Effects of electroacupuncture on rats with cognitive impairment: An iTRAQ-based proteomics analysis / 中西医结合学报

OBJECTIVE@#The study explores the effects of electroacupuncture (EA) at the governing vessel (GV) on proteomic changes in the hippocampus of rats with cognitive impairment.@*METHODS@#Healthy male rats were randomly divided into 3 groups: sham, model and EA. Cognitive impairment was induced by left middle cerebral artery occlusion in the model and EA groups. Rats in the EA group were treated with EA at Shenting (GV24) and Baihui (GV20) for 7 d. Neurological deficit was scored using the Longa scale, the learning and memory ability was detected using the Morris water maze (MWM) test, and the proteomic profiling in the hippocampus was analyzed using protein-labeling technology based on the isobaric tag for relative and absolute quantitation (iTRAQ). The Western blot (WB) analysis was used to detect the proteins and validate the results of iTRAQ.@*RESULTS@#Compared with the model group, the neurological deficit score was significantly reduced, and the escape latency in the MWM test was significantly shortened, while the number of platform crossings increased in the EA group. A total of 2872 proteins were identified by iTRAQ. Differentially expressed proteins (DEPs) were identified between different groups: 92 proteins were upregulated and 103 were downregulated in the model group compared with the sham group, while 142 proteins were upregulated and 126 were downregulated in the EA group compared with the model group. Most of the DEPs were involved in oxidative phosphorylation, glycolipid metabolism and synaptic transmission. Furthermore, we also verified 4 DEPs using WB technology. Although the WB results were not exactly the same as the iTRAQ results, the expression trends of the DEPs were consistent. The upregulation of heat-shock protein β1 (Hspb1) was the highest in the EA group compared to the model group.@*CONCLUSION@#EA can effect proteomic changes in the hippocampus of rats with cognitive impairment. Hspb1 may be involved in the molecular mechanism by which acupuncture improves cognitive impairment.

Sintomas de ansiedade generalizada entre estudantes de graduação: prevalência, fatores associados e possíveis consequências / Symptoms of generalized anxiety among undergraduate students: prevalence, associated factors and possible consequences

RESUMO Objetivo Medir a prevalência de sintomatologia ansiosa entre estudantes de graduação de uma universidade pública no Sul do Brasil, seus fatores associados e possíveis consequências. Métodos Trata-se de um estudo transversal com uma amostra aleatória de graduandos, realizado em 2019. Foi utilizado um questionário com questões sociodemográficas, acadêmicas e de saúde mental e física. Sintomas de ansiedade generalizada foram avaliados por meio do Generalized Anxiety Disorder - 7 (GAD-7). Fatores associados e possíveis consequências foram analisados por meio da regressão de Poisson com ajuste robusto da variância. Resultados Entre os 994 participantes, a prevalência de sintomas de ansiedade generalizada severa foi de 30,9% (IC 95%: 27,9%-33,9%). As variáveis independentemente associadas a sintomas de ansiedade generalizada foram: sexo feminino, orientação não heterossexual, sentir muito/muitíssimo medo de violência no bairro e perceber como difícil o acesso a serviço psicológico. Possuir alto nível de suporte social e apresentar algum nível de atividade física foi considerado fator de proteção para o desfecho. Possíveis consequências dos sintomas de ansiedade generalizada foram: pior qualidade de sono, utilizar algum medicamento sem prescrição nos últimos 30 dias e ser classificado como risco de suicídio moderado/severo. Conclusões Esses resultados demonstram a necessidade de um olhar mais atento para alguns subgrupos populacionais. Recomenda-se a implementação de intervenções psicológicas que incluam um número maior de indivíduos simultaneamente (como a criação de grupos terapêuticos), com o intuito de reduzir a carga de ansiedade no contexto universitário. O fortalecimento do setor de assistência estudantil da universidade também se torna uma importante possibilidade de melhora para esse cenário.

