Search for Snail Repellents: Antimollusc Activities from Stemona parviflora and Six Other Chinese Stemona Species.

Snails are important agricultural pests difficult to control, but data regarding molluscicidal assays are scant. Stemona alkaloids are typical secondary metabolites for the taxa and have been broadly investigated for their pharmacological and toxicological effects. This makes it possible for us to further develop the toxicities of these compounds to snails. In this work, we tested the antifeedant properties of leaves from seven Chinese Stemona species against the land snail species Bradybaena ravida in choice and non-choice feeding assays. The tested leaves Stemona parviflora exhibited the most deterrent effects, and a further phytochemical investigation of aerial parts led to the identification of 16 alkaloids. Among them, three novel alkaloids could be identified. The alkaloidal fraction and single alkaloids were further assayed against this snail species, and the results suggest a cocktail effect because the impact of the alkaloidal fraction was higher than the effects caused by single alkaloids. The study can promote the search process of natural antimollusc products from plants to control snails.

Discovery of ent-labdane derivatives from Andrographis paniculata and their anti-inflammatory activity.

The plant Andrographis paniculata has a long history of cultivation in Southeast Asia, especially its extensive anti-inflammatory activity, and the famous natural antibiotic andrographolide comes from this plant. In China, A. paniculata, as the main crop, has become a major source of traditional Chinese medicine (TCM) for the clinical treatment of inflammation. To further explore the diverse diterpene lactones with better anti-inflammatory activity from A. paniculata, twenty-one ent-labdanes, including six undescribed compounds (andropanilides D-I), were isolated. Their structures with absolute configurations were thoroughly determined by comprehensive NMR spectroscopic data, HRESIMS analysis and quantum chemical calculations. All isolated compounds were evaluated for anti-inflammatory activities based on the Griess method. Meanwhile, after structure-activity relationships analysis, the anti-inflammatory activity of andropanilide D (1) (IC50 = 2.31 µM) was found to be better than that of the positive control drug (dexamethasone, IC50 = 6.52 µM) and andrographolide (IC50 = 5.89 µM). Further mechanisms of activity indicated that andropanilide D significantly reduced the secretion of TNF-α, IL-6 and IL-1ß and downregulated the protein expression of COX-2 and iNOS in LPS-induced RAW264.7 macrophages in a concentration-dependent manner based on Western blot and ELISA experiments. In conclusion, andropanilide D possesses potential medicinal value for the treatment of inflammation and further expands the material basis of the anti-inflammatory effect of A. paniculata.

Integrative pharmacology reveals the mechanisms of Erzhi pills, A traditional Chinese formulation, stimulating melanogenesis.

ETHNOPHARMACOLOGICAL RELEVANCE: Erzhi pills (EZP), a traditional Chinese medicine formula prescribed for the treatment of vitiligo, has shown promising efficacy. However, the oral bioactive components and mechanisms underlying the promotion of melanogenesis by EZP remain unclear. AIM OF THE STUDY: This study aimed to investigate the pharmacological basis and mechanism of EZP in promoting melanogenesis. MATERIALS AND METHODS: UHPLC-TOF-MS analysis was used to identify absorbed phytochemicals in serum after oral administration of EZP. Network pharmacology methods were used to predict potential targets and pathways involved in the melanogenic activity of EZP, resulting in the construction of a "compound-target-pathway" network. Zebrafish and B16F10 cells were used to evaluate the effects of EZP on tyrosinase activity and melanin content. Western blot and ELISA analyses were used to validate the effects of EZP on melanogenesis-related proteins, including MITF, TYR, CREB, p-CREB, and cAMP. RESULTS: UHPLC-TOF-MS analysis identified 36 compounds derived from EZP in serum samples. Network pharmacology predictions revealed 89 target proteins associated with the identified compounds and closely related to vitiligo. GO and KEGG analyses indicated the involvement of the cAMP/PKA signaling pathway in the promotion of melanogenesis by EZP. Experimental results showed that EZP increased tyrosinase activity and melanin content in zebrafish and B16F10 cells without inducing toxicity. Western blot and ELISA results suggested that the melanogenic effect of EZP may be related to the activation of the cAMP/PKA signaling pathway. These results confirm the feasibility of combining serum pharmacological and network pharmacological approaches. CONCLUSIONS: EZP have the potential to increase tyrosinase activity and melanin content in zebrafish and cells possibly through activation of the cAMP/PKA pathway.

Long-term efficacy of hydrotherapy on balance function in patients with Parkinson's disease: a systematic review and meta-analysis.

