Objective To elucidate the role of human bone marrow stromal cells (hBMSC) in combination with exogenous cytokines such as stem cell factor(SCF) and FLT 3 ligand (FL) in supporting the in vitro expansion of CD 34 + cells purified from cord blood.Methods CD 34 + cells were isolated from umbilical cord blood by using a high gradient magnetic cell sorting system(MACS),expanded in combination with SCF,IL3 (interleukin 3),IL 6 (interleukin 6),FL,EPO (erythropoietin),and were planted onto the pre established irradiated (20Gy) stroma layer(hBMSC plus combinations of cytokines) respectively.On day 10,total cells were counted, and hematopoietic progenitor cells were assessed by semisolid culture assay,CD 34 + cells were quantitated by FACS.Results After separation by MACS,the frequency of CD 34 + cells was 92%?0.04%.On day 2,almost all the inoculated cells adhered to the stroma layer,with only a small number in the supernatant.Then,cells in the supernatant increased gradually, but the percentage of CD 34 + cells decreased.Compared with control, expansion fold of CD 34 + cells, CFU GM, BFU E were significantly higher in hBMSC group (P 0.5).Conclusion ① CD 34 + cells from cord blood formed foci adherent to the monolayer and colony forming cells remained in the culture of 10days,indicating that hBMSC can support hematopoiesis in vitro;②using human bone marrow stromal cells in cooperation with exogenous cytokines may be a feasible way to expand hematopoietic stem/progenitor cells.

Objective　To construct an anti-LPS single chain phage antibody library in mice for further biomedical works. Methods　Total RNA was extracted from splenic cells for reverse transcription after BALB/C mice had been immunized with pure LPS for 4 weeks. The designed primers were used to amplify the variable region genes of both heavy and light chain (VH,VL) with polymerase chain reaction. The VH and VL were then conjugated to form a single chain of variable fragment (ScFv) by a linker. The ScFv was cut by NotⅠ and SfiⅠ and then ligated into a pCANTAB5E phagemid vector which was digested with the same restriction enzymes. The ligated vector was then tranfected into the competent E.coli TG1 cells. Four TG1 clones were randomly selected to detect the exotic DNA. Results　The titer of anti-LPS in murine sera was 1∶12 800. The concentration of total RNA was 12.3813 μg/ml. The length of the fragments were 340 bp for VH, 320 bp for VL and 800 bp for ScFv respectively. 1.9×107 clones were grown after transfection and one from the four randomly selected clones was identified to contain the exotic DNA. Conclusion　A 4.75×106 murine anti-LPS single chain phage antibody library was successfully constructed.

