The human leucocyte differentiation antigens (HLDA) workshops: the evolv-ing role of antibodies in research, diagnosis and therapy

The 8^th International Workshop on Human Leucocyte Differentiation Antigens （chaired by HZ and managed by BS） was run over a 4-year period and culminated in a conference in December 2004. Here we review the achievements of the HLDA Workshops and provide links to information on CD molecules and antibodies against them, including the 93 new CDs assigned in the 8^th Workshop. We consider what remains to be achieved （including an estimate of the number of leucocyte surface molecules still to be discovered）, and how the field can best move forward.

CD4^+ T cell-mediated presentation of non-infectious HIV-1 virion antigens to HIV-specific CD8^+ T cells

Background The mechanism of chronic immune activation and impairment of HIV-specific immune responses during chronic infection is not fully understood. However, it is known that high immune activation leads to more rapid progression to AIDS. We hypothesize that CD4^＋ T cell-mediated viral antigen presentation contributes to this pathologic immune activation in HIV-infected individuals. Methods HIV-specific T cells, responding to noninfectious HIV-1 virions as antigen, were measured by flow cytometric assays. These experimental conditions reflect the in vivo condition where noninfectious HIV-1 represents more than 99% of the antigens. Results CD4^＋ T cells purified from HIV-infected individuals were capable of cross presenting exogenous noninfectious HIV-1 virions to HIV-1-specific CD8^＋ T cells. Cross presentation required the entry of HIV-1 to CD4^＋ T cells and antigen translocation from endoplasmic reticulum to the Golgi complex. Blocking CD4^＋ mediated activation of HIV-specific CD8^＋ T cells and redirecting the viral antigens to antigen presenting cells improved HIV-specific T cell responses. Contusions One possible cause of chronic immune activation and impairment of HIV-1 specific T cell responses is represented by HIV-1 harboring CD4^＋ T cells cross presenting HIV-1 antigen to activate CD8^＋ T cells. This new mechanism provides the first evidence that cross presentation of noninfectious HIV-1 virions play a role in the immunopathogenesis of HIV-1 infection.

CD19 CAR-T细胞治疗难治/复发急性B淋巴细胞白血病儿童及青少年患者的疗效及安全性

目的探讨CD19嵌合抗原受体T细胞(CAR-T)治疗对于儿童及青少年难治/复发急性B淋巴细胞白血病(B-ALL)的疗效及安全性。方法回顾性分析2017年6月至2021年3月接受CD19 CAR-T治疗的<25岁难治/复发B-ALL患者的临床资料,评估该疗法的疗效及安全性。结果共纳入64例难治/复发B-ALL患者,男35例、女29例,中位年龄8.5(1.0~17.0)岁。CD 19 CAR-T回输后1个月进行短期疗效评估,64例患者均获得完全缓解(CR)/完全缓解兼部分血细胞计数缓解(CRi),其中有62例患者达骨髓微小残留病灶(MRD)阴性。细胞因子释放综合征(CRS)及免疫效应细胞相关神经毒性综合征(ICANS)发生率分别为78.1%及23.4%。共22例患者复发,中位复发时间10.1个月,4年总生存(OS)率为(66.0±6.0)%,4年无白血病生存(LFS)率为(63.0±6.0)%。长期随访结果显示桥接异基因造血干细胞移植(allo-HSCT)患者的LFS和OS率均优于未桥接移植患者(4年LFS率:81.8%±6.2%对24.0%±9.8%,4年OS率:81.4%±5.9%对44.4%±11.2%;均P<0.01)。结论CD 19 CAR-T可有效治疗难治/复发B-ALL,输注后桥接allo-HSCT能进一步改善患者的长期生存情况。

Effect of a cancer vaccine prepared by fusions of hepatocarcinoma cells with dendritic cells

