A comprehensive investigation on the chemical changes of traditional Chinese medicine with classic processing technology: Polygonum multiflorum under nine cycles of steaming and sunning as a case study.

The processing of traditional Chinese medicine (TCM) plays an important role in the clinical application, which usually has the function of "increasing efficiency and reducing toxicity". Polygonum multiflorum (PM) has been reported to induce hepatotoxicity, while it is believed that the toxicity is reduced after processing. Studies have shown that the hepatotoxicity of PM is closely related to the changes in chemical components before and after processing. However, there is no comprehensive investigation on the chemical changes of PM during the processing progress. In this research, we established a comprehensive method to profile both small molecule compounds and polysaccharides from raw and different processed PM samples. In detail, an online two-dimensional liquid chromatography coupled with quadrupole-orbitrap mass spectrometry (2D-LC/Q-Orbitrap MS) was utilized to investigate the small molecules, and a total of 150 compounds were characterized successfully. After multivariate statistical analysis, 49 differential compounds between raw and processed products were screened out. Furthermore, an accurate and comprehensive method for quantification of differential compounds in PM samples was established based on ultra-high performance liquid chromatography/Q-Orbitrap-MS (UHPLC/Q-Orbitrap-MS) within 16 min. In addition, the changes of polysaccharides in different PM samples were analyzed, and it was found that the addition of black beans and steaming times would affect the content and composition of polysaccharides in PM significantly. Our work provided a reference basis for revealing the scientific connotation of the processing technology and increasing the quality control and safety of PM.

Potential Involvement of Organic Anion Transporters in Drug Interactions with Shuganning Injection, a Traditional Chinese Patent Medicine.

Traditional Chinese medicine injections have been widely used in China for the treatment of various diseases. Transporter-mediated drug-drug interactions are a major contributor to adverse drug reactions. However, the research on transporter-mediated Traditional Chinese medicine injection-drug interactions is limited. Shuganning injection is a widely used Traditional Chinese medicine injection for treating various liver diseases. In this study, we investigated the inhibitory effect of Shuganning injection and its four main ingredients (baicalin, geniposide, chlorogenic acid, and oroxylin A) on 9 drug transporters. Shuganning injection strongly inhibited organic anion transporter 1 and organic anion transporter 3 with IC50 values < 0.1% (v/v), and moderately inhibited organic anion transporter 2, organic anion transporting-polypeptide 1B1, and organic anion transporting-polypeptide 1B3 with IC50 values < 1.0%. Baicalin, the most abundant bioactive ingredient in the Shuganning injection, was identified as both an inhibitor and substrate of organic anion transporter 1, organic anion transporter 3, and organic anion transporting-polypeptide 1B3. Oroxylin A had the potential to act as both an inhibitor and substrate of organic anion transporting-polypeptide 1B1 and organic anion transporting-polypeptide 1B3. In contrast, geniposide and chlorogenic acid had no significant inhibitory effect on drug transporters. Notably, Shuganning injection markedly altered the pharmacokinetics of furosemide and atorvastatin in rats. Using Shuganning injection as an example, our findings support the implementation of transporter-mediated Traditional Chinese medicine injection-drug interactions in the development of Traditional Chinese medicine injection standards.

Critical influence of cytokines and immune cells in autoimmune gastritis.

Gastric cancer (GC) is a type of the most common cancers. Autoimmune gastritis (AIG) and infection with Helicobacter pylori (HP) are the risk factors of triggering GC. With the emphasis on the treatment of HP, the incidence and prevalence of HP infection in population is decreasing. However, AIG lacks accurate diagnosis and treatment methods, which occupies high cancer risk factors. AIG is controlled by the immune environment of the stomach, including immune cells, inflammatory cells, and infiltrating intercellular material. Various immune cells or cytokines play a central role in the process of regulating gastric parietal cells. Abnormal expression levels of cytokines involved in immunity are bound to face the risk of tumorigenesis. Therefore, it is particularly important for preventing or treating AIG and avoiding the risk of gastric cancer to clarify the confirmed action mode of immune cells and cytokines in the gastric system. Herein, we briefly reviewed the role of the immune environment under AIG, focussing on describing these double-edged effects between immune cells and cytokines, and pointing out potential research challenges.

