Prokaryotic expression, subcellular localization and enzymatic activity analysis of DXS gene from Platycodon grandiflorum / 药学学报

1-Deoxy-D-xylulose-5-phosphate synthase (DXS), the first key enzyme in 2-methyl-D-erythritol-4-phosphate (MEP) pathway, catalyzes the condensation of glyceraldehyde-3-phosphate with pyruvate to 1-deoxy-xylose-5-phosphate (DXP). In this study, PgDXS1, PgDXS2, and PgDXS3 genes were cloned from the root of Platycodon grandiflorum (P. grandiflorum). The open reading frame (ORF) of PgDXS1, PgDXS2, and PgDXS3 were 2 160, 2 208, and 2 151 bp in full length, encoding 719, 735, and 716 amino acids, respectively. Homologous alignment results showed a high identity of PgDXSs with DXS in Hevea brasiliensis, Datura stramonium and Stevia rebaudiana. The recombinant expression plasmids of pET-28a-PgDXSs were constructed and transformed into Escherichia coli (E. coli) BL21 (DE3) cells, and the induced proteins were successfully expressed. Subcellular localization results showed that PgDXS1 and PgDXS2 were mainly located in chloroplasts, and PgDXS3 was located in chloroplasts, nucleus and cytoplasm. The expression of three DXS genes in different tissues of two producing areas of P. grandiflorum were assayed via real-time fluorescence quantitative PCR, and the results showed that all of them were highly expressed in leaves of P. grandiflorum from Taihe. Under methyl jasmonate (MeJA) treatment, the expression levels of three PgDXS genes showed a trend of first decreasing and then increasing at different time points (3 - 48 h), and the activity of DXS showed a trend of first increasing and then decreasing in three tissues of P. grandiflorum. This study provides a reference for further elucidating the biological function of PgDXS in terpenoid synthesis pathway in P. grandiflorum.

Inhibitory effect of three strains of biocontrol microbes on pathogens causing rhizome rot of Polygonatum cyrtonema / 中国中药杂志

Rhizome rot is one of the main disease in the cultivation of Polygonatum cyrtonema, and it is also a global disease which seriously occurs on the perennial medicinal plants such as Panax notoginseng and P. ginseng. There is no effective control method at present. To identify the effects of three biocontrol microbes(Penicillium oxalicum QZ8, Trichoderma asperellum QZ2, and Brevibacillus amyloliquefaciens WK1) on the pathogens causing rhizome rot of P. cyrtonema, this study verified six suspected pathogens for their pathogenicity on P. cyrtonema. The result showed that Fusarium sp. HJ4, Colletotrichum sp. HJ4-1, and Phomopsis sp. HJ15 were the pathogens of rhizome rot of P. cyrtonema, and it was found for the first time that Phomopsis sp. could cause rhizome rot P. cyrtonema. Furthermore, the inhibitory effects of biocontrol microbes and their secondary metabolites on three pathogens were determined by confrontation culture. The results showed that the three tested biocontrol microbes significantly inhibited the growth of three pathogens. Moreover, the secondary metabolites of T. asperellum QZ2 and B. amyloliquefaciens WK1 showed significant inhibition against the three pathogens(P<0.05), and the effect of B. amyloliquefaciens WK1 sterile filtrate was significantly higher than that of high tempe-rature sterilized filtrate(P<0.05). B. amyloliquefaciens WK1 produced antibacterial metabolites to inhibit the growth of pathogens, and the growth inhibition rate of its sterile filtrate against three pathogens ranged from 87.84% to 93.14%. T. asperellum QZ2 inhibited the growth of pathogens through competition and antagonism, and P. oxalicum QZ8 exerted the inhibitory effect through competition. The research provides new ideas for the prevention and treatment of rhizome rot of P. cyrtonema and provides a basis for the di-sease control in other crops.

