Abstract：There are 17 formulas containing Pinelliae Rhizoma in the Catalogue of Ancient Famous Classical Formulas (the First Batch), most of which are only labeled with " washing with decoction" or without any processing method, which is inconsistent with the current use requirements and brings confusion to research and development of related famous classical formulas. Through combing the records of famous classical formulas in the original book, contemporary works and later works, the usage of processed products of Pinelliae Rhizoma in the evolution of past dynasties was contrasted, and the efficacy, indications and compatibility significance of different formulas were analyzed, according to the principle of " respecting the ancient but not confining the ancient" , the authors suggested that the 17 famous classical formulas should use the processed products of Pinelliae Rhizoma in the 2015 edition of Chinese Pharmacopoeia, including Pinelliae Rhizoma Praeparatum, Pinelliae Rhizoma Praeparatum cum Zingibere et Alumine and Pinelliae Rhizoma Praeparatum cum Alumine. Among them, Pinelliae Rhizoma Praeparatum cum Zingibere et Alumine should be used in Banxia Xiexintang, Gancao Xiexintang, Huangliantang, Xuanfu Daizhe Tang, Zhuye Shigaotang, Banxia Houpotang, Maimendong Tang, Gualou Xiebai Banxiatang, Wendantang, Zhurutang, Jinshui Liujun Jian and Yangweitang, Pinelliae Rhizoma Praeparatum should be used in Shengyang Yiweitang, Houpo Mahuangtang, Banxia Baizhu Tianma Tang and Huopo Xialing Tang, and Pinelliae Rhizoma Praeparatum cum Alumine should be used in Sangbaipi Tang.
Keywords：famous classical formulas;Pinelliae Rhizoma;processing of traditional Chinese medicine;washing with decoction;Pinelliae Rhizoma Praeparatum cum Zingibere et Alumine;Pinelliae Rhizoma Praeparatum cum Alumine;Pinelliae Rhizoma Praeparatum
Abstract：Objective:To investigate the possible mechanism of Dabuyuan Jian to promote neurogenesis by studying the effect of Dabuyuan Jian on Wnt/β-catenin signaling pathway in hippocampus of amyloid precursor protein/presenilil(APP/PS1) mice.Method:Totally 30 5-month-old APP/PS1 mice and 10 wild mice were divided into control group, model group, donepezil group (6.5×10-4 g·kg-1·d-1) and Dabuyuan Jian group (13.2 g·kg-1·d-1), and given drugs by gavage for 30 days. The control group and the model group were given an equal volume of saline. The learning and memory of mice were evaluated by water maze. The pathological changes of hippocampal neurons were observed by hematoxylin-eosin (HE) staining. The immunohistochemistry (IHC) was used to detect label positive cells of 5-bromodeoxyuridine (Brdu), adrenocortical hormone (Dcx) and neuronal nuclear antigen (NeuN). Real-time fluorescence quantitative polymerase chain reaction (Real-time PCR) and Western bolt were used to detect the mRNA and protein expression levels of β-catenin and glycogen synthase kinase-3β(GSK-3β) in hippocampus of mice.Result:Compared with the control group, the latency and swimming total distance of the model group were significantly increased (P<0.01), while the number of crossing platforms and the target quadrant time were significantly reduced (P<0.01). Compared with the model group, the platform latency and the total swimming distance of the donepezil group and the Dabuyuan Jian group were decreased (P<0.05, P<0.01), while the number of crossing platforms and the target quadrant time were increased (P<0.05). Compared with the control group, the number of neurons in the hippocampal dentate gyrus (DG) area of the model group was decreased, and necrotic neurons were observed. Compared with the model group, the number of neurons in the hippocampal DG area of the mice in the donepezil group and the Dabuyuan Jian group was increased, while the number of necrotic neurons was decreased. Compared with the control group, the number of positive cells labeled with Brdu, Dcx and NeuN in the hippocampal DG area of the model group was significantly decreased (P<0.01). Compared with the model group, the number of positive cells labeled with Brdu, Dcx and NeuN in the hippocampal DG area of the donepezil group and the Dabuyuan Jian group was increased (P<0.05, P<0.01). Compared with the control group, gene and protein expression levels of β-catenin in the hippocampus of the model group were significantly decreased (P<0.01), whereas gene and protein expression levels of GSK-3β were significantly increased (P<0.01). Compared with the model group, gene and protein expression levels of β-catenin in hippocampus of donepezil group and Dabuyuan Jian group were increased (P<0.05, P<0.01), while gene and protein expression levels of GSK-3β were increased (P<0.05, P<0.01).Conclusion:Dabuyuan Jian could promote hippocampal neurogenesis in APP/PS1 double transgenic mice by regulating Wnt/β-catenin signaling pathway.
Abstract：Objective:To study the chemical constituents in Baihe Dihuangtang by ultra-performance liquid chromatography-quadrupole-time of flight-mass spectrometry (UPLC-Q-TOF-MS).Method:The separation was performed on an ACQUITY UPLC BEH C18 column (2.1 mm×50 mm, 1.7 μm) by a gradient elution of acetonitrile-0.1%formic acid solution. The flow rate was 0.4 mL·min-1 and the column temperature was 30 ℃, the injection volume was 5 μL. Electrospray ionization was applied and the data were collected via positive and negative ion modes. By using SCIEX OS 1.4 software, the chemical constituents were analyzed based on the retention time, excimer ion peak and fragment ion peak of the compounds, as well as comparison with reference substances and literature data.Result:A total of 49 chemical constituents in Baihe Dihuangtang were identified, including 5 phenolic glycerides, 15 phenylethanoid glycosides, 11 iridoids, 8 lonones, 3 phenylpropanoids, 2 nucleosides, 1 organic acid, and 4 other compounds. Among them, phenolic glycerides belonged to Lilii Bulbus, and other components mainly belonged to Rehmanniae Radix. Four chemical constituents (acteoside, isoacteoside, ferulic acid and caffeic acid) were identified by comparison of reference substances.Conclusion:The established detection method can quickly and accurately analyze the chemical constituents of Baihe Dihuangtang. The information of chemical constituents in Baihe Dihuangtang is comprehensively expounded. The study establishes a foundation for the research of quality control, material foundation of efficacy and the development of compound preparations of Baihe Dihuangtang.
Abstract：Objective:To investigate the neuroprotective effect and mechanism by Wutoutang (WTT) in the spinal nerve ligation (SNL) mice by neurotrophic factor (BDNF)/tyrosine kinase receptor B (TrkB) signaling BDNF/tyrosine kinase receptor B (TrkB).Method:The 40 mice were randomly divided into Sham group, SNL group, WTT group(126 mg·kg-1), ANA-12+ WTT group.The L5 spinal nerve ligation model mice were established in mice, After that, WTT was administrated from the first day to the 10th day, then the consecutive 7-day hippocampal injection of ANA-12(0.05 nmol·L-1)were lasted for 7 days.The levels of brain-derived BDNF, cAMP-response element binding protein(CREB), and protein kinase B(Akt)and the change of hippocampal glutamatergic and GABAergic neurons in mice were detected by tissue immunofluorescence.E14 pregnant ICR mice were sacrificed and the hippocampus were dissected which were divided into control group, glycine group, tumo necrosis factor(TNF)-α(5 mg·L-1)+ glycine group, TNF-α+ WTT(5 mg·L-1)+ glycine group, TNF-α+ WTT+ glycine+ BDNF-siRNA group, TNF-α+ WTT+ glycine+ Akt-siRNA group, TNF-α+ WTT+ glycine+ CREB-siRNA group, the primary and secondary dendrrictes, in which the arrowheads indicate the expression od postsynapti desity protein 95(PSD95) in the shafts and arrows were tested by cellular immunofluorescence.The neurons were divided into control group, glycine group, ANA-12 group(0.5 mmol·L-1), ANA-12+ glycine group, ANA-12+ WTT group, ANA-12+ WTT+ glycine group, the morphology of hippocampal neurons were tested by cellular immunofluorescence.Result:Compared with Sham group, BDNF, Akt and CREB positive cell of SNL group decreased significantly(P<0.01), the hippocampal glutamatergic and GABAergic neurons out of balance(P<0.01). Compared with SNL group, the BDNF, Akt and CREB positive cell of WTT group increased significantly(P<0.01), Glutamine-aminobutyric acid neurons regein balance(P<0.01). Compared with WTT group, BDNF and CREB positive cell of ANA-12+ WTT group decreased significantly(P<0.05), Glutamine-aminobutyric acid neurons was disorderedd(P<0.05). Comparaed with control group, the level of PSD95 of glycine group were increase significantly(P<0.01). The number of dendritic spine density apically and basally of glycine group were increase significantly(P<0.01), but the primary and secondary dendrites of ANA-12 group, ANA-12+ glycine group, ANA-12+ WTT group, ANA-12+ WTT+ glycine group were not change.Comparaed with glycine group, the level of PSD95 of TNF-α+ glycine group were decreased significantly(P<0.01). Comparaed with TNF-α+ glycine group, the level of PSD95 of TNF-α+ WTT+ glycine group were increase significantly(P<0.01). Comparaed with TNF-α+ WTT+ glycine group, the level of PSD95 of TNF-α+ WTT+ glycine+ BDNF-siRNA group, TNF-α+ WTT+ glycine+ Akt-siRNA group, TNF-α+ WTT+ glycine+ CREB-siRNA group were decreased significantly(P<0.01).Conclusion:In vivo and in vitro studies have shown that the WTT mediated remission of the primary hippocampal glutamatergic neurons were dependent on the BDNF/TrkB pathway.
