AIM:To observe the effect of folic acid(FA)on C2C12 myoblast proliferation and differentia-tion,and to explore its mechanism.METHODS:During the proliferation stage,C2C12 myoblasts were treated with vari-ous concentrations of FA(0,2.5,5,10 and 20 μmol/L).The cell status was observed under a microscope,cell viability was detected using the MTT method,and cell proliferation was assessed using the EdU method.In the differentiation stage,C2C12 cells were divided into control(Ctrl)group(0 μmol/L FA)and FA group(10 μmol/L FA).On day 2 or 4 of differentiation,immunofluorescence staining and Western blot were employed to detect the expression of myoblast differen-tiation-related proteins,myoblast determination protein 1(MyoD),myogenin(MyoG)and myosin heavy chain(MyHC).The myotubule formation in each group was analyzed.On day 4 of differentiation,C2C12 cells were treated with FA for 0,1,3 and 6 h,and the protein levels of p-JNK,JNK,p-p38 MAPK and p38 MAPK at each time point were detected by Western blot.Additionally,C2C12 cells after 4-day differentiation were divided into Ctrl group,FA group,FA+ SP600125(specific inhibitor of JNK)group,and FA+SB203580(specific inhibitor of p38)group.The cells in FA+ SP600125 and FA+SB203580 groups were treated with 10 μmol/L SP600125 or SB203580 for 1 h,followed by treatment with 10 μmol/L FA for 24 h.The cells in FA group were treated with 10 μmol/L FA for 24 h,while the cells in Ctrl group were left untreated.The protein levels of p-JNK,JNK,p-p38 MAPK,p38 MAPK and MyHC were detected by Western blot.RESULTS:(1)Compared with 0 μmol/L FA group,the number of the cells in other concentration groups in-creased,cell viability was raised(P＜0.05 or P＜0.01),and the rate of EdU positive cells increased(P＜0.05).(2)Com-pared with Ctrl group,the expression levels of MyoD,MyoG and MyHC in FA group were increased(P＜0.05),and the myotube fusion index was raised(P＜0.05 or P＜0.01).(3)Compared with 0 h group,the ratios of p-JNK/JNK and p-p38 MAPK/p38 MAPK were elevated after FA treatment for 1,3 and 6 h(P＜0.05 or P＜0.01),and showed a trend of gradual increase with the extension of treatment time.(4)After FA treatment,the ratios of p-JNK/JNK and p-p38 MAPK/p38 MAPK,and the expression of MyHC were elevated(P＜0.01).Treatment with SP600125 decreased the ratio of p-JNK/JNK and the expression of MyHC(P＜0.05),while SB203580 intervention cut down the ratio of p-p38 MAPK/p38 MAPK and the expression of MyHC(P＜0.05 or P＜0.01).CONCLUSION:Folic acid can promote the differentiation of C2C12 myoblasts by activating the JNK/p38 MAPK signaling pathway.

OBJECTIVE To optimize extraction technology of couplet medicinals of Astragalus membranaceus-Puerariae lobatae. METHODS With contents of puerarin，daidzin，calycosin-7-O-β-D-glucopyranoside，daidzein，calycosin and formononetin and the yield of dry extract as index，the analytic hierarchy method was used to determine the weight coefficient of each index and calculate the comprehensive score. The effects of solid-liquid ratio， extraction times and extraction time on the comprehensive score were investigated by single factor test. The level of each factor was determined. By multi-index comprehensive scoring method， using comprehensive scores of above 7 indexes as indexes，the extraction technology of couplet medicinals of A. membranaceus-P. lobata was optimized by orthogonal experiment，and the validation tests were conducted. RESULTS The weight coefficient for the contents of puerarin，daidzin，calycosin-7-O-β-D-glucopyranoside，daidzein，calycosin and formononetin and the yield of dry extract were respectively 0.304 7，0.065 2，0.185 8，0.185 8，0.107 8，0.107 8 and 0.042 7. The optimal extraction technology was determined as follows： solid-liquid of 1∶8（g/mL），extracting 3 times and for 1 h each time. RSD of each evaluation index in the validation test results was lower than 3.00% （n＝3）. CONCLUSIONS The optimized extraction technology for A. membranaceus-P. lobata is stable and feasible.

