Objective: To establish a quality control method for Gene-Viral Therapy system CNHK200-hEndostatin. Methods: According to "The Guideline on Quality Control Methods of Human Gene Therapy Products", a quality control method for adenovirus was set up, which consisted of adenovirus protein identification, genomic identification, hEndostatin gene identification, hEndostatin protein quantitation, adenovirus particle quantitation, virus titer quantitation, determination of particle to PFU ratio, purity detection, AdWT detection, host cell DNA residue detectiong bovine serum protein residue detection, etc. The purified adenovirus product of CNHK200-hEndostatin was subjected to the above process. Results: We set up a SDS-PAGE method to identify the adenovirus protein and identified adenovirus genomic DNA by restriction endonuclease enzyme analysis. Human endostatin gene was identified by PCR method and its expression product was quantitated by ELISA. The number of adenovirus particle was quantitated by D260 method and HPLC method. The purity of adenovirus was determined by D260/D280 method and HPLC method. A PCR reaction was introduced to detect AdWT and hybridization method was used in host cell DNA residue detection. Bovine serum protein residue was detected by reverse indirect hemagglutination assay. All these methods were confirmed feasible in adenovirus quality control and our purified adenovirus sample was eligible in all test items. Conclusion: A series of basic methods have been successfully established for quality control of gene-viral therapy system CNHK200-hEndostatin. Our modified method for adenovirus particle quantitation by HPLC is more rapid and accurate than traditional method. The established methods have been approved in testing the purified adenovirus samples.

Objective　To investigate clinical features of neonatal ventilator associated pneumonia.Methods　Retrospective analysis of clinical features was performed in twenty-two newborns with ventilator associated pneumonia.Results　Ventilator associated pneumonia were found in 22 ou t of 36 neonates with mechanical ventilation.Conclusion　The incidence of VAP is significantly increased if the time of mechanical ventilation in longer than 96 hours.

Objective By selecting the prescriptions of treating blood group incompatibility in modern literature, and exploring their regularity, to establish the overall thought of traditional Chinese medicine in treatment.Methods 90 articles on maternal-fetal blood group incompatibility were found.Statistical methods were adopted to summarize high frequently used medicines and formula. Results High frequently used medicines were Virgate Wormwood Herb, Baical Skullcap Root, Rhubarb, Cape Jasmine Fruit, White peony Alba, Chinese Angelica,Largehead Atractylodes Rhizome,Motherwort Herb,and Milkvetch Root, which were mentioned for 238 times in the literatures, occupying 72.79% of the total frequency. Conclusion Based on this result,heat-clearing and dampness-removing, activating blood circulation to eliminate blood stasis was setup as the principle for treating this disease.

Background Studies showed that macophenolic acid (MPA)down-regulates and inhibits the expression and secretion of tissue growth factor and inflammatory factor,and further impacts the proliferation and inflammation process.Pterygium is an inflammatory and proliferative lesion.Whether MPA has an inhibitory effect on pterygium is unclear.Objective This study was to investigate the antifibrotic effects of macophenolic acid on pterygium fibroblasts(PFBs) in vitro and discuss its mechanism.Methods Pterygium tissue was obtained from pterygium patient during the surgery.PFBs were cultured using explants and identified with vimentin immunohistochemisty.0,0.125,0.250,0.500,1.000 μmol/L MPA were added into the culture medium,respectively,and the cells were cultured in the medium without MPA as the control group.MTT colorimetry was used to find the optimization effective concentration of MPA and evaluate their inhibitory effect on PFBs,and BrdU fluorescence staining was used to assess the growth statue of PFBs.Expressions of nuclear factor-κB(NF-κB),p65 and inhibitor of NF-κB-α(IκB-α) in the cells were detected by Western blot.Results The cells was spindle in shape 3 days after cultured and showed the vortex and radial arrangement with the positive response to vimentin.With the increase of MPA,the proliferative value of PFBs (A560)showed gradually decline,with a significant difference among the five groups (F =42.874,P＜0.01).In addition,the proliferative value of PFBs (A560) significantly lowed as the prolong of MPA active time(F=26.038,P＜0.01).BrdU fluorescence staining showed a significant decrease of DNA synthesis of PFBs with the elevation of MPA dose among the five groups(F=175.279,P＜0.05),and the A560of PFBs DNA synthesis in different concentrations of MPA groups was lower than that of the control group (all at P＜0.05).No apoptotic and necrotic cell was found after MPA action by DAPI staining.The expression level of p65 in the PFBs was 0.886±0.072 and 1.542±0.124 in the MPA group and the control group,indicating a declined value in the MPA group(P＜0.05).However,the expression value of IκB-α in the cytoplasm PFBs was significantly higher in the MPA group compared with the control group(2.141 ±0.305 vs.1.559±0.267) (P＜0.05).Conclusions MPA has an inhibitory effect on the growth of PFBs,which probably is related to the arresting of NF-κB pathway.