Objective To evaluate the value and clinical application of cardiac structure Z scores in children with preoperative tetralogy of Fallot (TOF) by echocardiography. Methods Considering the result of 31 patients who underwent echocardiography in Shenzhen Children’s Hospital before TOF repair from January 2011 to September 2013, the median age was 10 months (range:3 days to 14 years). Other 31 normal children were selected as normal control group with age-and sex-matched, the median age was 10 months (range: 6 days to 13 years). Echocardiographic measurements included the diameter of right ventricular outflow tract (RVOT), pulmonary valve diameter (PVD), left pulmonary artery diameter (LPAD), right pulmonary artery diameter (RPAD), pulmonary valve peak velocity (PV-Vmax), tricuspid annulus diameter (TVD), mitral annulus diameter (MAD), left ventricular end-diastolic volume (LVEDV) and left ventricular mass (LVmass). Shenzhen Children’s Hospital pediatric echocardiography normal reference values and the regression equation are applied to obtain the forecast average, and using the Z-score formula into the standard value (Z=[M-y]/ MSE ). Results There was no significant difference in body surface area between the TOF group and control group (P>0.05). Compared with control group, the true value of RVOT reduced significantly in TOF group, its Z-score showed negative values increase (-4.87±1.86 vs 0.33±0.93, t=-13.90, P<0.05). PVD, LPAD and RPAD reduced significantly in TOF group, those Z scores showed a negative value increase (-4.15±2.93 vs 0.11±0.85, t=-7.76;-1.35±2.38 vs 0.66±1.17, t=-4.24;-1.47±2.03 vs 0.59±1.04, t=-4.94;all P<0.05). While PV-Vmax increased significantly, and it’s Z score showed a positive value increase (8.38±0.19 vs 0.24±0.98, t=29.50, P<0.05). TVD of TOF patients was slightly larger than the control group (P=0.10), but the Z score was significant greater than the control group (1.00±1.17 vs 0.29±0.52, t=3.06, P<0.05). MV-D, LVEDV and LVmass of TOF patients were slightly decreased than the control group (all P>0.05), but the Z scores of TOF were significantly reduced than the control group (0.25±0.78 vs 0.76±0.65, t=-2.75;-0.92±1.94 vs 0.03±1.01, t=-2.41;-0.83±1.59 vs 0.67±0.69, t=-4.71; all P<0.05). The Z scores of RVOT, PVD, LPAD and RPAD had no significant difference between transannular patch surgery and pulmonary valve-sparing repair patients (-4.89±2.03 vs -4.84±1.67, t=-0.08; -4.73±3.49 vs -3.34±1.75, t=-1.31; -1.88±2.54 vs -0.62±2.00, t=-1.49;-1.89±1.90 vs-0.84±2.15, t=-1.41;all P>0.05). For the Z scores of PV-Vmax, there were no significant difference between transannular patch surgery and pulmonary valve-sparing repair (8.58±1.20 vs 8.12±1.16, t=1.07, P>0.05). Conclusions The Z scores value of cardiac structure can be used in quantitative analysis of RVOT obstruction and development of left ventricular backward of TOF. The Z score of transannular patch surgery is more lower than pulmonary valve-sparing repair patients. The normalized Z score by body surface area correction is important for the preoperative diagnosis and the surgical planning.

Objective:To study the effects of glutamine on the rehabilitation and the immune function in patients with acute stroke. Methods: The study included 57 patients with cerebral ischemia and 29 patients with brain hemorrhage.Nutritional and immune function parameters were evaluated at admission and after 2 weeks following stroke respectively.Neurological deficit was also evaluated by the Chinese Stroke Scale at admission and after 28 days following stroke.The infective complications were investigated. Results:At the same duration after acute stroke,the extent of neurological deficit recovery was significantly lower in the control group than the glutamine group.The rate of the infective complications was significantly higher and it's duration was significantly longer in the control group than the glutamine group.The degree of nutriture and immune function deteriorating was significantly severer in the control group than the glutamine peptide group. Conclusion:Parenteral glutamine supplements is helpful for the rehabilitation and in preventing deterioration of nutrition status and immune function following acute stroke.

OBJECTIVE To develop a new type of piezoelectric immunoglobulin microarray immunosensor for(determination) of immunoglobulin.METHODS The energy converters were 10MHz AT-cut quartz crystals with gold-coated electrodes.The monoclonal antibodies of immunoglobulin were immobilized onto the surfaces of(crystals) gold(electrodes) by SPA to study the best immobilization yield of the antibodies at different condition.Gold-(protein) A complex was highly stable and was believed to be Van der Waals in nature.We did experiments to find the best temperature-time condition about the binding of SPA and the gold surface and to explore the influence(about) the immobilization of antibody by the concentration and the pH of the antibody solution.RESULTS At the same antibody concentration, it had the highest immobilization yield of SPA at a low-temperature-long-time(combination).The most immobilization yield was obtained at the 5mg/ml concentration of antibody solution.The optimum pH for antibody immobilization was 7.2.CONCLUSIONS SPA for the immobilization of antibody is of(simple) operation,and time-saving.It is a good way to immobilize antibody on the gold electric surface of the quartz(crystal).

