Treadmill exercise alleviates neuropathic pain by regulating mitophagy of the anterior cingulate cortex in rats / 生理学报

This study aimed to investigate the effect of treadmill exercise on neuropathic pain and to determine whether mitophagy of the anterior cingulate cortex (ACC) contributes to exercise-mediated amelioration of neuropathic pain. Chronic constriction injury of the sciatic nerve (CCI) was used to establish a neuropathic pain model in Sprague-Dawley (SD) rats. Von-Frey filaments were used to assess the mechanical paw withdrawal threshold (PWT), and a thermal radiation meter was used to assess the thermal paw withdrawal latency (PWL) in rats. qPCR was used to evaluate the mRNA levels of Pink1, Parkin, Fundc1, and Bnip3. Western blot was used to evaluate the protein levels of PINK1 and PARKIN. To determine the impact of the mitophagy inducer carbonyl cyanide m-chlorophenylhydrazone (CCCP) on pain behaviors in CCI rats, 24 SD rats were randomly divided into CCI drug control group (CCI+Veh group), CCI+CCCP low-dose group (CCI+CCCP0.25), CCI+CCCP medium-dose group (CCI+CCCP2.5), and CCI+CCCP high-dose group (CCI+CCCP5). Pain behaviors were assessed on 0, 1, 3, 5, and 7 days after modeling. To explore whether exercise regulates pain through mitophagy, 24 SD rats were divided into sham, CCI, and CCI+Exercise (CCI+Exe) groups. The rats in the CCI+Exe group underwent 4-week low-moderate treadmill training one week after modeling. The mechanical pain and thermal pain behaviors of the rats in each group were assessed on 0, 7, 14, 21, and 35 days after modeling. Western blot was used to detect the levels of the mitophagy-related proteins PINK1, PARKIN, LC3 II/LC3 I, and P62 in ACC tissues. Transmission electron microscopy was used to observe the ultrastructure of mitochondrial morphology in the ACC. The results showed that: (1) Compared with the sham group, the pain thresholds of the ipsilateral side of the CCI group decreased significantly (P < 0.001). Meanwhile, the mRNA and protein levels of Pink1 were significantly higher, and those of Parkin were lower in the CCI group (P < 0.05). (2) Compared with the CCI+Veh group, each CCCP-dose group showed higher mechanical and thermal pain thresholds, and the levels of PINK1 and LC3 II/LC3 I were elevated significantly (P < 0.05, P < 0.01). (3) The pain thresholds of the CCI+Exe group increased significantly compared with those of the CCI group after treadmill intervention (P < 0.001, P < 0.01). Compared with the CCI group, the protein levels of PINK1 and P62 were decreased (P < 0.001, P < 0.01), and the protein levels of PARKIN and LC3 II/LC3 I were increased in the CCI+Exe group (P < 0.01, P < 0.05). Rod-shaped mitochondria were observed in the ACC of CCI+Exe group, and there were little mitochondrial fragmentation, swelling, or vacuoles. The results suggest that the mitochondrial PINK1/PARKIN autophagy pathway is blocked in the ACC of neuropathic pain model rats. Treadmill exercise could restore mitochondrial homeostasis and relieve neuropathic pain via the PINK1/PARKIN pathway.

Protective Effect of Glycyrrhizae Radix et Rhizoma-Granati Pericarpium Formula on Ultraviolet-induced Skin Damage / 中国实验方剂学杂志

