抗原负载的树突细胞对细胞因子诱导的杀伤细胞的影响

目的：研究树突细胞(DC)负载不同形式的抗原对其成熟和功能的影响,及DC对细胞因子诱导的杀伤细胞(CIK)增殖和功能的影响。方法：外周血单个核细胞(PBMC)经不同细胞因子作用后培养成DC和CIK细胞,将Bel 7402细胞制备成肿瘤细胞冻融物、坏死肿瘤细胞及正常生长的肿瘤细胞,并作为不同状态的肿瘤抗原刺激DC,以流式细胞仪检测DC表面分子的表达。应用MTT法检测肿瘤抗原负载的DC刺激的CIK对肿瘤细胞的杀伤力。结果：负载肿瘤细胞冻融物的Dc其表面cDla、CD80、CD83、HLA-DR高表达,促进CIK增殖和杀伤力的功能最强;与正常肿瘤细胞共培养的DC细胞成熟程度受到抑制;负载Bel 7402抗原的DC刺激的CIK对Bel 7402有较强杀伤力,而对HerpG2杀伤力弱。结论：肿瘤细胞冻融物预刺激的DC成熟程度最高,诱导抗原特异性CIK细胞的功能最强,为临床DC瘤苗和CIK的应用提供了实验依据。     

抗原负载的DC和CIK共培养后对MGC-803的杀伤活性

【目的】研究负载抗原后的树突状细胞（DC）对细胞因子诱导的杀伤细胞（CIK）增殖活性、表型和功能的影响。【方法】分离外周血单个核细胞（PBMC），经不同细胞因子作用后定向分化成DC和CIK细胞，以胃癌细胞株MGC-803冻融物作为肿瘤抗原刺激DC后与CIK共培养，以流式细胞仪检测CIK细胞膜表面分子表达，MTT法检测共培养后CIK细胞对MGC-803杀伤活性的变化。【结果】CIK与负载MGC-803抗原的DC共培养后3d增殖活性开始显著提高，最高增殖倍数可达1179．5±1．29；CD3^＋CD8^＋和CD3^＋CD56^＋细胞较共培养前增多，且与负载抗原的DC共培养的CIK具有比单独培养的CIK更强的杀瘤活性。【结论】以肿瘤细胞冻融物诱导成熟的DC能进一步促进CIK增殖，提高其对胃癌细胞的杀伤活性，为指导临床应用DC和CIK联合治疗消化道肿瘤提供了实验依据。     

负载自体肿瘤抗原的树突状细胞联合细胞因子诱导的杀伤细胞在乳腺癌的临床研究

目的：探讨负载自身肿瘤裂解物的树突状细胞（DC）与细胞因子诱导的杀伤细胞（CIK）共培养后，对CIK体外肿瘤细胞杀伤活性的影响，进一步观察负载自身肿瘤裂解物的DC联合CIK治疗乳腺癌患者的临床免疫学疗效、毒副反应。方法：选择20例乳腺癌患者，分离外周血单个核细胞，其中贴壁细胞经GM—CSF和IL-4诱导产生DC，并负载自体肿瘤细胞裂解物；悬浮细胞经IFN-γ、IL-2、抗CD3单抗、IL-1α体外诱导产生CIK细胞。将负载抗原的DC与CIK共培养，观察CIK在体外对自身肿瘤细胞和乳腺癌细胞株SKBR-3的杀伤活性。此外，20例患者在切除原发病灶后，接受Ag—DC＋CIK细胞的过继免疫治疗，观察临床免疫疗效。结果：①CIK细胞培养后可见CD3^＋、CD8^＋和CD3^＋CD56^＋细胞较培养前显著增加；②将负载自身肿瘤抗原的DC与CIK共培养后，CIK的体外特异性和非特异性杀瘤活性明显提高；③细胞治疗后患者外周淋巴细胞Ag—NORs的IS值明显增高；CD3^＋、CD8^＋、CD56^＋升高；PBMC对自身肿瘤细胞的杀伤活性增强（P〈0．05）；④除一过性发热、畏寒外未见其它不良反应。结论：负载自体肿瘤细胞裂解物的DC疫苗联合CIK细胞在体外能明显增强对乳腺癌细胞的杀伤作用，亦能增强乳腺癌患者的细胞免疫功能，取得较理想的临床免疫疗效。     

