1.Construction of Smac siRNA lentiviral vector and its silencing effect on Smac gene in human LECs
Yun, LI ; Guangying, ZHENG ; Yu, JIANG
Chinese Journal of Experimental Ophthalmology 2016;34(3):199-204
Background Research comfirmed that second mitochondrial activator of caspase (Smac) is a promoting tumor cell apoptosis protein.Our previous study showed that the expression level of Smac in LECs is obviously higher in cataract than that in normal eyes.We assumed that silencing Smac gene in LECs can inhibit the apoptosis of LECs.The way to transfect Smac siRNA into LECs is a key step.Objective This study was to construct siRNA lentiviral vector of Smac and identify its silencing efficiency in human lens epithelial B3 cell line (HLE-B3) and establish low-expressed Smac HLE-B3 line.Methods Based on the genebank and our previous study,siRNA sequence of Smac was designed and composed.The synthetic double-stranded DNA was linked to the lentiviral vector GVll8 by T4 DNA ligase and then transformed DH5α competent cells.The plasmids were transformed into the DH5α competent cells.Recombinant colonies were screened by PCR and sequenced.Recombinant plasmids and two other auxiliary plasmids were used to infect 293T cells.Cell culture supernatant was collected for the measurement of viral titer.Recombinant lentiviral vector was used to infect HLE-B3 cells to calculate the viral multiplicity of infection (MOI) under the fluorescence microscope.Transfection efficiency was examined by calculating the GFP-positive cells.HLE-B3 cells were divided into negative control group,siRNA plasmid tranfected group and GV118-Smac-siRNA1 tranfected group.The relative expression levels of Smac mRNA in the cells were detected and compared among the three groups by real-time fluorescent quantitative PCR.Results GV118-Smac-siRNA was successfully constructed with the positive colonies 340 bp and blank vector colonies 299 bp,and viral titer was 3.0× 108 TU/ml.At a MOI of 100,the infecting efficiency of the vector on HLE-B3 cells was about 82% and the cytotoxicity was low.The relative expression levels of Smac mRNA were (101.290±8.349)%,(92.330±6.320)% and (32.540±4.221)% in the negative control group,siRNA plasmid tranfected group and GV118-Smac-siRNA1 tranfected group,respectively,showing a significant difference among the three groups(F =32.871,P<0.01),and the relative expression level of Smac mRNA was significantly lower in the GV118-Smac-siRNA1 tranfected group than that in the negative control group (P =0.000).However,no significant difference was found in the Smac mRNA expression between the blank plasmid group and the negative control group (P=0.535).Conclusions GV118-Smac-siRNA lentiviral vector is successfully constructed.Smac-siRNA can effectively inhibit the expression of Smac mRNA in human LECs.
2.Prediction of GTV/LV for radiation pneumonitis in patients with stage Ⅲ non-small cell lung cancer after radical radiotherapy
Hongmei LIN ; Anhui SHI ; Xin SUI ; Rong YU ; Huiming YU ; Huimin MA ; Guangying ZHU
Chinese Journal of Radiological Medicine and Protection 2016;36(4):272-277
Objective To analyze relevant clinical and dosimetric factors associated with radiation pneumonitis in patients with stage Ⅲ non-small cell lung cancer after they received radical radiotherapy.Methods A total of 126 patients with stage Ⅲ non-small cell lung cancer who received precision radiotherapy in Peking University Cancer Hospital were analyzed from January 2013 to December 2014.Data were collected including various clinical factors (including sex,age,histological type,tumor location,history of diabetes,history of hypertension,history of smoking,the season patients received treatment,ECOG performance status before treatment,chemotherapy before radiotherapy,concurrent chemotherapy and the classification of radiation pneumonitis),as well as related dosimetric parameters [including GTV,lung volume (LV),bilateral V5,V10,V20,V30 and MLD].SPSS 19.0 software was used to analyze the relation between correlation factors and radiation pneumonitis (RP≥2).Results Among the patients,31 cases (24.6%) had occurrance of radiation pneumonitis ≥ 2.Univariate analysis showed that age,ECOG performance status before treatment,concurrent chemotherapy and GTV/LV ratio were significantly correlated with RP ≥ 2 (R =0.157-0.222,P < 0.05).Further multivariate Logistic regression showed that age,concurrent chemotherapy and GTV/LV ratio were significantly correlated (Wald =4.754,6.422,14.79,P < 0.05).Conclusions In patients with stage Ⅲ non-small cell lung cancer after receiving thoracic radical radiotherapy,increasing age and GTV/LV ratio≥3.2% are risk factors of RP≥2.The concurrent chemotherapy with low-dose paclitaxel might also increase the risk of RP≥2.
