Influence of Helicobacter pylori infection on autologous DC-CIK maintenance therapy in the advanced gastric cancer patients

目的:对比幽门螺杆菌(HElICObACTEr PylOrI,HP)感染阴性和阳性的晚期胃癌患者接受自体树突状细胞联合细胞因子诱导的杀伤(dEndrITIC CEllS-CyTOkInE InduCEd kIllEr,dC-CIk)细胞维持治疗的疗效差异。方法:收集2010年6月至2012年6月中国人民解放军第174医院肿瘤中心收治的72例晚期胃癌患者,年龄29~90岁,中位年龄56岁,通过胃镜检查进行胃癌确诊并检测HP,分为HP阳性组(45例)及HP阴性组(27例),在接受手术或/和放化疗后,接受2疗程自体dC-CIk维持治疗,比较两组外周血培养的dC分化成熟情况及临床疗效差异。结果:两组dC成熟过程形态变化无差异;HP阳性组dC表面分子Cd83、Cd86表达显著高于HP阴性组(P0.05)。HP阳性组治疗后生活质量评分(kPS)、外周血T细胞亚群(Cd3+、Cd4+、Cd8+)比例较治疗前显著提高(P0.05)。HP阳性组治疗后kPS评分、CEA和CA199表达以及Cd3+和Cd4+T细胞数量等的改善均优于HP阴性组(P0.05)。共随访2年,阳性组患者中位生存期为12.64个月,长于阴性组的11.42个月(P 0.05).Levels of CD83 and CD86 on the surface of DCs were significantly higher in the Hp-positive group than those in the Hp-negative group( P 0.05).In the Hp-positive group,the quality of life KPS score and levels of T lymphocyte markers( CD3+,CD4+and CD8+) were significantly increased( P 0.05).Hp-positive patients had better KPS score,higher levels of CEA and CA199,and more abundant CD3+CD4+T cells than Hp-negative patients( P 0.05).At the end of 2-years follow-up,the median survival time was 12.64 months in the HP-positive group and 11.42 months in the HP-negative group( P < 0.05).Conclusion: Hp infection may help stabilize the tumor size,improve the quality of life,and prolong the survival time in patients with advanced gastric cancer undergoing maintenance therapy with DC-CIK cells.南京军区医学科技创新课题资助(No.10MA068); 厦门市科技计划创新项目资助(No.3502z20134026;No.3502z20144034)~

pJAK2 polypeptide,an antagonist of suppressors of cytokine signaling-1,can enhance the antitumor effect of dendritic cells

