肾移植受者血他克莫司浓度对外周血自然杀伤细胞及其受体的影响

目的 研究肾移植受者血他克莫司(Tac)浓度对外周血自然杀伤(NK)细胞及其受体的影响.方法 将2007年12月至2009年7月间的60例受者纳入研究,术后受者均采用以Tac为基础的免疫抑制方案.根据术后6个月监测到的血Tac浓度将受者分为低浓度组和高浓度组[各为30例,术后6个月时血Tac浓度分别为(6.84±1.72)和(11.88±2.59)μg/L],另以20名健康志愿者作为对照组.术前和术后6个月,采用流式细胞术检测NK细胞及其抑制性受体(CD85j和CD158d)和活化性受体( CD94、NKG2D)的表达情况,采用酶联免疫吸附试验法检测免疫耐受分子分泌型HLA-G5( sH LA-G5)的表达水平.结果 术前低浓度组和高浓度组受者外周血NK细胞绝对值均较对照组显著降低(P＜0.05),术后6个月时低浓度组和高浓度组NK细胞比例及绝对值较对照组均显著降低(P＜0.05),低浓度组NK细胞绝对值显著高于高浓度组(P＜0.05).术前两组间CD85j、CD158d、CD94、NKG2D表达的差异均无统计学意义(P＞0.05);术后6个月时低浓度组和高浓度组CD85j和CD158d的表达较术前升高,CD94和NKG2D的表达下降,而低浓度组CD85j和CD158d的表达显著高于高浓度组(P＜0.05).经Spearman系数统计,CD85j和CD158d与sHLA-G5呈正相关(P＜0.01),NKG2D与sHLA-G5呈负相关(P＜0.01).结论 肾移植受者血Tac浓度与外周血NK细胞数量及其受体的表达具有相关性,低血Tac浓度受者的NK细胞数量及其抑制性受体的表达升高,仍然能有效保护移植肾功能.     

Zeta chain expression in T and NK cells in peripheral blood of children with nephrotic syndrome

The idiopathic nephrotic syndrome (INS) has been related to cellular immune disturbances. The zeta (ζ) chain, a component of the T-cell receptor/CD3 (TCR) complex and CD16 heterodimer in NK cells, plays a crucial role in T and NK cell activation and proliferation. The aim of our study was to examine zeta chain expression in CD4+, CD8+ T lymphocytes and NK cells in the peripheral blood of children with INS and to evaluate the effect of anti-CD3+rIL-2 stimulation on the level of zeta chain expression in the INS pediatric population. The study group consisted of 15 children with INS in relapse, 16 patients with INS in clinical remission, and 17 controls. The percentage of zeta-positive cells and the values of mean fluorescence intensity (MFI) were determined by flow cytometry. Compared with that in the controls, the percentage of zeta+ freshly isolated NK cells in children with INS in relapse was significantly lower, whereas, in CD3+/CD4+ and CD3+/CD8+ populations, no alteration was observed. There were no differences in the MFI values between the populations of freshly isolated cells either. Stimulation with anti-CD3+rIL-2 decreased the percentage of zeta+/CD4+ T cells and NKzeta+ cells in a significant way in all the groups analysed, whereas the percentage of zeta+/CD8+ T cells decreased significantly only in patients with INS in relapse. The altered pattern of zeta expression in fresh NK cells from children with INS in relapse, and the disturbed response of zeta+/CD8+ T cells to anti-CD3+rIL-2 stimulation in relapse, suggests the possible role of this chain in immune dysregulation in INS, particularly with regard to cytotoxic cells.     

Sex differences in cytokine production and surface antigen expression of peripheral blood mononuclear cells after surgery

BACKGROUND: Several clinical and epidemiological studies have observed a better outcome after sepsis in women than in men. The purpose of this study was to determine if these sex differences are observed in cytokine responses and the surface antigen expression of monocytes. In addition, the clinical courses of male and female patients after gastrointestinal surgery were compared. METHODS: A total of 25 patients with gastric carcinoma who underwent gastrectomy were enrolled in this study. Tumor necrosis factor-alpha (TNF-alpha), interleukin-10, and interferon-gamma (IFN-gamma) production by lipopolysaccharide-stimulated peripheral blood mononuclear cells (PBMCs) as well as the expression of Toll-like receptor-4 (TLR-4), TLR-2, human leukocyte antigen-D related (HLA-DR), and CD16 on monocytes in 16 men and 9 women on the day before surgery were compared with measurements on postoperative day (POD) 1. Furthermore, postoperative infectious complications, the development of systemic inflammatory response syndrome, and serum C-reactive protein levels on POD3 were compared. RESULTS: TNF-alpha production of PBMCs and TLR-2 and CD16 expression on monocytes were significantly higher in women than in men before surgery. IFN-gamma production of PBMCs and HLA-DR expression on monocytes were significantly lower in men than in women on POD1. Furthermore, TNF-alpha production of PBMCs on POD1 was significantly increased, and both IFN-gamma production and HLA-DR expression were significantly decreased compared with that observed before surgery in men, but no corresponding significant changes were observed in women. In addition, C-reactive protein levels on POD3 were significantly higher in men than in women. CONCLUSIONS: Both TNF-alpha and interleukin-10 production of PBMCs and both TLR-2 and CD16 expression on monocytes were significantly higher in women than in men on the day before surgery. Excessive TNF-alpha and suppressive IFN-gamma production of PBMCs, as well as a decrease in HLA-DR expression on monocytes, occurred more often in men than in women after surgery, suggesting that these factors all contribute to an increased susceptibility of men to develop systemic inflammatory response syndrome or postoperative infectious complications.     

