Objective To study the mutations of ERG11 gene which encodes P450 lanosterol 14-α demethylase, and to explore the possible role of ERG11 gene in inducing fluconazole resistance in Candida glabrata. Methods ERG11 genes of 9 fluconazole-resistant Candida glabrata isolates and 10 fluconazole-sensitive Candida glabrata isolates were cloned into pUC57-T vector. The open reading frame of ERG11 gene were sequenced by two directional sequencing using universal primers. All sequences were compared with the published sequence. Results Ten kinds of synonymous point mutation were found. Neither missense mutation nor frame-shifting mutation was found. Among the 10 kinds of synonymous point mutation, 5 were found in both fluconazole-resistant and fluconazolesensitive Candida glabrata isolates, and 3 were only found in fluconazole-resistant isolates, 2 were only found in fluconazole-sensitive ones. The majority of the point mutations were located between 1320-2200 base pair of ERG11 gene. Conclusions There are ERG11 gene polymorphisms in clinical strains of Candida glabrata. ERG11 gene mutations are not found to be involved in the development of fluconazole resistance in Candida glabrata.

Objective To investigate the clinical value of classic intrafascial supracervical hysterectomy(CISH) by including the clipping of the uterine artery.Methods A total of 60 cases of benign uterine diseases were included in the study.After the uterine artery had been dissected and clipped on both sides under laparoscope,classic intrafascial supracervical hysterectomy was performed.Results All the operations were performed successfully under laparoscope.No conversions to open surgery were needed.Operating complications happened in no case.The operating time was 72~186 min(91.4?26.3 min),the amount of blood loss was 50~150 ml(76.5?20.6 ml),the time to postoperative gastrointestinal function recovery was 18~30 h(22.7?5.8 h),and the volume of pelvic drainage within 24 hours,50~160 ml(80.5?31.8) ml.Postoperative body temperature was elevated to 38.5 ℃ in 2 cases,the postoperative pyrexia rate being 3.3%.The length of postoperative hospital stay was 4~7 d.Follow-up for 6~18 months(10.6?4.2 months) in the 60 cases showed 3 cases of small amount of vaginal bleeding at 1~3 months,which were cured with the use of antibiotics and hemostatics for 5~7 d.Conclusions Classic intrafascial supracervical hysterectomy by including the clipping of the uterine artery is a safe and effective improvement to CISH technique.

Objective To investigate the clinical efficacy of total laparoscopic hysterectomy(TLH) with separating and blocking uterine arteries.Methods From January 2004 to June 2006,68 cases of uterine benign diseases underwent TLH by blocking uterine arteries after separating and clipping uterine arteries with Titanic clip.Results All operations were performed successfully without conversion to open surgery and complications.1 patient,who had twice gastrohysterectomy histories,underwent bipolar coagulating uterine arteries as to the failure of separating bilateral uterine arteries.Pelvic adhesion release was performed in 18 cases,unilateral or bilateral adnexectomy in 14 cases,oophorocystectomy in 8 cases,appendectomy in 1 case,and cholecystectomy in 1 case simultaneously.The operation time was 90-185 min,(112.6?27.5)min.The time of separating uterine artery in one side was 3-15 min,(5.2?3.4)min.The intraoperative blood loss was 50-150 ml,(86.5?39.6)ml.The time to first bowel movement was 18-48 h,(27.3?4.8)h.The rate of postoperative pyrexia was 4.4%(3/68),and the hospital stay was 4-7 d,(5.1?1.8) d.A follow-up period of 2-6 months,(3.5?1.6) months,showed 3 cases of vaginal dropping hemorrhage 1-2 months after operation,which was cured with the use of antibiotics and hemostatics for 5-7 d.Conclusions TLH with separating and blocking uterine arteries is a safe,effective and feasible procedure with less complication,so it is worthy of being recommended.

To improve 3-ketosteroid-△1-dehydrogenase (KSDH) activity and the transformation level for androst-4-ene-3,17-dione,3-ketosteroid-△1 -dehydrogenase gene (ksdD) from Arthrobacter simplex was cloned into plasmid pWB980 and expressed in B. subtilis WB600 under the control of promoter P43. The molecular weight of expressed enzyme was about 55kDa by SDS-PAGE analysis. The activitities assayed by spectrophotometrical method of intracellular and extracellular soluble enzyme were 110 ± 0.5mU and 15 ± 0.6mU per milligram of protein respectively. The transformation rate of androst-4-ene-3,17-dione by the B. subtilis recombinant cells was 45.3%. Compared with Arthrobacter simplex, the enzyme activity of KSDH expressed in B. subtilis was improved about 30 fold, and the transformation level of androst-4-ene-3,17-dione by the B.subtilis recombinant cells was improved about 10 fold. The recombinant B. subtilis cells used in biotransformation of steroids provided a new way for steroid medicines production.