AIM: To prepare a cancer vaccine (H(22)-DC) expressing high levels of costimulatory molecules based on fusions of hepatocarcinoma cells (H(22)) with dendritic cells (DC) of mice and to analyze the biological characteristics and induction of specific CTL activity of H(22)-DC. METHODS: DCs were isolated from murine spleen by metrizamide density gradient centrifugation, purified based on its characteristics of semi-adhesion to culture plates and FcR-,and were cultured in the medium containing GM-CSF and IL-4. A large number of DC were harvested. DCs were then fused with H(22) cells by PEG and the fusion cells were marked with CD11c MicroBeads. The H(22)-DC was sorted with Mimi MACS sorter. The techniques of cell culture, immunocytochemistry and light microscopy were also used to test the characteristics of growth and morphology of H(22)-DC in vitro. As the immunogen, H(22)-DC was inoculated subcutaneously into the right armpit of BALB/C mice, and their tumorigenicity in vivo was observed. MTT was used to test the CTL activity of murine spleen in vivo. RESULTS: DC cells isolated and generated were CD11c+ cells with irregular shape, and highly expressed CD80, CD86 and CD54 molecules. H22 cells were CD11c- cells with spherical shape and bigger volume, and did not express CD80, CD86 and CD54 molecules.H(22)-DC was CD11c+ cells with bigger volume, being spherical, flat or irregular in shape, and highly expressed CD80, CD86 and CD54 molecules, too. H(22)-DC was able to divide and proliferate in vitro, but its activity of proliferation was significantly decreased as compared with H(22) cells and its growth curve was flatter than H(22) cells. After subcutaneous inoculation over 60 days, H(22)-DC showed no tumorigenecity in mice, which was significantly different from control groups (P【0.01). The spleen CTL activity against H(22) cells in mice implanted with fresh H(22)-DC was significantly higher than control groups (P 【 0.01). CONCLUSION: H(22)-DC could significantly stimulate the specific CTL activity of murine spleen, which suggests that the fusion cells have already obtained the function of antigen presenting of parental DC and could present H(22)specific antigen which has not been identified yet, and H(22)-DC could induce antitumor immune response; although simply mixed H(22) cells with DC could stimulate the specific CTL activity which could inhibit the growth of tumor in some degree, it could not prevent the generation of tumor. It shows that the DC vaccine is likely to become a helpful approach in immunotherapy of hepatocarcinoma.

鼻内翻性乳头状瘤中Ki-67的表达及与微血管密度的关系

1.1临床资料。收集1999年1月～2005年7月手术切除确诊为鼻内翻性乳头状瘤（nasal inverted papilloma,NIP）标本存档蜡块36例,男22例,女8例;鼻息肉（nasal polyps,NP）14例,男9例,女5例：鼻腔鳞状细胞癌（nasal squamous cell caminoma，NSCC）10例，男77例，女3例。

透明质酸和标准型白细胞分化抗原44在嗜酸性粒细胞型和非嗜酸性粒细胞型鼻息肉发病过程中的意义

1资料与方法1.1临床资料。随机选取我科2013~2014年行手术治疗的嗜酸性粒细胞型鼻息肉(eosinophilic chronic rhinosinusitis with nasal polyps,ECRSw NP)和非嗜酸性粒细胞型鼻息肉(non-eosinophilic chronic rhinosinusitis with nasal polyps,n ECRSwN P)标本各15例,取同期因行鼻中隔偏曲手术切除的下鼻甲黏膜标本15例作为对照组。所有标本均经病理学证实,其中鼻息肉分型标准为[1]:

Serum concentration of sFas and sFasL in healthy HBsAg carriers,chronic viral hepatitis B and C patients

