ABSTRACT
Background@#Somatic cell nuclear transfer (SCNT) is used widely in cloning, stem cell research, and regenerative medicine. The type of donor cells is a key factor affecting the SCNT efficiency. @*Objectives@#This study examined whether urine-derived somatic cells could be used as donors for SCNT in pigs. @*Methods@#The viability of cells isolated from urine was assessed using trypan blue and propidium iodide staining. The H3K9me3/H3K27me3 level of the cells was analyzed by immunofluorescence. The in vitro developmental ability of SCNT embryos was evaluated by the blastocyst rate and the expression levels of the core pluripotency factor. Blastocyst cell apoptosis was examined using a terminal deoxynucleotidyl transferase dUTP nick end-labeling assay. The in vivo developmental ability of SCNT embryos was evaluated after embryo transfer. @*Results@#Most sow urine-derived cells were viable and could be cultured and propagated easily. On the other hand, most of the somatic cells isolated from the boar urine exhibited poor cellular activity. The in vitro development efficiency between the embryos produced by SCNT using porcine embryonic fibroblasts (PEFs) and urine-derived cells were similar.Moreover, The H3K9me3 in SCNT embryos produced from sow urine-derived cells and PEFs at the four-cell stage showed similar intensity. The levels of Oct4, Nanog, and Sox2 expression in blastocysts were similar in the two groups. Furthermore, there is a similar apoptotic level of cloned embryos produced by the two types of cells. Finally, the full-term development ability of the cloned embryos was evaluated, and the cloned fetuses from the urine-derived cells showed absorption. @*Conclusions@#Sow urine-derived cells could be used to produce SCNT embryos.
ABSTRACT
Objectives To investigate characterization of imipenem resistance among imipenem non susceptible Pseudomonas aeruginosa isolated from patients who treated without imipenem and explore risk factors of imipenem resistance.Methods From April,2006 to March,2008,a total of 37 non-susceptible to imipenem without imipenem therapy isolates were collected from affiliated 3201st Hospital of Medical College of Xi'an Jiaotong University.The minimum inhibition concentration (MIC) to 11 antimicrobial agents were determined by the broth dilution method.We also tested imipenem MIC combined with efflux pump inhibitor PAβN.PCR was performed to check for the presence of carbapenem-hydrolylzing MBL genes and oprD gene.The expression level of oprD2 and ampC were evaluated by qRT-PCR.Molecular typing was performed using PFGE.Results There is significant difference ( t =- 2.9004,P ＜ 0.01 ) of the average number days of therapy between with two or more antibiotics in the 16 patients (20.0 ± 9.5 ) d and that with only one antibiotic in the other 21 patients ( 12.6 ± 4.4 ) d before imipenem-non-susceptible strains were isolated.In all 37 strains,32 strains showed resistance to more than three antibiotics.The MBL gene ( IMP-9 ) was only found in one strain,but its phenotype is negative,oprD2 gene from the 29 strains were found forward inserted by ISpa1328.Thirty-five isolated were considered to have no oprD expression.The patterns of the total DNA of 37 strains appeared six PFGE types.The 26 strains belonged to C2 PFGE type.In the presence of PAβN,all 37 strains increased sensitivity to meropenem.Conclusion Fluoroquinolones and cephalosporins treatment could play an important role in imipenem non-susceptible production in the research isolates.