Objective Determining the characteristics of human fetal hepatocytes in vitro. Methods Isolating and culturing human fetal hepatocytes by stepwise trypsinization of liver fragment in vitro; collecting the culture medium to determine the secretion of AFP, ALB and the functional enzymes (including ALT, AST, GGT, ALP and LDH) of different generations in culture; determining the expression of cytochrome C by immunohistochemistry; testing the effects of sodium byturate on human fetal hepatocytes. Results Human fetal hepatocytes were polygonal epithelial cells in DMEM medium. They could be maintained for 5.5 months (about 30 passages) in vitro. They secreted ALB and functional enzymes all over their cultivation. Conclusion Human fetal hepatocytes can be maintained keeping function in vitro for several months.

Objective Study the concentration of the hepatocyte growth factor(HGF) the expression product of recombinant human hepatocyte growth factor naked plasmid ,and whether the body generates immune responseafter wsing HCF. Methods Selected 21 patients with severe ischemic disease of lower extremity (Rutherford classification 4-6 class) , after signing the informed consent and divided them into four dosage groups( 4 mg, 8 mg, 12 mg and 16 mg) random.In each group the dosage was the lower limbs partly intramuscular injection following the feeding artery in experimental stage Dl and D15. The plasma samples were collected to determine the HGF concentration in Dl ( before administration), D8, D15 ( before administration) , D21, D59, and the HGF-antibody concentration in Dl, D15, D28, D59, D91. Results (1)The concentrations of HGF in the subjects ranged from 216 pg/mL to 1189.75 pg/mL.(2) HGF-antibodies were not dectected in the subjects' plasma. Conclusions After using recombinant human HGF naked plasmid through intramuscular injection, the concentration of HGF expression product in subjects' peripheral blood does not have abnormally changed and using the drug the human body immune response does not generate.

Objective To discuss the application effect of peer support education in improving theself-management ability of old patients with diabetes. Methods A total of 160 cases of old patients with type 2 diabetes who were treated and hospitalized from June 2014 to August 2015 were selected, then equally divided into observation group and control group through using random number method.Finally, 76 cases were studiedin observation group, while 75 cases in control group. The health education guidance of conventional diabetes had been used in two groups for six months, and the peer support education was also conducted for the patients in observation group on the basis of conventional education. The self-management ability, insulin standard injection knowledge-belief and attitude-behavior/practice (KAP) and glycosylated hemoglobin (HbA1c) value of old patients with diabetes were observed before intervention, 3 months and 6 months later, respectively. Also,the hypoglycemia frequency in two groups was compared during intervention. Results After three months of intervention, the score of diet, exercise management, glucose monitoring, foot care, pharmaceutical administration and smoke for self-management ability in observation group was 4.63 ± 0.92, 4.18 ± 0.97, 2.86 ± 0.96, 2.35 ± 0.59, 4.01 ± 1.43 and 0.62 ± 0.29, respectively, while that of these indicators in control group was 3.69 ± 0.83, 3.52 ± 0.67, 1.75 ± 0.67, 1.63 ± 0.72, 3.05 ± 1.39 and 0.59 ± 0.38. Except smoke (P > 0.05), the differences between two groups were statistically significant (t=2.987-5.965, P < 0.05). After six months of intervention, the score of diet, exercise management, glucose monitoring, foot care, pharmaceutical administration and smoke for self-management ability in observation group was 5.86±0.93, 5.37±0.61, 3.97±0.94, 5.06±0.92, 5.81±0.73 and 0.89 ± 0.31, respectively, while that of these indicators in control group was 3.68 ± 0.96, 3.65 ± 0.95, 1.86 ± 0.88, 2.65±0.88, 3.21±1.13 and 0.81±0.39. The difference was statistically significant (t=9.336-15.633, P<0.01), except smoke (P>0.05). Then, the insulin was normally injected in observation group after six months of intervention, the score of knowledge, attitude and behavior was 5.63 ± 1.87, 43.17 ± 6.03 and 16.57 ± 1.59, while that in control group was 3.76 ± 0.83, 23.71 ± 0.59 and 10.16 ± 0.95. The differences between two groups were statistically significant (t= 8.569, 15.623, 6.751, P<0.01). Also, the level of glycosylated hemoglobin was decreased from (8.17 ± 0.32) % to (7.16 ± 0.41) %, and the difference was statistically significant (t=-3.560, P=0.000). Conclusions Peer support education couldenhance the self-management consciousness of old patients with diabetes, standardize the insulin injection, improve the ability of self-management and improve glucose metabolism index so that it is worthy of being clinically popularized and applied.