Application of 6-minute walk and grip strength tests in nursing evaluation of early arterial stiffness in community-dwelling elderly / 中国实用护理杂志

Objective:To explore the application of 6MWT and handgrip strength test in the nursing evaluation of arterial stiffness in community-dwelling elderly.Methods:From September 2019 to January 2020, 129 community-dwelling elderly people (age 60 years or older) were selected as the research subjects. Arterial stiffness was detected by pulse wave detector, and handgrip strength test and 6MWT were performed. Pearson correlation analysis and multiple linear regression methods were used to analyze the relationship between handgrip strength and 6MWT test indicators and aortic pulse wave velocity (aPWV) and augmentation index75 (AIx75).Results:The community-dwelling older people of aPWV was (10.50 ± 1.36) m/s and was negatively correlated with 6MWT distance and 6WMT work, of which the correlation coefficients were -0.404 and -0.285 respectively ( P<0.05). The community-dwelling older people of AIx75 was (28.51 ± 10.81) % and was negatively correlated with handgrip strength, relative grip strength, 6MWT distance and 6MWT work, of which the range of correlation coefficients were from -0.261 to -0.226 ( P<0.05). After adjusting for age, gender, and blood pressure, 6MWT distance and grip strength were independently related to aPWV and AIx75, respectively. Conclusions:In community-dwelling elderly people, both the grip strength test and 6MWT can be used for nursing evaluation of early arterial stiffness, but the specific evaluation role is different.

Cerebrospinal Fluid β2-Microglobulin for Prognostic Evaluation of Patients with Central Nervous System Infiltration in Acute Lymphoblastic Leukemia / 中国实验血液学杂志

OBJECTIVE@#To investigate the prognostic value of serum and cerebrospinal fluid β2-microglobulin (β2-MG) in acute lymphoblastic leukemia (ALL) with central nervous system invasion after chemotherapy.@*METHODS@#40 patients with leukemia who had been confirmed to have central nervous system infiltration were selected for treatment at the Second Affiliated Hospital of Chongqing Medical University from January 2015 to May 2017, and the serum levels of β2-MG and CSF-β2MG were dynamically monitored and performed statistical analysis.@*RESULTS@#After chemotherapy, the changes in serum β2-MG were not statistically significant (P>0.05); the absolute level of CSF-β2MG and the percentage of relative baseline changes were statistically different in different clinical outcome groups(P<0.05), and the decreasing CSF-β2MG levels suggest a better prognosis, with cut-off values of 1.505 and -25%, respectively.@*CONCLUSION@#The best cut-off point may be a predictor of complete remission; the reduction of the absolute and relative levels of CSF-β2MG can suggest a good prognosis for patients.

Q-markers of Yuquan Capsules based on serum pharmacochemistry of Chinese medicine / 中国中药杂志

This study analyzed the quality markers(Q-markers) of Yuquan Capsules(YQC) based on serum pharmacochemistry of Chinese medicine and detected the components and metabolites of YQC absorbed into the blood by UPLC-Q-TOF-MS and UNIFI systems. As a result, 32 components of YQC were detected, including 17 prototype components and 15 metabolized components. Among them, 12 prototype components(ginsenoside Rh_2, genistein, formononetin, puerarin, daidzein, schizandrin A, schizandrin B, schizandrin C, schizandrol A, schizandrol B, gomisin D, and ononin) and 12 metabolized components(ginsenoside Rg_1, ginsenoside Rg_2, ginsenoside Rg_3, ginsenoside Ro, 3'-methoxypuerarin, daidzin, astragaloside Ⅱ, astragaloside Ⅳ, glycyrrhizic acid, liquiritigenin, isoliquiritin, and verbascoside) showed inhibitory effects and pharmacological activities against diabetes, and these 24 blood-entering components against diabetes were identified as Q-markers of YQC.

Cloning and prokaryotic expression of 3-ketoacyl-CoA thiolase gene AIKAT from Atractylodes lancea / 中国中药杂志

In this study, the gene encoding the key enzyme 3-ketoacyl-CoA thiolase(KAT) in the fatty acid β-oxidation pathway of Atractylodes lancea was cloned. Meanwhile, bioinformatics analysis, prokaryotic expression and gene expression analysis were carried out, which laid a foundation for the study of fatty acid β-oxidation mechanism of A. lancea. The full-length sequence of the gene was cloned by RT-PCR with the specific primers designed according to the sequence information of KAT gene in the transcriptomic data of A. lancea and designated as AIKAT(GenBank accession number MW665111). The results showed that the open reading frame(ORF) of AIKAT was 1 323 bp, encoding 440 amino acid. The deduced protein had a theoretical molecular weight of 46 344.36 and an isoelectric point of 8.92. AIKAT was predicted to be a stable alkaline protein without transmembrane segment. The secondary structure of AIKAT was predicted to be mainly composed of α-helix. The tertiary structure of AIKAT protein was predicted by homology modeling method. Homologous alignment revealed that AIKAT shared high sequence identity with the KAT proteins(AaKAT2, CcKAT2, RgKAT and AtKAT, respectively) of Artemisia annua, Cynara cardunculus var. scolymus, Rehmannia glutinosa and Arabidopsis thaliana. The phylogenetic analysis showed that AIKAT clustered with CcKAT2, confirming the homology of 3-ketoacyl-CoA thiolase genes in Compositae. The prokaryotic expression vector pET-32 a-AIKAT was constructed and transformed into Escherichia coli BL21(DE3) for protein expression. The target protein was successfully expressed as a soluble protein of about 64 kDa. A real-time quantitative PCR analysis was performed to profile the AIKAT expression in different tissues of A. lancea. The results demonstrated that the expression level of AIKAT was the highest in rhizome, followed by that in leaves and stems. In this study, the full-length cDNA of AIKAT was cloned and expressed in E. coli BL21(DE3), and qRT-PCR showed the differential expression of this gene in different tissues, which laid a foundation for further research on the molecular mechanism of fatty acid β-oxidation in A. lancea.