Keywords：Wutoutang;neuropathic pain;neurotrophic factor (BDNF)/tyrosine kinase receptor B (TrkB) neurotrophic factor (BDNF)/tyrosine kinase receptor B (TrkB);neuroprotection
Abstract：Objective:Duanteng Yimu decoction(DTYMD)is effective in treatment of rheumatoid arthritis (RA) by relieving joint inflammation and down-regulating some inflammatory factors in a short period of time, but the mechanism is still unclear. We aimed to investigate upstream kinase of mitogen activated protein kinases(MAPK) and define the anti-inflammatory mechanism of DTYMD.Method:Fibroblasts-like synovial cells(FLSs) were divided into blank group, model group (IL-1β), high-dose DTYMD group (1 000 mg·L-1), medium-dose DTYMD group (800 mg·L-1), low-dose DTYMD group (600 mg·L-1) and armour ammonia butterfly(MTX) group (20 μmol·L-1). The protein and mRNA expressions of mitogen-activated protein kinase kinase kinase 2 (MEKK2) were analyzed by real-time fluorescence quantitative PCR(Real-time PCR). Totally 42 male DBA/1J mice were randomly divided into 6 groups, with 7 mice in each group, namely normal group, model group and MTX group (2 mg·kg-1), low-dose DTYMD group (6.25 mg·kg-1), medium-dose DTYMD group (12.5 mg·kg-1), and high-dose DTYMD group (25 mg·kg-1). Except for the normal group, the other five groups were included in collagen-induced arthritis(CIA) model by secondary immunoassay. After administration, the posterior limbs and ankle joints were stained with htoxylin-eosin(HE), and the pathological scores of the joints were evaluated.Result:Compared with the model group, DTYMD inhibited the activity of FLSs in a concentration-dependent manner (P<0.01). Compared with the blank control group, the cell proliferation rate of the model group increased (P<0.01). Compared with the model group, high and middle-dose DTYMD groups could inhibit protein and mRNA expressions of MEKK2 (P<0.01), but there was no significant difference in low-dose group. However, the expression of DTYMD protein in high/medium/low-dose groups was significantly higher than that in blank group (P<0.01), but there was no significant difference in MTX group. Compared with the model group, the expressions of matrix metalloprotease-1 (MMP-1), tumor necrosis factor-α(TNF-α) and interleukin(IL)-6 were negatively regulated in different DTYMD groups(P<0.01), and the expressions of MMP-1, IL-6, TNF-α in the model group were significantly higher than those in the blank group (P<0.05, P<0.01). In the animal experiment, compared with the model group, high/middle-dose DTYMD groups could decrease the degree of joint swelling in CIA mice (P<0.01), but there was no significant difference in the low dose group, and the joint swelling in the model group was significantly higher than that in the blank group (P<0.05). In HE staining of ankle joint of CIA mice, the pathological scores of high/small-dose DTYMD groups were significantly lower those of model group (P<0.05, P<0.01), and the pathological score of model group was higher than that of blank group (P<0.01).Conclusion:DTYMD might down-regulate MEKK2 to negatively regulate inflammatory cytokines IL-6, TNF-α and MMP-1, thereby alleviating the inflammatory response in rheumatoid arthritis.
Keywords：Duanteng Yimu decoction;rheumatoid arthritis;mitogen-activated protein kinase kinase kinase 2;inflammatory factor;collegen induced arthritis(CIA) mice model
Abstract：Objective:To observe the effect of compound Kushen injection on the expressions of transforming growth factor-β1 (TGF-β1), drosophila mothers against decapentaplegic protein 3 (Smad3), glycogen synthase kinase-3β (GSK-3β) and β-catenin mice models with radiation-induced pulmonary injury (RIPI), in order to explore its possible mechanism of action.Method:On XStrahl precision radiation research platform for small animals (SARRP), a single 20 Gy bilateral lung field irradiation was performed to establish a mice model of RIPI. Thirty-two mice were randomly divided into normal control group, model group, compound Kushen injection group and dexamethasone injection group. The normal control group and the model group were given an equal volume of 0.9%sodium chloride solution and injected intraperitoneally for 4 weeks. The pathology of lung tissue tissues was observed by using hematoxylin-eosin (HE) staining. Immunohistochemical(IHC) was used to detect the expressions of E-cadheren and Vimentin proteins in mice lung tissues.Real-time polymerase chain reaction (Real-time PCR) was used to detect the mRNA expressions of TGF-β1, Smad3, GSK-3β and β-cateninin.Western blot was used to detect the protein expressions of TGF-β1, Smad3, GSK-3β and β-cateninin.Result:Compared with the normal group, the pulmonary coefficient of the model group was significantly decreased (P<0.01). Inflammatory cell infiltration, pulmonary interstitial edema, congestion, destruction of alveolar structure and partial alveolar atrophy were observed in the lung tissues of the model group. Compared with the model group, in the compound Kushen injection group, the levels of infiltration of lung inflammatory cells and pulmonary interstitial lesions in mice, the expression of Vimentin in lung tissues (P<0.01), and the expressions of TGF-β1, Smad3, GSK-3β and β-cateninin were significantly decreased (P<0.01), whereas the expression of E-cadheren was significantly increased (P<0.01). However, compared with the dexamethasone injection group, in the compound Kushen injection group, the pathological changes of lung tissues were similar, and the expression levels of E-cadheren, Vimentin, TGF-β1, Smad3, GSK-3β and β-cateninin were not significantly different.Conclusion:Compound Kushen injection can alleviate pulmonary fibrosis of lung in the treatment of RIPI, and the mechanism may be associated with inhibiting the mRNA and protein expressions of TGF-β1, Smad3, GSK-3β and β-catenin related to epithelial-mesenchymal transition(EMT), promoting the expression of E-cadheren, and inhibiting the expression of Vimentin, so as to inhibit the occurrence of EMT.
Keywords：radiation-induced pulmonary injury (RIPI);compound Kushen injection;drosophila mothers against decapentaplegic protein 3 (Smad3);transforming growth factor-β1 (TGF-β1);glycogen synthase kinase-3β (GSK-3β);β-catenin (β-catenin)
Abstract：Objective:To investigate the protective effect of Longxue Tongluo capsule (LTC) on H2O2-induced injury of PC12 cells and its potential mechanism.Method:An in vitro injury model in PC12 cells was established with 500 μmol·L-1 H2O2.The experiment was divided into control group, injury model group (H2O2 500 μmol·L-1), and Longxuetongluo capsule group (LTC, 1, 2, 4 mg·L-1). Cell counting kit-8 (CCK-8) assay, intracellular reactive oxygen species (ROS), mitochondrial membrane potential, apoptosis of PC12 cells, and Western blot were used to evaluate the protective effect of LTC on PC12 cells induced by H2O2.Result:Compared with the control group, the cell viability was significantly decreased in the injury model group (P<0.01), intracellular ROS level was significantly increased (P<0.01), mitochondrial membrane potential was decreased, while apoptosis of PC12 cells was significantly increased (P<0.01), and the expression of cleaved poly adenosine diphospho ribose polymerase (PARP) was also increased significantly (P<0.01). Compared with the injury model group, pretreatment with LTC at the concentrations of 2 and 4 mg·L for 6 h significantly increased cell viability in PC12 cells exposed to H2O2 (P<0.01). Moreover, pretreatment with LTC reduced intracellular ROS level (P<0.05), maintained mitochondrial membrane potential, and inhibited apoptosis of PC12 cells induced by H2O2 in a dose-dependent manner (P<0.01). The results of western blotting showed that pretreatment with LTC significantly reduced the expression of cleaved PARP (P<0.01).Conclusion:LTC exerts a significant protective effect against H2O2-induced PC12 cells injury through inhibition of neuronal apoptosis by suppressing intracellular oxidative stress, maintaining mitochondrial function, and promoting DNA repair.
