RESUMEN

Enterovirus A71 (EV-A71) is a neurotropic human pathogen which mainly caused hand, foot and mouth disease (HFMD) mostly in children under 5 years-old. Generally, EV-A71-associated HFMD is a relatively self-limiting febrile disease, but there will still be a small percentage of patients with rapid disease progression and severe neurological complications. To date, the underlying mechanism of EV-A71 inducing pathological injury of central nervous system (CNS) remains largely unclear. It has been investigated and discussed the changes of mRNA, miRNA and circRNA expression profile during infection by EV-A71 in our previous studies. However, these studies were only analyzed at the RNA level, not at the protein level. It's the protein levels that ultimately do the work in the body. Here, to address this, we performed a tandem mass tag (TMT) peptide labeling coupled with LC-MS/MS approach to quantitatively identify cellular proteome changes at 24 h post-infection (hpi) in EV-A71-infected 16HBE cells. In total, 6615 proteins were identified by using TMT coupled with LC-MS/MS in this study. In the EV-A71- and mock-infected groups, 210 differentially expressed proteins were found, including 86 upregulated and 124 downregulated proteins, at 24 hpi. To ensure the validity and reliability of the proteomics data, 3 randomly selected proteins were verified by Western blot and Immunofluorescence analysis, and the results were consistent with the TMT results. Subsequently, functional enrichment analysis indicated that the up-regulated and down-regulated proteins were individually involved in various biological processes and signaling pathways, including metabolic process, AMPK signaling pathway, Neurotrophin signaling pathway, Viral myocarditis, GABAergic synapse, and so on. Moreover, among these enriched functional analysis, the "Proteasome" pathway was up-regulated, which has caught our attention. Inhibition of proteasome was found to obviously suppress the EV-A71 replication. Finally, further in-depth analysis revealed that these differentially expressed proteins contained distinct domains and localized in different subcellular components. Taken together, our data provided a comprehensive view of host cell response to EV-A71 and identified host proteins may lead to better understanding of the pathogenic mechanisms and host responses to EV-A71 infection, and also to the identification of new therapeutic targets for EV-A71 infection.

Asunto(s)

Enterovirus Humano A , Infecciones por Enterovirus , Enterovirus , Enfermedad de Boca, Mano y Pie , Niño , Humanos , Preescolar , Enterovirus Humano A/fisiología , Cromatografía Liquida , Complejo de la Endopetidasa Proteasomal , Proteómica , Reproducibilidad de los Resultados , Espectrometría de Masas en Tándem , Infecciones por Enterovirus/metabolismo , Replicación Viral/fisiología , Células Epiteliales , Péptidos , Proteoma

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Objective:To analyze differentially expressed microRNAs (miRNAs) and target genes in human respiratory epithelial cells (16HBE) after enterovirus 71 (EV71) and coxsackievirus A16 (CVA16) infection using high-throughput sequencing.Methods:TargetScan and miRDB databases were used to predict the target genes of miRNAs that were both up-regulated or down-regulated after EV71 and CVA16 infection. The genes that were both up-regulated and down-regulated were screened out. GO and pathway analysis of the target genes were conducted to screen immune-related target genes and their corresponding miRNAs. The target genes and their corresponding miRNAs that were up-regulated or down-regulated in both immune-related GO and pathway were further screened. Some miRNAs and their target genes were selected for qRT-PCR verification.Results:There were 598 target genes of up-regulated miRNAs and 1 311 target genes of down-regulated miRNAs and 62 target genes that might be up-regulated or down-regulated simultaneously were screened out. The number of up-regulated target genes involved in immune-related GO and pathway were 17 and 13, respectively, and the number of corresponding miRNAs were 15 and 17, respectively. There were 58 and 47 down-regulated target genes involved in immune-related GO and pathway, respectively, and the number of corresponding miRNAs were 30 and 42, respectively. Three up-regulated target genes were involved in both immune-related GO and pathway and regulated by four miRNAs. Nine down-regulated target genes were involved in both immune-related GO and pathway and regulated by 13 miRNAs.Conclusions:This study was conducive to elucidate the host-pathogen interaction after EV71 and CVA16 infection, and provided reference for studying the pathogenesis of hand, foot and mouth disease.

