Objective:To construct Ewing's sarcoma EWS-FLI1 gene prokaryotic expression vector.Methods:The target gene of EWS-FLI1 was obtained by RT-PCR method after the total RNA was extracted from Ewing's sarcoma A673 cells.The site sequences of restrictive endonuclease SacⅠand Hind Ⅲ were introduced into the upstream and downstream of target gene respectively.The target gene fragment were cloned into pMD18-T and transformed into E.Coli JM109.Screened positive clones were confirmed by PCR,restrictive endonuclease digestion and DNA sequencing.The EWS-FLI1 gene was sequentially subcloned into prokaryotic expression vector pQE30,and the recombinant plasmid pQE30-EWS-FLI1 was confirmed by restrictive endonuclease digestion and DNA sequencing.The proteins,expressed in E.coli JM109 transformed with EWS-FLI1recombinant plasmid under IPTG induction,were characterized by SDS-PAGE and Western-blot.Results:PCR result indicated that an amplified DNA fragment was in size of 1.5 kb.Restrictive endonuclease digestion analysis indicated that the target gene was in size of 1.5 kb.DNA sequencing analysis demonstrated that sequence of target gene accorded with anticipated one.The EWS-FLI1 with a molecular weight of 54 kD was highly expressed in pQE30-EWS-FLI1.Western blot proved that the expressed product had the antigenicity of EWS-FLI1.Conclusion:The recombinant prokaryotic expression vector pQE30EWS-FLI1 is constructed successfully,which will contribute to the further research of EWS-FLI1.

A human diploid fibroblast cell line derived from normal abdominal skin of a 17-year-old female patient with ovary carcinoma was established and designated as HSF 79-2. The cultivation of skin fragments was employed in the primary explantation. The medium used was McCoy 5A supplemented with 15～25% calf serum. The grown fibroblasts were succesively subcultured at 1 to 2 split and stopped at the 45th to 54th passage.Having reached confluence the cells still had the ability to divide during 16 months observation if the medium was changed periodically. They displayed the property of dense growth and forming heavy, multilayer sheets, which became a visible membrane to the naked eyes. Under histological examination the membrane had the appearance of connective tissue. Part of the cells were subcultured again after 3,6, and 12 months maintenance in the same culture flask. Their growth character, ploidy, and the generation time were similar to that of the cells passaged normally.The characteristics of this cell line mentioned above appear to be shared by other diploid fibroblasts. It might be used for preserving cell lines and as a model for studying cell motility and differentiation.

Objective To evaluate the value of helical CT scan in the diagnosis of small renal cell carcinoma.Methods The helical CT features of small renal cell carcinoma proved by operation and pathology in 14 cases were studied retrospectively.Results CT showedisodense mass in 7 cases,slightly low density mass in 5 cases,slightly hyperdense mass in 1 case,cystic and solid mass in 1 case,and among them 2 cases showed punctural calcification.The lesions showed marked enhancement in cortical phase in 12 cases,slight to moderateenhancement in 2 cases,in parenchymal phase,the tumor density dropped promptly,and in the pelvic phase the tumor density droppedfurther. Conclusion Small renal cell carcinoma is of certain characteristic on multi-phasic spiral CT contrast-enhanced scan,it is a very valuable method in diangosis of small renal cell carcinoma.

Objective To examine the difference in myocardial protection provided by continuous and intermittent warm blood cardioplegia during coronary artery bypass (CAB) .Methods Thirty ASA Ⅰ-Ⅱ patients undergoing CAB with warm CPB were randomly divided into two groups : (A) continuous warm blood cardioplegia ( n = 15) and (B) intermittent warm blood cardioplegia ( n = 15) . During CPB the body temperature was maintained at 33℃ -34℃ . Arterial blood samples were taken before skin incision (T0) , 1 h after going on CPB (T1 ) and 6h , 24h after coming off CPB (T2, T3 ), for determination of plasma concentration of cardiac troponin T (cTnT) using ELISA method. A small piece of myocardium was obtained from right ventricle (about 1g ) before aortic crass-clamping and after the aortic clamp was removed for determination of myocardial ATP content and ultrastractural examination. Results The demographic data were comparable between the two groups. Plasma cTnT level increased significantly at T1 and T2 as compared with the baseline values (T0) and then returned to normal level at T3 in both groups. The cTnT level was significantly higher in group B than that in group A at T2 (6h after weaning from CPB)The myocardial ATP content decreased significantly after aortic clamp was removed as compared with that before cross-clamping of aorta, but myocardial ATP content in group A was significantly higher than that in group B after release of arotic clamp. Mitochondria score was significantly higher after release of aortic cross-clamp than that before aortic cross-clamping.Conclusion Continuous warm blood cardiaplegia is superior to intermittent warm blood cardioplegia during CPB in terms of myocardial protection.

