Abstract：ObjectiveTo investigate the effects of 4 weeks of aerobic exercise combined with traditional Chinese medicine （TCM） on the regulation of phosphatidylinositol 3-kinase （PI3K）/protein kinase B （Akt）/glycogen synthase kinase-3β （GSK-3β） signaling pathway and related apoptotic proteins in AD rats， and to explore its mechanism of action in improving cognitive function of AD rats.MethodFifty male SD rats were randomly divided into Sham group， exercise （EX） group， Alzheimer's disease （AD） group （model）， TCM group （Dabuyuan Jian）， and EX+TCM group， 10 in each group. Aβ25-35 oligomer solution was injected into the hippocampal region to establish the AD rat model， and the EX group underwent 6 days/week of running table training. The TCM group was given Dabuyuan Jian （5.36 g∙kg-1） by gavage， and the EX+TCM group received running table training combined with intragastric administration of Dabuyuan Jian. An equal volume of purified water was given to the Sham， AD， and EX groups by gavage. The rats' learning memory was evaluated by Morris water maze. The morphological changes and ultrastructural changes of hippocampal neurons were observed by hematoxylin-eosin （HE） staining and transmission electron microscopy， respectively. Western blot was performed to detect the protein expression levels of Akt， p-Akt， GSK-3β， p-GSK-3β， β-catenin， Bcl-2， Bax， and cleaved Caspase-3 in the hippocampus of rats.ResultCompared with the Sham group， the AD group showed an increase in escape latency and total swimming distance， and a decrease in the residence time in the target quadrant （P<0.01）， with loose and disorganized arrangement of hippocampal pyramidal cells， a decrease in density， myelin-like changes in hippocampal ultrastructure， and a blurring of mitochondrial swelling. In addition， the expression of Bcl-2， p-Akt， p-GSK-3β and β-catenin was down-regulated （P<0.05， P<0.01） while the expression of cleaved Caspase-3 was up-regulated （P<0.05）. Compared with the conditions in the AD group， the escape latency and swimming distance were reduced in the EX， TCM， and EX+TCM groups， while the residence time in the target quadrant was increased （P<0.05，P<0.01）， with regularly arranged pyramidal cells， elevated number of cells， and reduced myelin-like changes and improved mitochondrial swelling as seen by transmission electron microscopy. Moreover， the expression of Bcl-2， p-Akt and p-GSK-3β was up-regulated， while the expression of cleaved Caspase-3 was down-regulated（P<0.05，P<0.01）. The EX+TCM group had increased level of β-catenin while the EX and TCM groups were not statistically significant compared with the AD group. Compared with the EX+TCM group， the EX and TCM groups had increased escape latency and swimming distance and decreased residence time in the target quadrant（P<0.05，P<0.01）， with loosely arranged pyramidal cells and reduced synaptic integrity under transmission electron microscopy（P<0.01）. Furthermore， the levels of Bcl-2， p-Akt， p-GSK-3β and β-catenin were lowered （P<0.05，P<0.01）while the level of cleaved Caspase-3 was elevated（P<0.05）.ConclusionAerobic exercise combined with Dabuyuan Jian could improve the cognitive dysfunction of AD rats， and the mechanism might be related to the elevated expression of key proteins in the PI3K/Akt/GSK-3β signaling pathway. The combined use was better than the use of exercise or drugs alone.
Abstract：ObjectiveTo investigate the effect of Gegen Qinliantang （GGQLT）-medicated serum on free fatty acid （FFA）-induced nonalcoholic steatohepatitis （NASH） in vitro model of human hepatoma cells HepG2.MethodNASH model of HepG2 cells was established in vitro， and the cells were intervened with different volume fractions of GGQLT-medicated serum and resveratrol. Intracellular lipid deposition in each group was detected by oil red O staining， the level of reactive oxygen species （ROS） in each group were detected by flow cytometry， the levels of glutathione peroxidase （GSH-Px）， superoxide dismutase （SOD）， triglyceride （TG） and malondialdehyde （MDA） in each group were detected by kits. Real-time fluorescence quantitative polymerase chain reaction （Real-time PCR） was used to measure the mRNA expression levels of nuclear transcription factor （NF）E2-related factor 2 （Nrf2）， heme oxygenase-1 （HO-1）， quinone oxidoreductase 1 （NQO1）， Kelch-like epichlorohydrin-associated protein-1 （Keap1）， NF-κB， thioredoxin interacting protein （TXNIP）， interleukin-1β （IL-1β） in HepG2 cells of each group. The protein expression of Nrf2， TXNIP in cells of each group was detected by Western blot.ResultFFA induced large accumulation of intracellular lipids. Compared with the normal group， the activities of GSH-Px and SOD were significantly decreased （P<0.01） and the contents of TG， ROS and MDA were significantly increased （P<0.05， P<0.01） in the model group. Compared with the model group， all GGQLT groups and resveratrol group could elevate intracellular SOD activity to different degrees （P<0.05， P<0.01） and significantly reduce the levels of intracellular ROS and MDA （P<0.05， P<0.01）， GGQLD high- and medium-dose groups and resveratrol group significantly elevated GSH-Px activity （P<0.01）， GGQLD medium- and low-dose groups and resveratrol group significantly decreased TG content （P<0.05， P<0.01）. Compared with the model group， GGQLT high- and medium-dose groups and resveratrol group could significantly upregulate the mRNA expression levels of Nrf2， HO-1 and NQO1 （P<0.01）， all GGQLT groups and resveratrol group could significantly downregulate the TXNIP protein expression level， as well as significantly downregulate the mRNA expression levels of Keap1， NF-κB （P<0.05， P<0.01）. Nrf2-siRNA transfection of cells revealed that Nrf2 expression was significantly downregulated （P<0.01） in the Nrf2-siRNA group of cells by comparing with NC-siRNA group at the corresponding dose of drugs， and the inhibitory effects of GGQLT and resveratrol on TXNIP， IL-1β were attenuated.ConclusionFFA induces the production of ROS and inflammatory factors in HepG2 cells， and GGQLT can improve the anti-inflammatory and antioxidant capacities of cells， and its mechanism may be related to the regulation of Nrf2/TXNIP signaling pathway， so as to improve NASH.
Abstract：ObjectiveLipopolysaccharide （LPS）-induced zebrafish inflammation model was used to evaluate the anti-inflammatory activity of different extracts from Lianggesan （LGS） and its component Glycyrrhiza Radix et Rhizoma.MethodDifferent polar fractions of LGS and Glycyrrhiza Radix et Rhizoma were obtained by the principle of similar miscibility. For toxicity observation， the zebrafish （3 day-post-fertilization） was exposed to different concentrations of extracts for 24， 48 and 72 h. The yolk sac of the zebrafish was microinjected with 0.5 g·L-1 LPS to establish the inflammation model， and then the embryos were soaked with different concentrations of extracts to observe their survival status at 72 h and the aggregation of neutrophils in yolk sac at 12 h after treatment. Hematoxylin-eosin staining was used to analyze the yolk sac of the zebrafish microinjected with LPS. Quantitative Real-time polymerase chain reaction （Real-time PCR） was performed to further investigate the anti-inflammatory effects and mechanisms of LGS and Glycyrrhiza Radix et Rhizoma.ResultThe toxicity of LGS and Glycyrrhiza Radix et Rhizoma was decreased with the increase of polarity， and the descending order was petroleum ether>ethyl acetate>n-butanol>water. Compared with model group， the extracts from different fractions of LGS and Glycyrrhiza Radix et Rhizoma prolonged the survival time of the zebrafish， and inhibited the recruitment and aggregation of neutrophils and decreased the infiltration of inflammatory cells in the yolk sac， among which the water fraction of LGS and the ethyl acetate fraction of Glycyrrhiza Radix et Rhizoma had the most significant effect （P<0.01）. In addition， compared with model group， the water fraction of LGS and the ethyl acetate fraction of Glycyrrhiza Radix et Rhizoma down-regulated the mRNA expression of interleukin-6 （IL-6） and tumor necrosis factor-α （TNF-α）， and suppressed the expression of toll like receptor 4 （TLR4） and nuclear factor kappa-B （NF-κB） in LPS-stimulated zebrafish （P<0.01）.ConclusionThe extracts from different fractions of LGS and Glycyrrhiza Radix et Rhizoma exerted protective effects in LPS-induced zebrafish by inhibiting the TLR4 and NF-κB signaling pathways. Moreover， in zebrafish model， the method of administration by soaking was applicable to the high-throughput screening of anti-inflammatory Chinese medicine， which was suitable for the evaluation of anti-LPS activity of Chinese medicine and the different extracts.
