Study on the toxic mechanism of Mahuang xixin fuzi decoction based on fecal metabolomics / 中国药房

OBJECTIVE To study the toxic mechanism of Mahuang xixin fuzi decoction （MXF） on normal mice. METHODS Totally 48 SPF grade BABL/C mice were randomly divided into blank group， MXF low-dose， medium-dose and high-dose groups， with 12 mice in each group. MXF low-dose， medium-dose and high-dose groups were given drug intragastrically at the dose of 11.262， 33.786， 45.050 g/kg， respectively. Blank group was administered with equal volume of normal saline， once a day， for consecutive 7 d. The body weight， anal temperature and survival rate were recorded， organ index and serum biochemical factors were detected. After the last administration， fecal samples of mice were collected and detected by UHPLC-QE/MS. RESULTS Compared with blank group， the body weight was decreased significantly from the 3rd to the 5th day after administration in MXF medium-dose group， and from the 2nd to the 7th day after administration in MXF high-dose group significantly （P＜0.05）. There was no significant difference in anal temperature among the treatment groups； the average survival rates of MXF medium-dose and high-dose groups were 58.33% and 50.00%， respectively. Compared with blank group， there were significant difference in the indexes of spleen， lung， thymus， adrenal gland and creatine kinase in MXF low-dose， medium-dose and high-dose groups， the testis index in MXF low-dose and high-dose groups， the creatine kinase isoenzyme/creatine kinase ratio in MXF low-dose group， the α-hydroxybutyrate dehydrogenase， lactate dehydrogenase and alkaline phosphatase in MXF medium-dose group， the urine and cystatin C in MXF medium-dose and high-dose groups （P＜0.05）. The fecal metabonomic analysis showed that 19 biomarkers such as phenylpyruvate， L-tyrosine， phosphatidylcholine， glycerol 3-phosphate in MXF low-dose， medium-dose and high-dose groups were significantly different from those in the blank group. CONCLUSIONS When MXF reaches a certain dose， it will have adverse effects on the body weight， multiple organs and serum biochemical indicators of mice， thus showing a certain toxic effect. Its mechanism may be related to disrupting the intestinal flora metabolism， causing inflammatory reaction and immune disorders.

Protective effect of Asarum polysaccharide on H1N1 influenza virus infection and expression of inflammatory factors / 中国中药杂志

In this paper, Asarum polysaccharides(AP) were extracted, and its composition was analyzed to study the activity against H1 N1 influenza virus in vitro and its intervention effect on mice with kidney Yang deficiency syndrome. AP was prepared by the strategy of water extraction and alcohol precipitation, the content was determined, and its monosaccharide composition was analyzed. The cell Real-time monitoring system and Reed-Muench model were adopted to evaluate the antiviral activity of AP in vitro. And the mouse model of kidney Yang deficiency syndrome was established in vivo to compare the efficacy of Mahuang Xixin Fuzi Decoction(MXF) and AP. MXF group and AP group were treated with clinical equivalent doses of 1.8 g·kg~(-1)·d~(-1) and 0.077 g·kg~(-1)·d~(-1) respectively, once a day for 6 consecutive days. Real-time PCR was used to detect the relative expression of M gene of H1 N1 influenza virus and cytokines in lung tissue. The content of AP in Asarum was 25.22%, and the protein content was 0.8%. And the monosaccharide composition was identified as L-rhamnose, D-arabinose, D-xylose, D-glucose, D-galactose and D-mannose. TI values of Tamiflu, MXF and AP were 30.00, 8.06 and 10.33, respectively. Three different doses of AP could significantly reduce the concentration of virus in supernatant. Compared with the model mice, lung indexes of MXF group and AP group decreased significantly(P<0.05), and the relative expression of M gene decreased significantly(P<0.05). The relative expressions of IL-10 and IFN-γ were up-regulated to varying degrees, while the relative gene expressions of IL-1β, IL-6 and MCP-1 were down-regulated to different degrees. In addition, AP could significantly enhance the expression of TNF-α(P<0.01). AP had a good anti-influenza virus activity in vitro, and could protect mice with kidney Yang deficiency syndrome by reducing the viral load in lung tissue, decreasing inflammation damage in lung tissue, and regulating the expression of inflammatory cytokines. Compared with the prescription of MXF, AP had a better antiviral activity.