Abstract：Objective:To compare the pharmacodynamic effects of pith-nodecayed and pith-decayed products of Scutellariae Radix on rats with large intestine damp-heat syndrome, and to demonstrate the scientificness of dividing Scutellariae Radix into pith-nodecayed and pith-decayed products as medicines by modern pharmacological test.Method:Rats were randomly divided into blank group, model group, low-and high-dose group of pith-nodecayed products (0.9, 3.6 g·kg-1), low-and high-dose group of pith-decayed products (0.9, 3.6 g·kg-1), Scutellariae Radix group (0.9 g·kg-1), compound berberine tablets group (positive drug group, 0.045 g·kg-1), and 8 rats in each group. Taking model rats with large intestine damp-heat syndrome, the body temperature, thymus index, spleen index, pathological sections of colon and ileum, inflammatory factors and Secretory immunoglobulin (SIg) A content were selected as indexes to evaluate the therapeutic effect of pith-nodecayed and pith-decayed products on large intestine damp-heat syndrome, and make comprehensive evaluation of the difference in efficacy between them. Partial least squares-discriminant analysis (PLS-DA) was employed to analyze the pharmacological indexes of these two products against large intestine damp-heat syndrome.Result:Pith-nodecayed and pith-decayed products of Scutellariae Radix with different doses could reduce the body temperature, thymus index, spleen index, contents of interleukin (IL)-2, IL-6, IL-1β in serum and SIgA content in intestinal mucosa, and most of them had significant differences (P<0.05, P<0.01). Compared with the isodose group of pith-decayed products, the effect of corresponding dose group of pith-nodecayed products was better, and most of them had significant differences (P<0.05, P<0.01). PLS-DA results indicated that there were significant differences in the pharmacological effects of pith-nodecayed and pith-decayed products, and they were clustered on one side, respectively.Conclusion:Both of pith-nodecayed and pith-decayed products of Scutellariae Radix have therapeutic effect on large intestine damp-heat syndrome with distinctly different strength of action, and pith-nodecayed products is superior to pith-decayed products, which verify the scientific nature of pith-nodecayed products was specializedly used to treat bowel disease in ancient times.
Keywords：Scutellariae Radix;pith-nodecayed products of Scutellariae Radix;pith-decayed products of Scutellariae Radix;large intestine damp-heat syndrome;partial least squares-discriminant analysis;inflammatory factors;secretory immunoglobulin A
Abstract：Objective:To observe the synergistic effect and mechanism of Yiqi Jianpi Huayu (YQJPHY) recipe combined with 5-fluorouracil (5-FU) on inhibiting gastric cancer growth, and investigate its possible mechanism through polarization direction of tumor-associated macrophages (TAMs) in tumor micro-environment.Method:A total of forty 615 mice were randomly divided into four groups including control group, YQJPHY(25 g·kg-1) group, 5-FU (25 mg·kg-1) group and combined group (YQJPHY plus 5-FU), with 10 mice in each group. The gastric cancer transplantation model was induced by axillary inoculation of MFC gastric cancer cells. Changes in tumor pathology were observed with hematoxylin-eosin staining (HE) method. The contents of M1 and M2 TAMs in tumor tissues were determined with flow cytometry, while the expression of CD206 in tumor was observed with immunohistochemistry. The mRNA expressions of TAM-related genes including interleukin-1β(IL-1β), interleukin-12(IL-12), tumor necrosis factor-α(TNF-α), arginase-1(Arg1), and chitinase 3-like 3(Ym1) were detected by using quantitative real-time polymerase chain reaction (Real-time PCR). Furthermore, the mRNA and protein expressions of epithelial-mesenchymal transition (EMT) related epithelial calcium adhesion protein(E-cadherin), nervous calcium adhesion protein(N-cadherin), Zinc finger transcription factor Slug, Snail and Waveform protein (Vimentin) were detected with Real-time PCR and Western blot, respectively.Result:The gastric cancer cells in tumors were distributed in nest and/or flaky shape, with fibroblastic connective tissue reaction and inflammatory cells infiltrated in interstitium. Coagulative necrosis of tumor was observed in various treatment groups. The amount of residual cancer cells in mouse was as follows from largest to smallest: control group>YQJPHY group>5-FU group>combined group. Immunohistochemical studies showed that all treatment groups can reduce the expression of CD206 in the tumor, and the effect was most obvious in the combined group. The contents of M2-TAM in all treatment groups were lower than those in control group(P<0.05, P<0.01), and the most significant difference in the content of M2-TAMs was shown between the control group and the combined group. The contents of M1 TAMs were increased in YQJPHY and combined groups(P<0.05, P<0.01), with the largest incremental range in YQJPHY group. The expressions of Arg1 and Ym1 in YQJPHY and combined groups were lower, while IL-1β, TNF-α and IL-12 levels were higher than those in control group(P<0.05). The mRNA expressions of IL-1β, IL-12 and TNF-α in the combined group were significantly higher, while Arg1 and Ym1 mRNA expression levels were lower than those in the 5-FU group(P<0.05). As compared with the control group, the mRNA and protein expression levels of E-cadherin were up-regulated(P<0.05), while the mRNA expression levels of N-cadherin and Vimentin were down-regulation in the YQJPHY group (P<0.05), E-cadherin mRNA and protein expression levels were up-regulated, while N-cadherin, Slug, Snail, Vimentin mRNA expression as well as N-cadherin and Slug protein expression levels were down-regulated in the combined group(P<0.05). As compared with the 5-FU group, the mRNA and protein expression levels of E-cadherin were up-regulated(P<0.05), while the mRNA and protein expression levels of N-cadherin, Slug and Vimentin were down-regulated in the combined group(P<0.05).Conclusion:YQJPHY has an inhibitory effect on the growth of gastric cancer, and after being combined with 5-FU, it has a good synergistic effect on inhibiting the growth of gastric cancer and decreasing EMT. The mechanism may be related to promoting the conversion from M2-TAMs to M1-TAMs in gastric tumor micro-environment.
Abstract：Objective:To investigate the effect of drug-containing serum of Jianpi Xiaoai prescription on protein kinase B (Akt)/mammalian target of rapamycin (mTOR) signaling pathways in colorectal cells HCT116.Method:The HTC116 cells were treated by 15%concentration of drug-contained serum, and then the cell migration and invasion were detected by Transwell assay, the protein expression levels of Akt, phosphorylated protein kinase B (p-Akt), mTOR, phosphorylated mammalian target of rapamycin (p-mTOR), ribosomal protein S6 kinase, polypeptide1(S6K1), phosphorylated ribosomal protein S6 kinase, polypeptide1 (p-S6K1), 4E-binding protein1(4EBP1), and phosphorylated 4E-binding protein1(p-4EBP1) in HCT116 cells were detected by Western blot. The control group was treated by untreated serum (15%), and 10%fetal bovine serum(FBS).Result:As compared with the control group, the number of migration and invasion cells was significantly reduced in drug-contained serum group (P<0.01), the expression of Akt had no obvious decrease, p-Akt protein expression was significantly lowered in the drug-contained serum group (P<0.01), the expression of mTOR had no obvious decrease, but p-mTOR protein expression was significantly lowered in drug-contained serum group (P<0.01), the expression of S6K1 had no obvious decrease, but p-S6K1 protein expression was significantly lowered in the drug-contained serum group (P<0.01), the protein expression of 4EBP1 had no obvious decrease, but p-4EBP1 protein expression was significantly lowered in the drug-contained serum group (P<0.01).Conclusion:The anti-tumor mechanism and transfer of Jianpi Xiaoai prescription may be related to inhibiting the activation of Akt/mTOR signaling pathways in colorectal cancer.