Keywords：Lianggesan;Glycyrrhiza Radix et Rhizoma;zebrafish;lipopolysaccharide （LPS）;inflammation
Abstract：ObjectiveTo establish the quality standard of Liangditang benchmark samples.MethodUltra performance liquid chromatography-quadrupole time-of-flight mass spectrometry （UPLC-QTOF-MS） was used to qualitatively analyze the chemical composition of Liangditang on the basis of molecular and fragment ion peak information with cracking law. The mobile phase was methanol （A）-0.05% phosphate aqueous solution （B） for gradient elution （0-10 min， 5%-23.5%A； 10-20 min， 23.5%A； 20-58 min， 23.5%-63%A； 58-60 min， 63%-90%A）， the flow rate was 0.8 mL·min-1， and the detection wavelength was 254 nm. Electrospray ionization was employed under positive ion mode， the detection range was m/z 100-1 700. Key quality attributes and sources were determined by comparing with single medicine and reference substances. Through mass transfer analysis of multiple batches from decoction pieces to benchmark samples， high performance liquid chromatography （HPLC） for determining the contents of index components and HPLC detection of characteristic maps were established. Through the determination of 15 batches of benchmark samples， the content range of the index components and the common peaks of the characteristic map were determined. Thin layer chromatography （TLC） was applied to the identification of 5 medicines in the formula. Moisture and dry extract yield of the benchmark samples were determined by drying method.ResultA total of 27 compounds were inferred from the benchmark samples of Liangditang， among which 9 compounds were confirmed by comparison with the control， including catalpol， harpagide， gallic acid， albiflorin， paeoniflorin， verbascoside， angoroside C， cinnamic acid and harpagoside. A method for determining the characteristic maps of the benchmark samples were established and 13 peaks were assigned， and the characteristic peaks were mainly derived from wine-processed products of Rehmanniae Radix， Scrophulariae Radix and wine-processed products of Paeoniae Radix Alba. The similarity between the characteristic map of 15 batches of benchmark samples and the control characteristic map was >0.9. Methods for the determination of paeoniflorin， harpagoside， L-hydroxyproline and glycine were established， and the contents of these four components in 15 batches of benchmark samples were within ±30% of the corresponding mean value， and the transfer rate of decoction pieces to the benchmark samples was stable and controllable. TLC was established to identify 5 prescription drugs （except Ejiao） with two kinds of test solutions， and the results showed that the method had good specificity. The average dry extract yield was 48.06%， and the average moisture was 5.58%， which were within the range of ±10% and ±30% of their mean values， respectively.ConclusionThe quality standard of Liangditang benchmark samples was as follows：the similarity between the benchmark samples and the control characteristic map is >0.9， the contents of paeoniflorin， harpagoside， L-hydroxyproline and glycine are 217-403， 24-46， 634-1 178， 1 253-2 328 mg per dose， the dry extract yield is 43.0%-53.0%， the moisture is 4.0%-7.0%， under the set detection conditions， the benchmark samples have corresponding characteristic spots by comparing with the control herbs of 5 medicines. This quality standard is stable and reliable， which fills the gap in the quality control of Liangditang， and can provide a reference for the establishment of the quality standard of Liangditang granules.
Abstract：ObjectiveTo observe the protective effect of Shenlian prescription on acute lung injury induced by particulate matter （PM） exposure in rats and explore the mechanism.MethodFifty male SD rats were randomly divided into the control group， model group， Shenlian low-dose group （4.32 g·kg-1）， Shenlian high-dose group （8.64 g·kg-1）， and roflumilast group （3.46 mg·kg-1）， with 10 in each group. Pre-administration with drugs by gavage was performed for one week. On the 8th and 11th days， the control group was instilled with normal saline in the trachea and the other groups with PM suspension to establish a rat model of acute lung injury induced by PM exposure. After modeling， drugs were given continuously until the end of the experiment. Forty-eight hours after the last exposure， the lung function of rats was detected. Then the rats were sacrificed and the lung morphological changes and pathological changes by hematoxylin-eosin （HE） staining were observed. CD68 expression in lung was detected by immunohistochemistry， and the levels of lung injury markers surfactant protein A （SP-A） and Clara cell protein16 （CC16） in serum were detected by enzyme-linked immunosorbent assay （ELISA）. The mRNA expression of interleukin-1α （IL-1α）， IL-6， IL-18， and monocyte chemoattractant protein-1 （MCP-1） in lung tissue was measured by Real-time fluorescence quantitative polymerase chain reaction （Real-time PCR）.ResultCompared with those in the control group， the rats in the model group had decreased lung function and obvious structural damage of lung tissue， PM deposition， and infiltration of CD68 positive cells. The expressions of IL-1α， IL-6， IL-18， and MCP-1 in lung tissue were increased （P<0.01）. Compared with the model group， Shenlian prescription low and high doses restored the rats' lung function injury（P<0.05，P<0.01）， improved lung morphological and pathological structure， and reduced PM deposition. Infiltration of CD68 positive cells in lung was not significantly decreased. The levels of inflammatory factors IL-1α， IL -6， IL-18， and MCP-1 in lung were lowered （P<0.01）.ConclusionShenlian prescription could protect the rats' lung injury caused by PM exposure， improve lung morphology， and reduce PM deposition and inflammatory factor expression.
Abstract：ObjectiveTo investigate the effect of Zishen Huoxue prescription on promoting neurogenesis in hippocampal CA1 region of vascular dementia （VD） rats by regulating mitophagy.MethodThe 2-VO method was used to establish the VD rat model and 60 SD rats were randomly divided into sham operation group， model group， donepezil hydrochloride group， and Zishen Huoxue prescription low-dose（8.9 g·kg-1）， medium-dose（17.8 g·kg-1） and high-dose（35.6 g·kg-1） groups. Morris water maze test was performed to detect the escape latency and the number of crossing platform in each group. The expression of phosphatase and tensin homology-induced kinase 1 （PINK1） and Parkinson protein （Parkin） mRNA in hippocampal CA1 region was detected by Real-time fluorescent quantitative polymerase chain reaction（Real-time PCR）. Western blot was used to determine the expression of mitochondrial autophagy signaling pathway-related proteins Parkin， prohibitin 2 （PHB2）， mitofusin 2 （Mfn2） and dynamin-related protein 1 （Drp1） in hippocampal CA1 region. The neurogenesis in hippocampal CA1 region was tested by Brdu method.ResultCompared with the conditions in the sham operation group， the learning and spatial memory abilities of the model group were decreased （P<0.05）， with damaged mitochondrial structure and autolysosome formation in the hippocampal CA1 region. The expressions of Parkin， Pink1 mRNA and Parkin， PHB2， and Drp1 proteins were up-regulated （P<0.05）， while the expression of Mfn2 protein and the neuronal regeneration in hippocampal CA1 region were reduced （P<0.05， P<0.05）. Compared with the conditions in the model group， Zishen Huoxue prescription enhanced the learning and spatial memory abilities of VD rats （P<0.05）， increased the number of autophagosomes in hippocampal CA1 region and improved the mitochondrial structure. The expression of Parkin， Pink1 mRNA and Parkin， PHB2， and Drp1 proteins in hippocampal CA1 region was up-regulated （P<0.05， P<0.01）while the expression of Mfn2 protein was down-regulated（P<0.05， P<0.01）. The number of new neurons in hippocampal CA1 region was also increased（P<0.05， P<0.01）.ConclusionThe promoting effect of Zishen Huoxue prescription on the neurogenesis in hippocampal CA1 region of VD rats was related to the mitophagy mediated by Pink1/Parkin signaling pathway.
Abstract：ObjectiveTo explore the underlying mechanism of Bushen Huatan prescription in alleviating postmenopausal osteoporosis （PMOP） by maintaining the balance of osteogenesis and adipogenic differentiation in ovariectomized rats with osteoporosis.MethodSeventy-five 6-month-old non-pregnant female SD rats were randomly divided into sham-operation group， model group， atorvastatin group， liviol group， and Bushen Huatan prescription group. Bilateral ovaries were removed in the four groups except the sham-operation group， while only the same mass of adipose tissue around the ovaries was removed in the sham-operation group. On the 5th week after surgery， drugs were consecutively administrated for 8 weeks. Rats in the Bushen Huatan prescription group received 9.4 mg·kg-1 of the prescription， rats in the atorvastatin group received 0.92 mg·kg-1 of atorvastatin， rats in the Liviol group received 0.23 mg·kg-1 of liviol， and rats in the model group and the sham-operation group received saline once a day. Micro-computed tomography （Micro CT） was used to detect bone mineral density （BMD） of rat tibia in each group. Hematoxylin-eosin （HE） staining was used to detect the relative area of rat bone marrow adipose tissue （BMAT） in each group. Real-time fluorescence quantitative polymerase chain reaction （Real-time PCR） and Western blot were used to detect the relative expression levels of Runt-related transcription factor 2 （Runx2）， peroxisome proliferator-activated receptor （PPARγ）， leptin （LPN）， and leptin receptor （OBR） in bone tissues.ResultAs compared with the sham operation group， the BMD of rats in the model group decreased （P<0.05）， while the relative area of BMAT increased （P<0.05）. In addition， the expression levels of LPN， OBR， and Runx2 decreased in the model group （P<0.05）， while the level of PPARγ increased （P<0.05）. As compared with the model group， the BMD of rats in the atorvastatin group， the Livial group， and the Bushen Huatan prescription group increased （P<0.05）， and the relative area of BMAT decreased （P<0.05）. The expression levels of LPN， OBR， and Runx2 in these groups increased （P<0.05）， while the expression level of PPARγ decreased （P<0.05）.ConclusionBushen Huatan prescription plays the anti-osteoporosis role in the rat model of PMOP through up-regulating LPN and OBR in bone tissues and maintaining the balance of osteogenesis and adipogenic differentiation， thereby reducing postmenopausal bone loss and playing a role in the prevention and treatment of PMOP.