AIM:To estimate the amount of apoptosis among healthy HBsAg carriers,patients with chronic HBV infection treated wibh lamivudine and patients with chronic HCV infection treated with interferon alpha and ribavirin.Activity of apoptosis was evaluated by serum sFas/sFasL concentration measurement. Moreover dependence between apoptosis and HBV-DNA or HCV-RNA levels was studied. METHODS:Eighty-six persons were included into study:34 healthy HBsAg carders,33 patients with chronic HBV infecl^on and 19 patients with chronic HCV infection.Serum levels of sFas/sFasL were measured by ELISA assay.HBV-DNA and HCV-RNA were measured by RT-PCR assay.Levels of sFas/sFasL were determined before and 2 and 12 wk after therapy in patients with chronic hepatitis B and C infection. HBV-DNA or HCV-RNA was detected before treatment and 6 mo after treatment. RESULTS:Twenty-four (71%) healthy HBsAg carders showed HBV-DNA over 10~5/mL,which was comparable to the patients with chronic hepatitis B.independently from HBV-DNA levels, the concentration of sFas among healthy HBsAg carders was comparable to healthy persons.Among patients with chronic hepatitis B and C,the concentration of sFas was significantly higher in comparison to healthy HBsAg carriers and healthy persons.In chronic hepatitis B patients the concentration of sFas was decreased during lamivudine treatment.Among chronic hepatitis C patients the concentration of sFas was increased during IFN alpha and ribavirin treatment,sFasL was not detected in control group.Furbhermore sFasL occurred more frequently in chronic hepatitis C patients in comparison to chronic hepatitis B patients. CONCLUSION:There are no correlations between apoptosis and HBV-DNA levels.However ther is an association between apoptosis and activity of inflammation in patients with chronic HBV infection.Apoptosis can be increased in patients with chronic hepatitis C by effective treatment which may be a result of apoptosis stimulation by IFN-α.

胎盘来源细胞与母血、脐血细胞的嵌合

背景:胎盘中含有丰富的细胞群,因其中CD34+细胞含量高而受到越来越多学者的关注,有望成为临床造血干细胞移植治疗血液病及其他恶性疾病的新来源。目的:探索胎盘来源细胞的数量、集落形成能力及其与母体的嵌合程度。方法:采用健康足月剖腹产妇的胎盘,灌注法及组织块消化法收集胎盘中细胞;流式细胞仪检测胎盘及脐血中CD34+细胞比例;培养细胞集落,定期观察集落形态并计数;序列特异性引物PCR扩增技术及序列特异性寡核苷酸探针方法检测胎盘、脐血和母亲外周血的HLA类型;短串联重复PCR方法检测胎盘细胞、脐血细胞和母亲外周血细胞的嵌合情况。结果与结论:胎盘中CD34+细胞数量明显优于脐血,且胎盘具有体外多种集落形成能力,集落形成良好,同时胎盘提取细胞中含有母体来源成分。提示胎盘中细胞数量及其基本生物学特性使其有潜力成为临床造血干细胞移植的新来源。

胃肠间质肿瘤免疫组化及细胞增殖活性研究

目的：研究胃肠间质肿瘤（ＧＩＳＴｓ）的组织起源、分化以及细胞增殖。方法：应用ＬＳＡＢ法观察ＣＤ３４等系列标记物在２８例ＧＩＳＴｓ中的表达。结果：ＧＩＳＴｓ中ＣＤ３４和Ｖｉｍ的阳性率高达８８．５％和９４％，且阳性程度强；肌源性和神经源性标记为部分阳性且相对偏弱；ＣＤ３４和ＰＣＮＡ在良恶性ＧＩＳＴｓ中表达无差异，Ｋｉ６７有一定区别。结论：ＧＩＳＴｓ是一类独立的非定向分化的肿瘤，可能起源于胃肠道原始间叶组织，部分可伴有不完全的平滑肌或（和）神经鞘细胞分化，组织学上类似平滑肌肿瘤或神经鞘肿瘤，ＣＤ３４和Ｖｉｍ是较为特异而敏感的标记物。

CD_(44V6)在胃癌中表达及其临床意义

目的：探讨ＣＤ４４Ｖ６与胃癌发生及胃癌生物学行为的关系。方法：应用免疫组化Ｓ－Ｐ方法研究１０例胃粘膜肠化及４６例胃癌的ＣＤ４４Ｖ６表达。结果：胃粘膜肠化及胃癌阳性率分别为１０％和６３％，淋巴结转移组阳性率（７５％）明显高于非转移组（４５％，Ｐ＜０．０５），且ＣＤ４４Ｖ６表达与Ｌａｕｒｅｎ分型相关。结论：ＣＤ４４Ｖ６分子参与肿瘤发生。

