Exploring the mechanism of MP gel against skin photoaging based on network pharmacology, molecular docking, and experimental validation.

OBJECTIVE: Long-term and high exposure to UV radiation can lead to the development of skin photoaging diseases. Therefore, there is an ongoing need for more natural and safe drugs to prevent or treat skin photoaging diseases. METHODS: The Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform database were used to collect the active compounds and corresponding targets of Cnidii Fructus, Arnebiae Radix, Angelicae Sinensis Radix, Poria, and Borneolum. The GeneCards database and the NCBI Gene database were used to collect the targets of skin photoaging diseases. The STRING database was used to construct a protein-protein interaction network formed by the intersecting targets of drugs and diseases. The Metascape database was applied for Gene Ontology and Kyoto Encyclopedia of Genes and Genomes pathway enrichment analysis of the targets. Molecular docking between active compounds and targets was verified by Autodock. After that, the skin photoaging model of mice was established and treated with MP gel. The skin characterization on the back of mice was observed, and the ameliorative effect of MP gel on skin photoaging was evaluated by histological and epidermal thickness assays. The MDA content and SOD activity were measured. Caspase-3 expression in mouse skin tissues was detected by immunohistochemistry, quantitative real-time polymerase chain reaction assay, and Western blot. RESULTS: The results of network pharmacology experiments showed that the natural drugs have multi-component, multi-target therapeutic disease characteristics. The results of animal studies showed that MP gel improved the health of photoaged skin, promoted skin structural integrity, had antioxidant properties and significantly inhibited caspase-3 expression. CONCLUSION: The experimental validation of the results of the preliminary network pharmacology analysis was carried out in animal experiments, which confirmed part of the mechanism of action of MP gel in the prevention and treatment of skin photoaging.

Piperine treating sciatica through regulating inflammation and MiR-520a/P65 pathway.

Although the etiology of sciatica remains uncertain, there is increasing evidence that the disease process of sciatica is associated with the levels of inflammatory factors. Piperine, an alkaloid isolated from Piper nigrum, has previously been demonstrated to inhibit inflammation and analgesic effects. The purpose of this study is to verify the regulatory relationship between miR-520a and p65 and to explore how miR-520a/P65 affects the level of cytokines under the action of piperine, so as to play a therapeutic role in sciatica. Through ELISA experiment, we confirmed that four inflammatory factors (IL-1ß, TNF-α, IL-10, TGF-ß1) can be used as evaluation indexes of sciatica. The differentially expressed miRNA was screened as miR-520a, by microarray technology, and the downstream target of miR-520a was P65 by bioinformatics. Real-time fluorescence quantitative PCR confirmed that the expression of miR-520a was negatively correlated with pro-inflammatory cytokines, positively correlated with anti-inflammatory cytokines and negatively correlated with p65 expression at mRNA level. The expression of p65 was positively correlated with pro-inflammatory cytokines and negatively correlated with anti-inflammatory cytokines at the protein level verified by ELISA and Western blot. HE staining analysis showed that the nerve fibers were repaired by piprine, the vacuoles were significantly reduced, and the degree of nerve fiber damage was also improved. Immunohistochemical analysis showed that the expression of p65 decreased after administration of piperine. Dual-luciferase reporter gene assay confirmed that the luciferase signal decreased significantly after cotransfection of miR-520a mimics and p65 3'UTR recombinant plasmid. To sum up, in the rat model of non-compressed lumbar disc herniation, piperine plays a significant role in analgesia. MiR-520a can specifically and directly target P65, and piperine can promote the expression of miR-520a, then inhibit the expression of p65, down-regulate the pro-inflammatory factors IL-1ß and TNF-α, and up-regulate the effects of anti-inflammatory factors IL-10 and TGF-ß1, so as to treat sciatica.

Interaction between piperine and genes associated with sciatica and its mechanism based on molecular docking technology and network pharmacology.