Objective:To explore the potential synergistic protective mechanism of Glycyrrhizae Radix et Rhizoma-Granati Pericarpium formula compound by using the methods and tools of network pharmacology，and provide a basis for the modernization of traditional Chinese medicine（TCM） compounds and the discovery of new drugs. Method:Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform （TCMSP） was used to obtain the active components of Glycyrrhizae Radix et Rhizoma-Granati Pericarpium formula and their corresponding targets. The obtained targets were input to the UniProt database to inquire the gene names corresponding to the targets. By searching the CTD database，Genecards database and OMIM database of disease-related websites，the anti-sunburn targets were obtained. The interaction of the active targets was analyzed with online STRING database to screen the predicted core targets. The gene ontolog（GO） gene function enrichment analysis and Kyoto encyclopedia of genes and genomes（KEGG） pathway enrichment analysis of the predictive targets were performed by using DAVID database. Cytoscape 3.6.1 software was used to make "drug-component-target" network diagram，"protein-protein interaction" network diagram and "component-target-pathway" network diagram. Online website Draw Venn Diagram was used to show the relationship between disease targets and drug predicted targets. R Studio software was used to draw the functional enrichment analysis diagram of GO gene and KEGG pathway. Molecular docking between the active ingredients and the core targets was performed using GOLD software. Result:The 16 active compounds were collected，such as liquiritin，glycyrrhizin，kaempferol and quercetin. The active components mainly acted on 5 core targets：protein kinase B1（AKT1），interleukin（IL）-6，vascular endothelial growth factor（VEGFA），tumor necrosis factor（TNF） and tumor suppressor gene （TP53） and played a role in anti-sunburn effect primarily through these pathways such as hepatitis B，pathways in cancer，toxoplasmosis，chagas disease（American trypanosomiasis），and TNF signaling pathway. Conclusion:Based on the method of network pharmacology，the present study has preliminarily explored the anti-sunburn targets and pathways of Glycyrrhizae Radix et Rhizoma-Granati Pericarpium formula，and further verified the characteristics of multi-component and multi-target treatment of diseases in TCM，so as to provide certain scientific ideas for the modernization research of Chinese herbal compound prescriptions.

Anti-inflammatory Mechanism of Epimedii Folium in Treatment of Depression Based on Network Pharmacology and Molecular Docking / 中国实验方剂学杂志

Objective:To predict the anti-inflammatory targets and relevant signaling pathways of Epimedii Folium in the treatment of depression by network pharmacology，in order to explore the potential mechanism of its anti-depression effect. Method:The active constituents of Epimedii Folium were collected and screened out through the Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform（TCMSP） database. PharmMapper server and TCMSP database were used to predict and screen out protein targets. OMIM database，CTD database and GeneCards database were used to screen out relevant targets and anti-inflammatory targets of depression. Enrichment analysis of the gene ontology （GO） function and Kyoto Encyclopedia of genes and genomes（KEGG） signaling pathway for the key anti-inflammatory targets of Epimedii Folium were carried out by DAVID database. Cytoscape 3.6.0 was used to construct the network diagram of "active component-action target-signal pathway" of Epimedii Folium and analyze the topological structure of the network. GOLD molecular docking software was used to verify the results of active components and key anti-inflammatory targets. Result:A total of 12 active components，30 targets and 5 key anti-inflammatory targets of Epimedii Folium were screened out， 65 biological processes，4 cell components and 1 molecular function were enriched with GO function， and 41 KEGG pathways were enriched and analyzed，including 9 inflammation-related signaling pathways. Molecular docking verified that icariin and key anti-inflammatory targets could form the optimal binding structure. Conclusion:The study preliminarily reveals the molecular mechanism of Epimedii Folium on depression through its anti-inflammatory target and its relevant signaling pathway network，so as to provide a basis for further study on the antidepressant effect of Epimedii Folium.

Quantitative Analysis of Index Components in Glycyrrhizae Radix et Khizoma by Near Infrared Spectroscopy Based on R Software / 中国实验方剂学杂志