接受恩替卡韦治疗的慢性乙型肝炎患者外周血单个核细胞B7-H1表达的动态变化

近年的研究逐渐揭示免疫学因素影响着慢性乙型肝炎（CHB）的临床进展。B7-H1（又称PD-L1,CD274）及其受体PD-1是近年发现在T细胞激活中介导负性共刺激信号的分子,已有众多研究证实其在外周免疫耐受、自身免疫性疾病及慢性感染中发挥着重要作用。

复方黄黛片诱导急性早幼粒细胞白血病细胞凋亡的研究

为探讨复方黄黛片治疗急性早幼粒细胞白血病 (APL)的作用机制 ,观察了 2 1例经复方黄黛片治疗的APL患者 ,取不同治疗阶段的患者骨髓细胞 ,用流式细胞仪检测APL骨髓细胞分化抗原 (CD33和CD1 1b)、细胞周期 ,以及与细胞凋亡相关的Fas和Apo2 .7蛋白的表达。结果显示 ,服用复方黄黛片治疗后 ,患者骨髓的APL细胞CD33表达明显下降 ,CD1 1b表达升高 ,对细胞周期的影响主要表现为G2 /M期的阻滞 ,服药后Fas蛋白及Apo2 .7蛋白表达增加。提示复方黄黛片对APL细胞有凋亡和分化的双重作用 ,且细胞凋亡可能与Fas和Apo2 .

CD_(44)表达与乳腺癌淋巴结转移及预后相关因素的关系

目的：探讨ＣＤ４４表达与乳腺癌淋巴结转移与预后的关系。方法：采用ＬＳＡＢ法对７６例乳腺癌伴淋巴结转移和无转移组ＣＤ４４表达进行了免疫组化检测，并结合临床资料、增殖细胞核抗原（ＰＣＮＡ）、雌激素受体（ＥＲ）表达情况进行综合分析。结果：ＣＤ４４阳性率在乳腺癌伴淋巴结转移组为７０６％，无转移组为４５２％，两组间存在显著性差异（Ｐ＜００５），随着组织学分级的增高ＣＤ４４表达呈递增趋势，Ⅰ级与Ⅱ、Ⅲ级间阳性率差异存在显著性（Ｐ＜００５）；ＣＤ４４与ＰＣＮＡ表达存在平行关系，与ＥＲ表达呈负相关趋势。结论：ＣＤ４４表达与乳腺癌淋巴结转移及预后有关。

CD_(14)^+单核细胞人类白细胞抗原-DR预测脓毒症预后及指导免疫调理治疗的初步临床研究

目的 :验证 CD+ 1 4 单核细胞人类白细胞抗原 DR( HL A DR)对于评估严重脓毒症患者免疫功能的作用 ;探讨胸腺 5肽 ( TP 5 )治疗免疫抑制的有效性。方法 :符合严重脓毒症标准 ,CD+ 1 4 单核细胞 HL A DR<30 %的患者被定义为脓毒症免疫抑制和代偿性抗炎症反应综合征 ( CARS)者纳入本研究 ,并接受 1m g TP 5肌肉注射 ,1次 / d,直至 CD+ 1 4 单核细胞 HL A DR>5 0 %或死亡。在 TP 5治疗前及结束治疗时分别测量 CD+ 1 4单核细胞 HL A DR和肿瘤坏死因子α( TNFα)、白介素 6 ( IL 6 )、IL 10和 IL 13。结果 :15例患者存活 ,5例死亡。经 TP 5治疗后所有患者的 CD+ 1 4 单核细胞 HL A DR均有不同程度提高 ,但仅存活者有显著差异。所有患者细胞因子均高于健康对照 ,治疗后存活患者的 TNFα、IL 6明显下降 ;死亡者各种细胞因子变化不明显。结论 :用 CD+ 1 4 单核细胞 HL A DR鉴别脓毒症免疫抑制并指导免疫刺激治疗是安全可靠的 ,且对评估预后有重要价值 ;TP 5可能对逆转免疫抑制有效 ,但需要严格的对照治疗研究确认 ;脓毒症的免疫抑制发生和逆转与促炎 /抗炎细胞因子平衡无关 ,确切机制有待深入探讨。