Background: Hydrotherapy can improve the motor and non-motor symptoms of Parkinson's disease (PD), but the long-term effects of hydrotherapy on PD are still unclear. Objective: The purpose of this systematic evaluation and meta-analysis was to explore the long-term effects of hydrotherapy on balance function in PD patients. Methods: A systematic search of five databases was conducted to identify appropriate randomized controlled trials (RCTs) according to the established inclusion and exclusion criteria. The general characteristics and outcome data (balance, exercise, mobility, quality of life, etc.) of the included studies were extracted, and the quality of the included studies was evaluated using the Cochrane risk of bias assessment tool. Finally, the outcome data were integrated for meta-analysis. Results: A total of 149 articles were screened, and 5 high-quality RCTs involving 135 PD patients were included. The results of the meta-analysis showed positive long-term effects of hydrotherapy on balance function compared to the control group (SMD = 0.69; 95% CI = 0.21, 1.17; p = 0.005; I2 = 44%), However, there were no significant long-term effects of hydrotherapy on motor function (SMD = 0.06; 95% CI = -0.33, 0.44; p = 0.77; I2 = 0%), mobility and quality of life (SMD = -0.21; 95% CI = -0.98, 0.57; p = 0.6; I2 = 71%). Interestingly, the results of the sensitivity analysis performed on mobility showed a clear continuation effect of hydrotherapy on mobility compared to the control group (SMD = -0.80; 95% CI = -1.23, -0.37; p < 0.001; I2 = 0%). Conclusion: The long-term effects of hydrotherapy on PD patients mainly focus on balance function, and the continuous effects on motor function, mobility, and quality of life are not obvious.

Xiao Chai Hu Tang alleviates the pancreatic tumorigenesis via improving the mtDNA N6-Methyladenine modification mediated mitochondrial dysfunction in Syrian hamster model.

BACKGROUND: Pancreatic intraepithelial neoplasia (PanIN) is the most common precursor lesion of pancreatic ductal adenocarcinoma (PDAC), which is a highly malignant tumor and lack of effective treatment. Although Xiao Chai Hu Tang (XCHT) has a good therapeutic effect on pancreatic cancer patients with advanced stage, the effect and mechanism of XCHT remains unclear in pancreatic tumorigenesis. PURPOSE: To assess the therapeutic effects of XCHT on the malignant transformation from PanIN to PDAC and to reveal its mechanisms of pancreatic tumorigenesis. METHODS: Syrian golden hamster were induced by N-Nitrosobis (2-oxopropyl) amine (BOP) to establish the pancreatic tumorigenesis model. The morphological changes of pancreatic tissue were observed by H&E and Masson staining; the Gene ontology (GO) analysis the transcriptional profiling changes; the mitochondrial ATP generation, mitochondrial redox status, mitochondrial DNA (mtDNA) N6-methyladenine (6mA) level and relative mtDNA genes expressions were examined. In addition, immunofluorescence detect the cell localization of 6mA in human pancreatic cancer PANC1 cell. Using the TCGA database, the prognostic effect of mtDNA 6mA demethylation ALKBH1 expression on pancreatic cancer patients was analyzed. RESULTS: We confirmed the mtDNA 6mA levels were gradually increased with the mitochondrial dysfunction in PanINs progression. XCHT showed the effect to inhibit the occurrence and development of pancreatic cancer in Syrian hamster pancreatic tumorigenesis model. In addition, the lack of ALKBH1 mediated mtDNA 6mA increase, mtDNA coded genes down-expression and abnormal redox status were rescued by XCHT. CONCLUSIONS: ALKBH1/mtDNA 6mA mediated mitochondrial dysfunction to induce the occurrence and progression of pancreatic cancer. XCHT can improve ALKBH1 expression and mtDNA 6mA level, regulate the oxidative stress and expression of mtDNA coded genes. This study investigated a new molecular mechanism of pancreatic tumorigenesis, and revealed the therapeutic efficacy of XCHT in pancreatic tumorigenesis for the first time.

[Constructing of clinical thinking of acupuncture and moxibustion, improving the medical record writing ability of standardized training physicians: teaching experience of the standardized training textbook Acupuncture and Moxibustion for national TCM resident physicians].

To explore the methods of cultivating the clinical thinking ability of acupuncture and moxibustion in the standardized training of resident physicians, so as to improve the medical record writing ability of the regular training physicians. The clinical diagnosis and treatment of acupuncture and moxibustion has its own characteristics and can't copy the syndrome differentiation and treatment mode of TCM internal medicine. In the treatment section, Acupuncture and Moxibustion, a standardized training textbook for national TCM resident physicians, takes clinical cases as the breakthrough point and uses the problem as the guide, guides the training physicians to cultivate acupuncture and moxibustion clinical diagnosis and treatment from three aspects: disease diagnosis, syndrome diagnosis, and treatment ideas, forms a complete understanding of the disease, and improves the standardization, logicality and systematicness of medical record writing through repeated practical training.

[Consolidating the foundation, highlighting the practice and strengthening the training of clinical thinking of acupuncture and moxibustion: the thoughts of compiling the China national standardized training textbook Acupuncture and Moxibustion for residents of traditional Chinese medicine].

Based on the clinical needs and examination requirements of standardized training students, the China national standardized training textbook Acupuncture and Moxibustion for residents of traditional Chinese medicine has made innovations in the textbook content and form. In the part of meridians and acupoints, the classic original text is introduced and the main indications and operation methods of 200 commonly-used acupoints are summarized in the form of tables. In the part of acupuncture and moxibustion technique, the operating procedures are standardized and the core technical points of 20 commonly-used acupuncture and moxibustion techniques are summarized in the form of flow chart. In the part of acupuncture and moxibustion treatment, 48 typical diseases are introduced in the form of case discussion, highlighting the problem orientation and demonstrating the diagnosis and treatment procedures.