Abstract：ObjectiveTo explore the mechanism of Jianpi Yishen Huazhuo prescription in the improvement of ovarian function in polycystic ovary syndrome （PCOS）.MethodSeventy female SD rats in SPF grade were randomly divided into 6 groups， 15 in the blank group and 15 in the model group， 10 in the metformin group （0.1 g·kg-1·d-1）， and 10 in the low （1.275 g·kg-1·d-1）， medium （2.55 g·kg-1·d-1）， and high-dose （5.10 g·kg-1·d-1） Jianpi Yishen Huazhuo prescription groups. The blank group was given normal saline （10 mL·kg-1·d-1） by gavage and ordinary feed， and the other groups were given letrozole （1 mg·kg-1·d-1） by gavage combined with high-fat feed for 21 days to induce the model of PCOS. After modeling， the blank group and model group were given equal volume normal saline by gavage， and each drug group was given the corresponding dose of the drug by gavage for 30 days. The changes in body mass and fasting blood glucose （FPG） of rats before and after modeling were compared. Hematoxylin eosin （HE） staining was used to observe the morphological change in the ovaries of rats in each group. The serum levels of follicle stimulating hormone （FSH）， luteinizing hormone （LH）， testosterone （T）， anti-Mullerian hormone （AMH）， and estradiol （E2） were measured by enzyme-linked immunosorbent assay （ELISA）， and the LH/FSH ratio was calculated. Immunohistochemical staining （IHC） and Western blot were used to detect the protein expression levels of nucleoside binding oligomerization domain protein like receptor 3 （NALP3）， apoptosis-associated speck-like protein （ASC）， cysteine protease-1 （Caspase-1）， nuclear transcription factor-κB （NF-κB）， interleukin-1β （IL-1β）， interleukin-18 （IL-18）， and interleukin-6 （IL-6） in the rat ovaries.ResultAs compared with the blank group， large follicles with polycystic expansion were found in the ovaries of the model group， no dominant follicles were found， the granular layer of follicles decreased and arranged loosely， and the number of corpus luteum decreased significantly. Serum T， LH， AMH and LH/FSH increased in the model group （P<0.05， P<0.01）， while FSH and E2 decreased （P<0.05， P<0.01）. The relative protein expression levels of NALP3， ASC， Caspase-1， NF-κB， IL-1β， IL-18， and IL-6 increased （P<0.05， P<0.01） in the ovaries of the model group. Compared with the model group， the low， medium， and high-dose Jianpi Yishen Huazhuo prescription groups and the metformin group showed growing follicles and corpus luteum at all levels， the number of cystic expanding follicles decreased， the thickness of follicular granular layer increased， the number of follicular fluid increased， mature follicles were visible， and the local morphology of oocytes was complete. Serum T， LH， AMH， and LH/FSH in these groups decreased （P<0.05， P<0.01）， while E2 and FSH increased （P<0.05）. The relative protein expressions of NALP3， ASC， Caspase-1， NF-κB， IL-1β， IL-18， and IL-6 in the ovaries of these groups decreased （P<0.05， P<0.01）. There was no significant difference among the treatment groups.ConclusionBy inhibiting the activation of NLRP3 inflammasome， Jianpi Yishen Huazhuo prescription reduces the release of NALP3， ASC， Caspase-1， NF-κB， IL-18， IL-1β， and IL-6 inflammatory factors in ovarian tissues， regulates endocrine level， and effectively reduces PCOS inflammatory statu， so as to play a role in improving ovarian function.
Keywords：Jianpi Yishen Huazhuo prescription;polycystic ovary syndrome;nucleoside binding oligomerization domain protein like receptor 3 （NLRP3）;rats
Abstract：ObjectiveTo investigate the effect of pulsatilla saponin A （PSA） on proliferation and apoptosis of human Burkitt lymphoma （BL） cell line Raji cells and expression of related pathway proteins.MethodWith Raji cells as the research object， the cell proliferation was detected by cell counting kit-8 （CCK-8） method， and the half-maximal inhibitory concentration （IC50） values of 24 h， 48 h and 72 h were calculated to be 19.77， 18.31， 16.70 μmol·L-1， respectively. In subsequent related experiments， 0， 8， 16， 32 μmol·L-1 PSA were selected according to the IC50 value of Raji cells treated with PAS for 72 h. After 0， 8， 16， 32 μmol·L-1 PSA acted on Raji cells for 24， 48， 72 h， the optical density values of cell growth curve were detected by CCK-8 method. The zymogen activities of cysteine aspartate-specific protease （Caspase）-3， Caspase-8 and Caspase-9 in Raji cells treated with 0， 8， 16 and 32 μmol·L-1 PSA for 24 h were measured by Caspase-3， Caspase-8 and Caspase-9 colorimetric assay kit. The apoptosis rate and cell cycle of Raji cells treated with different concentrations of PSA after 24 h were detected by flow cytometry. The expression of B-cell lymphoma 2 （Bcl-2）， Bcl-2-associated X protein （Bax）， cleaved poly（ADP-ribose） polymerase （cleaved PARP）， cleaved cysteinyl aspartate-specific protease-3 （cleaved Caspase-3） apoptosis related protein and Janus kinase 2 （JAK2）， signal transducer and activator of transcription 3 （STAT3）， phosphorylated-JAK2 （p-JAK2）， and phosphorylated- STAT3 （p-STAT3） pathway proteins in Raji cells after 24 h of treatment with 0， 8， 16 and 32 μmol·L-1 PSA were tested by Western blot.ResultCompared with control group， decreased cell survival rate， inhibited cell proliferation， activated zymogens of Caspase-3， Caspase-8 and Caspase-9 （P<0.01）， increased apoptosis （P<0.05， P<0.01）， and enhanced cell cycle arrest in Gap phase 2 （G2） were observed in 8， 16 and 32 μmol·L-1 PSA groups（P<0.05， P<0.01）. Compared with control group， cells treated with 8， 16 and 32 μmol·L-1 PSA had lower expression of Bcl-2， p-JAK2， p-STAT3 proteins （P<0.05， P<0.01）， and higher expression of Bax， cleaved PARP and cleaved Caspase-3 protein （P<0.01）， while no significant change was found in the expression of JAK2 and STAT3 proteins.ConclusionPSA could inhibit proliferation and induce apoptosis of Raji cells， and its potential mechanism might be related to the regulation of JAK2/STAT3 signaling pathway.
Keywords：pulsatilla saponin A;Burkitt lymphoma;Raji cells;proliferation;apoptosis;janus kinase 2/signal transducer and activator of transcription 3 （JAK2/STAT3） signaling pathway
Abstract：ObjectiveTo study the effects of Toddalia asiatica alcohol extract on autophagy and apoptosis of non-small cell lung cancer A549 cells， and to explore its possible mechanism.MethodA549 cells were cultured in vitro. Cell counting kit-8 （CCK-8） was used to detect the proliferation of A549 cells， and cell survival rate was calculated to screen the drug concentration. The apoptosis in each dose group and that after the use of 3-methyladenine （3-MA）， an autophagy inhibitor， were detected by flow cytometry combined with Annexin V-FITC/PI double staining. Western blot was used to detect the expression levels of apoptosis-related proteins such as B cell lymphocytoma-2（Bcl-2）， Bcl-2-associated X protein（Bax）， microtubule-associated protein 1 light chain 3 （LC3）， cleaved cysteinyl aspartate-specific protease-3 （cleaved Caspase-3）， activated poly （Adenosine diphosphate） ribonucleotide polymerase （cleaved PARP1）， PARP1， activated death activator （t-Bid）， Bid， and ubiquitin-binding protein p62 in each group and those after the use of 3-MA.ResultCompared with the conditions in the control group， the cell survival rate in 0.25 g·L-1 group （P<0.05）， and 0.5， 1， 2， 4 g·L-1 groups （P<0.01） was decreased after 24 h intervention. Additionally， the cell survival rate was reduced in a concentration-dependent manner at 48 h and it was less than 10% at 4 g·L-1 （P<0.01）. Compared with the conditions in the control group， the total apoptosis rate in 0.5 g·L-1 group was increased （P<0.05）， and the apoptosis rate in 1 and 2 g·L-1 groups was also increased （P<0.01）. Compared with the 2 g·L-1 group and 3-MA group， the 3-MA combined with T. asiatica alcohol extract had significantly decreased apoptosis rate （P<0.01）. Compared with the conditions in the control group， elevated expression of pro-apoptotic proteins cleaved PARP1， Bax and t-Bid in 1 and 2 g·L-1 groups （P<0.05， P<0.01）， and reduced expression of Bid in the 2 g·L-1 group （P<0.01） were found. Compared with the conditions in the control group， the expression of anti-apoptotic protein Bcl-2 （P<0.05， P<0.01） and the level of p62 （P<0.01） were down-regulated in 0.5， 1， 2 g·L-1 groups， while the level of LC3 Ⅱ protein was up-regulated （P<0.01）， with certain concentration dependence.ConclusionT. asiatica alcohol extract could significantly inhibit the proliferation of A549 cells， which might be related to promoting autophagy and inducing apoptosis.
