of NLRP3, Caspase-1 and GSDMD and the mRNA expression levels of IL-1ß and IL-18 in the SN were up-regulated (P<0.01) in the PD model group. Following EA intervention, the behavioral score was decreased(P<0.01), the total exercise time, total distance and average speed were increased (P<0.01), the positive expression of TH and the mRNA and protein expressions of Nrf2 in SN were up-regulated (P<0.01, P<0.05), while the contents of IL-1ß and IL-18 in serum, the mRNA and protein expression levels of NLRP3, Caspase-1 and GSDMD as well as the mRNA expression levels of IL-1ß and IL-18 in the SN were down-regulated (P<0.01, P<0.05) in the EA group relative to the PD model and sham-EA groups. There were no significant differences in the above indicators between the PD model and sham-EA groups. CONCLUSIONS: EA stimulation of GV36, LR3 and ST36 can improve motor deficits, reduce the loss of dopamine neurons in the SN, and inhibit neuroinflammatory responses in mice with PD, which may be related to its effects in regulating the Nrf2/NLRP3/Caspase-1 pathway mediated pyroptosis.

Effects of electroacupuncture on the intestinal thioredoxin interaction protein/Nod-like receptor 3 signaling pathway in mice with Parkinson's disease. / ç”µé’ˆå¯¹å¸•é‡‘æ£®ç— å°é¼ è‚ é“ç¡«æ°§è¿˜è›‹ç™½äº’ä½œè›‹ç™½/NODæ ·å—ä½“3信号通路的影响.

OBJECTIVES: To observe the effects of electroacupuncture (EA) at "Fengfu" (GV16), "Taichong" (LR3) and "Zusanli" (ST36) on α-synuclein (α-syn), Occludin, Claudin-1, thioredoxin interaction protein (TXNIP) and Nod-like receptor 3 (NLRP3) in Parkinson's disease (PD) mice, so as to investigate the mechanisms of EA on intestinal barrier function and inflammation in PD mice. METHODS: Thirty six C57BL/6 mice were randomly divided into control, model and EA groups, with 12 mice in each group. PD mice model was induced by rotenone intragastric administration for 28 days. Mice in the EA group were treated with EA (2 Hz, 1 mA) at GV16, LR3 and ST36 for 30 min, once a day for 14 days. The behavioral scores were observed. The total distance of autonomic movement was measured by open field test. The expression level of α-syn in substantia nigra and colon tissue was determined by immunohistochemistry. The colonic morphology and goblet cell distribution were observed by Alcian blue staining. The expression levels of Occludin, Claudin-1, TXNIP and NLRP3 mRNA in colon tissue were detected by real-time fluorescence quantitative PCR. RESULTS: Compared with the control group, the behavioral scores of rats were increased (P<0.01)；the total distance of autonomous movement was decreased (P<0.01)；the positive expression level of α-syn in the substantia nigra and colon was increased (P<0.01)；the goblet cells and crypts in colon tissue were reduced, and the muscular layer was thinner；the expression levels of Occludin and Claudin-1 mRNAs in colon tissue were decreased (P<0.01) while TXNIP and NLRP3 mRNAs were increased (P<0.01) in the model group. Compared with the model group, the surface villi of colon tissue was more complete, the goblet cells and crypts were increased, and the muscular layer was thickened；the other indexes were reversed (P<0.01, P<0.05) in the EA group. CONCLUSIONS: EA at GV16, LR3 and ST36 can reduce the abnormal accumulation of α-syn in the substania nigra and colon tissue of PD mice, alleviate the damage of intestinal barrier, regulate TXNIP/NLRP3 signaling pathway, so as to delay the occurrence and development of PD.

Vitamin D alleviates non-alcoholic fatty liver disease via restoring gut microbiota and metabolism.

