IL-2-TNFα融合基因与B7.1基因联合修饰的肿瘤的疫苗作用

目的　改进基因修饰的肿瘤细胞疫苗的效力。方法　在小鼠乳腺癌细胞系EMF6 和一个已被含有IL - 2和TNFα融合基因的逆转录病毒载体XdF转染的EMF6 亚系 (T2 -EMF6)中表达B7 1基因。结果　免疫 /侵袭实验表明 ,与单基因转导的肿瘤细胞相比 ,IL - 2-TNFα/B7 1共同修饰的肿瘤细胞具有较低的致瘤性和较优的疫苗效力 (P <0 0 5)。在各种肿瘤细胞的免疫接种后三周 ,进行混合淋巴细胞培养实验和51 Cr释放实验来检测动物的细胞免疫活性 ,IL - 2 -TNFα和B7 1共同作用诱导了比其中的任意一个单独作用更强的抗肿瘤应答 (比IL - 2 -TNFα高 2 5% ,比B7 1高 2 0 % )。结论　IL - 2 -TNFα和B7 1协同作用可提高他们所修饰的肿瘤细胞的疫苗效力。     

腺病毒介导人IL-2基因治疗的抗肿瘤作用及机理分析

目的　观察重组人白介素 2腺病毒 (advh IL- 2 )在小鼠肝癌 H2 2中的转染效率、表达时程、体内抗肿瘤作用及其免疫机理。方法　小鼠皮下接种肝癌 H2 2 ,将肿瘤生长至体积为 10 0～ 2 0 0 mm3的荷瘤小鼠随机分组 ,经肿瘤局部注射给药。用 X- gal染色法检测重组腺病毒载体 adv L ac Z的转染效率 ;用 RT- PCR检测人 IL- 2基因的持续表达时间 ;以 H2 2瘤体生长及荷瘤小鼠存活时间评价 advh IL - 2抗肿瘤作用 ;以 51 Cr 4小时释放法测定治疗小鼠脾细胞的 L AK与 CTL 杀伤活性 ,免疫荧光法分析肿瘤组织内 CD4 +与 CD8+ T细胞浸润情况。结果　肿瘤局部单次注射 1× 10 9pfu重组腺病毒 ,可在肿瘤组织中进行有效的转染及表达 ,人 IL- 2基因的表达时间长于 12 d。AdvhIL- 2的抗肿瘤作用具有剂量依赖性 ,与 PBS组比较 ,2× 10 9pfu advh IL- 2可非常明显地抑制 H2 2皮下瘤的生长、延长荷瘤小鼠的生存时间 (P<0 .0 1) ,同时明显增强脾细胞 L AK与 CTL杀伤活性 (P<0 .0 1) ,增加肿瘤组织内CD4 + 与 CD8+ T细胞的浸润。结论　 Advh IL - 2通过介导人 IL - 2基因在小鼠 H 2 2肿瘤组织中的持续表达 ,产生明显的体内抗肿瘤作用 ,其机理与激活荷瘤小鼠抗肿瘤免疫反应有关     

肿瘤生物治疗新方法的研究

本文主要简述“九五”攻关期间《肿瘤生物治疗新方法的研究》课题组经协作攻关主要完成的研究工作;（1）重组人TNF、IL－2及B7重组腺病毒表达载体的构建及其包装体系的建立;（2）腺病毒介导的人TNF－α基因转染对人肝财亡和MHC－1类分子表达的影响;（3）瘤体内／瘤周注射TNF－α组腺病毒和（或）IL－2重组腺病毒对肝癌小鼠的治疗作用及其免疫机理研究;（4）粘附LAK细胞的制备、体外扩增、冻存及复苏;（5）转染B7、IL－2、TNF－α基因重组腺病毒的肿瘤细胞及A－LAK的生物学特性研究;（6）人肝癌组织中MAGE基因的表达及MAGE基因修饰树突细胞体外抗肿瘤作用;（7）转基因瘤苗临床应用安全性的初步检测。这些研究工作的完成为肿瘤基因治疗的临床应用打下基础。     