Effects of Mahuang Xixin Fuzi Decoction on TLRs Response and Cyt-CO-mediated Apoptosis Regulation in Mice with Influenza Disease of Kidney-yang Deficiency / 中国药房

OBJECTIVE：To study the effects of Mahuang xixin fuzi decoction on Toll-like receptors （TLRs）response and cytochrome C oxidase （Cyt-CO）-mediated apoptosis regulation in mice with influenza disease of kidney-yang deficiency. METHODS：Totally 48 male Balb/c mice were randomly divided into normal group （n＝12）and modeling group （n＝36）. The modeling group was intraperitoneally injected with estradiol benzoate solution （8 mg/kg）and intranasally injected with influenza virus H 1N1（20 μL/mice）to establish the influenza disease compound model of kidney-yang deficiency. After modeling ，the mice were randomly divided into model group ，positive drug group （Oseltamivir phosphate capsules ，0.195 g/kg），Mahuang xixin fuzi decoction group （1.802 g/kg，by crude dru g），with 12 mice in each group. Each group was given relevant medicine intragastrically，normal group and model group were given， corresponding volume of normal saline intragastrically 20 mL/kg，once a day ，for consecutive 6 days. During admi-nistration，body weight and anal temperature of mice were mail：xsy407861520@163.com measured daily ；the percentage of initial body weight was calculated. After last medication ，the organ （spleen，thymus and lung ）indexes were calculated ；the pathological changes of lung tissue were observed. The viral load of influenza A virus H 1N1 in lung tissue was detected （reflected by M gene mRNA expression）；mRNA expressions of TLR3，TLR7，myeloid differentiation factor （MyD88）and Caspase- 3 in cardiac tissue as well as the activity of Cyt-CO and the content of cytochrome C （Cyt-C）were also determined. RESULTS ：Compared with normal group，initial body weight percentage and anal temperature of the model group continued to decrease （P＜0.05）；the spleen and thymus indexes were decreased significantly （P＜0.05），while lung index was increased significantly （P＜0.05）；the lung tissue lesions were serious. Viral load in lung tissue ，mRNA expressions of TLR 3，TLR7，MyD88 and Caspase- 3 in cardiac tissue as well as the content of Cyt-C were increased significantly （P＜0.05 or P＜0.01），while the activity of Cyt-CO in cardiac tissue was significantly decreased （P＜0.01）. Compared with model group ，initial body weight percentage and anal temperature of mice in Mahuang xixin fuzi decoction group showed an increasing trend from the fourth day of administration （P＜0.05 or P＜0.01）. The spleen and thymus indexes were increased significantly （P＜0.05），while the lung index was significantly decreased （P＜0.05）；the pathological injury of lung tissue was significantly improved ；viral load in lung tissue ，mRNA expressions of TLR 3 and Caspase- 3 as well as the content of Cyt-C in cardiac tissue were decreased significantly （P＜0.05 or P＜0.01），while the activity of Cyt-CO was increased significantly in cardiac tissue （P＜0.01）. CONCLUSIONS ：Mahuang xixin fuzi decoction can improve influenza disease of kidney-yang deficiency in mice ，the effect may related to inhibit TLRs response and apoptosis regulation pathway mediated by Cyt-CO.

Efficacy-toxicity Evaluation of Mahuang Xixin Fuzi Decoction on Mice with Kidney-yang Deficiency / 中国药学杂志

OBJECTIVE: To study the pharmacodynamic effects of Mahuang Xixin Fuzi Decoction (MXF) on mice with kidney-yang deficiency, its toxic effects on normal mice and effect-toxic mechanism. METHODS: BABL/C male mice were randomly divided into normal groups; normal groups with low, medium and high toxicity; kidney Yang deficiency model group. The low, medium and high dose groups were treated with infected Influenza virus of kidney-yang deficiency. After continuous gavage for 6 days, the body weight anal temperature and other pharmacological indicators were recorded, and the general signs, biochemical indicators and inflammatory factors of the mice were detected. RESULTS: Compared with the normal group, the biochemical parameters of the kidney yang deficiency model group, which means the creatine kinase (CK), creatine kinase-isoenzyme (CK-MB), alpha-hydroxybutyrate dehydrogenase (α-HBDH), lactate dehydrogenase (LDH), alanine aminotransferase (ALT), aspartate transferase (AST), alkaline phosphatase (ALP) and urea (urea) were significantly increased (P<0.05), inflammatory factors interleukin-6 (IL-6), monocyte deproteinization protein (MCP-1), tumor necrosis factor-α (TNF-α), interleukin-10 (IL- 10), interferon-γ (INF-γ) and interleukin 12 (IL-12p70) increased significantly (P<0.05). After administration, the levels of biochemical markers such as CK, ALT and urea in each administration groups were slightly reduced compared with the model group. The levels of inflammatory factors such as IL-6, IL-10 and IFN-γ in each administration groups showed different degrees of callback compared with the model group. In the toxicity study, the biochemical indicators of CK, urea, and cysteine (CYS-C) in each drug-administered group were significantly higher than those in the normal group (P<0.05), and the inflammatory factor interleukin-10 (IL-10) increased significantly, and the content of interleukin-12 (IL-12p70) decreased significantly (P<0.05). CONCLUSION: After treatment with MXF, the above biochemical indicators of mice with kidney yang deficiency and external sensation have different degrees of callback. MXF can achieve therapeutic effects by regulating IL-6, IL-10, IFN-γ, MCP-1, TNF-α and other factors on the body′s inflammatory response. While normal mice give different doses of MXF, the heart, liver and kidney function are dysfunctional, the balance of immune response of the body are broken through cell transfer, thus making normal mouse developed an inflammatory response.