ABSTRACT Objective To measure the anxious symptoms of undergraduate students at a public university in southern Brazil, the factors associated with it and the possible consequences. Methods This was a cross-sectional study with a random sample of undergraduate students in 2019. A questionnaire that evaluated sociodemographic, academic, mental and physical health aspects was used. Symptoms of generalized anxiety were evaluated through the Generalized Anxiety Disorder - 7 (GAD-7). Associated factors and possible consequences were analyzed using the Poisson regression, with robust variance adjustment. Results Among the 994 participants, the prevalence of symptoms of severe generalized anxiety was 30.9% (95% CI: 27,9%-33,9%). The variables independently associated with symptoms of generalized anxiety were: female gender, non-heterosexual orientation, feeling very afraid of violence in the neighborhood, perceiving access to psychological service as difficult. Having a high level of social support, and with any level of physical activity were considered protective factors for the outcome. Possible consequences of symptoms of generalized anxiety were: worse sleep quality, non-medical use of prescription drugs in the last 30 days, and being classified with moderate or severe suicide risk. Conclusions These results indicate the need to a closer attention to subgroups within this population. Implementation of psychological interventions that includes a broader range of individuals simultaneously are recommended (such as therapeutic groups) to reduce the burden of anxiety in the university context. The strengthening of the university's student assistance sector is also important to improve this situation.

Specific PCR Identification of Hibisci Cortex and Its Adulterants Based on DNA Signature Sequence Tags / 中国实验方剂学杂志

ObjectiveUncommon medicinal herbs are valuable medicinal resources， but their identification is a difficult problem in Chinese medicine due to their particularity and complexity. It is， therefore， urgent to establish a method for the identification of uncommon medicinal herbs. In this study， DNA signature sequence （DSS） tags were used to establish a specific polymerase chain reaction （PCR） identification method for Hibisci Cortex， the origin plant of Hibisci Cortex， and its adulterants. MethodThe candidate DSS tags were obtained from the chloroplast genome sequence analysis， and the DSS tags were verified by DNA sequencing. The specific identification primers for H. syriacus were designed based on the obtained reliable DSS tags. The PCR reaction conditions were optimized， and the tolerance and feasibility were investigated. ResultA DSS tag for identification of H. syriacus was obtained from the comparison of sequencing results of the amplified products with DSS， which revealed the distinguishing characteristics of Hibisci Cortex and its adulterants. A pair of specific primers for H. syriacus was designed according to the DSS tag. After PCR amplification and gel electrophoresis with the primers， a single bright band of about 270 bp was observed from H. syriacus， which did not appear in the four adulterants. ConclusionA DSS tag obtained in this study can be used to identify H. syriacus. The specific primers designed based on this DSS tag can accurately and simply identify the original plant of Hibisci Cortex and its adulterants， which provides a new method and idea for the molecular identification of genuine and counterfeit products of Hibisci Cortex.

Proteomic analysis of extraocular muscles from patients with thyroid-associated ophthalmopathy by tandem mass tag / 中华实验眼科杂志

Objective:To identify the proteins differentially expressed in extraocular muscles between restrictive strabismus patients with thyroid-associated ophthalmopathy (TAO) and concomitant esotropia patients by proteomic analysis using tandem mass tag (TMT).Methods:Extraocular muscles samples from 5 restrictive strabismus patients with TAO and 5 concomitant esotropia patients were collected at Peking University People's Hospital from August 2019 to December 2020.All the patients received strabismus surgery.Differentially expressed proteins (DEPs) in extraocular muscles samples were identified by quantitative proteomic analysis and bioinformatic analysis based on TMT.Fold change≥1.2 or≤0.83 and P value<0.05 was regarded as the threshold to screen DEPs.GO annotation, KEGG pathways enrichment analysis and protein-protein interaction (PPI) network of DEPs were conducted through UniProtGOA and STRING.This study protocol was approved by the Ethics Committee of Peking University People's Hospital (No.2021PHB058-001). Results:A total of 53 DEPs were identified, 34 of which were up-regulated and 19 were down-regulated.The biological processes DEPs mainly participated included response to stimulation, multicellular organismal process, metabolism, developmental process, intracellular signal transduction, and positive regulation of biological process.DEPs were involved in pathways including focal adhesion, tight junction, regulation of action cytoskeleton, and apoptosis.Six key proteins identified using PPI network were myosin heavy chain 2, myosin heavy chain 7, myosin regulatory light chain, α-actinin-2, fibrinogen alpha chain and fibrinogen beta chain.Conclusions:There are DEPs in extraocular muscles between restrictive strabismus patients with TAO and concomitant esotropia patients.Myosin, actinin and filamin may be involved in the pathogenesis of TAO through regulation of actin cytoskeleton and focal adhesion.