Background: Non-alcoholic fatty liver disease (NAFLD) represents a severe public health problem. Dysbiosis of gut microbiome has been identified as one of the key environmental factors contributing to NAFLD. As an essential nutrition, Vitamin D (VD) plays an important role in regulating gut microbiota based on its receptor (Vitamin D Receptor, VDR) which is highly expressed in the gastrointestinal tract. Methods: Rats were fed with HFD (high-fat diet) for 12 weeks. And the rats were treated with VD two times a week by intraperitoneal injection for 12 weeks. H&E staining combined with plasma biochemical index was performed to characterize pathological changes and function of the liver. Fecal microbiota 16S rRNA gene sequencing and metabolomics were taken to reveal the altered gut microbiota and metabolites. Result: The VD alleviates the HFD-induced lipid accumulation in the liver as well as decreases the levels of amlodipine besylate (ALT) and amlodipine aspartate (AST). VD supplement decreased the ratio of phylum Firmicutes/Bacteroidetes (F/B) but increased alpha diversity. In addition, the VD treatment improved the HFD-induced gut microbiota by increasing the Prevotella and Porphyromonadaceae and decreasing Mucispirillum, Acetatifactor, Desulfovibrio, and Oscillospira abundance. Furthermore, the capability of tyrosine metabolism, tryptophan metabolism, arginine biosynthesis, and sphingolipid metabolism was enhanced after VD treatment. Consistently, Prevotella positively correlated with tryptophan metabolism and sphingolipid metabolism. Importantly, the Prevotella abundance was positively associated with serotonin, melatonin, tryptamine, L-arginine, and 3-dehydrosphinganine which synthesize from tryptophan, tyrosine, arginosuccinate, and serine, respectively. Conclusion: VD treatment inhibited HFD-induced NAFLD accompany by dysbiosis gut microbiota and metabolites, suggesting that VD supplement could be a potential intervention used for NAFLD treatment by targeting the specific microbiota.

[Effect of electroacupuncture on GLP-1R/PI3K/Akt protein pathway in mice with Parkinson's disease].

OBJECTIVE: To observe the effect of electroacupuncture(EA) on glucagon-like peptide-1 receptor (GLP-1R)/ phosphatidylinositol 3-kinase (PI3K)/ protein kinase B(Akt) protein pathway in the substantia nigra of mice with Parkinson's di-sease (PD)，so as to explore its underlying mechanisms in treatment of PD. METHODS: Forty-eight C57BL/6 male mice were randomly divided into normal, model, EA and inhibitor groups, with 12 mice in each group. PD mouse model was established by intragastrical administration of rotenone for 4 weeks. In the EA group, EA was applied at "Fengfu"(GV16), "Taichong"(LR3) and"Zusanli"(ST36) for 30 min, once daily, for 2 weeks. The mice of the inhibitor group received gavage of dipeptidyl peptidase-4 inhibitor ligliptin (10 mg·kg-1·d-1) once a day for 2 weeks. The behavioral scores of mice in each group were observed. The levels of tyrosine hydroxylase (TH) in serum and substantia nigra were detected by ELISA, and the protein relative expression levels of GLP-1R, phosphorylation of PI3K (p-PI3K) and phosphorylation of Akt (p-Akt) in substantia nigra of midbrain of mice were detected by Western blot. RESULTS: Compared with the normal group, the behavioral scores were significantly increased (P<0.01), TH levels in serum and substantia nigra, protein expression levels of GLP-1R, p-PI3K and p-Akt of the substantia nigra in the model group were significantly decreased (all P<0.01). After intervention and in comparison with the model group, the behavioral scores were significantly decreased (P<0.01), TH levels and the protein expression levels of GLP-1R, p-PI3K and p-Akt in both EA and inhibitor groups were significantly increased (all P<0.01). There were no significant differences in the abovementioned indexes between EA group and inhibitor group (all P>0.05), except for TH levels which were considerably down-regulated in the EA group relative to the inhibitor group (P<0.01, P<0.05). CONCLUSION: EA at GV16, LR3 and ST36 may increase the level of TH in serum and substantia nigra by up-regulating the activity of GLP-1R/PI3K/Akt protein pathway, and improve the behavioral performance of PD induced by rotenone.

[Effect of electroacupuncture preconditioning on cell apoptosis mediated by mitochondrial reactive oxygen species in myocardial ischemia/reperfusion injury rats].