Cytochrome P450-mediated herb-drug interaction (HDI) of Polygonum multiflorum Thunb. based on pharmacokinetic studies and in vitro inhibition assays.

BACKGROUND: Polygonum multiflorum Thunb. (PM) is well known both in China and other countries of the world for its tonic properties, however, it has lost its former glory due to liver toxicity incidents in recent years. PURPOSE: The purpose of this study is to determine whether the occurrence of herb-drug interaction (HDI) caused by PM is associated with cytochrome P450 (CYP450) based on pharmacokinetic studies and in vitro inhibition assays. The objective was to provide a reference for the rational and safe use of drugs in clinical practice. METHODS: In this study, raw PM (R), together with its two processed products which included PM by Chinese Pharmacopoeia (M) and PM by "nine cycles of steaming and sunning (NCSS)" ("9"), were prepared as the main research objects. A method based on fluorescence technology was used to evaluate the inhibition levels of raw and processed PMs, as well as corresponding characteristic compounds on seven recombinant human cytochrome P450s (rhCYP450s). The pharmacokinetics of sulindac (a representative of commonly used nonsteroidal anti-inflammatory drugs) and psoralen (a major compound of Psoralea in combination with PM) in rat plasma were studied when combined with raw and different processed products of PM. RESULTS: The inhibitory level order of the three extracts on major different subtypes of CYP450 (CYP1A2, CYP2B6, CYP2C8, CYP2C19, CYP2D6, and CYP3A4) was: R > M > "9". However, the inhibition level of R and "9" is higher than that of M on CYP2C9. Further studies showed that trans-THSG and emodin could selectively inhibit CYP3A4 and CYP1A2, respectively. Epicatechin gallate mainly inhibited CYP3A4 and CYP1A2, followed by CYP2C8 and CYP2C9. Genistein mainly inhibited CYP3A4, followed by CYP2C9 and CYP2C8. CYP3A4 and CYP2C9 were also inhibited by daidzein. The inhibitory effects of all the PM extracts were associated with their characteristic compounds. The results of HDI showed that R increased sulindac exposure to rat blood, and R and M increased psoralen exposure to rat blood, which were consistent with corresponding metabolic enzymes. Overall, the in vitro and in vivo results indicated that PM, especially R, would be at high risk to cause toxicity and drug interactions via CYP450 inhibition. CONCLUSION: This study not only elucidates the scientific connotation of "efficiency enhancement and toxicity reduction" of PM by NCSS from the perspective of metabolic inhibition but also contributes to HDI prediction and appropriate clinical medication of PM.

Characterization of volatile organic compounds in Polygonum multiflorum and two of its processed products based on multivariate statistical analysis for processing technology monitoring.

Herein we describe a comprehensive analysis of the volatile organic compounds (VOCs) of raw Polygonum multiflorum Thunb. (PM) and two of its processed products, as well as an effective and simple method based on volatile markers to determine to which extent the PM had been processed. Sixty-five VOCs were identified by headspace-solid-phase microextraction-gas chromatography-mass spectrometry (HS-SPME-GC-MS), along with headspace-gas chromatography-ion mobility spectrometry (HS-GC-IMS). Principal component analysis (PCA) of the HS-SPME-GC-MS spectra and fingerprint analysis of the HS-GC-IMS spectra allowed the identification of raw PM from its processed products based the VOCs identified. Furthermore, the content and distribution of VOCs in the samples were easily analyzed visually based on clustering-kernel density estimation (Cluster-KDE). Finally, exploratory factor analysis (EFA) allowed the screening of significant markers to identify the processing method and consequently distinguish the three studied groups of PM.

Studies on Chemical Characterization of Ginkgo Amillaria Oral Solution and Its Drug-Drug Interaction With Piceatannol 3'-O-ß-D-Glucopyranoside for Injection.