Proteomic analysis of lung injuries induced by paraquat in mice / 中华急诊医学杂志

Objective:To explore the molecular mechanism of paraquat (PQ)-induced lung injuries.Methods:Male C57BL/6 mice aged 6 to 8 weeks were randomly divided into four groups. Mice in the experimental groups (three groups, nine rats in each group) were intraperitoneally injected with 40 mg/kg PQ to establish an infection model, and mice in the control group ( n=9) were intraperitoneally injected with the same dose of saline. Mice were sacrificed at day 2, 7 and 14 after PQ administration. Pathological changes of lung tissues from mice model were observed by Hematoxylin-eosin staining. The expression of different proteins in the lung tissues at different time points were detected and identified by tandem mass spectrometry tag technology (TMT), and the functional analysis was performed. Results:Compared with the control group, there were 91 (69 up and 22 down), 160 (103 up and 57 down) and 78 (45 up and 33 down) proteins in the PQ-2 d, 7 d, and 14 d groups, respectively, and there was significant difference of protein expression . The subcellular localization analysis showed that compared with the control group, the differentially-expressed proteins in the PQ-2 d and -7 d groups were mainly distributed in the extracellular space, while in the PQ-14 d group were mainly distributed in the nuclear. GO analysis showed that compared with the control group, the differentially-expressed proteins in the PQ-2 d and PQ-7 d groups were mainly involved in humoral immunity and coagulation-related reactions, while in the PQ-14 d group were mainly involved in chemotactic and regulatory responses such as neutrophil aggregation. The KEGG signaling pathway analysis showed that the complement and coagulation cascades was the most important pathway in the PQ-2d and PQ-7 d groups, while metabolism of xenobiotics by cytochrome P450 was the most important pathway in the PQ-14 d group.Conclusions:It is the first time that TMT was used to analyze PQ-induced lung injuries in mice model at different time points. This study demonstrates the molecular mechanism of PQ-induced lung injuries at protein levels, and elucidates that humoral immunity and complement-coagulation pathways charge the main role of PQ-induced lung injuries. This study may provide an important theoretical basis for further research and clinical treatment.

Analysis of protein differences in aortic aneurysm/dissection based on tandem mass tag proteomics / 中国胸心血管外科临床杂志

@#Objective    To analyze the differences in proteins between aneurysm/dissection patients and healthy subjects, and subsequently figure out differential proteins related to medial degeneration of aortic aneurysm/dissection. Methods    Aortic wall samples were collected from 6 male aortic aneurysm patients (an aortic aneurysm group, mean age 56.50±8.19 years), 6 male aortic dissection patients (an aortic dissection group, mean age 54.17±6.68 years) and 6 male healthy subjects (a normal group, mean age 40.50±9.31 years) between December 2019 and May 2020 in West China Hospital of Sichuan University. Quantitative proteomics was performed using tandem mass tag (TMT) techniques, followed by gene ontology (GO) analysis and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis. Results    A total of 63 differential proteins were obtained both in the aortic aneurysm group and the aortic dissection group compared with the normal group, with 30 up-regulating and 33 down-regulating. The differential proteins were involved in multiple biological processes and clusted on peroxisome proliferators-activated receptor (PPAR) signaling pathway, extracellular matrix-receptor interaction signaling pathway and complement and coagulation cascades signaling pathway. Conclusion    The identified proteins may help to demonstrate new molecular mechanisms related to medial degeneration of aortic aneurysm/dissection.