3.Patients' satisfaction on catastrophic medical insurance of New Rural Cooperative Medical Scheme:Analysis of 484 patients in three districts of B city
Chunxia NA ; Guangying GAO ; Chengyu MA ; Ting DUAN ; Qianhui MA ; Jirong JIA ; Jinzhi YU
Chinese Journal of Health Policy 2015;8(3):34-39
Objective:To understand the patients'satisfaction on catastrophic medical insurance of New Rural Cooperative Medical Scheme ( NRCMS ) , and provide references for effective management and policy improvement. Methods:According to the principles of representative sampling, a questionnaire survey conducted. 484 patients got compensation for catastrophic insurance in 2013 were surveyed, and information of medical costs was analyzed. Re-sults:The average medical cost accounts for 84. 57% of total household expenditure. Mandatory medical expenditure is still high and economic burden of diseases is heavy. This policy has increased the compensation rate, and the actu-al compensation rate increases 4. 79% in three districts. The average score of patients' attitude toward catastrophic medical insurance is (3.95±1.05) points, 88. 43% of patients continue to participate in medical insurance. Con-clusions and suggestions:Patients' satisfaction on catastrophic medical insurance of NRCMS is high. It is suggested that the existing financing modes should maintain stability in the short term, and the compensatory rate should be im-proved reasonably.
4.Dosimetric comparison between RapidArc and fixed gantry dynamic IMRT for central-type lung cancer radiotherapy
Jian GONG ; Rong YU ; Hao WU ; Shukui HAN ; Bo XU ; Guangying ZHU ; Fan JIANG
Chinese Journal of Radiological Medicine and Protection 2010;30(4):448-451
Objective To compare the dosimetric difference between RapidArc and fixed gantry angle dynamic IMRT (dIMRT) for central-type lung cancer radiotherapy. Methods Therapy for 10 patients previously treated with dIMRT was replanned with RapidArc. Dose prescription was 66 Gy/33 fraction. Comparative endpoints were planning target volume (PTV) dose, doses to surrounding structures,number of monitor units, and treatment delivery time. Results There was no significant dosimetric difference between RapidArc and dIMRT. Compared with dIMRT, RapidArc slightly elevated target volume dose, lung V5, V10. The average values of lung V20, V30 and heart V30 were larger in dIMRT than those in RapidArc. The number of monitor units was reduced by 32% and the treatment time by 66% in RapidArc.Conclusions Both RapidArc and dIMRT plans could meet the clinical therapy needs. RapidArc could achieve similar target coverage and sparing of organs at risk, with fewer monitor units and shorter delivery time than dIMRT.
5.Consensus and controversies on delineation of radiotherapy target volume for a patient with locally advanced non-small cell lung cancer
Dan ZHAO ; Xiaolong FU ; Lyuhua WANG ; Baolin QU ; Baosheng LI ; Lujun ZHAO ; Xiangying XU ; Jianhua WANG ; Yaqin QU ; Shuchai ZHU ; Zhilong YU ; Guang LI ; Hong YU ; Yongjing YANG ; Jie LI ; Bo XU ; Weibo YIN ; Guangying ZHU
Chinese Journal of Radiation Oncology 2017;26(9):985-991
Objective To investigate the consensus and controversies on the delineation of radiotherapy target volume for patients with locally advanced non-small cell lung cancer (LA-NSCLC).Methods Questionnaires including 15 questions on the delineation of radiotherapy target volume of NSCLC were sent to 12 radiation departments in China in November 2015.A patient with LA-NSCLC was selected by Fudan University Shanghai Cancer Center, and simulation CT images and medical history data were sent to the 12 radiation departments.Twelve radiation oncologists from the 12 radiation departments showed and explained the delineation of radiotherapy target volume of their own, and the patient was discussed by all experts in the sixth multidisciplinary summit forum of precise radiotherapy and chemotherapy for tumor and lung cancer.Results All receivers of the questionnaire answered the questions.The standard lung window width/level for the delineation of lung cancer was 800-1600/-600 to-750 HU, and the mediastinum window was 350-400/20-40 HU.Respiratory movement was measured by stimulator, 4D-CT, and stimulator+4D-CT with 2-5 mm expansion based on experience.The primary