ObjectiveTo explore the role of osteopontin (OPN) in hyperoxia-induced acute lung injury and its relationship with nuclear factor-κB (NF-κB),matrix metalloproteinase 2 and 9 (MMP-2,MMP-9). MethodsNinety-six mice were randomly divided into a phosphate buffer solution intranasal inhalation group (PBS group) and a recombinant OPN intranasal inhalation group. The mice were exposed in sealed cages >95% oxygen for 24-72 hours to induce lung injury or room air as control. The severity of lung injury was evaluated. The expression of NF-κB,MMP-2,MMP-9,TIMP-1 and TIMP-2 mRNA in lung tissue at 24,48 and 72 hours under hyperoxia were examined by reverse transcript-polymerase chain reaction (RT-PCR). Immunohistochemistry (IHC) was performed for detection of NF-κB protein in lung tissue. ResultsPBS group mice developed more severe acute lung injury at 72 hours under hyperoxia.TIMP-1 and TIMP-2 mRNA expressions were significantly increased in r-OPN group than their matched PBS group when exposed to hyperoxia. IHC study showed higher expression of NF-κB protein in lung tissue of PBS group at 72 hours of hyperoxia. ConclusionOPN can protect against hyperoxia-induced lung injury by inhibiting the expressions of NF-κB,MMP-2 and MMP-9.
ObjectiveTo identify the expression functions of human NF-κBp65 nuclear localization signals' deletion mutant plasmids(namely pcDNA3.1(+)-NF-κBp65ΔNLS, NF-κBp65ΔNLS, for short) and the changes of proliferation, migration and adhesion ability of A549 lung cancer cells with low expression of NF-κBp65 (namely A549/NF-κBp65 shRNA cells). MethodsHuman A549/NF-κBp65 shRNA cells were cultivated and divided into a control group, a transfection pcDNA3.1 (+) group, and a transfection NF-κBp65ΔNLS group. Indirect immunofluorescence, real-time fluorescent quantitative PCR and Western blot techniques were used to detect the NF-κBp65 intracellular localization and the change of NF-κBp65 mRNA and protein expression level. MTT, Transwell and cell adhesion experiments were used to analyze the changes of proliferation, migration and adhesion ability of A549/NF-κBp65 shRNA cells. ResultsThe human NF-κBp65ΔNLS eukaryotic expression plasmid was successfully constructed. Compared with the control group and the transfection pcDNA3.1(+) group, NF-κBp65 mRNA expression level in A549/NF-κBp65 shRNA cells was increased in the transfection NF-κBp65ΔNLS group(10.63±0.84 vs. 1.04±0.21 and 1.23±0.22, P < 0.01) and NF-κBp65 protein expression level was also increased (1.07±0.06 vs. 0.53±0.02 and 0.59±0.04, P < 0.01). NF-κBp65 protein mainly located in the cytoplasm, and did not significantly transferred into the nucleus after stimulated by TNF-α. At the same time, A549/NF-κBp65 shRNA cells' proliferation, migration and adhesion ability were enhanced compared with the control group and the transfection pcDNA3.1(+) group. ConclusionsThrough gene mutation technology to build the human NF-κBp65ΔNLS eukaryotic expression plasmid and transfect into A549/NF-κBp65 shRNA lung cancer cell lines, both mRNA and protein expression levels of NF-κBp65 were increased significantly, and NF-κBp65 protein mainly located in the cytoplasm. The overexpressed NF-κBp65 in cytoplasm can obviously enhance the A549/NF-κBp65 shRNA cell's proliferation, migration and adhesion ability. It suggests that NF-κBp65 stranded in the cytoplasm can still regulate biological behavior of lung cancer cells by influencing the NF-κB signaling pathway related proteins.
Objective To investigate the pathogen distribution and drug resistance in ICU patients, provide reference for prevention of severe infection and empirical antibacterial treatment. Methods The patients admitted in ICU between January 2013 and December 2014 were retrospectively analyzed. The pathogenic data were collected including bacterial and fungal culture results, the flora distribution and drug resistance of pathogenic bacteria. Results A total of 2088 non-repeated strains were isolated, including 1403 (67.2%) strains of Gram-positive bacteria, 496 (23.8%) strains of Gram-negative bacteria, and 189 (9.0%) strains of fungus. There were 1324 (63.42%) strains isolated from sputum or other respiratory specimens, 487 (23.33%) strains from blood specimens, 277 (13.27%) strains from other specimens. The bacteria included Acinetobacter baumannii (17.2%), Klebsiella pneumoniae (14.8%), Pseudomonas aeruginosa (9.9%), C. albicans (6.3%), E. coli (5.6%), E. cloacae (5.4%), Epidermis staphylococcus (5.0%) and Staphylococcus aureus (4.7%). There were 15 strains of penicillium carbon resistant enterobacteriaceae bacteria (CRE) accounting for 2.3%, including 5 strains of Pneumonia klebsiella, 4 strains of E. cloacae. In 117 strains of E. coli, drug-resistant strains accounted for 86.4% including 85.5% of multiple drug-resistant strains (MDR) and 0.9% of extremely-drug resistant (XDR) strains. In 359 strains of Acinetobacter baumannii, drug-resistant strains accounted for 75.2% including 72.1% of XDR strains and 3.1% of MDR strains. MDR strains accounted for 10.6% in Pseudomonas aeruginosa. Detection rate of methicillin resistant Staphylococcus aureus (MRSA) and methicillin resistant coagulase-negative Staphylococci (MRCNS) was 49.0% and 95.5%, respectively. There were 4 strains of vancomycin resistant Enterococcus faecalis. There were 131 (69.3%) strains of C. albicans, 23 (12.2%) strains of smooth candida. C. albicans was sensitive to amphotericin and 5-fluorine cytosine, and the resistance rate was less than 1% to other antifungle agents. The resistance rate of smooth ball candida was higher than C. albicans and nearly smooth candida, but still less than 15%. Conclusions The predominant pathogens in ICU was gram-negative bacteria. The top eight pathogenic bacteria were Acinetobacter baumanni, Klebsiella pneumoniae, Pseudomonas aeruginosa, C. albicans, E. coli, E. cloacae, Epidermis staphylococcus and S. aureus. Sputum and blood are common specimens. CRE accounts for 2.3%. Drug-resistant strains are most common in E. coli mainly by MDR, followed by Acinetobacter baumannii mainly by XDR, and least in Pseudomonas aeruginosa. C. albicans is the most common fungus with low drug resitance.