Identification of a potential novel biomarker in intervertebral disk degeneration by bioinformatics analysis and experimental validation.

Background: Intervertebral disk degeneration (IVDD) is a major cause of low back pain and one of the most common health problems all over the world. However, the early diagnosis of IVDD is still restricted. The purpose of this study is to identify and validate the key characteristic gene of IVDD and analyze its correlation with immune cell infiltration. Methods: 3 IVDD-related gene expression profiles were downloaded from the Gene Expression Omnibus database to screen for differentially expressed genes (DEGs). Gene Ontology (GO) and gene set enrichment analysis (GSEA) were conducted to explore the biological functions. Two machine learning algorithms were used to identify characteristic genes, which were tested to further find the key characteristic gene. The receiver operating characteristic curve was performed to estimate the clinical diagnostic value of the key characteristic gene. The excised human intervertebral disks were obtained, and the normal nucleus pulposus (NP) and degenerative NP were carefully separated and cultured in vitro. The expression of the key characteristic gene was validated by real-time quantitative PCR (qRT-PCR). The related protein expression in NP cells was detected by Western blot. Finally, the correlation was investigated between the key characteristic gene and immune cell infiltration. Results: A total of 5 DEGs, including 3 upregulated genes and 2 downregulated genes, were screened between IVDD and control samples. GO enrichment analysis showed that DEGs were enriched to 4 items in BP, 6 items in CC, and 13 items in MF. They mainly included the regulation of ion transmembrane transport, transporter complex, and channel activity. GSEA suggested that the cell cycle, DNA replication, graft versus host disease, and nucleotide excision repair were enriched in control samples, while complement and coagulation cascades, Fc γ R-mediated phagocytosis, neuroactive ligand-receptor interaction, the NOD-like receptor signaling pathway, gap junctions, etc., were enriched in IVDD samples. Furthermore, ZNF542P was identified and tested as key characteristic gene in IVDD samples through machine learning algorithms and showed a good diagnostic value. The results of qRT-PCR showed that compared with normal NP cells, the expression of ZNF542P gene was decreased in degenerated NP cells. The results of Western blot suggested that compared with normal NP cells, the expression of NLRP3 and pro Caspase-1 was increased in degenerated NP cells. Finally, we found that the expression of ZNF542P was positively related to the proportions of T cells gamma delta (γδT cells). Conclusion: ZNF542P is a potential biomarker in the early diagnosis of IVDD and may be associated with the NOD-like receptor signaling pathway and the infiltration of γδT cells.

A Study on the Potential Mechanism of Shujin Dingtong Recipe against Osteoarthritis Based on Network Pharmacology and Molecular Docking.

Background: With the aging of the social population, Osteoarthritis (OA) has already become a vital health and economic problem globally. Shujin Dingtong recipe (SJDTR) is an effective formula to treat OA in China. Although studies have shown that SJDTR can significantly alleviate OA symptoms, its mechanism still remains unclear. Purpose: This study is aimed at investigating the potential mechanism of SJDTR for the treatment of OA based on network pharmacology and molecular docking. Methods: Main ingredients of SJDTR were retrieved from the Traditional Chinese Medicine Systems Pharmacology (TCMSP) database. OA disease targets were obtained from the Gene Expression Omnibus (GEO) database. The overlapped targets and signaling pathways were explored using Protein-Protein Interaction (PPI) network, Gene Ontology (GO), and Kyoto Encyclopedia of Genes and Genomes (KEGG). Following this, the core targets were employed to dock with corresponding components via molecular docking in order to further explore the mechanism of SJDTR in the treatment of OA. Results: From network pharmacology, we found 100 active components of SJDTR, 31 drug and OA-related targets, 1161 GO items, and 91 signaling pathways. Based on the analysis with PPI network and molecular docking, TP53, CCNB1, and MMP-2 were selected for the core targets of SJDTR against OA. Molecular docking demonstrated that Quercetin, Baicalein, and Luteolin, had good binding with the TP53, CCNB1, and MMP-2 protein, respectively. Conclusion: To conclude, our study suggested the main ingredients of SJDTR might alleviate the progression of OA through multiple targets and pathways. Additionally, network pharmacology and molecular docking, as new approaches, were adopted for systematically exploring the potential mechanism of SJDTR for the treatment of OA.

Entropy-Driven Self-Assembly of Single Quantum Dot Sensor for Catalytic Imaging of Telomerase in Living Cells.