Objective: To establish a better near infrared quantitative model for quality control of Glycyrrhizae Radix et Rhizoma of components (moisture,total ash,liquiritin and glycyrrhizic acid) in liquorice,in order to realize rapid detection.Method: The contents of moisture,total ash,liquiritin and glycyrrhizic acid were determined in 97 samples based on the methods set forth in Chinese Pharmacopoeia.Meanwhile,the near infrared spectrum was scanned using near infrared spectroscope.R software was used to screen out the spectral pretreatment and build the quantitative models.Result: The optimum spectral pretreatment method for establishing the near infrared quantitative model of moisture and liquiritin was the first order derivative.For moisture,the correlation coefficients of test and validation were 0.930 0 and 0.929 9,and the root mean square errors were 0.243 2 and 0.203 8,respectively.For liquiritin,the correlation coefficients of test and validation were 0.930 3 and 0.907 6,and the root mean square errors were 0.093 9 and 0.128 9,respectively.The optimum spectral pretreatment method for establishing the near infrared quantitative model of total ash was MSC.The correlation coefficients of test and validation were 0.926 5 and 0.917 7,and the root mean square errors were 0.109 6 and 0.103 7,respectively.The optimum spectral pretreatment method for establishing the near infrared quantitative model of glycyrrhizic acid was SNV.The correlation coefficients of test and validation were 0.918 1 and 0.915 7,and the root mean square errors were 0.274 8 and 0.236 0,respectively.Conclusion: In this study,a better near infrared quantitative models for quality control of components of Glycyrrhizae Radix et Rhizoma were established,with a high accuracy,which laid a foundation for rapid detection of the components in Glycyrrhizae Radix et Rhizoma.

Correlation of Nucleostemin with Programmed Death-ligand-1 in Multiple Myeloma U266 Cells / 中国实验血液学杂志

OBJECTIVE@#To investigate the correlation of nucleostemin (NS) gene with programmed death ligand-1 (PD-L1) in myeloma cells and the effect of NS expression down-regulation on the apoptosis of multiple myeloma cells, and to evaluate the associations among NS, PD-L1 and biological behavior of MM cells, and the feasibility of both NS and PD-1 as markers reflecting the status of MM cells.@*METHODS@#The NS gene expression in U266 cells was down-regulated by NS-RNAi-GV248 recombinant lentivirus, the real-time PCR was used to detect the mRNA expression of NS, PD-L1 and PI3K/AKT/mTOR. The Western blot and flow cytometry were used to detect the expression of NS and PD-L1. The Annexin V-APC/7-AAD staining method was used to detect the apoptosis of U266 cells before and after knocking out the NS gene.@*RESULTS@#Under the condition of MOI=10, the transfection efficiency was more than 75% by means of the fluorescent microscopy; real-time PCR showed that compared with the negative control group (1.002±0.026), the mRNA expression of NS, PD-L1 and PI3K/AKT/mTOR gene in the transfection group (0.415±0.089) was significantly reduced (P＜0.05). The results of flow cytometry and Western blot showed that the protein expression of PD-L1 was significantly down-regulated after transfection. After down-regulation of NS gene expression, the apoptosis of U266 cells increased (P＜0.05).@*CONCLUSION@#The abnormal high expression of NS and PD-L1 genes exists in U266 cells, moreover, the down-regulation of PD-L1 and the related PISK/AKT/mTOR pathway gene expression appears after down-regulation of NS gene expression, which suggest that the cell biological changes resulted from above-mentioned results, show a synergestic effect on U266 cells.

Network pharmacology of flavonoids in Sophora alopecuroides / 中国中药杂志

The aim of this paper was to investigate the potential pharmacological effect of flavonoids in Sophora alopecuroides by network pharmacology. This study predicted the potential targets of 11 flavonoids of S. alopecuroides with help of reversed pharmacophore matching target recognition service platform (PharmMapper). The pathway information was acquired from DAVID and KEGG databases. Cytoscape software was used to construct the "ingredient-target-pathway" network of flavonoids active components of S. alopecuroides. The flavonoids active components of S. alopecuroides play anti-inflammatory, blood sugar regulating and other pharmacological effects by regulating 62 targets (such as INSR，KDR，MET) and intervening 44 pathways, such as B cell receptor signaling pathway, insulin signaling pathway, neurotrophin signaling pathway, and T cell receptor signaling pathway. In this study, the mechanism of "muti components-multitargets-multiple pathway" of flavonoids was studied. It reflects the multi-components, multi-targets and multiple pathway features of traditional Chinese medicine. Meanwhile, it provides a scientific basis for the elucidation the mechanism of S. alopecuroides as a medicine, and the development and utilization resources of S. alopecuroides.