OBJECTIVE To establish the fingerprint of Guiqi tongmai mixture and evaluate its quality in combination with chemical pattern recognition . METHODS Using narirutin as reference peak ，HPLC fingerprint of Guiqi tongmai mixture was established with Similarity Evaluation System of Chromatographic Fingerprint of TCM （2012 edition）. Common peaks identification and similarity evaluation were conducted . Hierarchical cluster analysis （HCA），principle component analysis （PCA）and orthogonal partial least square -discriminate analysis （OPLS-DA）were performed ，and the value of variable importance in the projection （VIP） greater than 1 was taken as the standard to screen the differential markers affecting the quality of Guiqi tongmai mixture . RESULTS Nineteen common peaks in the HPLC fingerprints of thirteen batches of Guiqi tongmai mixture were calibrated ，and the similarities of them were 0.928-0.990；eight common peaks were identified ，including the peak 7（hydroxysafflor yellow A ），peak 8（albiflorin），peak 9（paeoniflorin），peak 10（calycosin-7-O-glucoside），peak 12（liquiritin），peak 14（ammonium glycyrrhizinate ）， peak 16（narirutin）and peak 19（hesperidin）. According to the results of HCA ，thirteen batches of samples could be clustered into three categories ：S5，S9 and S 11-S13 were clustered into one category ；S4 and S 8 were clustered into one category ；S1-S3，S6- S7 and S 10 were clustered into one category . The results of PCA indicated that the accumulative variance contribution rate of the principle components 1-7 was 92.115%. The results of OPLS -DA showed that VIP values of narirutin ，paeoniflorin，liquiritin， ammonium glycyrrhizinate ，the components corresponding to peak 11 and peak 1 were greater than 1. CONCLUSIONS HPLC fingerprint of Guiqi tongmai mixture is established ，and it could be used for quality evaluation of Guiqi tongmai mixture by combining with chemical pattern recognition techniques . Six components such as narirutin are differential markers affecti ng the quality of Guiqi tongmai mixture .

OBJECTIVE：To study the effects of gentiopicroside on the apo ptosis o f human pancreatic cancer cells PANC- 1，and to explore its mechanism from the perspective of IL- 6/JAK2/STAT3 signaling pathway. METHODS ：Using PANC- 1 cells as model ， the proliferation inhibition rate of cells was tested by MTT assay after treated with 0（negative contro ），2，4，8，16，32，64，128 mg/L gentiopicroside for 72 h and IC 50 were calculated. The cells were divided into negative control group ，gemcitabine group （positive control，4 mg/L）and gentiopicroside low-concentration ，medium-concentration and high-concentration groups （15，30，60 mg/L）. After cultured for 1，3，5，7 d，Trypan blue exclusion staining was used to count the survival cell ，and the growth of cells was investigated. After cultured for 72 h，colony formation assay was used to observe colony formation rate of cells ；the apoptotic rate of cells was detected by Hoechst 33258 staining；the mRNA and protein expressions of IL- 6，JAK2，STAT3 in cells were detected by RT-PCR and Western blotting assay. RESULTS ：4-28 mg/L gentiopicroside could inhibit the proliferation of cells （P＜0.05 or P＜ 0.01），in concentration dependent trend ；IC50 was 9.54 mg/L. Compared with negative control group ，survival cell count （cultured from 3，5，7 d），mRNA and protein expressions of IL- 6，JAK2 and STAT 3 in cells were decreased significantly in gemcitabine group ， gentiopicroside medium-concentration and high-concentration groups （P＜0.05 or P＜0.01），while the apoptotic rate was increased significantly （P＜0.01）. The colony formation rate of cellswere decreased significantly in gemcitabine group and gentiopicroside high-concentration group （P＜0.01）. mail：hb.gz@163.com Compared with gemcitabine group ，there was no statistical significance in above indexes of gentiopicroside high- 6716008。 concentration group （P＞0.05）. CONC LUSIONS：30，60 mg/L gentiopicroside could inhibit the proliferation and induce apoptosis of PANC- 1 cells，and 60 mg/L gentiopicroside is similar to gemcitabine in the effects. Its mechanism may be related to inhibiting the activation of IL- 6/JAK2/STAT3 signaling pathway.