Telomerase is a highly valuable cancer diagnosis biomarker and a promising cancer therapy target. So far, most telomerase assays are limited by the involvement of tedious procedures, multiple enzymes, and complicated reaction schemes. Sensitive monitoring of low-abundant telomerase in living cells remains a challenge. Herein, we demonstrate an entropy-driven catalytic assembly of quantum dot (QD) sensors for accurate detection and imaging of telomerase activity in living cells. In this sensor, target telomerase specifically catalyzes extension of telomerase primer, and the extended primer subsequently acts as a catalyst to continuously initiate entropy-driven catalytic reaction, generating a large number of fluorophore- and biotin-labeled DNAs that can be self-assembled on the QD surface to induce an efficient Föster resonance energy transfer signal. The proposed sensor requires a single step for both recognition and amplification of the telomerase signal, eliminating the use of either protein enzymes or laborious procedures. Taking advantage of the inherent superiority of single-molecule fluorescence detection and high amplification efficiency of the entropy-driven reaction, this sensor demonstrates single-cell sensitivity for the in vitro assay. Moreover, it is capable of screening the telomerase inhibitor, discriminating different tumor cells from normal ones, and even real-time imaging telomerase in living cells, providing a novel platform for telomerase-associated cancer diagnosis and drug screening.

Network Pharmacology-Based Strategy and Molecular Docking to Explore the Potential Mechanism of Jintiange Capsule for Treating Osteoporosis.

BACKGROUND: With the advent of ageing population, osteoporosis (OP) has already become a global challenge. Jintiange capsule is extensively applied to treat OP in China. Although recent studies demonstrate that it generates significant effects on strengthening bone, the exact mechanism of the jintiange capsule for treating OP remains unknown. PURPOSE: To understand the main ingredients of the jintiange capsule, predict the possible targets and the relevant signal transduction pathways, and explore the mechanism of the jintiange capsule for the treatment of OP. METHODS: Main ingredients of the jintiange capsule, drug targets, and potential disease targets for OP were obtained from public databases. Molecular biological processes and signaling pathways were determined via bioinformatic analysis, containing protein-protein interaction (PPI), Gene Ontology (GO), and Kyoto Encyclopedia of Genes and Genomes (KEGG). Subsequently, the disease-drug-ingredient-targets-pathways networks were constructed using Cytoscape. According to CytoNCA, core targets were acquired. Finally, the present study conducted molecular docking for better testing the abovementioned results. RESULTS: In the current work, we found that 4 main ingredients of the jintiange capsule, 33 drug targets, 4745 potential disease targets for OP, and 12 overlapping targets were identified. PPI network containing 12 nodes and 25 edges proved that there existed a complex relationship. As revealed by GO functional annotation, the intersected targets were mostly associated with BP, CC, and MF. The targets were enriched to 368 items in BP, 27 items in CC, and 42 items in MF. They mainly included calcium ion homeostasis, calcium channel complex, and calcium channel regulator activity. According to KEGG pathway analysis, the intersected targets were mostly associated with Rap 1, cGMP-PKG, Ras, cAMP, calcium pathways, and so on. Based on the analysis with CytoNCA, we acquired 4 core targets, respectively-CALR, SPARC, CALM1, and CALM2. Besides, 2 core targets, CALR and CALM1, were selected for molecular docking experiments. Molecular docking revealed that the main ingredient, calcium phosphate, had good binding with the CALR protein and CALM1 protein. CONCLUSION: To conclude, the main ingredient of the jintiange capsule, particularly calcium phosphate, may interact with 2 targets, CALR and CALM1, and regulate multiple signaling pathways to treat OP. Additionally, this also benefits us in further understanding the mechanism of the jintiange capsule for treating OP.

Triterpenic Acids from Potentilla parvifolia and Their Protective Effects against Okadaic Acid Induced Neurotoxicity in Differentiated SH-SY5Y Cells.

Six triterpenic acids were separated and purified from the ethyl acetate extractive fraction of ethanol extracts of Potentilla parvifolia FISCH. using a variety of chromatographic methods. The neuroprotective effects of these triterpenoids were investigated in the present study, in which the okadaic acid induced neurotoxicity in human neuroblastoma SH-SY5Y cells were used as an Alzheimer's disease cell model in vitro. The cell model was established with all trans-retinoic acid (5 µmol/L, 4 d) and okadaic acid (40 nmol/L, 6 h) treatments to induce tau phosphorylation and synaptic atrophy. Subsequently, the neuroprotective effects of these triterpenic acids were evaluated in vitro by this cell model. Results from the Western blot and morphology analysis suggested that compounds 3-6 had the better neuroprotective effects. Furthermore, we tested the level of mitochondrial reactive oxygen species and mitochondrial membrane potential of these compounds in SH-SY5Y cells by flow cytometry technology to investigate the potential neuroprotective mechanism of these compounds. All of the results indicated that maybe the mechanism of compounds 5 and 6 is to protect the cell from mitochondrial oxidative stress injuries.