OBJECTIVETo investigate the effect of Shenkang Injection (SI) on hypertrophy and expressions of p21 and p27 in glomerular mesangial cells of rats cultured in high glucose fluid.

METHODSCultured rat glomerular mesangial cells were divided into 6 groups: the normal control group, mannitol-treated group, high glucose-treated group, high, middle and low dose SI treated groups. The synthesis of cell protein and DNA were determined with incorporation of 3H-TdR and 3H-Leu. The changes of p21 mRNA and expressions of p21 and p27 protein were determined by reverse transcriptase polymerase chain reaction and Western blot.

RESULTSHigh glucose caused incorporation of 3H-TdR reduced, incorporation of 3H-Leu increased. SI could decrease incorporation of 3H-Leu, increase incorporation of 3H-TdR. Compared with the high glucose-treated group, SI could inhibit the overexpression of p2l mRNA and protein,p27 protein induced by high glucose.

CONCLUSIONSI could inhibit hypertrophy of glomerular mesangial cells cultured in high glucose partly through inhibiting overexpressions of p2l mRNA, p21 and p27 protein.

Animals ; Cell Enlargement ; Cells, Cultured ; Culture Media ; Cyclin-Dependent Kinase Inhibitor p21 ; metabolism ; Cyclin-Dependent Kinase Inhibitor p27 ; metabolism ; Drugs, Chinese Herbal ; pharmacology ; Glucose ; pharmacology ; Mesangial Cells ; cytology ; drug effects ; metabolism ; Rats

Objective To investigate the effect of gypenoside on lipopolysaccharide (LPS)-mediated inflammatory response. Methods The BV2 microglia cell line was cultured in vitro. The BV2 microglia cells were divided into four groups: normal control, LPS (10 ng/ml), GP + LPS (GP 20 μg/ml, LPS 10 ng/ml), and GP (20 μg/ml). After 24 h cultivation, ELISA was used to detect the levels of tumor necrosis factor α(TNF-α), interleukin (IL)-1β, and IL-6. Immunocytochemistry staining and Western blot were used to detect the expression levels of nuclear factor (NF-κB) and suppressor of cytokine signaling 1 (SOCS-1). Results Compared with the normal control group, the release of TNF-α, IL-1β and IL-6, as well as the expression level of NF-κB in the LPS group were increased significantly (all P < 0. 001). Compared with the LPS group, the release of TNF-α, IL-1β and IL-6, as well as the expression level of NFκB were decreased significantly, while the expression level of SOCS-1 was increased significantly (P < 0. 001). There were no significant differences in the release of TNF-α, IL-1β and IL-6, as well as the expression levels of NF-κB and SOCS-1 between the GP group and normal control group (all P > 0. 05 ). Conclusions GP can significantly inhibit the LPS-mediated microglial inflammatory response. SOCS-1 protein may be involved in GP inhibiting LPS-mediated microglial inflammatory response.

