Network Pharmacology-Based Analysis on the Potential Biological Mechanisms of Sinisan Against Non-Alcoholic Fatty Liver Disease.

Non-alcoholic fatty liver disease (NAFLD) has become the most prevalent liver disease in China. Sinisan (SNS) is a traditional Chinese medicine formula that has been widely used in treating chronic liver diseases, including NAFLD. However, its underlying biological mechanisms are still unclear. In this study, we employed a network pharmacology approach consisting of overlapped terms- (genes or pathway terms-) based analysis, protein-protein interaction (PPI) network-based analysis, and PPI clusters identification. Unlike the previous network pharmacology study, we used the shortest path length-based network proximity algorithm to evaluate the efficacy of SNS against NAFLD. And we also used random walk with restart (RWR) algorithm and Community Cluster (Glay) algorithm to identify important targets and clusters. The screening results showed that the mean shortest path length between genes of SNS and NAFLD was significantly smaller than degree-matched random ones. Six PPI clusters were identified and ten hub targets were obtained, including STAT3, CTNNB1, MAPK1, MAPK3, AGT, NQO1, TOP2A, FDFT1, ALDH4A1, and KCNH2. The experimental study indicated that SNS reduced hyperlipidemia, liver steatosis, and inflammation. Most importantly, JAK2/STAT3 signal was inhibited by SNS treatment and was recognized as the most important signal considering the network pharmacology part. This study provides a systems perspective to study the relationship between Chinese medicines and diseases and helps to discover potential mechanisms by which SNS ameliorates NAFLD.

A network pharmacology study on analgesic mechanism of Yuanhu-Baizhi herb pair.

BACKGROUND: Millions of people are suffering from chronic pain conditions, such as headache, arthritis, cancer. Apart from western medicines, traditional Chinese medicines are also well accepted for pain management, especially in Asian countries. Yuanhu-Baizhi herb pair (YB) is a typical herb pair applied to the treatment of stomach pain, hypochondriac pain, headache, and dysmenorrhea, due to its effects on analgesia and sedation. This study is to identify potentially active compounds and the underlying mechanisms of YB in the treatment of pain. METHODS: Compounds in YB were collected from 3 online databases and then screened by bioavailability and drug likeness parameters. Swiss target prediction was applied to obtain targets information of the active compounds. Pain-related genes were conducted for Gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis. Protein-protein interaction (PPI) networks of the genes were constructed using Cytoscape software. In addition, the hub genes were screened using maximal clique centrality (MCC) algorithm. RESULTS: In total, 31 compounds from Yuanhu were screened out with 35 putative target genes, while 26 compounds in Baizhi with 43 target genes were discovered. Hence, 78 potential target genes of YB were selected for further study. After overlap analysis of the 78 genes of YB and 2408 pain-associated genes, we finally achieved 34 YB-pain target genes, as well as 10 hub genes and 23 core compounds. Go enrichment and KEGG pathway analysis indicated that YB had a strong integration with neuro system, which might significantly contribute to antinociceptive effect. CONCLUSION: Our data provide deep understanding of the pharmacological mechanisms of YB in attenuating pain. The discovery shed new light on the development of active compounds of YB for the treatment of pain.

Development of an Enzyme-Linked Immunosorbent Assay and Immunoaffinity Column Chromatography for Saikosaponin d Using an Anti-Saikosaponin d Monoclonal Antibody.

This work developed a novel immunochemical approach for the quality control of saikosaponin d using an enzyme-linked immunosorbent assay. Splenocytes from mice immunized with the saikosaponin d-bovine serum albumin conjugate were fused with the hypoxanthine-aminopterin-thymidine-sensitive mouse myeloma SP2/0 cell line, and a hybridoma secreting monoclonal antibody against saikosaponin d was successfully obtained. The prepared anti-saikosaponin d monoclonal antibody 1E7F3 has a novel characteristic, showing weak reactivity with compounds that are structurally related to saikosaponin d. Using monoclonal antibody 1E7F3, a specific and reliable enzyme-linked immunosorbent assay was developed to detect saikosaponin d. The system shows a full measurement range from 156.25 to 5000.00 ng × mL(-1). Both intra-assay and inter-assay repeatability and precision were achieved, with relative standard deviations lower than 10.00%. The recovery rates ranged from 92.36% to 101.00%, meeting the requirements for biological samples. There was a good correlation between the enzyme-linked immunosorbent assay and high-performance liquid chromatography analyses of saikosaponin d, and the saikosaponin d levels in formulated Chinese medicines were successfully determined. Furthermore, immunoaffinity column chromatography was established using this anti-saikosaponin d monoclonal antibody, and the elution profile of saikosaponin d was detected by a Bio-Rad QuadTec UV/Vis detector at 203 nm. The results demonstrate that we generated a reliable and more efficient assay system for measuring saikosaponin d and provide a potential approach for purifying and separating saikosaponin d.

Development of a Fluorescence-Linked Immunosorbent Assay for Baicalin.

Previously, we developed an indirect competitive enzyme-linked immunosorbent assay (icELISA) for baicalin (BAL) and used this assay to investigate the pharmacokinetics of BAL in mice. In this study, an anti-BAL monoclonal antibody (MAb) was purified by the caprylic acid method and then labelled with fluorescein isothiocyanate (FITC). Subsequently, an indirect competitive fluorescence-linked immunosorbent assay (icFLISA) was developed to detect baicalin (BAL) using FITC-labelled anti-BAL MAbs. Characterization of the assay demonstrated an effective BAL measurement range of 6.4 ng/mL to 500 µg/mL (R(2) = 0.997). The relative standard deviations (RSDs) for both intra-assay and inter-assay repeatability and precision were below 10 %. This icFLISA for BAL is simple, rapid and sensitive, with a 390-fold larger linear range and a 2-fold lower limit of detection (LOD) compared with the previously developed icELISA. We observed a strong correlation between the results determined by the icFLISA and icELISA methods. Overall, this study provides a useful method for detecting BAL in medicines, enabling in vivo visualization research.

Establishment of an enzyme-linked immunosorbent assay and application on determination of ginsenoside Re in human saliva.

This work describes an immunochemical approach for the quality control of Panax ginseng and a pharmacological study of ginsenoside Re, a major bioactive constituent in P. ginseng, using an enzyme-linked immunosorbent assay. A hybridoma secreting monoclonal antibody against ginsenoside Re was produced by fusing splenocytes immunized with a ginsenoside Re-bovine serum albumin conjugate with the hypoxanthine-aminopterin-thymidine-sensitive mouse myeloma SP2/0 cell line. The method, at an effective measuring range of 7.8-500 ng ·â€ŠmL(-1) of ginsenoside Re, successfully detected ginsenoside Re in Chinese traditional herb prescriptions. The results demonstrate that we generated a novel and reliable assay system for measuring ginsenoside Re in Chinese medicines more efficiently. Futhermore, we determined the ginsenoside Re concentrations in the saliva of six healthy adults after the oral administration of a ginseng capsule to study the pharmacokinetics of ginsenoside Re in human saliva.