Production of antimicrobial peptide (Oxysterlin 1) in Escherichia coli with ELP self-cleavage tag / 生物工程学报

Antimicrobial peptides are the most promising alternatives to antibiotics. However, the strategy of producing antimicrobial peptides by recombinant technology is complicated and expensive, which is not conducive to the large-scale production. Oxysterlin 1 is a novel type of cecropin antimicrobial peptide mainly targeting on Gram-negative bacteria and is of low cytotoxicity. In this study, a simple and cost-effective method was developed to produce Oxysterlin 1 in Escherichia coli. The Oxysterlin 1 gene was cloned into a plasmid containing elastin-like polypeptide (ELP) and protein splicing elements (intein) to construct the recombinant expression plasmid (pET-ELP-I-Oxysterlin 1). The recombinant protein was mainly expressed in soluble form in E. coli, and then the target peptide can be purified with a simple salting out method followed by pH changing. The final yield of Oxysterlin 1 was about 1.2 mg/L, and the subsequent antimicrobial experiment showed the expected antimicrobial activity. This study holds promise for large-scale production of antimicrobial peptides and the in-depth study of its antimicrobial mechanism.

De novo transcriptome assembly and mining of EST-SSR markers in Gloriosa superba

Gloriosa superba is an economical source of pharmaceutical colchicine, which is a mitotic poison used to treat gout, cancer and inflammatory diseases. It is important to study the genetic variations in this plant, but the progress is impeded due to limited number of molecular markers. In this study, we developed the expressed sequence tag-derived simple sequence repeat (EST-SSR) markers from the transcriptome sequence of the leaf samples of three different ecotypes of G. superba. De novo assembly was performed on these sequencing data to generate a total of 65,579 unigenes and 38,200 coding sequences (CDSs). These CDSs were annotated using NCBI Nr protein database, gene ontology terms and KEGG pathways. Differential gene expression was studied to yield differences in these ecotypes at the molecular level. Finally, a total of 14,672 potential EST-SSRs were identified from these unigenes, among which the dinucleotide (5754, 39.22%) and trinucleotide (5421, 36.95%) repeats were most abundant types followed by mononucleotides (3213, 21.83%). The most frequent motifs were CT/GA (1392, 9.48%), AG/TC (1219, 8.31%), and GA/CT (1146, 7.82%) among the dinucleotide repeats and CCG/ CGG (1487, 10.13%), AGG/CCT (1421, 9.68%), AGC/CTG (697, 4.75%) and AAG/CTT (621, 4.23%) among the trinucleotide repeats. Polymorphism study using a random set of 20 newly developed EST-SSRs revealed polymorphic information content value ranging from 0 to 0.5926 with an average of 0.4021. The large-scale ESTs developed in the current study will be useful as a genomic resource for further investigation of the genetic variations in this species

Comparison of protein profiles of the pellicle, gingival crevicular fluid, and saliva: possible origin of pellicle proteins

BACKGROUND: The pellicle, the acellular organic material deposited on the surface of tooth enamel, has been thought to be derived from saliva. In this study, protein compositions of the pellicle, gingival crevicular fluid, and saliva collected from healthy adults were compared to elucidate the origin of pellicle proteins. RESULTS: The pellicle, gingival crevicular fluid, and saliva from the parotid gland or mixed gland were collected; subsequently, protein expression in samples from the respective individual was compared by SDS-PAGE and mass spectrometry. Following SDS-PAGE, proteins in the major bands were identified by mass spectrometry. The band pattern of pellicle proteins appeared different from those of gingival crevicular fluid, or saliva samples. Using mass spectrometry, 13 proteins in these samples were identified. The relative abundance of the proteins was quantitatively analyzed using mass spectrometry coupled with stable isotope labeling and by western blot. Cystatin S and α-amylase detected in pellicle were enriched in saliva samples, but not in gingival crevicular fluid, by western blot, and their abundance ratios were high in saliva and low in gingival crevicular fluid when analyzed by stable isotope labeling. Serotransferrin, however, was found only in the pellicle and gingival crevicular fluid by western blot and its abundance ratio was low in saliva. CONCLUSIONS: Our study revealed that the gingival crevicular fluid appears to contribute to pellicle formation in addition to saliva.

