Rheum palmatum L. and Salvia miltiorrhiza Bge. Alleviates Acute Pancreatitis by Regulating Th17 Cell Differentiation: An Integrated Network Pharmacology Analysis, Molecular Dynamics Simulation and Experimental Validation.

OBJECTIVE: To identify the core targets of Rheum palmatum L. and Salvia miltiorrhiza Bge., (Dahuang-Danshen, DH-DS) and the mechanism underlying its therapeutic efficacy in acute pancreatitis (AP) using a network pharmacology approach and validate the findings in animal experiments. METHODS: Network pharmacology analysis was used to elucidate the mechanisms underlying the therapeutic effects of DH-DS in AP. The reliability of the results was verified by molecular docking simulation and molecular dynamics simulation. Finally, the results of network pharmacology enrichment analysis were verified by immunohistochemistry, Western blot analysis and real-time quantitative PCR, respectively. RESULTS: Sixty-seven common targets of DH-DS in AP were identified and mitogen-activated protein kinase 3 (MAPK3), Janus kinase 2 (JAK2), signal transducer and activator of transcription 3 (STAT3), protein c-Fos (FOS) were identified as core targets in the protein interaction (PPI) network analysis. Gene ontology analysis showed that cellular response to organic substance was the main functions of DH-DS in AP, and Kyoto Encyclopedia of Genes and Genomes analysis showed that the main pathway included Th17 cell differentiation. Molecular docking simulation confirmed that DH-DS binds with strong affinity to MAPK3, STAT3 and FOS. Molecular dynamics simulation revealed that FOS-isotanshinone II and STAT3-dan-shexinkum d had good binding capacity. Animal experiments indicated that compared with the AP model group, DH-DS treatment effectively alleviated AP by inhibiting the expression of interleukin-1ß, interleukin-6 and tumor necrosis factor-α, and blocking the activation of Th17 cell differentiation (P<0.01). CONCLUSION: DH-DS could inhibit the expression of inflammatory factors and protect pancreatic tissues, which would be functioned by regulating Th17 cell differentiation-related mRNA and protein expressions.

Metabolite profiling analysis of hepatitis B virus-induced liver cirrhosis patients with minimal hepatic encephalopathy using gas chromatography-time-of-flight mass spectrometry and ultra-performance liquid chromatography-quadrupole-time-of-flight mass spectrometry.

This study used gas chromatography-time-of-flight mass spectrometry (GC-TOFMS) and ultra-performance liquid chromatography-quadrupole TOFMS (UPLC-QTOFMS) metabonomic analytical techniques in combination with bioinformatics and pattern recognition analysis methods to analyze the serum metabolite profiling of hepatitis B virus (HBV)-induced liver cirrhosis patients with minimal hepatic encephalopathy (MHE), to find the specific biomarkers of MHE, to reveal the pathogenesis of MHE, and to determine a promising approach for early diagnosis of MHE. Serum samples of 100 normal controls (NC group), 29 HBV-induced liver cirrhosis patients with MHE (MHE group), and 24 HBV-induced liver cirrhosis patients without MHE [comprising 12 cases of compensated cirrhosis (CS group) and 12 cases of decompensated cirrhosis (DS group)] were collected and employed into GC-TOFMS and UPLC-QTOFMS platforms for serum metabolite detection; the outcome data were then analyzed using principal component analysis and orthogonal partial least squares-discriminant analysis (OPLS-DA). There were no significant differential metabolites between the NC group and the CS group. A series of key differential metabolites were detected. According to the variable influence in projection values and P-values, 60 small-molecule metabolites were considered to be dysregulated in the MHE group (compared to the NC group); 27 of these 60 dysregulated differential metabolites were considered to be the potential biomarkers (see Table 4, marked in bold); 66 small-molecule metabolites were considered to be dysregulated in the DS group (compared to the NC group); 34 of these 66 dysregulated differential metabolites were considered to be the potential biomarkers (see Table 5, marked in bold). According to the fold-change values, 9 of these 27 metabolites, namely valine, oxalic acid, erythro-sphingosine, 4,7,10,13,16,19-docosahexaenoic acid, isoleucine, allo-isoleucine, thyroxine, rac-octanoyl carnitine, and tocopherol (vitamin E), were downregulated in the MHE group (compared to the NC group); the other 18, namely adenine, glycochenodeoxycholic acid, fucose, allothreonine, glycohyocholic acid, glycoursodeoxycholic acid, tyrosine, taurocheno-deoxycholate, phenylalanine, 2-hydroxy-3-methyl-butanoic acid, hydroxyacetic acid, taurocholate, sorbitol, rhamnose, tauroursodeoxycholate, tolbutamide, pyroglutamic acid, and malic acid, were upregulated; 6 of these 34 metabolites were downregulated in the DS group (compared to the NC group), and the other 28 were upregulated, as shown in Table 5. (a) GC-TOFMS and UPLC-QTOFMS metabonomic analytical platforms can detect a range of metabolites in the serum; this might be of great help to study the pathogenesis of MHE and may provide a new approach for the early diagnosis of MHE. (b) Metabonomics analysis in combination with pattern recognition analysis might have great potential to distinguish the HBV-induced liver cirrhosis patients who have MHE from the normal healthy population and HBV-induced liver cirrhosis patients without MHE.

Spinal dI4 Interneuron Differentiation From Human Pluripotent Stem Cells.

Spinal interneurons (INs) form intricate local networks in the spinal cord and regulate not only the ascending and descending nerve transduction but also the central pattern generator function. They are therefore potential therapeutic targets in spinal cord injury and diseases. In this study, we devised a reproducible protocol to differentiate human pluripotent stem cells (hPSCs) from enriched spinal dI4 inhibitory GABAergic INs. The protocol is designed based on developmental principles and optimized by using small molecules to maximize its reproducibility. The protocol comprises induction of neuroepithelia, patterning of neuroepithelia to dorsal spinal progenitors, expansion of the progenitors in suspension, and finally differentiation into mature neurons. In particular, we employed both morphogen activators and inhibitors to restrict or "squeeze" the progenitor fate during the stage of neural patterning. We use retinoic acid (RA) which ventralizes cells up to the mid-dorsal region, with cyclopamine (CYC), an SHH inhibitor, to antagonize the ventralization effect of RA, yielding highly enriched dI4 progenitors (90% Ptf1a+, 90.7% Ascl1+). The ability to generate enriched spinal dI4 GABAergicINs will likely facilitate the study of human spinal IN development and regenerative therapies for traumatic injuries and diseases of the spinal cord.