抗原负载的树突状细胞介导的抗自身膀胱癌作用研究

目的:研究膀胱癌肿瘤细胞冻融抗原负载的树突状细胞(DC)对特异性抗自身膀胱癌作用研究。方法:将膀胱癌肿瘤病人外周血单个核细胞来源的DC在体外用冻融负载,再与膀胱癌患者的经细胞因子诱导的杀伤细胞(CIK)共同培养,应用乳酸脱氢酶释放法观察其对自身膀胱癌细胞杀伤活性,与未负载肿瘤抗原的CIK组进行比较。结果:DC经抗原负载后介导的CIK对自身膀胱癌细胞杀伤性活性明显增强。结论:用肿瘤抗原负载可进一步提高DC介导的CIK细胞对膀胱癌细胞的杀伤活性。     

中药复方肺瘤平膏对树突状细胞功能影响的拆方研究

目的观察肺瘤平膏及其拆方干预体外培养体系中树突状细胞(DC)抗原递呈功能的作用并探讨其分子机制。方法建立体外人外周血单个核细胞(PBMC)诱导分化成熟DC的诱导培养体系,从健康人外周血中分离获得PBMC,采用多种细胞因子(TNFα、IL-4、GM-CSF)联合诱导,获得了分化与功能相对成熟的DC,用流式细胞仪检测肺瘤平膏及其拆方含药血清干预DC与抗原递呈功能相关表面分子表达,及其对DC分泌IL-12含量的影响,进行初步研究。结果肺瘤平膏可上调DC与抗原递呈功能相关膜分子MHC-Ⅱ、CD80、CD83、CD86及CD40的表达,并促进DC分泌IL-12含量。结论肺瘤平膏能够调节DC抗原递呈功能,提高机体的抗肿瘤免疫监视功能。     

DC-CIK对卵巢癌细胞杀伤作用及临床治疗安全性评价

目的 树突状细胞(dendritic cells,DC)和细胞因子诱导的杀伤细胞(cytokine-induced killer cells,CIK)经体外共培养后,探讨其增殖能力和对卵巢癌细胞的杀伤能力,并对其治疗卵巢肿瘤的安全性进行评价.方法 提取健康志愿者外周血单核细胞(peripheral blood mononuclear cell, PBMC),通过不同细胞因子组合,分别诱导培养DC和CIK细胞,并共培养,用苔盼蓝染色计算活细胞扩增倍数,流式细胞术检测细胞表型,乳酸脱氢酶法(LDH)检测细胞毒性,酶联免疫剂吸附法(ELISA)检测细胞分泌因子水平.在卵巢癌患者签署知情同意书后,提取卵巢癌患者PBMC,诱导DC、CIK,并共培养,将质检合格的DC、CIK细胞通过腹腔注射的方式回输入患者体内,根据美国国立癌症研究所发布的《常见不良反应标准》,对不良反应进行判定,评价DC-CIK细胞治疗的近期安全性.结果 DC和CIK细胞共培养后,CIK增殖速度明显快于CIK单独培养,DC和CIK成熟分型明显增加,对肿瘤的杀伤能力比CIK单独培养时更强,且随着效靶细胞比的增加,杀伤肿瘤能力逐渐增强;DC-CIK回输早期,患者并未出现明显副反应.结论 DC-CIK共培养后,细胞增殖能力、成熟分型及协同抗瘤作用均增强;此外,DC-CIK相对安全可行.     

白介素-13受体α2抗原肽致敏的DC-CIK细胞对人胶质瘤U251细胞的杀伤效应

目的：观察人脑胶质母细胞瘤特异性抗原（IL-13Rα2）致敏的同源细胞因子诱导的杀伤细胞（cytokine induced killer,CIK）在体外对U251的细胞毒效应。方法：用密度梯度分离法获取HLA-A＊0201＋健康志愿者的人外周血树突状细胞（dendritic cells,DC）的前体细胞与淋巴细胞,并在体外诱导而获得成熟的DC细胞和CIK细胞。收获IL-13Rα2抗原负载的DC与CIK细胞共培养,使DC递呈肿瘤抗原予CIK细胞,并激活CIK细胞。激活的CIK细胞与U251细胞在96孔板内混合培养24 h后,以CCK-8试剂间接检测其对U251细胞杀伤率。结果：IL-13Rα2抗原肽成功负载DC,CIK细胞被负载抗原的DC激活并增殖;特异性抗原肽激活的CIK细胞对人胶质瘤U251细胞有杀伤效应（P〈0.01）。结论：IL-13Rα2抗原肽激活的CIK细胞对人胶质瘤U251细胞有杀伤效应。     