Objective:To artificially design and express a recombinant protein named as ScFv-pLLO by fusing ScFv gene of Rituximab(C2B8)and dominant antigen epitopes from listeriolysin O(LLO),and studying its anti-tumor activity.Methods:VH and VL gene sequences of C2B8 against CD20 were acquired by searching United States Patent database,and ScFv sequence was constructed by linking VL and VH with a short peptide linker.Two CD4+T cell epitopes from LLO were selected and designed to splice ScFv sequence.The recombinant gene of ScFv-pLLO was cloned into prokaryotic expression vector and purified after induction.The capacity of ScFv-pLLO target-binding to B-cell lymphomas was evaluated by flow cytometry ( FCM ) and co-immunoprecipitation ( Co-IP ) .The effects of ScFv-pLLO on B-cell lymphomas proliferation and apoptosis were detected respectively.The immunogenicity of ScFv-pLLO was assessed by lymphocyte proliferation assay.Results: ScFv-pLLO was successfully expressed.It could bind to different B-cell lymphomas cell lines and obviously inhibit the growth of Raji cells as well as inducing apoptosis.Moreover,ScFv-pLLO was able to stimulate proliferation of spleen lymphocytes of immunized mice.Conclusion: The recombinant protein ScFv-pLLO can target-bind to B-cell lymphomas,and perform inhibitory effect and induce apoptosis on Raji cells that indicate ScFv-pLLO retain the capacity of ScFv derived from monoclonal antibody against CD20.Besides, ScFv-pLLO can induce immune response.This study provides a basis for further research about the role of ScFv-pLLO on simulating tumor cell antigens as well as being tumor vaccine adjuvant.

significant for the precaution of gastric cancer risk population. Aims:To explore gastric cancer susceptibility gene profiling in an area in Shanghai,and to assess the risk of gastric cancer susceptibility. Methods:A total of 152 patients with primary gastric cancer at Shanghai Pudong Hospital were enrolled,and 152 demographic characteristics matched patients with non-gastrointestinal diseases,non-tumor were served as controls. Gene polymorphism was determined by allele specific polymerase chain reaction. Susceptibility gene of gastric cancer was screened. Multiple genes interactions were analyzed and multiple genes risk was evaluated by DEMCHUK model. Results:Univariate analysis showed that CYP2E1,NAT2M1,NAT2M2,NAT2,XRCC1194,MTHFRA1298C,VDR TaqⅠ were susceptibility genes of gastric cancer. Multivariate analysis showed that CYP2E1(C1/ C1),NAT2M1(T/ T),NAT2M2(A/ A),XRCC1194( T/ T)and MTHFRA1298C(A/ C)were susceptibility genotypes. Synergistic effect was found between genes except MTHFRA1298C (A/ C)with NAT2M1(T/ T)and NAT2M2(A/ A)(P ﹤0. 05). Multiple genes risk analysis showed that combination OR of multiple genes was highly correlated with gene frequency,and the risk of gastric cancer was increased with the increasing number of susceptibility genes. Conclusions:CYP2E1(C1/ C1),NAT2M1(T/ T),NAT2M2( A/ A),XRCC1194( T/ T) and MTHFRA1298C( A/ C) are susceptibility genotypes of gastric cancer. Carrying multiple susceptibility genes can significantly increase the risk of gastric cancer.