Piperine is the main active component of Piper longum L., which is also the main component of anti-sciatica Mongolian medicine Naru Sanwei pill. It has many pharmacological activities such as anti-inflammatory and immune regulation. This paper aims to preliminarily explore the potential mechanism of piperine in the treatment of sciatica through network pharmacology and molecular docking. TCMSP, ETCM database and literature mining were used to collect the active compounds of Piper longum L. Swiss TargetPrediction and SuperPred server were used to find the targets of compounds. At the same time, CTD database was used to collect the targets of sciatica. Then the above targets were compared and analyzed to select the targets of anti-sciatica in Piper longum L. The Go (gene ontology) annotation and KEGG pathway of the targets were enriched and analyzed by Metascape database platform. The molecular docking between the effective components and the targets was verified by Autodock. After that, the sciatica model of rats was established and treated with piperine. The expression level of inflammatory factors and proteins in the serum and tissues of rat sciatic nerve were detected by ELISA and Western blot. HE staining and immunohistochemistry were carried out on the sciatica tissues of rats. The results showed that Piper longum L. can regulate the development of sciatica and affect the expressions of PPARG and NF-kB1 through its active ingredient piperine, and there is endogenous interaction between PPARG and NF-kB1.

The mechanism underlying hypaconitine-mediated alleviation of pancreatitis-associated lung injury through up-regulating aquaporin-1/TNF-α.

BACKGROUND/AIMS: Acute pancreatitis-associated lung injury (APALI) is one of the most common and most dangerous form of extra-pancreatic organ damage in severe acute pancreatitis (SAP). The treatment options for SAP were limited thus far; as a result, approximately 60%-80% of patients with SAP would die within a week. Hypaconitine (HC), one of the most important active ingredients in a Mongolian traditional medicine Radix Aconiti Kusnezoffii has an excellent anti-inflammatory effect. MATERIALS AND METHODS: To ascertain whether HC has a protective effect against APALI, we investigated the therapeutic effects and the underlying mechanisms in vivo and in vitro and attempted to elucidate the mechanism in detail. In this study, APALI rats and human pulmonary microvascular endothelial cells were treated with therapeutic doses of HC after establishing a model with sodium taurocholate and lipopolysaccharide, respectively. RESULTS: Serum amylase and lipase activity, lung wet/dry weight ratio, lung myeloperoxidase activity, and pancreatic and lung histopathological changes showed that HC alleviated APALI in a dose-dependent way, which can be abolished by an aquaporin-1 (AQP-1) knockdown. The results of the reverse transcriptase polymerase chain reaction, Western blot, and immunohistochemical staining confirmed the expression of AQP-1, a kind of transmembrane protein that mainly distributed in the membranes of pulmonary cells and contributed to maintain water balance in the body by interacting with tumor necrosis factor-alpha (TNF-α), is negatively associated with APALI. On the contrary, HC treatment up-regulated AQP-1 expression and down-regulated the TNF-α expression as a consequence in APALI. CONCLUSION: These results suggest that HC has a good anti-inflammatory therapeutic effect on APALI with a possible underlying mechanism that affects the AQP-1/TNF-α pathway.

Exploring the active compounds of traditional Mongolian medicine in intervention of novel coronavirus (COVID-19) based on molecular docking method.

OBJECTIVE: This article intends to use molecular docking technology to find potential inhibitors that can respond to COVID-19 from active compounds in Mongolian medicine. METHODS: Mongolian medicine with anti-inflammatory and antiviral effects is selected from Mongolian medicine prescription preparations. TCMSP, ETCM database and document mining methods were used to collect active compounds. Swiss TargetPrediction and SuperPred server were used to find targets of compounds with smiles number. Drugbank and Genecard database were used to collect antiviral drug targets. Then the above targets were compared and analyzed to screen out antiviral targets of Mongolia medicine. Metascape database platform was used to enrich and analyze the GO (Gene ontology) annotation and KEGG pathway of the targets. In view of the high homology of gene sequences between SARS-CoV-2 S-protein RBD domain and SARS virus, as well as their similarities in pathogenesis and clinical manifestations, we established SARS-CoV-2 S-protein model using Swiss-Model. The ZDOCK protein docking software was applied to dock the S-protein with the human angiotensin ACE2 protein to find out the key amino acids of the binding site. Taking ACE2 as the receptor, the molecular docking between the active ingredients and the target protein was studied by AutoDock molecular docking software. The interaction between ligand and receptor is applied to provide a choice for screening anti-COVID-19 drugs. RESULTS: A total of 253 active components were predicted. Metascape analysis showed that key candidate targets were significantly enriched in multiple pathways related to different toxins. These key candidate targets were mainly derived from phillyrin and chlorogenic acid. Through the protein docking between S-protein and ACE2, it is found that Glu329/Gln325 and Gln42/Asp38 in ACE2 play an important role in the binding process of the two. The results of molecular docking virtual calculation showed that phillyrin and chlorogenic acid could stably combine with Gln325 and Gln42/Asp38 in ACE2, respectively, which hindered the combination between S- protein and ACE2. CONCLUSION: Phillyrin and chlorogenic acid can effectively prevent the combination of SARS-CoV-2 S-protein and ACE2 at the molecular level. Phillyrin and chlorogenic acid can be used as potential inhibitors of COVID-19 for further research and development.