Abstract：ObjectiveTo investigate the role and mechanism of Panax notoginseng saponins （PNS） in inhibiting transforming growth factor-β1 （TGF-β1）-induced renal tubular epithelial cell injury.MethodNRK-52E renal tubular epithelial cells were cultured and divided into control group， TGF-β1 group，TGF-β1+12.5 mg·L-1 PNS group，TGF-β1+25 mg·L-1 PNS group and TGF-β1+50 mg·L-1 PNS group. After 48 hours of PNS intervention， the cells and the supernatant were collected， and cell morphology was observed by inverted microscope. Western blot was used to detect the expression of epithelial-mesenchymal transition （EMT）-related proteins and autophagy-related proteins. Flow liquid chromatography for multiple protein quantification and flow cytometry were employed to determine the content of inflammatory factors and apoptosis rate， respectively.ResultCompared with the conditions in the control group， after TGF-β1 induction， the cells showed a spindle-shaped change and the expression of E-cadherin was down-regulated （P<0.05）， while the expression of α-smooth muscle actin （α-SMA） was up-regulated （P<0.05）. After PNS treatment， most of the cells tended to be normal and reversed the occurrence of EMT. In addition， compared with the conditions in the control group， the level of TNF-α was increased while that of IL-10 was decreased， with elevated apoptosis rate （P<0.05） in the TGF-β1 group. After PNS treatment， the level of TNF-α was lowered while that of IL-10 was boosted with the increase of the dose， with reduced apoptosis rate （P<0.05）. Moreover， after TGF-β1 induction， the expression of autophagy-related proteins Beclin 1 and LC3Ⅱ/Ⅰ in renal tubular epithelial cells were up-regulated， while PNS inhibited their expression（P<0.05，P<0.01）.ConclusionPNS had a protective effect on TGF-β1-induced renal tubular epithelial cells， and the mechanism might be that it reduced inflammation and apoptosis by inhibiting autophagy， thus alleviating TGF-β1-induced injury.
Abstract：Continuous cropping obstacle is the bottleneck of medicinal plant cultivation, which seriously affects the quality and yield of medicinal materials. The research on the mechanism of continuous cropping obstacle has evolved from soil physical and chemical properties and allelopathy in the 1970s to the changes of rhizosphere microenvironment and plant response mechanism at present. According to the available studies in this field and our previous research work, we systematically analyzed the mechanism of rhizosphere exudate-mediated microbial community reconstruction in the soil of the medicinal plants in continuous cropping. Specifically, rhizosphere exudates, providing the carbon source and energy for microbial growth, act as inducers or repellents to induce microbial growth or transfer, thereby changing the physicochemical properties (such as acidity) of rhizosphere soil and further altering the structure of rhizosphere microbial community. Further, we comprehensively discussed the ways of synergism between rhizosphere exudates and soil microorganisms in causing harm to the medicinal plants in continuous cropping. That is, rhizosphere exudates mediate the infection of the rhizosphere by pathogenic microorganisms, increase the susceptibility of the nearby plants, inhibit the defense of the host plants, and protect the pathogens to occupy the dominant niche. The synergistic interaction results in the release of more pathogenic factors such as mycotoxins by rhizosphere pathogens, enhanced toxicity of rhizosphere allelochemicals, and deterioration of soil physical and chemical properties. This paper summarizes the role of interaction between rhizosphere exudates and soil microorganisms in the formation of continuous cropping obstacles, aiming to provide a new research idea for revealing the formation mechanism as well as the theoretical support for overcoming continuous cropping obstacles of medicinal plants.
Abstract：ObjectiveTo compare the effects of different drying methods on volatile components of Pseudostellariae Radix.MethodThe samples were dried by different methods， including air drying， sun drying， hot air drying （40， 60， 80 ℃） and vacuum freeze drying. Gas chromatography-ion mobility spectrometry （GC-IMS） was used to compare the changes of volatile components in the samples after different treatments. The samples were incubated at 80 ℃ and 500 r·min-1 for 15 min， the injection temperature was 85 ℃， the injection volume was 200 μL， the flow rate of carrier gas was from 2 mL to 150 mL during 20 min， and the temperature of IMS detector was 60 ℃. SE-54 capillary column （0.32 mm×30 m， 0.25 μm） was used， the column temperature was 60 ℃， and the analysis time was 35 min. The differential spectra of volatile components were constructed and analyzed by principal component analysis （PCA）.ResultA total of 37 volatile components were identified from dried Pseudostellariae Radix. The number of compounds in descending order was ketones， aldehydes and alcohols. There were some differences in the volatile components in samples dried by different methods. And the volatile components in samples with sun drying， air drying and hot air drying at 40 ℃ were similar， compared with other drying methods， vacuum freeze drying and hot air drying at 80 ℃ had great effects on the volatile components of Pseudostellariae Radix， and the compounds in the samples with vacuum freeze drying were the least.ConclusionIn this study， GC-IMS for the detection and analysis of volatile components in Pseudostellariae Radix is established， which has the characteristics of high efficiency， nondestructive inspection and simple sample processing. This method can be used for the distinction of Pseudostellariae Radix dried by different methods. And hot air drying at 40 ℃ can effectively retain the volatile components of Pseudostellariae Radix， and achieve similar flavor to samples with sun drying and air drying.
Abstract：Objective ： To study the effect of temperature and light intensity on photosynthetic fluorescence parameters, volatile oil content, and growth of Atractylodes lancea and provide reference for the rational selection of cultivation environment for A. lancea.MethodWe determined the photosynthetic indexes (such as net photosynthetic rate, water use efficiency, and carboxylation rate), light response curve, CO2 response curve, fluorescence parameters, and the content of four volatile oils in A. lancea under two temperature treatments (32 °C and 22 °C) and two light treatments (full light and shade).ResultThe net photosynthetic rate and water use efficiency of A. lancea under high temperature + strong light were significantly higher than those under high temperature + weak light and low temperature + strong light. The ability of A. lancea to use weak light at low temperature was the strongest, while the utilization rate of weak light under strong light significantly reduced. The photosynthetic rate of A. lancea at low temperature was more susceptible to light intensity and CO2 concentration than that at high temperature. The maximum photosynthetic rate and apparent quantum efficiency under weak light were significantly higher than those under strong light. The photoreaction efficiency at high temperature was higher than that at low temperature. The total amount of volatile oil in A. lancea treated with high temperature + weak light was the highest, reaching 4.582%. Compared with high temperature + strong light, high temperature + weak light significantly increased the content of hinesol and β-eudesmol in A. lancea by 91.7% and 35.7%, respectively, and low temperature + strong light significantly increased the content of hinesol by 87.5%. The content of β-eudesmol in low temperature + weak light treatment was significantly lower than that in high temperature + weak light treatment.ConclusionTThe growth of A. lancea was affected by the interaction between temperature and light. The light and temperature conditions required for the accumulation of volatile oil were not consistent with those suitable for the growth and development of A. lancea. A. lancea responded to the changes of light and temperature conditions by regulating the synthesis and accumulation of volatile oil.
Abstract：ObjectiveTo analyze the flavor substances and change rules of Rhei Radix et Rhizoma during the process of nine-time repeating steaming and sun-drying.MethodThe flavor response values of Rhei Radix et Rhizoma samples were obtained by using PEN3 electronic nose system. The data were processed and analyzed by principal component analysis （PCA）， linear discriminant analysis （LDA） and Loadings analysis.ResultRhei Radix et Rhizoma processed with nine-time repeating steaming and sun-drying could be effectively distinguished into two categories as the sixth sample was the turning point. The samples steamed and dried for one to five times could be grouped into one category， the other four samples were obviously distinguished from them. The main flavor components reached the maximum response in the sample processed with six-time repeating steaming and sun-drying， and its response value of inorganic sulfur compounds was about 2.7 times that of the sample processed with one-time repeating steaming and sun-drying. In addition， compared with the raw products， the flavors of Rhei Radix et Rhizoma processed with nine-time repeating steaming and sun-drying and wine stewing changed significantly， and the response value of inorganic sulfur compounds in sample processed with nine-time repeating steaming and sun-drying was about 2.2 times that of raw products. From the perspective of flavor analysis， the response values of inorganic sulfur compounds and nitrogen-oxygen compounds in sample processed with nine-time repeating steaming and sun-drying were higher than those of wine-stewed products， and the two were not completely equivalent.ConclusionElectronic nose technology preliminarily clarifies the dynamic change rules of the flavor of Rhei Radix et Rhizoma during the process of nine-time repeating steaming and sun-drying from the flavor characteristics， and clarifies the difference between products processed with nine-time repeating steaming and sun-drying and wine-stewed products from the odor characteristics， which lays a foundation for revealing the processing principle of Rhei Radix et Rhizoma processed with nine-time repeating steaming and sun-drying.
