Evaluation of porcine urine-derived cells as nuclei donor for somatic cell nuclear transfer

Background@#Somatic cell nuclear transfer (SCNT) is used widely in cloning, stem cell research, and regenerative medicine. The type of donor cells is a key factor affecting the SCNT efficiency. @*Objectives@#This study examined whether urine-derived somatic cells could be used as donors for SCNT in pigs. @*Methods@#The viability of cells isolated from urine was assessed using trypan blue and propidium iodide staining. The H3K9me3/H3K27me3 level of the cells was analyzed by immunofluorescence. The in vitro developmental ability of SCNT embryos was evaluated by the blastocyst rate and the expression levels of the core pluripotency factor. Blastocyst cell apoptosis was examined using a terminal deoxynucleotidyl transferase dUTP nick end-labeling assay. The in vivo developmental ability of SCNT embryos was evaluated after embryo transfer. @*Results@#Most sow urine-derived cells were viable and could be cultured and propagated easily. On the other hand, most of the somatic cells isolated from the boar urine exhibited poor cellular activity. The in vitro development efficiency between the embryos produced by SCNT using porcine embryonic fibroblasts (PEFs) and urine-derived cells were similar.Moreover, The H3K9me3 in SCNT embryos produced from sow urine-derived cells and PEFs at the four-cell stage showed similar intensity. The levels of Oct4, Nanog, and Sox2 expression in blastocysts were similar in the two groups. Furthermore, there is a similar apoptotic level of cloned embryos produced by the two types of cells. Finally, the full-term development ability of the cloned embryos was evaluated, and the cloned fetuses from the urine-derived cells showed absorption. @*Conclusions@#Sow urine-derived cells could be used to produce SCNT embryos.

Difference Analysis of Pharmacokinetic Behavior of Active Ingredients in Wujiwan Between Normal Rats and Rats with Irritable Bowel Syndrome / 中国实验方剂学杂志

Objective:To establish an ultra-performance liquid chromatography-mass spectrometry （UPLC-MS/MS） for determining the plasma concentrations of 10 active ingredients in Wujiwan at different time points after oral administration， and to compare the pharmacokinetic characteristics between normal rats and rats with chronic visceral hypersensitive irritable bowel syndrome （CVH-IBS）. Method:CVH-IBS rat model was prepared by the neonatal rat colon percutaneous transluminal coronary angioplasty （PTCA） balloon stimulation method. After intragastric administration of Wujiwan （0.245 g·kg^-1）， blood was collected from the jugular vein at different time points， and the plasma concentrations of 10 active ingredients （berberine hydrochloride， palmatine hydrochloride， coptisine hydrochloride， jatrorrhizine hydrochloride， epiberberine， dihydroberberine， evodiamine， evodine， paeoniflorin， albiflorin） in Wujiwan was detected simultaneously by UPLC-MS/MS， the pharmacokinetic parameters of each component in normal rats and CVH-IBS rats were calculated. Result:The established UPLC-MS/MS could sensitively and accurately detect the plasma concentrations of 10 active ingredients of Wujiwan in rats. Compared with the normal group， the absorption rates of these 10 active ingredients of Wujiwan in the blood of CVH-IBS rats all decreased to a certain extent， and the peak time （<italic>t</italic>_max） was prolonged. Among them， the <italic>t</italic>_max of berberine hydrochloride and jatrorrhizine hydrochloride were significantly prolonged from 54 minute and 39 minute to 90 minute， respectively （<italic>P</italic><0.05， <italic>P</italic><0.01）. Area under the plasma concentration-time curve （AUC_0-<italic>_t</italic>） of each component increased， and evodiamine and paeoniflorin were significantly different （<italic>P</italic><0.05，<italic> P</italic><0.01）. The clearance rates （CL/<italic>F</italic>） of these 10 active ingredients were all decreased， among which berberine hydrochloride， palmatine hydrochloride and evodiamine had significant differences （<italic>P</italic><0.05， <italic>P</italic><0.01）. Conclusion:There are significant differences in the pharmacokinetic behavior of the active ingredients in Wujiwan between normal rats and CVH-IBS rats， which may be related to the destruction of microstructure of intestinal epithelial cells and the change of activity of liver enzymes under the pathological state of IBS.

