Analysis on Antidepressant Mechanism of Verbascoside Based on RNA-Seq Technology / 中国实验方剂学杂志

Objetive： To investigate the mechanism of antidepressant effect of verbascoside based on high-throughput sequencing technology （RNA-Seq），and to explore the possible targets and signaling pathways. MethodsForty C57BL/6 mice were randomly divided into control group，model group，fluoxetine group and verbascoside group，10 mice in each group. Except for the control group，all the other three groups were constructed with chronic unpredictable mild stimulation （CUMS） combined with solitary feeding for four weeks. Control group，fluoxetine group and verbascoside group were administered by gavage once daily for three weeks during the second week of modeling. The mice were assessed by sugar-water preference test，forced swimming test，open field test，elevated cross maze test，and water maze test. Enzyme linked immunosorbent assay（ELISA） was performed to detect the levels of major neurotransmitters and inflammatory factors in mice serum，and mRNA high-throughput sequencing was performed in the nucleus accumben and colon to screen differentially expressed genes and perform pathway enrichment analysis. Real-time polymerase chain reaction（Real-time PCR） was used to detect the mRNA expression of Gad，Slc32a1（VGAT） and brain-derived neurotrophic factor (BDNF） in nucleus accumbens. ResultCompared with the control group，the anxiety and depression-like behaviors in the model group increased，while the learning and memory ability decreased significantly. The content of neurotransmitter in serum decreased significantly. The content of pro-inflammatory factors increased significantly. The mRNA expression of Gad and Slc32a1（VGAT） in nucleus accumbens decreased significantly，while that of BDNF increased significantly. Compared with the model group，the anxiety and depression-like behavior of mice in verbascoside group was significantly relieved. The neurotransmitter content increased significantly，and the pro-inflammatory factors decreased significantly. The mRNA expression of Gad and Slc32a1（VGAT） in nucleus accumbens increased significantly，while that of BDNF decreased significantly.A total of 48 differentially expressed genes in nucleus accumbens and 43 differentially expressed genes in colon were screened by high-throughput sequencing. Differential genes in nucleus accumbens mainly focus on neuroactive ligand-receptor interaction， γ-aminobutyric acid（GABA）ergic synapse，synaptic vesicle cycle and other pathways. Colonic differential genes are mainly concentrated in GABAergic synapses，synaptic vesicle circulation，cyclic adenosine monophosphate（cAMP） signal pathway and other signal pathways. Compared with the control group，the mRNA expression of Gad and Slc32a1（VGAT） in nucleus accumbens of model group decreased significantly，while the mRNA expression of BDNF increased significantly. ConclusionVerbascoside has significant antidepressant effects. Its antidepressant effect may be related to the increase of monoamine neurotransmitters，the decrease of pro-inflammatory factors and the restoration of neurotransmitter homeostasis by increasing GABA，and it mainly acts through the signaling pathways such as neuroactive ligand-receptor interaction，GABAergic synapses，synaptic vesicle cycle and cAMP signaling pathway.

Application of reactive soft tissue preservation in extraction sockets of periodontitis: recent progress and perspectives / 口腔疾病防治

@#Chronic periodontitis is a prevalent disease; if left untreated, it is a main indication for tooth extraction and can lead to tooth loss. The reactive soft tissue, formed as a result of the immune response to chronic inflammation, is left in the compromised socket. The major concern is how to deal with the residual reactive soft tissue. Conservative thought states that the reactive soft tissue should be completely debrided. In addition, novel practices concerning the reactive soft tissue were proposed in recent trials, which demonstrated that there might be merits for soft and hard tissue regeneration with preservation of the reactive soft tissue. Studies have shown that mesenchymal stem cells exist in inflammatory reactive soft tissue, stressing their potential in tissue regeneration. Although the therapeutic value is highly promising, the specific components of the reactive soft tissue and the standard on whether it should be preserved need further investigation.

Metagenomic study on the composition and function of oral microorganisms in healthy adults / 口腔疾病防治