人B7-1转基因肝癌瘤苗的制备

目的:制备人B7-1(hB7-1)转基因肝癌瘤苗.方法:用逆转录病毒转移系统将hB7-1基因导入人肝癌细胞株HepG2,并用RT-PCR、PCR-Southern杂交法检测转染空载体 pLXSN的HepG2/neo细胞和转染重组质粒pLXSN/hB7-1的HepG2/hB7-l细胞中hB7-1 mRNA表达;间接免疫荧光法检测HepG2细胞、HepG2/neo细胞、HepG2/hB7-1细胞形态,计算当日细胞绝对数,绘制生长曲线.结果:所建立的肝癌瘤苗(HepG2/hB7-1)细胞可高效表达hB7-1分子.基因转染对HepG2细胞的生长形态及生长曲线无明显影响.结论:逆转录病毒基因转移系统是使肝癌细胞高效表达hB7-1分子的有效途径.     

pEgr-angiostatin基因辐射诱导表达特性及其抗肿瘤作用

目的 :检测 p Egr- angiostatin重组质粒在 B1 6细胞中的辐射诱导表达和观察 p Egr-angiostatin基因 -放射治疗的抗肿瘤作用。方法 :用 RT- PCR方法从小鼠肝脏中扩增出 angiostatinc DNA,经测序证实后 ,构建 p Egr- angiostatin重组质粒 ,脂质体介导的转染法转染小鼠 B1 6细胞 ,检测 B1 6细胞内 angiostatin m RNA的辐射诱导表达 ,体内观察 p Egr- angiostatin基因 -放射治疗的抑瘤作用。结果 :测序表明扩增的 angiostatin c DNA序列与报道基本一致 ,Egr- 1启动子和 angiostatinc DNA正确插入表达载体 pc DNA3.1 ,转染 B1 6细胞内 angiostatin m RNA的表达具有辐射诱导特性 ,p Egr- angiostatin基因 -放射治疗荷瘤小鼠具有明显的抑瘤作用。结论 :成功地克隆小鼠angiostatin基因 ,并证实 p Egr- angiostatin的辐射诱导表达规律及其基因 -放射治疗的抗肿瘤作用     

B7—1基因转染小鼠EL—4淋巴瘤协同IL—2作为瘤苗诱导的免疫治疗试验

目的　观察B7 1基因转染小鼠EL 4淋巴瘤协同IL 2作为瘤苗诱导的免疫治疗作用。方法　首先以 2×10 5 EL 4Wt接种C5 7BL 6小鼠左前肢腋下 ,然后进行免疫治疗。结果　用B7 1 的EL 4淋巴瘤细胞免疫组小鼠的成瘤时间推迟 ,荷瘤鼠存活期也能延长 ,但经EL 4B 7 1 IL 2免疫治疗的小鼠 ,其瘤结节出现的时间和存活的时间明显延长 (P <0 .0 1) ,且瘤结节大小也有明显差别。结论　B7 1基因转染小鼠EL 4淋巴瘤细胞具有免疫治疗作用 ,B7 1基因转染小鼠EL 4淋巴瘤协同IL 2作为瘤苗诱导的免疫治疗作用更显著。     

大鼠心肌肌凝蛋白a重链基因逆转录病毒载体的构建

目的 :克隆编码大鼠心肌肌凝蛋白α重链 aa736～ 96 0的 c DNA片段 ,构建携带 hu IL - 2 / m yosin融合基因的逆转录病毒载体。方法 :以 RT- PCR方法从幼龄大鼠心肌组织中扩增出 6 81bp的目的基因 ,与 hu IL - 2 c DNA序列拼接 ,构建融合基因 hu IL - 2 / m yosin及其逆转录病毒载体。采用脂质体法转染 NIH 3T3细胞 ,经 G418筛选后 ,用 RT- PCR和免疫组化法分析重组逆转录病毒的表达。 结果 :核苷酸序列测定显示 ,克隆基因的核苷酸及推导的氨基酸序列与报告序列一致 ,开放阅读框正确 ;逆转录病毒载体 p L NC- hu IL - 2 - m yosin酶切结果与预期一致。RT- PCR检测提示转染细胞有 p L NC- hu IL - 2 - myosin m RNA转录 ;免疫组化分析表明转染细胞胞质内有大量肌凝蛋白表达。 结论 :大鼠心肌肌凝蛋白α重链基因逆转录病毒载体构建成功 ,并可在 NIH3T3细胞中表达 ,为心肌肌凝蛋白基因转移诱导机体特异性免疫耐受对心肌免疫损伤的防治研究打下了基础     