OBJECTIVE: To observe the effect of electroacupuncture (EA) preconditioning on cell apoptosis and the content of mitochondrial reactive oxygen species (ROS) in rats with myocardial ischemia/reperfusion injury (MIRI), so as to explore their mechanisms underlying improvement of MIRI. METHODS: Eighty male Wistar rats were randomly divided into control, sham, ischemia reperfusion (IR) and EA groups, with 20 rats in each group. Rats of the control, sham and IR groups were just banded on the boards for 30 min, once daily for 7 days. Before modeling, EA (2 Hz, 1 mA) was applied to "Neiguan" (PC6), "Zusanli" (ST36), "Guanyuan"(CV4) for 20 min, once daily for 7 successive days in the EA group. The MIRI model was established by occlusion of the left anterior descending branch of the left coronary artery for 20 min, followed by reperfusion for 30 min. Ventricular arrhythmia (VA) score was used to evaluate arrhythmia. Enzyme-linked immunosorbent assay (ELISA) was used to detect the level of CK-MB. DHE staining was used to detect the content of reactive oxygen species (ROS). The gene expression levels of cytochrome C (Cyt-C), Caspase-9 and Caspase-3 were detected by real-time fluorescent quantitative PCR. RESULTS: Compared with the sham group, VA score, serum CK-MB content, ROS content in heart tissue and Cyt-C, Caspase-9 and Caspase-3 gene expression were significantly up-regulated in the MIRI group (all P<0.01). Following the intervention, the increased VA score, serum CK-MB content, ROS content in heart tissue, Cyt-C, Caspase-9 and Caspase-3 gene expression were all reversed in the EA group compared with the MIRI group (all P<0.01). CONCLUSION: The protective effect of EA preconditioning on MIRI may be based on the regulation of ROS mediated-apoptosis pathway.

[Protective effect and regulating effect on FXR/SHP gene of electroacupuncture preconditioning on myocardial ischemia-reperfusion injury in rats].

OBJECTIVE: To explore the protective mechanism of electroacupuncture preconditioning on myocardial ischemia-reperfusion injury in rats. METHODS: A total of 88 SPF-grade Wistar male rats were randomized into a normal group, a sham-operation group, a model group and an electroacupuncture (EA) group, 22 rats in each one. The rats in the normal group received normal diet and no intervention was given for 7 d. The rats in the sham-operation and the model group were bound for 20 min, once a day for 7 d. In the EA group, EA, continuous wave, 2 Hz and 1 mA, was applied at "Neiguan" (PC 6), "Zusanli" (ST 36) and "Guanyuan" (CV 4) for 20 min, once a day for 7 d. In the 8th day, the blood was collected by abdominal aortic method 60 min after intraperitoneal injection of 10% urethane anesthesia by 10 mL/kg in the normal group, and then the tissue of heart was collected. In the model group and the EA group, ligating the left anterior descending coronary artery (LAD) was adopted for 20 min and reperfusion was applied for 40 min, then the samples were collected. The rats in the sham-operation group recevied open-chest operation and no other intervention, the samples were collected in 60 min. Nitro Blue Tetrazolium Chloride monohydrate (NBT) stain was used to detected myocardial infarct size and weight, ELISA was used to measured myocardial injury markers and inflammatory factors (LDH、CK、cTnI), and the fluorescent quantitative PCR method was used to determine the expressions of FXR and SHP gene. RESULTS: ①There was no significant difference between the normal group and the sham-operation group in myocardial infarct size and weight (P>0.05) .Compared with the normal group, myocardial infarct size and weight were higher in the model group (P<0.01). Compared with the model group, myocardial infarct size and weight were lower in the EA group (P<0.01). ②There was no significant difference between the normal group and the sham-operation group in serum contents of LDH、CK、cTnI (P>0.05). Compared with the normal group, the serum contents of LDH、CK、cTnI were higher in the model group (P<0.05). Compared with the model group, the serum contents of LDH、CK、cTnI were lower in the EA group (P<0.05). ③The expressions of FXR and SHP gene in the model group were higher than those in the normal group (P<0.05). The expressions of FXR and SHP gene in the EA group were lower than those in the model group (P<0.05). CONCLUSION: Electroacupuncture preconditioning could significantly improve cardiac function of rats with myocardial ischemia-reperfusion, reduce infarct size and inflammatory factors, and down-regulate the expressions of FXR and SHP gene. The protective effect may generate based on regulation of FXR/SHP apoptosis signal pathway.

Radix Ilicis Pubescentis total flavonoids ameliorates neuronal damage and reduces lesion extent in a mouse model of transient ischemic attack.

Flavonoids are a major component in the traditional Chinese medicine Radix Ilicis Pubescentis. Previous studies have shown that the administration of Radix Ilicis Pubescentis total flavonoids is protective in cerebral ischemia. However, to our knowledge, no studies have examined whether the total flavonoids extracted from Radix Ilicis Pubescentis prevent or ameliorate neuronal damage following transient ischemic attacks. Therefore, Radix Ilicis Pubescentis total flavonoids question and the potential underlying mechanisms. Thus, beginning 3 days before the induction of a mouse model of transient ischemic attack using tert-butyl hydroperoxide injections, mice were intragastrically administered 0.3, 0.15, or 0.075 g/kg of Radix Ilicis Pubescentis total flavonoids daily for 10 days. The results of spectrophotometric analyses demonstrated that Radix Ilicis Pubescentis total flavonoids enhanced oxygen free radical scavenging and reduced pathological alterations in the brain. Hematoxylin-eosin staining results showed that Radix Ilicis Pubescentis total flavonoids reduced hippocampal neuronal damage and cerebral vascular injury in this mouse model of transient ischemic attack. These results suggest that the antioxidant effects of Radix Ilicis Pubescentis total flavonoids alleviate the damage to brain tissue caused by transient ischemic attack.