Ginkgo Amillaria oral solution (GAO) is commonly used for the treatment of cardiovascular and cerebrovascular diseases in China. Piceatannol-3'-O-ß-D-glucopyranoside for injection (PGI) is mainly used for the prevention and treatment of ischemic cerebrovascular diseases. With the spread of cerebrovascular disease, the possibility of combining the two drugs has increased; however, there is no research on the drug-drug interaction (DDI) between these two medicines. In this paper, an ultrahigh-performance liquid chromatography/quadrupole-orbitrap mass spectrometry (UHPLC/Q-Orbitrap MS) method was established to characterize the chemical constituents of GAO first; 62 compounds were identified or tentatively identified based on their retention time (RT), MS, and MS/MS data. Nine main compounds were determined by ultrahigh-performance liquid chromatography/triple quadrupole mass spectrometry (UPLC-QQQ-MS). Furthermore, incubation with liver microsomes in vitro was fulfilled; the results showed that GAO had a significant inhibitory effect on UGT1A9 and UGT2B7 (p < 0.05), and PGI was mainly metabolized by UGT1A9. The identification results of in vivo metabolites of PGI showed that PGI mainly undergoes a phase II binding reaction mediated by UDP-glucuronosyltransferase (UGT) and sulfotransferase (SULT) in vivo. Therefore, pharmacokinetic studies were performed to investigate the DDI between GAO and PGI. The results showed that the AUC (p < 0.05) and T1/2 (p < 0.05) of PGI in vivo were significantly increased when administered together with GAO, whereas the CL was significantly decreased (p < 0.05). The exploration of in vitro and in vivo experiments showed that there was a DDI between GAO and PGI.

A strategy based on gene sequencing and molecular docking for analysis and prediction of bioactive peptides in Shuxuetong injection.

Peptides are a class of protein fragments with relatively high biological activity and intense specificity, which play crucial role in the treatment of Shuxuetong injection (SXT). However, the extraordinary complexity of Chinese medicinal formulates and the lack of systematic identification methods are primary challenges for study of pharmacodynamic peptides. In addition, infinitesimal peptides contents further hinder the identification and structural characterization of polypeptide by traditional means. In this paper, we described a strategy that LC-MS combined with molecular docking to systematically illustrate the peptide components of SXT. The key to this research was used of gene sequencing to establish a SXT protein database to further achieve the separation and enrichment of chemical methods. Moreover, the ADRA2A, PAR4 and DRD3 were precisely docked with the identified peptides. The result indicated that 12 compounds had stable binding ability and were speculated to be the latent bioactive monomers for the treatment of stroke. Additionally, 12 peptides were verified by cell-based experiment. The results showed that only YLKTT could indeed protect astrocytes from oxygen glucose deprivation/reoxygenation (OGD/R). The YLKTT showed higher activity than the others in vitro. It might be a completely new compound that has never been reported before, providing the basis for further research and a new paradigm for stroke.

Biomarkers and Mechanism Analysis for Polygoni Multiflori Radix Preparata-Induced Liver Injury by UHPLC-Q-TOF-MS-Based Metabolomics.

BACKGROUND: Polygonum Multiflorum Radix Preparata (PMP), prepared from Polygonum multiflorum Thunb. (PM), is traditionally valued for its liver and kidney-tonifying effects. However, the previous studies showed that PMP was hepatotoxic, which limited its clinical use. Unfortunately, the potential hepatotoxic ingredients and the molecular mechanism are still uncertain. OBJECTIVE: The aim of this study was to find out potential biomarkers of hepatotoxicity using metabolomics profile. MATERIALS AND METHODS: 60% ethanol extract of PMP (PMPE) was prepared. Subsequently, an untargeted metabolomics technology in combination with ROC curve analysis method was applied to investigate the alteration of plasma metabolites in rats after oral administration of PMPE (40 g/kg/d) for 28 days. RESULTS: Compared to the control group, the significant difference in metabolic profiling was observed in the PMPE-induced liver injury group, and sixteen highly specific biomarkers were identified. These metabolites were mainly enriched into bile acids, lipids, and energy metabolisms, indicating that PMPE-induced liver injury could be related to cholestasis and dysregulated lipid metabolism. CONCLUSIONS: This study is contributed to understand the potential pathogenesis of PMP-induced liver injury. The metabonomic method may be a valuable tool for the clinical diagnosis of PMP-induced liver injury.