共刺激分子B7-2基因转染抑制乳腺癌生长和转移的实验研究

目的:研究B7-2基因转染乳腺癌细胞对人乳腺癌肺转移的影响.方法:设计B7-2内引物,在内引物的5'端引入XbaⅠ、KpnⅠ酶切位点;在基因组B7-2编码基因两端设计外引物.分离人外周血单个核细胞,孵育成簇后提取总RNA,通过RT得到DNA,分别以外引物和内引物PCR扩增出B7-2基因,PCR产物经双酶切装入真核载体pcDNA3.1( )构建成重组质粒B7-2pcDNA3.1,对重组质粒进行扩增.将重组质粒和空载体转染MD-MBA-231细胞,用FITC anti-human B7-2 antibody标记两组转染细胞和原细胞,流式细胞检测B7-2表达.将B7-2基因转染乳腺癌MD-MBA-231细胞,将MD-MBA-231/B7-2、MD-MBA-231/pcDNA3.1和MD-MBA-231分别接种于BALB/c小鼠皮下,观察B7-2基因转染对肺转移的影响.结果:获得B7-2重组基因,重组基因转染组的B7-2表达阳性率比空载体组和原细胞组分别高出36.77%和51.88%,种植MD-MBA-231/B7细胞株的BALB/c小鼠成瘤率及肺转移率显著低于对照组,HE染色发现淋巴细胞广泛浸润于MD-MBA-231/B7种植的肿瘤及其肺转移灶.结论:B7-2基因通过提高免疫原性抑制乳腺癌的生长及肺转移.     

B7-1基因转染小鼠EL-4淋巴瘤细胞体外诱导免疫小鼠脾细胞产生的白细胞介素-2

目的　研究B7 1基因转染小鼠EL 4淋巴瘤体外诱导免疫小鼠脾细胞产生的白细胞介素 2。方法　以CTLL 2细胞检测EL 4B7 1 免疫小鼠的脾细胞与瘤细胞共育的MLTC培养上清中的IL 2活性。结果　EL 4B7 1 免疫小鼠的SPL经瘤细胞在体外刺激可诱生出明显的IL 2活性 ,而EL 4免疫小鼠的SPL则无此作用 ,两者具有较明显的差异 (P <0 0 1)。结论　B7 1基因转染小鼠EL 4淋巴瘤可在体外诱导免疫小鼠脾细胞产生较高水平的白细胞介素 2。     

肿瘤生物治疗新方法的研究

本文主要简述"九五"攻关期间<肿瘤生物治疗新方法的研究>课题组经协作攻关主要完成的研究工作(1)重组人TNF、IL-2及B7重组腺病毒表达载体的构建及其包装体系的建立;(2)腺病毒介导的人TNF-α基因转染对人肝癌细胞凋亡和MHC-1类分子表达的影响;(3)瘤体内/瘤周注射TNF-α重组腺病毒和(或)IL-2重组腺病毒对肝癌小鼠的治疗作用及其免疫机理研究;(4)粘附LAK细胞的制备、体外扩增、冻存及复苏;(5)转染B7、IL-2、TNF-α基因重组腺病毒的肿瘤细胞及A-LAK的生物学特性研究;(6)人肝癌组织中MAGE基因的表达及MAGE基因修饰树突状细胞体外抗肿瘤作用;(7)转基因瘤苗临床应用安全性的初步检测.这些研究工作的完成为肿瘤基因治疗的临床应用打下基础.     

Value of D-Dimers in patients with acute aortic dissection

Objective: To evaluatel the value of D-dimers in patients with acute aortic dissection (AAD). Methods: This study consisted of 16 patients with AAD and 27 non-AAD patients. Serum D-dimets were measured by Sta-Liatest D-DI immunoturbidimetric assay. Results: D-dimer level was higher (P ＜ 0.001) in patients with AAD(7.91 ± 5.52 μg/ml) than that in non- AAD group(1.57±1.24 μg/ml). D-dimer was positive (＞0.4 μg/ml) in all patients with AAD and in 10 control group patients (37%). Among patients with acute AAD, D-dimers tended to be higher in Stanford A than in Stanford B (8.67 ± 4.31 μg/ml vs. 3.24±1.27 μg/ml, P ＜0.01). D-dimer values tended to be higher in more extended disease(3.84 ± 1.65 μg/ml, 8.57 ± 3.58 μg/ml and 11.87 ± 5.69 μg/ml in thoracic aorta, thoracic and abdominal aorta, thoracic and abdominal aorta and iliacal arteries, respectively, P ＜ 0.05 for both 8.57 ± 3.58 and 11.87 ± 5.69 vs. 3.84 ± 1.65 ). Including the control group into the analysis, we found a sensitivity of 100%, a negative predictive value of 100%, and a specificity of 66% and a positive predictive value of 64% for D-dimer in diagnosis of AAD in our patients with suspected AAD. Conclusion: D-dimer was elevated in patients with AAD. A negative D-dimer test result could be useful in excluding AAD.     

