[Effect of Erjing Pills on alleviating neuroinflammation of AD rats based on TLR4/NF-κB/NLRP3 pathway and its mechanism].

This paper aimed to study the effect of Erjing Pills on the improvement of neuroinflammation of rats with Alzheimer's di-sease(AD) induced by the combination of D-galactose and Aß_(25-35) and its mechanism. SD rats were randomly divided into a sham group, a model control group, a positive drug group(donepezil, 1 mg·kg~(-1)), an Erjing Pills high-dose group(9.0 g·kg~(-1)), and an Erjing Pills low-dose group(4.5 g·kg~(-1)), with 14 rats each group. To establish the rat model of AD, Erjing Pills were intragastrically administrated to rats for 5 weeks after 2 weeks of D-galactose injection. D-galactose was intraperitoneally injected into rats for 3 weeks, and then Aß_(25-35) was injected into the bilateral hippocampus. The new object recognition test was used to evaluate the learning and memory ability of rats after 4 weeks of intragastric administration. Tissues were acquired 24 h after the last administration. The immunofluorescence method was used to detect the activation of microglia in the brain tissue of rats. The positive expressions of Aß_(1-42) and phosphory protein Tau~(404)(p-Tau~(404)) in the CA1 area of the hippocampus were detected by immunohistochemistry. The levels of inflammatory factors interleukin-1ß(IL-1ß), tumor necrosis factor-α(TNF-α), and interleukin-6(IL-6) in the brain tissue were determined by enzyme-linked immunosorbent assay(ELISA). Toll-like receptor 4(TLR4)/nuclear factor kappa B(NF-κB)/nucleotide-binding oligomerization domain-like receptors 3(NLRP3) pathway-associated proteins in the brain tissue were determined by Western blot. The results showed that as compared with the sham group, the new object recognition index of rats in the model control group decreased significantly, the deposition of Aß_(1-42) and p-Tau~(404) positive protein in the hippocampus increased significantly, and the levels of microglia activation increased significantly in the dentate gyrus. The levels of IL-1ß, TNF-α, and IL-6 in the hippocampus of the model control group increased significantly, and the expression levels of TLR4, p-NF-κB p65/NF-κB p65, p-IκBα/IκBα, and NLRP3 proteins in the hippocampus increased significantly. Compared with the model control group, the Erjing Pill groups enhanced the new object recognition index of rats, decreased the deposition of Aß_(1-42) and the expression of p-Tau~(404) positive protein in the hippocampus, inhibited the activation of microglia in the dentate gyrus, reduced the levels of inflammatory factors IL-1ß, TNF-α, and IL-6 in the hippocampus, and down-regulated the expression levels of TLR4, p-NF-κB P65/NF-κB P65, p-IκBα/IκBα, and NLRP3 proteins in the hippocampus. In conclusion, Erjing Pills can improve the learning and memory ability of the rat model of AD presumably by improving the activation of microglia, reducing the expression levels of neuroinflammatory factors IL-1ß, TNF-α, and IL-6, inhibiting the TLR4/NF-κB/NLRP3 neuroinflammation pathway, and decreasing hippocampal deposition of Aß and expression of p-Tau, thereby restoring the hippocampal morphological structure.

Tubeimoside I Ameliorates Myocardial Ischemia-Reperfusion Injury through SIRT3-Dependent Regulation of Oxidative Stress and Apoptosis.

Myocardial ischemia-reperfusion injury (MIRI) is a phenomenon that reperfusion leads to irreversible damage to the myocardium and increases mortality in acute myocardial infarction (AMI) patients. There is no effective drug to treat MIRI. Tubeimoside I (TBM) is a triterpenoid saponin purified from Chinese traditional medicine tubeimu. In this study, 4 mg/kg TBM was given to mice intraperitoneally at 15 min after ischemia. And TBM treatment improved postischemic cardiac function, decreased infarct size, diminished lactate dehydrogenase release, ameliorated oxidative stress, and reduced apoptotic index. Notably, ischemia-reperfusion induced a significant decrease in cardiac SIRT3 expression and activity, while TBM treatment upregulated SIRT3's expression and activity. However, the cardioprotective effects of TBM were largely abolished by a SIRT3 inhibitor 3-(1H-1,2,3-triazol-4-yl) pyridine (3-TYP). This suggests that SIRT3 plays an essential role in TBM's cardioprotective effects. In vitro, TBM also protected H9c2 cells against simulated ischemia/reperfusion (SIR) injury by attenuating oxidative stress and apoptosis, and siSIRT3 diminished its protective effects. Taken together, our results demonstrate for the first time that TBM protects against MIRI through SIRT3-dependent regulation of oxidative stress and apoptosis. TBM might be a potential drug candidate for MIRI treatment.

