BACKGROUND: Human granulocyte-macrophage colony stimulating factor (hGM-CSF) is a kind of cytokine which can stimulate the differentiation and proliferation of haematopoictic precursor cells and plays an important role in the process of immunoloregulation. Escherichia coli (E.coli) expression system has advantages,such as low cost and high output, over other systems.Therefore, E.coli is still the most commonly used exogenous gene expression system.OBJECTIVE: To observe the expression of hGM-CSF in the E.coli swain DH5α.DESIGN, TIME AND SETTING: The present observational experiment was performed at the Central Laboratory of Biotechnology Research,China Three Gorges University between February and June 2007. MATERIALS: Plasmid pBR322hGM-CSF was purchased from American Type Culture Collection, USA.hGM-CSF antibody was sourced from Sigma Company, USA.IPTG, X-gal,and vector pMGT-18 were purchased from Shanghai Bioengineering Technology Company, China. METHODS: Primer was designed according to hGM-CSF gene. Taking cloning vector plasmid pBR322-hGM-CSF as template, hGM-CSF gene was acquired and recombined into prokaryotic expression vector pGEX-4T-1. Recombinant plasmid confirmed by enzyme digestion and sequencing was transformed into E.coli strain DH5α and induced by isopropy-β-D-thiogalactoside (IPTG). Expression product was identified by sodium dodecyl sulfate polyacrylamide gel electropheresis (SDS-PAGE) and detected by Western blotting assay. MAIN OUTCOME MEASURES: hGM-CSF expression in the E.coli strain DH5α. RESULTS: SDS-PAGE results revealed that the recombinant hGM-CSF protein with relative molecular weight of 40 500 was produced up to approximately 17.9% of total protein. Western blotting detection results indicated that there was a 40 500 bp brand. CONCLUSION: The present study reconstructed prokaryotic expression vector pGEX-hGM-CSF, induced its expression in the E.coli strain DH5α, and obtained hGM-CSF fusion protein.

Objective To study the clinical factors of sclerema neonatorum complicated with pneumorrhagia and propose therapy and control measures in order to.improve clinical treatment.Methods27 cases of sclerema neonatorum complicated with pneumorrhagia treated in Chaozhou Central Hospital from January 2005 to December 2007 were investigated retrospectively.Results In all 96 cases of sclerema neonatorum,27 Were complicated with pneumorrhagia.mortality of which was 67%.9 cases of pneumorrhagia without mechanical ventilation died totally.Early tracheal intubation discovered 9 cases of pneumorrhagia,mortality was 56%.5 cases died within the 6 oronasal emissing blood,taking up 83%in mortality.There were significant differences between them(P＜0.05).Conclusion The incidence of pneumorrhagia in sclerema neonatorum was closely related to fetal month,age,scleredema degrees,acidosis,birth weight ect.The more severe scleredema,the lower birth weight and the fewer fetal month were,the higher incidence rate was.

Objective To further explore the relationship between increasing genetic material and clinical manifestation of partial trisomy 5p .Methods G‐banding karyotypes of peripheral blood lymphocytes in the patient and his parents ,and at the same time to summary the partial trisomy 5p clinical performance .Results patient ,46 ,XX ,der(6)t(5 ;6)(p13;q25) mat ;partial trisomy for 5p13→pter resulting from the balanced translocation of the mother .Mother:46 ,XX ,t(5;6)(p13 ;q25);carrier of a balanced 5/6 translocation .Father :46 ,XY .Conclusion The phenotype of trisomy 5p may be associated with express and function of gene at spe‐cial chromosome region .

Objective To investigate the therapeutic effects and side effects of FLAG and MEA regimen in the treatment of relapsed and refractory adult acute myeloid leukemia.Methods Use retrospective analysis to Observe the therapeutic effects and side effects of the 51 cases of relapsed and refractory adult acute myeloid leukemia (M3 except) from January 2009 to June 2012 in our hospital,which are divided into FLAG group (23 cases) and MEA group (28 cases) according to chemotherapy.Results In FLAG group,the rate of complete remission was 30.4 ％ (7/23),the rate of partial remission was 17.4 ％ (4/23),the effective rate was 47.8 ％ (11/23).In MEA group,the rate of complete remission was 35.7 ％ (10/28),the rate of partial remission was 21.4 ％ (6/28),the effective rate was 57.1％ (16/28),difference between two groups was not statistically significant. Both groups appeared Ⅳ degrees myelosupression,and there were no significant differences between them on incidences of secondary infection [95.7 ％ (22/23) vs 89.3 ％ (25/28)] and haemorrhagia [82.6 ％ (19/23) vs 85.7 ％ (24/28)].Difference on cardiac toxicity was statistically significant.Conclusions Compared with MEA regimen, FLAG regimen are similar effective and can be well tolerated,which has lower cardiac toxicity. Thus, FLAG regimen can be used as first-line treatment for relapsed and refractory adult acute myeloid leukemia.