Objective:To investigate the effect of MT01 on the differentiation of osteoblasts under infected condition through determining the expression level of collagen Ⅰ (Col Ⅰ )mRNA in MG63 cells treated with Porphyromonas gingivalis (Pg).Methods:The cultured MG63 cells were divided into blank control,MT01,Pg, and MT01+Pg groups.MT01 at a concentration of 1 mg·L-1 was added into the MG63 cells,and the cells were incubated for 3 h.The cells treated with PBS (1 mg·L-1 )were used as control group.Then Pg (MOI=100∶1) was added.Real-time PCR was used to detect the expression levels of ColⅠ mRNA in MG63 cells at 2,4,6,8, 12 and 24 h after incubation.Results:Compared with blank control group,the levles of ColⅠ mRNA in the MG63 cells in MT01 and MT01 + Pg groups had no significant changes at 2 and 4 h (P > 0.05);the Col Ⅰ mRNA expression levels in MT01 group at 8,12 and 24 h were increased (P < 0.05 or P < 0.01).Compared with Pg group,the expression levels of ColⅠ mRNA in MT01+ Pg at 2 and 4 h were decreased,but the expression levels of ColⅠ mRNA were increased at 6,8,12 and 24 h (P <0.05 or P <0.01).Conclusion:MT01 can up-regulate the expression level of Col Ⅰ mRNA in the infected MG63 cells; MT01 could promot the differentiation of osteoblasts under infected condition.

Objective To explore the application effect of interactive health education in insulin injections of aged patients with type 2 diabetes in the incipient stage of diagnosis and treatment. Methods 100 cases of aged patients with type 2 diabetes were hospitalized into endocrinology department from January 2014 to December 2014, the pen insulin injections were conducted for them at the beginning. They were equally divided into experimental group and control group through random number method. And, the general education guidance was conducted in control group. Meanwhile, the diabetes health education team was established in observation group on the basis of control group. Then, it was educated through the interactive lectures, interviews, teaching, repeated teaching, psychological intervention and individual assessment. Before and after education, the KAP survey of standard insulin injections were used for the patients in two groups to evaluate their degree of proficiency in the knowledge and skills about insulin injections, and the incidence of glucose metabolism level and low blood sugar. Results After intervention, the insulin injection skill in observation group was better than that of in the control group, (64.6%,31/48 vs. 24.5%, 12/149,χ2=16.441, P<0.01), while the level of glycosylated hemoglobin was reduced from (7.95±0.65)%to (6.58±0.62)%(t=7.85, P<0.01). Conclusions In the incipient stage of diagnosis and treatment, the ability of aged patients with type 2 diabetes in learning new knowledge and technology was improved through interactive health education mode. Besides, the insulin injections were standardized. Also, the glucose metabolism was improved, and the incidence of hypoglycemia was significantly reduced to ensure the patients′safety when the insulin was injected outside hospital, which was worth being clinically applied and promoted.

Objective To identify the species of cultured cells and to detect inter-species cross contamination by polymerase chain reaction (PCR).Methods From references and NCBI database,we outsourced 32 pairs of species-specific primers targeted to genomic DNA of ten common species of cultured cells.Then we screened for optional primers with high specificity and high sensitivity.PCR amplification with these primers,the genomic DNAs isolated from the tested cell line and agarose gel electrophoresis of the PCR products were done.Mixed DNA of 10 species was used as positive-control template,and water as negative-control template.Results Ten pairs of species-specific and highly sensitive primers were selected.By PCR amplification with these primers and agarose gel electrophoresis,we may easily identify the origin of cell lines and find whether the tested cell lines are contaminated by cells of other species.Conclusion This PCR assay provides a simple,rapid,sensitive,and cost-effective method to identify cell species and detect interspecies cross-contamination.

OBJECTIVETo study the effects of hypoxia and hyperoxia on the expression and activity regulation of matrix metalloproteinase-2 (MMP-2) of the hepatic stellate cell (HSC).

METHODSThe expression of MMP-2, tissue inhibitor of matrix metalloproteinase-2 (TIMP-2) and membrane type matrix metalloproteinase-1 (MT1-MMP) in cultured rat HSC under hypoxic or hyperoxic conditions were detected with immunocytochemistry (LSAB method), the contents of MMP-2, TIMP-2 in culture supernatant with ELISA and the activity of MMP-2 in supernatant with zymography.

