Objectives:To compare the transcriptional activities of tumor-specific hTERT promoters and survivin promoter in human lung cancer A549 cells,in order to lay some groundwork for targeting gene therapy in human lung cancer.Methods:The 1 084 bp fragment of hTERT promoter and 980bp survivin promoter from A549 cells were acquired by PCR amplification.The eukaryotic expression plasmids(pGL-H-RED,pGL-LH-RED and pGL-S-RED)which have Red Fluorescin reporter gen(eRed)and are modulated by 263 bp hTERT promoter,1084bp hTERT promoter and 980bp survivin promoter respectively were constructed based on the pGL3-hTERT which has 263bp hTERT promoter from HepG2 cells.These recombinant plsmids were transfected into A549 and MRC-5 cells with liposome.After 72 hours,the expression level of red fluorescin target protein modulated by 3 promoters at the same time was analyzed by Imagepro-Plus6.0 software.Results:The red fluorescin target protein was observed in transfected A549 cells,but there was no red fluorescence in transfected MRC-5 control cells.The pGL-S-RED had the highest expression activity in A549 cells and the strength rate(IOD)of red fluorescence of A549 cells transfected by pGL-S-RED,pGL-LH-RED and pGL-H-RED were 201.17,171.70,and 136.34 respectively.Conclusion:Our data reveals that the cloned hTERT and survivin promoter are tumor-specific,and the survivin promoter has the highest transcriptional activities in lung cancer A549 cells,and may serve as a useful tool for transcriptional targeting gene therapy of human lung cancer.

OBJECTIVE:To study tanshinone ⅡA inhibiting the proliferation and migration induced by homocysteine(Hcy)of rat aortic vascular smooth muscle cells(VSMCs)and its signal pathway. METHODS:VSMCs were selected for the following ex-periments. In order to validate TanshinoneⅡA inhibiting the proliferation and migration induced by Hcy of VSMCs,VSMCs were di-vided into control group,Hcy group(1 000μmol/L),TanshinoneⅡA low-dose,medium-dose and high-dose groups(5,10,20 μg/L). In mechanism study test,VSMCs were divided into control group,TanshinoneⅡA group(20 μg/L),rapamycin group(20 nmol/L), MHY1485 group(10μmol/L),TanshinoneⅡA+rapamycin group(TanshinoneⅡA,20 μg/L+rapamycin,20 nmol/L),TanshinoneⅡA+MHY1485 group (TanshinoneⅡA,20 μg/L+MHY1485,10 μmol/L). In validation test of P70S6K and p-P70S6K pathway expres-sion,rapamycin and MHY1485 were used for inhibitory test and activation test,respectively. The proliferation of VSMCs was de-termined by ELIASA,and Transwell chambers and wound healing test were employed to test the migratory ability of VSMCs. West-ern blotting were used to investigate the expressions of P21,P27,MMP-2,MMP-9,P70S6K and p-P70S6K in VSMCs. RE-SULTS:In validation test,compared with control group,24,48 h absorbance,migration area,the number of VSMCs penetration and the expression of MMP-2,MMP-9 and p-P70S6K increased significantly,while the expression of P21 and P27 decreased sig-nificantly(P<0.01). Compared with Hcy group,48 h absorbance of TanshinoneⅡA low-dose group,24,48 h absorbance and the expression of MMP-2 of TanshinoneⅡA medium-dose and high-dose groups,migration area,the number of VSMCs penetration, the expression of MMP-9 and p-P70S6K in low-dose,medium-dose and high-dose groups all decreased significantly;the expres-sion of P21 and P27 increased significantly in TanshinoneⅡA medium-dose and high-dose groups(P<0.01);there was no statisti-rapamycin group,while 24,48 h absorbance increased significantly in MHY1485 group(P<0.01). Compared with TanshinoneⅡA group,24,48 h absorbance decreased significantly in TanshinoneⅡA+rapamycin group,while 12,24,48 h absorbance,migration area and the number of VSMCs penetration increased significantly in TanshinoneⅡA+MHY1485 group (P<0.01). In inhibitory test,compared with control group,the expression of p-P70S6K decreased significantly in TanshinoneⅡA group (P<0.01);com-pared with TanshinoneⅡA group,the expression of p-P70S6K decreased significantly in rapamycin group and TanshinoneⅡA+rapa-mycin group(P<0.01);in activation test,the expression of p-P70S6K decreased significantly in TanshinoneⅡA group(P<0.01), compared with TanshinoneⅡA group,the expression of p-P70S6K increased significantly in MHY1485,TanshinoneⅡA+MHY1485 group (P<0.01). CONCLUSIONS:TanshinoneⅡA can inhibit the proliferation and migration of VSMCs by suppressing mTOR/P70S6K signal pathway.