Therapeutic Efficacies of Wujiwan at Different Compatibilities in Treatment of Irritable Bowel Syndrome / 中国实验方剂学杂志

Objective:To compare the therapeutic efficacies of Wujiwan at two different compatibilities （No.1 and No.2） on irritable bowel syndrome （IBS） based on neuro-endocrine-immune network， and provide a theoretical basis for the treatment based on syndrome differentiation in traditional Chinese medicine （TCM）. Method:The chronic animal model of IBS with visceral hypersensitivity was established by colon irritation via percutaneous transluminal coronary angioplasty （PTCA） in suckling rats. The animals were randomly divided into a control group， a model group， a dicetel group （0.01 g·kg^-1）， low- （0.335 g·kg^-<bold>¹</bold>）， medium- （0.67 g·kg^-<bold>¹</bold>）， and high-dose （1.34 g·kg^-<bold>¹</bold>） No. 1 Wujiwan groups， and low- （0.385 g·kg^-<bold>¹</bold>）， medium- （0.77 g·kg^-<bold>¹</bold>）， and high-dose （1.54 g·kg^-<bold>¹</bold>） No. 2 Wujiwan groups. The thresholds of abdominal elevation and bow back elevation were evaluated to detect the effect of Wujiwan on intestinal sensitivity of IBS. The density of mast cells （MC） in the colonic tissue of model rats was detected by the modified toluidine blue staining method. The concentrations/positive expression of 5-hydroxytryptamine （5-HT）， substance P （SP）， somatostatin （SS）， and vasoactive intestinal peptide （VIP） in the blood/colon tissue were detected by enzyme-linked immunosorbent assay （ELISA） and immunohistochemistry （IHC） assay. Result:There was no significant difference in body weight among different groups. Compared with the control group， the model group exhibited decreased thresholds of abdominal elevation and bow back elevation （<italic>P<</italic>0.01）， increased density of MCs in the colon tissue （<italic>P<</italic>0.05）， up-regulated levels of 5-HT， SP， and SS in the blood and colon tissue （<italic>P<</italic>0.05， <italic>P<</italic>0.01）， and elevated VIP level in the colon tissue （<italic>P</italic><0.05）. Compared with the model group， Wujiwan at different compatibilities could increase the thresholds of abdominal elevation and bow back elevation （<italic>P</italic><0.01）， diminish the count of MC in the colon tissue （<italic>P</italic><0.05）， and reduce the levels of 5-HT， SP， SS， and VIP （<italic>P</italic><0.05）. As demonstrated by the comparison of No. 1 and No. 2 Wujiwan， No. 1 was superior to No. 2 in reducing the concentrations of 5-HT， SP， and SS in the blood， especially in 5-HT （<italic>P</italic><0.01）. No significant difference between No. 1 and No. 2 in reducing 5-HT positive expression in the colon tissue was observed. Compared to the No. 1 Wujiwan， No. 2 significantly reduced SP expression， and the intensity and range of SS expression in the colon tissue in the No. 2 groups were smaller than those in the No. 1 groups （<italic>P</italic><0.05）. Conclusion:Wujiwan at different compatibilities was capable of improving gastrointestinal hormone disorder of IBS to reduce intestinal sensitivity. In terms of systemic effect， No. 1 was superior to No. 2， while in terms of local effect， No. 2 was advantageous. No. 1 Wujiwan was superior to No. 2 in the effect on intestinal dynamics， while No. 2 had an advantageous effect on intestinal sensation over No. 1.

Shenlian extract attenuates TNF-α-induced ECV304 injury by regulating Nrf2/Keap1 signaling pathway / 中国中药杂志

This study aimed to investigate the effect and the possible mechanism of Shenlian( SL) extract on tumor necrosis factor-α( TNF-α)-induced ECV304 injury. After the establishment of TNF-α-induced ECV304 cells injure model,MTT assay was used to detect cell viability and the level of reactive oxygen species( ROS) was measured by flow cytometry. The contents of superoxide dismutase( SOD),malondialdehyde( MDA),nitric oxide( NO),endothelin-1( ET-1) and interleukin-1β( IL-1β) in the supernatant were detected by biochemical method and enzyme linked immunosorbent assay( ELISA). The expression levels of apoptosis-related proteins B-lymphoma-2 gene( Bcl-2),Bcl-2 associated X protein( Bax),caspase-3,caspase-9 and nuclear factor E2 associated factor2( Nrf2)/Kelch like epichlorohydrin associated protein-1( Keap1) signaling pathway related proteins Nrf2,Keap1,quinone oxidoreductase( NQO1) and heme oxygenase 1( HO-1) were detected by Western blot. The results showed that 50 μg·L-1 TNF-α significantly damaged ECV304 cells,induced the impairment of cell viability( P<0. 01),the increase of ROS production,the decrease of SOD activity,and the increase of MDA,NO,ET-1 and IL-1β( P<0. 01),meanwhile,it caused the up-regulation of Keap1,caspase-9 and Bax protein expression,and down-regulation of NQO1 and Bcl-2 protein expression( P<0. 05) compared with the control group.Compared with the model group,SL extract reduced the damage of ECV304 cells induced by TNF-α,improved cell viability,reduced ROS production,increased SOD activity and decreased MDA,NO,ET-1,IL-1β content( P<0. 01 or P<0. 05). In addition,SL extract also down-regulated the protein expression levels of Keap1,caspase-3,caspase-9 and Bax,and increased the protein expressions of Nrf2,NQO1,HO-1 and Bcl-2( P<0. 01 or P<0. 05). The above results indicate that SL extract can provide protective effect on ECV304 cells injury induced by TNF-α,alleviate oxidative stress injury,inflammation and apoptosis,and its mechanism may be related to regulating Nrf2/Keap1 signaling pathway.