Efficacy and safety of Chinese herbal medicine Jianpi formulas for atopic dermatitis: Evidence from 11 randomized controlled trials.

BACKGROUND: Jianpi formulas have been widely used to treat atopic dermatitis (AD) by traditional Chinese medicine (TCM) practitioners, but the efficacy and safety need further evaluation. A meta-analysis of randomized controlled trials was performed to assess the efficacy and tolerability of Jianpi formulas for AD. METHODS: We searched the following databases up to October 2020: PubMed, Embase, China National Knowledge Infrastructure database (CNKI), and Wanfang database. All randomized controlled trials (RCTs) evaluated Jianpi formulas add-on effect for AD were included for meta-analysis. The primary outcome was minor remission. Secondary outcomes include complete remission, marked remission, and decrease of SCORAD. RESULTS: Totally eleven studies were included. Nine studies were included in qualitative analysis and seven were included in quantitative analysis. The common herbs were poria, glycyrrhiza uralensis, rhizoma atractylodis, atractylodes macrocephala, and semen coicis. The Jianpi formulas add-on therapy showed consistent higher effectiveness rate in complete remission (RR1.528, 95%CI 1.176-1.984. p<0.01; Q=2.31, I2 =0%), marked remission (RR 1.388, 95% CI 1.234-1.562. p<0.01; Q=3.682, I2 =0%), and minor remission (RR 1.170, 95% CI 1.105-1.239, p<0.01; Q=14.355, I2=44.27%). The decrease of the scoring atopic dermatitis (SCORAD) in Jianpi formulas was significantly higher than in control group (SMD=0.671, 95% CI 0.018 to 0.937). Jianpi formulas were generally well tolerated, with no significantly increased risk of adverse events (AE). CONCLUSIONS: The combinations of Jianpi formulas showed moderate positive add-one effects in AD patients. Further well-designed clinical trials are needed to evaluate long-term effect of this therapy.

[Effect of electroacupuncture combined with Schwann cell transplantation on remyelination of axons and neuregulin1 expression in rats with compressed spinal injury].

OBJECTIVE: To explore the effect of electroacupuncture (EA) combined with Schwann cell (SC) transplantation (SCT) on remyelination of axons and neuregulin (Nrg1) in rats with compressed spinal cord injury(CSCI),so as to explore the mechanism of EA and SCT underlying improvement of CSCI. METHODS: SD female rats were randomly divided into normal, mo-del, EA, SCT, and EA+ SCT groups (n=40 per group). A self-developed model of spinal compressed injury was adopted in this study. Rats of the model group were administrated laminectomy without treatment. Rats in the EA group were administrated EA stimulation at "Dazhui"(GV14), "Mingmen"(GV7), bilateral "Zusanli" (ST36) and "Taixi" (KI3) on the second day post-surgery for 10 min. Rats in the SC group were administrated SCT at 1 week post-surgery, and in the EA+SC group were given EA stimulation combined with SCT. The injured spinal cord tissue was obtained 0, 2, 4 and 8 weeks after compressed spinal injury. The functional recovery was assessed by Basso-Beattie-Bresnahan (BBB) score. The survivals and migration of SC after transplantation, myelination were observed by immunofluorescence. The ultrastructure of myelin in injured site was observed by transmission electron microscope，and the expression levels of glial fibrillary acidic protein (GFAP), protein zero(P0), and Nrg1 and Nrg1-ntf (cleavage protein of Nrg1) proteins of the spinal cord were detected by Western blot. RESULTS: Compared with the normal group, BBB scores in the model group was significantly decreased（P<0.05）,nervous fibers were demyelinated, numbers of normal and newborn myelination were decreased(P<0.05)，expression of P0 was significantly increased (P<0.05)，expression of GFAP was significantly increased(P<0.05)，and the expression levels of Nrg1 and Nrg1-ntf proteins were decreased(P<0.05). In comparison with the model group, the BBB scores in the EA, SCT and EA+SCT groups were significantly increased（P<0.05,P<0.01）, demyelination was improved, numbers of normal and newborn myelinations were increased(P<0.05,P<0.01)，expressions of P0 were significantly increased (P<0.05,P<0.01)，expressions of GFAP were significantly decreased(P<0.05)，and the expression levels of Nrg1 and Nrg1-ntf proteins were increased (P<0.05, P<0.01).The differences were most significant in the EA+SCT group among the three groups. CONCLUSION: EA can improve the locomotor function in CSCI rats, which may be rela-ted to its functions in promoting the survival and migration of transplanted SC and remyelination, and increasing the expressions of Nrg1 and its cleavage protein after SC transplantation.

Decreased Sexual Desire among Middle-Aged and Old Women in China and Factors Influencing It: A Questionnaire-Based Study.