负载胃癌干细胞抗原的树突细胞对胃癌细胞免疫作用的研究

目的探讨胃癌干细胞抗原负载树突细胞(DC)联合细胞因子诱导杀伤细胞(CIK)对胃癌细胞的影响。方法分离人胃癌干细胞,冻融法制备抗原,将胃癌干细胞负载DC-CIK细胞,用流式细胞仪检测胃癌干细胞和DC、CIK细胞表型;将胃癌细胞与DC组、DC-CIK组、胃癌细胞抗原组、负载胃癌干细胞抗原组联合培养,用四甲基偶氮唑蓝法(MTT)检测不同组别对胃癌细胞的杀伤作用。结果经胃癌干细胞抗原刺激的DC细胞,DC表面成熟标志CD83和CD86表达明显增加,CD83和CD86表达率为80.4%,高于DC组、DC-CIK组、胃癌细胞抗原组,差异有统计学意义(P<0.01);MTT检测结果显示:负载胃癌干细胞抗原组、胃癌细胞抗原组、DC-CIK组、DC组对胃癌细胞杀伤率分别为(80.6±0.8)%、(72.3±0.6)%、(58.4±0.2)%和(49.7±0.8)%,差异有统计学意义(P<0.01)。结论胃癌干细胞作为抗原刺激DC细胞,可增强树突状细胞免疫原形表达,促进细胞增殖,提高对胃癌细胞的杀伤作用。     

SKOV3冻融抗原负载的DC-CIK细胞对SKOV3的杀伤作用

目的 探讨负载人卵巢癌细胞株SKOV3冻融抗原(Ag)的树突状细胞(DC)与细胞因子诱导的杀伤细胞(CIK)共培养后对SKOV3杀伤作用的影响.方法 取12例卵巢癌患者外周血,常规诱导出DE及CIK细胞,以SKOV3冻融Ag负载DC,经Ag负载与未经Ag负我的DC分别和CIK细胞共培养,流式细胞技术分析DC、Ag-DC、Ag-DC-CIK 3组DC细胞表型,CIK、DC-CIK、Ag-DC-CIK组CIK细胞表型;乳酸脱氢酶释放法测CIK组、DC-CIK组、Ag-DC-CIK组对SKOV3杀伤活性.结果 Ag-DC-CIK组中DC细胞成熟表型高于DC及Ag-DC组(P<0.01);Ag-DC-CIK组中CIK细胞成熟表型高于CIK及DC-CIK组(P<0.01);Ag-DC-CIK组对SKOV3杀伤率高于CIK及DC-CIK组(P<0.01).结论 负载SKOV3冻融Ag的DC与CIK共培养可促进DC、CIK的成熟,且可提高对SKOV3的杀伤作用.     

细胞因子诱导的杀伤细胞与树突细胞联合治疗晚期实体肿瘤40例临床分析

细胞因子诱导的杀伤细胞(cytokine induced killer,CIK)是以CD3 CD56 、CD3 CD8 T细胞为主要效应细胞的异质细胞群,具有增殖快、杀瘤活性高、杀瘤谱广等优点[1～3]。树突状细胞(dendritic cell,DC)是体内功能最强大的专职抗原递呈细胞,是惟一能激活幼稚T细胞(naive T cell)的抗     

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.     

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.     

Expression of OPGmRNA and ODFmRNA in the patients of hip fracture due to osteoporosis

Objective：To investigate the gene expression of osteoprotegerin（OPG） and osteoclast differentiation factor（ODF） in the bone tissue of patients with hip fracture due to osteoporosis. Methods：OPGmRNA and ODFmRNA in the bone tissue in 50 cases of osteoporosis sufferers（over 50 years old） with hip fracture（Observer Group） and 30 cases of hip facture sufferers with no osteoporosis（Control group） were analyzed with the Semi-Quantitative RT-PCR method. Results：The mRNA expressed of ODF, OPG were both high in the patients with hip fracture. In the control group, the expression of OPG mRNA was observed, while the expression of ODF mRNA was very slight. Conclusion：Aged patients contained all signals including OPG, ODF that are essential for inducing osteoclastogenesis and promoting bone resorption.     