Keywords：electronic nose;Rhei Radix et Rhizoma;nine-time repeating steaming and sun-drying;intelligent sensory technology;Chinese medicine processing;principal component analysis （PCA）;linear discriminant analysis （LDA）
Abstract：ObjectiveTo analyze the production of Poria in 5 major producing areas and the problems through field investigation and literature review and thus to lay a foundation for healthy development of Poria industry.MethodEach production process of Poria in Anhui， Hubei， Hunan， Yunnan， and Sichuan was probed and related research was retrieved for the analysis.ResultSichuan， Guangxi， Guizhou， and other areas have been the emerging producers of Poria. However， a few Poria cocos lines were used and the variety breeding lagged behind. Different cultivation techniques were adopted in different production areas. For example， the "induction" method failed to be widely used. Moreover， the harvest time was mainly dependent on the market demand not matter it was suitable or not. Furthermore， steaming has replaced the traditional diaphoretic processing in the processing of this medicinal material in production areas. In addition， fumigation with sulphur was still used in the processing of Poria.ConclusionAn excellent variety is the key to the quality of Poria. Efforts should be made to strengthen the evaluation of the germplasm resources of P. cocos and variety breeding and standardize the processing in production areas， thereby ensuring the safety and effectiveness of Poria and the decoction pieces. In addition， the contradiction between artificial cultivation and woodland ecology should be fully coordinated to ensure the sustainable development of Poria industry in China.
Keywords：Poria;resources;strains;cultivation;processing in production area;analysis of the status quo
Abstract：ObjectiveTo identify Dendrobium flexicaule and its related species， and analyze the differences in polysaccharide composition and D-mannose content， so as to provide theoretical basis for the accurate identification and quality control of Dendrobium medicinal materials.MethodNine samples of Dendrobium （S1-S9） were identified by DNA barcoding and infrared spectroscopy， and the contents of polysaccharides and D-mannose were determined by ultraviolet spectrophotometry （UV） and high performance liquid chromatography （HPLC）， respectively. UV detection condition was 488 nm， HPLC detection conditions were the mobile phase of 20 mmol·L-1 ammonium acetate solution-acetonitrile （81.5∶18.5） and the detection wavelength at 250 nm.ResultDNA barcoding results showed that samples S1-S3 were D. nobile， samples S4-S5 were D. officinale， sample S6 was D. huoshanense， and S7-S9 were D. flexicaule. One-dimensional infrared spectroscopy showed that only D. nobile had stable characteristics at the wavenumber of 1 570-1 467 cm-1， showing a "W" shape， while no absorption peak was found at the wavenumber of 842-740 cm-1， but the other Dendrobium samples had stable absorption peaks at the wavenumber of 842-740 cm-1. In the first derivative spectrum， at the wavenumber of 785 cm-1， D. huoshanense presented a "V" shape， while the rest of Dendrobium presented a "W" shape. At the wavenumber of 1 110 cm-1， D. flexicaule had a stable characteristic peak. In the second derivative spectrum， at the wavenumber of 1 125 cm-1， D. officinale presented an "M" shape， and the rest of Dendrobium was approximately "W" shape. The results of determination showed that the contents of polysaccharides in samples S1-S9 were 9.35%， 9.12%， 32.78%， 49.38%， 48.97%， 32.48%， 32.95%， 39.41% and 25.32%， and their contents of D-mannose were 1.39%， 0.47%， 13.57%， 3.04%， 33.85%， 23.57%， 16.64%， 17.47% and 19.49%， respectively. Among them， D. flexicaule had high polysaccharide and D-mannose contents.ConclusionBoth DNA barcoding and infrared spectroscopy can be used to identify D. flexicaule and its related species， and infrared spectroscopy is cost-effective and easy to operate. At the same time， D. flexicaule has high contents of polysaccharides and D-mannose， which can provide a scientific basis for rapid identification of D. flexicaule and its relatives， and provides a reference for its quality control， and resource development and utilization.
Abstract：ObjectiveTo evaluate the methodological quality of traditional Chinese medicine （TCM） diagnosis and treatment guidelines/consensus of constipation with Appraisal of Guidelines for Research and Evaluation Ⅱ （AGREE Ⅱ）tool， and to study the attention situation of the included Chinese patent medicines in China's National Reimbursement Drug List in the guidelines/consensus.MethodThe data of CNKI，VIP，Wanfang Data，SinoMed，PubMed and Cochrane from the inception of the databases to October 2021 were searched to collect the TCM diagnosis and treatment guidelines/consensus of constipation. Then，the diagnosis and treatment standards and recommended Chinese patent medicines were extracted. Two researchers assessed the methodological quality of the guidelines/consensus with AGREE Ⅱ tool independently. The quality of reports was evaluated by Reporting Items for practice Guidelines in HealThcare （RIGHT） Statement. The recommended Chinese patent medicines in the guidelines/consensus were compared with those in the National Reimbursement Drug List.ResultEleven consensus and 2 guidelines were included，involving 794 experts. The scores of AGREE II were clarity of presentation（59.0%），scope and purpose（44.0%），stakeholder involvement（23.1%），rigor of development （12.1%），applicability （11.1%），and editorial independence （8.3%） from high to low. Five articles were recommended at B level（recommended after revision） and 8 articles were at C level （not recommended）. The average scores of RIGHT Statement were as follows：basic information （93.59%），background （57.69%），evidence （18.46%），recommendations （20.88%），review and quality assurance （19.23%），funding，declaration and management of interests （0.00%）， and other information （0.00%）. The included guidelines/consensus recommended a total of 27 Chinese patent medicines，among which 20 were included in the National Reimbursement Drug List，with 4 species of Class A and 16 species of Class B， accounting for 74.1% of all recommended Chinese patent medicines. Ten purgative Chinese patent medicines in the National Reimbursement Drug List were recommended by the guidelines/consensus，accounting for 50% of all purgative drugs， and 8 were not recommended. There were prescriptions for purgation， for promoting digestion and removing food stagnation， for clearing heat and purging fire，and for warming the middle and dissipating cold，Tibetan medicine and Mongolian medicine.ConclusionBy the AGREE Ⅱ assessment，the methodological quality of the TCM diagnosis and treatment guidelines/consensus of constipation included in this study needed to be improved in the future. The report quality evaluated with RIGHT Statement was low. Most drugs included in the National Reimbursement Drug List were paid attention in the TCM diagnosis and treatment guidelines/consensus of constipation. Moreover，the drugs included in the National Reimbursement Drug List could basically fulfill the clinical needs reflexed from the guidelines/consensus recommendations. However， the reasons of some drugs failing to be included in the National Reimbursement Drug List needed to be studied in the future.
Keywords：traditional Chinese medicine guidelines;constipation;Appraisal of Guidelines for Research and Evaluation Ⅱ （AGREE Ⅱ）;Reporting Items for practice Guidelines in HealThcare （RIGHT） Statement;National Reimbursement Drug List
Abstract：This study systematically and comprehensively sorted out the application status of Chinese patent medicines for the treatment of constipation in the clinic， to provide scientific evidence for future research directions and clinical drug use in this field. Based on the method of scoping review， the Chinese patent medicines for the treatment of constipation were retrieved from three drug lists to obtain the medicines that needed to be evaluated. A comprehensive and systematic search was carried out on the included studies on the treatment of constipation by Chinese patent medicines through eight Chinese and English databases， and the included studies were integrated and analyzed. The results were displayed in combination with charts. Thirty-four Chinese patent medicines and 118 studies were included in this study. According to the efficacies， Chinese patent medicines were divided into 4 categories， namely eliminating accumulation， purging fire， promoting Qi， and moistening bowels， involving 125 Chinese medicines. The overall attention of constipation research is on the rise. Marenwan （granules or soft capsules） gained the highest attention， with 42 studies， followed by Qirong Runchang oral liquid， with 21 studies， and Biantong tablets （capsules）， with 19 studies. There are 10 studies on Congrong Tongbian oral liquid. Seventeen Chinese patent medicines had no corresponding clinical research. There were 8 study design types， and all drugs were mainly randomized controlled trials except Danggui Longhui pill. Among the intervention types， the comparison between Chinese patent medicines and western medicines was the most. The highest outcome indicators were efficacy， safety evaluation， and main symptoms or scores， and there was a lack of a unified core outcome indicator set. There were few studies on adverse reactions and the economy. Only 11.86% of the studies were funded. Clinically， Chinese patent medicines are widely used in the treatment of constipation， and the amount of related research shows an increasing trend. However， some Chinese patent medicines lack corresponding clinical evidence， and the published research has problems such as unrigorous design， ununified criteria for efficacy evaluation， lack of comprehensive evidence studies， and insufficient funds. It is hoped that more investment will be made in this field in the future， and more attention will be paid to drugs with relatively blank research and constipation syndromes with few treatments. Comprehensive evidence studies such as systematic reviews should be carried out actively. And the study design should be standardized to provide reliable evidence for the treatment of constipation with Chinese patent medicines.