Objective@#Metagenomic sequencing was used to explore the species composition and internal functional metabolic pathway of saliva and supragingival plaque microbial communities in healthy adults to provide a theoretical reference for the biological prevention and treatment of oral diseases.@*Methods@#Saliva and supragingival plaque samples were collected from healthy adults, total DNA was extracted, and a metagenomic library was constructed. The qualified library was sequenced via metagenomics, and the sequencing data were analyzed using bioinformatics and statistics. @*Results @#The main bacterial phyla in healthy oral samples were Proteobacteria (32.51%), Bacteroidetes (30.81%), and Actinobacteria (16.23%), and the main bacterial species were Corynebacterium matruchotii (3.84%), Haemophilus parainfluenzae (2.91%), and Prevotella melaninogenica (2.76%). The alpha diversity of the supragingival plaque group was higher than that of the saliva group, and there was a significant difference in the composition of the microbial community between the two groups (P<0.05). At the species level, Prevotella melaninogenica, Fusobacterium periodonticum, and Prevotella intermedia were more abundant in saliva samples than in supragingival plaque samples, while Corynebacterium matruchotii, Propionibacterium acidifaciens, and Rothia dentocariosa were more abundant in supragingival plaque samples than in saliva samples (P<0.05). High-quality gene sets of saliva and supragingival plaque in healthy adults were constructed based on metagenomic sequencing. The results of KEGG pathway functional metabolic differences showed that starch and sucrose metabolism, leucine and isoleucine degradation, and arginine biosynthesis in salivary microorganisms were more abundant than in supragingival plaque, while glycolysis/gluconeogenesis and carbon metabolism in supragingival plaque were more abundant than in saliva.@* Conclusion@#There are significant differences in the species composition and functional gene metabolic pathways of saliva and supragingival plaque microecology in healthy adults. The sensitivity of dominant species in different microecological regions to the identification of oral diseases may be different. In the microbiological study of oral diseases, appropriate samples should be selected according to different diseases.

Research strategies for endophytes in medicinal plants based on high-throughput sequencing and traditional culture and isolation methods / 中国中药杂志

The research on endophytes of medicinal plants mainly relies on the traditional culture and isolation methods. Because of their functions such as promoting host growth, improving stress resistance, promoting the accumulation of medicinal active ingredients or directly producing medicinal active ingredients, the endophytes of medicinal plants have gradually attracted wide attention. However, it was found that the strains isolated by traditional methods were not the true dominant endophytes of medicinal plants by comparing the results of traditional culture isolation with high-throughput sequencing. The blind and random nature of traditional methods leads to the lack of standards in terms of medium selection, culture time and interaction between species. On the contrary, high-throughput sequencing technology is an emerging molecular biology technology developed in recent decades. Due to its high resolution level and indepen-dent culture, it can be used for thorough analysis of the community structure and diversity of environmental microorganisms. Therefore, we proposed the strategy of using high-throughput sequencing technology to guide the traditional culture and isolation of endophytes from medicinal plants. Firstly, the endophytic structure and diversity of medicinal plants were completely clear by high-throughput sequencing technology, and the dominant endophytes of the host were unequivocal. Then according to the characteristics of each dominant endophytes design or query suitable medium for its growth to culture and isolation. Finally, the function of the isolates was studied. This method can prevent researchers from missing out on the important functional strains of the host, expand the research scope of endophytes of medicinal plants, and facilitate the in-depth excavation and utilization of endophytes of medicinal plants.

Effect of active peptides of Eupolyphaga on intestinal flora in rats with hyperlipemia / 中国药理学通报

Aim To investigate the effect of active peptide of Eupolyphaga (APE) on changes of intestinal flora and blood lipid in rats with hyperlipidemia. Methods SD rats fed with high-fat diet were induced to hyperlipemia model. Simvastatin being a positive drug, ELISA was used to investigate the effect of APE on blood lipids in hyperlipidemic rats. At the same time, the results of the microbial flora about APE model rats were measured using 16rS high-throughput assay technology (V4-V5). Results APE could significantly reduce the serum lipid index in model rats and improve the degree of liver pathological changes. Meanwhile, the intestinal flora results showed that when the relative abundance of hymenoscyphus, lactobacillus and bifidobacterium in firmicutes were enriched, the relative abundances of rumenococcus, plasmodium and prevotella in bacteroidetes and tenericutes were reduced. Conclusions APE could obviously reduce the level of blood lipid and repair the disordered intestinal flora of hyperlipidemia rats. It can be speculated that APE might play a role in lowering blood lipids through the intervention of lipid metabolism pathway by intestinal flora.

Airway Dysbacteriosis Exacerbated Murine Airway Allergic Inflammation / 中山大学学报(医学科学版)