Effects of different decocting methods on quality of Mahuang Xixin Fuzi decoction / 中国中药杂志

To compare the quality difference between Mahuang Xixin Fuzi decoction(MXF) prepared by traditional decocting method and that prepared by two commonly used decocting methods, and explore the scientific nature of the traditional decocting method. By taking effect-toxic components in MXF as the research object, this article investigated these three different decocting methods from the quantitative determination of effect-toxic components in MXF. By using multivariate statistical analysis methods, three characteristic constituents were identified as kakoul, mesaconitine (MA) and hypaconitine (HA) respectively. As compared with two commonly used decocting methods, MXF decoction prepared by traditional decocting method had the shortest boiling time, but with the lowest dissolution rates of MA and AC and the higher dissolution rates of mono-ester aconitum alkaloids. In addition, the traditional decocting method increased the dissolution of ephedra alkaloid and accelerated the hydrolysis of diester diterpenoid alkaloids. There were differences in the content of effect-toxic components in MXF decoctions prepared by three different decocting methods, which can provide a reference for use of the classical prescriptions.

Action mechanism of Mahuang Xixin Fuzi decoction for mice with influenza based on metabolomics information / 中国中药杂志

This study aimed to analyze the endogenous metabolite changes in the serum of mice infected with H1N1 virus after intervention by Mahuang-Xixin-Fuzi decoction (MXF) based on metabolomics method, investigate potential biomarkers and related metabolic pathways, and explore the therapeutic mechanism of MXF through metabolomics technology. Thirty-six Kunming (KM) mice were randomly divided into three groups: normal group, model group and MXF group. Influenza virus H1N1 was used by nasal drip to establish influenza mice model. The mice in MXF group were orally administrated with MXF for 6 consecutive days after inoculation, and the other two groups were given with equal volume of saline solution in the same way. Body weight, rectal temperature, morbidity and mortality were recorded daily. Serum samples were collected 24 hours after the last administration for HPLC-TOF-MS analysis. The results showed that as compared with the normal group, the body weight and rectal temperature were decreased in model group, and their lung index and mortality rate were significantly increased (P<0.05); MXF had good therapeutic effects on the abnormity of body weight, rectal temperature, lung index and high mortality rate of mice infected with H1N1 virus. The original data collected from the serum samples were analyzed with R language, MPP, SIMCA-P and other software, and significant changes were found in 14 kinds of endogenous substances from mice serum (P<0.05). As compared with model group, the potential metabolic markers in MXF group recovered to normal levels to a certain degree after being intervened by MXF. Further analysis with MetPA data platform showed that, the pathways involved in 14 metabolites included glucose metabolism, arachidonic acid metabolism, glycerophospholipids and sphingolipids metabolism etc. The metabolomics study and pharmacological experiment showed that MXF might play a role of efficacy by improving glucose metabolism, regulating arachidonic acid metabolism, glycerophospholipid and sphingolipid metabolic pathways.

Effects of Mahuang-Xixin-Fuzi decoction on plasma levels of interlcukin-6 and tumor necrosis factor-α in allergic rhinitis in rats / 国际中医中药杂志

Objective To investigate the effects of Mahuang-Xixin-Fuzi decoction on the plasma levels of interlcukin-6 (IL-6) and tumor necrosis factor-α (TNF-α) in allergic rhinitis in rats. Methods Thirty-two Wistar rats were randomly divided into a control group, a model group, a chlorpheniramine group, and a Mahuang-Xixin-Fuzi decoction group, with 8 rats in each group. Allergic rhinitis was induced by ovalbumin sensitization. Rats in the chlorpheniramine and Mahuang-Xixin-Fuzi decoction groups received intragastric administration of 0.01%chlorpheniramine and Mahuang-Xixin-Fuzi decoction 0.4 g (twice daily for 21 days), respectively. The plasma levels of IL-6 and TNF-αwere measured with enzyme-linked immunosorbent assay. Results The frequencies of sneezing (2.4 ± 0.9/30 min vs.18.5 ± 3.6/30 min) and nasal rubbing (4.2 ± 1.5/30 min vs. 31.2 ± 3.7/30 min) in the Mahuang-Xixin-Fuzi decoction group were significantly lower than those in the model group (all P<0.05). The plasma levels of IL-6 (153.00 ± 13.87 pg/ml vs. 230.38 ± 33.92 pg/ml) and TNF-α(21.25 ± 4.62 pg/ml vs. 56.38 ± 7.93 pg/ml) in the Mahuang-Xixin-Fuzi decoction group were significantly lower than those in the model group (all P<0.05). Conclusions Mahuang-Xixin-Fuzi decoction can reduce the plasma levels of IL-6 and TNF-α, and alleviate symptoms in allergic rhinitis in rats.