Objective To study the impact of the resolution of reference dose distribution and evaluated dose distribution to Gamma rate in the verification of IMRT. Methods Designed four fields, the resolution of dose distribution calculated in water phantom and exported from TPS are 1 ,2,3 ,4,5,6 mm. To calculate Gamma index by IBA's OmniPro-I'mRT software in different sampling resolution. Results When the resolution of evaluated dose distribution was fixed, the change of reference dose distribution's resolution has little effect on Gamma rate (5％ to IMRT field) ;When the resolution of reference dose distribution was fixed , the Gamma rate increased as the evaluated dose distribution's resolution ( ＞ 1 mm) raise ( the Gamma rate increase ( 15. 2 ±6. 2) ％ ( t = 11. 99 ,P < 0. 01 ) and ( 14. 9 ± 5. 5) ％ ( t = 13. 24 , P ＜ 0. 01 ) while the resolution of evaluated dose distribution changed from 6 mm t0 3 mm and from 3 mm to 1 mm　respectively) .Conclusions To use Camma method for verification of IMRT, the measured data can be as a reference dose distribution without　interpolation;the computed data by TPS can be as a evaluated dose distribution and it is more appropriate for the resolution of 1 mm.

Human T-cell leukemia virus type 1 (HTLV-1) is a retrovirus demonstrated to be associated with human disease. Infection by the HTLV-1 can cause T-cell leukemia (ATL) in adults. HTLV-1 bZIP factor (HBZ) is a viral protein encoded by the minus strand of the HTLV-1 provirus. Among the regulatory and accessory genes of HTLV-1, HBZ is the only gene that remains intact and which is expressed consistently in all patients with ATL. Moreover, HBZ has a critical role in the leukemogenesis of ATL. Here, we review the function of HBZ in the oncogenesis of HTLV-1 and its molecular mechanism of action.
Animals ; Basic-Leucine Zipper Transcription Factors ; genetics ; metabolism ; Carcinogenesis ; HTLV-I Infections ; pathology ; virology ; Human T-lymphotropic virus 1 ; genetics ; metabolism ; Humans ; Leukemia, T-Cell ; pathology ; virology ; Retroviridae Proteins ; genetics ; metabolism

Objective:To inspect the dosimetry characteristic of a 2-D ionization chamber array and its response to oblique incident electron beam.And to discuss feasibility to using it to validate the electron arc therapy plan.Materials and methods:(1)The 2-D ionization chamber array was calibrated referring to a themble chamber,comparing the response to electron beams with incidence angle ranged from-50°to 50°.(2)Phantom test plans of 6 and 10MeV electron beam arc therapy were designed,with the beam rotation arcs of 30°,60°and 90° respectively,symmetric to the Central axis of 0°gantry angle.Dose distribution of each plan was verifled with the 2-D array.Results:(1)Measurement differences between the central detector of the array and the thimble chamber were less than 2%,under the planned oblique inddent 6/10MeV electron beam.(2)For the all test plans,dose error was less than 3%at dmax in the symmetric axis of the electron beam arcs.Off-axis dose error was less than 2%within the central 70%area of the profile in non-rotating direction,and smaller than 1.5%in the rotational direction.The measured dose distribution had good agreement with the TPS calculation in isodose curves between 100%and 20%.The gammapass rates(△D=5%,△d=5cm)were 99.98%、99.89%、99.74%、98.64%、99.16%and 99.44%respectively for 6 and 10 MeV plans with electron beam arcs of 30°,60°and 90°.Conclusion:it is practicable using the tested 2-D ionization chamber array to verify and validate the electron beam arc therapy plan.