Mechanism of Chuanxiong Rhizoma intervention on central sensitization of Panx1-Src-NMDAR-2B signaling pathway in neuropathic pain model rats / 中国中药杂志

Excitatory toxicity(ET) is an important factor of neuropathic pain(NPP) induced by central sensitization(CS), and the association of pannexin-1(Panx1)-Src-N-methyl-D-aspartate receptor subunit 2 B(NMDAR-2 B) is an important new pathway for ET to initiate CS. The present study confirmed whether the central analgesic effect of Chuanxiong Rhizoma extract(CRE) was achieved through the synchronous regulation of the brain and spinal pathways of Panx1-Src-NMDAR-2 B. In this study, dynamic and simulta-neo-us microdialysis of the brain and spinal cord in vivo combined with behavioristics, high performance liquid chromatography(HPLC)-fluorescence detection, microdialysis analysis(ISCUS~(flex)), ultrasensitive multifactorial electrochemiluminescence immunoassay, ELISA, and Western blot was employed to investigate the protein expression of NMDAR-2 B, Src, and Panx1, extracellular excitatory amino acids, cytokines, energy metabolites, and substance P in spinal dorsal horn(SDH) and anterior cingulate cortex(ACC) after CRE intervention with the rat model of spared sciatic nerve injury(SNI) as the experimental tool. Compared with the sham group, the SNI group exhibited diminished mechanical withdrawal threshold(MWT)(P<0.01), increased cold spray scores(P<0.01), glutamate(Glu), D-serine(D-Ser), and glycine(Gly) in extracellular fluids of ACC, and Glu, D-Ser, interleukin-1β(IL-1β), and lactic acid(Lac) in extracellular fluids of SDH(P<0.05), dwindled tumor necrosis factor(TNF-α)(P<0.05), and elevated protein levels of NMDAR-2 B, Src, and Panx1 in ACC(P<0.05). Compared with the SNI model rats, high-and medium-dose CRE(CRE-H/M) could potentiate the analgesic activity as revealed by the MWT test(P<0.05) and CRE-M enabled the decrease in cold spray scores(P<0.05). CRE-H/M could inhibit the levels of Glu, D-Ser and Gly in the extracellular fluids of ACC(P<0.05), and the levels of Glu in the extracellular fluids of SDH(P<0.05) in SNI rats. CRE-M significantly increased the levels of glucose(Gluc), Lac, interferon-gamma(IFN-γ), keratinocyte chemoattractant/human growth-regulated oncogenes(KC/GRO), and IL-4 in extracellular fluids of SDH in SNI rats(P<0.05). CRE-H/M/L could also inhibit the levels of NMDAR-2 B, Src and Panx1 in ACC and SDH in SNI rats(P<0.05). The central analgesic effect of CRE is presumedly related to the inhibited release of excitatory amino acid transmitters(Glu, D-Ser and Gly) in ACC and SDH of SNI rats, decreased protein expression of NMDAR-2 B, Src and Panx1 in the two regions, and the regulation of the Panx1-Src-NMDAR-2 B pathway in the spinal cord and brain. The above findings partially clarified the scientific basis of clinical analgesic effect of Chuanxiong Rhizoma.

Relationship Between Immune Checkpoint and Autoimmune Disease / 中国实验方剂学杂志

The normal immune system has the ability to distinguish between "self" and "non-self". Because of its dynamic balance of "immune activity-immune tolerance", it will produce immune response to the non-self antigen, but with no response or weak response to the self-antigen. However, if the balance was broken, T cell in the abnormal immune activation state will respond continually to the self-antigen, with an abnormal immune response, which caused autoimmune disease. Pathologically, "invalid" immune recognition and immune response become the main causes for autoimmune diseases. Co-stimulatory molecule is an important link between Attach antigen presenting cells（APC） and immune cells (T cell and B cell). Studies have proved that excessive co-stimulation and/or insufficient co-inhibition could cause detect of self-tolerance and induce autoimmunity. Although co-stimulatory and co-inhibitory pathways have a significant impact on all ADS, this paper focuses on their effect on two systemic autoimmune diseases [systemic lupus erythematosus （SLE） and rheumatoid arthritis（RA）] and two organ-specific autoimmune diseases [multiple sclerosis （MS） and type 1 diabetes （T1DM）], in order to discuss the pathogenesis and relationship between co-stimulatory molecules and autoimmune diseases.

