0.05) in results. The positive rate was 26.99% for these 3 kits, and any two of them could achieve a positive rate of 53.95%. The total positive rate was 80.95%, which was significantly different as compared with non-tuberculosis group(P

Objective To study two new factor Ⅸ mutations Cys82Ser and Ile288Ser in vitro and research the molecular mechanism of haemophilia B. Methods PcDNA3. 1 ( - ) FⅨwt expression plasmid was prepared. The mutated FⅨcDNA expression plasmids, PcDNA3.1 ( - ) FⅨM1 (Cys82Ser) and PcDNA3. 1 ( - ) F Ⅸ M2 (Ile288Ser) were constructed by megaprimer method respectively. Transient expression experiments were performed using HEK293 cells transfected with the expression vectors containing the wild-type or the mutation recombinant cDNA. PcDNA3. 1 ( - ) was used as a blank control. The expression proteins were detected by ELISA, factor activity assay and flourescence stain. Results The results suggested that the two FⅨ gene mutations did not induce the reduction of the mutant FⅨ mRNA compared with the wild-type FⅨ mRNA. The FⅨ:Ag in culture media and cell lysate of wild type conduct were assigned as 100. 0%. The results of PcDNA3.1 ( - ) FⅨ M1 mutation protein were (27. 1 ± 5. 2)% and (99.4 ±4. 1)% respectively. For PcDNA3. 1( - )FⅨM2, the results were (5.3 ± 1.8)% and (31.7 ±2. 5)% respectively. The FⅨ: C in culture media of wild type conduct was also assigned as 100. 0%. Then the two types of mutant protein were ( 8. 5 ± 3.2 ) % and ＜ 1%, respectively. Immunofluorescence microscopy result suggested that the intensity of perinuclear spot was reduced in cells transfected with PcDNA3.1 ( - ) FⅨM2 while staining for PcDNA3. 1 ( - ) FⅨM1 was predominantely diffuse without perinnclear enhancement. Conclusions These results strongly suggest that the FⅨ Cys82Ser mutation protein is not been correctly folded, by any possibility. The mutation protein has secretion defect. The secretion dysfunction and the protein degradation intracellular are possiblely the molecular pathology of Ile288Ser mutant protein.

Objective To detect the expression of hypoxia-inducible factor 1α (HIF-1α) and bcl-2 ovarian serous carcinoma and their clinical significances. Methods Paraffin specimens including 61 cases of ovarian serous carcinoma and 50 normal ovarian tissues were selected. The expressions of HIF-1α and bcl-2 proteins were detected by immunohistochemical EnVision method and their relationship between them was analyzed. Results The positive rate of HIF-1α and bcl-2 proteins expression in 61 ovarian serous carcinoma was 68.9%and 54.1%, respectively. There was a significant difference between the two groups (χ2=55.381, P< 0.05; χ2= 38.493, P< 0.05). The clinical pathological parameters showed that the positive expression of HIF-1αand bcl-2 proteins were not related with the age (P>0.05). HIF-1αpositive expression was correlated with tumor grades, the state of lymph node metastasis and FIGO stages (χ2=4.931, 25.008, 5.610, P<0.05). Bcl-2 was significantly associated with tumor grades and lymph node metastasis (χ2= 6.956, 33.869, P<0.05), but not with FIGO stages (χ2=3.391, P>0.05). The expression of bcl-2 was positively correlated with HIF-1α in ovarian serous carcinoma (r= 0.304, P= 0.017). Conclusions The expressions of HIF-1α and bcl-2 play a synergic role in the progression of ovarian serous carcinoma. The combined detection of HIF-1αand bcl-2 is effective for patients'prognosis judgment.