RESULTS(1) In the situation of hypoxia for 12 h, the expression of MMP-2 increased (hypoxia group positive indexes: 5.7 +/- 2.0; control: 3.2 +/- 1.0; P < 0.01), while TIMP-2 decreased (hypoxia group positive indexes: 2.5 +/- 0.7; control: 3.6 +/- 1.0; P < 0.05) in HSC, and the activity of MMP-2 in supernatant declined obviously (hypoxia group: 7.334 +/- 1.922; control: 17.277 +/- 7.424; P < 0.01). At the different time courses of hypoxia, the change of expression and activity of MMP-2 was most notable at 6 h. (2) In the situation of hyperoxia for 12 h, the protein contents of MMP-2, TIMP-2 in supernatant were both higher than those of the control, especially the TIMP-2 (hyperoxia group A(450): 0.050 +/- 0.014; control: 0.022 +/- 0.010; P < 0.01), and so was the activity of MMP-2 (hyperoxia group total A: 5.252 +/- 0.771; control: 4.304 +/- 1.083; P < 0.05). The expression of MT1-MMP was also increased.

CONCLUSIONSThe HSC is sensitive to the oxygen. Hypoxia accelerates the expression of MMP-2 and the effect is more marked at the early stage. Hyperoxia increases the activity of MMP-2.

Animals ; Cell Hypoxia ; physiology ; Hyperoxia ; enzymology ; Liver ; cytology ; enzymology ; Male ; Matrix Metalloproteinase 2 ; analysis ; metabolism ; Rats ; Rats, Sprague-Dawley ; Tissue Inhibitor of Metalloproteinase-2 ; analysis

Objective To explore the effect of peer support education on self-management ability and survival quality of urbanized type 2 diabetes patients in rural areas. Methods From January to September 2015, a total of 117 patients with type 2 diabetes were selected from three large-scale living communities of Fengxian District.Then,they were randomly divided into observation group(58 cases)and control group (59 cases) using random number method, respectively. And the same health education contents were conducted for the patients in both two groups.The patients in control group were educated by the diabetic nurse in classroom. Additionally, the peer support education was conducted for these patients in observation group for 6 months on this basis.The diabetes patient's self-management behavior scale and 36-item Short-Form Health Survey Questionnaire(SF-36)were used to assess the patients before intervention and 6 months later,respectively.Besides,the level of glycosylated hemoglobin was measured by the automatic biochemical analyzer. The self- management ability and survival quality and glycometabolism level were assessed as well. Results 6 months later after intervention, the survival quality score in observation group was as following:physical function(PF) 73.6±11.5,role physical(RP) 86.7 ± 14.5, body pain (BP) 76.3 ± 12.3, general health (GH) 64.5 ± 13.6, vitality (VT) 67.5 ± 12.3, social function (SF) 87.1 ± 16.2, role emotional (RE) 81.1 ± 14.2, mental health (MH) 68.9 ± 12.6, while that in control group was as following:physical function(PF)51.3±10.6,role physical(RP)62.1±12.1,body pain (BP)51.3±10.8,general health(GH)51.1±11.6,vitality(VT)56.4±12.1,social function(SF)70.1±14.1, role emotional (RE) 71.2 ± 13.9, mental health (MH) 56.1 ± 10.8 (t=2.316-6.365, P<0.05). The self-management score in observation group was following:diet management 5.79±0.98,exercise management 5.41 ± 0.57, blood glucose monitoring 4.01 ± 0.90, foot care 5.26 ± 0.89, drug administration 4.47 ± 0.90, smoking 0.86 ± 0.34, while that in control group was following: diet management 3.62 ± 0.99, exercise management 3.66 ± 0.94, blood glucose monitoring 1.90 ± 0.84, foot care 2.54 ± 0.62, drug administration 3.73 ± 0.80, smoking 0.76 ± 0.42 (t=4.659-19.137, P of rest was less than 0.01 except that of smoking equaled to 0.172 and t=1.375). Conclusions Peer support education can improve the quality of life, enhance self-management ability, control the level of glucose metabolism and has a strong operability. Therefore,it is worthy of promotion.