目的:探讨细胞因子信号转导抑制因子-1(SuPPrESSOrS Of CyTOkInE SIgnAlIng1,SOCS1)拮抗物PJAk2多肽(氨基酸序列号为1001-1013)参与树突状细胞(dEndrITIC CEllS,dCS)的体外诱导培养后对dCS抗肿瘤作用的影响。方法:采集健康人外周血,离心获得单个核细胞,用重组人粒细胞巨噬细胞集落刺激因子(rECOMbInAnT HuMAn grAnulOCyTE-MACrOPHAgE COlOny STIMulATIng fACTOr,rHgM-CSf)及重组人白细胞介素-4(rECOMbInAnT HuMAn InTErlEukIn-4,rHIl-4)诱导dCS,第5天分为4组:单纯dCS培养(对照)组、抗原负载(lySATE-dCS)组、多肽修饰(PJAk2-dCS)组和抗原与多肽共培养(lySATE+PJAk2-dCS)组,第6天各组加入肿瘤坏死因子-α(TuMOr nECrOSIS fACTOr-AlPHA,Tnf-α)促成熟。倒置显微镜下观察dCS形态;fCM法检测dCS表型;乳酸脱氢酶(lACTATE dEHydrOgEnASE,ldH)细胞毒实验检测各组细胞毒性T淋巴细胞(CyTOTOXIC T lyMPHOCyTE,CTl)对胃癌细胞bgC-823的靶向杀伤作用;ElISA法检测白细胞介素-12(InTErlEukIn-12,Il-12)和γ干扰素(InTErfErOn-γ,Ifn-γ)的水平。结果:与未加入诱导剂组相比,各组均成功诱导出成熟dCS,均高表达Cd80、Cd83、Cd86和人类白细胞dr抗原(HuMAn lEukOCyTE AnTIgEn dr,HlA-dr),但以lySATE+PJAk2-dCS组的表达水平最高。在10:1~30:1的效靶比范围内,CTl杀伤作用与效靶比呈正相关。当效靶比为30:1时,对照组的CTl杀伤率达(19.77±2.34)%,低于其他3组(P<0.01),而lySATE+PJAk2-dCS组较lySATE-dCS组及PJAk2-dCS组都高(P<0.05)。lySATE+PJAk2-dCS组培养上清液中Il-12及Ifn-γ的分泌水平明显高于对照组(P<0.01)。结论:SOCS1拮抗物PJAk2多肽(1001-1013)可增强dCS对胃癌细胞的抗原递呈及特异性抗肿瘤作用。Objective:To investigate the effect of pJAK2 polypeptide,an antagonist of SOCS1(suppressors of cytokine signaling 1),on antitumor effect of in vitro cultivation-induced DCs(dendritic cells).Methods:Peripheral blood was collected from the healthy volunteers,and the PBMCs(peripheral blood mononuclear cells)were isolated.DCs were induced by rhGM-CSF(recombinant human granulocyte-macrophage colony-stimulating factor)and rhIL-4(recombinant human interleukin-4).On the fifth day,DCs were divided into four groups:control group,Lysate-DCs group,pJAK2-DCs group,and Lysate + pJAK2 DCs group.On the sixth day,TNF-α(tumor necrosis factor-alpha)was added into each group.The morphological features of DCs were observed under an inverted microscope;the phenotypes were detected by FCM(flow cytometry);the killing effect of CTLs(cytotoxic T lymphocytes)on gastric cancer BGC-823 cells was evaluated by LDH(lactate dehydrogenase)cytotoxicity test;the concentrations of IL-12(interleukin-12)and IFN-γ(interferon-γ)were detected by ELISA(enzyme-linked immuno sorbent assay).Results:Mature DCs presented typically morphological and phenotypic features;the DCs in Lysate + pJAK2-DCs group had the highest expression levels of CD80,CD83,CD86 and HLA-DR(human leukocyte antigen DR).When the ratio of effectors to target cells ranged from 10:1 to 30:1,the killing activity of CTLs had a positive correlation with the ratio.When the ratio of effectors to target cells was 30:1,the killing activity of CTLs in the control group was(19.77±2.34)%,which was lowest as compared with the other groups(P < 0.01),meanwhile the killing activity of CTLs in Lysate + pJAK2-DCs group was higher than those in Lysate-DCs and pJAK2-DCs groups(P < 0.05).The levels of IL-12 and IFN-γ secretion in Lysate + pJAK2-DCs group were apparently higher than those in the control group(P < 0.01).Conclusion:An antagonist of SOCS1,pJAK2 polypeptide,can enhance the ability of antigen presentation and specific antitumor effect of DCs on gastric cancer cells.南京军区医学科技创新课题资助项目(编号:10MA068); 福建省自然科学基金面上项目(编号:2010D013); 厦门市科技计划创新项目(编号:3502z20104014

Effect of Helicobacter pylori on the function of peripheral blood monocyte-derived dendritic cells in gastric cancer patients

分析HP阳性和HP阴性的胃癌患者外周血单核细胞来源树突状细胞(MOdCS)功能的差异性及其临床意义。方法:用尿素14C呼气试验对解放军第一七四医院2011年1月至2012年10月收治的84例胃癌患者进行HP感染状态鉴定,分别采集HP阳性和阴性胃癌患者外周血,分离外周血单个核细胞,采用经典方法(rHgM-CSf、rHIl-4联合rHTnf-α)诱导产生dCS,采用流式细胞仪检测dCS表型,采用ldH释放法检测dCS致敏T细胞对胃癌细胞的毒性杀伤作用,采用ElISA方法检测细胞因子Il-12、Ifn-γ的分泌水平。结果:两组MOdCS成熟过程形态变化无差异,HP阳性组MOdCS表面标记分子Cd1A、Cd80、Cd83、Cd86和HlA-dr平均表达百分率均高于HP阴性组,其中Cd80、Cd83、Cd86的表达水平差异有统计学意义,Cd1A、HlA-dr差异无统计学意义。HP阳性组dCS致敏T淋巴细胞对胃癌细胞杀伤率和Il-12、Ifn-γ的分泌水平均高于HP阴性组(P<0.05)。结论:HP感染状态不影响胃癌患者外周血MOdCS成熟过程形态变化,HP感染的胃癌患者MOdCS成熟和活化水平更高。Objective:This study aimed to compare and analyze the functional differences between peripheral blood monocyte-derived dendritic cells(DCs) of Helicobacter pylori-positive and H.pylori-negative patients with gastric cancer.Methods:H.pylori infection was detected in 84 patients with gastric cancer in our hospital from January 2011 to October 2012 by the14C-urea breath test.DCs were generated from monocytes isolated by an adherent method from the two groups of patients and cultured in the presence of rhIL-4,rhGM-CSF,and rhTNF-α.Furthermore,the expression of surface marker molecules was determined by fluorescence-activated cell sorting analysis.The cytotoxicity of DCs pulsed T cells against gastric carcinoma cell was assessed by the lactate dehydrogenase-releasing assay.The secretion of IL-12 and IFN-γ in the supernatant was determined by enzyme-linked immunosorbent assay.Results:No difference was observed in the morphological change of the maturation process.The mean expression of CD1a,CD80,CD83,CD86,and HLA-DR molecules in DCs of H.pylori-infected patients was higher than that in DCs of H.pylori-negative group,and the differences were statistically significant except for CD1a and HLA-DR.The cytotoxicity activities,IL-12 release,and IFN-γ release in the H.pylori-positive group were significantly higher than those in the H.pylori-negative group(P<0.05).Conclusion:H.pylori infection has no effect on the morphological change of the maturation process of monocyte-derived DCs.These data clearly demonstrate that monocyte-derived DCs of H.pylori-infected patients with gastric cancer can induce stronger maturation and activation than those of H.pylori-negative patients.南京军区医学科技创新课题(编号:10MA068); 福建省自然科学基金资助项目(编号:2010D013); 厦门市科技计划创新项目(编号:3502z20104014)资助~