Objective To investigate the calculation method and its inlfuencing factors of Z scores in the aortic root diameters measured by echocardiography in children. Methods A total of 105 children with median age 19 months, who came to Shenzhen Children′s Hospital from March 2012 to October 2012 were included. The diameters of aortic ring (ARD) and aortic sinus (ASD) were measured by two dimension echocardiography, Z scores of ARD and ASD were calculated using two different normal reference values regression equation and mean square error derived from Shenzhen children′s hospital (C method) and Pettersen et al (P method). The regression equation from C method and body surface area (BSA) formula from P method were adopted to calculate Z scores of ARD and ASD (ZH method). The Z results of ARD and ASD calculated by those three methods were compared and were analyzed for their normality probability distributions. Results Z scores of ARD and ASD derived from C method were all showed as normal distribution (P=0.067 and 0.650). Z scores of ARD and ASD derived from P method were all showed as normal distribution (P=0.208 and 0.970). Z score of ARD derived from ZH method was showed as non-normal distribution (P=0.027), but Z score of ASD was normal distribution (P=0.430). There were no significant differences in ARD-Z calculated by C method (0.41±0.89), P method (0.23±0.85) and ZH method (0.36±0.94) (F=1.117, P=0.309). There were signiifcant differences in the Z scores of ASD calculated by C method (0.38±0.89), P method (0.58±0.71) and ZH method (0.36±0.84) (F=5.443, P=0.005). Z scores of ARD (r=0.917, P=0.000) and ASD (r=0.900, P=0.000) calculated by C method correlated well with that by P method. Conclusions Calculation method of BSA and normal reference values regression equation were the main influencing factors of Z score value in quantifying children aortic root diameters by echocardiography. For the clinical applications. The normal reference value should be used which is suitable for the Chinese children.

Objective To investigate the effect of health education on enhanced curative effect of motor imagination therapy of patients with hyperbaric oxygen(HBO). Methods Thirty patients undergoing HBO combined with motor imagination therapy were given health education including contents of motor imagination,time for motor imagination,intervention of bad emotions and time for motor imagination after leaving the treatment cabin.The understanding of motor imagination-associated knowledge after education was compared to that before education. Results The understanding of motor imagination-associated knowledge after education was significantly better than before education(χ2=9.017,P=0.003). Conclusion Health education may help improve the curative effect of hypterbaric oxygen treatment(HBO)by way of improving the understanding of motor imagination-related knowledge.

Objective To evaluate the value of lung ultrasound score (LUS) in the quantitative assessment of the severity of neonatal respiratory distress syndrome (NRDS) and the value of clinical diagnosis and treatment.Methods Totally 74 NRDS cases and 30 normal neonates were studied.LUS was compared with X-ray examination,clinical data,ventilator assisted ventilation and ventilator parameters.ROC curve was used to calculate sensitivity and specificity of LUS to predict the severity of NRDS and application of invasive ventilator treatment.Results The main findings of the lung ultrasound in NRDS included diffuse distribution of dense B line,disappeared A line,pleural line abnormalities,decreased pulmonary slip sign and pulmonary consolidation.LUS in patients with NRDS was significantly correlated with X ray grades,clinical grades,assisted ventilation mode grades,number of days on ventilator and ventilator parameters (all P＜0.05).LUS value to predict mild,moderate and severe NRDS were 13.0,22.5,and 29.5,respectively.The best cutoff point for LUS to predict the adoptation of invasive assisted ventilation was 22.5,which had sensitivity of 86.0 ％ and specificity of 64.5 ％.Conclusion LUS can be used to diagnose and evaluate the severity of the desease,and to guide the clinical diagnosis and treatment.

