Objective　To study the interference effects of siRNA and dsRNA in Aedes albopictus and their impact on mosquito survival, providing reference and technical support for the study of mosquito gene function, the research and development of new insecticides, the control of vector mosquitoes, and the interruption of arboviruses transmission. 　　Methods　For genes of interest in Ae. albopictus, siRNAs targeting the desired genes were designed and commercially synthesized, while dsRNAs were obtained by amplifying the target gene fragments, followed by in vitro transcription and purification, with verification by agarose gel electrophoresis. The interfering RNA was injected into Ae. albopictus by thoracic microinjection. Then the expression of targeted genes on the 3rd day post-microinjection was detected by quantitative PCR to verify the interfering effect, and the survival curve of the mosquitoes was recorded at the same time. Results　In the siRNA interference experiments targeting 10 genes with 22 siRNA pairs, only two siRNA pairs, SR-C-2 siRNA and ENR-1 siRNA, successfully interfered with the target gene expression compared to the control group, with interference rates of 58% and 33%, respectively, while the remaining siRNAs did not show interference effects, with some even causing increased expression of the target gene. In the dsRNA interference experiments, Mesh dsRNA significantly reduced the expression of mesh gene at the concentration of 3×1011 RNA molecules per mosquito, with an interference effect lasting 5 days and an interference rate of 55%-60% on the third day and 34% on the fifth-day post-injection. Mosquito survival within 3 days post-dsRNA injection was significantly higher than in the siRNA injection group. Conclusions　Ae. albopictus on the fifth-day post-emergence is the most suitable to receive thoracic microinjection. dsRNA is more stable than siRNA for interference, with higher post-injection survival rates, making it more suitable for in-depth studies of mosquito gene function. Although siRNA-based gene interference is simpler and more feasible, attention should be paid to the purification method of siRNA and its homology with other genes in the mosquito species to avoid non-specific experimental results due to the off-target combinations.

Objective: To detect the expression of serum exosomal miR⁃30d⁃5p in occupational dermatitis medicamentosa⁃like of trichloroethylene(OMDT) patients and its correlation with liver function , then perform bioinformatics analysis and verify the target gene. Methods : Serum exosomes were extracted from 6 OMDT patients and 6 healthy controls , and miRNA was extracted from exosomes which were identified by transmission electron microscopy, nanoparticle tracking analysis and western blotting. The expression of serum exosomal miR⁃30d⁃5p was detected by real⁃time fluorescence quantitative PCR , then the correlation with liver function was analyzed. The target genes of miR⁃30d⁃5p were predicted by the miRWalk and miRBD databases. Gene ontology analysis and KEGG pathways analysis were performed. Finally , RHOB was verified by Dual⁃luciferase reporter assay. Results: The expression of serum exosomal miR⁃30d⁃5p significantly decreased in 6 OMDT patients at the peak of the disease(P < 0. 05) , and it was negatively correlated with the level of AST , ALT and GGT (correlation coefficient : rs = - 0. 943 ,P = 0. 005 ;rs = - 0. 886 , P = 0. 019 ; rs = - 0. 886 , P = 0. 019 ) . Bioinformatics analysis and dual⁃luciferase reporter assay showed that RHOB was the target gene of miR⁃30d⁃5p. Conclusion The expression of serum exosomal miR⁃30d-5p decreases in OMDT patients , which is negatively correlated with the level of liver function , and the target gene RHOB may be involved in the process of liver injury induced by trichloroethylene.

Objective: To investigate the association of SNPs in TET2 gene with the susceptibility and prognosis of sepsis. Methods: Ninety-nine patients diagnosed with sepsis and 107 controls were enrolled in the study. The septic patients were further divided into survivors (56 cases) and non-survivors (43 cases) according to the outcome of 28-day hospitalization. Patients without sepsis after major surgery were enrolled as the controls. The genotypes of the five loci (rs6839705, rs7670522, rs7679673, rs7698522 and rs10010325) with high minor allele frequency in the TET2 were screened according to the existing research reports and the SNP database of the NCBI website. The five loci were detected by TaqMan probe based allelic discrimination assays using real-time polymerase chain reaction (PCR). The data were calculated for genetic association study through χ² test and Fisher’s exact probability method. Results: There was no significant difference in genotype frequencies of the five tested SNPs in TET2 gene between septic patients and controls or between survivors and non-survivors in septic patients (P > 0.05). Furthermore, the allelic frequencies of the five SNPs between septic patients and controls or between survivors and non-survivors in septic patients also had no significant difference (P > 0.05). Conclusions This study showed that the five SNPs in TET2 gene (rs6839705, rs7670522, rs7679673, rs7698522, and rs10010325) were not associated with the susceptibility and prognosis of sepsis, which needs to be further confirmed by large-sample studies.