GLYX-13, a NMDA receptor glycine-site functional partial agonist, induces antidepressant-like effects without ketamine-like side effects.

Recent human clinical studies with the NMDA receptor (NMDAR) antagonist ketamine have revealed profound and long-lasting antidepressant effects with rapid onset in several clinical trials, but antidepressant effects were preceded by dissociative side effects. Here we show that GLYX-13, a novel NMDAR glycine-site functional partial agonist, produces an antidepressant-like effect in the Porsolt, novelty induced hypophagia, and learned helplessness tests in rats without exhibiting substance abuse-related, gating, and sedative side effects of ketamine in the drug discrimination, conditioned place preference, pre-pulse inhibition and open-field tests. Like ketamine, the GLYX-13-induced antidepressant-like effects required AMPA/kainate receptor activation, as evidenced by the ability of NBQX to abolish the antidepressant-like effect. Both GLYX-13 and ketamine persistently (24 h) enhanced the induction of long-term potentiation of synaptic transmission and the magnitude of NMDAR-NR2B conductance at rat Schaffer collateral-CA1 synapses in vitro. Cell surface biotinylation studies showed that both GLYX-13 and ketamine led to increases in both NR2B and GluR1 protein levels, as measured by Western analysis, whereas no changes were seen in mRNA expression (microarray and qRT-PCR). GLYX-13, unlike ketamine, produced its antidepressant-like effect when injected directly into the medial prefrontal cortex (MPFC). These results suggest that GLYX-13 produces an antidepressant-like effect without the side effects seen with ketamine at least in part by directly modulating NR2B-containing NMDARs in the MPFC. Furthermore, the enhancement of 'metaplasticity' by both GLYX-13 and ketamine may help explain the long-lasting antidepressant effects of these NMDAR modulators. GLYX-13 is currently in a Phase II clinical development program for treatment-resistant depression.

Effect of carbonate on anaerobic acidogenesis and fermentative hydrogen production from glucose using leachate as supplementary culture under alkaline conditions.

Carbonate was added into a co-culture of glucose and fresh leachate under alkaline condition to enhance batched acidogenesis and fermentative hydrogen production simultaneously. Results indicated carbonate has positive effect on both H(2) production and acetic acid generation. The highest hydrogen yield (about 1.40 mol/mol glucose) was obtained at [CO(3)(2-)] = 280 mg/L with pH 8.0 and [CO(3)(2-)] = 560 mg/L with pH 9. The dominant liquid metabolites were ethanol, acetic and butyric acid. The highest total volatile fatty acid yield (0.38 g/g glucose) was achieved at [CO(3)(2-)] = 560 mg/L with pH 9. In this case, the acetic acid yield reached 0.13 g/g glucose. Verification tests using simulated wastewater as substrate were also carried out at pH 9. Results demonstrated calcium ions inhibit hydrogenogens activity while carbonate addition can alleviate the suppression effect caused by Ca(2+).

No primexine and plasma membrane undulation is essential for primexine deposition and plasma membrane undulation during microsporogenesis in Arabidopsis.

Primexine deposition and plasma membrane undulation are the initial steps of pollen wall formation. However, little is known about the genes involved in this important biological process. Here, we report a novel gene, NO PRIMEXINE AND PLASMA MEMBRANE UNDULATION (NPU), which functions in the early stage of pollen wall development in Arabidopsis (Arabidopsis thaliana). Loss of NPU function causes male sterility due to a defect in callose synthesis and sporopollenin deposition, resulting in disrupted pollen in npu mutants. Transmission electronic microscopy observation demonstrated that primexine deposition and plasma membrane undulation are completely absent in the npu mutants. NPU encodes a membrane protein with two transmembrane domains and one intracellular domain. In situ hybridization analysis revealed that NPU is strongly expressed in microspores and the tapetum during the tetrad stage. All these results together indicate that NPU plays a vital role in primexine deposition and plasma membrane undulation during early pollen wall development.