The most difficult field in gene therapy is that vector system should offer both a means of successful transfection and a maximum of safety for the patient. Viral vectors and plasmid vectors are traditional vectors; they may cause unwanted immunological side effects resulting from the expression of nontherapeutic genes. Our aim is to develop a new general gene therapy vector which is suggested to be called as Micro-Linear Vector. The gene expression cassette is capped by our designed cap, including promoter, enhancer, objective gene, and RNA-stabilizing sequence, so it can defend the exnuclease in the eukaryotic cell, at the same time, DNA not encoding the objective gene is reduced to a minimum. The GFP gene is separated from the pEGFP-N3 plasmid, and acts as a reporter gene to construct the Micro-Linear Vector, then both the new vector and the plasmid are transfected to cells, the results are tested by fluorescence microscope and flow cytometry. The results show that the Micro-Linear Vector has a high effective of transfection and safety in 293, 3T3, CNE2 and B95-8 cell lines, at the same time it is less toxicity than the plasmid. We can get the rudiments of conclusion that Micro-Linear Vector has high effection of the transfection and more safety than tradition plasmid in eukaryotic cell.
3T3 Cells ; Animals ; CpG Islands ; Genetic Therapy ; methods ; Genetic Vectors ; genetics ; Green Fluorescent Proteins ; genetics ; Humans ; Kidney ; cytology ; Mice ; Promoter Regions, Genetic ; genetics ; Transfection ; methods

Acute respiratory infection is a common infectious diseases in children.Rapid and accurate identification of pathogens is critical for targeted treatment and prognosis.Therefore, taking sensitive and effective detection methods is the key to pathogenic diagnosis.Now, the progress in the application and research of detection for respiratory pathogens in children were reviewed.

Sepsis is a life-threatening organ dysfunction caused by the host disordered response to infections.As one of the most important innate immune cells in the body，macrophages can maintain the immune homeostasis by recruiting other immune cells，clearing pathogens，presenting antigens，and play important regulatory roles in infectious diseases such as sepsis by releasing inflammatory factors.As a critical neurotransmitter，dopamine not only participates in the neurological processes such as learning and cognition，but also regulates the immune processes，including regulating the activation，proliferation and functional changes of immune cells such as neutrophils，lymphocytes，monocytes and macrophages.Current studies demonstrate that during the infection and inflammation process of sepsis，the phagocytosis，polarization，and release of inflammatory factors of macrophages are regulated by dopamine.This review summarized the recent research progress on the regulatory functions and the underlying mechanisms of dopamine on macrophages in sepsis.

Objective:To evaluate the clinical significance of molecular detection testing multiple pathogens in children with viral central nervous system infections.Methods:We retrospectively included 176 children who were suspected with central nervous system infection at Shanghai Children′s Medical Center from January 2017 to May 2021.Film Array Meningitis/Encephalitis Panel(FA-M/E) was used to test cerebrospinal fluid samples of these children.The results were analyzed compared with clinical symptoms and cerebrospinal fluid indices.Results:There were 34 samples with positive FA-M/E virus detection(19.32%, 34/176). Among the 34 samples, enterovirus was the most common pathogen(27 cases, 79.41%). In different combinations, the sensitivity and positive predictive value were all less than 90%.The median time for antiviral drugs used in FA-M/E virus-positive and negative children was 4.5(0, 8.5)d and 2.6(0, 2.0)d, respectively.The difference was statistically significant( P<0.05). Conclusion:Molecular tests of multiple pathogens can quickly and sensitively detect pathogens.It can improve the efficacy of clinical diagnosis of viral central nervous system infection.

Objective: To investigate the interaction between the gut microbiota and the sepsis in children by comparing the difference of gut microbiota between septic children and healthy children. Methods: Genome was extracted from excrements of 18 cases of sepsis and 6 cases of healthy children.After genomic extraction, the hypervariable region of 16S rDNA gene were amplified and a small fragment library was constructed, and high-throughput sequencing was carried out, then the data of the lower machine was effectively sequenced by biological information processing.We could seek for the species that had changed significantly due to sepsis by comparing the diversity and the differences in the composition of intestinal flora between the two groups. Results: The gut microbiome of the sepsis group was distinct from that of the health group.The operational taxonomic units in the sepsis group were significantly reduced compared with healthy group(P=0.001). The gut microbiome of children with sepsis had significantly lower diversity and richness compared with healthy group(P<0.05). A total of 7 species were shown to be differentially abundant between septic patients and healthy controls.The genus Pseudomonadales, Carnobacteriaceae and Granulicatella_elegans were significantly more abundant in the sepsis group; meanwhile the genus Pasteurellaceae, Ruminococcus, Lactobacillus_rogosae and Anaerostipes_butyratucus were less abundant in the sepsis group.In addition, the Granulicatella_elegans was characteristically present in the intestine of children with sepsis(P<0.001). Conclusion The microbial diversity and structure of the gut microbiome in children with sepsis are significantly different from those of healthy children.Our data suggest biomarkers identified in this study might participate in the pathogenesis or development process of sepsis.