OBJECTIVE:To establish HPLC-ELSD method for content determination of astragaloside Ⅳ in Qizhi jiangtang granules. METHODS:The separation was performed on Hypersil ODS2(250 mm?4.6 mm,5 ?m)column with acetonitrile-water(32 ∶ 68)as mobile phase. The flow rate was 1.0 mL?min-1. Injection volume was 10 ?L. The column temperature was 25 ℃. The temperature of drift tube for measurement was 110 ℃ and flow rate of N2 was 3.12 SLPM. RESULTS:Regression equation was A=1.699 C+6.181. The linear range of astragaloside Ⅳ was 0.259～1.295 mg?mL-1(r=0.999 6,n=5) with lowest detectable limit of 5.18 ?g?mL-1. An average recovery rate were 97.75%(RSD=2.17%). CONCLUSION:The method is simple,rapid and reliable for quality control of Qizhi jiangtang granules.

Objective To determine the effect of proinflammatory cytokines including interleukin-lbeta (IL-1β) and tumor necrosis factor-Alpha (TNF-α) on human serotonin transporter (SERT) in human enterocyte-like cell line Caco-2.Methods Caco2 cells were cultured for 5 days,then were divided into control group.IL-1β treated group (50 ng/ml) and TNF-α treated group (50ng/ml) for 2.24,48 and 72 hours.The expression of SERT mRNA was detected by RT-PCR at 2,24 and 48 hours and its protein expression was measured by Western blotting at 24,48 and 72 hours.Results The expression of SERT mRNA at 2,24 and 48 hours was lower both in IL-1β treated group (1.393±1.184,1.064±0.625 and 1.013±0.415,respectively) and TNF-α treated group (1.000±0.000,0.829±0.162and 0.945±0.147,respectively) in comparison with control group (2.282±1.367,1.586±0.421 and 1.86 ±O.496,respectively).There was significant difference among three groups (P<0.01).The expression of SERT protein at 24,48 and 72 hours was lower in IL-1β treated group and TNF-α treated group than in control group.Conclusions The IL-1β and TNF-α can inhibit the expressions of SERT mRNA and protein in Caco-2 cells,which indicates that IL-1β and TNF-α may change visceral sensitivity by influencing peripheral 5-hydroxytryptamine levels.

