Objective To investigate the immunophenotype and the cytotoxicity of cytokine-induced killer(CIK) against tumor cells in vitro.Methods Lymphocytes cells were isolated freshly from peripheral blood of healthy donors by Ficoll-Hypaque density centrifugation,and the cells obstained were induced by IFN?,IL-2 and CD3McAb.Phenotypes and cytotoxicity of CIK were analysed by FACS.Target cells were differentiated from effect cells by CFSE dying.The mortality of Target cells were determined by FACS.Results The maximum proliferation of CIK reached at the 22nd day.The phenotypes of CD3,CD11a,CD54,HLA-DR were expressed highly;CD25,CD28,CD69,FasL were expressed moderately on CIK.The expression of CD16 was not increased.CIK possessed the cytotoxicity against tumor cells of K562,HL-60,Hela,SMMC7721 and A375.Conclusion IFN?,CD3McAb,IL-2 can induce peripheral lymphocytes to produce CIK which own strongly proliferation and exert highly efficient cytotoxic effects on tumor cells.

Abstract Objective: To prepare high-purification and high-specific activity of recombinant human interleukin-6 (rhIL-6). Methods: The rhIL-6 was obstained from inclusion body expressed by IPTG-induced pT7.7hIL-6 expressed vector using extracting,denature and refolding techniques. The rhTL-6 was further purified by anionic-exchange chromatography. Activity of rhIL-6 was measured by 3H-TdR method. Results: After a single-step purification,the product purity reach 95% and it's specific activity was 3.0 x 10~8 U/mg. Conclusion:This scheme of puri-fication was an easy way requiring rhTL-6.

Objective To construct a recombinant expression vector of human interleukin-24(hIL-24) gene and express it in E.coli M15,and to evaluate the bioactivity of IL-24 fusion protein.Methods The human IL-24 cDNA fragment was amplified from plasmid by polymerase chain reaction(PCR),and sequenced.PQE/hIL-24 was constructed by gene rearrangement,then it was transformed into E.coli M15.The expression of the target protein was induced with IPTG and purified by Ni2+-NTA agarose column.The expressed recombinant hIL-24(rhIL-24) was identified by SDS-PAGE and Western blotting.Normal peripheral blood mononuclear cells(PBMCs) were cultivated with the expression protein for 48 and 72 h,the levels of IL-6,IFN-? and TNF-? of PBMCs stimulated with rhIL-24 were detected by ELISA.Results The recombinant prokaryotic expression vector PQE/IL-24 was constructed successfully and expressed stably in E.coli M15.At about 18 400 of molecular weight,there was an induced protein band.The levels of IFN-?,IL-6 and TNF-? in the group of cultivated with the expression protein were obviously higher than those in the groups without the expresson protein(P

Objective To get the protein expression of sFlt-1 by transfection and to investigate the effects of sFlt-1 gene transfection on the growth of K562 cells.Methods The recombinant plasmid pcDNA3-sFlt-1D4 was constructed.The recombinant plasmid pcDNA3-sFlt-1D4 was transfected into the bone marrow stromal cells by Lipofectamine 2000,which was identified by RT-PCR,ELISA and MTT.Results The transfection efficiency identified by flow cytometry was 9.27%.The protein expression of sFlt-1D4 was found in the culture supernatant 24 h,48 h and 4 weeks after transfetion by ELISA and the expression concentrations were(0.104?0.078),(0.158?0.022) and(0.171?0.069) ?g?L-1,respectively.The content of VEGF secreted by K562 culturing with transfectant cells culture supernatant was reduced compared with control.The inhibitoy rates on the proliferation of K562 cells via MTT assay were 9.41%?4.71%,23.63%?7.50%,and 33.13%?6.93%,respectively.Conclusion The bone marrow stromal cells transfected with recombinant plasmid pcDNA3-sFlt-1D4 could secrete sFlt-1D4 and inhibit the proliferation of K562 cells.

Objective:To investigate the expression of cyclooxygenase-2 in primary hepatocellular carcinoma, in cancer surrounding tissues and normal liver tissue and its relationship with clinical pathological features.Methods:The expression of COX-2 was detected in 30 cases of hepatocellular carcinoma, 20 cases of cancer surrounding tissue and 10 normal liver tissue by flow cytometry (FCM) and immunohistochemistry (SP). The clinical data were analyzed retrospectively.Results:(1)The expression of COX-2 in the HCC tissue was significantly higher than in cancer surrounding tissues and normal liver tissue (P0.05).Conclusion:The hyperexpression of COX-2 in tissue can reflex the biological behavior of HCC,and have very important role in the development of HCC.The specificness of COX-2 protein expression make it to be new target of tumor diagnosis and treatment.These results provide a theoretical basis for the chemoprevention of hepatoma.