Protective Effect of Artemisinin on Mice with Systemic Inflammatory Response Syndrome / 中国实验方剂学杂志

Objective:To study the protective effect and mechanism of artemisinin on systemic inflammatory response syndrome （SIRS）mice using endotoxin （LPS）-induced SIRS mouse model. Method:Male BALB/c mice aged 5-7 weeks were randomly divided into normal group， LPS model group， low， medium and high-dose artemisinin groups （25， 50， 100 mg·kg-1） and ibuprofen group （39 mg·kg-1）. LPS （10 mg·kg-1） was intraperitoneally injected at the 7th day after the prophylaxis. According to the SIRS clinical diagnostic criteria， the respiratory rate， rectal temperature， lung index， spleen index， glycolipid metabolism， brain tissue inflammatory factors， and phosphorylation of lung tissue inflammation-related proteins were measured. Result:Intraperitoneal injection of LPS significantly reduced the respiratory rate of mice （P<0.05）， body temperature decreased significantly （P<0.01）， spleen index increased significantly （P<0.01）， peripheral blood neutrophil percentage increased significantly （P<0.05）， percentage of monocytes decreased significantly （P<0.01）， thrombocyte decreased （P<0.01）， platelet specific ratio decreased （P<0.01）， total cholesterol content in plasma decreased （P<0.01）， plasma glucose content decreased （P<0.01）. The expression of interleukin-1β increased in hippocampus and cortex of brain tissue （P<0.01）， and the secretion of tumor necrosis factor-α increased in hippocampus and cortex of brain tissue （P<0.01）. The expression of phosphorylated protein STAT1 was increased （P<0.01）， and the expression of phosphorylated protein c-Jun was increased （P<0.01）. After the administration of artemisinin， the body temperature and the respiratory rate of mice induced by LPS were significantly increased， the pathological changes of various organs induced by LPS were alleviated， the hypoglycemia induced by LPS was significantly increased （P<0.05）， the levels of inflammatory factors in hippocampus and cortex was significantly reduced， and the expressions of phosphorylated proteins STAT1 and c-Jun in lung tissue were significantly reduced （P<0.05， P<0.01）. Conclusion:Artemisinin has a significantly protective effect on SIRS mice induced by intraperitoneal injection of LPS possibly by reducing the secretion of inflammatory factors TNF-α and IL-1β.

Potential therapies for COVID-19 cardiovascular complications using artemisinin and its derivatives intervene based on its cardiovascular protection / 中国中药杂志

Corona virus disease 2019(COVID-19) has brought untold human sufferings and economic tragedy worldwide. It causes acute myocardial injury and chronic damage of cardiovascular system, which has attracted much attention from researchers. For the immediate strategy for COVID-19, "drug repurposing" is a new opportunity for developing drugs to fight COVID-19. Artemisinin and its derivatives have a wide range of pharmacological activities. Recent studies have shown that artemisinin has clear cardiovascular protective effects. This paper summarizes the research progress on the pathogenesis the pathogenesis of COVID-19 in cardiovascular damage by 2019 novel coronavirus(2019-nCoV) virus from myocardial cell injury directly by 2019-nCoV virus,viral ligands competitively bind to ACE2 and then reduce the protective effect of ACE2 on cardiovascular disease, "cytokine storm" related myocardial damage, arrhythmia and sudden cardiac death induced by the infection and stress, myocardial injury by hypoxemia, heart damage side effects from COVID-19 drugs and summarizing the cardiovascular protective effects of artemisinin and its derivatives have activities of anti-arrhythmia, anti-myocardial ischemia, anti-atherosclerosis and plaque stabilization. Then analyzed the possible multi-pathway intervention effects of artemisinin-based drugs on multiple complications of COVID-19 based on its specific immunomodulatory effects, protective effects of tissue and organ damage and broad-spectrum antiviral effect, to provide clues for the treatment of cardiovascular complications of COVID-19, and give a new basis for the therapy of COVID-19 through "drug repurposing".