Tratamento do TAG nas terapias cognitivas de terceira geração / GAD treatment third generation cognitive therapies

Esta rápida revisão tem como objetivo levantara literatura sobre o tratamento do transtorno de ansiedade generalizada (TAG) pelas terapias cognitivas de terceira geração e, com isso, estimular o entendimento de clínicos e pacientes sobre essa abordagem recente de tratamento. Os estudos, em sua maioria, demonstram que as preocupações e ruminações são o processamento principal do TAG. É uma condição multifacetada em nível cognitivo, físico e emocional. O tratamento apresentado por essas técnicas integrativas mostrou-se relevante, tendo como consequência a diminuição dos sintomas. Já os efeitos do tratamento foram mantidos após o seu término por um período maior em comparação à TCC. Apenas a terapia do esquema e a terapia comportamental dialética não retornaram dados suficientes. As evidências fornecem, portanto, um arcabouço teórico para os clínicos utilizarem de intervenções sistematizadas quanto ao TAG, com técnicas cognitivas, meditativas e emocionais.(AU)

This brief review aims to review the literature on the treatment of generalized anxiety disorder (GAD) by third generation cognitive therapies and thereby stimulate the understanding of clinicians and patients about this recent approach to treatment. Studies generally showed that concerns and ruminations are GAD's main processes. It is a multifaceted condition on the cognitive, physical and emotional level. The treatment presented by these integrative techniques was relevant, with a decrease in symptoms and the treatment effects were maintained after the treatment ended for a longer period compared to CBT. Only schema therapy and dialectical behavior therapy did not show sufficient data. Therefore, the evidence provides a theoretical enough for clinicians to use systematic GAD interventions with cognitive, meditative and emotional techniques.(AU)

La presente breve revisión tiene por objetivo revisar la literatura sobre el tratamiento del Trastorno de Ansiedad Generalizada (TAG) por terapias cognitivas de tercera generación y, con ello, estimular el entendimiento de clínicos y pacientes sobre dicho abordaje reciente del tratamiento. En su mayoría, los estudios demuestran que las preocupaciones y reflexiones son el procesamiento principal del TAG. Es una condición multifacética a nivel cognitivo, físico y emocional. El tratamiento presentado por estas técnicas integrativas se mostró relevante, y su consecuencia trajo una disminución de síntomas. Por otro lado, los efectos del tratamiento se sostuvieron luego del término del tratamiento por un periodo mayor comparado al de la TCC. La Terapia de Esquema y la Terapia Dialéctico-Conductual fueron las únicas a no ofrecer datos suficientes. Así, las evidencias proveen un marco teórico para que clínicos utilicen intervenciones sistémicas para el TAG, con técnicas cognitivas, mediáticas y emocionales.(AU)

A new quantitative 16S rRNA amplicon sequencing method / 生物工程学报

In recent years, 16S rRNA amplicon sequencing technology has been widely used to study human gut microbiota and to detect unknown pathogens in clinical samples. However, its resolution to bacterial population can only reach the relative abundance of genus level, and different factors affect the final bacterial profile, such as sample concentrations, PCR cycle numbers and amplification primers. In order to solve these problems, we developed a quantitative 16S rRNA amplicon sequencing method by combining random tag and internal marker method. The new methods improved the accuracy of human gut microbiota, reduced the impact of experimental operation on the results, and improved the comparability between sequencing and other molecular biological methods.

