Objective To construct the tranfected cell expressing the human CXCR4 gene and to identify the effect on its immigration. Methods Total RNA was isolated from peripheral blood monouclear cell (PBMC),the full-length CXCR4 gene was amplified by RT-PCR and was inserted into plasmids PBudCE4.1 which have two promtors, after the identification by digestion and sequencing,the recombinant was transfected into K562 cell by lipofectamineTM2000. After screening culture by zeocin, stable transfected K562 cell line was established, and transcription and exression of CXCR4 were checked by flow cytometry; the chemotactic activity of K562 cell transfected and untrandfected CXCR4 was analyzed by Transell plate. Results The eukaryotic expression plasmid PBudCE4.1/ CXCR4 was constructed successfully. The stable trasfected K562/CXCR4 cell lines which highly express CXCR4 was established,the chemotactic activity of K562/CXCR4 was increased significiantly than K562. ConclusionCXCR4 transfected K562 cell line was successfully established, and it can make the basis for the further research on mechanism of extramedullary infiltration in leukemia.

BACKGROUND: Hematopoietic stem cells have an ability of multi-directional differentiation and can be differentiated into megakayocytes in the presence of suitable hematopoietic growth factors and lineage special growth factors.Inducing of CD34+ stem cells to megakaryocytes in vitro involves many changes in gene expression, especially the signal transduction genes.OBJECTIVE: To study the expression of signal transduction genes after differentiation from stem cells into megakaryocytes.DESIGN: Open experiment.SETTING: Department of Tumor Immunology, Fujian Provincial Tumor Hospital.MATERIALS: 60-145 mL of cord blood was collected from normal labor fetuses under sterile conditions by using the blood-bag collective method. Patients and their relatives provided the confirmed consent. Reagents and equipments were detailed as follows: VarioMACS segregation apparatus, CD34+ Isolation Kit, SCGM serum-free medium, CD monoclonal antibodies, cytokines, human genome Oligo Set Version 2.1 from the CapitalBio Corporation, and LuxScan 10K/A bi-passage laser scanning apparatus. The researchers obtained consent from the local ethic committee.METHODS: This experiment was carried out in the Department of Tumor Immuonlogy, Fujian Provincial Tumor Hospital and the CaptialBio Corporation. from July 2005 to June 2006. RNA was extracted separately from CD34+ cells purified from cord blood and CD41+ cells induced from the same cord blood. The cDNA probes were synthesized by reverse transcription and purified. CD34+ cells before culturing and CD41+ cells after culturing were labeled with Cy3-dCTP and Cy5-dCTP respectively. Labeled DNA was dissolved in 30 μL hybridization fluid and hybridized at 42 ℃ over night MAIN OUTCOME MEASURES: The differential gene expression between haematopoietic stem cells and megakaryocytes involving cell signal transduction were detected by using DNA microarray analysis. According to the gene chip results,17 differentially expressed genes were selected for further analysis by RT-PCR.RESULTS: In this experiment, 3 522 differentially expressed genes were screened, including 1705 up-regulated genes and 1 817 down-regulated genes. In the 3 522 differentially expressed genes, there were 343 genes involved in cell signaling, 150 genes involved in transcription regulation, 21 genes involved in cell differentiation. The expression of the CD61 gene increased 369.83 fold, the expression of the CD41 gene increased 27.38 fold, and the expression of PF4 gene increased 24.06 folds; The expression of mitogen-activated protein kinases (MAPKs), G protein-coupled receptors (GPCRs) and members of RAS oncogene family increased mostly. The expression of the genes involved in STAT pathways, both SOCS1 and JAK2 were up-regulated, but STAT5A was down-regulated. As determined by the gene chip results, 17 differentially expressed genes were selected for further analysis by RT-PCR. GAPDH was used as the internal control, and RT-PCR results were in agreement and confirmed the finding from the gene chip expression results.CONCLUSION: Thrombopoeitin (TPO) and other hematopoietic growth factors may enhance the proliferation and differentiation of megakaryocytes derived from cord blood stem cells by GPCRs-Ras-MAPK pathway.