Research of combined liver-kidney transplantation model in rats

Objective： To set up a simple and reliable rat model of combined liver-kidney transplantation. Methods： SD rats served as both donors and recipients. 4℃ sodium lactate Ringer＇s was infused from portal veins to donated livers,and from abdominal aorta to donated kidneys, respectively. Anastomosis of the portal vein and the inferior vena cava （IVC） inferior to the right kidney between the graft and the recipient was performed by a double cuff method, then the superior hepatic vena cava with suture. A patch of donated renal artery was anastomosed to the recipient abdominal aorta. The urethra and bile duct were reconstructed with a simple inside bracket. Results： Among 65 cases of combined liver-kidney transplantation, the success rate in the late 40 cases was 77.5%. The function of the grafted liver and kidney remained normal. Conclusion： This rat model of combined liver-kidney transplantation can be established in common laboratory conditions with high success rate and meet the needs of renal transplantation experiment.     

APPLICATION OF LORNOXICAM TO PATIENT-CONTROLLED ANALGESIA IN PATIENTS UNDERGOING ABDOMINAL SURGERIES

FOR anesthesiologis s ,treatingpostoperativepainhas alwaysbeen a problem.Althoughopioidshave been provedtobe effective,theirsideeffectscouldnotbeignored.With thedevelopmentofscienceand pharmacology,many drugs with aspectsof satisfactoryanalgesicefficacyand couldbe welltoleratedby patientshave been developed.And lornoxicamisone of them, which isa non-steroidalanti-inflammatorydrug (NSAID ), with analgesic, anti-infl-ammatory,andantipyreticproperties.Itseliminationhalf-time(3 to 5 hours) isle…     

BLOOD PRESSURE CHANGE WITH AGE IN SALT-SENSITIVE TEENAGERS

Objective To observe blood pressure change with age in salt-sensitive teenagers whose salt sensitivity were determined by repeated testing.Methods Salt sensitivity was determined through intravenous infusion of normal saline combined with volume-depletion by oral diuretic furosemide in 55 teenagers. After five years, salt sensitivity was re-examined and subject blood pressure was followed up. Blood pressure changes in salt-sensitive teenagers were compared to that of non-salt sensitive teenagers over five years.Results After 5 years, the repetition rate of salt sensitivity determined by intravenous saline loading is 92.7%. In teenagers with salt sensitivity on the baseline, both the systolic blood pressure increments and increment rates were much higher than non-salt sensitive teenagers (12.7±12.1 mmHg vs. 2.8±5.2 mmHg, P＜ 0.01; 12.2%± 12.0% vs. 2.5% ±4.4%, P＜ 0.001,respectively). There was a similar trend for diastolic blood pressure (8.4 ± 6.4 mmHg vs. 3.7 ± 6.4 mmHg, P = 0.052; 13.2% ±10.6 % vs. 6.8%± 10.1%, P = 0.053, respectively).Conclusions Salt sensitivity determined by intravenous saline loading showed good reproducibility. Blood pressure increments with age were much higher in salt-sensitive teenagers than non-salt sensitive teenagers, especially in terms of systolic blood pressure.     

Acupuncture Combined with Spinal Tui Na for Treatment of Primary Dysmenorrhea in 30 Cases

Objective: To observe the therapeutic effects in acupunture treatment of primary dysmenorrhea combined with spinal Tui Na, and study its mechanism. Methods: Thirty cases of the treatment group were treated by acupuncture combined with spinal Tui Na, and thirty cases in the control group were treated by routine acupuncture. Results: The total effective rate was 93.3% in the treatment group, and 73.3% in the control group, with a significant difference between the two groups (P<0.05). Conclusions: Acupuncture combined with spinal Tui Na has good prospects for treatment of primary dysmenorrhea.     