Objective:To explore the protective effect of emodin on D-galactosamine (D-GalN)/lipopolysaccharide (LPS)-induced acute liver injury and its mechanism.Methods:A total of 40 male BALB/c mice were randomly (random number) divided into 5 groups ( n=8 in each group): the control group, the emodin group, the D-GalN/LPS group, the emodin+D-GalN/LPS group and the 3-MA+emodin+D-GalN/LPS group. D-GalN (700 mg/kg) and LPS (10 μg/kg) were intraperitoneally injected to induce acute liver injury in mice. Autophagy inhibitor 3-MA (15 mg/kg) and/or emodin (20 mg/kg) were intraperitoneally injected 30 min before the liver injury model. The animals were sacrificed under anaesthesia 6 h after D-GalN/LPS challenge, blood samples and liver tissues were collected. The levels of alanineaminotransferase (ALT) and aspartateaminotransferase (AST) in serum, and myeloperoxidase (MPO) activity of liver tissues were determined by colorimetric quantitative method; the levels of tumor necrosis factor-α (TNF-α), and interleukin-6 (IL-6) were measured by ELISA; the expression of LC3-II and Beclin 1 in the liver tissues were evaluated by Western blot; the pathological changes of liver was evaluated by HE staining. Animal survival rate was also analyzed. The one-way ANOVA was use to compare quantitative data, SNK- q test was used for pairwise comparison between two groups, and Games-Howell test was used when homogeneity of variance were not met. Results:Compared with the control group, the levels of ALT, AST, TNF-α, IL-6 and MPO activity [(2 476.80 ± 263.14) U/L, (271.71 ± 47.15) U/L, (537.92 ± 89.35) pg/mL, (169.74 ± 25.52) pg/mL, and (1.37 ± 0.22) U/mg] were obviously increased in the D-GalN/LPS group ( P<0.05). Compared with the D-GalN/LPS group, the levels of ALT, AST, TNF-α, IL-6 and MPO activity [(1 248.01 ± 380.70) U/L, (142.59 ± 34.63) U/L, (288.91 ± 67.21) pg/mL, (61.83 ± 13.64) pg/mL, and (0.80 ± 0.21) U/mg] were obviously decreased in the emodin+ D-GalN/LPS group ( P<0.05). Compared with the D-GalN/LPS group, the histopathological abnormalities in liver tissue were significantly alleviated and the survival rate of mice was improved in the emodin+ D-GalN/LPS group. Compared with the control group, the expression of LC3-II and Beclin1 was decreased in the liver tissue in the D-GalN/LPS group, while compared with the D-GalN/LPS group, the expression of LC3-II and Beclin1 was increased in the emodin+ D-GalN/LPS group. With co-administration of 3-MA, the protective effects of emodin in acute liver injury were reversed, the levels of AST, ALT, TNF-α, IL-6, and MPO [(2 398.78 ± 233.57) U/L, (242.79 ± 43.46) U/L, (505.07 ± 67.89) pg/mL, (151.46 ± 14.11) pg/mL, and (1.27 ± 0.15) U/mg] were increased, and the pathological damage of liver tissue was aggravated. Conclusions:Emodin alleviates D-GalN/LPS-induced acute liver injury in mice, which may be related to the activation of protein LC3-II, Beclin1 and restored autophagy.

Objective:To study the clinicopathologic characteristic of the primary extranodular non-Hodgkin's lymphoma in head and neck.Method:Clinical manifestation and the characteristic of clinicopathology of 216 extranodular non-Hodgkin's lymphoma patients in head and neck were analyzed retrospectively.Result:The age of thirty-one to sixty and seventy-one to eighty years old were more common age range in the group of patients. Nasal cavity was the most commonly primary involved site(95 cases, 44.0% ) , secondly was tonsil ( 47 cases, 21.8%). The most common histologic subtype was NK/T cell lymphoma, which accounted for 49.5%(107 cases)of cases, secondly was diffuse large B cell lymphoma (DLBCL, 58 cases, 26.7%). The most common histologic subtype in different swtach groups respectively is: NK/T cell lymphoma in nasal cavity(74 cases,77.9%), DLBCL in paranasal sinus(6 cases,50.0%), DLBCL in tonsil(27 cases,57.4%),NK/T cell lymphoma in nasopharynx(17 cases,44.7%), and DLBCL in lingual root(5 cases,45.4%).Conclusion:We conclude that primary extranodular non-Hodgkin's lymphoma is common in head and neck patients. There is characteristic in age, primary involved site and histologic subtype, which is helpful to understand these characteristic for pathologic diagnosis.