Objective To explore the related factors influencing the prognosis of children with Tourette syndrome ( TS) . Methods We collected 420 outpatient cases of TS children in the department of pediatrics of Guangdong Provincial Hospital of Traditional Chinese Medicine from January of 2007 to October of 2010 as the research objects. Using the unified survey questionnaire, we observed the influencing factors of TS prognosis such as sex, onset age, mother’s pregnancy situation, the perinatal period, partiality for food intake, family relationship, the first symptom, the severity of disease, underlying diseases, comorbidities ( such as attention deficit hyperactivity disorder, anxiety disorder, and obsessive-compulsive disorder) , family history of psychiatric or neurological diseases, trace elements levels, electroencephalogram (EEG), antistreptolycin O (ASO), course of disease, and maintenance treatment. The related factors of prognosis was analyzed with single factor and multiple factors Logistic regression analysis. Results Among the 420 cases of TS children, 396 cases were included into the final analysis, 24 were lost and the follow-up lost rate was 5.7%. The remission rate of TS was 78.3%, and the uncured rate was 21.7%. The results of preliminary screening of the influencing factors by single factor Logistic analysis showed that the related influencing factors for TS prognosis were 12, and they were course of diseases, abnormal birth history, father’s education level, mother’s education level, upbringing methods, family history of psychiatric or neurological diseases, underlying disease history, comorbidities history, abnormal ASO, the severity of disease, the frequency of disease relapse, and the medication history of western medicine (P<0.05) . And then the obtained 12 factors were analyzed by the multiple fac tors Lo gistic regression analysis, the results showed that upbringing methods, comorbidities history, the severity of disease, and the frequency of disease relapse were correlated with TS prognosis ( P < 0.05) . Conclusion TS children will have poor prognosis when their parents spoil, indiscipline, beat and scold, and dictate them, or when the children have severe illness state, frequent recurrence of the disease, and the history of comorbidities ( atten tion deficit hyperactivity disorder, anxiety disorder, obsessive-compulsive disorder, etc.) .

Objective:To observe inhibitory effrects of DNA vaccine co-expressing CEA tandem repeat epitopes and FL on cancer cells in mice.Methods:The encoding sequences for CEA tandem repeats and FL were inserted into plasmid pcDNA3.0 using gene recombinant technique.BALB/c mice were immunized intramuscularly with the co-expressing DNA vaccine.The survival time and tumor size were measured and specific CTL cytotoxicity was detected by ~(125)I-UdR release method.Results:Compared with that of the control,the survival time was prolonged (P＜0.01)and the tumors were significantly inhibited in the mice immunized with the vaccine peDNA-triCEA_(625-667)-sFL(P＜0.01).The splenic cells from mice immunized with the vaccine pcDNA-triCEA_(625-667)-sFL induced strongly cytotoxicity against tumor cells H22-CEA ~+(P＜0.01).Conclusion:The recombinant DNA vaccine co-expressing pcDNA-triCEA_(625-667)-sFL can suppress the growth of tumor expressing CEA in mice and enhance CTL response against CEA antigen.

