Objective: To isolate Mycobacterium tuberculosis H37Rv and H37Ra genes especially,and to construct two cDNA libraries by using suppression subtractive hybridization (SSH).Methods: We used suppression subtractive hybridization (SSH) to clone the differential genomic genes between Mycobacterium tuberculosis virulence strain H37Rv and attenuated strain H37Ra.After two times of subtractive hybridization and two times of PCR,the products of last PCR amplification were inserted into pGEM-T Easy vector and be transformed into the E.coli DH5α and screened of blue and white clones of the transformants.The subtracted cDNA library of differentially expressed genes were identified by RT-PCR.Results:High or especially expressed genes as tester had been obtained by SSH in correctitude reaction (H37Rv as tester) and reverse reaction (H37Ra as tester),the cDNA libraries of A and B of Mycobacterium tuberculosis H37Rv and H37Ra were successfully constructed.90% of the colonies were white clones,the single band of the colonies was 75% and 80%.Conchision:The cDNA libraries of Mycobacterium tuberculosis H37Rv and H37Ra were successfully constructed using SSH technique,which lay a solid foundation for screening and cloning new specific differentially expressed genes in them.

Objective To investigate the relationship between the levels of total serum IgE and IL-4 in patients with seasonal contact dermatitis. Methods Total serum IgE was tested by immunoradiometric assay(IRMA) in 29 cases of seasonal contact dermatitis, 35 cases of allergic contact dermatitis and 24 cases of healthy controls. Production of IL-4 was detected by ELISA from peripheral blood mononuclear cells (PBMC) cultured with PHA for 48 hours. Results The levels of IL-4 secreted by PBMC and total serum IgE were significantly higher in patient with seasonal contact dermatitis than those in control group (P

Objective To investigate the status of social support and job satisfaction of community nurses and their relationship in Wuhan. Methods A total of 230 community nurses from 20 community health service centers in Wuhan were investigated by general information questionnaire, job satisfaction scale of community nurse and social support scale. Results The total score of job satisfaction was (96.14±14.76) points, the highest score of dimension was leadership and management (23.64±4.60) points, and the lowest was pay and welfare (10.83±3.00) points. The total score of social support was (37.17±7.05) points, the highest score of dimension was perceived social support (20.15 ± 4.48) points, the lowest was availability (8.05±1.57) points. There was a positive association between community nurses′social support and job satisfaction (r=0.248, P<0.05). Conclusions Social support is one of the predictors of job satisfaction. In order to make sure the sustainable development of community health services, it is necessary to know the status and take corresponding measures to improve the levels of social support and job satisfaction.

Objective To investigate the effects of recombinant human pigment epithelium derived factor (rhPEDF)on in vitro proliferation of and expressions of interleukin 6(IL-6), IL-8 and vascular endothelial growth factor (VEGF)in cultured human HaCaT keratinocytes. Methods Some cultured HaCaT cells were treated with rhPEDF at various concentrations(25, 50, 100 μg/L)for different durations, and some treated with RPMI 1640 medium only served as the control group. Cell counting kit-8(CCK8)assay was performed to evaluate cell proliferation after 24-, 48- and 72-hour treatment, reverse transcription (RT)-PCR to measure the mRNA expressions of IL-6, IL-8 and VEGF in HaCaT cells after 24-hour treatment, and Western blot to detect the protein expressions of IL-6, IL-8 and VEGF in HaCaT cells after 48-hour treatment. Statistical analysis was carried out by two- and one-way analysis of variance, Student-Newman-Keuls(SNK)-q test and Pearson correlation analysis. Results After treatment with rhPEDF of 25-100 μg/L for 24 - 72 hours, the proliferation of HaCaT cells was significantly inhibited to different extents compared with the control group(all P < 0.05), and the inhibition rate significantly increased with the increase in treatment duration and concentrations of rhPEDF(F = 1115, 329.9, respectively, both P < 0.001). Moreover, there was a significant decrease in the expressions of IL-6, IL-8 and VEGF mRNAs(at 24 hours)and proteins(at 48 hours)in HaCaT cells after treatment with rhPEDF of 25 - 100 μg/L compared with the control group(all P < 0.05). The expression levels of VEGF mRNA as well as IL-6 and IL-8 proteins all significantly decreased with the increase of rhPEDF concentrations (all P < 0.05). The mRNA expressions of IL-6 and IL-8 were significantly lower in the 100-μg/L rhPEDF group than in the 25-μg/L rhPEDF group (both P < 0.05), and the protein expression of VEGF was significantly weaker in the 100-μg/L rhPEDF group than in 25-and 50-μg/L rhPEDF groups (both P < 0.05), but similar between the 25- and 50-μg/L rhPEDF groups (P > 0.05). Conclusions rhPEDF can inhibit the proliferation of HaCaT cells, and down-regulate the mRNA and protein expressions of IL-6, IL-8 and VEGF.