OBJECTIVE: This survey was designed and conducted with an aim to present data on sexual desire and activity in Chinese women. METHODS: Between October 2013 and December 2013, we surveyed 3000 women (aged 40-65 years) at Beijing No. 2 Hospital and the Yuetan Community Health Service Center using a questionnaire. The primary outcomes included determination of sexual desire in the past 4 weeks, reasons for stopping sexual activity, and postmenopausal syndrome. The secondary outcome was determination of factors for low sexual desire. RESULTS: A total of 2400 women (mean age 54.33 ± 6.25 years; mean menopausal age 50.11 ± 3.31 years) returned the questionnaire, with 58% of women reporting lowered sexual desire and 39.3% reporting stoppage of sexual activity. Compared with the postmenopausal group, the incidence of anxiety, depressive, somatic, and vasomotor symptoms was higher in the perimenopausal group. Muscle and joint pain (45.8%) and vaginal pruritus (21.5%) were the most commonly reported menopausal and vulvovaginal symptoms, respectively. The odds of decrease in sexual desire were significantly higher with older age, menopause, presence of gynecological disease, menopausal depression symptoms, menopausal vasomotor symptoms, and vulvovaginal atrophy; only cesarean delivery (odds ratio = 0.887, P=0.018) was associated with lesser reduction in sexual desire compared with the aforementioned factors. CONCLUSION: This survey showed that a high proportion of Chinese middle-aged and old women have lowered sexual desire and activity. Lack of sexual desire is associated with multiple factors and affects the quality of life of women.

Network Pharmacology-Based Analysis on the Mechanism of Action of Ephedrae Herba-Cinnamomi Ramulus Couplet Medicines in the Treatment for Psoriasis.

BACKGROUND This study explored the mechanism of action of Ephedrae Herba-Cinnamomi Ramulus couplet medicine (MGCM) at the pharmacological level in the treatment of psoriasis. MATERIAL AND METHODS The active ingredients in MGCM were mined through literature retrieval and the BATMAN-TCM database, and potential targets were predicted. In addition, targets associated with psoriasis were acquired using multiple disease-related databases. Thereafter, an interaction network between candidate MGCM targets and the known psoriasis-associated targets was constructed based on the protein-protein interaction (PPI) data, using the STRING database. Then, the topological parameter degree was determined for mining the core targets for MGCM in the treatment of psoriasis, which also represented the major hubs within the PPI network. In addition, the core networks of targets and ingredients were constructed using Cytoscape software to apply MGCM in the treatment for psoriasis. These core targets were then analyzed for Gene Ontology biological processes and Kyoto Encyclopedia of Genes and Genomes pathway enrichment using OmicShare. RESULTS The ingredient-target core network of MGCM for treating psoriasis was constructed; it contained 52 active ingredients and corresponded to 19 core targets. In addition, based on enrichment analysis, these core targets were majorly enriched for several biological processes (immuno-inflammatory responses, leukocyte differentiation, energy metabolism, angiogenesis, and programmed cell death) together with the relevant pathways (Janus kinase-signal transducer and activator of transcription, toll-like receptors, nuclear factor kappaB, vascular endothelial growth factor, and peroxisome proliferator-activated receptor), thus identifying the possible mechanism of action of MGCM in treating psoriasis. CONCLUSIONS The present network pharmacology study indicated that MGCM alleviates various pathological factors of psoriasis through multiple compounds, multiple targets, and multiple pathways.

H2 O2 -induced oxidative stress disrupts mitochondrial functions and impairs migratory potential of human epidermal melanocytes.

Reactive oxygen species (ROS) have already been demonstrated to impede the migratory ability in non-melanocytic cell lines by depleting mitochondrial ATP production. Therefore, understanding the mitochondrial metabolic response to migration in the presence of ROS should be a key to understanding repigmentation in vitiligo. This study aimed to investigate the energy mechanism associated with the ROS-mediated attenuation of melanocyte migration. After melanocytes were pretreated with H2 O2 , their ATP production, migratory ability, ultrastructural changes and Mitochondrial Permeability Potential were analysed. The results showed that, in parallel with the decreased ATP production, the migratory ability of melanocytes was significantly inhibited by oxidative stress. Supplementation with exogenous ATP reversed the suppressed ATP-dependent migration of melanocytes. Melanocytes were then stressed with H2 O2 and Agilent Whole Human Genome microarray analysis identified 763 up-regulated mRNAs and 1117 down-regulated mRNAs. Among them, 11 of the encoded proteins were involved in mitochondrial ATP production and their expression levels were verified. The decreased expression of NADH dehydrogenase 2(ND2) , cytochrome c oxidase 1(COX1) and cytochrome c oxidase 3(COX3) was shown to be involved in the depletion of mitochondrial ATP production, which was coupled with the impaired migratory potential. These results indicate that the migration of melanocytes relies heavily on an inexhaustible supply of ATP from mitochondria.

The relationship between obstructed defecation and true rectocele in patients with pelvic organ prolapse.