Evidence-based complementary and alternative medicine bioinformatics approach through network pharmacology and molecular docking to determine the molecular mechanisms of Erjing pill in Alzheimer's disease.

Erjing pill, a Traditional Chinese Medicine (TCM) formulation composed of Polygonatum sibiricum and Lycium chinense, has an important role in the treatment of Alzheimer's disease (AD). However, the underlying mechanisms of the action of Erjing pill in AD have remained elusive. In the present study, the key ingredients of Erjing pill were investigated and the active components and their mechanisms of action on AD were analyzed based on networks pharmacology. By using the TCM and TCM Systems Pharmacology and databases, the components of Erjing pill were screened and the data were captured using Discovery Studio. The SwissTarget webserver database was used to predict the potential protein targets of Erjing pill components for pathologies related to AD. The data were further analyzed with the disease targets of AD based on analysis of the Online Mendelian Inheritance in Man, DiGSeE and Therapeutic Target Database. Subsequent analysis of mechanistic pathways of the screened components and protein targets allowed us to construct a network by using Gene Ontology and Kyoto Encyclopedia of Genes and Genomes, which revealed potential molecular mechanisms of Erjing pill against AD. Finally, the protective effect of three active components on neurons was verified using an in vitro injury model of PC12 cells induced by Aß25-35. The results indicated that 65 bioactive components of Erjing pill, including lauric acid and zederone, and 6 targets, including acetylcholinesterase, butylcholinesterase and amyloid protein precursor, were closely associated with the prevention and treatment of AD. The molecular components of Erjing pill were indicated to be involved in various biological signaling processes, mainly in synaptic signal transmission, transsynaptic signal transmission and chemical synaptic transmission. Furthermore, related pathways targeted by Erjing pill in AD included the regulation of neuroactive ligand-receptor interactions, the PI3K-Akt signaling pathway, serotoninergic synapses, calcium signaling pathways and dopaminergic synapses. A cell viability assay indicated that the compounds (polygonatine A, polygonatine C and 4',5-dihydroxyflavone) assessed were able to significantly improve the survival rate and increase the Ca2+ level in a PC12 cell model of AD induced by amyloid-ß25-35. The present study revealed that the mechanisms of action of Erjing pill to prevent and treat AD included a multicompound, multitarget and multipathway regulatory network.

Tubeimoside I promotes angiogenesis via activation of eNOS-VEGF signaling pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: Tubeimoside I (TBM) is a triterpenoid saponin purified from tubeimu (tuber of Bolbostemma paniculatum (Maxim.) Franquet). In traditional Chinese medicine, tubeimu had been used to treat acute mastitis, snake bites, detoxication, inflammatory diseases, and tumors for over 1000 years. AIM OF THE STUDY: This study aimed to investigate whether TBM could promote angiogenesis and how to promote angiogenesis. MATERIALS AND METHODS: In vivo, the pro-angiogenic effects of TBM were examined using the hindlimb ischemia model. After the ischemia operation, 1 mg/kg/day TBM was given via intraperitoneal injection for 28 days and the recovery of blood flow was monitored by Doppler scanner every 7 days. The capillary density in gastrocnemius muscle was detected by immunofluorescence. Expression of related proteins were determined by western blotting. In vitro, the pro-angiogenic effects of TBM on HUVECs were examined by Cell Counting Kit-8, scratch assay, endothelial cell tube formation assay and western blotting. RESULTS: TBM improved recovery from hindlimb ischemia in C57BL/6 mice. TBM promoted endothelial cell viability, migration and tube formation in HUVECs. TBM could activate eNOS-VEGF signaling pathway by enhancing expression of eNOS. And TBM's pro-angiogenesis effects could be abolished by L-NAME (an inhibitor of eNOS). CONCLUSIONS: TBM promoted angiogenesis via the activation of eNOS-VEGF signaling pathway and TBM could be a novel agent for therapeutic angiogenesis in ischemic diseases.

Network pharmacology-based strategy to investigate pharmacological mechanisms of Tinospora sinensis for treatment of Alzheimer's disease.