AIM: To understand the epidemiology and related factors of strabismic amblyopia of students of primary school, and to provide guidances for the prevention and control strategy.
METHODS: A total of 600 cases of primary school students of Leshan City, Jiajiang County were given vision, oblique incidence and ocular and other screening. The prevalence rate of poor eyesight of strabismus, amblyopia prevalence rate of different sexes, ages were compared, and the degree of amblyopia and strabismus of children with different types of amblyopia and whether or not had stereoscopic vision were counted.
RESULTS: The prevalence rate of amblyopia and strabismus prevalence rate were respectively 4. 0% and 2.5%;With the growth of all age, low vision of students was significantly decreased, the difference of comparison of low vision rate of each age had statistical significance (P<0. 05), but different ages, strabismus prevalence of different sexes, amblyopia prevalence were compared, the difference had no significant differences ( P>0. 05 );Ametropic amblyopia was the main type, accounting for 55. 6%, and the degree of amblyopia mainly was light, moderate; ametropic amblyopia, most of ametropic amblyopia and strabismus had stereo vision, but there were no stereopsis of most of the strabismic amblyopia and all esotropia.
CONCLUSION:Ametropic is mainly type of amblyopia, the prevalence of relationship between the incidence of strabismic amblyopia of primary school students and sexes is not obvious, but the oblique amblyopia treatment effect, such as the establishment of stereoscopic vision and the age, eye position has a close relationship, should be early discovered, early treatment.

Objective To identify and characterize uveal melanoma associated protein variants with two-dimensional electrophore- sis and mass spectrometry.Design Experiment study.Participants 4 cases of specimens of uveal melanoma and 4 cases of normal con- tributed uveal tissue.Methods Proteins from uveal melanoma and normal urea were separated with two-dimensional eleetrophoresis (2-DE)and visualized with Coomassie G-250.Gels were analyzed by Image Master 5.0 software.The mass spectra were measured by matrix-assisted laser desorption/ionizatian time-of-flight mass spectrometry(MALDI-TOF MS)and searched against NCBI database using Mascot software.Main Outcome Measures Differential proteins.Results A set of 30 proteins were differentially expressed in uveal melanoma compared to nomal urea.Twenty-four types of protein only expressed in uveal melanoma.Five types of protein were up-regu- lated and 1 type of one down-regulated.These proteins can be subdivided into groups according to cellular function,such as enzyme, signal transduction,signal regulation,cytoskehton,immune,etc.Conclusions There is significant difference in protein profilings be- tween uveal melanoma and normal uvea.The differentially expressed proteins may be associated with the development of uveal melanoma.

Objective To evaluate the influence and significance of preoperative design of surgical approach on stem cell transplantation via stereotactic surgery. Methods Six patients with stroke in the basal ganglia region were selected. The transplantation target and transcranial approach point were designed by magnetic resonance examination before stem cell transplantation via stereotacfic surgery to guarantee that the line connecting the transplantation target and transcranial approach point could avoid the important functional areas, the ventricular system and the softening focus. Postoperative magnetic resonance examination was performed to observe whether the practical target and surgical approach coincided with the preoperative design or not. Results The practical transplantation target was coincided with the designed transplantation target, distributed around the softening focus without implanted cells in the softening focus. Surgical approach was coincided with the preoperative design and it successfully avoided the important brain functional area, ventricular system and softening focus.Conelnsion The preoperative design of surgical approach can not only ensure the cells being exactly transplanted into the reservation target and guarantee the curative effect, but also promise the surgical approach successfully avoiding the important brain functional area, ventricular system and softening focus and reduce the operative injury.

Animals ; Captopril ; pharmacology ; Heart Failure ; metabolism ; Male ; Myocytes, Cardiac ; metabolism ; Proto-Oncogene Proteins c-bcl-2 ; metabolism ; Rats ; Rats, Sprague-Dawley ; bcl-2-Associated X Protein ; metabolism