Effect of Mongolian warm acupuncture on the gene expression profile of rats with insomnia.

BACKGROUND: The mechanism of Mongolian warm acupuncture (MWA) for the treatment of insomnia has not been previously reported. OBJECTIVE: To investigate the effect of MWA on gene expression profile in the p-chlorophenylalanine (PCPA)-induced rat model of insomnia. METHODS: A rat model of insomnia was established and the animals were divided into five groups: control, PCPA (untreated), PCPA+estazolam, PCPA+MA (manual acupuncture), and PCPA+MWA. The rats were euthanased at 7 days after treatment, and hypothalamic tissue was harvested to extract total RNA for the analysis of gene expression profile. Micro-array and Partek Genomics Suite analysis system were used to analyse differential expression of genes between groups. Furthermore, ingenuity pathways analysis was used to analyse the main regulators. RESULTS: After treatment, in rats with improved sleep, micro-array data from the follow-up phase compared with baseline showed that MWA down-regulated 11 genes compared with the control group and 16 genes compared with the PCPA group. Six genes were selected following the micro-array detection to perform quantitative polymerase chain reaction (qPCR) verification, and the results showed that the coincidence rate was up to 90%, which verified the reliability of the microarray results. Compared with the PCPA group, transcription levels of Egr 1, Btg2 and BDNF in the PCPA+MWA group were up-regulated (P<0.05). CONCLUSION: In combination, the findings of this study suggests that MWA is efficacious at improving sleep in an experimental rat model of insomnia.

Research on Roles of Mongolian Medical Warm Acupuncture in Inhibiting p38 MAPK Activation and Apoptosis of Nucleus Pulposus Cells.

BACKGROUND: Mongolian medical warm acupuncture has a desirable therapeutic effect on sciatica. Apoptosis of the nucleus pulposus cells is considered to play an important role in sciatica. Evidence has demonstrated that oxidative stress and its induced activation of the signaling pathways play important roles in sciatica. However, further research is expected to reveal whether Mongolian medical warm acupuncture can inhibit the apoptosis of nucleus pulposus cells and oxidative stress. OBJECTIVE: To study the effect of the p38 MAPK pathway activated by the generated ROS on apoptosis and the expression of the genes related to the balance of the extracellular matrix metabolism during treatment of sciatica with Mongolian medical warm acupuncture. METHOD: The volume of the active oxygen generated in the nucleus pulposus cells was detected following intervention of Mongolian medical warm acupuncture. The p38 MAPK phosphorylation level was detected with Western blot. The genes are related to the metabolism of the nucleus pulposus extracellular matrix. RESULT: Mongolian medical warm acupuncture reduced the active oxygen within the nucleus pulposus cells and inhibited the activation of the p38 MAPK pathway (P=0.013). Meanwhile, it upregulated the gene expression of Type II collagen, aggrecan, Sox-9, and tissue matrix metalloproteinase reagent 1 (P-0.015; P=0.025; P=0.031; P=0.045) and downregulated the gene expression of matrix metalloproteinase 3 (P=0.015). CONCLUSION: Mongolian medical warm acupuncture may inhibit apoptosis of nucleus pulposus cells and activation of the extracellular matrix decomposition metabolism pathway and promote its anabolism. This process may rely on the oxidative stress matrix of the p38 MAPK pathway.