Keywords：Jianpi Xiaoai prescription;colorectal cancer;migration and invasion;protein kinase B (Akt)/mammalian target of rapamycin (mTOR);phosphorylated
Abstract：Objective:To evaluate the intervention effect of Bushen Tongluo formula on zebrafish osteoporosis model and to clarify its regulation of autophagy mechanism on osteoclast differentiation.Method:The 260 young zebrafishes (3 dpf) were selected, the zebrafish osteoporosis model was established with 25 mmol·L-1 prednisolone (Pred) for 3 days, confirming the successful model by calcein staining. The zebrafishes were divided into control group, Pred group (25 mmol·L-1), etidronate disodium (ED) group (300 mg·L-1), Bushen (BS) group (180 mg·L-1), Tongluo (TL) group (30 mg·L-1), and Bushen Tongluo (BSTL) group (210 mg·L-1), 40 tails per group. After intervened with medicine for 4 days, the calcein staining was adopted to count the vertebral bone fluorescence area of zebrafish, Real-time PCR was adopted to detect the mRNA expression of akaline phosphatase (ALP), bone morphogenetic protein 2b (BMP-2b), runt-related protein 2 (Runx2) and cathepsin K (CTSK), phosphorus and tartrate-resistant acid phosphatase (TRAP), nuclear factor of activated T-cells, cytoplasmic-1 (NFATC-1). Divided RAW264.7 cells into control, Rankl induction (10 μg·L-1), BS (180 mg·L-1), TL (30 mg·L-1), and BSTL group (210 mg·L-1) after they were cultured to 80%-90% density. The expression of actin ring was detected by phalloidin cytoskeleton staining. The mRNA expression of TRAP, CTSK and autophagy-related genes 5 (ATG5), autophagy-related genes 7 (ATG7), and ubiquitin-binding protein p62 (p62) were detected by Real-time PCR.Result:Compared with the control group in vivo, the vertebral area and ALP, BMP-2b, and Runx2 expression of zebrafish in the Pred group were significantly decreased (P<0.01), and CTSK, TRAP, and NFATC-1 expression of zebrafish were significantly increased (P<0.01). Compared with the Pred group, the vertebral area of the TL (P<0.05) and BSTL group (P<0.01) increased significantly. The expressions of ALP, BMP-2b, and Runx2 in the BS, TL, and BSTL group were significant increased (P<0.01). The expression of CTSK, TRAP, and NFATC-1 in BSTL group were significant decreased (P<0.05, P<0.01). Compared with the control group in vitro, the number of actin rings (P<0.01) and CTSK, TRAP, ATG5 (P<0.01) expression and ATG7, p62 (P<0.01) expression were significantly increased in the Rankl-induced group. Compared with the Rankl induction group, the number of actin rings and CTSK and TRAP expression were significantly decreased in the BS, TL, and BSTL group (P<0.05, P<0.01), while the expression of ATG5, ATG7, and p62 were significant decreased in the BSTL group (P<0.05).Conclusion:BSTL can significantly improve prednisolone-induced zebrafish osteoporosis and inhibit osteoclast differentiation by reducing autophagy-related gene expression.
Abstract：Objective:To observe effect of addition and subtraction therapy of Huaihuasan combined with Taohuatang to ulcerative colitis with cold-heat complicated syndrome at active stage, and to study regulation effect to immune function and inflammatory response.Method:One hundred and twelve patients were randomly divided into control group and observation group by random number table. Patients with light and middle symptoms in control group got mesalazine slow release tablets, 1.0 g/time, 3 times/days, patients with severe symptoms or whose symptoms were not changed after getting for 4 weeks in control group got prednisone acetate tablets, 0.75 mg·kg-1·d-1 for 3 times. Based on the treatment in control group, patients in observation group added Huaihuasan combined with Taohuatang, 1 dose/day. The course of treatment was 4 weeks. At remission period, mesalazine slow release tablets were used for maintain long-term maintenance therapy, 0.5 g/times, 3 times/days. Scores of disease activities were graded by improvement mayo, and clinical remission and clinical efficacy were recorded, scores of cold-heat complicated syndrome, mucous membrane under enteroscopy and histology of mucosa belongs to Geboes were graded. And levels of tumor necrosis factor-α(TNF-α) in peripheral blood, interleukin-8 (IL-8), IL-10, T lymphocyte subsets (CD4+, CD8+), and adverse reactions, 6 months' follow-up and recurrence were also be recorded.Result:Therapeutic effect of traditional Chinese medicine syndromes were analyzed by rank sum test, which in observation group was better than that in control group (Z=1.915, P<0.05). Clinical effect in observation group was 98.04%(50/51) higher than 84.00%(42/50) in control group, clinical remission rate was 94.12%(48/51) higher than 78.00%(39/50) in control group, and mucosal healing rate was 96.08%(49/51) higher than 82.00%(41/50) in control group (P<0.05). Scores of mayo, cold-heat complicated syndrome, colonic mucosa and index scores of Geboes were all lower than those in control group (P<0.01). And levels of TNF-α, IL-8 and CD8+ were lower than those in control group (P<0.01), and levels of IL-10, CD4+ and CD4+ /CD8+ were higher than those in control group (P<0.01). Recurrence rate during 6 months in observation group was 10.42%(5/48) lower than 30.77%(12/39) in control group (χ2=5.669, P<0.05), as for adverse reactions, there was no significant difference between two groups.Conclusion:Huaihuasan combined with Taohuatang can induce UC to remission period, inhibit the activity of disease, alleviate clinical symptoms, regulate immune function and expression of inflammatory factors, alleviate inflammatory reaction, promote intestinal mucosal healing, and can maintain clinical remission and reduce recurrence. The clinical efficacy is superior to that of 5-ASA/glucocorticoid in Western medicine.
Abstract：Objective:To observe the clinical efficacy of dialectical therapy of Bufeitang combined with Shengesan and Fujiu application on chronic obstructive pulmonary disease (COPD) and lung-kidney Qi deficiency syndrome, and its effect on inflammatory damage and airway remodeling.Method:One hundred and thirty-four patients were randomly divided into control group (66 cases) and observation group (68 cases) by random number table. Patients in control group got spiriva by powder inhaler, 1 grain/time, 1 time/day, and salmeterol xinafoate and fluticasone propionate powder for inhalation for spray as appropriate, 1 suction/time, 1-2 times/days, for a continued 12 months. In addition to the therapy of control group, patients in observation group were also given Fujiu application at two-tailed acupoints of Feiyu, Piyu and Shenyu for the first day of the every San Fu and San Jiu, and dialectical therapy of Bufeitang combined with Shengesan were given at the first day of San Fu and San Jiu for 2 months. The course of treatment was 12 months. Before and after treatment, FEV1% of self-assessment questionnaire of patients with COPD (CAT), 6-min walking distance, St George's respiratory questionnaire (SGRQ), severity of dyspnea (mMRC) and index of BODE were assessed. And levels of tumor necrosis factor-alpha (TNF-alpha), interleukin-6 (IL-6), matrix metalloproteinase-9 (MMP-9) and matrix metalloproteinase inhibitor-1 (TIMP-1) were detected.Result:After treatment, scores of CAT, the total score of SGRQ, scoring of each dimension and index of BODE in observation group were all lower than that in control group (P<0.01), while FEV1% was higher than that in control group (P<0.01). And 6-min walking distance was more than that in control group (P<0.01), and the numbers of acute exacerbations were less than that in control group (P<0.01). The severity of dyspnea was lighter than that in control group (Z=2.047, P<0.05). And levels of MMP-9, TNF-α, IL-6 and ratio of MMP-9/TIMP-1 were lower than those in control group (P<0.01), whereas the level of TIMP-1 was higher than that in control group (P<0.01).Conclusion:Dialectical therapy of Bufei decoction combined with Shenge powder and Fujiu application can alleviate the current symptoms of dyspnea, improve exercise tolerance, quality of life and pulmonary function, reduce the number of acute exacerbations, relieve inflammation damage and airway remodeling. The comprehensive clinical efficacy is better than that of conventional western medicine.
Abstract：Objective:To observe the clinical efficacy of flavored Sanxitang combined with celecoxib in treating gonarthromeningitis caused by dampness and cold.Method:Totally 120 case were randomly divided into the control group and the treatment group, with 60 cases in each group. The control group was given celecoxib, and the treatment group was treated with flavored sanxitang combined with celecoxib for 30 d. The visual analogue scale (VAS), American knee association knee function scale (AKS), arthritis impact scale (AIMS), Western Ontario and McMaster Universities Osteoarthritis Index (WOMAC) were observed before and after treatment. The circumferential diameter, articular cavity effusion and synovial hyperplasia thickness were measured. The serum tumor necrosis factor-α(TNF-α), matrix metalloproteinase (MMP)-3, serum interleukin-1 (IL-1) and erythrocyte sedimentation rate (ESR) were detected. The effective rate and adverse reactions were observed in two groups.Result:The total effective rate in treatment group was 94.8% (55/58), which was higher than 80.7% (46/57) in control group (P<0.05). Compared with before treatment, VAS, AIMS, WOMAC, knee circumference, articular cavity effusion and synovial hyperplasia thickness were significantly reduced in two groups (P<0.05), while the AKS score was significantly increased (P<0.05). Compared with after treatment, VAS, AIMS, WOMAC, knee circumference, articular cavity effusion and synovial hyperplasia thickness were significantly reduced in observation group (P<0.05), whereas the AKS score was significantly increased (P<0.05). TNF-α, MMP-3, IL-1 and ESR in two groups were significantly lower than those group before treatment (P<0.05). After treatment, TNF-α, MMP-3, IL-1 and ESR levels in observation group were significantly reduced compared with those in control group (P<0.05). The incidence rate of adverse reactions in treatment group was 5.89% (4/58), which was lower than 22.81% (13/57) in control group (P<0.05).Conclusion:Flavored Sanxitang combined with celecoxib could alleviate the clinical symptoms of gonarthromeningitis caused by dampness and cold, with a low incidence of adverse reactions.
