Humans ; Hypoxia-Ischemia, Brain ; diagnosis ; therapy ; Infant, Newborn ; Infant, Premature

Asphyxia Neonatorum ; therapy ; Humans ; Infant, Newborn ; Infant, Newborn, Diseases ; Neonatology ; methods ; Resuscitation ; methods

Asphyxia Neonatorum ; complications ; Humans ; Hypothermia, Induced ; methods ; trends ; Hypoxia-Ischemia, Brain ; diagnosis ; therapy ; Infant, Newborn ; Neurologic Examination

Bronchoalveolar Lavage ; Bronchopulmonary Dysplasia ; etiology ; therapy ; Humans ; Infant, Newborn ; Infant, Premature ; Infant, Very Low Birth Weight ; Intubation, Intratracheal ; adverse effects ; Male ; Positive-Pressure Respiration ; Respiratory Insufficiency ; etiology ; therapy ; Treatment Outcome

Electrocardiography ; Female ; Fetal Hypoxia ; complications ; Humans ; Infant, Newborn ; Myocardial Infarction ; etiology ; therapy ; Myocardium ; metabolism ; pathology ; Treatment Outcome

0.05),but the injury was more serious in hilar bile duct compared with those of the proximal and distal common bile ducts(P

Objective To construct eukaryotic expression plasmid pEE14.1-dsFv?pr+,and detect the expression of the recombined gene in eukaryotic CHO-K1 cells.Methods The cationic DNA fragment was cloned into the 3' of VH gene by overlapping extension PCR,and the 6?His tab was inserted to the 3' of VL and human IFN-? gene by the same way.The above mentioned recombinant VH and VL genes were inserted into a pCI-GPI vector first,and then cloned into the pEE14.1 vector to construct the recombinant plasmid pEE14.1-dsFv?pr+.Finally,the recombinant plasmid was transfected into the CHO-K1 cells by LipofectamineTM 2000,and the expression was detected by RT-PCR,ELISA and Western blotting.Results The enzyme digestion and sequencing analysis showed that the recombinant plasmid was successfully constructed.RT-PCR showed that only the cells with transfected plasmid can generate the specific 1700bp fragment.ELISA analysis showed that the production of IFN-?expressed in the supernatant of transfected cells was about 1.1ng/ml.Also,Western blotting could reveal the characteristic band of HBsAg dsFv?pr+ protein.Conclusion The antibody targeting to human IFN-?genes has been successfully expressed in a single open reading frame.Changing the electricity of the antibody may provide the necessary condition for the study of the a new type of anti-HBV drug in nanoscale in the future.

Objective To construct the prokaryotic expression plasmid of human prostate stem cell antigen (PSCA),to induce the expression of GST-PSCA fusion protein in E. coli BL21,and to identify the purified recombinant fusion protein. Methods The fragment of PSCA gene was amplified by PCR,and then cloned into the pGEM-T Easy vector. The transitional plasmid Teasy-PSCA was identified by DNA sequencing. The PSCA gene was digested from the plasmid Teasy-PSCA by restrictive enzyme BamH I and Sal I,and then inserted into the pET42a vector which contains a glutathione s-transterase (GST) tag. Following the double restriction enzyme digestion,the recombinant plasmid pET42a-PSCA was obtained and transformed into E. coli BL21 (DE3). The expression of GST-PSCA fusion protein was induced with IPTG. The recombinant fusion protein was purified by passing over a Glutathione Sepharose 4B column,and was identified by SDS-PAGE and Western blotting. Results The length of amplified PSCA gene fragment was consistent with that expected,and the sequence was correct as exemplified by the PSCA gene reported in GenBank. The result of enzyme digestion indicated that the prokaryotic expression plasmid pET42a-PSCA was successfully constructed. After transformation with pET42a-PSCA and induction with IPTG,the recombinant target protein of about 43kD was obtained. The GST-PSCA fusion protein was correctly identified by SDS-PAGE and Western blotting. Conclusions The prokaryotic expression plasmid of human PSCA gene has been successfully constructed. The GST-PSCA fusion protein may express and be purified in E. coli BL21,and it lays a foundation for further study on the anti-prostate cancer gene vaccine.

BACKGROUND: Studieshave shown that long time of warm ischemia or cold preservation would injury the biliary tract in liver transplantation. However, whether relative warm ischemia (RWI) of biliary tract would result in bile component changes is unclearly. OBJECTIVE: To establish auto-liver transplantation bile ducts RWI models, observe the effects of RWI on the bile salts and phospholipid concentration secreted by the donor liver, and to study the correlation between the total bile salt/phosphoUpid ratio (TBA, PL ratio) and billary tract injury. METHODS: A total of 32 SD rats were selected for auto-liver transplantation models with bile ducts RWI, and the rats were randomly divided into 4 groups (n=8). In Group Ⅰ (sham operation group), rats only received liver dissociation without any cold reperfusion. The RWI time of Group Ⅱ,Ⅲ, and Ⅳ were 0 minutes, 30 minutes and 60 minutes, respectively. The concentration of TBA in bile was measured with enzymatic cycling assay, andPL with enzymic colorimetric. Pathological observation with light microscope and ultrastructural observation with transmission electron microscope were performed on the hilar bile duct. The endothelial cell apoptosis was detected with TUNEL assay. The correlation between TBA, PL ratio and biliary injury was analyzed. RESULTS AND CONCLUSION: One rat died, the other 31 rats were included in the final analysis. RWI could change the composition of bile secreted by donor liver, raise the TBA/PL ratio, and increase the bile toxicity. These changes had a positive correlation to RWI time, and the changes were obviously with time prolonged. In addition, the changes are closely related to the biliary tract injury. This study shows an important mechanism of the biliary tract injury caused by RWI-injury.

Objective To develop a method for rapid and accurate detection and identification of bacterial pathogens directly from body fluid specimens and to evaluate its application in clinical laboratory.Methods Bacteria DNA was extracted from 205 body fluid specimens with column-based kit,and the high variable V1 and V3 regions of bacterial 16S rRNA gene were amplified with broad-range primers.Amplicons were analyzed by pyrosequencing and the generated sequences were searched in the bacterial identification database.Traditional culture-biochemical method was also used for these specimens and the results were taken as the golden standard.SPSS 11.0 was used to calculate the sensitivity,specificity,false positive/negative rate,positive/negative predictive value and positive/negative likelihood rate of pyrosequencing method.Results The positive rate of bacteria culture was 39.5％ (81/205),among which 71 were infected with single bacterium,and 10 were infected with two species of bacteria.Compared with the culture identification results,pyrosequencing had a 100.0％ (71/71) concordance when applied to detect and identify bacterial pathogens from specimens with single specie bacterium infected.To specimens with two species bacteria infected,7 out of 10 specimens were in concordance with the culture identification results.Besides,pyrosequencing detected 10 positive specimens and identified bacterial pathogens infected in the 124 culture-negative specimens.Taken bacteria culture as the standard method,the sensitivity of pyrosequencing for identifying bacterial pathogen in body fluid was 100.0％,and with a specificity of 91.9％,the false positive rate was 8.1％,the false negative rate was 0.0％,the positive predictive value was 89.0％,the negative predictive value was 100.0％,and the positive and the negative likelihood rate were 12.4 and 0,respectively.Conclusion Pyrosequencing can be used to detect and identify bacterial pathogens directly from body fluid specimens with the advantages of rapidity,high sensitivity,high accuracy and high throughput.