Triterpenoids with antioxidant activities from Myricaria squamosa.

A new triterpenoid, namely myricarin C (compound 1), together with three known compounds myricarin A (compound 2) and myricarin B (compound 3), 3α-hydroxy-D-friedoolean-14-en-28-oic acid (compound 4), was isolated from the overground part of Myricaria squamosa. Compound 2 and compound 3 existed in the solution by the form of cis-trans isomers. Their structures were elucidated by means of extensive spectroscopic methods, including 1D-NMR, 2D-NMR, and HR-ESI-MS. The antioxidant properties of all compounds were calculated based on the DPPH radical scavenging activities. Results showed that myricarin A and myricarin C had general antioxidant activities with EC50 values of 40.90 µg/ml, 42.22 µg/ml, respectively, compared to the control, rutin (5.17 µg/ml). The EC50 values of myricarin B was 195.81 µg/ml. Compound 4 had no antioxidant activities.

[GC-MS analysis of low polar components from Ainsliaea yunnanensis].

OBJECTIVE: To analyze the chemical constituents of low polar components from ethanol extract of Ainsliaea yunnanensis. METHODS: The air-dried stems and leaves of Ainsliaea yunnanensis were extracted with ethanol using ultrasonic extration. After removal of solvent under reduced pressure, the residue was suspended in H2O, and then partitioned sequentially with petroleum ether, EtOAc and n-BuOH, respectively. Then the petroleum ether fraction was chromatographed on normal phase silica gels eluting with a gradient increasing Me2 CO in petroleum ether. Then the components of low polarity were separated from the petroleum ether-Me2CO(50:1 - 20: 1) fraction and identified by GC-MS, and the relative contents of the components were determined with area percentage method. RESULTS: 50 constituents were identified from the low polar components of Ainsliaea yunnanensis. CONCLUSION: The research provides a theoretical basis for the study of chemical constituents and pharmaceutical activities of Ainsliaea yunnanensis.

[Triterpene compounds of Ainsliaea yunnanensis].

The compounds of Ainsliaea yunnanensis were isolated and purified by various kinds of column chromatography methods and their structures were determined by spectroscopic data analysis. Twelve compounds were obtained from the petroleum ether of ethanolic extract of A. yunnanensis and elucidated as bauerenyl acetate (1), bauerenol (2), alpha-amyrin (3), psi-taraxasterol (4), beta-amyrin (5), echinocystic acid (6), multiflorenol (7), 3beta-hydroxy-olean-18-ene germanicol (8), 3beta-hexadecanoyl-12-oleanen-11-one (9), fernenol (10), fern-7-en-3beta-ol (11), and lupeol (12). All compounds were isolated from this genus for the first time except compound 1, 3, 5 and 10, and they were all isolated from this plant for the first time.

Biomechanical properties of peripheral nerve after acellular treatment.

BACKGROUND: Peripheral nerve injury causes a high rate of disability and a huge economic burden, and is currently one of the serious health problems in the world. The use of nerve grafts plays a vital role in repairing nerve defects. Acellular nerve grafts have been widely used in many experimental models as a peripheral nerve substitute. The purpose of this study was to test the biomechanical properties of acellular nerve grafts. METHODS: Thirty-four fresh sciatic nerves were obtained from 17 adult male Wistar rats (age of 3 months) and randomly assigned to 3 groups: normal control group, nerve segments underwent no treatment and were put in phosphate buffered saline (pH 7.4) and stored at 4°C until further use; physical method group, nerve segments were frozen at -196°C and then thawed at 37°C; and chemical method group, nerve segments were chemically extracted with the detergents Triton X-200, sulfobetaine-10 (SB-10) and sulfobetaine-16 (SB-16). After the acellularization process was completed, the structural changes of in the sciatic nerves in each group were observed by hematoxylin-eosin staining and field emission scanning electron microscopy, then biomechanical properties were tested using a mechanical apparatus (Endura TEC ELF 3200, Bose, Boston, USA). RESULTS: Hematoxylin-eosin staining and field emission scanning electron microscopy demonstrated that the effects of acellularization, demyelination, and integrity of nerve fiber tube of the chemical method were better than that of the physical method. Biomechanical testing showed that peripheral nerve grafts treated with the chemical method resulted in some decreased biomechanical properties (ultimate load, ultimate stress, ultimate strain, and mechanical work to fracture) compared with normal control nerves, but the differences were not statistically significant (P > 0.05). CONCLUSION: Nerve treated with the chemical method may be more appropriate for use in implantation than nerve treated with the physical method.