Research on network pharmacology of alkaloids in Sophora alopecuroides / 中国中药杂志

It was aimed at exploring the potential pharmacological effects of alkaloids in Sophora alopecuroides by means of network pharmacology in this study. The main alkaloids in S. alopecuroides were collected for analysis of drug properties, prediction of potential targets and screening of signaling pathways. DAVID analysis tool combined with KEGG database was used to annotate and analyze the signaling pathway. The alkaloids-targets-signaling pathways network was built through Cytoscape software. Results showed that 17 alkaloids in S. alopecuroides involved 49 targets (170 times in all) and 22 important signaling pathways. Three nodes in model of network pharmacology were cross-linked, and the metabolic pathways were coordinated and regulated by each other. It indicated that alkaloids in S. alopecuroides may have therapeutic effect on diseases of cancer, metabolic disorder, endocrine system, digestive system, nervous system and so on.

Effects of different drying methods on content of bioactive component and antioxidant activity in Lycium ruthenicum / 中国中药杂志

To compare the appearances, tastes, contents of bioactive components and antioxidant activity of Lyceum ruthenicum under different drying methods, so as to direct its production practice. The folin-phenol colorimetric method, UV, extinction coefficient method and DPPH, as well as fluorescence recovery after photobleaching (FRAP) method to determine the contents of polyphenols, proanthocyanidins, total anthocyanin and antioxidant activity under different drying methods： vacuum freeze drying, low-temperature oven drying and air drying for L. ruthenicum. The results showed that the drying methods had certain effects on its appearances, tastes, contents of bioactive components and antioxidant activity. The appearances and tastes were best after the L. ruthenicum was dried by vacuum freeze drying, with significantly lower moisture than air drying method. The contents of total polyphenols, anthocyanin and proanthocyanidins were highest by air-drying but lowest by low temperature oven drying in L. ruthenicum. The scavenging ability to DPPH was strongest by freeze-drying and lowest by low temperature oven drying, while the antioxidant activity was strongest by air-drying in the FRAT method. In addition, the appearances and tastes were poor in air drying, with higher moisture but highest contents of the three bioactive components. Therefore, the drying methods for L. ruthenicum shall be comprehensively considered.

Comparative identification studies on wild and cultivated Glycyrrhiza uralensis produced Ningxia based on index components and near-infrared spectroscopy / 中国中药杂志

This study is to construct a rapid and effective method for identification of wild and cultivated Glycyrrhiza uralensis (hereinafter referred to as Glycyrrhizae Radix et Rhizoma) from Ningxia by comparison of the difference in chromatography identification based on index components and near-infrared spectroscopy identification. HPLC and UV methods were used to determine the content of liquiritin, glycyrrhizate and total flavonoids for 9 wild Glycyrrhizae Radix et Rhizoma and 14 cultivated Glycyrrhizae Radix et Rhizoma samples,and the near-infrared spectroscopy was also,collected. The results illustrated that the chromatography identification based on index components could not identify wild and cultivated Glycyrrhizae Radix et Rhizoma from Ningxia, while near-infrared spectroscopy could quickly and effectively achieve it. It provides an effective method for the growth pattern identification and application of Glycyrrhizae Radix et Rhizoma.

