Objective To investigate the influence of the purging fire and removing toxin method on chemokines and adhesion factors related to vascular endothelialitis injury induced by toxin of trimeresurus stejnegeri bite.Methods ① Animal experiment:50 healthy New Zealand white rabbits were chosen.According to random numbers generated by statistical software,they were divided into normal control group,model group,low,middle and high dose Sheshang capsule groups,10 in each group.Trimeresurus stejnegeri bite model was replicated by injecting 0.75 mL/kg snake venom into subcutaneous tissues of rabbits' right hind legs.And the same volume of normal saline was injected into the rabbit in the normal control group.After the model was established for 6 hours,the rabbits in low,middle and high dose Sheshang capsule groups received 174,348 and 522 mg· kg-1 · d-1 of Sheshang capsule solution respectively (the content of capsules was dissolved in normal saline to make liquid with 17.4,34.8 and 52.2 g/L Sheshang solution respectively,so the volume of gavage of each group was 10 mL· kg-1 · d-1);in the model and normal control groups,the same amount of normal saline was given by gavage,once daily for consecutive one week.24 hours after the last gavage,the blood of the rabbits was collected through an auricular border vein and the serum was separated by centrifuge ready for use.Meanwhile,the whole abdominal aorta segment of the rabbit was harvested and kept them in liquid nitrogen ready for use.② Cell experiment:human umbilical vascular endothelial cell (HUVEC) was cultured with MEM for 24 hours.The solution was replaced and according to the random number generated by statistical software,the cells were divided into blank control group,model group and low,middle,high dose Sheshang capsule medicinal serum groups,10 wells in each group.Trimeresurus stejnegeri toxin cell model was reproduced by addition of 5 mg/L snake venom into the cell culture medium.After 6-hour culture,the cells of model group and blank control group received 10％ normal rabbit serum,and the cells of low,middle and high dose Sheshang medicinal serum capsule groups received serum containing 5％,10％ and 15％ drug,respectively.After culture for 72 hours,the cells were collected and the total RNA was extracted.The real-time fluorescent quantitative polymerase chain reaction (qPCR) was used to detect the levels of mRNA of interleukin-8 (IL-8),monocyte chemoattractant protein-1 (MCP-1),intercellular adhesion molecule-1 (ICAM-1) and vascular endothelial cell adhesion molecule-1 (VCAM-1) in the vascular endothelial cells of rabbit aorta abdominalis and human umbilical vein,and the content of serum E-select element (CD62E) was measured by enzyme linked immunosorbent assay (ELISA).Results In model group,the expression levels of mRNA in IL-8,MCP-1,ICAM-1,VCAM-1 and the content of CD62E were all increased significantly in the endothelial cells of rabbit aorta abdominalis and HUVEC compared with those in control group [when the mRNA expression levels of IL-8,MCP-1,ICAM-1 and VCAM-1 in normal and blank control group were all being 1,the mRNA expression levels (2-△ △Ct) of the above mentioned inflammatory factors and adhesion molecule in animal model group were 3.96 ± 0.39,3.07 ± 0.27,3.71 ± 0.26,3.94 ± 0.26,and the mRNA expression levels (2-△ △Ct) of the above mentioned inflammatory factors and adhesion molecule in HUVEC model group were 3.53±0.70,2.24±0.48,3.13±0.44,2.80±0.13,respectively,all P ＜ 0.01].The content of CD62E in serum was increased significantly in model group compared with that in normal control group (μg/L:1.31 ± 0.22 vs.0.82 ± 0.13,P ＜ 0.01),the mRNA expression levels of IL-8,MCP-1,ICAM-1 and VCAM-1 were decreased significantly in low,middle,high dose Sheshang capsule groups compared with those in model group in endothelial cells of aorta abdominalis of rabbits and HUVEC [abdominal aorta:IL-8 mRNA (2-△ △Ct) were 1.13 ± 0.19,1.26 ± 0.16,1.27 ± 0.17 vs.3.96 ± 0.39,MCP-1 mRNA (2-△ △ Ct) were 1.79 ± 0.24,2.22 ± 0.38,1.76±0.19 vs.3.07±0.27,ICAM-1 mRNA (2 △△Ct) were 2.05±0.11,1.68±0.09,2.37±0.48 vs.3.71±0.26,VCAM-1 mRNA (2-△△Ct) were 1.59±0.08,1.40±0.11,1.84±0.11 vs.3.94±0.26;HUVEC:IL-8 mRNA (2-△△Ct) were 2.33±0.59,2.82±0.82,2.51±0.77 