OBJECTIVE：To establish HPLC fingerprint of Schisandra sph enanthera and S. chinensis，and to analyze chemical pattern recognition. METHODS ：HPLC method was adopted. Using schizandrin A as reference ，HPLC fingerprints of 10 batches of S. sphenanthera and S. chinensis （N1-N10，S1-S10） were drawn. Similarity Evaluation System of TCM Chromatographic Fingerprint（2012 edition）was adopted for similarity evaluation to determine the common peaks. SPSS 20.0 and SIMCA 14.1 software were used for HCA ，unsupervised madel of PCA ，supervised model of OPLS-DA. Using variable importance projection （VIP）value greater than 1 as the standard ，the differential markers that affected the quality of S. sphenanthera and S. chinensis were screened. RESULTS ：S. sphenanthera and S. chinensis were identified 32 and 33 common peaks ，respectively. The similarity of 10 batches of S. sphenanthera and 10 batches of S. chinensis were all higher than 0.9，and the similarity of S. sphenanthera and S. chinensis was 0.05. A total of 19 characteristics peaks were identified ，among which five common peaks were identified as schisandraol A ，schisandraol B ，schisantherin A ，schizandrin A and schisandrin B by reference. HCA results showed that N 1-N10 were clustered into one category ，and S 1-S10 were clustered into one category ，of which N 1，N3，N8，and N 9 were clustered into one category ，and the rest were clustered into one category ；S1，S3，S6，and S 9 were grouped together ，and the rest were grouped together. The results unsupervised model of PCA showed that the cumulative variance contribution rate of the first two principal component factors was 87.20％. Supervised model of OPLS-DA showed that schizandrin A ，schisandraol A ，schisantherin A and schisandrin B were the differential markers that affected 、the quality of S. sphenanthera and S. chinensis （VIPs were 2.29，2.24，1.73，1.48，respectively）. CONCLUSIONS ：The established fingerprint is accurate ，scientific，simple and easy to use ，combined with multivariate statistical analysis can be 话：0395-3356116。E-mail：wangrui56116@163.com used to evaluate the quality of S. sphenantherae and S. chinensis. The components of S. sphenanthera and S. chinensis were different ，schisanolrin A is differential marker.

OBJECTIVE： To study analgesic and anti-inflammatory effects of sophocarpine （SC） on inflammatory pain model mice and related COX-2/PGE2 signaling pathway. METHODS： （1）Analgesic experiment. Totally 50 mice were randomly divided into blank control group （normal saline）， positive control group （aspirin， 100 mg/kg） and SC high-dose， medium-dose and low-dose groups （40， 20， 10 mg/kg）， with 10 mice in each group. They were given relevant medicine once a day intragastrically for consecutive 5 d. 2 h after last medication， mice in each group was given glacial acetic acid solution intraperitoneal injection； writhing times of mice within 15 minutes were recorded. Other 50 mice were collected； they were grouped and given medicine as above. The response pain threshold （Tr） of mice was determined by intelligent hot plate instrument 15， 30， 60， 120 min after last administration. （2）Anti-inflammatory and mechanism experiment. Other 60 mice were randomly divided into blank control group （normal saline）， model control group （normal saline）， positive control group （aspirin， 100 mg/kg）， SC high-dose， medium-dose and low-dose groups （40， 20， 10 mg/kg）， with 10 mice in each group； they were given relevant medicine intragastrically， once a day， for consecutive 5 d. 60 min after last medication， except for blank control group， other groups were given 1％ carrageenan to induce inflammation. 1， 3， 5 h after inducing inflammation， the degree of paw swelling were determined in each group. Other 30 mice were randomly divided into blank control group （normal saline）， model control group （normal saline）， SC group （40 mg/kg）， with 10 mice in each group； they were given relevant medicine intragastrically once a day， for consecutive 5 d. 60 min after last medication， except for blank control group， other groups were given 1％ carrageenan to induce inflammation in other groups. 5 h later， the levels of SOD， MDA， GSH-Px and T-AOC in paw swelling tissue of mice were determined by biochemical method. The level of PGE2 in paw swelling tissue was determined by ELISA. The mRNA and protein expressions of COX-1 and COX-2 in paw swelling tissue of mice were detected by RT-PCR and Western blot method. RESULTS： In analgesic experiment， compared with blank control group， writhing times of mice were decreased significantly in administration groups （P＜0.05 or P＜0.01）， Tr were increased significantly 30， 60， 120 min after last medication （P＜0.05 or P＜0.01）. In anti-inflammatory and mechanism experiment， compared with blank control group， the degree of paw swelling were increased significantly in model control group 1， 3， 5 h after inducing inflammation （P＜0.01）； the levels of SOD， GSH-Px and T-AOC in paw swelling tissue were decreased significantly （P＜0.01）； the levels of MDA and PGE2 were increased significantly （P＜0.01）， and mRNA and protein expressions of COX-2 were increased significantly （P＜0.01）. Compared with model control group， the degree of paw swelling were decreased significantly in positive control group， SC high-dose and low-dose groups 3 and 5 h after inducing inflammation （P＜0.05 or P＜0.01）. The levels of SOD， GSH-Px and T-AOC in paw swelling tissue were increased significantly in SC group （P＜0.05 or P＜0.01）， while the levels of MDA and PGE2 were decreased significantly （P＜0.01） as well as mRNA and protein expressions of COX-2 were decreased significantly （P＜0.05）. There was no statistical significance in other indexes （P＞0.05）. CONCLUSIONS： SC possesses good anti-inflammatory and analgesic effects， and its mechanism may be related to anti-oxidative stress and inhibition of COX-2/PGE2 signaling pathway.