Objective To select the aptamer to an extracellular soluble fragment of recombinant human TGF-? receptor Ⅱ (TGF-? RⅡ) in order to antagonize TGF-? effectively by using systematic evolution of ligants by exponential enrichment (SELEX). Methods Initial random RNA library was transcripted in vitro from ssDNA 5′-TAATACGACTCACTATAGGGAGGACGATGCGG-N60-CAGACGACTCCCCGA-3′; rhTGF-? RⅡ was used as target protein. Totally,selection of 8 times was carried out in SELEX experiment. Membrane binding assay was performed to detect the evolution of enriched RNA library; Electrophoretic mobility shift assay (EMSA) was done to determine the affinity between the selected nucleic acid sequence and TGF-? RⅡ. Results Evolution of the enriched RNA library along the increased affinity to TGF-? RⅡ was observed with the development of selection. Two types of dominant sequences were isolated and named as sequence A and sequence B. In membrane binding assay,both sequences A and B showed obvious affinity to TGF-? RⅡ. However,no retarded bands were seen in EMSA. Conclusion The affinity of sequences A and B to TGF-? RⅡ is beyond satisfaction. However,possible sequences with improved affinity to TGF-? RⅡ can be selected by post-SELEX on the basis of candidate sequences A and B.

Objective To investigate the effect of the artificial cycle on the prognosis after transcervical resection of adhesion (TCRA) and its mechanism .Methods 80 patients with intrauterine adhesion were randomly divided into the observation group and the control group .The observation group were performed TRCA and postoperative artificial cycle for successive 3 months :oral es-tradiol valerate(9 mg once daily) for 21 d ,adding oral medroxyprofgesterone acetate(10 mg once dialy) on 15 d ,then the medication discontinuation for 7 d ,which was taken as 1 cycle;the control group received only TCRA .The re-examination was performed after 3 months .The levels of follicle stimulating hormone(FSH) ,luteinizing hormone(LH) ,estradiol(E2 ) ,prolactin(PRL) ,progesterone (P) and testosterone(T) were examined by ELISA after TCRA three months .The expression of MMP-9 and TGF-β1 in adhesive endometrium were detected by real-time PCR and Western blot .Results Compared with the control group ,the artificial cycle thera-py combined with the operation could significantly increase the effective rate of TCRA for treating mild and moderate intrauterine adhesion(P<0 .05);the seum levels of FSH ,LH ,E2 ,PRL ,P and T had no statistical differences between the observation group and the control group(P>0 .05);the artificial cycle significantly improved the adhesive degree of intrauterine adhesion ,increased the MMP-9 expression and decreased the TGF-β1 expression .Conclusion Artificial cycle could improve the degree of uterine cavity ad-hesion after TCRA ,which the mechanism might be related to the increase of MMP-9 expression and decrease of TGF-β1 expres-sion .

Objective To discuss the application value of DWI and ADC on predicting therapeutic effect of radiotherapy treatment in NPC. Methods Twenty four local recurrent cases and 38 non-recurrent cases after radiotherapy treatment in NPC were reviewed. MRI and DWI-MRI were performed at pre-radiotherapy and 3, 6, 12 months after treatment, the ADC values of the lesions were analyzed by SPSS 18.0 statistical software. ROC curves based on the ADC values were measured in 3, 6, 12 months after treatment plotted to analyze the threshold ADC value for confirming recurrence. Results The recurrent group and newly diagnosed group showed significantly high signal on DWI, while the non-recurrent group acquired low or mixed signal. The ADC values of the primary tumor in the recurrent group and the non-recurrent group were (0.709 ± 0.078) × 10-3 and (0.693 ± 0.089) × 10-3mm2/s, respectively, t=-0.717,P>0.05, respectively.The ADC values of the primary and recurrent tumor in the recurrent group were (0.730± 0.068) × 10-3mm2/s and (0.709 ± 0.078) × 10-3mm2/s, t=-1.000,P>0.05 , respectively.There were statistical differences between the recurrent group and the non-recurrent group for ADC in 3, 6, 12 months after treatment:(1.128 ± 0.179) × 10-3 and (1.358 ± 0.145) × 10-3mm2/s, t=5.567,P<0.01;(1.164 ± 0.174) and (1.450 ± 0.102) × 10-3mm2/s, t=7.310,P<0.01;(1.107 ± 0.180) × 10-3 and (1.584 ± 0.125) × 10-3mm2/s, t=11.189,P<0.01;respectively. Take 1.29 × 10-3 mm2/s,1.32 × 10-3mm2/s,1.37 × 10-3mm2/s respectively in 3, 6, 12months after treatment as the diagnostic threshold to predict tumor recurrence. The sensitive , specificity, and accuracy were (83.3%, 73.7%, 77.4%), (83.3%, 89.5%, 87.1%), (100.0%, 94.7%, 96.3%).Conclusions Both DWI and ADC value are important for diagnosing and predicting recurrent NPC after radiotherapy treatment, DWI and ADC can be used to regular follow-up after radiotherapy, to further improve the rate of early diagnosis of recurrent NPC.