Objective A intervention study was conducted with systemic health education in patients with internet addiction disorder (IAD),and evaluate the intervention effect of systemic health education.Methods 120 patients with IAD were collected,and then divided into the observation group andthe control group with 60 cases in each group.On the base of conventional treatment,the observation groupwas given systemic health education,and the control group was given general health education.The differences between the two groups were analyzed.Results The self-evaluated addiction scores,family cohesion scores,ideal adaptability scores,reality adaptability scores in the observation group before the intervention were significantly different from those after the intervention.The addiction scores,family cohesion scores,ideal adaptability scores,reality adaptability scores in the observation group were significantly different from those of the control group after the intervention.The scores of addiction ranged between 86～90,systemic health education had protection effect on IAD,the higher family cohesion scores,the better effect of IAD with systemic health education.When the ideal adaptability scores ＞46,the observation group has approximately two times the positive effect of the control group.When the reality adaptability scores ＞46,the effect of systemic health education in the observation group was better than the control group.Concl~ons Systemic health education has a positive effect on treatment of IAD,and it can improve family cohesion and family adaptability,so that the parent-kid relationship will improve.

Objective To systematic evaluate the efficacy of paricalcitol on estimated glomerular filtration rate (eGFR) and proteinuria in non-dialysis chronic kidney disease (CKD) patients.Methods According to the collaborative search strategy,PubMed,the clinical control test database of Cochrane Library,Embase,Chinese Wanfang database,CNKI,VIP database (form the date of database establishment to March 2014) were searched.Published and unpublished literature,abstracts in academic meetings (ASN,WCN,CSN) were also searched by hand.The randomized controlled trials (RCTs) about the efficacy paricalcitol on eGFR and proteinuria in non-dialysis CKD patients were selected.Review Manager Software 5.2 was used for statistical analysis.Results Seven RCTs with a total of 834 patients were included (508 in experimental group,326 in placebo group).No statistical difference of the efficacy on eGFR[SMD=-0.10,95％ CI:(-0.28-0.07),P=0.26] between lower dose paricalcitol (＜ 2 μg/d) group and placebo group,while higher dose (2 μg/d) group reduced eGFR significantly [SMD=-0.45,95％ CI:(-0.63--0.27),P ＜ 0.01].Compared with placebo,paricalcitol reduced proteinuria significantly [OR(95％C1):2.09(1.52-2.58),P ＜ 0.01],and there was no difference between different dose groups [OR(95％CI):1.09(0.62-1.91),P=0.77].Lower dose group [OR(95％C1):0.93(0.57-1.52),P=0.76] and higher dose group [OR(95％ CI):2.08(0.70-6.18),P=0.19] did not significantly increase the risk of adverse events.Conclusions Lower dose paricalcitol (＜ 2 μg/d)has no effect on eGFR in non-dialysis CKD patients meanwhile reduces proteinuria.The higher dose (2μg/d) may reduce eGFR without farther reduction in proteinuria.

Objective To investigate the influence of internet addictive disorder(IAD) patient' s family cohesion and adaptability effect with Satir family therapy.Methods According with table of random number,the patients (n =120) with IAD were divided into two groups,test group with the 60 patients and the control group with 60.All of subjects were given Linyi mental health center conventional interventions,test group with satir family therapy and the control group without the therapy.Measurements with the addiction self-test scale and the family cohesion and adaptability scale for five months before and after the intervention.The differences of the two groups were analyzed,and then the correlation analysis were used.Results After the intervention of the test group with Satir family therapy,compared to control group,the IAD score (54.28 ± 4.69) and family ideal cohesion (74.64 ±3.22),real cohension (70.42 ± 3.66),ideal adaptability (54.08 ± 5.78),cohesion dissatisfaction degree (5.07 ±1.64) and adaptability dissatisfaction degree (2.23 ± 0.85) score were all had statistically significant (P ＜ 0.05 or 0.01).IAD score,ideal cohesion,real cohension,ideal adaptability and real adaptability score,before and after the intervention between the control group and the test group had statistically significant (P ＜ 0.05 or 0.01).Conclusion The Satir family therapy can improve family cohesion and the adaptability,and also effectively improve the parent-child relationship.