Screening combination ratio and exploring mechanism of Momordicae Semen and Epimedii Folium / 中国中药杂志

The aim of this paper was to obtain low toxicity and high efficiency anti-tumor Chinese medicine through screening the combination ratios of Momordicae Semen and Epimedii Folium, and to explore the anti-tumor mechanism of the combination of two drugs by observing their effect on apoptosis-related proteins in cancer cells. Methyl thiazolyl tetrazolium(MTT) assay was used to observe the effect of drug combination on the proliferation of tumor cells from different tissue sources. The effects of the combination of the two drugs on tumor cells were analyzed by Compusyn software. Plate cloning assay was used to observe the effect of combination of these two drugs on the proliferation of A549 cells in vitro. The expression of reactive oxygen species(ROS) and apoptotic proteins p53, Bcl-2 and Bax were compared by using ROS kit and Western blot. Lewis lung cancer model was used to observe the anti-tumor effect of drugs in vivo. The results showed that the anti-tumor effect of their ethanol extract was more significant than that of water extract, and the anti-proliferation effect was strongest when the ratio was 1∶1(P<0.05). Compusyn analysis showed that the combination of the two drugs had synergistic effect. Further studies showed that after combined use, the number of clonogen formation in A549 cells was significantly reduced(P<0.01); ROS production was increased; the expression of apoptosis-related protein p53 was up-regulated, and the ratio of Bcl-2/Bax was decreased. In vivo animal study showed that the tumor inhibition rate was 53.06%(P<0.05) in the high dose group. As compared with the single use of the two drugs, the combination of the two drugs had more significant anti-proliferative effect on tumors, and the optimum ratio was 1∶1. The combination of the two drugs at a ratio of 1∶1 inhibited the proliferation of various tumor cells, and had no significant effect on normal liver cells LO2 when compared with other ratios. Therefore, it can be preliminarily inferred that the combination of the two drugs may have the effect of synergism and detoxification. Further studies showed that the combination of the two drugs can significantly inhibit the proliferation of A549 cells, and its mechanism may be related to the activation of endogenous apoptotic pathway. In vivo experiments also showed that the tumor inhibition rate increased with the increase of drug concentration.

Protective Effect of Phenolic Compound of Osmundae Rhizoma on Systemic Inflammatory Response Syndrome in Mice / 中国实验方剂学杂志

Objective: To investigate the protective effect of phenolic compounds 4-hydroxybenzylideneacetone and (HBAc)3,4-dihydroxybenzylideneacetone (DHBAc) of Osmundae Rhizoma on the systemic inflammatory response syndrome (SIRS) in mice by establishing the mice model of SIRS.Method: BALB/c mice were randomly divided into the normal group,the SIRS model group and the different doses of HBAC and DHBAc group (25,50,100 μg·kg-3).Lipopolysaccharides (LPS) was injected intraperitoneally after 7 days of prophylactic administration.After 5 hours of modeling,the anus temperature,respiratory rate,the number of white blood cell (WBC) and platelets (PLT),WBC classification,glycolipid metabolism,inflammatory factor and signal transducing phosphorylated protein of lung were measured.Result: Intraperitoneal injection with LPS (6 mg·kg-1) in mice can significantly reduce the respiratory rate (PPPPPβ(PPPPPPPPPβ(PConclusion: The SIRS model can be established through intraperitoneal injection of LPS.HBAc and DHBAc have protective effects on endotoxin-induced SIRS in mice,and may exert anti-inflammatory effects through IκB and c-JUN pathways.

Effect of Shenlian Extract on Macrophage Function and Inflammatory Resolution in Lipid Peroxidation Inflammatory Injury Models / 中国实验方剂学杂志

Objective:To study the effect of Shenlian extract (SL extract) on macrophage function and inflammatory resolution in lipid peroxidation inflammatory injury models. Method:The effects of different concentrations of SL extract (2.5, 5.0, 10.0, 20.0 mg·L-1) on the polarization type, foam formation and chemotactic function of macrophages were detected with RAW264.7 cells induced by oxidized low-density lipoprotein(ox-LDL). Western blot was used to detect pro-inflammatory resolution factor arachidonate 5-lipoxygenase(ALOX5) and inducible inducible nitric oxide synthase(iNOS), and phosphorylated p65 (p-p65) and phosphorylated IκK (p-IκK) in nuclear factor(NF)-κB related signaling pathways. Result:Compared with the control group, ox-LDL enhanced the expressions of M1 macrophage markers TNF-α, IL-1β, iNOS (PPPα, IL-1β, and iNOS (PPPPκK was inhibited significantly (PConclusion:In the inflammatory damage model of lipid peroxidation, SL extract can regulate the polarization of macrophage, inhibit the chemotaxis and foaming of ox-LDL, increase the inflammatory resolution molecular expression, and improve the state of lipid peroxidation, which may be related to the inhibition of NF-κB signaling pathway.