clinical target volume (CTV) was defined as gross target volume (GTV) plus 5-6 mm for squamous carcinoma/5-8 mm for adenocarcinoma.The metastatic lesion of mediastinal lymph nodes was delineated as 5 mm plus primary lesion in 6 departments and as primary lesion in another 6 departments.Of the 12 departments, 10 applied 5 mm of set-up error, 1 applied 3 mm, and 1 applied 4-6 mm.For V20 of the lungs, 10 departments defined it as<30%, 1 as<35%, and 1 as 28%.Nine departments defined the radiation dose of concurrent chemoradiotherapy (CCRT) for LA-NSCLC as 60 Gy in 30 fractions, 62.7 Gy in 33 fractions in 1 department, 50-60 Gy in 25-30 fractions in 1 department, and 60-70 Gy in 25-30 fractions in 1 department.For the delineation of target volume for the LA-NSCLC patient treated with CCRT, the primary planning target volume (PTV) was defined as GTV plus organ movement (IGTV) and set-up error (GTV→IGTV→PTV) in 3 departments, as CTV plus organ movement (ITV) and set-up error (GTV→CTV→ITV→PTV) in 8 departments, and as CTV plus set-up error/IGTV plus 5-6 mm for squamous carcinoma/5-8 mm for adenocarcinoma (CTV) and set-up error (GTV→CTV→PTV/GTV→IGTV→CTV→PTV) in 1 department.For the delineation of PTV in the mediastinal lymph node, GTV→IGTV→PTV was performed in 3 departments, GTV→CTV→ITV→PTV in 8 departments, and GTV→CTV→PTV in 1 department.For 10%-100% patients with LA-NSCLC, the radiation field needed to be replanned when 38-50 Gy was completed.There was no unified standard for the optimal standardized uptake value (SUV) of positron emission tomography (PET)-computed tomography (CT) simulation and delineation.Seven departments had applied magnetic resonance imaging (MRI) simulation and 10 departments had applied stereotactic body radiation therapy (SBRT) for the treatment of early-stage NSCLC.For the delineation of PTV for early-stage NSCLC (T1-2N0M0), GTV→IGTV→PTV was performed in 5 departments, IGTV→PTV in 3 departments, and GTV→CTV→ITV→PTV in 2 departments.In all the 12 departments, peripheral early-stage NSCLC was given 6.0-12.5 Gy/fraction, 3-12 fractions and central early-stage NSCLC was given 4.6-10.0 Gy/fraction, 5-10 fractions.The results of discussion on the delineation of target volume for the patient were as follows:respiratory movements should be measured by 4D-CT or simulator;the lung window width/level is 1600/-600 HU and the mediastinal window width/level is 400/20 HU;the primary controversy is whether the involved-field irradiation or elective nodal irradiation should be used for the delineation of CTVnd in the mediastinal lymph node.Conclusions Basic consensus is reached for the delineation of target volume in LANSCLC in these aspects:lung window width/level, respiratory movements and set-up error, primary lesion delineation, the radiation dose in CCRT, and the optimal time for replanning the radiation field.There are controversies on the optimal SUV in the delineation of target volume based on PET-CT simulation, the optimal dose fractionation in SBRT for early-stage NSCLC, and the delineation of CTVnd.
6. A preliminary study on the reform of clinical skill curriculum based on the phased examination of TCM medical practitioners qualification
Juan WU ; Wenjun SU ; Jianqiang QI ; Ying LOU ; Yun LIU ; Guangying YU
Chinese Journal of Medical Education Research 2019;18(9):893-896
The 2013 National Practitioners Qualification Examination Work Conference proposed to reform the traditional one-stop test into two stage. In May 2017, Fujian University of Traditional Chinese Medicine participated in this empirical study as a pilot for the first time. This paper made a preliminary summary and analysis of the results of the phased examination. Because of the absence and deficiency of clinical skills of students in our school, we carried out a series of reform measures for clinical skill courses: enhancing teaching facilities, enriching teaching contents and methods, reforming assessment mode and strengthening the monitoring of the teaching quality of clinical skills. According to the students, reform measures are conducive to consolidating theoretical knowledge and improving clinical skills, laying a good foundation for successfully passing the national examination of medical practitioners and becoming a qualified medical talent.