Keywords：flavored Sanxitang;celecoxib;dampness and cold syndrome;gonarthromeningitis
Abstract：Objective:To observe the effect of modified Erxian decoction on the physical condition, pain in the lower back and joints, limb activity, bone density, bone metabolism and biochemical indexes in patients with osteoporosis caused by Yang deficiency, in order to explore the possible mechanism.Method:Totally 100 cases of osteoporosis with Yang deficiency were randomly divided into treatment group and control group, with 50 cases in each group. The control group was treated with basic anti-osteoporosis therapy, and the treatment group was give modified Erxian decoction combined with basic therapy for 6 weeks. Short physical performance battery (SPPB) scores of low back and joint pain and Yang deficiency symptom score before and after treatment were recorded and analyzed. before and after treatment, the changes of blood serum calcium (Ca), phosphorus (P), L1-4, femoral neck bone mineral density(BMD), osteocalcin (BGP) and type I collagen peptide amino end level (P1NP) were measured.Result:The scores of lumbar back and joint pain, Yang deficiency symptom score, limb function and activity, lumbar spine 1-4 (L1-4), femoral neck BMD, Ca, P, BGP and P1NP between two groups before treatment had no statistically significant difference. After 6 weeks of treatment, limb activity score, L1-4, femoral neck BMD, serum Ca and P levels in treatment group were significantly higher than those in control group (P<0.05). The scores of the pain in the lower back and joints, Yang deficiency symptom score, BGP and P1NP in treatment group were significantly lower than those in control group (P<0.05).Conclusion:Modified Erxian decoction can significantly alleviate the pain in the lower back and joints of patients with osteoporosis caused by Yang deficiency, enhance the limb function and activity status, improve the physical condition and bone density of patients, reduce bone conversion, with a good effect in treating and alleviating symptoms of osteoporosis caused by Yang deficiency.
Keywords：modified Erxian decoction;osteoporosis;Yang deficiency;bone mineral density;bone metabolism
Abstract：Objective:To study the pharmacokinetics of sinapic acid from stir-fried Raphani Semen in normal rats and the correlation between pharmacokinetics-pharmacodynamics (PK-PD) in asthma rats.Method:Normal rats received 4.5, 9, 18 g·kg-1 of stir-fried Raphani Semen by oral administration, respectively. Blood was taken from ophthalmic venous plexus at different time points according to the experimental design, the plasma concentration of sinapic acid was analyzed by UHPLC-MS/MS, and data analysis was performed using DAS 3.2.8 software to obtain the pharmacokinetic parameters. Rat asthma model was established by intraperitoneal injection of ovalbumin with aluminum hydroxide, and treated with ethanol extract of stir-fried Raphani Semen (low and high doses of 4.5, 9 g·kg-1). After treatment for 3 weeks, taking blood at different time points, plasma and serum were separated. UHPLC-MS/MS was established for the determination of plasma concentration of sinapic acid, contents of interleukin-5 (IL-5), immunoglobuin E (IgE), tumor necrosis factor-α (TNF-α) in serum at different time points were detected by enzyme-linked immunosorbent assay (ELISA), DAS 3.2.8 software was used for PK-PD model fitting and data analysis.Result:After normal rats were administrated with low, medium and high doses of stir-fried Raphani Semen, the peak concentration (Cmax) of sinapic acid in plasma were (29.35±10.32), (62.70±27.47), (137.33±40.95) μg·L-1, its area under the curve (AUC0-t) were (92.83±27.16), (240.74±75.09), (633.95±195.88) μg·L-1·h, its peak time (Tmax) were (2.58±0.80), (3.00±0), (5.50±1.23) h, respectively. Compared with the low dose group, AUC0-t and mean retention time (MRT0-t) were all increased in the medium and high dose groups, showing statistical differences (P<0.05, P<0.01). The linear relationship of AUC0-t in sinapic acid was good within the dose range of 4.5-18 g·kg-1. After treating with ethanol extract of stir-fried Raphani Semen for 0.083, 0.167 h, compared with the model group of asthmatic rats, serum levels of IL-5, IgE, TNF-α of the medication groups were decreased to different degrees (P<0.05, P<0.01). Cmax of sinapic acid in the low and high dose groups were (58.43±29.94), (61.16±18.79) μg·L-1, its AUC0-t were (188.75±37.07), (247.90±36.89) μg·L-1·h, respectively. AUC0-t, apparent volume of distribution (Vz/F) and clearance rate (CLz/F) all increased significantly with the increase of dose. The best pharmacokinetic model of sinapic acid was fitted as a one-compartment model for extravascular administration, PK-PD model may be applicable to indirect connection model.Conclusion:The plasma concentration of sinapic acid is correlated with contents of IL-5, IgE and TNF-α, dosage and functional state (pathological or physiological state) can affect the pharmacokinetic behavior of sinapic acid from stir-fried Raphani Semen in rats, and it has a certain correlation with the anti-asthmatic effect.
Abstract：Objective:To prepare 15 batches of Banxia Xiexintang substance benchmark and lyophilized powder from different places, and the lyophilized powder was analyzed by ultra-high performance liquid chromatography with diode array detection (UHPLC-DAD) and desorption electrospray ionization-mass spectrometry imaging (DESI-MSI) in order to investigate the advantages of DESI-MSI in quality control of famous classical formulas.Method:Taking Banxia Xiexintang as the research model, fingerprints of the substance benchmark and lyophilized powder were established by UHPLC-DAD, and the content of index components and the yield of dry extract were also investigated. Meanwhile, as the research carrier, the lyophilized powder corresponding to Banxia Xiexintang was dissolved in methanol and dotted on qualitative filter paper with 5 μL quantitative capillary, and fixed it on the slide to make samples. The samples were analyzed on a DESI-MSI system in positive and negative ion mode with methanol-formic acid (1 000∶1, flow rate of 3 μL·min-1) as spray solvent, N2 as spray gas (pressure of 0.5 MPa). The scanning range was 100-1 200 Da, the spatial resolution was 300 μm, the spray voltage was 3 kV, the sampling cone voltage was ±40 V, incidence angle of sprayer was 60 degree, its collection angle was 10 degree, the ion source temperature was 120 ℃.Result:DESI-MSI could not only detect the index components of liquiritin, baicalin and wogonoside, as well as the common peaks of liquiritin apioside, berberine and glycyrrhizic acid, but also analyzed them semi-quantitatively, the analysis results were basically consistent with UHPLC-DAD. At the same time, DESI-MSI could detect 16 other components from Glycyrrhizae Radix et Rhizoma, Coptidis Rhizoma, Scutellariae Radix, Jujubae Fructus and Ginseng Radix et Rhizoma, such as licoricesaponin G2, palmatine, coptisine, rutin and ginsenoside Rg1, and present their relative content visually. The qualitative analysis ability of DESI-MSI was much better than UHPLC-DAD.Conclusion:DESI-MSI can be used as the quality control method for substance benchmark and lyophilized powder and dispensing granules of famous classical formulas with advantages of high sensitivity, strong analytical ability, no complex sample processing, qualitative and relative content analysis of complex samples without reference substance.