We aimed to investigate the prevalence of true rectocele and obstructed defecation (OD) in patients with pelvic organ prolapse (POP), to investigate the correlation between true rectocele and OD, and to understand the diagnostic value of translabial ultrasound (TLUS) in the diagnosis of true rectocele. The patients who scheduled for POP surgery were enrolled in this study. Patients who had previous reconstructive pelvic surgery or repair of rectocele were excluded. Birmingham Bowel and Urinary symptoms questionnaires and Longo's obstructed defecation syndrome scoring system were used to assess the bowel symptoms of patients. TLUS was used to evaluate anatomical defects. P value <0.05 was considered statistically significant, and confidence intervals were set at 95%. 279 patients were included into this study. The prevalence rate of OD was 43%, and the average value of ODS score was 6.67. 17% patients presented straining at stool, 33% presented incomplete emptying, 13% presented digitations, and 12% required laxatives or enema. The prevalence rate of true rectocele was 23%. Defecation symptoms were significantly correlated with age, levator-ani hiatus, levator-ani muscle injury and true rectocele. Logistic regression showed that true rectocele and increased levator-ani hiatus were independent risk factors of OD. True rectocele was significantly correlated with straining at stool, digitation, incomplete emptying and requirement of laxatives or enema.In POP patients, the prevalence rate of true rectocele and OD was 23% and 43%, respectively. True rectocele was related to OD. TLUS was a valuable approach in anatomical evaluation of POP.

[Evidence-based Clinical Practice Guideline of Acupuncture and Moxibustion for Primary Dysmenorrhea: development and characteristics].

Evidence-based Clinical Practice Guideline of Acupuncture and Moxibustion for Primary Dysmenorrhea (Guideline) is clarified from three aspects: the development method, contents and clinical application, which could provide clinical practical strategies suitable for most patients for clinicians. Based on comprehensive literature search and summary of acupuncture practice, the best clinical research evidence is obtained according to the evidence-based method, the evidence-quality evaluation and recommendation grading are introduced. With ancient literature, famous experts' experience and expert consensus, the recommendations of acupuncture-moxibustion for primary dysmenorrhea are developed, which are safe, effective and applicable for clinical treatment. The Guideline could standardize the acupuncture treatment plan for primary dysmenorrhea.

[Electroacupuncture improves locomotor function by regulating expression of inflammation and oxidative stress-related proteins in mice with spinal cord injury].

OBJECTIVE: To observe the effect of electroacupuncture (EA) on the expression of apolipoprotein E (ApoE) and related proteins of inflammation and anti-oxidative stress in spinal cord in mice with spinal cord injury (SCI), so as to explore its mechanisms underlying function repair. METHODS: Thirty-six female C57BL/6 mice were equally randomized into 3 groups: sham operation, model and EA. The SCI model was established by clamping the spinal cord for 25 s with a serrefine after laminectomy of the 1st lumbar vertebra (L1). EA (1.5 Hz/7.5 Hz, 1.0 mA) was applied to bila-teral "Zusanli" (ST36) and "Sanyinjiao" (SP6) for 10 min, once a day for 7 days. The hindlimb locomotor function was assessed according to the state of the range of motion, coordination, claw gesture of the hind leg ankle-joint, trunk stabi-lity and the tail posture by using Basso Mouse Scale(BMS). The histopathological changes of the injured area of the spinal cord were determined by H.E. staining. The expression levels of ApoE, phosphorylated nuclear transcription factor-κB(p-NF-κB), interleukin 1 beta(IL-1ß), phosphorylated extracellular regulatory protein kinase(p-ERK1/2), extracellular regulatory protein kinase(ERK1/2), nuclear factor erythroid 2-related factor 2(Nrf2) and heme oxidase-1(HO-1) in the spinal cord were detected by Western blot, and the glial fibrillary acidic protein (GFAP)-positive astrocytes were displayed by immunofluorescence staining. RESULTS: After modeling, the BMS scores were significantly decreased in the model group compared with the sham operation group (P<0.05). Following EA, the BMS scores were markedly increased in the EA group relevant to the model group (P<0.05), suggesting an improvement of the hindlimb locomotor function. H.E. stain showed structural disorder with lots of cavities, severe inflammatory infiltration with large quantity of inflammatory cells, and apparent reduction of normal neurons in the injured spinal cord tissue of model group, which was milder in the EA group. The expression levels of ApoE, p-NF-κB, IL-1ß, p-ERK1/2 (not ERK1/2), Nrf2 and HO-1 were significantly increased in the model group than those in the sham operation group (P<0.05). Compared with the model group, the expression levels of ApoE, p-ERK1/2, Nrf2 and HO-1 were further notably up-regulated (P<0.05), and those of p-NF-κB and IL-1ß proteins obviously down-regulated in the EA group (P<0.05). Immunoflorescence staining showed that the number of GFAP-positive cells was apparently increased in the model group compared with the sham operation group and observably decreased in the EA group relevant to the model group (P<0.05). CONCLUSION: EA can significantly improve locomotor function in SCI mice, which is associated with its effects in reducing inflammation, oxi-dative stress reactions and reactive astrocyte proliferation via up-regulating expression of ApoE, p-ERK1/2, and Nrf2/HO-1 (antioxidant pathway) and inhibiting IL-1ß and NF-κB expression.