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Objective To investigate the role of CXC chemokine ligand 5 (CXCL5) in the patho-genesis of dextran sulfate sodium (DSS)-induced inflammatory bowel disease (IBD). Methods A mouse model of IBD was established by giving 3% DSS in drinking water. Influences of CXCL5 knockout on mouse body weight, clinical symptoms, survival rate, pathological injury and the secretion of inflammatory cyto-kines were analyzed. Results CXCL5 levels in serum of mice with DSS-induced IBD were significantly higher than those of the normal control group. DSS-induced weight gain, death, pathological damages and inflammatory cytokine secretion were alleviated in mice after knocking out CXCL5. Conclusion CXCL5 might promote the secretion of inflammatory cytokines in mice with DSS-induced acute colitis and aggravate pathological damages,suggesting that CXCL5 might be a potentially important candidate target for the treat-ment of IBD.

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Objective To provide a comprehensive reference index for different mouse models of herpes simplex virus 1 (HSV-1) infection by investigating the related clinical manifestations, pathological features and characteristics of viral distribution in tissues and organs of BALB/c mice infected with different HSV-1 strains by using different strategies.Methods Acute infection models were established by challenging BALB/c mice at age three or six weeks with HSV-1 17+ and McKrae strains via intranasal and corneal administrations.Correspondingly, chronic infection models were established with BALB/c mice through subcutaneous and foot pad injections.Results Although all experimental mice showed trichiasis and roachback, there were differences in weight and fatality rate among different groups.Results of the quantitative PCR detection indicated that the proliferation of HSV-1 in the nervous tissues (brain, spinal cord, trigeminal ganglion) varied among different groups.The pathological examination indicated that in the acute infection groups, significant pathological changes only occurred in the brain tissues, while in the chronic infection groups, pathological injuries only occurred in the trigeminal ganglia.Although a key index latency-associated transcript (LAT) was not detected in the trigeminal nerve tissues of mice in the chronic infection groups, co-culturing the tissues with Vero cells resulted in infectious lesions in the cells.Conclusion This study indicates that there are significant differences in weight and fatality rate among different BALB/c mouse models of HSV-1 infection.Varied replication dynamics of HSV-1 were observed in different tissues or organs of the BALB/c mice in different groups.Therefore, different indexes should be adopted to evaluate different HSV-1 infection models.

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Employee motivation is to meet their needs and improve their productivity and work enthusiasm for the organization.Therefore,an accurate understanding of their needs is a prerequisite for the implementation of effective motivation.In view of this,we conducted a questionnaire survey of incentive factors for young medical staff at a public hospital.This study aimed at analyzing different incentive factors among medical staff with different seniority and job categories as well as their differences,in an effort to provide references for fine human resource management and motivation implementation at public hospitals.

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The unsatisfactory operation of the existing mechanism for resolving medical disputes by doctors and patients may be contributed to the instability of this mechanism, as lack of regulation constraints results in huge discrepancies of outcomes between identical cases. Specifically, the uncertainty for doctor-patient consultation in the guidelines, evaluation and decision mechanism,consultation process, and compensation calculation. In this consideration, it is recommended to elevate the mechanism stability of this consultation by means of uniform guidelines, third-party evaluation and decision, standardized consultation process, and unified compensation standard.

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Aim To investigate the effects of antioxidant probucol on vascular smooth muscle cells(VSMCs) apoptosis induced by H2O2.Methods H2O2 (1 mmol?L-1) was used to induce VSMCs apoptosis.The VSMCs were treated with probucol(100,10,1 ?mol?L-1) for 6 hours.For the evaluation of apoptosis,Annexin V-FITC staining,Hoechest33258 staining and the TUNEL assay were used.The expressions of ASK-1 and Trx-1 were detected by Western blot analysis.Results H2O2 could promote the apoptosis of VSMCs,increase the expression of ASK-1 and decrease the expression of Trx-1.Probucol could attenuate the apoptosis induced by H2O2 in a dose-dependent,down-regulate ASK-1 expression and increase Trx-1 expression.Conclusion Probucol can antagonize the apoptosis of VSMCs induced by H2O2.The mechanism may be correlated with a decreased expression of ASK-1 and an increased expression of Trx-1.