BACKGROUND: Some researches suggest that placenta is regarded as a new source of mesemchymal stem cells (MSCs). Adherent cells derived from placenta tissue have similar morphological characteristics and surface markers to MSCs; meanwhile, they can differentiate into osteoblasts and nerve cells.OBJECTIVE: To find a new source and way for MSCs separation.DESIGN: Observational study.SETTING: The Third People's Hospital of Wuxi.MATERIALS: Placenta was sourced from cesarean in our department of obstetrics and gynecology and provided confirmed consent from the relatives and ethics committee. The main reagents contained culture medium, sABC kit, DAB staining kit, caprine-anti-rat FITC, recombinant human basic fibroblast growth factor (rh-bFGF), rabbit-anti-human glial fibriliary acidic protein (GFAP).METHODS: The experiment was carried out in the Laboratory of Cells & Molecular Biology, the Third People's Hospital of Wuxi from May 2005 to August 2006. Placenta entity tissue was digested, adherent cells were cultured, morphological characteristics were observed, and growth curves of placenta-derived multipotent cells (PDMCs) in various generations were drawn. On the 1st and 7th days, supernatant was derived from cells in primary culture to measure content of β-glycerophosphate disodium (β-HCG) with chemiluminescence technique. In addition, expression of surface antigen and differentiating potency were detected at the same time. At 24 hours (at phase of nerve cells) and 2 weeks (at phase of osteoblasts) after inducible differentiation, cultured cells were dealt with routine immunocytochemical stain, and then they were observed under routine microscope or fluorescence microscope.MAIN OUTCOME MEASURES: ① Morphological indexes and growth curves of PDMCs; ② measurement of β-HCG in supernatant of PDMCs and expression of antigen of PDMCs with flow cytometer; ③ immunohistochemical analysis of PDMCs during and after inducible differentiation.RESULTS: ① Primary culture of PDMCs: After digestion of placenta tissue, a few of adherent cells were obtained and gradually formed thin and flat monolayer cells two weeks later. The monolayer cells grew like whirlpool or cluster. With the increase of cell density, soma was slenderer and slenderer and like fibroblast. ② Growth curves of PDMCs: Growth latency ranged from 2 to 8 days after cell inoculation. During this period, adherence was observed gradually but not obviously amplified. Eight days later, cells entered log growth phase. During this period, proliferation was active and cell process surrounding extended under phase contrast microscope. A lot of MSCs were observed in division phase between two nuclei. In addition, density was increased and cells connected to each other. Within 11-14 days, growth curve gradually entered platform phase. MSCs covered the bottom of bottle, cells slowly expanded, and primary culture stopped. ③ Measurement of β-HCG: Expression of β-HCG was not detected in supernatant at two time points. ④ Characteristics of surface antigen of PDMCs: PDMCs could express CD29, CD44 and CD105, but not CD34, CD45,CD19 and CD106. ⑤ Inducible differentiation of PDMCs: At 24 hours after inducing to nerve cells, form of PDMCs obviously changed, soma rebounded, refraction of nucleus was partially reinforced, structure was similar to dendrite and axis-cylinder, and positive neuro-specific enolase and GFAP were observed after staining.CONCLUSION:Placenta tissue contains PDMCs whose morphological function is similar to MSCs; in addition, placenta is regarded as an effective source of MSCs.

In order to better fulfill the tasks of research,to turn out more quality papers,to produce outstanding results,and to further strengthen management and supervision of scientific research,the"quantitative economic management of scientific research quotas" was established in the hospital.Applying of the measure in scientific research management in the past eight years it was shown that the desired results were achieved,the academic advancement and the personnel growth were greatly promoted.

Objective: To study the expression of negative costimulatroy molecule B7-H4 in non-small cell lung cancer (NSCLC) tissues and its relationship with the clinical features of NSCLC. Methods: Fifty-two NSCLC specimens from pa-tients who were pathologically diagnosed in our hospital during January 2008 to April 2009 were included in the present study. B7-H4 expression and infiltration of CD3~+ T cells in NSCLC tissues were detected by immunohistochemistry. The correlation between B7-H4 expression, CD3~+ T infiltration, and the clinical features of NSCLC was studied. Results: The positive rate of B7-H4 in 52 NSCLC tissues was 48.08% (25/52), and B7-H4 expression in normal lung tissues was neg-ative or low (P <0.05). B7-H4 expression was positively correlated with the clinical tumor stages and lymph node metas-tasis of NSCLC (P < 0.05), and negatively correlated with tumor infiltration of CD3~+ T cells (P < 0.05), but had no re-lationship with clinicopathologie parameters of NSCLC (P > 0.05). Conclusion: Negative costimulatroy molecule B7-H4 may play important roles in the development of NSCLC. Positive expression of BT-H4 is correlated with the clinical tumor stages and lymph node metastasis of NSCLC, which provides a foundation for diagnosis and therapy of NSCLC.

Objective To identify the clinical phenotypic diagnosis and gene mutation detection of two kindreds with PS deficiency. MethodsPS: A was measured by chromogenic substrate method;TPS:Ag, FPS: Ag levels were measured by ELISA method; PS gene(PROS1 gene)was detected by amplifying 15 exons and flanking intron sequences from the propositus with PCR method. PCR products were purified and directly sequenced. Results For propositus 1,PS: A was 48.6% ,TPS: Ag was 136 mg/L, FPS : Ag was 41 mg/L, PROSI gene exon 2 was in c. Heterozygous base substitutions was detected in C121T locus, which led to Arg-1Cys (R-1C) heterozygous roissense mutation encoded in PS proteins. For propositus 2, PS: A was 29.2%, TPS: Ag was 83 mg/L, FPS: Ag was 26 mg/L, PROSI gene exon 14 was in c. Heterozygous base substitutions was identified in CI687T locus, in which Gln.522Stop heterozygous nonsense mutation was encoded in PS proteins. Conclusions c. C121T is a novel mutation locus detected in PROS1 gene. This heterozygous mutation could lead to type Ⅱ PS hereditary deficiency, while c. C1687T heterozygous mutation could bring about type Ⅰ PS hereditary deficiency.