An effective strategy for distinguishing the processing degree of Polygonum multiflorum based on the analysis of substance and taste by LC-MS, ICP-OES and electronic tongue.

The efficacy of raw and processed products of Polygonum multiflorum (PM) varies greatly. "Nine cycles of steaming and sunning" (NCSS) is recognized as an effective technology in enhancing efficacy and reducing toxicity for PM. In this paper, PM was prepared differently into three groups (including group R, M, and "9"), which represent raw PM, PM processed using the method of Chinese Pharmacopoeia (ChP) and PM processed using traditional NCSS, respectively. The purpose is to establish an effective method to distinguish raw PM from different processed products and highlight the rationality of processing technology. The main organic compounds that could distinguish these three groups of samples were identified by in-depth mining of mass spectral information and various chemometric methods. Level of related metal cations have been quantified and used as another important distinguishing markers. The electronic tongue was utilized to determine the taste traits of aqueous extract from PM. Furthermore, the material basis that caused the difference in taste was discovered according to correlation analysis. In detail, saltiness has the most important contribution associated with the concentrations of K+ and Na+, however, bitterness and astringency were mainly associated with the contents of epicatechin gallate, catechin, procyanidin B1, procyanidin B2 and epicatechin. This study proposed a novel and effective strategy for identification of processing technology of PM. It lays the foundation for clarifying the modern scientific recommendations of processing technology to PM. On the other hand, it also provides a reference for related researches on other traditional Chinese medicine (TCM).

Multiple component-pharmacokinetic studies on 10 bioactive constituents of Peiyuan Tongnao capsule using parallel reaction monitoring mode.

Peiyuan Tongnao capsule (PTC) plays an important role in clinical application due to its excellent curative efficacy in the treatment of ischemic stroke and chronic cerebral circulation insufficiency. To standardize and rationalize the clinical application of PTC, a rapid and sensitive method based on ultra-high performance liquid chromatography/quadrupole-Orbitrap mass spectrometry with parallel reaction monitoring (PRM) mode was developed and validated for the pharmacokinetic (PK) study. Ten bioactive compounds (aucubin, salidroside, echinacoside, paeoniflorin, verbascoside, liquiritin, 2,3,5,4'-tetrahydroxy stilbene-2-O-ß-d-glucoside, coumarin, glycyrrhizic acid, and emodin) were simultaneously determined in rat plasma. All calibration curves exhibited good linearity (r2 > 0.99). The lower limits of quantification were 0.082-13.291 ng mL-1 . The intra- and inter-day precision was 0.54-12.36%, whereas the intra- and inter-day accuracy ranged from 100.45 to 114.00%. The mean extraction recoveries were 81.77-117.66%, and the average matrix effects (MEs) were 86.23-109.96%. The high extraction recoveries and acceptable MEs indicated that the pretreatment method was feasible. And the stability was acceptable under various storage conditions and processing procedures. The validated method was successfully applied to the multiple components-PK studies, which lay the foundation for further pharmacological and clinical research of PTC and may provide a reference for other traditional Chinese medicines.

The composition differences between small black beans and big black beans from different habitats and its effects on the processing of Polygonum multiflorum.