[Effect of electroacupuncture combined with Schwann cell transplantation on limb locomotor ability, regional remyelination and expression of spinal CD4 and CD8 proteins in compressive spinal injury rats].

OBJECTIVE: To observe the effect of electroacupuncture (EA) combined with transplantation of Schwann cells (SCs) on limb locomotor, myelin sheath repair and expression of CD4 and CD8 in compressed spinal cord injury (CSCI) rats, so as to explore its mechanisms underlying improvement of CSCI. METHODS: A total of 45 female SD rats were randomly divided into normal control, model, EA, Schwann cell (SC) transplantation, and EA＋SC transplantation groups (n＝9 rats in each group). The CSCI model was established by laminectomy at T12-L2 and clip compression. Rats of the SC transplantation group accepted injection of the cultured SC suspension (2×106/6 µL) into the central, upper and lower sites of the injured spinal cord (5 mm in depth) 7-8 days after CSCI modeling. EA (2 Hz) was applied to bilateral "Zusanli" (ST36) and "Sanyinjiao" (SP6) for 10 min, once daily and 6 days a week for 3 weeks. The Basso, Beattie and Bresnahan locomotor rating scale (BBB scale) was used to evaluate the function state of CSCI. Morphological changes of the regional injured tissue were observed under light microscope after H．E. staining. The myelin sheath repair state and survival of SCs were detected by Luxol fast blue (LFB) staining and immunofluorescence histochemistry, and the expression of CD4, CD8 and P0 of the injured spinal cord was detected by Western blot. RESULTS: Compared with the normal control group, the BBB scores at the time-points of 0 d, and 1, 2, and 3 weeks were significantly decreased in the model group (P<0.001), and those of the EA＋SC transplantation group at the 2nd and 3rd week were significantly higher than those of the model group (P<0.05). No significant changes of BBB scores were found after EA and SC transplantation relevant to the model group (P>0.05). LFB staining showed a disordered arrangement of the nerve fibers in the white matter, myelinociasis and obvious decrease of the medullated fibers in the model group, and these situations were relatively milder in both EA and SC transplantation groups and obviously milder in the EA＋SC transplantation group. H．E. staining displayed that the structure of the injured region of the spinal cord was incomplete, accompanied with a large number of defect cavities and neuronal karyopyknosis in the model group, while the structure was relatively clear, with an increase of the normal neurons and fewer neuronal karyopyknosis in the EA＋SC transplantation group. Compared with the normal control group, MBP in the model group was significantly decreased (P<0.001)，and P0 was significantly increased (P<0.001). Compared with the model group, the expressions of MBP and P0 were significantly increased in the EA, SC transplantation, and EA＋SC transplantation groups (P<0.01, P<0.001), and was significantly higher in the EA＋SC transplantation group than in both EA and SC transplantation groups (P<0.001). The average immunofluorescence intensity of Hoechst33342-labeled SCs was significantly higher in the EA＋SC transplantation group than in the SC transplantation group (P<0.05). After CSCI, the expression levels of spinal CD4, CD8 and P0 proteins had no significant changes in comparison with the normal control group (P>0.05), while after the intervention and in comparison with the model group, the expression levels of P0 protein were significantly increased in the EA, SC transplantation and EA＋SC transplantation groups (P<0.05), and was significantly higher in the EA＋SC transplantation group than in both EA and SC transplantation groups (P<0.05). The expression levels of CD4 and CD8 proteins were significantly lower in the EA＋SC transplantation group than in the SC transplantation group (P<0.05)．. CONCLUSION: EA＋SCs transplantation can improve the locomotor function in CSCI rats, which may be related to its effects in increasing the survival of transplanted SCs to promote the remyelination and in reducing the immune rejecting reaction.

[Effect of electroacupuncture of acupoints on the healthy limb (opposing needling) on acute skeletal muscle contusion in rats].