Effects of sinew-regulating bone-setting manipulations on chondrocytes, IL-1β and NO in rabbits with knee osteoarthritis / 针灸推拿医学（英文版）

Objective: To observe the therapeutic effect of sinew-regulating bone-setting manipulations for knee osteoarthritis (KOA) model rabbits and its impacts on the chondrocyte apoptosis rate and the levels of interleukin (IL)-1β and nitric oxide (NO). Methods: According to the random number table method, 30 New Zealand white rabbits were divided into a normal group (n=9) and a modeling group (n=21). Rabbits in the modeling group were used to establish KOA models with the modified Hulth method. At the 8th week, three rabbits were sacrificed to verify the model and the remaining 18 rabbits were randomly divided into a model group (n=9) and an intervention group (n=9). Rabbits in the normal group and model group were bred routinely without any intervention. Rabbits in the intervention group were treated with the sinew-regulating bone-setting manipulations, 10 min/time, once every other day for a total of 20 times. The Lequesne MG knee function rating was used to evaluate the behavioral differences of the rabbits in each group. The Pelletier score was used to evaluate the general changes of the rabbits. The Mankin score was used to evaluate the pathology of knee cartilages. The enzyme-linked immunosorbent assay and nitrate reductase methods were used to determine the levels of IL-1β and NO in serum and synovial fluid of each group, respectively. In situ terminal deoxynucleotidyl transferase-mediated nick and labeling method was used to determine the apoptosis of chondrocytes in each group. Results: Compared with the normal group, the scores of Lequesne MG, Pelletier and Mankin, and the levels of IL-1β and NO in the model group were increased (P<0.05), which indirectly indicated the success of the model. Compared with the model group, the scores of Lequesne MG, Pelletier and Mankin, IL-1β and NO levels, and chondrocyte apoptosis rate of the intervention group were decreased, and the differences were statistically significant (P<0.05). Conclusion: The sinew-regulating bone-setting manipulations can reduce the levels of IL-1β, NO, and chondrocyte apoptosis rate, and delay the articular cartilage degeneration, therefore, having a good therapeutic effect on KOA.

Different development phase of transcriptome analysis from Crataegus pinnatifida Bge. and cloning, structure and function prediction of squalene epoxidase involved in triterpenic acid biosynthesis / 药学学报

italic>Crataegus pinnatifida is a traditional Chinese medicine, which contains organic acids, triterpenoid acids and other active components, has important medicinal and edible value. In order to study the difference of gene expression level in different developmental stages of hawthorn and explore the genes of active ingredient biosynthesis in Crataegus pinnatifida, high-throughput Illumina HiSeq 2000 technology were used to conduct transcriptome sequencing and bioinformatics analysis on Crataegus pinnatifida fruits from the same origin at different developmental stages. 78 496 Unigenes with an average length of 941 nt were obtained by Trinity software. Among them, 58 395 Unigenes can be annotated by NR, NT, Swiss prot, KEGG, COG, GO and other public databases. KEGG pathway analysis showed that 52 Unigenes encoding 15 key enzymes involved in the citric acid cycle. There are 62 Unigenes were involved in the triterpene biosynthesis pathway of Crataegus pinnatifida. Two key enzymes SQE of triterpenoid metabolism pathway in Crataegus pinnatifida were cloned and performed bioinformatic analysis. The results showed that ORF of CpSQE1 and CpSQE2 were 1 594 bp and 1 597 bp, respectively, encoding 530 and 531 amino acids. The molecular weight of proteins was 57.6 kDa and 57.5 kDa. Bioinformatics analysis showed that both CpSQE1 and CpSQE2 proteins have a PLN02985 superfamily conserved domain, belonging to the squalene monooxygenase superfamily. The phylogenetic tree shows that CpSQE1 and CpSQE2 are clustered together with SQE with squalene epoxidase function in other plants. This study provides a research basis for further exploring the key genes in the biosynthesis process of hawthorn active ingredients and analyzing the regulation pathway of its active ingredient biosynthesis.