Luo Lingjie's Experience in Treating Chronic Nephropathy

In treating chronic nephropathy,Luo Lingjie,a chief physician,pays attention to regulating the balance between yin and yang,treating infection if present,and removing pathogenic factors.He prescribes gentle drugs and uses carefully strongly warming-tonifying ones,emphasizes the importance of persuading the patient to persist in treatment with medication and nurse one's health for recuperation,and is good at combined use of TCM and western medicine therapy and brings the merits of various therapies into full play,with obvious theraoeutic effects.     

Dr.Zhang Ren's Experience in the Acupuncture Treatment of Different Diseases with the Same Therapeutic Principle

Dr.Zhang Ren,the chief physician,is the chairman of Shanghai Acupuncture and Moxibustion Association.Having been engaged in medicine for about 40 years,he is experienced in treating various intractable diseases.In his long years of clinical practice,he advocates taking the TCM differentiation as the basis to seek for the acupuncture method for treatment of modern intractable diseases.The author of this essay had the fortune to follow Dr.Zhang in study.The following is a summary of Dr.Zhang's experience in the acupuncture treatment for different intractable diseases with the same therapeutic principle.     

安心颗粒对急诊经皮冠状动脉介入术后生活质量影响的研究

目的：评价使用安心颗粒对急诊经皮冠状动脉介入术（PPCI）术后生活质量的影响.方法：将160例接受PPCI的急性ST段抬高型心肌梗死患者随机分为安心颗粒组（术前顿服安心颗粒8.8g,术后安心颗粒4.4 g/次,每日2次）和对照组（仅接受基础药物治疗）.所有患者均服用阿司匹林、氯吡格雷和阿托伐他汀.分别在入院时、出院前1d、出院后180 d时,应用心肌梗死多维度量表（MIDAS）、中文版SF-36评价量表对患者生活质量评分.并观察术后30 d以内的出血并发症、血小板减少症发生情况.结果：入院时和出院前1d,两组患者的心肌梗死MIDAS、SF-36量表评分比较无差异（P＞0.05）;出院后180 d时,与对照组比较,安心颗粒组MIDAS、SF-36评分明显减低（P＜0.05）;组内与入院时比较,两组出院前1d、出院后180 d时,MIDAS、SF-36评分均降低（P＜0.05）.两组患者在随访期间均无大量出血、少量出血、重度和极重度血小板减少症发生,安心颗粒组有4例、对照组有7例发生不明显出血（P＞0.05）.两组发生轻度血小板减少症的患者数比较无差异（P＞0.05）.结论：PPCI使用安心颗粒,能改善急性ST段抬高型心肌梗死患者的生活质量,且不增加出血风险.     

The comparative studies of the influences of Urapidil and Nicardipine on sino-atrial node function, atrio-ventricular node function and hemodynamics

Objective:To investigate the influences of urapidil and nicardipine on rabbit sinus function,atrio-ventricular node function and hemodynamics.Methods:Thirty-two Angora's rabbits were selected and randomly divided into four groups.U1 group:urapidil 0.25 mg/kg;U2 group:urapidil 0.5 mg/kg;N1 group:nicardipine 10 μg/kg;N2 group:nicardipine 20 μg/kg.All these medicine were administrated within 30 seconds.Measurements were taken before and after the administration of urapidil or nicardipine for the following data:mean blood pressure(MAP),heart rate(HR),sino-atrial conduction time(SACT),maximal sinoatrial recovery time(SNRTmax)corrected sinus node recovery time(CSNRT),index of sinus node recovery time(SNRTI),Wenckebach A-V conduction frequency (WB),and P-R interval.Results:Significant MAP and HR changes were identified in all of the four groups before and after administration of both urapidil and nicardipine.No significant changes could be found in the rest of the parameters.Intergroup analysis showed that SACT and CSNRT of N1 and N2 groups were shorter than those of the U2 group(P＜0.01);the MAP decreased(P＜0.01)and the HR increased drastically(P＜0.01).Conclusions:Neither urapidil(0.25 mg/kg,0.5 mg/kg)nor nicardipine(10μg/kg,20μg/kg)has any significant influence on rabbit sinus function or rabbit atrio-ventricular node function.Nicardipine could be a better choice than urapidil for parafunctional sinus node patients.