5-FC/CD系统对荷大肠癌裸鼠肿瘤的杀伤效应

目的研究ｃｄ基因对大肠肿瘤的特异杀伤作用，探索自杀基因靶向治疗大肠癌的有效途径．方法逆转录病毒感染法将Ｇ１ｃｅａｃｄＮａ及ｐｃｄ２分别转导入高分泌ＣＥＡ的大肠癌细胞Ｌｏｖｏ中，再分别接种到裸鼠皮下．成瘤后腹腔给予５ＦＣ（５００ｍｇ／ｋｇ）治疗，观察肿瘤重量的变化及病理学特点．结果含Ｇ１ｃｅａｃｄＮａ及ｐｃｄ２逆转录病毒载体的病毒滴度分别为：１３×１０７及２１×１０８ＣＦＵ／Ｌ．所有转基因成功并接种动物均成瘤．腹腔给予５ＦＣ治疗后发现，转由ＣＥＡ基因顺式转录调控序列（ＴＲＳ）驱动ｃｄ基因的组织特异性重组逆转录病毒载体Ｇ１ｃｅａｃｄＮａ的肿瘤对５ＦＣ的敏感性明显高于转非ｃｅａ调控ｃｄ基因的肿瘤，治疗结束后肿瘤重量分别为３１ｍｇ±８ｍｇ及１１３ｍｇ±２３ｍｇ（Ｐ＜００１）．结论本实验提示ｃｅａ转录调控序列可控制ｃｄ基因在ＣＥＡ阳性的大肠癌组织中高效表达。

传染性单核细胞增多症患儿免疫功能的动态观察

目的 :探索传染性单核细胞增多症 (IM)患儿免疫功能的动态变化。方法 :应用流式细胞仪分别对 5 0例传染性单核细胞增多症 (IM)患儿不同病期外周血淋巴细胞 CD3、 CD4 、 CD8、 CD1 6 + 56 抗原进行检测 ,并以同样例数的健康儿童作对照。结果 :入院初检测 ,IM患儿外周血 CD3+ 、 CD8+ 淋巴细胞百分率明显高于对照组而 CD4 + 、 CD1 6 + 56 + 淋巴细胞百分率及 CD4 +细胞与 CD8+细胞的比值 (CD4 + / CD8+ )明显低于对照组 (P<0 .0 1)。出院前患儿的临床症状已基本消失 ,但上述各项检测指标却仍明显异于对照组 (P<0 .0 1) ,直至出院一周各项指标才基本恢复正常。结论 :IM患儿疾病过程中外周血淋巴细胞各功能群明显异常 ,提示机体体液免疫和细胞免疫功能均发生紊乱 ,需临床症状消失一周才逐渐恢复。该病的护理中预防细菌继发感染尤为重要。

国产胸腺肽治疗慢性乙肝疗效及其机理初探

本文观察大剂量胸腺肽对慢性乙肝患者的治疗作用并研究其作用机理。用ＥＬＩＳＡ、ＰＣＲ 和免疫化学染色法等比较分析胸腺肽治疗三个月后，ＡＬＴ、ＨＢＶ－ Ｍ 和ＨＢＶ ＤＮＡ 等指标变化，发现患者ＡＬＴ 复常率、ＨＢｅＡｇ 转阴率、ＨＢＶ ＤＮＡ 阴转率和ＨＢＶ ＤＮＡ 有效下降率分别为５８．３３ ％ 、３３．３３ ％ 、４１．６７ ％ 和９１．６７ ％ ；同时患者外周血ＣＤ４ ＋淋巴细胞亚群上调，ＣＤ８ ＋ 亚群下调，二者比值趋于正常。说明国产胸腺肽治疗慢性乙肝有较好的应用前景，其治疗机理与免疫调节作用有关。