Objective To probe the relationship between the expression of TL1A and the level of IFN-γ secreted by T cells in the acute stage of Guillain-Barre syndrome (GBS). Methods ① Six-week female Bal b/c mice were immunized by purified recombinant human soluble TNF-like molecular 1A (rhsTL1A) protein. The polyclonal antibody against rhsTL1A was identified by immunofluorescence using human umbilical vein epithelial cells (HUVEC). ② To detect the biologic activity of rhsTL1A, the peripheral blood mononuclear cells (PBMC) from the healthy donors were separated by Ficoll gradient centrifugation and were seeded on 96-well plates with medium containing 2 μg/ml PHA (control group), 2 μg/ml PHA + 25 ng/ml rhsTL1 A, 2 μg/ml PHA + 100 ng/ml rhsTL1A and 2 μg/ml PHA + 400 ng/ml rhsTLlA respectively. T cell proliferation assay was carried out using ~3H-TdR. ③ IFN-γ productions in the sera of the children with GBS in the acute stage were detected by ELISA. ④ The ratio of CD_3~+ TL1A~+ T cells to CD_3~+ T cells in the peripheral blood of the children with GBS in acute stage was detected with flow, cytometry. ⑤PBMC from the children in acute GBS were separated and cultured in the environment adding 2 μg/ml PHA and 400 ng/ml rhsTL1A in vitro. Then, the IFN-γ in the supernatant was determined by ELISA kit after 72 hours. Results ① hTL1A A expressed by eukaryotic HUVECs was recognized by rhsTL1 A polyclonal antiserum. ② The result of T cell proliferation assay showed that SI of 25 ng/ml rhTL1A, 100 ng/ml rhTL1A A and 400 ng/ml rhTL1A group was increased compared with control group. The SI of 2 μg/ml PHA +400 ng/ml rhsTL1 A group was the highest (2. 65) among them. ③ IFN-γ productions in the sera of the children with GBS in the acute stage ((102. 25±22. 17) pg/ml) were increased significantly compared with healthy control ((28.75 ± 1.31) pg/ml, t = 3. 309, P < 0. 05). ④ The ratio of CD_3~+ TL1A~+ T cells to CD_3~+ T cells in the peripheral blood of the children with GBS in acute stage (18.22%± 1.83%) was enhanced significantly compared with healthy control (5. 17% ±0. 48%, t = 6. 884, P < 0. 01). ⑤ PBMC both in healthy control and the acute GBS secreted more IFN-γ markedly ((43.56± 4.41) pg/ml and (180.64 ± 38.39) pg/ml) after being incubated in 2 μg/ml PHA and 400 ng/ml rhsTL1A (t =4. 523 and 2. 600, P <0. 01 and 0. 05 respectively). Moreover, PBMC in acute GBS secreted more IFN-γ, than that of the healthy group markedly (t = 3. 545, P < 0. 05). Conclusions ① The mouse antiserum recognizing rhsTL1A is successfully obtained. ② In this study, 400 ng/ml rhsTL1A promotes the proliferation of T cells activated by 2 μg/ml PHA, indicating that rhsTL1A has biological activity. ③ The expression of hTL1A of activated T cells in the peripheral blood of the children with acute GBS is up-regulated. These TL1A proteins promote the secretion of IFN-γ through binding to their receptors DR_3.

Objective To explore the impact of family education methods and parental rearing or not on the psychological problem tendency of primary school students.Methods The mental health test (MHT) was used on getting mental health data of the 2 838 primary school students from Dongguan city.Results (1) The results of MHT showed that 94.28％ of primary school students had no psychological problems,the tendency rate of psychological problems was 4.29％,and 1.43％ of them were suffering from serious psychological problems.Learning anxiety,physical symptoms and allergy tendency were more prominent.(2) The total MHT scores of primary school students with authoritarian,democratic,permissive and neglected family education methods were (34.84±14.58),(31.04±13.60),(35.19±12.82) and (41.19±13.10)respectively.There was no statistically significant difference in the terrorist tendency dimension scores of primary school students with different education methods (F=2.33,P=0.054),and the differences in the MHT total score and other dimensions were statistically significant (F=4.35-16.88,P＜0.01).(3) There were statistically significant differences in the scores of the total score of psychological problem tendency,dimensions of learning anxiety,anxiety to people,allergy tendency and impulse tendency of primary school students who were parental rearing or not (t=2.09-3.67,P＜0.05).However,there was no statistically significant difference in the dimensions of loneliness tendency,self-accusation tendency,physical symptoms and terrorist tendency (P＞0.05).Conclusion Family education methods and parenting styles has a impact on the mental health of primary school students.

Objective To explore the clinical effect of psychological nursing pathway for patients with hepatocellular carcinoma (HCC) who are receiving 125I particle implantation therapy.Methods A total of 130 HCC patients,who were scheduled to receive 125I particle implantation therapy,were randomly and equally divided into the study group and the control group with 65 patients in each group.Traditional routine psychological nursing method was conducted for patients of the control group,while routine psychological nursing that was based on psychological nursing pathway was carried out for patients of the study group.The patient's conditions of both groups were evaluated by using self-rating anxiety scale (SAS),trait coping style questionnaire (TCSQ) and the quality of life assessment scale (SF-36).The results were statistically analyzed with t test.Results After nursing intervention,SAS scores of the study group and the control group were (43.76±6.25) and (46.16±7.19) points respectively,the difference between the two groups was statistically significant (t=-3.92,P＜0.001).The PC and NC values of the TCSQ score in the study group were (41.61±4.82) points and (20.53±5.88) points respectively,while those in the control group were (37.32±4.62) and (22.80±6.93) respectively,the differences between the two groups were statistically significant (t=5.944 and t=-3.316 respectively,P＜0.01).One month after discharge,the mental health scores of the study group and the control group were (74.58±8.68) points and (71.53±8.16) points respectively,the emotion function scores were (79.46±10.05) points and (75.13±15.09) points respectively,and the differences in both mental health score and emotion function score between the two groups were statistically significant (P＜0.05).Conclusion Standardized psychological nursing pathway can make nurses more clear about the content of psychological nursing work and enable nurses to actively plan and complete psychological nursing care measures,so that patients can get prospective,individual and comprehensive psychological care to alleviate his or her negative emotion,and to improve the quality of life.