ETHNOPHARMACOLOGICAL RELEVANCE: Tinospora sinensis (Lour.) Merr. belongs to the family Menispermaceae. It is called LeZhe and is widely used as a kind of folk medicine especially in the Tibetan Plateau of China. T. sinensis has the functions of clearing away heat and detoxification, dispelling wind and dredging collaterals, calming and soothing the nerves. T. sinensis is an effective medicine for the prevention and treatment of aging diseases such as Alzheimer's disease (AD) in the Tibetan Plateau of China, whereas its material basis and underlying mechanisms are not clear. The aim of this study was to investigate the material basis and potential mechanisms of T. sinensis in the treatment of AD by using network pharmacology and molecular docking. MATERIALS AND METHODS: In this study, targets were collected from DrugBank database, Therapeutic Target Database (TTD) and literatures reports for the treatment of AD. Compounds were searched by literatures and systematic separation from T. sinensis. The molecular docking experiment was carried out by using Autodock Vina software to screen the bioactive compounds in T. sinensis and target proteins for AD. Then, the "compound-target network" was constructed by Cytoscape software. The drug-like properties of the active compounds were analyzed by pKCSM performs, and the protein-protein interaction (PPI) network was constructed by Search Tool for the Retrieval of Interacting Genes/Proteins (STRING). The Kyoto Encyclopedia of Genes and Genomes (KEGG) target pathway enrichment analysis was carried out by Database for Annotation, Visualization and Integrated Discovery (DAVID). Furthermore, the protective effect of neurons of two active compounds were verified with the injury cell model of PC12 and primary hippocampus neurons induced by Aß25-35. Finally, the key proteins of related pathways were quantitatively analyzed with Western blot method. RESULTS: In total, 105 compounds and 38 targets have been screened. The main active compounds contained berberine, which belongs to alkaloids, Aurantiamide acetate, N-P-coumaroyltyramine, which belongs to amides, Trans-syringin and 3-demethyl-phillyrin, which belongs to phenylpropanoids. The targets covered inflammation-related proteins, including Protein kinase B (AKT), Phosphoinositide 3-kinase (PI3K), Tyrosine-protein kinase JAK1 (JAK1), mammalian target of rapamycin (mTOR), tumor necrosis factor alpha (TNF-α), Neuronal NOS (NOS1), and cholinergic function-related proteins, including α4-Nicotinic acetylcholine receptor (α4 nAChR), Muscarinic acetylcholine receptor M1 (Muscarnic M1). Inflammation and cholinergic dysfunction were the center of the network and occupy a dominant position. And the results of enrichment analysis shown the pathways mainly contained phosphoinositide-3-kinase/Akt (PI3K/Akt) signal pathway, neurotrophic factors (NTFs) signal pathway, Hypoxia-inducible factor 1 (HIF-1) signal pathway, mechanistic Target of Rapamycin (mTOR) signal pathway, Tumor necrosis factor (TNF) signal pathway, insulin resistance (IR). The results of in vitro assays showed that the tested compounds could significantly improve the survival rate and inhibit the apoptosis of PC12 cells and primary hippocampal neurons injured by Aß25-35. Western blot results showed that T. sinensis had a significant effect on the expression of protein PI3K and Akt. CONCLUSION: Our results revealed that T. sinensis could prevent and treat AD through a multi-compound-multi-target-multi-pathway regulatory network. Our work also expected to provide new ideas and theoretical bases for searching for the active compounds in T. sinensis and potential mechanism in the prevention and treatment of AD by the network pharmacology and molecular docking. The results of in vitro assay and in vivo assay supported the results of molecular docking.

Tubeimoside I improves survival of mice in sepsis by inhibiting inducible nitric oxide synthase expression.

Sepsis is a disease with high mortality rate worldwide and inducible nitric oxide (iNOS) induced vascular hyporeactivity plays a key role in it. There is no effective drug to treat vascular hyporeactivity specifically. Tubeimoside I (TBM) is a triterpenoid saponin isolated from Rhizoma Bolbostemmatis. In this study, we found that 4 mg/kg TBM intraperitoneally injected 1 h before cecal ligation and puncture (CLP) partially improved survival, ameliorated mean arterial pressure (MAP) and enhanced vascular responsiveness to norepinephrine (NE) and KCl in wild-type septic mice. CLP activated TLR4-MyD88-NF-κB-iNOS pathway was also inhibited by TBM both in vitro and in vivo. However, iNOS gene knockout counteracted the protection provided by TBM. We conclude that TBM protects mice in sepsis by reducing excessive NO production through inhibiting the TLR4-MyD88-NF-κB-iNOS pathway. Our study suggests a possible therapeutic application of TBM in sepsis.