INTRODUCTION: The roots of Polygonum multiflorum (PM) serve as a classical traditional Chinese medicine (TCM), which has multiple biological activities. However, many cases of hepatotoxicity in PM have been reported in recent years. Processing PM with black beans decoction is one of the typical processing methods to reduce the hepatotoxicity of PM since ancient times. OBJECTIVES: To find potential effective constituents, as well as the optimal variety and origin of black beans for the processing of PM. METHODS: Based on ultrahigh-performance liquid chromatography Q-Orbitrap mass spectrometry (UHPLC-Q-Orbitrap-MS) analysis, we measured the contents of the two potential toxic compounds (emodin-8-O-glucoside and torachrysone-O-hexose) in raw PM (R-PM), PM processed with big black beans (B-PM) and PM processed with small black beans (S-PM). The flow cytometry method analysed the effects of different processed products of PM on apoptosis of L02 cells in different drug concentration. Proton nuclear magnetic resonance (1 H-NMR) and UHPLC-Q-Orbitrap-MS together with multivariate statistical analysis were used to systematically analyse the different components between small black beans (Small-BB) and big black beans (Big-BB) from 30 different habitats. RESULTS: The toxicity was ranked from small to large: S-PM < B-PM < R-PM. Processing PM with black beans could significantly decrease the apoptosis rate of L02 cells, especially when the drug concentration is 80 µg/mL. Besides, we find five differential compounds (α-arabinose, α-galactose, proline, isomer of daidzein and isomer of genistein) may be potential active ingredients. In terms of the black beans collected from 30 producing areas, we find that Small-BB from Weifang in Shandong province was optimum to processing PM, followed by Shangqiu in Henan province, Jilin and Liaoning province. CONCLUSION: The ingredients that affect the processing of PM may be attributed to α-arabinose, α-galactose, proline, isomer of daidzein and isomer of genistein in black beans. When the drug concentration is higher, the effect of reducing the hepatotoxicity of PM is better. Besides, Small-BB was more effective than Big-BB for reducing the toxicity of PM, especially Small-BB from Weifang in Shandong, Shangqiu in Henan province and northeast China.

Rapid identification of chemical composition and metabolites of Pingxiao Capsule in vivo using molecular networking and untargeted data-dependent tandem mass spectrometry.

Pingxiao capsule (PXC) is a herbal medicine used for adjuvant therapy in breast cancer. However, the constituents and absorbed components of the formula and their related metabolites have not been elucidated to date. PXC is a typical traditional Chinese medicine formula consisting of Strychnos nux-vomica L., Curcuma wenyujin Y. H., Agrimonia pilosa Ledeb., Toxicodendron vernicifluum, Trogopterus dung, alumen, potassium nitrate (saltpeter) and Citrus aurantium L. In this study, a ultra-high performance liquid chromatography system equipped with high resolution Q-Orbitrap mass spectrometry (MS) and comparative Global Natural Product Social molecular networking together with the Compound Discoverer software were used to identify metabolites of PXC in vitro and in vivo. Based on untargeted data-dependent MS2 and data-mining techniques, 89 peaks of alkaloids, flavonoids, organic acid and phenolic compounds were identified in a PXC 70% methanol extract. Furthermore, 15 absorbed prototype compounds and their metabolites were rapidly confirmed in rat blood. Glucuronidation, oxidation, methylation and hydroxylation were the main metabolic pathways. We fully clarified the chemical constituents of PXC and provided a scientific and efficient strategy for rapid discovery and identification of prototypes and their metabolites in rat plasma using high-resolution MS aided by Global Natural Product Social and Compound Discoverer software.

Pharmacokinetic studies unveiled the drug-drug interaction between trans-2,3,5,4'-tetrahydroxystilbene-2-O-ß-d-glucopyranoside and emodin that may contribute to the idiosyncratic hepatotoxicity of Polygoni Multiflori Radix.

Polygoni Multiflori Radix (PMR) has been a reputable tonifying traditional Chinese medicine for a long history. However, clinical side effects regarding its idiosyncratic hepatotoxicity are occasionally reported. The containing anthraquinones, particularly emodin, could cause liver injury in both in vitro and in vivo experiments. It is well-known that some compounds could influence other compounds' pharmacokinetic parameters significantly. In this work, the influence of trans-2,3,5,4'-tetrahydroxystilbene-2-O-ß-d-glucopyranoside (TSG) on the pharmacokinetic behavior of emodin in rats was evaluated by an ultra-high performance liquid chromatography/triple quadrupole mass spectrometry (UHPLC/MS-MS) approach. Pharmacokinetic parameters of emodin, PMR extract, and TSG-free PMR extract (prepared by a component "knock-out" strategy with TSG eliminated), in rats after one-day and seven-day administration were determined and compared. We found that, after seven-day administration of the whole PMR extract (rather than TSG-free extract), emodin in rats was accumulated. And accordingly, the exposure of emodin in rats pre-treated with single TSG for seven days could be significantly enhanced. The results indicate that TSG was able to accelerate the exposure and metabolism of emodin. The effect of TSG on the metabolic activities of cytochrome P450 enzymes was further assessed by an LC-MS cocktail method. The accelerated exposure and metabolism of emodin could result from the up-regulation activity of CYP450s, in particular CYP1A2 isozyme. The findings obtained in this work firstly unveiled DDI between TSG and emodin in the administration of PMR, thus may provide a basis for unveiling the underlying mechanism of PMR-induced liver injury.