Objective To investigate the change of the distribution of peripheral blood lymphocyte subsets,before and after treated by infliximab (IFX) in patients with inflammatory bowel disease (IBD).Methods From September 2008 to January 2013,a total of 20 patients with IBD accepted more than three times of IFX treatment and on time follow-up were collected,11 cases of ulcerative colitis (UC) and nine of Crohn's disease (CD).At same time,20 healthy individuals were enrolled as healthy control group.The efficacy of IFX on patients with UC or CD was evaluated according to Mayo score and simplified Crohn's disease active index (CDAI) before and after treatment.Fasting blood of healthy control group,one day before IFX treatment and in 24 to 72 hours after the third time of IFX injection of IBD patients was collected.The percentage of total T lymphocyte,total B lymphocyte,CD4+ T lymphocyte,CD8+ T lymphocyte and natural killer (NK) cell in lymphocyte was determined by fluorescent labeled monoclonal antibodies and flow cytometry.Independent sample t-test was performed for comparison between two groups.Analysis of variance was for comparison among three groups.Results Three of 11 patients with UC achieved clinical remission,three cases were clinical improved and five cases were ineffectiveness.Among nine CD patients,one achieved clinical remission,six cases were clinical improved and two cases were ineffectiveness.Of UC patients,the percentage of total T lymphocyte before and after treatment ((84.2±8.1) ％ and (82.1±6.2)％),the percentage of CD8+ T lymphocyte before treatment ((40.0± 13.2)％) were all higher than that of healthy control group ((74.7±10.7)％ and (30.5±11.9) ％),while the percentage of NK cell before and after treatment ((5.1±2.8)％ and (7.8±4.3)％) were all lower than that of healthy control group ((13.7 ±7.8)％) and the differences were statistically significant (t=2.540,2.074,2.251,3.464 and 2.063,all P＜0.05).Compared with healthy controls,there were no significant differences in the percentage of total B lymphocyte and CD4+ T lymphocyte of UC patients before and after treatment,the percentage of total T lymphocyte,total B lymphocyte,CD4+ T lymphocyte,CD8+T lymphocyte and NK cell of CD patients before and after treatment (all P＞0.05).Before treatment,there were significant differences among effective treatment group,ineffective treatment group and healthy control group in the percentage of total T lymphocyte,CD8+ T lymphocyte and NK cell (F=4.095,4.571 and 7.432,all P＜0.05),of those there were significant differences between ineffective treatment group ((88.3 ± 8.2) ％,(44.4 ± 11.5) ％ and (4.6 ± 3.2) ％) and healthy control group (t=2.902,2.105 and 3.647,all P＜0.05).Conclusion The percentage of peripheral blood total T lymphocyte and CD8+T lymphocyte increase in active IBD patients with ineffective IFX treatment,however the percentage of NK cell decreases.

Objective To analyze the efficacy and debase the toxicity of radiotherapy combined with composite Kushen injection in treatment of esophageal carcinoma patients. Methods A total of 126 cases of esophageal cancer were treated by radiotherapy were analysed. All patients were treated with the continuative hyperfractionated radiotherapy. The total dose was 66 ~ 74 Gy/5 ~ 7 wk, 2.2 ~ 2.4 Gy/d of esophageal carcinoma patients. Two groups were given respectively delivered with the same radioactive dosages and methods.But the synthetic group was treated by radiotherapy with composite Kushen injection. composite Kushen injection was given at the dosage of 20 ml with 500 ml of saline infusion for 20 consecutive days. Results X-ray grades of esophageal cancer after radiotherapy. The grade I of X-ray manifestation after radiotherapy for patients in synthetic group was superior to that patient in the radiotherapy alone group. (60.4 % vs 41 %; ?2 =4.57, P

Objective To evaluate the clinical features of local advanced glottic laryngeal cancer patients with beyond five years survival and to provide help for the treatment of advanced glottic laryngeal cancer patients.Methods The data of 129 cases of patients with local advanced glottic laryngeal cancer treated in our hospital from January 1999 to August 2005 were analyzed retrospectively.They were divided into two groups based on their survival time.Forty six patients with beyond five years survival were included in the first group and the other eighty three patients were in the second group.The clinical features were compared and factors associated with survival were identified.Results Compared with the group with under five years survival,the lymph node metastatic rate before treatment was lower,patients treated concurrently with operation were more,and complete remission rate after treatment were higher in the group with beyond five years survival.The differences between two groups had statistical significant.Cox regression analysis identified only operation and complete remission rate were independent prognostic factors.Conclusion For local advanced glottic laryngeal cancer,rational comprehensive treatment and complete remission after treatment and complications prevention are important.But they need to be validated by the large-scale clinical trials.