The clinicopathological and immuohistochemical analysis of solid-pseudopapillary tumor of the pancreas： report of 9 cases

Objective：To investigate the clinical features, pathological characteristics and immunophenotype of solid-pseudopapillary tumor of the pancreas（SPTP）. Methods：Nine surgically treated cases of SPTP were retrospectively reviewed. Hematoxylin and Eosin（HE） staining and immunohistochemical staining were used to analyze all cases, and the general clinical data was collected. Results：Six patients were asymptomatic except for a palpable mass. Two patients complained of vague-epigastric pain. One patient appeared jaundice. The tumor was encapsulated and solid tissues alternately with cystic tissues. Histologically, the histological structure of solid portion was pseudopapillary with a fibrovascular core. Tumor cells were uniform and medium-sized which were arranged in sheets ets or nests or pseudopapillary patterns. Immunohistochemical studies demonstrated that SPTP proved positive in vimentin（9/9 cases）, AAT（9/9 cases）, NSE（9/9 cases）, ACT（7/9 cases）, CK20（2/9 cases）, CgA（1/9 cases）, S-100（3/gcases）, PR（4/gcases）, Syn（3/9 cases） and CD56（5/9cases）, negative in CEA and ER. Conclusion：SPTP is a tumor predominantly occurring in young women frequently without special symptoms. This tumor has various characteristical histological patterns with different immunophenotype.     

Electro-Acupuncture Combined with the Trigger Point Needle-Embedding for Treatment of Primary Trigeminal Neuralgia in 31 Cases

In recent years, the author of this essay has applied electro-acupuncture combined with the trigger point needle-embedding for treatment of primary trigeminal neuralgia in 31 cases, yielding satis- factory results as reported in the following.     

Investigation of Matrix Metalloproteinase -1, 2 and tissue inhibitor of matrix metalloproteinases-1 in Endometriosis

Objective： To explore the role of matrix metalloproteinase-1,2 （MMP-1, MMP-2） and tissue inhibitor of matrix metalloproteinases-1 （TIMP-1） in endometriosis. Methods： The eutopic and ectopic endometria from 40 subjects suffering from endometriosis and regular.endometria from 40 subjects （excluding endometriosis） were collected and examined by in situ hybridization technology and western blot assay. Results： Both expressions of MMP-1 and -2 were stronger in ectopic endometrium and eutopic endometrium than in normal endometrium. On the contrary, the expression of TIMP-1 in ectopic endometrium and eutopic endometrium was lower. The differences were significant （P 〈 0.01 ）. Moreover, there was no relationship among the expressions of MMP-1, 2 and TIMP-1 in ectopic endometrium. Conclusion： The expressions of MMP-1, 2 and TIMP-1 lose balance and lack of periodic changes in ectopic endometrium , which explains the biological invasive behavior of endometriosis. It was suggested-that regulating the balance between the MMPs and TIMP-1 should be an ideal therapeutic target to endometriosis.     

Shi Da-zhuo (史大卓) —An outstanding cardiologist of integrative medicine

Prof. SHI Da-zhuo, Ph.D., male, was born on March 20, 1960. Prof. SHI entered the Ph.D. program in 1990 at the China Academy of Chinese Medical Sciences under the supervision of Prof. CHEN Ke-ji, majoring in the treatment of cardiovascular diseases. After receiving his Ph.D. degree in 1993, Prof. SHI started working at the Cardiovascular Center in Xiyuan Hospital affiliated to China Academy of Chinese Medical sciences.     

Persist in and Carry Forward the Integrative Medical Research——Speech at the 6th National General Congress of Chinese Association of Integrative Medicine by Academician CHEN Zhu,Minister of Ministry of Health

The 6th National General Congress of Chinese Association of Integrative Medicine （CALM） was convened at 19-20, April 2008 in Beijing. Academician CHEN Zhu, the minister of Ministry of Health indicated at the congress that the integration of Chinese and Western medicine is very well in keeping with the situation of our country and the general rule of development in medical science; and as a good integration of Chinese medicine and Western medicine, it is mutually beneficial and advantageous to both of them. Seeing the creativity shown in integrative medical investigation in theoretic and methodological sides, we should and must persist in and develop it.