鄂尔多斯高原草地灌丛化不同阶段主要植物水分利用特征

草地灌丛化是全球草地退化的普遍现象,理解草地灌丛化的过程和机制对灌丛化草地的恢复十分必要。目前有多种假说解释草地灌丛化现象,尽管大部分假说认为灌木和草本植物对土壤水分的不同利用方式在草地灌丛化过程中发挥着重要作用,但有关直接实验证据仍十分缺少。本文通过对鄂尔多斯高原流动沙地沙米群落、半固定沙地油蒿群落、固定沙地油蒿群落和固定沙地本氏针茅群落4个演替阶段植物群落的调查,采用稳定同位素分析的方法,分析不同阶段主要物种对不同水源的利用比例。结果表明:在不同演替阶段,草本植物对土壤水分的利用主要集中在土壤表层(0～20 cm),而灌木植物对中深层(50～200 cm)土壤水分的利用较多;灌木植物水分利用的可塑性强,在遭受水分胁迫时,灌木植物能够根据土壤水分的可利用性而改变对不同土层水分的利用。以上结果对于认识鄂尔多斯高原不同演替阶段主要植物水分利用策略具有一定意义,同时表明降低放牧等人为因素对表层土壤的干扰,改善表层土壤结构,提高表层土壤含水量是鄂尔多斯高原油蒿灌丛化草地恢复为地带性本氏针茅草原的关键

长芒草(Stipa bungeana)种子萌发与出苗对关键环境因子的响应

种子萌发为植物生活史的关键阶段,对植物的定居、群落演替与动态起着决定作用。通过室内控制实验的方法,研究长芒草(Stipa bungeana)种子萌发对光照和温度,水势和温度,以及幼苗出土对沙埋深度和供水的响应。使用光照、温度和湿度自动控制的生长箱,设置光照、黑暗和持续黑暗3个光处理和5个变温处理(5℃:15℃、10℃:20℃、15℃:25℃、20℃:30℃和25℃:35℃)(夜:昼);设置11个水势处理(0、-0.2、-0.4、-0.6、-0.8、-1、-1.2、-1.4、-1.6、-1.8 MPa和-2 MPa)和5个恒温处理(10、15、20、25℃和30℃),研究长芒草种子萌发对光照和温度、水势和温度的响应。设置6个沙埋深度(0.5、1、1.5、2、3、5 cm)和10个供水处理(2.5、5、7.5、10、15、20 mm和30 mm 7个单次供水处理,2.5、5、7.5 mm/3d 3个多次供水处理),研究长芒草幼苗出土对沙埋和供水的响应。结果表明:光照对长芒草种子萌发具有促进作用,黑暗与持续黑暗条件下的最终萌发百分率无显著差异。在10:20℃—15:25℃温度范围内,长芒草种子萌发快速且集中,15:25℃为种子萌发最适温度,超过最适温度时,最终萌发百分率随温度升高而下降。水势低于-1 MPa时长芒草种子不能萌发,最终萌发百分率和萌发速率均随水势的降低而降低。当温度达到30℃时,种子基本不能萌发。单次供水条件下,长芒草的最终出苗百分率和出苗速率接近为零;5 mm/3d及7.5 mm/3d供水条件下,最终出苗百分率可达到50%。5 mm/3d及7.5 mm/3d的多次供水与单次供水及2.5 mm/3d的多次供水相比更有利于长芒草出苗。适宜出苗的沙埋深度为0.5—2 cm,沙埋深度超过3 cm时最终出苗百分率和出苗速率均率接近于零。长芒草种子出苗情况随沙埋深度增加而降低,在供水量为7.5 mm/3d,沙埋深度为0.5—2 cm时,出苗最好。降低人为干扰对表层土壤的破坏,提高表层土壤含水量,是促进长芒草种子萌发和幼苗出土的有效措施