Objective To approach the therapeutic effects and complication of hyperfractionated radio-therapy for nasopharyngeal carcinoma. Methods From January 2000 to November 2002,283 patients with na-sopharyngeal carcinoma were treated with hyperfraetionated radiotherapy. For hyperfractionated radiotherapy,γ-ray from 60Co was performed and two fractions of 1.2 Gy were given daily,with an interval of≥6 h,5 days per week to a median dose of 72 Gy for nasopharynx and 68.4 Gy for neck. For some patients, carboplatin or cispla-tin and 5-fluorouracil were transfused. Results The incidence rate of≥3 grade acute mueositis was 12. 4%.The complete remission rate after treatment was 93.6%. The 1,2 and 5 year survival rate was 93.6% ,82. 7%and 60. 1% respectively. Conclusion patients treated with hyperfractionated radiotherapy have good tolerance and therapeutic effect. It provids a treatment means for nasopharyngeal carcinoma. But it must be validated by the large-scale clinical trials.

Objective To investigate the influencing factors on re-examination compliance of patients with breast cancer Who were in the stage of recovery.Methods By clinic services and telephones,we investigated the reexamination compliance of 189 cases of breast cancer.Results 66.1% of the cases took periodic re-examinations and on the opposite,33.9% of the patients didn't do so.Statistical meaning could be found from the difference of ages,living sites and survival time limits(P＜0.05).Conclusions The main factors to influence the re-examination compliance of patients with breast cancer are age,living site and survival time limit.And it is possible to improve the re-examination compliance of patients by enhancing health education,raising medicsl levels and service qualities,and creating comfortable environment for patients.

Objective To assess the expression of TNFα, IL-1β and the serotonin transporter (SERT) in adult rats of chronic visceral hypersensitivity induced by colon irritation during postnatal development, and to provide evidence to clarify the complex relationship between inflammatory cytokines and SERT among visceral hypersensitivity. Methods Sixteen neonatal male Sprague-Dawley rats were randomly separated into two groups undergoing different treatments. The irritated group ( n = 8 ) was received intracolonic injections of acetic acid(0.5%) between postnatal days 8 and 21 and another group ( n = 8 )was received colonic infusion with 0. 9% saline served as control. When they became adults( ages 7 weeks),the threshold intensity for a visually identifiable contraction of the abdominal wall and body arching were recorded during rectal distention (RD) to evaluate visceral hypersensitivity. Histological evaluation and MPO activity assay were performed to quantify the inflammation. The expression of cytokine of colon was assessed through immunohistochemistry. The expression of SERT was examined by Western blot. Results Histological examination of the tissues showed no significant structural damage or loss of crypts. The MPO levels in both groups were similar[(0. 497 ±0. 570) unit/g vs (0. 623 ±0. 739) unit/g, P =0.724]. The threshold to elicit a distinctive abdominal muscle contraction in response to RD decreased from (0.40 ±0. 14) ml in the control group to (0. 19 ±0.06)ml in the irritated group. And the threshold for bodying arching decreased from (0.91 ± 0. 26 ) ml in the control group to ( 0. 47 ± 0. 13 ) ml in the irritated group (P ＜ 0. 01 ). Cytokine immunoreactivity was increased in the irritated group when compared to the control group (TNFα: 0. 194 ±0. 001 vs 0. 182 ±0. 001, P ＜0. 01; IL-1β: 0. 196 ±0. 002 vs 0. 185 ±0.001, P＜0. 01 ), while SERT expression were reduced in the irritated group (0. 298 ±0. 038 vs 0. 634 ±0. 200, P＜0. 05). Conclusion There is an increase in the proinflammatory cytokines and a decrease in the SERT expression associated with the presence of chronic visceral hypersensitivity, both of them may play an important role in the pathogenesis of visceral hypersensitivity.