Comparative studies on HPLC fingerprint and near-infrared spectra of cultivated and reference crude Scutellaria baicalensis / 中国中药杂志

Scutellaria baicalensis is a common and important medicinal plant in China, facing with reducing sharply in wild resources. To meet the needs in Chinese herbwouls medicine market and clinical application, S. baicalensis has been widely cultivated in Ningxia, Hebei, Shanxi, and Gansu et al. HPLC finger-print and near-infrared were studied in the research to evaluate quality difference of S. baicalensis in four districts. The results showed that the similarity of HPLC finger-print of 12 cultivated S. baicalensis and reference crude herb is more than 0.961, and the other is more than 0.983. On the other hand, paired sample t-test indicated there has no significant difference between the common peaks' area of 12 cultivated S. baicalensis and reference crude herb. It was verified that 12 cultivated S. baicalensis has highly consistency with reference crude herb. On the basis of chromatographic finger-print and near-infrared spectrum, the study applied paired sample t-test to verify analysis results, which could avoid erroneous judgment induced by indefinite threshold values in the similarity of chromatographic finger-print and provide reliable basis for the analysis results. Meanwhile, it also provides a new idea for improving the quality control method of Chinese medicinal materials by comparative study about two comprehensive detection means.

Hyperammonemia-induced hepatic injury in rats: characterization of a new animal model / 中华肝脏病杂志

<p><b>OBJECTIVE</b>To establish an accurate new rat model of hyperammonemia-induced liver injury for use in studies of the molecular mechanisms underlying acute liver failure (ALF).</p><p><b>METHODS</b>Twenty-six Sprague-Dawley rats were administered D-galactosamine (400 mg/kg) and endotoxin (50 mug/kg) via intraperitoneal injection to induce ALF and sacrificed at 12 h post-injection (ALF-12 group, n = 10) or 24 h post-injection (ALF-24 group, n = 16). Ten rats administered physiological saline served as the control group. In addition, 20 rats were given serial oral administrations of 10% NH4Cl solution (10 ml/kg, every 8 hrs) to establish the hyperammonemia-induced liver injury model; an additional 20 rats were prepared in parallel to serve as the ALF control group (n = 10; D-galactosamine at 800 mg/kg every 6 d for 30 days) and the physiological saline control group (n = 10). Serum samples were collected from each mouse and used to detect markers of liver function, including alanine aminotransferase (ALT), aspartate aminotransferase (AST), alpha-fetal protein (AFP), and gamma-glutamyltransferase (GGT), as well as blood ammonia (BA) level and prothrombin time activity (PT-A). Affects on liver histology was assessed by hematoxylin and eosin staining of resected liver tissues, and on apoptosis by TUNEL assay and calculating the apoptotic index (AI).</p><p><b>RESULTS</b>ALF rats showed elevated levels of ALT (1202.51+/-282.00 U/L), AST (1560.14+/-298.98 U/L), and BA (165.9+/-23.6 mumol/L) as early as 6 hrs after model establishment; these levels peaked at 12 hrs after model establishment (ALT: 774.40+/-207.65 U/L; AST: 967.60+/-121.94 U/L; BA: 143.4+/-18.1 mumol/L; P less than 0.05). No significant variations were detected in the levels of AFP (except for the ALF-24 group) or GGT. Liver tissues of the ALF-12 and ALF-24 groups showed large or diffuse hemorrhagic necroses with sinusoidal congestion or spotty bleeding, as well as increased AI. Hyperammonemia-induced liver injury rats showed elevated levels of ALT and BA as early as 6 hrs after model establishment. Similar to the ALF rats, AFP and GGT were unaffected and AI increased. However, in contrast to the ALF rats, the liver tissues of the hyperammonemia-induced liver injury rats showed no signs of hepatocyte swelling, necrosis, or inflammatory cell invasion.</p><p><b>CONCLUSION</b>ALF rats and hyperammonemia-induced liver injury rats have elevated BA and marked hepatocyte necrosis. Given that reducing the level of ammonemia can improve the animal's biochemistry indexes, it is likely that hyperammonemia plays a role in acute liver injury or ALF consequent to repeated injury. The pathogenic mechanisms of repeated injury may involve promotion of hepatocyte apoptosis in conjunction with inhibition of cellular regeneration.</p>