vs.3.53±0.70,MCP-1 mRNA (2-△△Ct) were 1.59±0.35,1.48±0.36,1.54±0.29 vs.2.24±0.48,ICAM-1 mRNA (2-△△Ct) were 1.46±0.38,1.77±0.65,1.73±0.50 vs.3.13±0.44,VCAM-1 mRNA (2-△△Ct) were 2.49±0.24,2.18±0.19,2.45±0.24 vs.2.80±0.13,all P ＜ 0.05].The contents of CD62E were decreased significantly in middle,high dose Sheshang capsule groups compared with the content in model group (μg/L:1.01 ±0.14,1.04±0.13 vs.1.31 ±0.22,all P ＜ 0.01),but there were no statistical significant differences among the three drug group (all P ＞ 0.05).Conclusion The therapy of purging fire and removing toxin can treat vascular endothelial injury by inhibiting the inflammatory response induced by Trimeresurus stejnegeri bites.

Objective To observe the effect of Astragalus injection on the expressions of inflammatory cytokines in human primary macrophages stimulated by lipoteichoic acid (LTA) and lipopolysaccharide (LPS),and investigate its effects on inflammatory reactions of Gram-positive (G+) and Gram-negative (G-) bacteria sepsis and its mechanisms.Methods Percoll density gradient centrifugation was used to isolate the human peripheral blood mononuclear cells,then they were purified by immune Anti-Biotin Microbeads with magnetic character and under the induction of recombinant human granulocyte-macrophage colony stimulating factor (rhGM-CSF),the cells were cultivated for 12 days in vitro,eventually the human monocyte-derived macrophage was formed.The cultured human macrophages were inoculated in 96-well plates (each group 3 wells) and 6-well plates (each group 3 wells).The cells were divided into control group (200 μL DMEM added in each well),LTA 1 mg/L group,LPS 0.1 mg/L group and low astragalus injection (0.1 mg/L) and high astragalus injection (0.2 mg/L) dose groups.After the incubator plates were put in an incubator for 24 hours,the protein content of IL-8 and IL-10 in supernatant were detected by enzymelinked immunosorbent assay (ELISA),and the mRNA expression levels of IL-8 and IL-10 were detected by real time fluorescence quantitative reverse transcription-polymerase chain reaction (RT-qPCR).Results LTA and LPS all can obviously up-regulate the expression levels of pro-inflammatory factor IL-8 and anti-inflammatory factor IL-10 of macrophage.The expressions of IL-8 and IL-10 protein and mRNA in LTA group and LPS group were significantly higher than those in control group after cuhure for 8 hours and 24 hours,the degrees of increment were more significantly at 24 hours [LTA stimute group:IL-8 protein (ng/L,× 103):41.57± 1.90 vs.1.58 ±0.24,IL-8 mRNA (A value):21.49±8.05 vs.1.00±0.16;IL-10 protein (ng/L):5.90±3.02 vs.2.91 ± 1.54,IL-10 mRNA (A value):1.35±0.34 vs.0.95±0.14;LPS stimute group:IL-8 protein (ng/L,× 103):345.00±22.80 vs.5.60±0.31,IL-8 mRNA (A value):29.84 ± 8.93 vs.1.00 ± 0.16,IL-10 protein (ng/L):122.37 ± 39.26 vs.44.79 ± 3.67,IL-10 mRNA (A value):7.38 ± 1.58 vs.1.35 ± 0.34,all P ＜ 0.05].The Astragalus injection could regulate LTA and LPS to stimulate the macrophage to decrease the expression levels of pro-inflammatory factor IL-8 protein and mRNA and increase the expression levels of anti-inflammatory factor IL-10 protein and mRNA in the macrophage;the changes of regulatory effect in the 24 hour-culture of Astragalus injection high dose group was the most significant [LTA stimute group:IL-8 protein (ng/L,×103):22.63±1.91 vs.41.57±1.90,IL-8 mRNA (A value):12.10±1.93 vs.21.49±8.05,IL-10 protein (ng/L):14.03±2.22 vs.5.90±3.02,IL-10 mRNA (A value):10.37±6.08 vs.1.35±0.34;LPS stimute group:IL-8 protein (ng/L,× 103):167.75 ± 19.90 vs.345.01 ±22.80,IL-8 mRNA (A value):15.61 ± 3.63 vs.29.84±8.93;IL-10 protein (ng/L):243.22±14.41 vs.122.37±39.26,IL-10 mRNA (A value):16.14±4.10 vs.7.38± 1.58,all P ＜ 0.05].Conclusions In the process of inflammatory response,the pro-inflammatory and anti-inflammatory factors co-exist simultaneously.Astragalus injection can inhibit the expression levels of pro-inflammatory factor gene and protein in the inflammatory response of G+ and G-bacteria sepsis and in the mean time,it can promote the expression levels of anti-inflammatory factor gene and protein,thus the immune mechanism of sepsis is affected,achieving the balance between pro-inflammation and anti-inflammation.