Three-dimensional chromosome conformation capture and its derived technologies / 生物工程学报

Linear chromatin is compacted into eukaryotic nucleus through a complex and multi-layered architecture. Consequently, chromatin conformation in a local or long-distance manner is strongly correlated with gene expression. Chromosome conformation capture (3C) technology, together with its variants like 4C/5C/Hi-C, has been well developed to study chromatin looping and whole genome structure. In this review, we introduce new technologies including chromosome capture combined with immunoprecipitation, nuclei acid-based hybridization, single cell and genome sequencing, as well as their application.

Construction of full-length cDNA library from Antrodia cinnamomea mycelium and EST sequence analysis / 中草药

Objective: To excavate the terpenoid synthesis and metabolism-related gene function and screen the interaction protein and fingerprint analysis of Antrodia cinnamomea mycelium, a cDNA library from A. cinnamomea mycelia was constructed and the EST sequences were analyzed. Methods: The cDNA library from the A. cinnamomea mycelium was constructed by the Gateway technique. A part of EST sequences about the bioinformatics, functional annotation and EST-SSR were analyzed. Results: The cDNA library of the A. cinnamomea mycelium was constructed successfully. The recombinant rate of the cDNA library was 95%, the titer of the library was 6.1 × 106 cfu/mL, the total cloning number was 1.2 × 107 cfu, the length of cDNA was between 300-2 000 bp with an average length of 1 000 bp. The clones were randomly sequenced and 65 valid ESTs were obtained. After being compared in the Genbank database, 45 ESTs had a definite annotation, and 18 ESTs were unnamed and hypothetical protein. The results with GO functional annotation showed that the ESTs involved the cell composition, transport, catalytic activity, regulation functions and etc. It contained 271 SSRs of all the ESTs in total. The nucleotide repeats in A. cinnamomea were abundant, among which dinucleotide and trinucleotide repeat units were more common accounting for 94.23%. Conclusion: The cDNA library from the A. cinnamomea mycelium and its ESTs related biological information were preliminarily identified, which will provide a theoretical foundation for research the mycelium genomics of A. cinnamomea.

Proteomic analysis of vascular access in patients with chronic kidney disease at stage 5 / 西安交通大学学报(医学版)

Objective: By applying the proteomic method, to analyze differences of protein expressions and signal pathways in veins at arteriovenous fistula in chronic kidney disease stage 5 (CKD5) patients with or without diabetes so as to explore the pathogenesis of high incidence of arteriovenous fistula functional incapacitation in CKD5 patients with diabetes. Methods: The protein expression RAW datasets of vascular access from CKD5 patients with or without diabetes in Proteomexchange Database were screened out and downloaded. Then the identified features were searched from UniProt/SwisssProt human proteins database through the software ProteomeDiscovery (PD). Then, the PD generated a file of quantitative proteins data. The significantly different proteins between two groups were screened out and analyzed by T-test or Adj T-test depending on homogeneity of variance. These significantly different proteins were enriched into different biological pathways through IPA analysis, GO enrichment analysis, and KEGG pathway analysis, which signifies these biological pathways are significantly different. Ultimately, the correlation of proteins in different biological pathway was analyzed by Spearman correlation test. Results: TheiTRAQ labeled proteins RAW datasets comparing the vessels from CKD5 patients with diabetes and without diabetes (PXD010883) were collected at first. After searching identified features again and disposing the data, a total of 120 significantly different proteins including 89 up-regulated proteins and 31 down-regulated proteins were screened out finally. Through GO, KEGG and IPA analyses, there were two significantly different biological pathways. On the one hand, the purine nucleoside monophosphate metabolic change and the ratio of ATP/AMP decrease were reflected in oxidative phosphorylation receding and AMP metabolism enhancing in CKD5 patients with diabetes. On the other hand, the muscle system process which played a vital role in VSM cells receded in CKD5 patients with diabetes. It was specific in the decrease of MYH11, CNN1 and so on. Excitingly, the significantly differential genes in the two pathway had a strong correlation. The results explained why CKD5 patients with diabetes always have cardiovascular disease. Conclusion: The ratio of ATP/AMP reduction caused by diabetes led to muscle function disorder of VSM cells, which explains the reason for the internal fistula functional incapacitation in CKD5 patients with diabetes.