Plasma Transfusion in Patients With Cirrhosis in China: A Retrospective Multicenter Cohort Study.

Patients with cirrhosis used to be associated with frequent use of blood components because of their complex disorder of hemostasis and bleeding complications. Recent findings have indicated that patients with cirrhosis have a state of "rebalanced" or even procoagulant hemostasis and have questioned the prophylactic use of plasma. To evaluate the current status of plasma use in patients with cirrhosis, we conducted a retrospective survey in 11 tertiary-care hospitals in China from September 1 to October 31, 2013. All patients admitted with cirrhosis during the study period were included in the study. The survey collected information including patients' diagnostic and demographic data, clinical course including bleeding complications and invasive procedures, laboratory results, and plasma transfusion data. Among 1595 patients with cirrhosis admitted to the 11 hospitals, 236 (14.8%) patients received 1 or more plasma transfusions during the study period. The number of plasma transfusions is defined as the number of transfusion orders. A total of 1037 plasma transfusions were administered to these patients, with a mean of 4.4 transfusions per transfused patient, ranging from 1 to 22 transfusions per transfused patient. Most plasma transfusions (760/1037; 73.3%) were given to patients without bleeding, for treatment of coagulopathy either without planned invasive procedures (70.4%) or before invasive procedures (2.9%). The median dose of plasma transfusion was 3.8 mL/kg. The rate of plasma transfusion of participating hospitals varied from 5.3% to 31.8%. It is encouraging to see that in one teaching hospital, 85.7% plasma transfusions were given to patients with bleeding indication, showing a promising sign in appropriate transfusion. Prophylaxis or empirical plasma transfusion is still a common problem in managing patients with liver cirrhosis. Wide variations are found in plasma transfusion practice among hospitals. Effective measures to control and reduce empirical correction of abnormal coagulation tests through transfusing plasma should be strengthened urgently.

A multicenter study of blood component transfusion in patients with liver cirrhosis in China: Patient characteristics, transfusion practice, and outcomes.

BACKGROUND: Cirrhosis is a complex acquired disorder of hemostasis and patients frequently receive blood transfusions. But there is very limited data on patterns of blood use at a patient level. AIMS: To characterize blood use in cirrhotic patients in China and compare with recommendations to help identify areas where quality improvement strategies can be targeted. We also compared findings to a similar study undertaken in UK. METHODS: A cross-sectional study was conducted in 11 hospitals over a 2-month period. Data were collected prospectively on each hospitalized cirrhotic patient to day 28. RESULTS: 1595 cirrhotic patients were included and 20.6% were transfused. 48.2% of transfused patients received transfusion for bleeding, most commonly gastrointestinal bleeding (65.8%). The remaining 51.8% were transfused for non-bleeding indications. 32.5% of patients transfused for gastrointestinal bleeding with red blood cells had a pre-transfusion haemoglobin >7g/dL. 89.1% of patients transfused frozen plasma for non-bleeding indications received them in the absence of a planned procedure. The patterns of blood transfusion in cirrhosis were different between China and UK. Of note, empirical prophylactic use of frozen plasma was more common in the Chinese study (89%) than in the UK (24%). CONCLUSION: Education and research should be implemented to improve patient blood management, especially in prophylactic frozen plasma use area.

Immunosuppressive effects of mesenchymal stem cell transplantation in rat burn models.