Aim To explore whether total flavonoids of Verbena officinalis L(TFV)can induce apoptosis of HepG-2 cells through targeting topoisomerase Ⅱ.Methods HepG-2 cells were cultured with TFV at 200,100,50 mg·L-1.Cell apoptosis was evaluated by TUNEL-DAPI double staining.TopⅡ activity was detected by supercoiled pBR322 DNA relaxation assay.The levels of the mRNA of TOP Ⅱα,TOP Ⅱβ were analyzed by real time PCR.Expression of Bcl-2,Bax,TOP Ⅱα,TOP Ⅱβ and caspase-3 was analyzed by Western blot.

Aim To observe the ameliorating effects of sophocarpine on mice with neuropathic pain induced by spared nerve injury(SNI), and to explore the analgesic mechanism.Methods Sixty male ICR mice were randomly divided into six groups: sham-operated, neuropathic-pain model(SNI), SNI+Pregabalin(Pre) 10 mg·kg-1, SNI+SC 40 mg·kg-1, 20 mg·kg-1 and 10 mg·kg-1 group.Neuropathic-pain mouse models were established by SNI of the sciatic nerve.The paw withdrawal mechanical threshold(PWMT), paw withdrawal thermal latency(PWTL), and cold withdrawal times(CWT) were detected to assess the analgesic effects of sophocarpine on mice with neuropathic pain induced by spared nerve injury.The expressions levels of TLR4, p38 MAPK, IL-1β and TNF-α mRNA and protein in spinal cord tissue were detected by reverse transcription-polymerase chain reaction(RT-PCR) and Western blot methods.Results Compared with sham group, the SNI group′s PWMT decreased, PWTL was shorten, CWT increased, and the expression levels of p38 MAPK, TLR4, TNF-α and IL-1β mRNA and protein in spinal cord tissue were up-regulated.Compared with SNI model group, the sophocarpine 40 mg·kg-1 group′s PWMT increased, PWTL was lengthened, CWT decreased, and the expression levels of p38 MAPK, TLR4, TNF-α and IL-1β mRNA and protein in spinal cord tissue were down-regulated.Conclusion Sophocarpine has the analgesic effects on neuropathic pain induced by SNI, and its mechanism may be related to the down-regulation of TLR4, p38 MAPK, IL-1β and TNF-α expression levels.

Objective:To amendment a tool of theory of mind (TOM) tests that can be applied to children with autism spectrum disorder (ASD) in mainland China,and assess the ability of ToM of children with ASD.Methods:The items of the ToM tests were revised by observing and recording events of children's life.Totally 200 normal children were selected in line with the standard from kindergarten to Grade 6 for formal tes-ting.With 156 valid data,Pearson correlation coefficient and Cronbach α coefficient were established for the test.Three experts were invited to make sure the content validity.Researchers randomly selected 30 normal children for retest purpose after 2 months.Twenty five children with ASD were tested and compared with normal children's test scores.Results:The revised test included 39 entries,which was divided into three sub-tests,42 points in total.The correlation coefficient of three subtests of the tests was 0.54-0.77,the correlation coefficient between the test and the subtest was 0.62-0.93 (Ps ＜ 0.01).The scores of three experts for the test were 114,108,and 105.The total scores and subtest scores were lower in children with ASD than in normal children (Ps ＜0.01).The Cronbach a coefficient of the test was 0.84,Cronbach a coefficients for three subtest were 0.83,0.80,and 0.78,respectively.The retest reliability was 0.84,and reliabilities for three subtest were 0.75,0.74,and 1.00.Conclusion:The revised Theory of Mind Tests for Children with Autism Spectrum Disorder are fulfilled mostly in line with psychometric testing requirement.It might be a selection to measure the ability of theory of mind of children with ASD in mainland China.

Objective To investigate the effects of fluvastatin (Flu) on the levels of estradiol(E2 ) ,tumor necrosis factor-α(TNF-α) ,insulin-like growth factor-1(IGF-1) and bone glaprotein(BGP) in experimental osteoporosis rats .Methods The model of osteo-porosis was established ,all rats were ovariectomized .The rats were given Flu(20 ,10 ,5 mg/kg) through intragastric administration (ig) .After treating for 12 weeks ,weighed all rats ,then sacrificed all rats ,removed the uterus and weighed its wet weight ,and calcu-lated uterus index ;the levels of E2 ,TNF-α,IGF-1 and BGP in serum were detected .Results Flu did not affect the uterus weight in experimental osteoporosis rats ;20 mg/kg Flu could increased the osteoporosis rat′s serum levels of E2 and IGF-1(P<0 .05) ,de-creased the levels of TNF-αand BGP(P<0 .01);10 mg/kg Flu could decreased the osteoporosis rat′s serum levels of BGP .Conclu-sion Flu could increased the levels of E2 ,IGF-1 ,and decreased the levels of TNF-α,BGP in experimental osteoporosis rats .