Objective To investigate the effects of protein kinase C (PKC) on proliferation, extracelluar matrix synthesis and transcriptional factor SOX9 (SRY-related high mobility group-box gene 9) expression of rat growth plate chondrocytes in vitro. Methods Rat costochondral growth plate chondrocytes (RGC) were isolated and cultured. The 1st serum free cultured passage RGCs were treated with 1, 10 and 100 nmol/L phorbol 12,13-dibutyrate (PDBu), PKC agonist, cell morphology were observed with inverted microscopy, cell proliferation, COLLAGEN and GAG synthesis were detected by isotope incorporation COLLAGEN TYPEⅡ and AGGRECAN mRNA transcription and SOX9 expression were revealed by RT-PCR and Western blot.Results 100 nmol/L PDBu treatment made the cell morphology of serum free cultured RGC closed to serum group and inhibited proliferation but promoted COLLAGEN and AGGRECAN synthesis, 3H-TdR、3H-proline and 35S-sulfate incorporation of 100nmol/L PDBu group were as 83%, 52.6% and 146.5% as those of serum free control(P

AIM: To establish the methods for rat costochondral growth plate chondrocyte (RGC) separation and culture and investigate their biological features, thereby provide experimental bases for studying the regulation of chondrocyte proliferation and differentiation. METHODS: RGC were obtained by microdissection and digestion, and cultured in monolayer. Morphological changes of the serial passage of RGC and the cell growth kinectics were observed. The cellular GAG and collagen type Ⅱ expression were detected by histochemistry and ICC. RESULTS: There were more than 98% viable cells in the obtained RGC. The morphology of primary cultured RGC was round or polygon. In this experiment, the sixth passage RGC was still maintained and showed polygonal morphology. The index of duplicatings/day increased in the preceding fourth passage RGC and decreased afterwards. There were more than 95% cells expressed collagen type Ⅱ and alcine blue stained positively in the primary RGC, as the passage number increased, the ratio of collagen type Ⅱ expression and alcine blue positive stained RGC dropped abruptly. CONCLUSION: The separation and culture methods adopted in this study can obtain high pure and viable RGC. The preceding three passage RGC maintains their in vivo phenotype, they are idle experimental materials for studying the regulation of proliferation and differentiation of chondrocyte and tissue engineering.

0.05). [3H]-proline incorporation in testing groups was 20% higher than that in control. CONCLUSION: The present study suggests EGF is able to enhance RGCs proliferation and collagen synthesis. Dedifferentiation caused by serial passage decreases proliferative effect of EGF on RGCs, but has no effect on collagen synthesis enhancement.

Objective To observe the effects of methylene blue photochemical(MB-P) inactivation of human cytomegalovirus in blood products.Methods Plasma and red blood cell suspensions containing 10% HCMV and 5?mol/L MB were illuminated by fluorescent light of 38000 Lux for 1 hour, HCMV was used as model viruses for validation of virus inactivation.Results The 50% tissue culture infective dose (TCID_ 50 ) contained in plasma and red blood cell suspensions decreased by 4~6 times.Conclusion MB-P treatment is effective in inactivating the infectivity of HCMV in plasma and red blood cell suspension.

Aim To investigate effects of genistein (5, 7, 4′-trihydroxyisoflavone) on rat costochondral growth plate chondrocyte (RGC) collagen and Sox9 expression. Methods Primary cultured RGC, effects of genistein on collagen synthesis, col2a1 mRNA transcription and Sox9 protein expression of 1st passage RGC were detected by isotope incorporation, RT-PCR and western blotting. Results Genistein inhibited collagen synthesis of 1st passage RGC (P

Objective To investigate the effects of laser irradiation on intracellular ROS(reactive oxidant species),intracellular calcium concentration(_i,and cell membrane integrity in the process of live cell imaging with confocal laser scanning microscopy. Methods The effects of a given laser irradiation on ROS,intracellular calcium concentration(_i and cell viability were revealed respectively by stained ECV-304 with H_2DCFDA,Fluo-4AM and calcein-AM/PI,and visualized and analyzed using ultra view LCI(live cell image)confocal microscopy. Results The irradiation of 488nm laser induced fluorescent intensity of DCF to increase abruptly and attain the climax in about 80 seconds,afterwards the fluorescent intensity fell and returned to the baseline.In the 70 minutes of the irradiation,the fluorescent intensity of intracellular Fluo-4 kept a slightly ascending tendency.The fluorescent intensity of calcein decreased 15minutes after the irradiation,and serval cells were PI positively stained.Conclusion 488nm laser irradiation induces intracellular reactive oxidant species(ROS) and calcium concentration to increase,but there is no significant influence on cell membrane integrity.