Objective To express and purify the envelope ( E) protein of Japanese encephalitis virus (JEV) in soluble form.Methods Various prokaryotic expression vectors , host strains and induction conditions including time and temperatures were screened to obtain an optimum prokaryotic expression system for JEV E protein in soluble form .The expressed protein was purified by using nickel column chromatography and gel filtration chromatography .Results The soluble JEV E protein , accounting for 23% of the totally bacteria soluble protein was effectively expressed by using the recombinant plasmid PBCX -E406 at low tem-perature.The purity of the expressed protein reached up to 85%after the purification by using nickel column and gel filtration chromatography .Conclusion Soluble JEV E protein was successfully expressed and puri-fied in a simple and efficient way .It would provide a useful tool for further investigation on JEV infection , attenuation mechanism of JE live vaccine strain SA 14-14-2 and the quality control of JE vaccine .It can also be used for the development of diagnosis assay for JEV .

AIM:As a factor that can improve cell growth,there are few studies about the effect of insulin-like growth factor Ⅰ(IGF-Ⅰ) on the apoptosis of endothelial cell.The study investigated the inhibition and mechanism of IGF-Ⅰ on the apoptosis of human umbilical vein endothelial cells(HUVEC) induced by oxidized low density lipoprotein(ox-LDL).METHODS:The experiment was performed in the Institute of Cardiovascular Disease,Union Hospital of Huazhong University of Science and Technology from December 2006 to July 2007.①Fresh human umbilical cord was obtained(the informed consent) to isolate and culture HUVECs.The cells were divided into four groups.Except the control group,HUVEC cells were cultured with IGF-Ⅰ(1?10-9mmol/L),ox-LDL(200 mg/L)+IGF-Ⅰ(1?10-9mmol/L),and ox-LDL(200 mg/L),respectively after cultured for 24 hours.②Cell viability was determined by MTT assay,morphology and apoptosis by DAPI fluorescence staining,and expressions of caspase-3 were analyzed.RESULTS:①Ox-LDL could significantly inhibit HUVEC cell proliferation.After treated with both IGF-Ⅰand ox-LDL,the cell proliferation increased obviously compared with the cells treated with ox-LDL(P

Objective To analyse the relationship between T lymphocyte subsets and NK cells expression and dynamic changes in lung cancer patients 'peripheral blood and the occurrence and development of cancer,and investigate their clinical significances.Methods Flow cytometry was applied to detect 66 patients with lung cancer,60 patients with pulmonary tuberculosis and 60 healthy persons peripheral blood CD+3,CD+3CD+8,CD+3CD+4,Th/Ts,CD+16CD+56 expression.Lung cancer group peripheral blood CD+3,CD+3CD+8,CD+3CD+4,Th/Ts,CD+16CD+56 expression were also detected on 3rd,7th and 20th day before and after chemotherapy.Results Lung cancer group CD+3,CD+3CD+4,Th/Ts,CD+16CD+56 expression decreased significantly [(54.23±10.37)％,(34.23±8.03)％,1.35±0.20,(25.18±4.34)％] and had significant differences compared with pulmonary tuberculosis group [(63.09±9.19)％,(39.46±12.74)％,1.51±0.41,(26.45±3.96)％] and healthy group [(69.68±8.31)％,(42.31±13.29)％,1.89±0.48,(29.44±2.51)％](P ＜ 0.05),but CD+3CD+8 expression showed no significant difference(P ＞ 0.05).In chemotherapy group,comparing with before chemotherapy,remission group CD+3,CD+3CD+4,Th/Ts and CD+16CD+56 expression decreased significantly (P ＜ 0.01)on 3rd day after chemotherapy,while CD+3CD+8expression increased significantly(P ＜ 0.01).On 7th day,each index recovered to the level of before chemotherapy basically.On 20th day,CD+3,CD+3CD+4,Th/Ts and CD+16CD+56 expression increased significantly(P ＜ 0.05)compared with before chemotherapy,while CD+3CD+8 expression significantly decreased(P ＜ 0.05).Chemotherapy unease group had no significant difference (P ＞ 0.05).Lung cancer of stage Ⅲ A and Ⅲ B compared with stage Ⅰ A,and lymph node metastasis in N3 group compared with N0 group,CD+3,CD+3CD+4,CD+3CD+8,Th/Ts and CD+16CD+56 expression had significant differences (P ＜ 0.05).Compared with their pathological types,each index had no significant difference(P ＞ 0.05).Conclusion Monitoring the peripheral blood T lymphocyte subsets and NK cells dynamic of lung cancer patients can guide the clinical diagnosis and treatment,and contribute to the assessment of immune function.