流式细胞术评价极高频电磁波对化放疗肿瘤患者免疫细胞表达的影响

目的探讨低功率极高频电磁波(extremely high frequency wave,EHFW)辐照对肿瘤化放疗患者免疫细胞表达的影响。方法62例恶性肿瘤化放疗患者按自愿和随机原则分为化放疗后极高频电磁复合波幅照组(联合治疗组)及单纯化放疗后观察组(单纯化放疗组)。联合治疗组患者于化放疗后配合以功率密度为10 mW/cm2的EHF波辐照。采用流式细胞术配比性(Matched-pair)检测患者EHF波辐照治疗前-后外周血免疫学指标。结果单纯化放疗组患者在化放疗后第8天,除CD3+、CD4+、CD8+T细胞表达率和CD4+/CD8+比值与化放疗后第3天水平相当外,其NK细胞、NKT细胞、B细胞表达率均明显低于化放疗后第3天水平(P<0.01);而EHF波辐照治疗5 d后,NK细胞表达水平显著高于化放疗后第3天(P<0.05);NKT细胞、B细胞、CD3+、CD4+、CD8+T细胞表达率和CD4+/CD8+比值均略高于化放疗后第3天水平,其前-后变化水平均未见统计学意义(P>0.05)。结论低功率EHF波配合化放疗治疗恶性肿瘤,可稳定和提高免疫细胞表达水平,对患者细胞免疫机能具有保护作用。

胃癌患者外周血T淋巴细胞亚群的测定及临床意义

目的 分析胃癌患者外周血T淋巴细胞亚群水平的特点、影响因素及临床意义。方法 采用流式细胞术 (FCM)检测 30 8例胃癌患者外周血中的T淋巴细胞亚群的水平与 4 8例正常对照比较研究。结果 患者外周血CD3 + 、CD4+ 明显减少 ,而CD8+ 明显增加 ,CD4+ /CD8+ 比值明显下降 ,与正常组比较差异具有高度显著性 (P <0 .0 1) ;患者性别、年龄、临床分期不影响其表达 ,差异无显著性 (P >0 .0 5 )。结论 胃癌患者存在细胞免疫功能紊乱 ,检测胃癌患者外周血T淋巴细胞亚群的表达对评估患者的细胞免疫功能及疾病的诊断具有一定的临床意义。

镉胁迫对拟南芥幼苗错配修复基因表达的影响

采用半定量反转录聚合酶链式反应(RT-PCR)技术研究了Cd胁迫对拟南芥(Arabidopsis thaliana)幼苗错配修复和增殖细胞核抗原基因表达的影响,并结合幼苗的形态和生理指标,选取Cd胁迫敏感的生物标记物。结果表明:不同浓度(0.25、0.5、1.0 mg.L-1)Cd处理对拟南芥幼苗叶片数、地上部鲜质量影响不大;Cd浓度为0.25 mg.L-1时,地上部分可溶性蛋白质含量明显升高(P<0.05),Cd浓度为0.5和1.0 mg.L-1时,可溶性蛋白质含量明显降低(P<0.05);叶绿素含量随着Cd浓度的增加而微弱增加(P>0.05)。Cd浓度为0.25 mg.L-1时,以18S rRNA为内参照,PCNA1、PCNA2、MSH2、MSH3、MSH6、MSH7 6个基因均出现了诱导表达,当Cd浓度增加到1.0 mg.L-1时,除了MSH6持续表达诱导及MSH3基因与对照相比表达抑制外,其他基因的表达依然出现诱导,但都低于0.5 mg.L-1Cd处理下的基因表达水平。以上结果表明,基因表达的改变可作为检测Cd污染对植物遗传毒性效应潜在有用的生物标记物。