OBJECTIVE: To observe the therapeutic effect of electroacupuncture (EA) of "Zusanli" (ST36) and "Ashi"-point on the healthy side (opposing needling) on muscular injury and expression of myogenin (myoG) and fast myosin skeletal heavy chain (Fast MyHC) proteins in the gastrocnemius muscle (GM) tissues in skeletal muscle contusion rats，so as to explore its mechanism underlying improvement of skeletal muscle injury. METHODS: A total of 54 male SD rats were divided into normal control (n ＝ 6)，model (n＝24) and opposing needling (EA, n＝24) groups. The latter two groups were further randomized into 3, 5, 7 and 14 d subgroups (n＝6 per subgroup). The skeletal muscle contusion model of the hind-limb was established by using a self-made striking device. EA (1 Hz/3 Hz，1－2 mA) was applied to ST36 and "Ashi"-point on the uninjured side of the hind-limb for 15 min every time, once a day for 3, 5, 7 and 14 days, respectively. The injured GM was harvested on the 3rd, 5th, 7th and 14th day after muscular contusion. The morphological changes of the injured GM and the mean cross-sectional areas (CSAs) of the neonatal muscle cells were observed by microscope after H．E. staining. The immunoactivity of desmin protein (myogenic marker protein of myoblast cell) of GM was detected by immunofluorescence stain on the 7th day after injury, and the expression levels of myoG (on the 3rd and 5th day after injury) and fast MyHC protein of GM tissues (on the 7thand 14th day after injury) were detected by Western blot. RESULTS: H．E. staining of GS tissue showed fewer neuronal myocytes with disordered arrangement at different sizes, and appearance of some collagenous fibers among the mesenchyme on day 7 and 14 after muscular contusion, which was relatively milder in the EA group. In the EA group, the CSA values of the neonatal muscle cells were significantly larger than those in the model group on the day 7th (P<0.05), 14th (P<0.001) after injury. On day 7 after muscular contusion, the desmin was found to express on the cellular membrane of GM in the normal control group, while in the model group, the desmin expressed mainly in the cellular plasma in the model group, and on the cellular membrane of neonatal myocytes in the EA group, respectively. The desmin positive myocytes showed disordered arrangement and different sizes after muscular contusion, whereas the situations of the EA group were close to those of the normal control group. Desmin expression was up-regulated in the EA group compared with the model group which was not significant difference (P>0.05). On the 3rd and 5th day after muscular contusion, the expression level of myoG protein was significantly up-regulated in the model group compared with the normal control group (P<0.001), and significantly up-regulated in the EA group than that in the model group (P<0.001). On the 7th and14th day after contusion, the expression level of fast MyHC protein was significantly down-regulated in the model group relevant to the normal control group (P<0.001), and markedly up-regulated in the EA group relevant to the model group (P<0.01)．. CONCLUSION: EA of ST36 and "Ashi"-point on the contralateral limb can up-regulate the expression of myoG and fast MyHC proteins of GM in acute skeletal muscle contusion rats, which may contribute to its effect in promoting the repair of skeletal muscle injury.

[Electroacupuncture improves denervated gastrocnemius atrophy by regulating expression of fork-head protein 3A，muscle atrophy F-box and myogenic differentiation antigen proteins in sciatic nerve injury rats].

OBJECTIVE: To observe the effect of electroacupuncture (EA) on morphological changes of denervated gastrocnemius(GS) and the expression of fork-head protein(FOXO3A), muscle atrophy F-box(MAFbx)and myogenic differentiation antigen (Myod1) in sciatic nerve injury rats, so as to reveal its mechanism underlying improvement of myoatrophy. METHODS: Eighteen male Sprague-Dawley rats were randomly divided into sham operation, model and EA groups (n＝6 per group). The model of gastrocnemius atrophy was established by crushing the right sciatic nerve. Then, EA (2 Hz) was applied to the right "Zusanli" (ST36) and "Huantiao" (GB30) for 10 min, once a day for 14 successive days. The wet weight of the GS on both sides was weighted to calculate the wet weight ratio (the injured side /the healthy side), and the cross-sectional area (CSA) and diameter of GS fibers were measured after H．E. staining. The expressions of FOXO3A, MAFbx and Myod1 protein and mRNA in the GS tissue were tested using Western blot and fluorescence quantitative PCR, separately. RESULTS: Following modeling, the GS wet weight ratio, CSA and fiber diameter were smaller in the model group than those in the sham group (P<0.01), and were significantly higher in the EA group than in the model group (P<0.01). H．E. staining showed that the GS fibers became smaller and the myocyte got round in the model group, while the GS fibers were bigger and the myocyte was relatively regular in morphology in the EA group. After modeling, the expression levels of FOXO3A, MAFbx and Myod1 mRNA and protein were evidently higher in the model group (P<0.01); Moreover, after EA treatment, modeling-induced increasing of expression levels of FOXO3A and MAFbx mRNA and protein were revised (P<0.01), while the increased expression level of Myod1 was further up-regulated relavant to that in the model group (P<0.01)．. CONCLUSION: EA of ST36 and GB30 can suppress the up-regulated expression of FOXO3A and MAFbx mRNA and protein and further promote the expression of Myod1 mRNA and protein in the GS tissue in rats with denervated GS atrophy, which may contribute to its function in relieving the myoatrophy, promoting the skeletal muscle protein hydrolysis and differentiation of satellite cells.

[Serum metabolic profile involving protective effect of "Neiguan"(PC6)-electroacupuncture preconditioning in rats with myocardial ischemia reperfusion injury].