OBJECTIVE:To evaluate the management of the free anti-TB drugs in He'nan province,and to analyze ex-periences and find out the gap.METHODS:Based on the requirements of the Chinese TB Control Program-Free anti-TB drug management manual,a questionnaire comprised of 17 items was developed and 20 TB drug storerooms at city or county level were randomly sampled for on the spot investigation.RESULTS:Of the total drug storerooms investigated,75% had yearly drug demand plan,but only 35% was up to the standard in inventory control,95% had no expired drug,90% had inventory/supply vouchers and detailed inventory records,only 25% achieved conformity between records and physical counts.The condition of drug storerooms and the storage of drugs were unable to meet the requirement.CONCLUSION:Tuberculosis Control Agency at different levels haven't paid due attention to the management of free anti-TB drugs.The personnel in this agency should raise their drug management responsibility from aspects of saving public belongings and ensuring patients' medication quality.

Objective To investigate the phenotype, genotype and molecular mechanisms in four Chinese pedigrees with venous thrombosis caused by hereditary PC deficiency. Methods The plasma activity of PC: A, TPS: A and FPS: A of the probands and their family members were detected with chromogenic and coagulation assay. The antigen of PC and FPS were identified with ELISA. Thrombin generation tests were applied to indicate the coagulation status. All of the nine exons and intron-exon boundaries of PC gene and PS gene were amplified by PCR and directly sequenced for mutaiton investigation. Results Compound heterozygous mutations of L-34P, K150del and A209V with 36% of PC: A and 57% of PC: Ag were identified in proband 1. PC: A was 46% , PC: Ag was 64. 4% while TPS: A, FPS: A and FPS: Ag were 36% , 19.5% and 20.9% respectively in proband 2. Two independent heterozygous mutations of R147W in PC gene inherited from his mother and T519stop in PS gene inherited from his father were identified. The anticoagulant activity of Proband 2 and his parents were declined in thrombin generation assay. In proband with PS defeciency and his father, the inhibition of thrombin generation capacity was decreased with exogenous APC, while his mother did not have significant difference. In Proband 3, PC: A was 32% while PC: Ag was 48.42% . Two independent mutations of R147W and R178W in Exon 7 were detected. Compound heterozygous mutations of R178W and D255H,with 21% of PC : A and 18. 36% of PC: Ag were identified in the Proband 4. Conclusions Hereditary PC deficiency or combined PC and PS deficiency result in venous thrombosis in four Chinese families. Mutants of L-34P, A209V, R178W, R147W and D255H might be the molecular mechanisms of PC deficiency.

Objective To investigate the clinical phenotype and genotype in three probands with antithmmbin(AT)deficiency and their families,and to identify the molecular mechanism of AT deficiency.Methods Chromogenic substrate method and immunoturbidimetry assay was used to detect the plasma levels of AT:A and AT:Ag,respectively.Genomic DNA was extracted from the peripheral blood.All 7 exons and the flanking sequences were amplified by PCR.and the abnormal mutant genes were analyzed by direct sequencing.Western blot was used to detect the AT levels and thrombin generation tests were used to detect coagulation status.Results The plasma levels of AT:A and AT:Ag of the three probands declined by 50%.G7386C(Trp225Cys)mutation in exon 4,C2591G(Ser36stop)in exon 2 and C9819T(Arg359stop)in exon 5 were characterized in the three prebands and they could result in W(Trp)225C(Cys)missense mutation,S(Set)36X(stop)nonsense mutation and R(Arg)359X(stop)nonsense mutation respectively,The testing results of phenotype and genotype from some of their family members showed consistent with results from the probands.Western blot results indicated that the Icyels of PC:Ag were lower compared with the normal pooled plasma.The hypercoagulative status was present in the probands using thrombin generation tests.Conclusions Type Ⅰ hereditary AT deficiency was found in these three families.The 3 heterozygous mutations.W225C,S36X and R359X are genetic defects of hereditary AT deficiency.W225C and S36X have not been described before.