Research progress of effect of artemisinin family drugs on T lymphocytes immunomodulation / 中国中药杂志

Artemisinin was isolated from traditional Chinese herb Artemisia annua for treating malaria. A series of derivatives,like dihydroartemisinin,artesunate,artemether,artether,had the same core chemical structure,and sesquiterpene lactone containing peroxide bridge constitute the basic chemical structure. Besides anti-malaria,artemisinin family drugs were found to ameliorate many different diseases,which have attracted wide attention in recent years. Among different diseases,artemisinin family drugs were found to have T lymphocytes immunomodulation effects,including activation,proliferation,differentiation,apoptosis and subsets function. Because T cell immunologic response is the key point of many diseases,and impact the pathogenic process,therapeutic effect and prognosis,the drug studies with it as the target have become hotspots in recent years. Studies of artemisinin family drug on T cell immunomodulation were still at the initial stage and involved in different disease; furthermore,T cell immune process involves complicated molecular mechanism,it is imperative to summarize the advance of current studies for further systematic explanation and exploration of their characteristics and mechanisms. This article will summarize the research progress of artemisinin family drugs for malaria,autoimmune disease,hypersensitivity reaction,tumor,schistosomiasis and AIDS relating to T cell immune modulation,so as to provide basic and professional reference for related research and application.

Research advancement in natural anti-cancer product / 中国中药杂志

Human health has been severely threatened by malignant tumors continuously.Rational and effective drug use provides an effective means for the treatment of malignant tumors,and is expected to become an important way to solve the problem of tumor treatment in the future.In recent years,with the escalation of new cancer theories and the emergence of clinical drug resistance,innovative research and development of anti-cancer drugs has always been a hot spot and focus in cancer research.Among them,the discovery of novel anti-cancer drugs from natural compound is of top priority due to its strong anti-cancer efficacy and the abundant drug resources.Therefore,it is imperative to systematically summarize the cutting-edge advancements of the natural products and their potential pharmacological mechanisms according to the characteristics of tumor progression,and put forward the new directions and trends for further development of anti-cancer natural products in the future.Specifically,the research advancements on anti-cancer effect of natural products were reviewed,focusing on both the traditional and innovative application.We hope this review could bring the light on the research path of the natural anti-cancer products clearly and comprehensively,and also provide inspirations for innovative,safer and more effective anti-cancer drug development and exploration.

Efficacy and mechanism of Lianhua Qingwen Capsules(LHQW) on chemotaxis of macrophages in acute lung injury (ALI) animal model / 中国中药杂志

This paper was mainly to discuss the potential role and mechanism of Lianhua Qingwen Capsules(LHQW) in inhibiting pathological inflammation in the model of acute lung injury caused by bacterial infection. For in vitro study, the mRNA expression of MCP-1 in RAW264.7 cells and THP-1 cells, the content of MCP-1 in cell supernatant, as well as the effect of LHQW on chemotaxis of macrophages were detected. For in vivo study, mice were randomly divided into 7 groups, including normal group, model group(LPS 5 mg·kg~(-1)), LHQW 300, 600 and 1 200 mg·kg~(-1)(low, middle and high dose) groups, dexamethasone 5 mg·kg~(-1) group and penicillin-streptomycin group. Then, the anal temperature was detected two hours later. Dry weight and wet weight of lung tissues in mice were determined; TNF-α and MCP-1 levels in alveolar lavage fluid and MCP-1 in serum were detected. In addition, the infiltration of alveolar macrophages was also observed and the infiltration count of alveolar macrophages was measured by CCK-8 method. HE staining was also used to observe the inflammatory infiltration of lung tissues in mice. Both of the in vitro and in vivo data consistently have confirmed that: by down-regulating the expression of MCP-1, LHWQ could efficiently decrease the chemotaxis of monocytes toward the pulmonary infection foci, thus blocking the disease development in ALI animal model.

Basic research of fibrosis on atherosclerotic plaque stability and related drug application / 中国中药杂志

In the background of the high incidence and high mortality of cardiovascular diseases,atherosclerosis is the main pathological feature of cardiovascular diseases and the core pathological basis for disease progression. In the evolution of atherosclerotic plaques,the rupture of unstable plaques,plaque shedding and formation of thrombosis are the most dangerous parts. In this process,the formation of plaque fibrosis is the core mechanism regulating plaque stability. Additionally,fibrosis reflects dynamic changes in the inflammatory processes and pathological changes. In view of the inflammation regulation and fibrosis regulation,this paper clarified the process of atherosclerotic plaque,explained the roles of relevant inflammatory cells and cytokines in plaque stability,and summed up drug researches related with stable plaque in recent years. In the future,improving the fibrosis will be a new idea for stabilizing plaque in atherosclerosis drug development.