A large fibroma polyp of labia majora

Fibroepithelial stromal polyps are benign proliferations which are usually polypoid or pedunculated, and less than 5cm in size. They are generally single lesions but can be multiple during pregnancy. They can be polypoid or pedunculated and are usually solitary. Symptoms usually include bleeding, discharge and general discomfort with sensation of a mass. They typically have a central fibrovascular core and contain stellate and multinucleated stromal cells which are best seen beneath the surface epithelium. True myxoid stroma is absent. Although vulvovaginal fibroepithelial stromal polyps are well documented, a giant variant such as the one authors report here is rather rare. To the knowledge, present case is the largest fibroepithelial stromal polyp compared to others reported in the literature.

Optimization of a microarray for fission yeast

Bar-code (tag) microarrays of yeast gene-deletion collections facilitate the systematic identification of genes required for growth in any condition of interest. Anti-sense strands of amplified bar-codes hybridize with ~10,000 (5,000 each for up- and down-tags) different kinds of sense-strand probes on an array. In this study, we optimized the hybridization processes of an array for fission yeast. Compared to the first version of the array (11 µm, 100K) consisting of three sectors with probe pairs (perfect match and mismatch), the second version (11 µm, 48K) could represent ~10,000 up-/down-tags in quadruplicate along with 1,508 negative controls in quadruplicate and a single set of 1,000 unique negative controls at random dispersed positions without mismatch pairs. For PCR, the optimal annealing temperature (maximizing yield and minimizing extra bands) was 58℃ for both tags. Intriguingly, up-tags required 3× higher amounts of blocking oligonucleotides than down-tags. A 1:1 mix ratio between up- and down-tags was satisfactory. A lower temperature (25℃) was optimal for cultivation instead of a normal temperature (30℃) because of extra temperature-sensitive mutants in a subset of the deletion library. Activation of frozen pooled cells for >1 day showed better resolution of intensity than no activation. A tag intensity analysis showed that tag(s) of 4,316 of the 4,526 strains tested were represented at least once; 3,706 strains were represented by both tags, 4,072 strains by up-tags only, and 3,950 strains by down-tags only. The results indicate that this microarray will be a powerful analytical platform for elucidating currently unknown gene functions.

Proteomic analysis of the cerebrospinal fluid from patients with amyotrophic lateral sclerosis based on tandem mass spectrometry technique / 南方医科大学学报

OBJECTIVE@#To investigate the differentially expressed proteins in the cerebrospinal fluid (CSF) of patients with amyotrophic lateral sclerosis (ALS) at the proteomics level using tandem mass spectrometry label (TMT) technique and explore the pathogenic mechanism and related pathways of ALS.@*METHODS@#Between November, 2017 and April, 2018, 5 patients with medulla oblongata onset ALS and 5 patients with limb onset ALS were selected from the Departments of Neurology of 928 Hospital of Army Joint Logistics Support Force of PLA and Xiangya Hospital of Central South University, with 5 patients with migraine and low intracranial pressure headache serving as the healthy controls.CSF samples were obtained from all the participants, and the differentially expressed proteins in the CSF were identified using tandem mass spectrometry (TMT) technique with bioinformatics analysis.@*RESULTS@#A total of 1530 proteins were identified and quantified in the CSF samples.The expression of 48 proteins was up-regulated and 6 proteins were down-regulated in medulla oblongata onset ALS patients; 16 proteins were up-regulated and 19 were down-regulated in limb onset ALS patients.GO analysis showed that these proteins, which were distributed both within and outside the cells, were involved in cell physiological process, single organ process and biological regulation and had binding function, catalytic activity, and receptor activity.KEGG pathway analysis showed that the up-regulated proteins in the CSF from patients with medulla oblongata onset ALS participated in 3 pathways involving the lysosomes, metabolism, and measles.The down-regulated proteins in the CSF from patients with limb onset ALS participated in 7 pathways involving the complement and coagulation cascade, infection and herpes simplex infection, and all the pathways contained complement components.@*CONCLUSIONS@#The CSF samples of ALS patients with medullary onset and limb onset have differentially expressed proteins.The lysosomal pathway is involved in the occurrence and progression of ALS with medullary onset, and the immune responses are involved in the occurrence and progression of ALS with limb onset.