Objective To analyze the risk factors of incision infection and spinal canal hematoma after lumbar spinal posterior surgery.Methods Data of 33 patients with incision infection and 25 patients with spinal canal hematoma after posterior surgery for lumbar spinal disease from January 2010 to December 2014 were retrospectively analyzed.For the patients with incision infection,20 of them were males and 13 were females,with an average age of 58.85±8.76 years (range 38-79 years).Of these patients,5 were diagnosed with lumbar disc herniation,9 with lumbar spondylolisthesis,15 with lumbar spinal stenosis and 4 with lumbar vertebral fracture.For patients with spinal canal hematoma,17 of them were males and 8 were females,with a mean age of 60.96±11.37 (range,38-77).The diagnoses of them were lumbar disc herniation in 18 patients,spondylolisthesis in 3 patients and spinal stenosis in 4 patients.From the same period database,patients who underwent lumbar posterior lumbar surgery with no postoperative complications were selected by 3:1 ratio as the control group according to age,gender and diagnosis.Results For patients with incision infection,it was found that diabetes mellitus,preoperative RBC,hemoglobin,total protein,albumin,serum calcium,operation time,number of segment fusion,intraoperative blood loss,postoperative WBC,RBC,hemoglobin and platelet were significantly different from those with non-infection group.Moreover,multivariate logistic analysis showed that diabetes mellitus (OR=3.716,P=0.032),preoperative serum calcium (OR＜ 0.001,P=0.001),intraoperative blood loss (OR=1.002,P=0.014),postoperative hemoglobin (OR=0.923,P=0.018) and postoperative platelet (OR=1.007,P=0.017) were independent risk factors for postoperative incision infection.For patients with spinal canal hematoma,it was found that patients' preoperative total protein,albumin,serum calcium,platelet,operation time,intraoperative blood loss and postoperative total protein were significantly different from non-hematoma group.Multivariate logistic analysis showed that preoperative serum calcium (OR＜ 0.001,P=0.001),preoperative total protein (OR=1.298,P=0.043),intraoperative blood loss (OR=1.003,P=0.021) and postoperative total protein (OR=1.080,P=0.028) were independent risk factors for postoperative spinal canal hematoma.Conclusion The preoperative diabetes mellitus,serum calcium,intraoperative blood loss,postoperative hemoglobin and platelet were important risk factors for lumbar incision infection.And preoperative serum calcium,total protein,intraoperative blood loss and postoperative total protein were the risk factors for spinal canal hematoma.

Objective To investigate the clinical efficacy and safety of testosterone supplement and Mreceptor blockers in the treatment of patients with lower urinary tract symptoms(LUTS)in late -onset hypogonadism(LOH). Methods 28 cases diagnosed as LOH with mild to moderate LUTS were collected.They were given testosterone supplementation (oral testosterone undecanoate capsules,80mg,2 times/d)and M receptor blocker (oral succinate solifenacin tablet,5mg,1 time /d)treatment for 1 -3 months.After treatment for 1 month and 3 months respectively, reviewd PSA,serum total testosterone (TT),international prostate of urinary storage symptoms score (urinary storage IPSS),quality of life (QOL)score,international index of erectile function (IIEF -5)score,maximum urinary flow rate (Qmax),residual urine (Ru)and rectal examination (DRE).The improvement of LUTS before and after treatment was evaluated.Results 1 month after treatment,1 patient was difficult to tolerate the solifenacin side effects and took testosterone supplementation alone.The rest 27 patients were able to take medicine for 3 months.After 1 month and 3 months treatment,the IPSS score had significant differences compared with before treatment[(10.3 ±2.1)points vs (14.2 ±3.3)points and (9.42 ±1.8)points vs (14.2 ±3.3)points,t =13.67,14.72,all P <0.05 ].After 3 months treatment,the QOL and IIEF -5 scores were (2.1 ±0.7)points vs (4.3 ±0.6)points and (16.8 ± 3.6)points vs (11.9 ±2.5)points,Qmax was (12.5 ±5.6)mL/s vs (9.8 ±4.8)mL/s(t =6.42,5.64,14.92,all P <0.05 ),the difference was statistically significant compared with before treatment.There were no significant changes in PSA and RU before and after treatment.All patients had no severe complications such as acute urinary retention.Conclusion Combination of testosterone and testosterone in the treatment of LUTS patients in LOH is safe and effective,and can significantly improve the LUTS and quality of life.

Objective High expression of the valosin-containing protein ( VCP) gene can enhance the metastasis of osteosar-coma via the AKT/PI3K/NF-KappaB/MMP-9 signaling pathway, but the molecular mechanisms underlying the up-regulation of VCP in osteosarcoma cells remains unknown .This study aimed to determine whether miRNA-129-5p can regulate the VCP expression and its targets in human osteosarcoma cells . Methods The microRNA target-predicting software TargetScanhuman 6.2 ( http://www.tar-getscan.org/) was used to predict the possible targets of miRNA-129-5p on the VCP gene.Then, two recombinant gene report vectors containing the wild VCP gene 3′UTR ( psi-VCP vector ) and mutant VCP gene 3′UTR ( psi-VCPmut vector ) were constructed , se-quenced, and identified.The human osteosarcoma U2-OS cells were co-transfected with miRNA-129-5p mimic and psi-VCP vector or psi-VCPmut vector, respectively.A non-specificity mimic transfection served as negative control , and the luciferase activity was detec-ted in each group. Results The software prediction showed only one conserved function site of miRNA-129-5p on the VCP gene 3′UTR163-169 bp.Luciferase activity was significantly lower in the psi-VCP vector +miRNA-129-5p transfection group (15.529 ± 1.902) than in the VCP control group (21.781 ±0.854), VCP mutation experimental group (19.978 ±1.377), and VCP mutation control group (21.952 ±1.516) (P<0.05), with no remarkable difference between the VCP mutation control and VCP control groups (P=0.276). Conclusion miRNA-129-5p can probably regulate the targets of the VCP gene in human osteosarcoma U 2-OS cells.