Mechanism of Mongolian medical warm acupuncture in treating insomnia by regulating miR-101a in rats with insomnia.

MicroRNAs (miRNAs or miRs) and the target genes before and after warm acupuncture at the genetic level were assessed, and the cytokines and neurotransmitters related to insomnia were studied. Male Sprague-Dawley rats were used to create PCPA insomnia rat models and randomly divided into the normal, model, warm acupuncture, and drug groups. The Dinghui Acupoint, Heyi Acupoint, and Xin Acupoint were inserted in the Mongolian medicine warm acupuncture group. The differential expression profile of microRNA in the brain tissue of the insomnia rats was determined before and after Mongolian medicine warm acupuncture for establishment of miR-101a mimics and inhibitor. qPCR was used to detect the expression level of miR-101a. Western blotting was used to detect the expression level of PAX8. The rats receiving Mongolian medicine warm acupuncture had 141 miRNAs with differential expression compared with the normal rats. The expression level of miR-101a in the cells of the hippocampus of the insomnia rats transfected with miR-101a mimics increased significantly at 72 h (P<0.05). The activity of the neuronal cells transfected with miR-101a inhibitor increased significantly at 72 h (P<0.05). The western blotting result indicated that the expression of the PAX8 protein in the neuronal cells of the insomnia model rats was inhibited and downregulated significantly at 72 h after addition of miR-101a mimics compared with that in the scramble added group (P<0.01). The levels of the interleukins IL-1, IL-2, and IL-6 and the tumor necrosis factor-α in the hypothalamus, hippocampus, and prefrontal cortex decreased significantly compared with those in the blank control group (P<0.05). The levels of noradrenaline, dopamine, and glutamic decreased significantly following warm acupuncture or western medicine treatment (P<0.05). In conclusion, this study demonstrates that the upregulation of miR-101a in the rats treated with warm acupuncture is directly associated with PAX8 regulation.

Hypolipidemic effects of flavonoids extracted from Lomatogonium rotatum.

Contained in the Mongolian volumes of Chinese Materia Medica, Lomatogonium rotatum Fries ex Nym. may reduce blood lipid levels and prevent obesity; however, its exact mechanism of action remains unclear. The present study investigated the hypolipidemic and obesity-inhibiting effects of four similarly structured flavonoids extracted from L. rotatum. According to a well-established method, flavonoids such as decussatin were extracted from the whole herb of L. rotatum, and male Wistar rats were subsequently fed a high-fructose diet supplemented with flavonoids (20 mg/kg) for 12 weeks. The levels of total cholesterol, triglyceride (TG), low-density lipoprotein-cholesterol and high-density lipoprotein-cholesterol (HDL-C) were detected. In addition, hepatic and epididymal adipose tissues were weighed, and levels of blood glucose, alanine aminotransferase, aspartate aminotransferase, non-esterified fatty acid, insulin and leptin were determined. The mRNA expression levels of fatty acid synthase (FAS) were analyzed using a reverse transcription polymerase chain reaction; whereas FAS, adenosine monophosphate-activated protein kinase (AMPK) and threonine-172 phosphorylated AMPK protein levels were detected by western blotting. The epididymal adipose tissues of rats fed with flavonoids were lighter, as compared with those fed with fructose in the model group. Following a 12-week administration of flavonoids, the serum levels of fasting blood glucose, feeding blood glucose and leptin were decreased. Furthermore, flavonoid treatment reduced TG and cholesterol levels in the blood and increased serum HDL-C levels, as compared with the model group. High-fructose diet administration significantly increased FAS mRNA and protein expression levels, whereas the FAS protein levels of flavonoid-treated rats were markedly reduced. The flavonoid compounds also enhanced threonine-172 phosphorylation of AMPK in the liver lysate, and all flavonoids successfully downregulated leptin levels and the majority decreased the relative weights of epididymal adipose tissue. Therefore, flavonoids may function in a similar way to epigallocatechin gallate, which has previously been shown to inhibit FAS activity by stimulating AMPK in hepatocyte cells via the liver kinase B1 pathway.