Objective To investigate the effects of rottlerin on in vitro proliferation of and expressions of interleukin (IL)-17C,CCL20 chemokine,and nuclear factor (NF)-κB in cultured human HaCaT keratinocytes.Methods Some HaCaT cells were divided into several test groups treated with rottlerin at concentrations of 0.5,1.0,2.0 and 4.0 μmol/L,a solvent group treated with RPMI 1640 culture solution containing the same volume of dimethyl sulfoxide (DMSO) as that of 4.0 μmol/L rottlerin,and a control group treated with RPMI 1640 culture solution.Cell counting kit-8 (CCK8) assay was conducted to estimate the proliferative activity of HaCaT cells after 24-,48-and 72-hour culture,RT-PCR to determine the mRNA expressions of IL-17C and CCL20 after 48-hour culture,and Western blot to measure the protein expressions of IL-17C,CCL20 and NF-κB after 48-hour culture.Statistical analysis was carried out by using repeated-measures analysis of variance,one-way analysis of variance and Pearson correlation analysis with the SPSS16.0 software.Results Rottlerin showed an inhibitory effect on the proliferation of HaCaT cells,and the inhibitory effect increased over time (F =126.936,P ＜ 0.05) and with the increase of rottlerin concentrations (F =838.308,P ＜ 0.05),with a significant interaction effect between rottlerin concentrations and treatment duration (F =15.961,P ＜ 0.05).After 48-hour treatment,a significant decrease was observed in the mRNA and protein expressions of IL-17C (F =206.041,233.887,respectively,both P ＜ 0.05) and CCL20 (F =143.883,162.431,respectively,both P ＜ 0.05),as well as in the protein expression of NF-κB (F =577.915,P ＜ 0.05) in the test groups with the increase in rottlerin concentrations.Conclusions Rottlerin can inhibit the proliferation of HaCaT cells in vitro,and decrease the mRNA and protein expressions of IL-17C and CCL20 likely by downregulating the protein expression of NF-κB.

Objective To explore the determination of dioxin in wastewater samples pretreated by solid phase microextraction(SPME). Methods The wasterwater samples were purified and enriched by SPME, then were determined with HRGC-HRMS for the concentration of dioxin in wastewater samples. Results The most suitable conditions of the pretreatment were 30 min microextraction at 45 ℃. The detection limit was 0.05 pg/?l. RSD was lower than 10%. The recovery rates were 99%-102%. The concentration of total dioxins in wastewater sample was 0.78 pg/?l. Conclusion The method of the determination of dioxins in wastewater by SPME was simple and quick, and presented a broad prospects for application.

Objective To investigate the endocrine disrupting chemicals contamination in source water and tap water in a city and to provide the scientific data for the water quality inspection,management and water treatment.Methods Fifteen samples of source water and tap water were collected in April,2006 and February,2007.Extraction of potential EDCs from samples was performed in columns packed with XAD-4 resins by using solid-phase extraction(SPE).Each sample was dried under a gentle air stream for 1 h and eluted with a mixture of acetone(20 ml) and dichloromethane(40 ml) for 3 times.The extracts were collected into a glass flask,evaporated under a gentle air stream and suspended again in 0.5 ml dichloromethane.Gas chromatography-mass spectrometry(GC-MS) was used for quantitative or qualitative analysis of the organic pollutants and the optimum conditions were determined.Results Seventy-eight organic pollutants were identified from the water samples.Most of them were phthalates,hydrocarbon,phenol,and benzene.Phthalates were found in all samples.Among them,bis(2-ethylhexyl) phthalate(CAS# 117817),di-n-butyl phthalate(CAS# 84742),and pentachlorophenol(CAS# 87865) were regarded as suspected endocrine disrupting chemicals.The concentration of bis-(2-ethylhexyl) phthalate and di-n-butyl phthalate ranged 0.004 75-4.450 ?g/L and 0.002 25-2.39 ?g/L,respectively.The concentration of pentachlorophenol was 0.727 ?g/L.Conclusion The source water and tap water in this city has been seriously polluted by organic pollutants,the main endocrine disrupting chemicals detected in the present investigation are phthalates and pentachlorophenol and the related health effects remain to be studied.