Pathological Mechanisms of Brain-Heart Mutual Damage and Treatment with Traditional Chinese Medicine: A review / 中国实验方剂学杂志

The brain and heart has a tight relationship upon physiology and pathology during the development of cardiovascular and cerebrovascular diseases with clinical conditions interacting with each other and complex pathological mechanisms. Clinical studies prove that cerebral diseases such as stroke usually happen with cardiac diseases as complications， and cardiac diseases such as atrial fibrillation can also cause cerebral diseases， which can even aggravate brain atrophy and lead to cognitive impairment. Traditional Chinese medicine （TCM） also believes that “the substance of Shenming is located in the brain and the function in the heart.” Specifically， Yuanshen is in the brain and Shishen is in the heart， which thus makes the heart and brain closely related. If Shenming in any of them is impaired， the other one would also be injured. Therefore， the pathological mechanisms of brain-heart mutual damage have become one of the current research hotspots. This paper， combined with the clinical research status of the brain-heart mutual damage， summarized its pathological mechanisms from the perspectives of inflammatory responses， dysregulation of autonomic nervous system， apoptosis， energy metabolism and oxidative stress. The necessity of “brain-heart concurrent regulation” was proposed and the research progress on the treatment of cerebral and cardiac diseases with TCM represented by Naoxintong capsule was profiled in the light of heart and brain function described by TCM and the holistic concept of TCM treating cerebral and cardiac diseases. This paper reviews the pathological mechanisms of brain-heart mutual damage and the research progress on its treatment with TCM， which can provide reference for the prevention and treatment of cardiovascular and cerebrovascular diseases and further research on them.

Application of Upper Arm Circumference and Calf Circumference in Screening and Diagnosis of Sarcopenia for Old Adults in Community / 中国康复理论与实践

Objective:To explore the correlation of upper arm circumference and calf circumference with muscle mass, and the screening effect and the cut-off values for low muscle mass and sarcopenia. Methods:From March to August, 2020, a total of 831 community-dwelling old people (≥ 60 years old) were recruited by using convenience sampling. Muscle mass and other body composition indicators were tested by body composition tester. The upper arm circumference, calf circumference, grip strength and gait speed were measured at the same day. Pearson cofficient was used to analyze the correlation of upper arm circumference and calf circumference with muscle mass, and receiver operation characteristic (ROC) curve was used to analyze the diagnostic effect of upper arm circumference and calf circumference on low muscle mass and sarcopenia, and the Youden index (YI) was used to determine the cut-off values. Results:Upper arm circumference and calf circumference were positively correlated with appendicular skeletal muscle mass (ASM) and skeletal mass index (SMI) (r = 0.6 to 0.8, P < 0.001). For diagnosing low muscle mass, the areas under the ROC curve (AUC) of upper arm circumference were 0.888 for male, 0.787 for female; and that of calf circumference were 0.933 for male, 0.854 for female, respectively. The cut-off values of upper arm circumference for low muscle mass were 26 cm for both male and female; and that of calf circumference were 34 cm for male and 33 cm for female, respectively. For diagnosing sarcopenia, the AUC of upper arm circumference was 0.902 for male, 0.744 for female; and that of calf circumference was 0.923 for male, 0.832 for female, respectively. The cut-off values of upper arm circumference and calf circumference for sarcopenia were 26 cm and 33 cm for both male and female. The AUC of combined diagnosis with age, upper arm circumference, calf circumference, grip strength and gait speed for low muscle mass and sarcopenia in males and females was all greater than 0.85. Conclusion:In the community old adults, upper arm circumference and calf circumference are positively correlated with muscle mass, and both indices can be used to diagnose low muscle mass and sarcopenia. multi-indices combination can improve the diagnosis effect of upper arm circumference in females.