Keywords：constipation;Chinese patent medicines;scoping review;randomized controlled trials;clinical research;rational use of drugs
Abstract：The method of scoping review was used to systematically search and sort out the clinical research of oral Chinese patent medicines for ischemic stroke，to understand the scope of relevant research and the distribution of evidence. Three medical catalogs were manually searched to obtain the oral Chinese patent medicines used for ischemic stroke，and 7 databases were retrieved to obtain the clinical research including these oral Chinese patent medicines. Then the clinical evidence results were visualized by description combined with chart analysis. A total of 68 oral Chinese patent medicines were retrieved，and 1 392 articles were included，with 367 published in core journals， involving 35 oral Chinese patent medicines. The research types included randomized controlled trials，cohort studies，case series，case reports，secondary studies，adverse drug reaction reports，pharmacoeconomic evaluations，drug interactions，consensus or guidelines，non-randomized intervention studies and cross-sectional studies，of which randomized controlled trials had the largest number （283， 77.1%），followed by secondary studies and case series （25， 6.7% for each）. Among the 283 randomized controlled trials，there were 159 clinical studies in the acute phase of ischemic stroke，65 in the non-acute phase，and 59 in the unclear phase. Ten intervention control types and 20 outcome index types were summarized. Among them， the composite outcome index and surrogate outcome index were used 217 times （76.7%） and 245 times （86.6%）， respectively，followed by the degree of neurological impairment （three scales）. Future clinical research of oral Chinese patent medicines for ischemic stroke should clarify the stage of the disease，and the research design should specify the advantages of oral Chinese patent medicines intervening in ischemic stroke. Furthermore， publicly-recognized positive controls should be employed，and important clinical outcome indexes should be selected.
Keywords：oral Chinese patent medicine;ischemic stroke;cerebral infarction;scoping review;evidence mapping
Abstract：ObjectiveTo evaluate the safety， efficacy， and economy of the four Chinese patent medicines （CPMs）， including Simotang oral liquid， Liuwei Anxiao capsule， Baohe pill， and Jianwei Xiaoshi oral liquid in the treatment of functional dyspepsia （FD） by a rapid health technology assessment （RHTA）， thus providing evidence support for clinical decision making.MethodChina National Knowledge Infrastructure （CNKI）， WanFang Database， VIP Chinese Technology Periodical Database （VIP）， China Biology Medicine disc （CBMdisc）， PubMed， EMBASE， Cochrane Library， and Web of Science were searched by computer from inception to March 2022. After literature screening， data extraction， and quality evaluation， the descriptive analysis of the results combined with visual charts was performed. Nineteen studies were included， involving 18 randomized controlled trials （RCTs） and 1 Meta-analysis. Neither economic studies nor health technology assessment （HTA） reports were retrieved.ResultThe four CPMs were safe and effective in the treatment of FD， but economic research was lacking. Among them， Simotang oral liquid could be used for children with FD and FD caused by qi and food stagnation， liver and spleen disharmony， and liver and spleen stagnation. Liuwei Anxiao capsule could be used for adult patients with FD caused by food stagnation. Baohe pill could be used for the elderly with FD. Jianwei Xiaoshi oral liquid could be used for children with FD caused by spleen and stomach weakness.ConclusionThe four CPMs have their advantages in the treatment of FD. Among them， the clinical universality of Simotang oral liquid is higher. However， the quality of clinical evidence is generally low， and comparative analysis among drug dosage forms is lacking. In the future， it is necessary to improve， apply， and promote RHTA for rapid evidence production while carrying out a more standardized and scientific evidence-based demonstration of the comprehensive clinical efficacy of CPMs.
Abstract：This paper aims to systematically retrieve and summarize the clinical evidence on oral Chinese patent medicine in otorhinolaryngology by scoping review and analyze the distribution of the evidence， which is expected to serve as a reference for clinical practice and healthcare decision-making. Seven databases were searched （from inception to March 2022） for the clinical evidence of oral Chinese patent medicine in the prevention and treatment of otorhinolaryngologic diseases， and the distribution of the evidence was discussed. A total of 248 papers from core journals/SCI were included： 238 clinical studies （185 randomized controlled trials， 46 semi-/non-randomized controlled trials， 7 case series studies）， 5 systematic reviews， 4 guidelines/expert consensuses， and 1 pharmacoeconomic study. The papers covered 26 oral Chinese patent medicines and 40 otorhinolaryngological diseases （5 ear diseases， 22 nose diseases， and 13 throat diseases）. The majority of the clinical studies included 100-300 cases. The combination of Chinese patent medicine and western medicine is the common intervention in the experimental group. The outcomes were mainly “cure rate” and improvement of clinical symptoms. Common adverse events were nausea， vomiting， rash， headache， gastrointestinal discomfort， fatigue， etc. In summary， there is a lack of high-quality clinical evidence on oral Chinese patent medicine in otorhinolaryngology. In addition， the available studies have such problems as seldom use of recognized outcomes， low quality of clinical studies， and lack of pharmacoeconomic study. In future， efforts should be made to carry out more rigorous primary and secondary research and enhance the pharmacoeconomic evaluation， in a bid to explore the advantages of Chinese patent medicine in the treatment of otorhinolaryngologic diseases and promote the more rational allocation and application of health resources.
Keywords：otorhinolaryngology;oral Chinese patent medicine;clinical evidence;scoping review
Abstract：To understand the development status and provide the basis for the construction and development of health technology assessment （HTA） institutions/organizations in China， this paper systematically reviewed the status of international HTA institutions/organizations and the HTA institutions/organizations in western developed countries and some Asian countries. This study was based on the results of the second round of global survey on HTA conducted by the World Health Organization （WHO） in 2020/2021. The websites of WHO， International HTA Database （INAHTA）， international HTA institutions/organizations， and the HTA institutions/organizations in different countries were searched. After data extraction and classification， we summarized the status of 9 international HTA institutions/organizations （including Decide： Health Decision Hub， Health Technology Assessment International， HTAsiaLink， International Network of Agencies for Health Technology Assessment， European Network for Health Technology Assessment， Health Technology Assessment Network of the Americas， International Society for Pharmacoeconomics and Outcomes Research， Southern African Health Technology Assessment Society， International Information Network on New and Emerging Health Technologies）， 11 HTA institutions/organizations in western developed countries and Asian countries （including Canada， France， Germany， the United Kingdom， Australia， the United States， Japan， Malaysia， South Korea， Singapore， and Thailand）， and 6 HTA organizations/teams in China. The use of HTA varies greatly because of different medical and health systems among different countries， and thus the role of HTA in health decision-making varies among different countries. The international HTA institutions/organizations have greatly strengthened the communication between HTA institutions/organizations and the pharmaceutical industry and facilitated the promotion and practice of evidence-based health decision-making.
Abstract：ObjectiveTo explore difference in the mechanism of Shoutaiwan， a representative kidney-tonifying and abortion-preventing formula， and Juyuanjian， a typical spleen-invigorating and abortion-preventing formula in reversing the pathology of decidua of spontaneous abortion （SA） patients and to expound the connotation of "uterine collaterals connecting kidney" and "fetal collaterals connecting spleen" theory.MethodThe targets of SA were retrieved from GeneCards， followed by gene ontology-biological process （GO-BP） annotation. Based on Cytoscape and previous research， the main processes and core targets were screened out. High-performance liquid chromatography-mass spectrometry （HPLC-MS） was used to identify the potential active components of Shoutaiwan and Juyuanjian and the regulatory networks were constructed. SA was induced in rats and the model rats were treated with Shoutaiwan and Juyuanjian at the same unit. Hematoxylin and eosin （HE） staining， transmission electron microscopy （TEM）， enzyme-linked immunosorbent assay （ELISA）， immunohistochemistry （IHC）， immunofluorescence （IF）， and other methods were employed to verify the mechanisms against miscarriage.ResultThe dysregulation of cell adhesion， inflammatory response， cell death， and angiogenesis was the core pathological process of SA. A total of 13 potential specific active components of Shoutaiwan and 14 active components of Juyuanjian were screened out. The regulatory networks showed that the potential active components of the two prescriptions modulated vascular endothelial growth factor （VEGF）， interleukin （IL）-2， estrogen receptor （ESR）-1， matrix metalloproteinase-9 （MMP-9）， and other targets to regulate the pathological process of SA. The two can significantly improve the pregnancy rate and the integrity rate and blood supply of decidua cells， control the apoptosis morphology and the expression of estrogen （E2）， progesterone （P）， and its receptor， and down-regulate the expression of MMP-2， MMP-9， IL-2， and IL-6 in decidua tissue of SA rats. At the same time， they up-regulated the expression of anti-apoptotic protein B-cell lymphoma-2 （Bcl-2） and IL-4. Shoutaiwan significantly up-regulated the expression of VEGF， and Juyuanjian significantly down-regulated the expression of E-cadherin （E-Cad）.ConclusionBoth Shoutaiwan and Juyuanjian regulate the core pathological process of SA to prevent miscarriage. At the same unit， Shoutaiwan is overall superior to Juyuanjian. Shoutaiwan is better than Juyuanjian in regulating angiogenesis and Juyuanjian is superior to Shoutaiwan in regulating cell adhesion. This conclusion can partly explain the biological basis of "treating the same disease with different methods"， and provide objective data reference for the identification of quality marker （Q-marker） of anti-miscarriage Chinese medicine and further study of formula-syndrome metabolome.