Objective To evaluate the biological characteristics of hip joint in infants with developmental dislocation of the hip (DDH).Methods From January 2013 to June 2016,30 patients (age from 1 to 8 months,25 females and 5 males) who were diagnosed as DDH,underwent ultrasound examination in Shenzhen Children's Hospital,in these patients 20 cases were treated surgically,and 10 cases were treated with non-surgical treatment.There were 34 hips (26 unilateral dislocation and 4 bilateral dislocation) dislocation in 60 hips.All the patients were confirmed by X-ray,magnetic resonance imaging (MRI) examination or operation.All the patients were examined by ultrasound through the coronal and transverse plane of the hip joint.The α angle,femoral head coverage ratio by acetabulum (FHC),femoral head length and width,distance from pubis to femoral head (P-H) and distance from ischium to femoral head (I-H) were measured.The dislocation joints were compared with contralateral joints.Results The α angle in the hip dislocation group was smaller than the contralateral group [(50.5±3.75)° vs (64.8±3.38) °],and there was significant difference between the two groups (t=-15.181,P ＜ 0.001).The FHC,femoral head length and width in the hip dislocation group were all smaller than the contralateral group [(23.4t17.63)％ vs (64.3±6.45)％,(0.98 ±0.15) cm vs (1.19 ±0.11) cm,(1.38±0.21) cm vs (1.61 ±0.16) cm],and there were significant differences between the two groups (t=-12.469,-6.034,-4.568,all P ＜ 0.001).The P-H and I-H in the hip dislocation group were larger than the contralateral group [(0.97±0.45) cm vs (0.27±0.05) cm,(0.75±0.30) cm vs (0.17±0.05) cm],and there were significant differences between the two groups (t=8.805,10.696,both P ＜ 0.001).The α angle,femoral head length and width in bilateral dislocation of hip group were slightly smaller than the unilateral dislocation of hip group [(50.3±2.75)° vs (51.3±4.77)°,(0.90 ±0.15) cm vs (0.97 ±0.12) cm,(1.25±0.20) cm vs (1.37 ±0.17) cm],but there were no significant differences between the two groups.The P-H and I-H in bilateral dislocation of hip group were slightly larger than the unilateral dislocation of hip group [(0.97 ± 0.49) cm vs (0.80±0.31) cm,(0.92±0.26) cm vs (0.68±0.18) cm],but there were no significant differences between the two groups.The value of acetabular index in the ultrasound group was slightly larger than the X-ray group [(33.13 ± 7.82)° vs (31.20 ± 8.31)°],and there were no significant differences between the two groups.Conclusions The characteristics of DDH includes acetabulum and femoral head dysplasia,and femoral head and acetabulum position relationship abnormalities.Quantitative ultrasonography of the hip can be used to quantitatively evaluate the structural features of DDH,and it is helpful to the early diagnosis and follow-up of DDH.

In order to construct a new type of piezoelectric quartz immunosensor for the determination of carcino-embryonic antigen (CEA), the sensor detection pools were consisted of plastic loops and crystals that were 10 MHz quartz AT-cut with gold coated electrodes and the immobilization of the monoclonal antibody against CEA onto gold electrode surface of the quartz crystal was accomplished via Thiol method. Then the immunosensors were used in clinical laboratory. The experimental results showed that the piezoelectric immunosensor had good response to CEA, the frequency shifts were linearly dependent on CEA concentration in the range of 1.56 to approximately 50.00 ng/ml, other antigens such as alpha fetoprotein (AFP), prostate specific antigen (PSA), human chorionic gonadotropin (hCG) did not interfere with the sensor's response. The results obtained from this method were in satisfactory accordance with those obtained by radio immunoassay(P>0.05), The correlation coefficient was 0.90. Piezoelectric immunosensor for determination of CEA has the advantage of high sensitivity, high specificity, unnecessary labeling, simple performing, rapid analysis, low cost, real-time detection and repeated use, etc. It can be used for detecting serum CEA in clinical laboratory.
Antibodies, Monoclonal ; Carcinoembryonic Antigen ; blood ; immunology ; Electrodes ; Equipment Design ; Humans ; Immunoassay ; instrumentation ; methods ; Quartz ; Sensitivity and Specificity