Research initiative of new thought on "main effect" of TCM formulae--new thinking on mechanism of compound action and compatibility mechanism of Chinese herbal compound formulae / 中国中药杂志

This article proposes a new thought on the study of "main effect" of traditional Chinese medicine (TCM) formulae. The blood concentrations of the pharmacodynamic substances of Chinese material medica（CMM）are usually very low, with lower toxic and side effects than western medicine. Therefore, according to a recent hypothesis of additive effect of multiple components for a single target, local targets in multi-component multi-target synergistic effect network of TCM may have the additive effect of similar components. Studies on the disposition of CMM showed that a constituent could bebio-transformed to many metabolites; these compounds with a similar structure are likely to have the same pharmacological effects on the same target, which could provide experimental evidences for the hypothesis of "additive effect". The authors of this article further believe that additive effect of TCM multi-components only comes up under a limited conditions/concentration. Because of the complexity of TCM-organism system, the complex effect of multicomponent addition and competition/antagonism is more likely to appear in single targets of drug effect. This complex effect may be the key to impact the synergistic effect of TCM multi-targets. In theory, choose and create a single target additive effect could realize the scientific compatibility of TCM and improve the curative effect and attenuate toxicity. According to the clinical demand and under the guidance of the above thought, we proposed the "main effect" of TCM formulae. Because traditional Chinese medicine (compounds) have diverse and complex effects, how to better study TCM formulae compatibility mechanism and improve the curative effect? Efforts shall be made to select one or several effects relating to clinical specific syndromes from the complex and diverse effects of TCM as the "main effect". The "main effect" of TCM formulae is the macroscopic manifestation of the synergistic effect of multi-component/multi-target. The study of the Formulae "main effect" can contain at least two aspects: one is the study of pharmacokinetic application of TCM formulae, and another is the study for pharmacodynamics effect. In the study of main effect, there are two main elements. First, which drug targets are directly related to the main effect? This requires identifying the target network. Second, which drug components positively or negatively control the single target of the target network? And what change in single target effect as well as the multi-target synergistic effect will be caused by the regulatory component concentration or the change in number? These two elements is the key to elucidate the mechanism of compound action and compatibility mechanism of Chinese herbal compound formulae. Through the study of the main effect, the clinical curative effect and the mechanism of the TCM formulae shall be improved.

Effect of dihydroartemisinin on permeability of human erythrocyte membrane infected plasmodium / 中国中药杂志

In view of the fact that the antimalarial effects of artemisinins are significant but the mechanism has not yet been clarified and there are many different opinions, it is possible that artemisinins can produce high anti-malarial efficacy through various mechanisms and multiple pathways. In addition, the researches on the pathogenesis of malaria "erythrocyte membrane plasmodial surface anion channel (PSAC)" in the past few years have provided more positive findings, which may confirm and discover the new antimalarial mechanism of artemisinins. This paper was as to study the effect of dihydroartemisinin (DHA) in vitro on erythrocyte membrane permeability of HB3 plasmodium infection, with using the mechanism of 5% sorbitol can be used to kill the Plasmodium falciparum in red blood cell membrane selectively, the effectual difference of sorbitol on the killing of P. falciparum with adding DHA or not was detected, so as to investigate whether DHA can affect the permeability of the erythrocyte membrane. Result showed that, Pre-stimulation with 10 nmol·L⁻¹ DHA (the final concentration of plasmodium in vitro culture system) for 30 min could significantly decrease the killing effect of sorbitol on the HB3 plasmodium in the P. falciparum erythrocytic cycle, and DHA may inhibit the permeability of the erythrocyte membrane for preventing sorbitol through the red blood cell membrane, thereby reducing the killing effect of sorbitol on the P. falciparum.

Protective effect and mechanism of compound Ginkgo biloba granules on oxidative stress injury of HUVEC / 中国中药杂志

