ABSTRACT
Objective To assay the level of neutrophil extracellular traps (NETs) in systemic lupus erythematosus (SLE) patients and healthy controls and compare the difference between SLE group and control group, and to analyze between the level of NETs and related laboratory parameters. Methods Forty-four females patients with SLE were recruited as research subjects, with 24 cases in the active stage and 20 in stable stage. Forty-two healthy female volunteers matched in age were enrolled as control subjects. The fluorescence intensity of NETs in neutrophils was detected by fluorospectrophotometry. The concentration of neutrophil elastase in plasma was quantitatively detected. Statistical analysis of the difference of level of NETs between the SLE group and control group was conducted. Then the correlation between the fluorescence intensity of NETs and erythrocyte sedimentation rate (ESR) and C-reactive protein (CRP), and anti-double-stranded DNA (dsDNA), and anti-histone, and anti-nucleosome, and systemic lupus erythematosus disease activity index (SLEDAI) was analyzed respectively. The same tests were conducted for the level of NE. The results of the two groups were compared using analysis of variance and the relevance was analyzed using Spearman correlation analysis. NETs morphosis was observated by immunoflurescence method, and the difference of NETs between SLE group and control group was analyzed. Results ① The fluorescence intensity of NETs was significantly increased in SLE group(241±139) than that in the control group (173±135), (t=2.31, P0.05] and NE concentration [(104±43) vs (96±48), t=0.50, P>0.05] between SLE active stage and stable stage was detected. ③ The fluorescence intensity of NETs in SLE patients was positively correlated with SLEDAI, but had no obvious correlation with anti-dsDNA, anti-histone, anti-nucleosome, CRP or ESR, respectively. ④Images under confocal microscope showed that more NETs generated by PMN from SLE patients than that from controls. Conclusion The generation of NETs is enhanced in female patients with SLE. And NETs may relate to disease activity. However, NETs may not induce the production of autoantibodies.
ABSTRACT
Objective To investigate the effect of neutrophil elastase (NE) inhibitor on the blood-brain barrier(BBB) permeabili-ty and hydrocephalus in rats with traumatic brain injury (TBI) .Methods 99 SD rats were randomly divided into the control group , the TBI group and the intervention group(dividing into 5 sub-groups:6 ,24 ,48 ,72 ,168 h) .The hydraulic impact model of rats was duplicated .Sivelestat sodium was given in the intervention group .The NE concentration in the brain tissue ,BBB permeability and brain water content were detected in each group and performed the comparative analysis .Results The NE concentration in the brain tissue ,BBB permeability and brain water content at each timepoint in the TBI group and the intervention groups were higher than those in the control group .The NE concentration at 24 ,48 ,72 ,168 h in the intervention group was lower than that in the TBI group .The BBB permeability and the brain water content at 48 ,72 ,168 h in the intervention group were lower than those in the TBI group(P<0 .05) .Conclusion Sivelestat sodium can inhibit the NE release in TBI rat brain tissue ,reduce the BBB permeability and the occurrence of hydrocephalus ,which indicating that sivelestat sodium has the protective effect on TBI secondary lesion in rat .
ABSTRACT
Objective To investigate the efficiency of biology function of ITIH4 gene silenced by small interfering RNA (siRNA) on ovarian cancer.Methods The four pairs ITIH4 gene siRNA interference fragments(ITIH4-546,ITIH4-795,ITIH4-917 and ITIH4-1568) were designed respectively,and transfected into HO8910pm cells with ITIH4 mRNA high expression by liposomal method transiently.Quantitative PCR method was used to detect the ITIH4 mRNA expression in HO8910pm cells transfected with interference fragment.The ITIH4 917 was selected as the best silencing effect of siRNA interference fragment and then the recombinant plasmid expression vector pGPU6/GFP/Neo-shRNA-ITIH4-917 was constructed and transfected into HO8910pm cells.The stably transfected cells-pGPU6/GFP/Neo-shRNA-ITIH4-917-HO8910pm cells was obtained by screening of aminoglycoside antibiotics (G418).The experiment was divided into three groups,namely ITIH4-917 transfection group,the HO8910pm cell group transfected with pGPU6/GFP/Neo-shRNA plasmid (empty vector group),and the HO8910pm cell group transfected with pGPU6/GFP/Neo-shRNA-ITIH4-NC the plasmid (negative control group).Fluorescence quantitative reverse transcription(RT)PCR and western blot were used to detect the ITIH4 mRNA and protein expression.The cell proliferation,the cell cycle,colony formation of cells,cells migration and invasion in vitro were determined by using methyl thiazolyl tetrazolium (MTT),flow cytometry,colony formation assay and transmembrane (transwell) small chamber method [value represented by absorbance (A)],respectively.Results The fluorescent quantitative PCR results showed that the ITIH4 mRNA expression levels in ITIH4-917 HO8910pm cells was significantly lower than that in the control cells,the relative copy number was only 0.26 ± 0.15.Also the relative copy number of ITIH4 mRNA in ITIH4-917 transfection group cells was 0.34 ±0.10,it significantly lower than that in empty vector group (1.87 ±0.12,P =0.008) and negative control group (1.58 ±0.21,P =0.032) ; Western blot results showed that the ITIH4 relative expression levels of the protein in ITIH4-917 HO8910pm group cells,empty vector group and negative control group were 0.51,1.64 and 1.74,respectively,there were statistically significant differences (0.51 vs.1.64,P =0.012; 0.51 vs.1.74,P =0.014).MTT colorimetric assay showed that the proliferation of ITIH4-917 HO8910pm group cells was significantly faster than that in the empty vector group and negative control group,and there were statistically significant differences among them (P =0.001).The S ± G2/M phase cell ratio in ITIH4-917 HO8910pm group cells was 54.2％,which was significantly higher than that in the empty vector group or negative control group (26.3％ and 31.3％,respectively,all P ＜ 0.05).The colony formation rate (55.7 ± O.7) ％ in ITIH4-917 HO8910pm group cells was also significantly higher than that in empty vector group (29.7 ±0.9) ％ (P =0.037) and negative control group (31.4 ± 0.3) ％ (P =0.043).Migration and invasion experiments showed that cell migration in ITIH4-917 HO8910pm group cells was 0.40 ± 0.18,whicht was significantly higher than that in the negative control group or empty vector (0.30 ±0.03,P =0.031 ;0.25 ±0.03,P =0.028,respectively).Although the invasive ability of ITIH4-917 HO8910pm group cells (1.31 ±0.34) was higher than that in the control cells (1.05 ±0.68) and empty vector group (1.14 ±0.08),while there were not significant difference (P ＞ 0.05).Conclusion It would be to promote the cell doubling time and increase the migration capability in HO8910pm cells that ITIH4 expression was down-regulating by ITIH4 mRNA interference.