To investigate the damage on macrophage of the commercial peritoneal dialysis solution(CDS). Methods Macrophages were seperated from peritoneal fluid remained overnight of seven CAPD patients and TNF-a level of supernatant was determined and compared with those macrophages from uremic patients not yet recieving peritoneal dialysis. Results TNF-a levels of different glucose concentration decreased obviously in experimental group compared with control group, especially lower in 2.5% and 4.25% group. Conclusion In vivo experiment confirms that CDS possesses a long time inhibition on macrophage and this inhibition varies with different glucose concentrations.

Objective To evaluate the current situation and problems of the application of modified Rankin scale (mRS) in the outcome assessment in Chinese stroke trials. Methods Randomised and quasi-randomised controlled tri?als on stroke therapy published before September 2013 in 3 Chinese databases were included. All clinical trials applied mRS as the method of outcome assessment. Subarachnoid hemorrhage and transient cerebral ischemia were excluded. Types of stroke, statistical methods used for data analysis, duration of follow up, blinding of outcome assessment, types of intervention and the significance of the results were evaluated. Results Two hundred and ninety-eight trials were includ?ed in this analysis. 71.14%was for ischemic stroke, 21.48%for hemorrhagic stroke, 7.38%for both ischemic and hemor?rhagic stroke and 91.28%was for acute stroke(onset time<14d). Regarding to statistical methods used for data analysis, 50.00%of the trials used t-test or variance analysis which treated the mRS score as continuous data, while 22.15%used rank sum test or Chi-square test which regarded the mRS score as ranked data or multiply variable data. Dichotomous data was applied in statistical analysis accounts for 25.50%of trials. 12.42%trials applied mRS with other scales as the methods of outcome assessment. Duration of follow up ranged from 10d to 2 years (median 90 d, interquartile range 30-90 d). Only 5.03%assessed outcome blindly. 60.07%of the trials were drug therapy, 7.72%was rehabilitation thera?py, 10.40%were surgical treatment, 14.43%were combined therapy, 2.35%were management mode, 0.67%were nurs?ing, and 4.36%other therapy. Results in 86.91%of the trials were favorable to the tested interventions. Conclusions In aspects of, there is large difference between domestic and foreign clinical stroke trials in methodology of mRS including duration of follow up, blinding of outcome assessment and statistical methods used for data analysis.

OBJECTIVE To establish a fingerprinting method by randomly amplified polymorphic DNA.Epidemiological study was carried out on Pseudomonas aeruginosa in Ruijin Hospital. METHODS To obtain optimum scheme on reaction system for randomly amplified polymorphic DNA(RAPD) of P.aeruginosa. RESULTS P.aeruginosa strains isolated from the same ward shared the same RAPD fingerprint type,except for pulmonary ward.Different ward was with different fingerprint type. CONCLUSIONS Prevalent strain was not found in the whole hospital,but within ward exists hospital-acquired infection phenomenon.

Aim To prepare the monoclonal antibodies (mAbs) against human CD28 and to study its biological feature. Methods The hybridoma cell lines were obtained by fusing spleen cells of Blab/c mice that had been immunized with murine lymphoma cells transfected with full-length huaman CD28 cDNA to myeloma cells Sp2/0. Ascites were induced to produce the mAbs. The specificity and affinity of the mAb 18G8 was verified by CD28 competitive inhibitory test and FACS. Reactivities of mAb 18G8 to PBTC, U266, 8226, Jurkat and Daudi cell were studied by indirect immunofluorescence staining. mAb 18G8-inducing proliferation of peripheral blood T cells (PBTCs) was determined by [3H]thymidine incorporation test. Results Five hybridoma cell lines were obtained. mAb 18G8 secreted by one of the them, belong to mouse IgG2a. It recognized a epitope different from which recognized by the standard mAb(clone CD28.2). The Reactivitrates of the mAb 18G8 to PBTC, U266, 8266, Jurkat and Daudi cells were 70.2％ , 99.3％ , 98.6％ , 76.4％ and 1.9％ , respectively, similar with CD28.2. It was indicated that different antigen epitopes expressed on all above cells. mAb 18G8 could promote the PBTC proliferation in vitro(SI=7). It was indicated that The substitution of mAb 18G8 for B7-1 molecule could also mediate the costimulatory signals. Conclusion 18G8 is a specific and functional anti-CD28 mAb it may be of significant value in basic studies and clinical application.