To reveal the protective and anti-apoptosis effect of compound Ginkgo biloba granules on oxidative stress injury of human umbilical vein endothelial cells (HUVEC). Negative control group, H2O2 model group and 4 drug pretreatment groups (80, 160, 320, 640 mg• L⁻¹) were established. The cell proliferation, morphological changes in each group after oxidative stress injury was detected by MTT assay and through microscope observation respectively. The content of LDH, MDA, SOD and NO and SOD activity in supernatant were detected to judge the protection effect of the drugs on endothelial cells. The protective effect on HUVEC apoptosis was analyzed by Caspase-3 activity test and Annexin V-FITC/PI staining. Western blot was used to observe the expression of apoptosis-related proteins Bcl-2 and Bax. Results showed that 1 200 μmol• L⁻¹ H2O2 can induce oxidative stress injury in endothelial cells and reduce the cell survival rate; cell proliferation inhibition degree is positively correlated with the effect time of H2O2. Besides, 80, 160, 320 640 mg•L⁻¹ compound Ginkgo biloba granules can protect HUVEC from oxidative stress injury, recover the normal proliferation level of cells, improve their state, prohibit cell apoptosis, and can up-regulate and down-regulate the expression level of Bcl-2 and Bax respectively. In conclusion, compound G. biloba granules can protect HUVEC from the oxidative stress injury induced by H2O2, its mechanism may be correlated with inhibition of the mitochondrial apoptotic pathway in HUVEC.

Establishment of vulnerable plaque model by mast cell activation in adventitia and evaluation of effectiveness of intervention of Shenlian tablet / 药学学报

This study was designed to investigate the activity of Shenlian tablet in stabilization of the atherosclerosis (As) plaque in apoE-/- mice and explore the mechanisms. Rat peritoneal mast cells were randomly allocated and treated with Shenlian tablet (100, 50, 25, 12.5 mg ·L-1) or cromoglicate sodium (200 μg·L-1) for 2 h before exposure to substance P. Histamine, tryptase, IL-1β and NF-κB were measured in the cell culture supernatant by ELISA assay. The plaque formation was induced by common carotid artery cannula method combined with high-fat diet in apoE-/- mice, and the plaque instability was induced by substance P through local mast cell degranulation. Mice were divided into eight groups that included the model 1 (M1, sham-operated group), M2 (carotid artery cannula combined with high-fat diet), M3 (M2 combined with substance P 0.5 μg/mouse), Shenlian extract (95, 190 and 380 mg·kg-1·d-1), atorvastatin (2.6 mg·kg-1·d-1) and normal control group. Total cholesterol (TC), high-density lipoprotein (HDL-C), high-sensitivity C-reactive protein (hs CRP), matrix metalloproteinases 9 (MMP-9) and histamine were measured by ELISA. Thickness, plaque area, mast cell degranulation were observed by hematoxylin and eosin staining, toluidine blue staining. CD117 antigen expression were observed by confocal microscopy. Intracellular phosphorylation was detected using the Bio-Plex 6-plex phosphoprotein assay kit. The results show that the mast cell membrane was stabilized by Shenlian tablet. Histamine, tryptase, interleukin l-β and NF-κB exhibited a significantly reduction in the Shenlian tablet-treated group (PP-/- mice model group. The proliferation, degranulation and inflammation of mast cell were significantly inhibited by Shenlian tablet. On the other hand, the same treatment decreased hs-CRP, MMP-9 and histamine in serum. IκB, p38 MAPK phosphorylation, intraplaque hemorrhage and collagen degradation were reduced in the presence of Shenlian tablet, which increased the stability of the As plaque. The results show that the vulnerable plaque model induced by mast cell activation in adventitia was established. Shenlian tablet exhibited a protective effect in this model. Shenlian tablet may increase the plaque stability via inhibition of mast cell-mediated inflammatory response.

Genetic characterization of bovine viral diarrhea virus strains in Beijing, China and innate immune responses of peripheral blood mononuclear cells in persistently infected dairy cattle

To acquire epidemiological data on the bovine viral diarrhea virus (BVDV) and identify cattle persistently infected (PI) with this virus, 4,327 samples from Holstein dairy cows were screened over a four-year period in Beijing, China. Eighteen BVD viruses were isolated, 12 from PI cattle. Based on genetic analysis of their 5'-untranslated region (5'-UTR), the 18 isolates were assigned to subgenotype BVDV-1m, 1a, 1d, 1q, and 1b. To investigate the innate immune responses in the peripheral-blood mononuclear cells of PI cattle, the expression of Toll-like receptors (TLRs), RIG-I-like receptors, interferon-alpha (IFN-alpha), IFN-beta, myxovirus (influenza virus) resistance 1 (MX1), and interferon stimulatory gene 15 (ISG15) was assessed by qPCR. When compared with healthy cattle, the expression of TLR-7, IFN-alpha, and IFN-beta mRNA was downregulated, but the expression of MX1 and ISG-15 mRNA was upregulated in PI cattle. Immunoblotting analysis revealed that the expression of interferon regulatory factor 3 (IRF-3) and IRF-7 was lower in PI cattle than in healthy cattle. Thus, BVDV-1m and 1a are the predominant subgenotypes in the Beijing region, and the strains are highly divergent. Our findings also suggest that the TLR-7/IRF-7 signaling pathway plays a role in evasion of host restriction by BVDV.