Objective To investigate the health behavior level of elderly patients with coronary heart disease (CHD) in order to instruct nurses how to give health education to them. Methods By con-venient sampling, 71 elderly patients with CHD in four hospitals in Wuhan were surveyed with the ques-tionnaire of health promoting lifestyle profile Ⅱ of FANG Heng-ying. The investigation results were ana-lysed. Results The findings showed that mean total score of health behavior of elderly patients with CHD obtained from the questionnaire was only 2.40,the level of health behavior was low. There were statistically significant differences among scores of elderly patients with CHD at different levels of family income and at different occupations before retirement. Conclusions The general level of health behavior of elderly pa-tients with CHD was low, especially in the aspect of health responsibility. Moreover, Patients with lower family income, peasants, and as a worker before retirement have lower health behavior level. This survey suggests that nurses should enhance the health education of the elderly patients with CHD, especially the poor and elderly farmer patients with CHD to improve their health behavior level.

Objective To investigate the effects of COX-2 antisense oligonucleotide (AsODN) on the proliferation and expression of COX-2 in human skin squamous cell carcinoma cell line Coio-16. Methods The COX-2 AsODN was synthesized artificially, and various concentrations (50, 100, 200, 400 nmol/L) of the AsODN were transfected into Colo-16 cells with lipofectin followed by additional culture for different durations. The transfection results were observed with fluorescence microscopy. Subsequently, MTT assay,Western blotting and reverse transcription PCR were used to detect the cell proliferation, protein and mRNA expression of COX-2 in Coio-16 cells, respectively. Restults Compared with untreated cells, the proliferation of Colo-16 cells was inhibited significantly at 24, 48, 72 and 96 hours after transfection with different concentrations of COX-2 AsODN (all P < 0.05), and the COX-2 AsODN of 400 nmol/L exerted the highest inhibition rate of 60.3% at 48 hour. The average gray scale was 0.763±0.070, 0.600±0.065, 0.430±0.074 and 0.251±0.045 for COX-2 protein, 0.778±0.025, 0.602±0.041, 0.417±0.031 and 0.297±0.051 for COX-2 mRNA in Colo-16 cells transfected with COX-2 AsODN of 50, 100, 200, and 400 nmol/L respectively,significantly lower than that in untreated Colo-16 cells (all P < 0.05); there was a significant difference in the expression of COX-2 protein and mRNA among the cells transfected with the four concentrations of COX-2 AsODN and untreated cells (F = 83.54, 132.48, respectively, both P < 0.05). Conehtsions COX-2 AsODN can inhibit the proliferation, as well as the expression of COX-2 protein and mRNA in Colo-16 cells, which suggests that COX-2 AsODN has a potential therapeutic effect on skin squamous cell carcinoma.

Objective To investigate the effects of a short hairpin RNA targeting epidermal growth factor recereceptor (EGFR-shRNA) on Colo-16 cell apoptosis and sensitivity to rapamycin. Methods The expression vector of EGFR-specific shRNA was constructed. Colo-16 cells were classified into 4 groups, normal control group remaining untreated, liposome group transfected with lipofectamine 2000, negative control group transfected with shRNA-NC/Iipofectamine 2000 and positive interference group transfected with the expression vector of shRNA-EGFR/Lipofectamine 2000. After additional culture, immunocytochemistry and Western blot were conducted to detect the protein expression of EGFR, and flow cytometry to measure the apoptosis in Colo-16cells. MTT assay was performed to measure the sensitivity of Colo-16 cells to rapamycin. Results Compared with the normal control group, the expression of EGFR was down-regulated by 43.3% in positive interference group (F= 44.6, P< 0.05), and the sensitivity to rapamycin was increased by 2.44 folds. The apoptosis rate in positive interference group was (12.65±0.091)%, significantly different from that in the normal control group (F = 2042.9, P < 0.05). Conclusion The plasmid expression vector containing shRNA targeting EGFR can effectively suppress the expression of EGFR by Colo-16 cells, enhance the sensitivity of Colo-16 cells to rapamycin and induce the apoptosis in Colo-16 cells.