To investigate the association between single nucleotide polymorphism (SNP) in the 11th exon of transient receptor potential melastatin 2 (TRPM2) gene with the susceptibility and outcome of sepsis.A total of 119 septic patients and 112 normal subjects were enrolled from the First Affiliated Hospital, Zhejiang University School of Medicine. Among 119 septic patients, 62 died (fatal group) and 57 survived (survival group) within 28 days of disease onset. The genotypes of these individuals were detected using TaqMan allelic discrimination assays, and its correlations with susceptibility and outcome of sepsis were analyzed.There was no significant difference in genotype frequencies and allelic frequencies of TRPM2 SNP rs1556314 between septic patients and the controls (all>0.05). And no significant difference in genotype frequencies and allelic frequencies of TRPM2 SNP rs1556314 was observed between the survivors and fatal cases of septic patients (all>0.05).The TRPM2 SNP rs1556314 does not have significant association with sepsis, but this result need to be confirmed by large scale studies.
Exons ; genetics ; physiology ; Female ; Gene Frequency ; Genetic Predisposition to Disease ; genetics ; Genotype ; Humans ; Male ; Polymorphism, Single Nucleotide ; genetics ; physiology ; Sepsis ; genetics ; mortality ; TRPM Cation Channels ; genetics

Objective To study the changes of trans-diaphragmatic pressure (Ptra) and its correlation with esophageal pressure (Peso) through ARDS piglet model.Methods Five piglets were enrolled in the study.Peso,gastric pressure (Pgas) and intra-thoracic pressure (Pint) was monitored through balloon inserted.The data before ARDS serve as control.ARDS was produced in the piglets through saline lavage.The pressure were observed and the Ptra were calculated.The pressure changes and correlation between Ptra and Peso were analyzed as well.Linear regression with the coefficient of determination and t-test were used as appropriate.Significance was assumed for P ＜ 0.05.Results Peso,Pgas and Pint before ARDS were 7.3 ± 1.9,25.5 ± 2.4,- 1.23 ± 0.21 cmH2O,Ptra was 18.2 ± 1.6 cmH2O.While after ARDS,the data were 4.7 ± 1.4,31.1 ± 3.1 and - 1.79 ± 0.28 cmH2O,and Ptra was 26.4 ± 2.1 cmH2 O,and all these changes were obviously ( P ＜ 0.05 ).The correlation between Pint and Peso,Pint and Ptra (A) and Ptra ( B ) were 0.93 ± 0.025,0.88 ± 0.023 and 0.87 ± 0.37 before ARDS.After ARDS,the correlation changed to be 0.82 ±0.21,0.81 ±0.20 and 0.78 ±0.31.Although a bit decreased,the correlation was still positive (P ＜ 0.01 ).Conclusions There existed good correlations between Peso and Ptra as well as between Pint and Peso before or after ARDS.Ptra was increased obviously after ARDS,which could lead to respiratory muscle fatigue.

This study examined the effects of a combined surgery of sleeve gastrectomy (SG) and modified jejunoileal bypass (JIB) on the body weight, food intake, and the plasma levels of active glucagon-like peptide-1 (GLP-1) and total ghrelin of rats. Rats were divided into 3 groups in terms of different surgical protocol: SG-JIB (n=12), SG (n=12), JIB (n=12) and sham surgery groups (n=10). In SG-JIB group, rats was subjected to sleeve gastrectomy and end to side anastomosis of part of the jejunum (25 cm from the ligament of Treitz) to the ileum 25 cm proximal to the cecum. The body weight and food intake were evaluated during 10 consecutive weeks postoperatively. The levels of active GLP-1 and total ghrelin in the plasma of the rats were measured by ELISA assay. The results showed that the SG-JIB treated rats relative to SG- or JIB-treated ones produced a sustained reduction in food intake and weight gain. The level of active GLP-1 was elevated and total ghrelin level decreased in SG-JIB-treated rats as compared with SG- or JIB-treated ones. It was concluded that SG-JIB could efficiently reduce the body weight and food intake, alter obesity-related hormone levels of the rats, indicating that SG-JIB may be potentially used for the treatment of obesity.