Objective To analyze the influence of peptidimer-c on the gene expression profiling of K562 cells and investigate the mechanism of peptidimer-c inducing the apoptosis and inhibiting proliferation of K562cells.Methods Trypan blue staining technique was performed for counting the number of living K562 cells treated with peptidimer-c.The ultrastructure changes of K562 cells treated with peptidimer-c was observed under transmission electron microscope.The Human U133 Plus 3.0 gene chips were used to detect the differentially expressed genes of K562 cells treated with peptidimer-c.Reverse transcription PCR was conducted to confirm some genes identified by gene chips.Results Peptidimer-c could induce the apoptosis and inhibit the proliferation of K562 cells.Peptidimer-c caused widely changes of the gene expression profiles of K562 cells.The chip data suggested that there were 529 differentially expressed genes,of which 455 genes were up-regulated and 74 genes were down-regulated.The relevant apoptotic genes were down-regulated markedly,including JUN,AXUD1,TNFRSF10B,etc.Fifteen of the differentially expressed genes were detected by RT-PCR,which was consistent with the chip data.Conclusion Peptidimer-c may induce aooptosis of K562 cells by activating the TNF/TNFR family and the JUN family.

Objective To analyze the characteristic of serum proteins in non-small cell lung cancer (NSCLC) patients,establish serum markers pattern for the diagnosis of NSCLC. Methods Surface enhanced laser desorption ionization time of flight mass spectormetry(SELDI-TOF-MS) technology was used to analyze serum samples. Biomarker Pattern Software (BPS) was used to detect the protein peaks. Results Sixteen significantly different protein peaks were found in serum samples in NSCLC patients and healthy controls. Eight up-regulated protein peaks and eight down-regulated protein peaks (P

Objective To identify and rename one strain stored in National Center for Medicine Culture Collections ( CMCC) and used for tracheitis vaccine production. Methods The test strain CMCC (B)29108 and the type strain DSM30007T were cultured on NA medium. Characteristics in morphology, physiology, biochemistry and fatty acid profile were compared between the two strains. Phylogenetic analysis was based on 16S rRNA and rpoB gene sequences, together with the DNA-DNA hybridization assay. Results A Comparative analysis of a partial sequence of the 16S rRNA gene sequence revealed that the CMCC( B) 29108 strain was closed to the Acinetobacter species and showed the highest similarity with the type strain Acinetobacter baumannii DSM30007T. Moreover, the CMCC(B)29108 strain was highly similar to type strain DSM30007T in morphology, physiology, biochemistry and fatty acid profile. On the phylogenetic tree based on 16S rRNA and rpoB gene of all Acinetobacter members, the CMCC(B)29108 strain steadily clustered into one independent branch only with the DSM30007 T strain with a DNA-DNA hybridization value of 100%. Conclusion The CMCC(B)29108 strain that is one of the strains used for the production of tracheitis vac-cine should be assigned to the species of Acinetobacter baumannii based on its phenotypic, phylogenetic and chemotaxonomic characteristics.

Objective To develop a simple method of determination of sorafenib in serum by reversed-phase high performance liquid chromatography (RP-HPLC) and to explore its application in sorafenib therapeutic drug monitoring (TDM).Methods Sorafenib extracted by ethyl ether-petroleum (9∶1) with internal standard of erlotinib from serum was wiped off in 60 ℃ water bath.Sorafenib was redissolved by mobile buffer and analyzed by 40 μl.Chromatographic column was Symmetry Rp18 (5 μm,4.6 mm×250 mm,waters) column in normal temperature.The mobile buffer was 28 mmol/L acetate buffer (pH 5.8)-acetonitrile (37∶63).Sorafenib and erlotinib were detected in 249 nm and 335 nm,respectively.Results The concentration range of sorafenib was 0.50-20.00 μg/ml (r =0.9999).The within-day and between-day accuracies of sorafenib were less than 4.77 ％ and 8.79 ％,respectively.The average recovery rate was 98.48 ％.Sorafenib was stable in serum or after extraction.The concentrations of sorafenib in two patients were detected.Conclusion Detection of sorafenib in serum by RP-HPLC is simple and accurate,which is available to determine sorafenib in serum.The TDM of sorafenib has clinical significance.