Predictive Value of Tumor Mutation Burden for PD-1/PD-L1 Inhibitors Treatment on Non-small Cell Lung Cancer: A Meta-analysis / 肿瘤防治研究

Objective To investigate the relation between TMB and the efficiency of PD-1/PD-L1 inhibitors treatment for non-small-cell lung cancer. Methods Studies were searched from PubMed, Embase, Cochrane Library database, Chinese Biomedical Literature Database and Wanfang Database up to March 25, 2020. RevMan 5.3 software and STATA15.0 were used for analysis. Results Twelve literatures were involved, including 1209 patients. TMB significantly improved PFS (HR=0.54, 95%CI: 0.42-0.70, P < 0.001) but reduced the ORR (OR=4.41, 95%CI: 2.54-7.63, P < 0.001) of NSCLC patients treated with PD-1/PD-L1 inhibitors. The subgroup analyses showed that the predictive value of TMB was significant in non-small cell lung cancer treated by PD-1/PD-L1 inhibitors combined with anti-CTLA-4 therapy or chemotherapy. No significant publication bias was observed by the Begg's test and Egger's test. Conclusion High tumor mutation burden may predict the improved PFS of non-small cell lung cancer by PD-1/PD-L1 inhibitors treatment, but its predictive value for OS, ORR and long-term survival need more exploration.

Tumor-derived neomorphic mutations in ASXL1 impairs the BAP1-ASXL1-FOXK1/K2 transcription network

Additional sex combs-like 1 (ASXL1) interacts with BRCA1-associated protein 1 (BAP1) deubiquitinase to oppose the polycomb repressive complex 1 (PRC1)-mediated histone H2A ubiquitylation. Germline BAP1 mutations are found in a spectrum of human malignancies, while ASXL1 mutations recurrently occur in myeloid neoplasm and are associated with poor prognosis. Nearly all ASXL1 mutations are heterozygous frameshift or nonsense mutations in the middle or to a less extent the C-terminal region, resulting in the production of C-terminally truncated mutant ASXL1 proteins. How ASXL1 regulates specific target genes and how the C-terminal truncation of ASXL1 promotes leukemogenesis are unclear. Here, we report that ASXL1 interacts with forkhead transcription factors FOXK1 and FOXK2 to regulate a subset of FOXK1/K2 target genes. We show that the C-terminally truncated mutant ASXL1 proteins are expressed at much higher levels than the wild-type protein in ASXL1 heterozygous leukemia cells, and lose the ability to interact with FOXK1/K2. Specific deletion of the mutant allele eliminates the expression of C-terminally truncated ASXL1 and increases the association of wild-type ASXL1 with BAP1, thereby restoring the expression of BAP1-ASXL1-FOXK1/K2 target genes, particularly those involved in glucose metabolism, oxygen sensing, and JAK-STAT3 signaling pathways. In addition to FOXK1/K2, we also identify other DNA-binding transcription regulators including transcription factors (TFs) which interact with wild-type ASXL1, but not C-terminally truncated mutant. Our results suggest that ASXL1 mutations result in neomorphic alleles that contribute to leukemogenesis at least in part through dominantly inhibiting the wild-type ASXL1 from interacting with BAP1 and thereby impairing the function of ASXL1-BAP1-TF in regulating target genes and leukemia cell growth.

Research on grade prediction of Spatholobi Caulis via components-anti-oxidant activity correlations / 中草药

Objective: The model for grade evaluation of Spatholobi Caulis medicinal slices was constructed based on the quality control idea of traditional Chinese medicines that “components reflect activity and activity points to efficacy”. Methods: A method to determinate catechin and epicatechin content by using ultra performance liquid chromatography (UPLC) was proposed. ABTS•+ clearance rate, hydroxyl radical clearance rate and DPPH• clearance rate were used as evaluation indexes of biological activity. Correlations between content and anti-oxidant activity were analyzed by the logistic algorithm. Finally, a “principal component analysis-logistic regression” model for grade prediction of Spatholobi Caulis was constructed. Results: Catechin and epicatechin content in Spatholobi Caulis medicinal slices from different origins was between 0.40-1.26 mg/g and 0.57-2.02 mg/g, respectively. The anti-oxidant indexes ABTS•+, hydroxyl radical and DPPH• clearance rate were between 12.96%-51.76%, 30.65%-66.65%, and 37.65%-60.33%, respectively. The binary logistic regression analysis results showed that five batches were evaluated as excellent, average and poor rank, and four batches were considered as good rank (P > 94%) among 17 batches of Spatholobi Caulis herbal pieces and its two kinds of counterfeit (Sargentodoxa cuneata and Mucuna sempervirens). Conclusion: Binary logistic regression model for grade evaluation of Spatholobi Caulis medicinal slices was constructed preliminarily. It is applicable to assess quality of Spatholobi Caulis herbal pieces. The grading evaluation model of Spatholobi Caulis via logistic regression analysis can be used to classification of Spatholobi Caulis from different sources.