Objective To observe the effect of monosialoteterahexosyl ganglioside (GM1) combined with Shuxuening injection on nerve function in patients with acute cerebral infarction (ACI) and its mechanism.Methods A total of 94 patients with ACI admitted to the Department of Neurology in Xiangyang First Peoples' Hospital Affiliated to Hubei Medical College from January 2013 to June 2016 were enrolled,and they were divided into two groups by random number table,each group 47 cases.The patients in two groups were all given conventional western medicine treatment;The patients in one group (single group) were treated by intravenous (Ⅳ) drip of GM1,100 mg once a day;and the patients in another group (combined group),by above GM1 Ⅳ drip combined with Shuxuening intramuscular injection,once 2 mL,twice a day;the therapeutic course in two groups was 14 days.Before and after treatment,the changes of China stroke clinical neurological impairment score (CSS score),glasgow coma score (GCS),nerve factor,oxidative stress index and hemodynamics index of two groups were observed.Results Compared with those before treatment,after treatment the CSS score,the levels of neuron specific enolase (NSE),and malondialdehyde (MDA) were significantly lower,while the GCS score,the levels of nerve growth factor (NGF),neurotrophic factor (NTF),maximum blood flow velocity (Vmax),minimum blood flow velocity (Vmin),mean blood flow velocity (Vmean),mean blood flow quantity (Qmean),glutathione peroxidase (GSH-Px),catalase (CAT) and superoxide dismutase (SOD) were all significantly higher in the combined group (all P ＜ 0.05).After treatment,the CSS score,levels of NSE and MDA in the combined group were significantly lower than those of the single group [CSS:11.20 ± 1.78 vs.16.24 ± 1.95,NSE (μg/L):13.17± 1.00 vs.17.68 ± 1.84,MDA (μmol/L):4.14±0.49 vs.5.61 ±0.50,all P ＜ 0.05],GCS score,NGF,NTF,GSH-Px,CAT,SOD,Vmax,Vmin,Vmean and Qmean of the combined group were all significantly higher than those of the single group [GCS:13.68± 1.85 vs.12.01±1.76,NGF (ng/L):88.10±8.83 vs.68.13±7.16,NTF (pg/L):5.13±0.38 vs.3.71±0.30,GSH-Px (U/L):128.13±8.07 vs.103.90±6.58,CAT (U/L):25.74±2.15 vs.19.43± 1.84,SOD (μU/L):94.36±8.00 vs.77.29±7.34,Vmax (cm/min):48.23±3.36 vs.43.17±2.88,Vmin (cm/'min):8.11±0.76 vs.6.85 ± 0.64,Vmean (cm/min):18.69 ± 1.37 vs.15.60 ± 1.24,Qmean (mL/min):9.10 ± 0.74 vs.7.79 ± 0.66,all P ＜ 0.05].Conclusions GM1 combined with Shuxuening injection can improve nerve function in patients with ACI by improving brain blood flow,secreting neurotrophic related factors and inhibiting oxidative stress reaction,thus it has important clinical significance for repairing the damaged nerve function.