Objective To analyze the characteristic of serum proteins in non-small cell lung cancer (NSCLC) patients, establish serum markers pattern for the diagnosis of NSCLC. Methods Surface enhanced laser desorption ionization time of flight mass spectormetry ( SELDI-TOF-MS) technology was used to analyze serum samples. Bio-marker Pattern Software (BPS) was used to detect the protein peaks. Results Sixteen significantly different pro-tein peaks were found in serum samples in NSCLC patients and healthy controls. Eight up-regulated protein peaks and eight down-regulated protein peaks ( P < 0. 001 ) were identified in serum samples of NSCLC patients. Three up-regulated protein peaks(P <0. 05) were identified in serum samples of patients of NSCLC with smoking history. Two up-regulated protein peaks(P <0. 01) were identified in serum samples of patients of squamous carcinoma comparing with adenocarcinoma. No significantly different protein peak was found in serum samples of NSCLC patients at different clinical stages . Conclusion SELDI - TOF - MS technology can identify different protein peaks and so function as a diagnostic tool with high sensitivity and specificity.

Objective To screen serum biomarkers in patients with hepatocelhlar carcinoma by using surface-enhanced laser desorption and ionization time-of-flight mass spectrometry(SELDI-TOF-MS)technique and to evaluate its clinical implication in the patients whose alpha-fetoprotein were negative in the sera. Methods Proteomic spectra were generated by mass spectroscopy in 112 cases, including 57 cases AFP-negative hepatocellular carcinoma,and 55 cases of healthy control. The consequence was analyzed and the characteristic preteomic peaks was selected by using Biomarker Wizard. Results Seven low expressed potential biomarker were indentified with the mass-to-charge ratio of 4.2×103, 4.1×103, 6.7×103, 5.7×103, 6.5×103, 6.9×103, 5.8×103. The sensitivity for diagnosing hepatic cancer were 88.23 % and specificity was 92.31%. These peaks were not correlated with age, sex, tumor mass size, pathology grading and cirrhosis in hepatocellular carcinoma. Conclusion SELDI-TOF-MS offers a unique platform for the proteomic detection of hepatocellular carcinoma.It also offers an auxiliary diagnosis method to the patients whose alpha-fetoprotein are negative in the serum.

Objective: To explore the influence of birth weight and growth patterns during infancy on overweight and obesity among first grade primary school pupils, so as to provide a theoretical basis for the formulation of early life prevention and intervention policies. Methods: In 2019, data related to routine physical examinations were collected for primary school pupils in the Minhang District of Shanghai, and information regarding birth and follow ups was collected retrospectively. Physical examination data of 4 434 pupils at 12 months of age were obtained. A multiple linear regression model was used to analyze the relationship between growth patterns during infancy and body mass index (BMI) in the first grade of primary school. A generalized linear model was employed to analyze the relationship between birth weight and growth patterns during infancy and overweight and obesity in the first grade of primary school. A hierarchical analysis was conducted. Results: A linear relationship was observed between growth patterns during infancy and BMI and the BMI Z score of first grade primary school pupils [ β(β 95%CI)=0.30(0.24-0.35),0.12(0.10- 0.15 ), P <0.01]. In addition to subjects classified as small for gestational age (SGA), catch up growth during infancy was identified among subjects who were classified as appropriate for gestational age (AGA) and large for gestational age (LGA). LGA at birth and catch up growth during infancy were independent risk factors for overweight and obesity among first grade primary school children ( RR =1.31-1.55, P <0.05). The hierarchical analysis showed that catch up growth increased the risk of overweight and obesity among first grade primary school pupils classified as AGA [ RR(RR 95%CI )=1.74(1.42-2.14),1.87(1.56-2.26)], and increased the risk of obesity among first grade primary school pupils classified as SGA and LGA [ RR(RR 95%CI )=3.74(1.04-13.49),3.24(1.62-6.46)]( P <0.05). Among those who exhibited catch up growth during infancy, LGA increased the risk of obesity among first grade primary school pupils ( RR= 2.60 , 95%CI=1.35-5.02, P <0.01), but not the risk of being overweight ( P =0.13). Conclusion Birth weight and growth patterns during infancy have an impact on overweight and obesity among children in the first grade of primary school. It is suggested that attention should be paid to growth and physical development in early life for those classified as LGA and AGA, and catch up growth in children should be closely monitored.