UPLC fingerprint-based analysis of differences in chemical components from different parts of Stephania tetrandra / 中草药

Objective: To analyze and compare UPLC fingerprints of root, rhizome, stem, and leaf of Stephania tetrandra, learn the differences in chemical component types and contents of main active components, and provide basis for rational development and utilization of S. tetrandra. Methods: UPLC was used to obtain characteristic chromatograms of different parts; The Similarity Evaluation System for Chromatographic Fingerprints of Traditional Chinese Medicine (Version 2012) was run to capture the common peaks of different parts and calculate their similarity and analyze the characteristic peaks of different parts. SPSS 23.0 was run to compare the difference in component contents of the roots and rhizomes using the paired sample t-test. Results: The similarity in chemical composition between root and rhizome was 0.928, indicating they have similar chemical composition, and both of them contained tetrandrine and fangchinoline, the index components. The similarity between rhizome and leaf was 0.947; The similarity was low between stem, leaf and root and rhizome, and there were no tetrandrine and fangchinoline in the first two parts. The results of paired samples t-test show that the total content of chemical components in rhizome was higher than that in roots, and the mainly difference came from other non-index components, but there was no significant difference between tetrandrine and fangchinoline. Conclusion: Significant differences are present in chemical composition types and contents of different medicinal parts of S. tetrandra; The type of chemical components in rhizome is similar to that in root, and the content of some components in rhizome is significantly higher than that in root, which means that rhizomes can be used as an equivalent of roots. Stems and leaves cannot be a substitute for roots because they do not contain tetrandrine and fangchinoline, but they contain many other chemical components which can be utilized as a new resource.

Therapeutic effect of cinnamaldehyde on ulcerative colitis in mice induced by dextran sulfate sodium with Candida albicans colonization and its effect on dectin-1/TLRs/NF-κB signaling pathway / 中国中药杂志

To observe the efficacy of cinnamaldehyde on dextran sulfate sodium(DSS)-induced ulcerative colitis(UC) with Can-dida albicans(Ca) colonization and its effect on dectin-1/TLRs/NF-κB signaling pathway in mice. C57 BL/6 mice were randomly divided into normal group, DSS group, DSS+Ca group, cinnamaldehyde group and mesalazine group. Mice in DSS+Ca group were given Ca(1×10~8 CFU per mouse) through intragastrical administration for 4 consecutive days and then distilled water with 3.0% DSS for 7 consecutive days. In cinnamaldehyde group and mesalazine group, in addition to the induction method of the DSS+Ca group, mice were given 75 mg·kg~(-1) cinnamaldehyde and 200 mg·kg~(-1) mesalazine accompanied with 3.0% DSS for 7 consecutive days, respectively. Mice in normal group and DSS group were correspondingly administered with distilled water. The general conditions of the mice were observed daily, the diseased activity index(DAI) score was calculated, and fungal loads of feces were detected by plate method. The mice were sacrificed on day 12, colon length was measured, colon mucosa damage index(CMDI) score was calculated, and histopathological analysis was carried out by HE staining. Anti-saccharomces cerevisiae antibody(ASCA) and β-1,3-glucan in serum, and TNF-α, IL-1β, IL-6, IL-8, IL-10 in serum and colon tissue were detected by ELISA. The contents of β-1,3-glucan and macrophage infiltration in colon tissues were examined by immunofluorescence staining. The protein expressions of dectin-1, TLR2, TLR4 and NF-κB were detected by Western blot and immunohistochemistry staining. The results showed that cinnamaldehyde could significantly improve the general conditions of UC mice with Ca colonization, decrease DAI and histopathological scores, reduce intestinal mucosal congestion, erosion and colon shortening, decrease Ca load in mouse feces and tissues, down-regulate the contents of ASCA and β-1,3-glucan in serum, reduce the contents of TNF-α, IL-1β, IL-6, IL-8 and increase IL-10 in serum and colon tissues, inhibit macrophages infiltration and down-regulate the protein expression of dectin-1, TLR2, TLR4 and NF-κB in colon tissue. These results suggested that cinnamaldehyde had a therapeutic effect on UC mice with Ca colonization, which might be related to the inhibition of Ca proliferation, the regulation of dectin-1/TLRs/NF-κB signaling pathways and the coordination of the balance between pro-inflammatory and anti-inflammatory factors.