OBJECTIVE: To observe the effect of "Neiguan" (PC6)-electroacupuncture (EA) preconditioning on serum metabolites in myocardial ischemia-reperfusion injury (MIRI) rats, so as to reveal its mechanism underlying improvement of ischemic myocardium from metabonomics. METHODS: A total of 48 male SD rats were randomly divided into control, model, EA "Neiguan"(PC6) and EA "Hegu"(LI4) groups (n＝12 rats/ group). Rats of the control group were just banded on animal boards for 30 min, once daily for 7 days. The MIRI model was established by occlusion of the left anterior descending branch of the left coronary artery for 40 min, followed by reperfusion for 1 h, and rats of the model group were also banded as those in the control group. Before modeling, EA (10 Hz/50 Hz, 1 mA) was applied to bilateral "Neiguan"(PC6) and "Hegu"(LI4) for 30 min, once daily for 7 successive days. After the treatment, serum samples were collected to be analyzed by proton nuclear magnetic resonance (1H NMR) spectroscopy. The orthogonal partial least squares discriminate analysis (PLS-DA) was employed to distinguish the serum differential metabolic profile of rats in different groups and identify potential biomarkers. RESULTS: After modeling, the ECG of model group and electroacupuncture groups showed T wave towering, and there was no obvious ST segment between R wave and T wave. The T wave decreased more than 0.2 mV after reperfusion, and there was no obvious ST segment. Compared with the control group, MIRI induced significant changes of metabolites in the serum including increase of acetoacetate acid, lectic acid, creatine, glycerol and glucose, and decrease of alanine, glutamine, glycerophosphoryl choline and phosphorylcholine. In comparison with the model group, PC6-EA preconditioning induced significant changes, including an increase of glucose, and a decrease of leucine，isoleucine, valine，3-hydroxybutyric acid，lactate，acetate，acetone，acetoacetate acid，pyruvic acid，glutamine，creatine and glycerol. There is no significant difference in metabolic patterns between "Hegu" group and model group. Metabolic pathway enrichment analysis indicated that the protective effect of PC6-EA pretreatment was realized mainly by regulating pathways of glycolysis, gluconeogenesis, citric acid metabolism, pyruvate metabolism, ketone body metabolism, etc. CONCLUSION: PC6-EA pretreatment has a role in regulating gluconeogenesis, pyruvate metabolism, amino metabolism, ketone body metabolism and energy metabolism in rats with MIRI, which maybe contribute to its protective effect on ischemic myocardium, but the specific metabolic pathways and mechanisms need being studied further.

[Effect of electroacupuncture on amyotrophia and expression of myogenic differentiation-related genes of gastrocnemius in rats with chronic constriction injury of sciatic nerve].

OBJECTIVE: To explore the effect of electroacupuncture(EA) on amyotrophia and expression of paired box7(Pax7), myogenic differentiation antigen-1 (Myod1), myogenin (Myog), myosin heavy chain- Ⅱa (Myh2), myosin heavy chain-Ⅱx (Myh1) and myosin heavy chain-Ⅰ (Myh7) of denervated gastrocnemius in rats with chronic constriction injury (CCI) of the right sciatic nerve, so as to explore its mechanisms underlying postponing development of amyotrophia. METHODS: Sixty-six SD adult male rats were randomly divided into sham operation (sham) group (n＝24), model group (n＝24) and EA group (n＝18). The denervated muscle (gastrocnemius) atrophy model was established by CCI of the right sciatic nerve. EA (2 Hz，1.0 mA) was applied to the right "Zusanli"(ST36) and "Huantiao"(GB30) for 10 min, once a day, six times a week and for 1, 2 and 4 weeks, respectively. After complete dissection of bilateral gastrocnemius muscles, their wet weight levels were measured and the ratio of wet weight (＝that of the operation side/that of the non-operation side) was calculated, and the cross-sectional area (CSA) and diameter of the gastronemius were detected after fixation in 4% paraformaldehyde, sectioning, and H．E. staining. The expression levels of Pax7, Myod1, Myog, Myh2, Myh1 and Myh7 mRNAs in the gastrocnemius tissue after 3 weeks of modeling were detected with quantitative real time-PCR (qPCR). RESULTS: After 1 week of modeling, the ratios of wet weight of gastrocnemius and the CSA and fiber diameter at the 2nd, 3rd and 5th week were significantly smaller in the model group than in the sham group (P<0.05). The expression levels of Myod1 and Myog mRNAs were significantly up-regulated (P<0.01), and those of Myh2, Myh1 and Myh7 considerably down-regulated in the model group compared with the sham group (P<0.05, P<0.01). No significant changes were found in the expression levels of Pax7 mRNA after modeling and EA intervention (P>0.05)．Following EA intervention, the CSA and diameterof the gastronemius were obviously increased (P<0.05), and the expression levels of Myod1, Myog and Myh7 further markedly or remarkably up-regulated in comparison with the model group (P<0.05, P<0.01). No significant changes were found in the ratio of wet weight of gastrocnemius at the 3 time-points, and the expression levels of Myh2 mRNA and Myh1 mRNA in the EA group relevant to the model group after 3 weeks of modeling (P>0.05). CONCLUSION: EA treatment may delay gastrocnemius atrophy in CCI rats, which is possibly associated with its effects in up-regulating the expression of Myod1, Myog and Myh7 mRNAs to control the differentiation of the satellite cells and the muscle fiber type transformation.