Lucy-tag promotes expression of sea lamprey (Petromyzon marinus) olfactory receptor on cell membrane / 中国药理学通报

Aim To elucidate the effect of Lucy-tag polypeptide label on the expression of olfactory receptor membrane in sea lamprey ( Petromyzon marinus). Methods Immunocytochemis-try, double luciferase system and calcium flow detection were used to verify the effect of polypeptide label Lucy-tag on olfactory receptors of sea lamprey. Results Polypeptide label Lucy-tag promoted the membrane expression of 13 olfactory receptors out of 20 olfactory receptors, and Lucy-tag did not affect the intrinsic activity of olfactory receptors, nor did it affect the IP3 signaling pathway of G protein coupled receptor (GPCR). Conclusions It provides a favorable technique for finding the ligand of olfactory receptor and lays a good theoretical foundation for the further study of the function of the GPCR and ligand.

Expression, purification and NETs degradation activity of human DNase I / 中国药科大学学报

@#In order to study the effect of human deoxyribonuclease I(DNase I)on neutrophil extracellular traps(NETs)degradation, genetic engineering bacteria E. coli Rosetta(DE3)/pET32a-His-DNase I was constructed. The fusion protein His-DNase I was induced by lactose, and purified by Ni-affinity chromatography. Mouse neutrophils were extracted and stimulated with phorbol-12-myristate-13-acetate(PMA)to form NETs. The degradation activity of the fusion protein on NETs was detected by SytoxGreen and fluorescence microscopy. The results showed that the purified His-DNase I had high nuclease activity. This study provided the research foundation for further exploration of the clinical application of DNase I.

Analysis of risk factors of decreased heart rate variability in patients with type 2 diabetes mellitus / 上海交通大学学报（医学版）

Objective • To analyze the related risk factors of decreased heart rate variability (HRV) in type 2 diabetes mellitus (T2DM) patients. Methods • A total of 210 cases with T2DM in Shanghai General Hospital, Shanghai Jiao Tong University School of Medicine from Jan. 2013 to Dec. 2014 were divided into two groups by the levels of urinary albumin creatinine ratio (UACR) and serum triacylglycerol (TAG), and with/without hypertension, respectively. By comparing the differences of clinical characteristics, HRV parameters and other general clinical data between the two groups, the correlations between HRV indexes and their risk factors were explored. Results • The average heart rate of the whole day was higher, and SDNN [normal RR intervals of sinus beats (NN intervals) standard deviation] was lower in the high UACR group than those in the normal UACR group (both P<0.05). The average heart rate of the whole day was higher, while SDNN and pNN50 (percentage of total heart beats with adjacent NN intervals greater than 50 ms) were lower in the hypertriglyceridemia group than those in the normal TAG group (all P<0.05). Minimum frequency domain power hour was lower in the hypertension group than that in the normotension group (P<0.05). Regression analysis showed that the linear correlativity between TAG and SDNN was very prominent (P<0.05), as well as between the diastolic blood pressure (DBP) and the average heart rate of the whole day, SDNN, frequency domain power 24 hours and minimum frequency domain power hour (all P<0.05). Similarly, the age was linearly correlated significantly with the average heart rate of the whole day, SDNN, SDANN (the standard deviation of the mean NN intervals every 5 min), frequency domain power 24 hours, minimum frequency domain power hour and maximum frequency domain power hour (all P<0.05). Conclusion • Age, TAG level and hypertension are the risk factors of decreased HRV in T2DM patients. Thus, early detection of the decrease of HRV in these patients can prevent the cardiovascular events of T2DM.