Clinical Trial Research on Mongolian Medical Warm Acupuncture in Treating Insomnia.

Objective. Insomnia is one of the most common sleep disorders. Hypnotics have poor long-term efficacy. Mongolian medical warm acupuncture has significant efficacy in treating insomnia. The paper evaluates the role of Mongolian medical warm acupuncture in treating insomnia by investigating the Mongolian medicine syndromes and conditions, Pittsburgh sleep quality index, and polysomnography indexes. Method. The patients were diagnosed in accordance with International Classification of Sleep Disorders (ICSD-2). The insomnia patients were divided into the acupuncture group (40 cases) and the estazolam group (40 cases). The patients underwent intervention of Mongolian medical warm acupuncture and estazolam. The indicators of the Mongolian medicine syndromes and conditions, Pittsburgh sleep quality index (PSQI), and polysomnography indexes (PSG) have been detected. Result. Based on the comparison of the Mongolian medicine syndrome scores between the warm acupuncture group and the drug treatment group, the result indicated P < 0.01. The clinical efficacy result showed that the effective rate (85%) in the warm acupuncture group was higher than that (70%) in the drug group. The total scores of PSQI of both groups were approximated. The sleep quality indexes of both groups decreased significantly (P < 0.05). The sleep quality index in the Mongolian medical warm acupuncture group decreased significantly (P < 0.01) and was better than that in the estazolam group. The sleep efficiency and daytime functions of the patients in the Mongolian medical warm acupuncture group improved significantly (P < 0.01). The sleep time was significantly extended (P < 0.01) in the Mongolian medical warm acupuncture group following PSG intervention. The sleep time during NREM in the Mongolian warm acupuncture group increased significantly (P < 0.01). The sleep time exhibited a decreasing trend during REM and it decreased significantly in the Mongolian warm acupuncture group (P < 0.01). The percentage of sleep time in the total sleep time during NREM3+4 in the Mongolian medical warm acupuncture group increased significantly. Conclusion. Mongolian medical warm acupuncture is efficient and safe in treating insomnia. It is able to better improve the patients' sleep time and daytime functions. It is better than that in the estazolam group following drug withdrawal in terms of improving the sleep time. It is more effective in helping the insomnia patients than hypnotics.

Hypolipidemic effects of a new piperine derivative GB-N from Piper longum in high-fat diet-fed rats.

CONTEXT: Long pepper, Piper longum Linn. (Piperaceae), is widely used in traditional Mongolian medicine for treating hyperlipidemia and coronary heart disease. OBJECTIVE: To investigate the hypolipidemic effects of a new piperine derivative GB-N isolated from long pepper in high-fat diet-fed rats. METHODS: The levels of serum total cholesterol, triacylglycerols (TG), low-density lipoprotein cholesterol (LDL-C) and high-density lipoprotein cholesterol (HDL-C) were determined by enzymatic colorimetric method. The levels of 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA), CYP7A1, lecithin cholesterol acyltransferase (LCAT) and LDL receptor (LDLR) mRNA and protein expression were detected by real-time polymerase chain reaction and western blot analysis. RESULTS AND DISCUSSION: Compared with model rats, oral administration of GB-N at doses of 2.5-10 mg/kg to hyperlipidemic rats could significantly decrease the levels of serum TG from 1.54 mmol/L in hyperlipidemic rats to 0.94-1.02 mmol/L, with an increase in serum HDL-C levels from 0.40 mmol/L in hyperlipidemic rats to 1.21-2.26 mmol/L. Treatment with GB-N (10 mg/kg) could also significantly upregulate levels of hepatic HMG-CoA reductase, CYP7A1, LCAT and LDLR mRNA and protein expression. CONCLUSION: GB-N had hypolipidemic activity via regulating lipid metabolism pathways in liver of hyperlipidemic rats and could be explored as a potential agent for the prevention of hyperlipidemia diseases.