Abstract：ObjectiveTo investigate the effect of Jiawei Bazhen Yimu capsule on serum sex hormones， sexual organs and estrogen signaling pathway in female rats with premature ovarian failure.MethodThe key target proteins of Jiawei Bazhen Yimu capsule in the treatment of premature ovarian failure were screened out by network pharmacology analysis. Female healthy SD rats were selected, and the rat model of premature ovarian failure was established by ovariectomy. Fifty ovariectomized rats were randomly divided into a model group, an estradiol (E2) valerate group, and Jiawei Bazhen Yimu capsule low, medium, and high-dose groups. Another 10 healthy female rats were set as a sham operation group. The sham operation group and the model group were given distilled water by gavage, and other administration groups were given corresponding doses of drugs by gavage. After 21 d, the serum hormone levels of female rats were measured, including E2, progesterone (P), follicle stimulating hormone (FSH), and luteinizing hormone (LH). Immunofluorescence staining (IF) was used to detect the protein expression levels of estrogen receptor 1 (ESR1), estrogen metabolism P4503A4 enzyme (CYP3A4), and P45019A1 enzyme (CYP19A1) in the uterine tissues of female rats.ResultAs compared with the model group, the serum E2 and P levels of female rats in the Jiawei Bazhen Yimu capsule low, medium, and high-dose groups were significantly increased. Jiawei Bazhen Yimu capsule improved the endometrial status of female rats and increased positive expression of ESR1, CYP3A4, and CYP19A1 in the uterine tissues of female rats (P<0.05).ConclusionThe mechanism of Jiawei Bazhenyimu capsule in the treatment of premature ovarian failure may be related to its hormone-like effect and activation of the estrogen signaling pathway.
Abstract：China ranks the first in the morbidity and mortality of malignant tumors in the world. Traditional Chinese medicine （TCM） has played a crucial part in preventing recurrence of malignant tumors after surgery， enhancing efficacy and reducing toxicity of radiotherapy and chemotherapy， and promoting the survival of patients with advanced tumors. According to the whole process of tumor occurrence and development and via the long-term clinical practice， we proposed the "deficiency-cold-toxin-blocking-failure evolution theory" of malignant tumors， with depression in the whole process and cancer toxin as the core. The theory is a summary of pathophysiological status at different stages of tumor development based on the disease and syndrome. This paper analyzes the biological basis of the theory. To be specific， we believe that the ''deficiency'' is mainly caused by the tumor immunoediting abnormality and thus the escape of clearance. In addition， the ''cold'' is in line with the metabolic reprogramming of the tumor microenvironment. ''Toxin'' is manifested as the stemness of cancer cells induced by the tumor microenvironment. ''Blocking'' is closely related to the enhanced permeability and retention （EPR） effect of tumor neovasculature. ''Failure'' is exactly the cachexia caused by change of dominant metabolism. ''Depression'' is essentially stress-the accelerated progress of the neuroendocrine system. In combination with clinical practice， we conclude that method of reinforcing healthy Qi and adjusting the balance can help recover tumor immunoediting and restore the immunity to regulate the "deficiency". Moreover， replenishing Qi and warming Yang can correct the tumor metabolic programming to avoid "cold". Removing toxin and reinforcing healthy Qi can reverse tumor cell stemness and thus suppress the ''toxin''. Eliminating the ''blocking'' source can suppress tumor angiogenesis and help regulate the EPR. Regulating Yin and Yang will help relieve the change of dominant metabolism and futile cycle to avoid ''failure''. The method of relieving depression and regulating Qi can relieve neuroendocrine stress and prevent tumor progression， thus alleviating the "depression". Moreover， effect of these methods was tested with effective formulas. This study is expected to lay a foundation for the establishment of a new system for the clinical prevention and treatment of malignant tumors.
Keywords：malignant tumor;five-phase evolution;biological basis;traditional Chinese medicine
Abstract：Anemia is one of the most common complications of rheumatoid arthritis （RA）. It is an important factor affecting the quality of life of patients and disease prognosis in addition to joint dysfunction. The pathogenesis of RA with anemia in western medicine is complex and diverse， and it is difficult to give consideration to medication. Traditional Chinese medicine （TCM） has certain therapeutic advantages because it pays attention to overall regulation and syndrome differentiation. In recent years， with the in-depth exploration of the pathogenesis of RA in modern medicine and the continuous improvement of the idea of diagnosis and treatment combined with disease and syndrome in TCM， the connotation of TCM pathogenesis of RA has gradually broken through the traditional theoretical framework of "arthralgia syndrome"， and TCM etiology research of toxin in RA and its complications has received increasing attention. By tracing and sorting out the theory of "toxin caused arthralgia"， combing with the research results of the pathological mechanism of modern medicine， this thesis expounds the importance of toxins in the pathogenic factors of RA in TCM. On this basis， it is further proposed that "deficiency caused by toxic heat" is the core pathogenesis of RA with anemia. Deficiency of healthy Qi， the six exogenous factors， the seven emotional factors， and improper diet can lead to the formation of toxic heat， and damage to the joint structure and viscera， resulting in blood deficiency. To treat RA with anemia， the syndrome differentiation by stages is conducted based on the principle of detoxification and dredging collaterals， and the mechanism of self-made prescription Ersi decoction on RA complicated with anemia is explored based on previous research， to enrich the theoretical basis for the treatment of RA from toxic heat and provide new ideas for TCM differentiation and treatment of RA with anemia and other complications.
Keywords：rheumatoid arthritis;anemia;traditional Chinese medicine therapy;etiology and pathogenesis;theoretical discussion
Abstract：Endangered animal medicine is an important part of traditional Chinese medicine， which is distinctive in the treatment of diseases. At present， the rare and endangered medicinal materials such as tiger bone， rhinoceros horn， pangolin， antelope horn， bear bile are listed as national key protected animals， so their clinical application is limited， the current solution is mainly based on the ideas and methods of similar pharmacological effects， close genetic relationships， artificial breeding， and artificial synthesis to find and develop alternatives for endangered animal medicinal materials. Although artificially cultured bear bile and musk， and artificially synthesized tiger bone， bezoar and musk can solve the shortage of endangered animal medicines to a certain extent， there are still some problems such as difficult breakthroughs in breeding technology and incomplete recognition in the substitute industry. According to this， based on summarizing the existing substitutes for endangered animal medicines， our group proposed the concept of homology， homogeneity and equivalent of substitutes， and constructed a new idea to develop and evaluate substitutes by combining frontier biotechnology with multi-omics detection， so as to provide some support for protecting rare and endangered animals and solving the shortage of endangered animal medicines.
Abstract：Stroke is a common cerebrovascular disease， characterized by high incidence， mortality and disability rate. Neuronal cells， the basic unit of the central nervous system， can be injured to varying degrees when stroke occurs. Neuronal cell injury after stroke is also the key cause leading to neurological dysfunction， affecting the prognosis and quality of life of patients. Therefore， reducing the neuronal cell injury and delaying the process of cell death are effective to decrease the nerve function injury in stroke patients and improve their prognosis， thus lowering the death and disability rate of stroke. Ferroptosis is a new form of cell death that has been widely concerned in recent years. Several studies have confirmed that there is ferroptosis in neuronal cells after stroke. Since ferroptosis is an adjustable form， its intervention can help regulate the injury and death of neuronal cells. Studies have shown that inhibiting ferroptosis plays a role in protecting neuronal cells. Traditional Chinese medicine （TCM）， with the multi-channel and multi-target treatment advantages， has been widely used in the whole stroke and has achieved good clinical efficacy. It might be a new direction taking TCM regulation of ferroptosis as the entry point for stroke treatment in the future. This review revealed the mechanism of ferroptosis， discussed the research status of TCM in intervening in neuronal cell ferroptosis， and provided reference for further improving the efficacy of TCM in stroke.