Clinical observation of sinew-regulating bone-setting manipulations plus exercise therapy for chronic non-specific low back pain / 针灸推拿医学（英文版）

Objective: To evaluate the clinical efficacy of sinew-regulating bone-setting manipulations plus exercise therapy in treating chronic non-specific low back pain (CNLBP). Methods: A total of 65 CNLBP patients were divided into two groups by the random number table method. Thirty-three cases in the treatment group were intervened by sinew-regulating bone-setting manipulations plus exercise therapy; 32 cases in the control group were intervened by medium-frequency electrotherapy plus exercise therapy. Before and after treatment, visual analog scale (VAS), dynamic and static muscle endurance of low back, median frequency (MF) of surface electromyography (sEMG) and Oswestry disability index (ODI) were used to evaluate the low back function. The therapeutic efficacy was estimated after treatment. Results: The two groups each had 2 dropouts during the study. The total effective rate was 90.3％ in the treatment group versus 66.7％ in the control group, and the between-group difference was statistically significant (P<0.05). After treatment, the VAS score, dynamic and static muscle endurance of low back, MF of sEMG and ODI score all changed significantly in both groups (all P<0.05); all the items in the treatment group were significantly different from those in the control group (all P<0.05). Conclusion: Sinew-regulating bone-setting manipulations plus exercise therapy can effectively release pain in CNLBP patients, increase muscle endurance of the low back and improve the quality of life, and its therapeutic efficacy is more significant than that of medium-frequency electrotherapy plus exercise therapy.

Quality Regionalization and Environmental Impact Factors Analysis Based on Main Active Ingredients of Stephaniae Tetrandrae Radix / 中国实验方剂学杂志

Objective:To investigate the quality regionalization and environmental impact factors of Stephaniae Tetrandrae Radix based on main active ingredients，and provide a reference for the determination of high-quality production areas and the dominant environmental factors affecting the content of Stephaniae Tetrandrae Radix. Method:Partial least squares regression analysis (PLS) and principal component analysis (PCA) were used to study the quality regionalization and environmental impact factors based on the main active ingredients of tetrandrine and fangchinoline, and investigate the environmental factors of the producing areas. Result:The content of fangchinoline was positively correlated with soil pH and annual average temperature，negatively correlated with latitude. The content of tetrandrine was positively correlated with soil pH，negatively correlated with annual rainfall and longitude. The total content was positively correlated with soil pH and annual average temperature，but negatively correlated with annual rainfall，latitude and longitude. Principal component analysis showed that the 50 production areas could be divided into four groups of quality formation. The groups with the highest scores were Shixing county in Guangdong，Shexian county in Anhui，Songxi county in Fujian，Nanxiong city in Guangdong and Xiangxiang city in Hunan，all of which were best areas for accumulation of the two main active ingredients. Conclusion:Soil pH，annual average temperature，annual rainfall，latitude and longitude are the main environmental factors affecting the main active ingredients of Stephaniae Tetrandrae Radix. The best areas for accumulation of tetrandrine and fangchinoline are Shixing county in Guangdong，Shexian county in Anhui，Songxi county in Fujian，Nanxiong city in Guangdong and Xiangxiang city in Hunan.

SSR-based Analysis of Genetic Diversity and Quality Trait-related Markers of 69 Picria felterrae Germplasm Samples / 中国实验方剂学杂志