Abstract：Objective:To identify the active constituents of Paidu Qingzhi tablets by UPLC-Q-TOF-MS technique, and to reveal its potential targets and molecular mechanisms by network pharmacology.Method:Chromatography separation was achieved on an ACQUITY UPLC HSS T3 column (2.1 mm×100 mm, 1.8 μm) with mobile phase consisted of 0.1% formic acid aqueous solution and acetonitrile for gradient elution (0-17 min, 95%-5%A; 17-17.01 min, 5%-95%A; 17.01-20 min, 95%A), the flow rate was 0.3 mL·min-1, the column temperature was 40 ℃. Data acquisition was carried out in electrospray ionization (ESI) under the negative ion mode, the scanning range was 50-1 200.Ingredients in Paidu Qingzhi tablets were identified according to reference substance, relative molecular weight, mass spectrometric cleavage rule and literature information. Multiple databases were used to retrieve the targets of these identified ingredients and related diseases treated by the tablets, and metabolic pathways. A visual network of " herbs-compounds-targets-pathways-diseases" was constructed by Cytoscape 3.7.1.Result:A total of 33 active compounds in methanol extract of Paidu Qingzhi tablets were identified by UPLC-Q-TOF-MS, including 27 compounds from Rhei Radix et Rhizoma, 4 compounds from Panacis Quinquefolii Radix, 2 compounds from Ophiopogonis Radix. Totally 18 direct targets and 58 indirect targets corresponding to 31 ingredients were obtained, 7 metabolic pathways including steroid hormone biosynthesis pathway, arachidonic acid metabolic pathway, insulin signaling pathway were strongly correlated with the treatment of hyperlipidemia, acne and simple obesity by Paidu Qingzhi tablets.Conclusion:Ingredients of Paidu Qingzhi tablets are revealed by UPLC-Q-TOF-MS, based on these identified ingredients, targets and related metabolic pathways are visualized by network pharmacology. The current research can provide theoretical basis for quality control and clinical application of Paidu Qingzhi tablets.
Abstract：Objective:To establish an ultra-high-performance liquid chromatography (UHPLC) method for simultaneous determination of seven components(chlorogenic acid, 3, 5-dicaffeoylquinic acid, 1, 5-dicaffeoylquinic acid, kaempferol-3-O-rutinoside, quercetin, kaempferol and thymol) in blossoms of Inula nervosa, and provide references for its quality control.Method:The separation was performed on an Agilent Poroshell 120 C18 column (3.0 mm×100 mm, 2.7 μm) with 0.1% formic acid(A) and methyl (B)as mobile phase for gradient elution(0-4 min, 2%B; 4-6 min, 2%-5%B; 6-10 min, 5%-10%B; 10-20 min, 10%-20%B; 20-30 min, 20%-27%B; 30-37 min, 27%-25%B, 37-45 min, 25%-32%B; 45-68 min, 32%-58%B; 68-75 min, 58%-25%B; 75-82 min, 25%-2%B; 82-90 min, 2%B). The flow rate was 0.3 mL·min-1 and the detection wavelength was 254 nm.Result:There was a good linear relationship between the concentration and peak area of all the seven components in the investigated concentration range (r>0.999). The average recoveries ranged from 97.80% to 101.28% with RSD≤3.0%. Cluster analysis of SPSS software and principal component analysis of SIMCA software can be used to intuitively classify samples from four different origins.Conclusion:The established method is simple and fast with high precision, which can be used to compare the differences of blossoms of Inula nervosa from different origins and efficiently control its quality.
Keywords：blossoms of Inula nervosa;UHPLC;index components;principal component analysis
Abstract：Objective:To analysis the chemical constituents in Sancao Baogan decoction by ultra-high performance liquid chromatography coupled with hybrid quadrupole-orbitrap mass spectrometry (UPLC-ESI-HRMSn).Method:The separation was performed on an ACQUITY UPLC BEH C18 column (2.1 mm×100 mm, 1.7 μm) by a gradient elution of methanol (A)-0.1% formic acid solution (B) (0-2 min, 5%A; 2-20 min, 5%-12%A; 20-35 min, 12%-40%A; 35-38 min, 40%A; 38-48 min, 40%-80%A; 48-50 min, 80%A). The flow rate was 0.3 mL·min-1 and the column temperature was set at 30 ℃, the injection volume was 10 μL. Electrospray ionization was applied and the data were collected via positive and negative ion modes. By using Xcalibur 3.0 software, the chemical constituents were analyzed based on the relative retention time, excimer ion peak and fragment ion peak of the compounds, as well as comparison with human metabolome database (HMDB), reference substances and literature data.Result:A total of 40 chemical components were identified from Sancao Baogan decoction, including 16 phenolic acids, 19 flavonoids, 2 anthraquinones, 1 triterpenoid, 1 sterol, and 1 monoterpenoid. Six compounds (dansensu, α-pinene, epigallocatechin, 2, 5-dihydroxybenzoic acid, naringenin and emodin) were reported for the first time from Sancao Baogan decoction.Conclusion:The established UPLC-ESI-HRMSn can quickly, accurately and comprehensively analyze the chemical constituents of Sancao Baogan decoction, which lays a foundation for the basic research of pharmacodynamic substances and quality control of this formula.
Abstract：Objective:To analyze and identify the chemical constituents of Citri Sarcodactylis Fructus by an ultra-high performance liquid chromatography coupled with hybrid quadrupole-orbitrap high-resolution mass spectrometry (UPLC-Q-Orbirap HRMS) method.Method:Compounds were separated on Thermo Scientific Accucore™ C18 column (3 mm×100 mm, 2.6 μm). The mobile phase was 0.1% formic acid solution and 0.1% formic acetonitrile solution. The flow rate was 0.3 mL·min-1, and the column temperature was set at 30 ℃. HRMS was performed using an electrospray ion source (ESI), and scanned in a positive ion mode by means of full scan/data dependent secondary scan (Full MS/dd-MS2). Compound Discoverer 3.0 software that could be linked to mzCloud network database and local high-resolution mass spectrometry database of traditional Chinese medicine components was used to analyze the data, based on accurate molecular mass, retention behaviors and characteristic ion fragmentation of the compounds, as well as literature information and relevant reference materials.Result:Totally 54 chemical constituents in Citri Sarcodactylis Fructus were identified, including 16 flavonoids, 17 coumarins, 3 limonoids, 6 nucleosides and nucleobases, 2 organic acids, 3 aromatic aldehydes, 1 amino acid and 6 other components.Conclusion:The established UPLC-Q-Orbitrap HRMS method can be used to effectively and rapidly identify main chemical constituents of Citri Sarcodactylis Fructus. The findings provide a methodological reference and theoretical foundation for defining the pharmacodynamic material base and optimizing quality control index of Citri Sarcodactylis Fructus, which is of guiding significance for further development and utilization of the resources.
Abstract：Objective:To screen out active fraction from Euphorbia fischeriana, separate active components from E. fischeriana and explore structure-activity relationships, in order to analyze and identify chemical compositions of petroleum ether fraction from E. fischerian ethanol extract by ultra-performance liquid chromatography with quadrupole-time-of flight mass spectrometry (UPLC-Q-TOF-MS).Method:The anti-tumor activities of petroleum ether, ethyl acetate, n-butanol and water extracts from E. fischeriana were tested by methyl thiazolyl tetrazolium(MTT)method. A variety of modern chromatographic separation methods were used to separate active compounds from petroleum ether layer. Compounds were isolated. Their structures were identified by NMR technique. The structure-activity relationships between anti-tumor activities and structures of compounds were investigated. UPLC-Q-TOF-MS technique was used to identify the structures of petroleum ether extract from E. fischeriana. Mass spectrometry was performed in the positive ion mode using ESI ion sources.Result:Six compounds were isolated from petroleum ether fraction. They were jolkinolide A, jolkinolide B, 17-hydroxyjolkinolide A, 17-hydroxyjolkinolide B, euphopilolide and atis-16-en-13(s)-hydroxy-3, 14-dione. A total of 23 peaks were identified based on the comparison of retention times, accurate masses and fragmentation patterns with available standard compounds and literatures. Among them, there were 19 diterpenoids, 2 polyphenols, 1 fatty acid and 1 triterpenoid. Peaks No.18 and No.21 were tentatively identified as new compounds.Conclusion:The petroleum ether fraction showed a potential anti-tumor activity. The structure-activity relationships were discussed. UPLC-Q-TOF-MS technology can be used to quickly and accurately identify the structures, so as to provide a reference for its quality evaluation and active ingredient research.