[Objective] To investigate the effects of airway dysbacteriosis on the development of murine atlergic airway diseases (AAD).[Methods] Female C57BL/6 mice were neubulized with Vancomycin for 10 days and then were sacrificed.The bacterial population in bronchoalveolar lavage fluid (BALF) were evaluated using 16S rRNA high-throughput sequencing technology,exploriug the method of establishing an airway dysbacteriosis mouse model.After the mouse model was established successfully,airway dysbacteriosis mouse models were established by the same method,and based on that,the mice were sensitized and challenged with ovalbumin (OVA) to induce airway allergic inflammation.The frequency of nasal rubbing behaviors per mice was counted;the total cell number and eosinophil relative abundance in BALF were evaluated;the lung tissue inflammation and goblet cell metaplasia were assessed according to histopathological features;and the IgE level in serum,IFN-γ,IL-4 and IL-5 levels in BALF,and IL-33 levels in serum,BALF and intestine tissue were measured by ELISA.[Results] Nebulization of Vancomycin increased Bradyrhizobium,Sphingopyxis,Cupriavidus,Pelomonas,and decreased Akkermansia and Prevotella_6 in airway,inducing significant airway dysbacteriosis.Using the animal model,further study found that airway dysbacteriosis exacerbated OVA-induced airway allergic inflammation,including increased nasal rubbing frequency,higher serun IgE level,more total cell count especially eosinophil infiltration,more serious lung tissue inflammation and goblet cell metaplasia.Additionally,compared to OVA group,mice in Dysbacteriosis and OVA group had significantly increased level of Th2 cytokine IL-4 and IL-5,and significantly decreased Thl cytokine IFN-γin BALF,which revealed that mice in Dysbacteriosis and OVA group had mote remarkable Thl/Th2 imbalance.Furthermore,IL-33 level showed a significant increase in BALF,but didn't change in serum or intestine tissue in Dysbacteriosis and OVA group compared to OVA group.Indicating that airway dysbacteriosis may only affect the local production of IL-33.[Conclusions] An airway dysbacteriosis mouse model was established by Vancomycin nebulization successfully.Airway dysbacteriosis may activate innate lymphoid cells (ILC) and Th2 cell by inducing local IL-33 secreting,which leads to the imbalance of Th1/Th2,and in turn promotes the development of AAD.

miRNA library and thermodynamic stability of Astragali Radix decoction pieces / 中草药

Objective: To establish a small RNA library of Astragalus membranaceus, in order to understand the intake and absorption of miRNAs in human body, and to lay the foundation for pharmacological research of Astragali Radix decoction pieces (ARP). Methods: The decoction of A. membranaceus after desiccation, high temperature and microwave treatment was used as sample, and using high-throughput sequencing technology, the unknown miRNA sequence was obtained. Results: Totally 9931049 small RNA sequences were identified from ARP by the high-throughput sequencing technology. After compared with all hairpin-forming precursors in miRBase 21 database, the miRNA library of ARP decoction was established, and one conserved miRNA and 9 novel miRNA sequences were confirmed. Twelve class I small interfering RNAs (siRNAs) were obtained by further bioinformatic analysis and we found that the 5' terminal was unstable while the 3' terminal was stable thermodynamically. Conclusion: The miRNAs expression profiles of ARP is first revealed by thermodynamic treatment. Our study suggests that class I siRNA can be used as a potential biochemistry components of phytomedicine, which might contribute to the epigenetic studies on Chinese materia medica.

Advances research in chromatin DNaseⅠhypersensitive sites and its transcriptional regulation function / 医学研究生学报

DNaseⅠhypersensitive sites (DHSs) are regions of extreme chromatin which is highly sensitive to DNaseⅠ.Ge-nome-wide mapping DHSs is a powerful method for identifying many different types of regulatory elements within nuclear chromatin .It can help systematically resolve gene and genomic information , and the dynamic molecular mechanism of transcriptional regulation .This paper reviews the distribution of DHSs , the technology of DNase-seq and the research progress in transcriptional regulation functions of DHSs.

Analysis of BCR CDR3 repertoire of peripheral blood with HBsAb titer higher than 10 000 mU/ml / 中国免疫学杂志

Objective:To acquire potential HBsAb sequences,we have analyzed the BCR CDR3 repertoire of the peripheral blood with HBsAb titer higher than 10 000 mU/ml,which could provide a data basis for follow-up study.Methods:Genomic DNA of pe-ripheral blood mononuclear cells was extracted from samples with HBsAb titer higher than 10 000 mU/ml.We have adopted Illumina Solexa high-throughput sequencing technology of the Adaptive Biotechnologies ImmunoSEQ platform to acquire sequence data.IMGT/High V-QUEST was used to preliminary analyze our sequence data,including usage of IGHV,IGHJ and IGHD gene subgroups,IGHV-J matching,distribution of CDR3 amino acid (AA) length and usage of total CDR3 AA.And these sequences were compared with the HBsAb sequences from NCBI database.Results:Experimental samples have highly selected gene subgroups IGHV3,IGHV4,IGHJ4, IGHJ6,IGHD3,IGHD6,and IGHV3-J4 pairing,IGHV3-J6 pairing.The AA length distributions of CDR3 region were normal distribution with the length of 14/15 AA as the midline.In the regard to amino acid usage in CDR3 region, each sample prior used Alanine, Tyrosine,Glycine,Alanine,Aspartic acid and Serine.The amino acid usages of 107,108,109,113,114 positions were diversified but 105,106,115,116,117 positions taking conservative amino acids usages.We have found 48 unique sequences that have same IGHV, IGHJ and CDR3 AA length with the HBsAb sequences from NCBI database.Conclusion:There were almost the same characteristics of BCR CDR3 repertoire of the peripheral blood with HBsAb titer higher than 10 000 mU/ml.The 48 unique sequences provided a solid data basis for the follow-up study.