OBJECTIVE: This study is to investigate the effects of mesenchymal stem cell (MSC) transplantation in burn treatment. METHODS: Wharton's Jelly was stripped from neonatal umbilical cord, and human umbilical cord MSCs were then cultured. Burn models were constructed in male SD rats weighted at 200±5 g, and the rats were randomly divided into control and MSCs transplantation groups. The rats in transplantation group were injected subcutaneously with MSCs (2×10(6)) at 24 h after burning. Blood samples were collected at 0 d, 1 d, 2 d, 3 d, 5 d and 7 d after burning and the contents of white blood cells (WBC), C-reactive protein (CRP), interferon-γ (IFN-γ), tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6) and interleukin-10 (IL-10 ) were detected. The wound healing rate at 7 d, 14 d, 21 d and 28 d together with the wound healing time were compared and analyzed statistically by analysis of variance (ANOVA). RESULTS: WBC and CRP in control group increased significantly at 1 d and 2 d, 2 d and 3 d, respectively. IFN-γ, IL-6 and IL-10 levels in serum showed increasing till 5th day and TNF-α arrived its peak value at 7th day. By contrast, WBC, CRP, TNF-α, IL-6 and IL-10 in the MSCs transplantation group showed slight increase after burning and the differences were verified by statistically analysis. IFN-γ showed no significant difference between the two groups. MSCs transplantation group showed significantly higher wound healing rate at 14 d, 21 d, 28 d and showed shorter wound healing time than control. CONCLUSIONS: MSCs transplantation could suppress secondary inflammatory reaction by lowering inflammatory cytokines after burning, thus promoting wound healing and scald repair in burn animal model.

Construction and analysis of the protein-protein interaction network related to essential hypertension.

BACKGROUND: Essential hypertension (EH) is a complex disease as a consequence of interaction between environmental factors and genetic background, but the pathogenesis of EH remains elusive. The emerging tools of network medicine offer a platform to explore a complex disease at system level. In this study, we aimed to identify the key proteins and the biological regulatory pathways involving in EH and further to explore the molecular connectivities between these pathways by the topological analysis of the Protein-protein interaction (PPI) network. RESULT: The extended network including one giant network consisted of 535 nodes connected via 2572 edges and two separated small networks. 27 proteins with high BC and 28 proteins with large degree have been identified. NOS3 with highest BC and Closeness centrality located in the centre of the network. The backbone network derived from high BC proteins presents a clear and visual overview which shows all important regulatory pathways for blood pressure (BP) and the crosstalk between them. Finally, the robustness of NOS3 as central protein and accuracy of backbone were validated by 287 test networks. CONCLUSION: Our finding suggests that blood pressure variation is orchestrated by an integrated PPI network centered on NOS3.

[Study on the indeterminate results of characterization and verification of HIV antibody from Western blot test].

OBJECTIVE: To study the serological characterization of indeterminate Western blot (WB) results of HIV antibody and to find a new way to verify the HIV antibody indeterminate results and provide references for editing "National Guideline for Detection of HIV/AIDS". METHODS: All of the 42 subjects who were confirmed as indeterminate HIV antibody in People' Liberation Army HIV Confirmation Laboratory from 2005 to 2006, were collected. Line immunoassay, HIV viral load test and HIV-1 p24 were tested and followed up for 3-6 months' to compare the changes of WB bands patterns. RESULTS: (1) For the 42 individuals with indeterminate HIV antibody, a total of 8 different patterns of bands were found in WB test including 45.2% of them were p24 mono band, 30.9% were gp160 mono band, 11.9% were gp160 with p24, 2.4% (only one case) were gp160gp120 +/-, gp41p24, p24p17, gp41 or gp120 respectively. It was noticed that the most patterns of common bands with indeterminate results were p24 mono band, gp160 mono band and gp160 with p24, which composed 88.0% of the whole indeterminate WB band patterns. (2) Twenty three cases had been followed up for more than 3 months with 22 giving no WB band image change and were confirmed as HIV sero-negative. The other one with case gp160 and p24 had developed to more bands in the period of 77 days follow-up with more bands, including gp160, gp120, p66, p31, p24 and p17,showed up and was confirmed as HIV primary infection. (3) Line immunoassay was applied to all of those 23 cases who had been followed up and the results showed that only one serological change was found and the case was confirmed to be HIV-positive. Among the other 22 cases without serological changes, 16 cases were proved to be HIV-negative, 6 cases were still indeterminate. The specificity was 72.7%. P24 antigen test showed negative in all the 23 cases, including the case which later was confirmed as HIV-positive. Of all the 23 originally indeterminate cases, viral loads were tested in 7 cases. Positive result was found in the case which was proved later to be HIV-positive. No viral loads were detected in the other 6 cases (< LDL). CONCLUSION: The most common band patterns of indeterminate HIV antibody were mainly p24 monoband, gp160 monoband or with p24. Most of them (95.6%) were not infected by HIV, the bands showed up in WB test and demonstrated as non-specific reactions. Line immunoassay could determine about 70% of the indeterminate reactions. Results from viral load test also suggested that it was an efficient method to discriminate indeterminate results. With these two techniques, HIV serology could be diagnosed without 3 months' follow-up in primary infection which gave indeterminate WB results.