Objective::Sixty-nine germplasm samples of Picria felterrae collected from the main producing areas in Guangxi were subject to genetic diversity and genetic relationship analyses using the simple seguence repeat(SSR) molecular marker technology and good germplasm genes associated with the content of picfeltarraenins were screened so as to provide references for germplasm resource evaluation, phylogenetic analysis, and molecular mark assisted breeding of that species. Method::20 pairs of randomly selected primers were amplified based on the transcriptome sequencing technology. The genetic diversity of and genetic relationship between the 69 samples were analyzed using the genetic polymorphic information for each marker locus, and one-variable linear regression and multiple stepwise regression analyses were performed to screen molecular markers associated with the content of picfeltarraenins. Result::The amplification using the 20 pairs of SSR primers produced 76 alleles, 3.8 alleles for each locus on average, higher than effective alleles (1.969 2), and the rare allele rate was 38.2%, suggesting that the alleles distributed unequally. The polymorphism rates of alleles varied between 0-59%, with an average of 38.24%, showing a great difference among loci. The polymorphic information content (PIC) varied between 0-0.621 1, with an average of 0.378 0.Shannon polymorphic information index varied between 0-1.240 1, with an average of 0.759.Nei's gene diversity index (Nei) varied between 0-0.682 3, with an average of 0.440 9.P21 had the highest level accompanied with the lowest P7 for the above three indexes, and significant genetic diversity differences were identified among the loci. For all loci, the mean observed heterozygosity was 0.382 4, lower than the average expected heterozygosity of 0.442 5, suggesting the loss of heterozygosity, the average genetic differentiation coefficient (Fst) was 0.365 9 and the average gene flow Nm was 0.433 2, suggesting a high genetic divergence and a low gene flow. The results of one-variable linear regression and multiple stepwise regression showed that there were 5 loci related to each of the picfeltarraenin IA and IB, and only 1 loci was associated with the content of both. Conclusion::There were significant differences in the genetic diversity of 20 SSR marker sites, and the 69 germplasm samples were greatly genetically differentiated and had low gene flow. From the selected 20 SSR markers 9 marker loci associated with the content of picfeltarraenin IA and IB were selected. The results can be used as a reference for phylogenetic analysis and selective breeding of Picria felterrae.

Differentially expressed genes of HepG2 cells treated with gecko polypeptide mixture / 中国药理学与毒理学杂志

OBEJECTIVE Gecko has been clinically used in China for many years. It has been proved that the gecko polypeptide mixture(GPM)extracted from gecko could inhibit the growth of multiple types of tumor cells.In order to investigate the possible anti-tumor molecular mechanisms of GPM,we used RNA-seq technology to identify the differentially expressed genes of human hepatocellular carci-noma(HCC)HepG2 cells treated with or without GPM.METHODS The HepG2 cells were treated with different concentration of GPM(0,0.1,0.2,0.3,0.4 mg·mL-1)for 6 h,12 h and 24 h,respectively.MTT assay was used to detect the viability of HepG2 cells. DAPI fluorescence staining was performed to observe nucleus morphological changes of HepG2 cells.Western blot analysis was applied to observe the expres-sion of apoptosis- related proteins and endoplasmic reticulum stress (ERs)-related proteins in HepG2 cells.Flow cytometry was also applied to detect reactive oxygen species(ROS)generation.In this report, we showed that GPM could induce HepG2 cells apoptosis and influence HepG2 cells proliferation in a dose-dependent manner.We applied many analysis methods,including differentially expressed genes analysis,Gene Ontology(GO)enrichment analysis,KEGG pathway enrichment analysis,protein-protein interaction network analysis to screen out possible molecular mechanisms.RESULTS ER-nucleus signaling pathway, cellular response to stress and apoptotic processes were identified the potential anti-cancer molecular biological process of GPM.GPM may also induce apoptosis in HepG2 cells via endoplasmic reticulum stress pathway. The GPM could induce ROS generation and up-regulate ERs-related proteins. CONCLUSION The present study revealed the potential anti-tumor mechanism of GPM.

Changes of Alternative Splice Variant of the SRSF12 by Cisplatin in Multiple Cells / 昆明医科大学学报

Objective To detect the changes of mRNA isoforms in multiple cancer cell lines with dose of cisplatin. Methods Total RNA of the cells treated by cisplatin were abstracted 24 h after the treatment. mRNA isoforms of SRSF12 gene were detected by semiquantitative RT-PCR. The ratios of mRNA isoforms were analyzed by gel image software and statistical analysis. Results Under the cisplatin treatment, 2 mRNA isoforms of SRSF12 were detected in five cells except Caski cells,their ratios and relative mRNA levels were changed. With the increase of dose of cisplatin, the ratio of isoform-a was slightly increased; but the ratio of isoform-b was different, the changes were not obvious in the A549 and 293FT cells, the weaker expression was expressed in the H1299 and C33A cells, and the two isoforms were gradually weakening in the Siha cells. Conclution Under the cisplatin treatment in multiple cancer cells, the expression of SRSF12 shows tissue-specific and cell type-specific patterns .