Abstract：Objective:To explore the correlation between bulb quality and rhizosphere soil factors of Fritillaria taibaiensis of different origins and years, in order to provide theoretical basis for the high quality and safe production of F. taibaiensis.Method:Totally 11 samples of bulb and rhizospheric soil of F. taipaiensis of different origins and years were taken as the research objects. Available N, available P, available K, organic matter, pH and six soil enzyme activities in rhizosphere soils were determined by soil agrochemical analysis method. Peimisine and nine nucleosides in F. taibaiensis bulbs were determined by HPLC, and total alkaloid content was determined by UV. SPSS 22.0 software was used to analyze the correlation of the measured data.Result:There were significant differences in rhizosphere soil factors and bulb quality between F. taibaiensis of different origins and years (P<0.05). In terms of soil factors, the contents of available N, available K, organic matter and six soil enzyme activities in rhizosphere soil of wild varieties were higher than those of cultivated varieties, while the contents of available P and pH were lower than those of cultivated varieties. With the increase of growth years, the soil nutrient index of cultivated varieties showed different change trends, while that of wild varieties did not change significantly. However, most of the soil enzymes in both groups decreased in varying degrees. In terms of bulb quality, the contents of nine nucleosides and alkaloids in F. taibaiensis bulbs decreased with the increase of growth years, with larger change trends of cultivated varieties, while that of wild varieties was not significant. The contents of nucleosides and alkaloids in most cultivated varieties were higher than those in wild varieties. The correlation analysis showed certain correlations between soil factors in rhizosphere as well as soil factors and bulb quality. In general, soil nutrient status and bulb quality decreased with the increase of years.Conclusion:The quality of F. taibaiensis is mainly affected by its rhizosphere soil factors. In the process of field conservation and artificial cultivation, attention shall be paid to increase or decrease of the content of soil nutrients and their proportional relationship according to actual situations, so as to ensure the quality of F. taibaiensis.
Abstract：Objective:Based on gene array technology, gene set enrichment analysis (GSEA) and immune infiltration analysis were performed on chip data of intracranial aneurysm (IA) mRNA expression profile, in order to provide theoretical basis for understanding the formation mechanism of IA.Method:The GSE75436 raw data were obtained from the gene expression omnibus (GEO). GSEA of biological process (BP) in gene ontology (GO) and Kyoto gene and genome encyclopedia (KEGG) signaling pathways were analyzed for gene expression profile by R software. The CIBERSORT deconvolution method was used to analyze the infiltration ratio of 22 types of immune cells in the expression profile. And COREMINE database was used to predict traditional Chinese medicines (TCMs), which were significant correlation with the enrichment result.Result:The GSEA results showed that the changes in gene expression of IA samples mainly involved in the regulation of cytokines, activation and differentiation of leukocyte, inflammatory immune response and other processes. The infiltration matrix analysis of immune cells showed that mast cells resting and neutrophils were significantly reduced in IA samples. The comparison of paired samples showed that mast cells and natural killer cells (NK cells) were significantly activated in the IA samples of the same individual, while neutrophils and T cells CD4 naive were significantly reduced. Through COREMINE prediction, it was found that Stephaniae Tetrandrae Radix was correlated with the activation of granulocytes, Sapindi Mukorossi Semen and Pistaciae Chinensis Cortex were correlated with the activation of neutrophils, Trichosanthis Semen, Paeoniae Radix Alba and Ligustri Lucidi Fructus were correlated with the cytotoxicity mediated by NK cells.Conclusion:Activation of mast cells and NK cells are closely associated with the occurrence and development of IA. The inflammatory immune processes and pathways such as nucleotide-binding oligomerization domain (NOD)-like receptor (NLR) signaling pathway and cytotoxicity mediated by NK cells may be important factors in the pathogenesis of IA, and TCMs such as Stephaniae Tetrandrae Radix may be the potential molecular drug sources.
Keywords：intracranial aneurysm;gene expression profile;gene set enrichment analysis;immune infiltration;inflammatory cells;subarachnoid hemorrhage;traditional Chinese medicine
Abstract：Objective:To discover a small molecule active ingredient of traditional Chinese medicine (TCM) with the inhibitory activity of histone deacetylase (HDAC) 3/8.Method:The molecular docking technique was performed by AutoDock 4.2.6 software. Trichostatin A (TSA) was used as a reference to screen 19 small molecular components from TCM, and the default docking conformation number was set to obtain the docking binding energy, active site amino acid residues and hydrogen bonds, and the biological activity was verified.Result:The binding energies of 19 small molecule components from TCM to HDAC3 and HDAC8 were different. Among them, ursolic acid, fangchinoline and tetrandrine have low binding energies to HDAC3 and HDAC8, and their binding activities were strong. The optimal binding energy of fangchinoline and HDAC3 at the site 1 was the lowest (-26.71 kJ·mol-1), and that of HDAC8 at the site 9 was the lowest (-26.84 kJ·mol-1). The optimal binding energy of tetrandrine and HDAC3 at the site 13 was the lowest (-26.38 kJ·mol-1), and that of HDAC8 at the site 12 was the lowest (-25.41 kJ·mol-1). In addition, the binding energy of ursolic acid and HDAC3 at the site 16 was the lowest (-25.83 kJ·mol-1), and that of HDAC8 at the site 8 was the lowest (-35.62 kJ·mol-1). Three kinds of amino acids at the docking site of small molecules were rendered by PyMOL 2.3.1.When ursolic acid was combined with HDAC3/8, the active sites produced two hydrogen bonds, and the interaction was strong, and many amino acids were connected at the active site. The fangchinoline formed two hydrogen bonds with the active site of HDAC3 and one hydrogen bond with the active site of HDAC8, and hydrophobic binding with some active site amino acids. There was no hydrogen bond between tetrandrine and HDAC3/8, and all docking sites were docked by 4 active amino acids. Three small molecules (ursolic acid, fangchinoline and tetrandrine) with the best docking effect had the inhibitory activity against HDAC3/8 at the concentration of 500 μmol·L-1 and 100 μmol·L-1, and the inhibitory activity was still optimal among the 10 selected small molecules.Conclusion:Among the screened 19 small molecules, ursolic acid, tetrandrine and fangchinoline may be the new anti-inflammatory drugs of HDAC3/8 inhibitory target, which provides a reference for further exploration and discovery of new anti-inflammatory drugs.
Abstract：Objective:Based on the protective effect of Guhong injection (GH) on cerebral ischemia, mechanism of GH against cerebral ischemia was identified using RNA-seq transcriptome and bioinformation analysis.Method:The model of middle cerebral artery occlusion (MCAO) was established through thread embolization. Sham group, model group, low-dose GH group (0.625 mL·kg-1·d-1), high-dose GH group (2.5 mL·kg-1·d-1), positive group (Ginaton, 8 mL·kg-1·d-1) were set up. Ludmila Belayev 12-point scoring method was applied to assess the protective effect of GH against MCAO-induced cerebral ischemia. And the differentially expressed genes after treatment with GH were identified by RNA-Seq technology. Enrichment analysis, cluster analysis and association analysis on disease targets of cerebral ischemia were carried out through such databases as DAVID, String and The Human Phenotype Ontology. Finally, the regulatory network was constructed by Cytoscape3.4.0.Result:Compared with the sham group, the neurological impairment was obvious in the model group (P<0.01), and the neurological impairment was alleviated in the GH group compared with the model group (P<0.05). RNA-Seq technology analysis showed that GH regulated genes involving such biological processes as cell apoptosis, inflammation, oxidative stress, toll-like signaling pathway and mitogen-activated protein kinase (MAPK) signaling pathway. Twenty disease targets and 64 MAPK signaling pathway genes were associated with differentially expressed genes after GH treatment, in which 23 genes were involved in apoptosis and inflammation.Conclusion:GH protected against cerebral ischemia in many ways, among which MAPK signaling pathway is an important way to exert its effect in inhibiting apoptosis and inflammation.
Keywords：Guhong injection;cerebral ischemia;RNA-Seq;mitogen-activated protein kinase (MAPK)
Abstract：Objective:To identify the main active components of Coptidis Rhizoma and Euodiae Fructus for the treatment of gastric cancer, predict the targets of the common active components in these two herbs, establish the network of active drug components-target genes, and further explore the potential mechanism and effect of Coptidis Rhizoma-Euodiae Fructus for the treatment of gastric cancer.Method:The active components of Coptidis Rhizoma-Euodiae Fructus were screened by Traditional Chinese Medicine Systems Pharmacology Database (TCMSP), gastric cancer targets were predicted and screened by Genecard database platform, " drug-active ingredient-disease-targets" networks were built by Cytoscape (3.7.1) software, and protein interaction networks were built by String database platform. Finally, Gene Ontology (GO) analysis and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis were performed by using Bioconductor platform and R language.Result:The 14 potential active components of Coptidis Rhizoma and 15 potential active components of Euodiae Fructus were obtained, involving 127 targets related to gastric cancer. There were 33 common targets for Coptidis Rhizoma-Euodiae Fructus-gastric cancer, which played a therapeutic role in gastric cancer mainly by regulating target genes such as PTGS2, PTGS1, AR, RXRA and NOS2, as well as cell apoptosis, p53 signaling pathway and interleukin (IL)-17 signaling pathway.Conclusion:The therapeutic mechanism of Coptidis Rhizoma-Euodiae Fructus reflects the multi-component, multi-target and multi-pathway characteristics of traditional Chinese medicines and provides the scientific basis for further study and the material basis of Coptidis Rhizoma-Euodiae Fructus against gastric cancer.