[Inhibition of L-arginine and cilostazol on activation of platelets in vitro].

The purpose of study was to investigate the effects of L-arginine and cilostazol on platelet-activation and aggregation reserve in vitro, so as to provide proof for selecting reversible activation-inhibitors for platelets lyophilization. Activation and function of platelets were investigated by using flow cytometry with the CD62p and PAC-1 expression and re-expression after being activated by thrombin, and by means of platelet aggregation reaction to thrombin, ADP and propyl gallate, as well as coagulation activity of platelets. The results showed that expression of CD62p and PAC-1 increased after being pretreated. Both L-arginie and cilostazol could inhibit CD62p and PAC-1 expression and related with their concentrations. Cilostazol had an intensive inhibition effect on expressions of CD62p and PAC-1 induced by thrombin, and the inhibition increased when concentration augmented. L-arginine had the same effects on PAC-1, but had no effects on CD62p. L-arginine and cilostazol inhibited aggregation induced by thrombin, ADP and propyl gallate, and the inhibitions were related directly with dosage. When L-arginine concentration was higher or equal to 15 mmol/L, or cilostazol concentration was in range of 1 - 4 mmol/L, the aggregation time were prolonged so much or even no aggregation. It is concluded that when L-Arginine concentration is 5 mmol/L and 10 mmol/L, platelet activation can be inhibited, but aggregation ability and characters keep intact. Concentration at 5 mmol/L may be the best. 1 mmol/L of cilostazol can inhibit activation in vitro and retain part of platelet ability of aggregation and reexpression.

[Experimental study on lyophilization of platelets].

The aim of this study was to search a procedure of platelet lyophilization and find a way of long-term storage of human platelets at normal temperature with smaller size and lighter weight, to be convenient to transport at long distance thus to meet the demands in accidents and war time. Human platelets were pretreated by freezing, the first and the second desiccation, and were added with reversible activation-inhibitors of platelets, DMSO and trehalose, then were rehydrated. At the same time, the recovery rate of platelets, platelet maximal aggregation induced by thrombin, coagulation of platelets, CD62p expression and PAC-1 expression were assayed. The results indicated that the recovery rate of the platelets was 56.29%. The platelet maximal aggregation induced by thrombin had no significant difference between lyophilized platelets and the fresh platelet-rich plasma (FPRP), but the aggregation of platelets induced by ADP or propyl gallate was decreased by 49.34% and 26.25%. Coagulation of the lyophilized platelets was not significantly different from FPRP. CD62p expression of the lyophilized platelets (42.36%) was higher than that in FPRP while PAC-1 expression was 2.12%. CD62p re-expression rate induced by thrombin was 50.88% and PAC-1 re-expression was 54.55%. It is concluded that the ability of recovered lyophilized platelets added with reversible activation-inhibitors, DMSO and trehalose to aggregate and coagulate has showed no significant difference as compared with FPRP. The reversible activation-inhibitors can decrease CD62p expression of lyophilized platelets, and may enhance their survival ability and prolongate survival time. Therefore the efficiency of lyophilizing platelets can be improved based on this freeze-drying procedure.