Keywords：ferroptosis;stroke;traditional Chinese medicine;intracerebral hemorrhage;ischemic stroke
Abstract：Osteoporosis is a chronic skeletal disease characterized by low bone mass， destruction of bone tissue microarchitecture， and imbalance of bone homeostasis， leading to increased bone fragility and increased risk of fractures. Oxidative stress caused by the disruption of the balance between excess reactive oxygen species （ROS） and the anti-oxidative system is an important factor in the occurrence and progression of osteoporosis. Nuclear factor E2-related factor 2 （Nrf2）/heme oxygenase-1 （HO-1） is an important anti-oxidative stress pathway. Nrf2 is a primary factor in regulating cellular oxidative stress. Activating Nrf2 can stimulate the expression of HO-1. HO-1 is a key enzyme whose metabolites are bile green Oxygen， carbon monoxide， and free iron. The metabolites can scavenge ROS， thereby exerting an antioxidant effect in cells. At present， domestic and foreign scholars have reported that the Nrf2/HO-1 signaling pathway is closely related to the occurrence and development of osteoporosis and the mechanism of drugs. Chinese medicine can effectively solve the insufficiency of western medicine with multi-target， multi-channel， and multi-level advantages. Chinese medicine can resist oxidative stress， inflammatory response， and apoptosis by regulating the Nrf2/HO-1 signaling pathway， thus treating osteoporosis. This article reviewed the relationship between Nrf2/HO-1 signaling pathway and its key target protein factors and osteoporosis， to clarify the important role of the Nrf2/HO-1 signaling pathway in osteoporosis. At the same time， a systematic summary of Chinese medicines targeting and regulating the Nrf2/HO-1 signaling pathway for the treatment of osteoporosis was conducted， to provide a theoretical basis for further precise treatment of osteoporosis.
Abstract：Osteoarthritis （OA） is also known as degenerative arthritis with prominent clinical manifestations of joint pain， joint swelling， and loss of mobility. With the aggravation of population aging in modern society， its incidence is increasing year by year worldwide， and it has become one of the main diseases that reduce the quality of life of middle-aged and elderly people and even lead to disability. Modern medical research on OA has shown that tumor necrosis factor-α （TNF-α） produced by activated macrophages and/or synoviocytes is the initiator of OA inflammatory response and can interact with multiple signal transduction pathways. It is considered to be one of the most important cytokines and inflammatory mediators that promote the degradation of cartilage matrix and the destruction of articular cartilage in the course of OA， and plays a key role in the course of OA. Under the guidance of the basic theories of traditional Chinese medicine （TCM）， Chinese medicine provides individualized treatment of OA following the principle of people-oriented， syndrome differentiation and treatment， and has the significant advantages of multiple targets， high efficacy， and low toxicity. In recent years， it has become a research hotspot to explore the specific mechanism of Chinese medicine and its active ingredients in the treatment of OA. This study described the role of TNF-α and its signal transduction pathways in the course of OA and reviewed the modern medical research advances in Chinese medicine acting on TNF-α and its signal transduction pathways in the treatment of OA， providing a certain reference basis for the clinical treatment of OA， basic research， and development of targeted drugs.
Abstract：Gastric cancer （GC）， originating from gastric mucosal epithelium， threatens the life and health of patients. The morbidity and mortality are high in developing countries including China. Despite the major headway in medical technology， methods such as surgery， chemotherapy， and targeted therapy fail to curb the progression. Thus， it is particularly important to clarify the etiopathogenesis and molecular mechanism of this disease and develop effective therapy. The continuous progression of GC is inseparable from the changes in the energy metabolism of tumor cells. Aerobic glycolysis （AEG）， as a unique metabolic method of tumors， directly or indirectly results in various malignant phenotypes of GC tissues. The tumor microenvironment promotes the AEG， as its disordered signaling molecules activate a large number of signaling pathways， key proteins， glycolysis-related enzymes， and various genes that initiate AEG and regulate its activity and ultimately improve the AEG level. In recent years， major progress has been made in research on the intervention of AEG in GC cells with Chinese medicinals， components of Chinese medicinals， and compound Chinese medicine prescriptions. Chinese medicine has shown multi-target and multi-pathway characteristics in the anti-GC process， thus attracting the interest of scholars in China and abroad. This study reviews the intervention of Chinese medicine in AEG of GC from the aspects of genes， proteins， key enzymes of glycolysis， and signaling pathways， in order to further clarify the exact role of AEG in the development of GC and the specific relationship of Chinese medicine with AEG and GC. In addition， the limitations of available research were summarized. This study is expected to provide a reference for future clinical and experimental research in related fields.
Abstract：Shenling Baizhusan is a traditional Chinese medicine compound prescription formulated on the basis of Si Junzitang （Ginseng Radix et Rhizoma， Atractylodis Macrocephalae Rhizoma， Poria， Glycyrrhizae Radix et Rhizoma）. It has excellent functions of replenishing Qi， invigorating spleen， draining dampness， and checking diarrhea， and is one of the classical prescriptions of ''reinforcing earth to generate metal''. This prescription is primarily used in clinical practice to treat malnutrition in children， chronic diarrhea， gastrointestinal dysfunction， and other disorders. In addition， it has a good effect on gastrointestinal adverse reactions associated with radiotherapy and chemotherapy. With the booming of molecular biology， researchers have revealed the role of Shenling Baizhusan in the treatment of diseases， especially the mechanism of regulating different signaling pathways. We retrieved 26 relevant papers （4 written in English and 22 in Chinese） published in recent 5 years from 6 databases including China National Knowledge Infrastructure （CNKI）， Wanfang Data， VIP， PubMed， Cochrance Library， and Excerpta Medica Database （Embase）. On the basis of these papers， we summarized the mechanisms of Shenling Baizhusan in disease treatment. In the animal model of inflammatory bowel disease， Shenling Baizhusan can protect gastrointestinal mucosa by regulating the activation of nuclear factor kappa-B （NF-κB）， mitogen-activated protein kinase （MAPK）， Janus kinase/signal transducer and activator of transcription （JAK/STAT）， and myosin light chain kinase-myosin light chain （MLCK-MLC） signaling pathways. In the animal model of non-alcoholic fatty liver disease， Shenling Baizhusan regulates the activation of phosphoinositide 3-kinase/protein kinase B/mammalian target of rapamycin （PI3K/Akt/mTOR） signaling pathway and Kelch-like ECH-associating protein 1/NF-E2-related factor 2/advanced glycation end-products （KEAP1/NRF2/AREs） signaling pathway， thus alleviating the lipid metabolism disorder induced by high-fat diet and reducing liver lipid accumulation and inflammatory response. In the animal model of lung cancer with bone metastasis， Shenling Baizhusan regulates the activation of PI3K/Akt/mTOR signaling pathway， thus playing an analgesic role. By summarizing the mechanisms of Shenling Baizhusan in treatment of different disease models from signaling pathways， we aim to provide clues for the in-depth study of this prescription.
Keywords：Shenling Baizhusan;disease models;signaling pathways;crosstalk of pathways
Abstract：Esophageal cancer ranks the seventh and sixth in morbidity and mortality among the malignant tumors， respectively. In traditional Chinese medicine， toxic medicinals are commonly used to enhance the efficacy on esophageal cancer. In recent years， as natural drugs have become the focus of research on anti-tumor drugs， toxic Chinese medicinals have received wide attention. It has been found that a variety of toxic Chinese medicinals have significant anti-esophageal cancer effect. In this study， articles on the treatment of esophageal cancer were retrieved from SinoMed， China National Knowledge Infrastructure （CNKI）， Wanfang Data， and VIP， and the toxic Chinese medicinals in the articles were summarized. It was found that the toxic Paridis Rhizoma， Gekko， Cremastrae Pseudobulbus Pleiones Pseudobulbus， Scolopendra， Hirudo， Sophorae Tonkinesis Radix et Rhizoma， Scorpio， and Bufonis Corium were mainly used for the treatment of this cancer. They can be classified into the heat-clearing and toxin-removing medicinals， toxin-counteracting medicinals， phlegm-resolving medicinals， and blood-activating and stasis-resolving medicinals. Most of them were pungent （19，52.78%） or bitter （17，47.22%）. The majority had the meridian tropism toward liver （25， 69.44%）， spleen （13， 36.11%）， and lung （12， 33.33%）. According to the research on the above commonly used toxic Chinese medicinals， most of them have anti-tumor effect and some have been reported to have anti-esophageal cancer effect. The mechanism is mainly the inhibition of proliferation. To be specific， they exert the anti-cancer effect by suppressing the proliferation， migration， and differentiation of cancer cells， inducing cell cycle arrest， and activating B-cell lymphoma/leukemia-2 （Bcl-2）-associated X protein （Bax）/Bcl-2/Caspase signaling pathway to induce apoptosis. In this paper， the commonly used toxic Chinese medicinals for the treatment of esophageal cancer were statistically analyzed， and the mechanisms were summarized， in order to provide a reference for the clinical rational use of toxic Chinese medicinals and the research on the mechanisms for their efficacy.
Keywords：esophageal cancer;toxic Chinese medicinal;mechanism;meridian tropism;advance