Abstract：Post-traumatic stress disorder (PTSD) is a mental disorder characterized by restitution of trauma and emotional irritability and avoidance caused by sudden, threatening or catastrophic life events. PTSD occurs not only abnormal behavioral indicators but also abnormal neurobiological indicators. Studies on the pathogenesis of PTSD have focused on neuroendocrine system, central nervous system and immune system, mainly finding abnormal levels of stress hormones, pro-inflammatory cytokines, neurotransmitters and neuroprotective factors. Traditional Chinese medicines(TCM) have many advantages in the treatment of PTSD due to their multi-target, multi-component and multi-pathway characteristics. Current studies have showed that a variety of TCM can improve the abnormal behaviors of animals in the models of post-traumatic stress disorder and reverse the neurobiological indicators. As many of these TCM compounds have been used in clinic and have good efficacy, TCM treatment of PTSD has a broad application prospect. However, there is no specific medicine for PTSD, and its mechanisms have not been fully understood. Therefore, this paper reviews the current studies on the treatment of PTSD with TCM, and focuses on the occurrence and treatment progress of PTSD in terms of TCM, mechanisms of action and animal models, so as to provide references for the treatment of PTSD.
Keywords：post-traumatic stress disorder;traditional Chinese medicine(TCM) monomers;single TCM extracts;TCM compounds
Abstract：Erigeron breviscapus, a species within the genus of Erigeron, is mainly distributed in Southwest China. It is cold in property, slightly bitter in taste, and has the effect of dispersing cold table, removing wind and dehumidification, promoting blood circulation and removing blood stasis, relieving pain and inflammation. Breviscapine is the extract of E. breviscapus. It is mainly consisted of flavonoids, lignans, coumarins, terpenes, phytosterols, etc. As the major components of breviscapine, the content of breviscapine b (4′-hydroxybaicalin-7-O-glucuronide) and breviscapine a (apigenin-7-O-glucuronide) is greater than 90%. Modern pharmacological studies have shown that breviscapine has a wide range of pharmacological effects, including anti-oxidation, anti-fibrosis, anti-inflammation, anti-aging, anti-platelet aggregation, lowering blood lipid, increasing blood flow, improving microcirculation, preventing and treating tumors, and resisting brain injury. In clinical, breviscapine has been widely used in the treatment of diabetes, cerebral insufficiency, sequelae caused by cerebral hemorrhage, hypermucolipemia, cerebral thrombosis, kidney disease, liver disease, Alzheimer's disease, and some other complex diseases. Specially, in the treatment of diabetes and its chronic complications, such as diabetic nephropathy, diabetic cardiomyopathy, diabetic foot, diabetic retinopathy, breviscapine has showed significant efficacy. In addition, studies have demonstrated that the combined application of breviscapine, mecobalamine, and micopol can improve the therapeutic effect. In this work, the application of breviscapine in the treatment of chronic complications of diabetes mellitus and its related combination drugs were reviewed, by which we attempted to provide some valuable clues for the clinical application of breviscapine in the treatment of diabetes mellitus and its chronic complications.
Keywords：breviscapine;diabetes;research progress;mechanism of action;combination
Abstract：The incidence and mortality of gastric cancer are higher among all kinds of tumors, and the number of deaths due to gastric cancer in China is as high as 498 000.Because the pathogenesis of early gastric cancer is not obvious, most patients who have been treated in hospitals are already in middle and advanced stages of gastric cancer, which brings a heavy burden to families and the society. The postoperative survival of patients of advanced gastric cancer is significantly lower than those of early gastric cancer. The most important reason is lymphatic metastasis of gastric cancer. Lymphatic metastasis is the main metastasis pathway of gastric cancer. The lymphatic metastasis rate of progressive gastric cancer is as high as 70%, and early gastric cancer can also have lymphatic metastasis. The lymph node metastasis rate of gastric cancer was positively correlated with the infiltration depth of the tumor. In addition to surgical resection, lymphatic chemotherapy and molecular targeted therapies, traditional Chinese medicine(TCM) has also played an important role due to the advantages of a low toxicity, good therapeutic effect and low price. According to the theory of TCM, the occurrence of gastric cancer is caused by unclean diet, emotional disorders, and deficiencies, which result in phlegm, Qi resistance, blood stasis, and attack the stomach. The pathogenesis is the asthenia in origin and asthenia in superficiality, and clinical TCM therapies mostly focus on phlegm and blood stasis, spleen and Qi, warm and cold. In recent years, a number of studies have proved that TCM has a significant and effective effect in the treatment of lymphatic metastasis of gastric cancer. It may inhibit the anoikis and the tumor by inhibiting the hydrolysis of matrix metalloproteinase on the basement membrane and extracellular matrix (ECM), stimulating tumor anoikis, suppressing lymphatic neovascularization, regulating tumor-associated gene expression and other pathways, so as to treat lymphatic metastasis of gastric cancer. This article reviews the advance of theoretical research, experimental research and clinical research of TCM in the treatment of lymphatic metastasis of gastric cancer in recent years, and reveals the mechanism of TCM and its targets, in order to provide scientific basis for further studies and clinical application of the effect of TCM on the lymphatic metastasis of gastric cancer.
Keywords：gastric cancer;lymphatic metastasis;traditional Chinese medicine
Abstract：Depression is a serious threat to human health. There are grave difficulties in its prevention and treatment. At present, Western medicine has not yet found a definitive and effective therapy for depression. By virtue of its holistic view, rich clinical practice plus less side effects, traditional Chinese medicine(TCM) may have potential advantages in the prevention and treatment of depression. Single-herbal TCM or TCM compound is one of the main methods for TCM to treat depression. The chemical composition of natural medicine is quite complicated, moreover, the composition of TCM compound is more complicated. Therefore, this paper focuses on the role of single-herbal TCM in preventing and treating depression. Many researchers have discovered more TCM with the effect in preventing depression from historical literatures or clinical experience of different doctors. There are four main causes in the pathogenesis of depression: the first is the abnormal expression of monoamine neurotransmitters and their receptors, the second is the depression of nerve cells and relevant signaling pathways, the third is the hypothalamus-pituitary-adrenal axis that continuously intensifies to cause dysfunction and depression, and the fourth is the abnormal secretion of cytokines. Which mechanism does TCM work through? The meridian tropism theory is an important guiding principle in the clinical use of TCM. Is there a certain relationship between the different meridian tropisms and different pharmacological mechanisms of TCM? To solve these two basic problems, this paper systematically summarizes all the literatures on the mechanism of TCM in the prevention and treatment of depression, and uses the chi-square test to analyze the relationship between the TCM and the four mechanisms of action. This exploration may contribute to further clinical and experimental research on the prevention and treatment of depression with TCM.
Keywords：depression;pathogenesis;prevention and treatment with traditional Chinese medicine
Abstract：There are 16 species and 7 varieties of medicinal plants of Trollius in China, with effects in clearing heat, detoxification and swelling, they are used for the treatment of acute and chronic tonsillitis, acute otitis media, chronic bronchitis and urinary tract infections. Through retrieval of literatures on nasturtium from 1972 to 2019 in a number of databases, such as CNKI, Wanfang Database, Baidu Academic and PubMed, the cultivation and quality control methods, extraction and purification processes, chemical composition, pharmacological effects, pharmacokinetics and metabolism of medicinal plants of the genus nasturtium were summarized. The researches of cultivation mainly focus on the establishment of seed treatment and tissue culture system. Quality control is mainly based on orientin and flavonoids. Research on extraction and purification technology mainly focused on the extraction and purification of total flavonoids. In terms of chemical composition, there are currently more than 100 major compounds isolated from this genus, including flavonoids, organic acids, alkaloids, coumarins and styrenes. Among them, there are 80 flavonoids, 12 organic acid monomers, 3 alkaloid monomers, 4 coumarin monomers and 14 styrene monomers. In terms of pharmacological action, there are mainly in vitro experiments of crude extracts of medicinal materials. The data of pharmacokinetics and metabolism mainly focus on the study of flavone monomers.
Keywords：Trollius;cultivation;quality control;extraction and purification;chemical composition;pharmacological action