This study examined the effects of a combined surgery of sleeve gastrectomy (SG) and modified jejunoileal bypass (JIB) on the body weight,food intake,and the plasma levels of active glucagon-like peptide-1 (GLP-1) and total ghrelin of rats.Rats were divided into 3 groups in terms of different surgical protocol:SG-JIB (n=12),SG (n=12),JIB (n=12) and sham surgery groups (n=10).In SG-JIB group,rats was subjected to sleeve gastrectomy and end to side anastomosis of part of the jejunum (25 cm from the ligament of Treitz) to the ileum 25 cm proximal to the cecum.The body weight and food intake were evaluated during 10 consecutive weeks postoperatively.The levels of active GLP-1 and total ghrelin in the plasma of the rats were measured by ELISA assay.The results showed that the SG-JIB treated rats relative to SG- or JIB-treated ones produced a sustained reduction in food intake and weight gain.The level of active GLP-1 was elevated and total ghrelin level decreased in SG-JIB-treated rats as compared with SG- or JIB-treated ones.It was concluded that SG-JIB could efficiently reduce the body weight and food intake,alter obesity-related hormone levels of the rats,indicating that SG-JIB may be potentially used for the treatment of obesity.

Objective To investigate the correlation between gene polymorphism within human β defensin 1 (DEFB1) and fungal susceptibility to severe sepsis through case-control association study.Methods A total of211 patients with severe sepsis in ICU were enrolled in the present case control study. Sepsis in this study was diagnosed according to the definition of American College of Chest Physicians/Society of Critical Care Medicine Consensus Conference in 1992 and 2002. Based on the development of fungal infection during ICU stay, all 211 patients were divided into fungal infection group (Group Ⅰ) and control group (Group C). Alleles and genotypes of-1816A/G, -390A/T, -52A/G, -44C/G and-20A/G within DEFB1 gene were assayed in all 211 patients by means of DNA direct sequencing, Allele-specific PCR amplifications or high-throughput site-specific TaqMan assay. Genetic analysis was employed to calculate the distribution frequency of haplotypes. The correlation between the genomic variations (allele,genotype and haplotype) and fungal infection was analyzed by Chi-square test or Fisher's exact test.Odds ratio (OR) was employed to reflect the correlation degree of genetic factor with fungal susceptibility to severe sepsis. Results Group Ⅰ enrolled 80 patients, of whom 43 pstients were male, at age of (60.81 ± 18.30) years. Group C enrolled 131 patients, of whom 80 patients were male, at mean age of (60.42 ± 17.03) years. No significant difference was found between two groups in aspect of gender and age (P＞0.05). The genetic locus of -1816A/G, -390A/T, -52A/G, -44C/G and -20A/G of both groups were in agreement with Hardy Weinberg equilibrium. No significant difference was found between two groups in the distribution of allelic frequencies and genotype frequencies (P ＞0.05). No significant difference was found in the distribution frequency of four common haplotypes of the above five genetic locus such as AAACG, ATGCA, GTGGG and ATACG (all P ＞ 0.05). Conclusions Genetic locus of -1816A/G, -390A/T, -52A/G, -44C/G and-20A/G within DEFB1 gene have no correction with fungal infections in severe sepsis, suggesting that DEFB1 gene polymorphism may not serve as a key genetic marker for the predisposition to fungal infection in severe sepsis.

Objective To observe the effect of LIGHT on synovial macrophage-osteoclast difier entiation in rheumatoid arthritis.Methods Synovial macrophages were collected from 8 synovial tissues har vested from RA patients by digestion with conagenase.The macrophages from each patient were divided into 5 subgroups:group 1 was co-cultured with Macrophage Colony Stimulating Factor(MCSF).group 2 was co cultured with MCSF and LIGHT.group 3 was co-cultured with MCSF and Receptor activator for Nuclear Fac tor kB ligand(RANKL),group 4 was co-cultured with MCSF and LIGHT and RANKL group 5 was co-cul tared with LIGHT.After two weeks of culture.tartrate resistant acid phosphatase(TRAP)and F-actin staining were used to detect the formation of osteoclast on cover slip(CS).Functional evidence of osteoclasts was as sessed by the formation of resorption pits ou dentine slice(DS).Results TRAP and F-actin were both nega tive in group 1 and group 5 and no pit on dentine slice could be observed.In group 2.there were some small round and ovoid osteoclasts with TRAP(+)and F-actin(+).Pits on DS were small and discrete.In group 3. there were many large irregular osteoclasts with TRAP(++)and F-actin(++).The number and volume of pits were both increased.In group 4.there were even more and larger osteoclasts with TRAP(+++)and F-actin (+++).The pits were even larger and became confluent,ie,pits(++++).Conclusion LIGHT can promote RANKL-mediated osteoclastogenesis in RA synovial macrophages.and it can induce osteoclast formation through a mechanism independent of RANKL.