Objective: To investigate the distribution of various types of screen time and examine the association of screen time with psychological and behavioral development problems in children aged 3-6 years, so as to provide scientific basis for children s screen use and mental health promotion. Methods: A total of 3 875 mother child dyads who completed the follow up in Shanghai Maternal-Child Pairs Cohort were included. The daily usage time of children s tablet, mobile phone, TV, projectors, and other types of screens were obtained in questionnaire survey. Children s psychological and behavioral development problems were evaluated by Age-Stage Questionnaire, Third Edition (ASQ-3) and Strengths and Difficulties Questionnaire (Parent version) (SDQ). The Chi-square test, Mann Whitney U and Kruskal Wallis rank sum test were used to compare the detection rate of psychological and behavioral development problems and screen time in children with different characteristics. Multivariate binary Logistic regression was used to analyze the association of screen time with psychological and behavioral development problems. Results: There were 49.91% of children having screen time more than 1 h/d. Children s TV, tablet and mobile phone screen time were 0.39(0.25, 0.96 ), 0.25(0,0.61) and 0.18(0,0.25) h/d. The detection rates of suspected developmental delay in fine motor, problem solving and personal-social domains and pro social behavior deficiency and externalizing behaviors in boys (8.54%, 6.77%, 5.46%, 30.07 %, 27.39%) were higher than that in girls (4.64%, 4.85%, 2.48%, 22.10%, 22.36%) ( χ 2=23.76, 6.49, 22.37, 31.81, 13.06, P <0.05). There were statistically significant differences in the detection rates of suspected developmental delay in communication, fine motor and problem solving, as well as internalizing behavior and externalizing behavior of children with different parents educational levels ( χ 2=14.37, 15.18, 21.10, 11.66, 9.27; 16.34, 26.75, 32.89, 16.97, 6.37, P <0.05). There were significant differences in the detection rates of suspected developmental delay in problem solving, prosocial behavior deficiency, internalizing behavior and externalizing behavior of children whose mothers had anxiety/depression symptoms during pregnancy ( χ 2= 5.61 , 9.05, 21.90, 7.17; 8.75, 6.06, 12.76 , 5.55, P <0.05). The average total screen time of boys was longer than that of girls (1.07, 1.00 h/d, Z=-2.08, P =0.04). Compared with children with other educational levels of their parents, the total screen time, mobile phone and TV screen time of children whose parents had college education or above were short (father: H =42.01, 44.49 , 21.24, mother: H =42.31, 39.21 , 26.47, P <0.01). Among all types of screen time, mobile phone screen time had the most impact on psychological and behavioral development. More mobile phone screen time increased the risk of suspected developmental delay and abnormal emotional behavior ( P < 0.05). Screen time of tablet, mobile phone and TV were positively correlated with externalizing behavior ( OR=1.36, 1.57, 1.27 , P <0.05). Conclusions Screen time is related to children s psychological and behavioral development problems and mobile phones affect the most. Parents should limit their children s screen time to avoid excessive screen time affecting their psychological and behavioral development.