肾移植术后外周血自然杀伤细胞的CD158b表达及意义

目的探讨肾移植术后外周血自然杀伤细胞(NK细胞)的CD158b表达及意义。方法测定62例患者肾移植前、术后第1d、术后第7d、肾功能正常时以及疑有排斥反应时外周血NK细胞的CD158b表达水平。结果62例中,术后38例肾功能恢复正常,观察期内无排斥反应发生,移植前后外周血CD3-CD16/CD56 细胞(NK细胞)及CD3-CD16/CD56 CD158b 细胞稳定,NK细胞中CD158b 细胞的比例也稳定;24例术后7～14d发生急性排斥反应,其外周血CD3-CD16/CD56 细胞呈上升趋势,CD3-CD16/CD56 CD158b 细胞呈下降趋势,NK细胞中CD158b 细胞的比例也呈下降趋势,经单因素方差分析,各指标在不同测定时点的差异均有统计学意义(P<0.05,P<0.01)。结论干扰NK细胞表达CD158b的因素较少,在临床上做出排斥反应诊断前,患者外周血中NK细胞的CD158b表达即呈下降趋势,因此术后监测NK细胞的CD158b表达可为评价患者的免疫状况提供依据。     

人骨肉瘤外周血T淋巴细胞亚群和自然杀伤细胞免疫变化的研究

目的研究人骨肉瘤患者外周血T淋巴细胞亚群和自然杀伤细胞(NK细胞)的变化。方法采用流式细胞术对42例骨肉瘤、41例骨髓炎、34例健康者的外周血CD4 、CD8 、CD4 、CD4 /CD8 、CD3 、CD16 56均显著性低于骨髓炎患者和健康对照组(P<0.01),而CD8 则升高(P<0.05)。骨髓炎患者外周血CD4 、CD4 /CD8 、CD3 较健康人降低,CD8 则升高,但差异均无统计学意义(P>0.05),而CD16 56与健康对照组相比降低(P<0.05)。结论骨肉瘤患者细胞免疫功能明显降低;骨肉瘤患者的免疫抑制远较骨髓炎的强;骨肉瘤患者NK细胞的降低较骨髓炎患者明显多。流式细胞术可作为检测骨肉瘤患者免疫功能的有效方法。     

Cell surface receptors in malignant glioma

Despite advances in surgery, radiation, and chemotherapy, malignant gliomas are still highly lethal tumors. Traditional treatments that rely on nonspecific, cytotoxic approaches have a marginal impact on patient survival. However, recent advances in the molecular cancer biology underlying glioma pathogenesis have revealed that abnormalities in common cell surface receptors, including receptor tyrosine kinase and other cytokines, mediate the abnormal cellular signal pathways and aggressive biological behavior among the majority of these tumors. Some cell surface receptors have been targeted by novel agents in preclinical and clinical development. Such cancer-specific targeted agents might offer the promise of improved cancer control without substantial toxicity. Here, we review these common cell surface receptors with clinical significance for malignant glioma and discuss the molecular characteristics, pathological significance, and potential therapeutic application of these cell surface receptors. We also summarize the clinical trials of drugs targeting these cell surface receptors in malignant glioma patients.     

IGG and complement receptor expression on peripheral white blood cells in uraemic children.

BACKGROUND: Phagocytosis of IgG- or complement-opsonized bacteria and antibody production by lymphocytes are regulated by cell surface receptors for IgG (FcgammaRI, FcgammaRII and FcgammaRIII) and complement (CR1 and CR3). We measured the effect of uraemia and dialysis treatment on FcgammaR and CR expression on leukocytes in blood. METHODS: Blood samples were obtained from children: 40 treated with peritoneal dialysis (PD), 23 with haemodialysis (HD), 46 not yet dialysed (CRF) and 33 healthy (HC). White blood cells, isolated from EDTA-blood by centrifugation after cell fixation with paraformaldehyde, were labelled with FITC-conjugated CD16 (FcgammaRIII), CD32 (FcgammaRII), CD64 (FcgammaRI), CD11b (CR3) and CD35 (CR1) monoclonal antibodies and analysed by flow cytometry. RESULTS: In PD, HD, CRF and HC, monocytes and neutrophils were all positive for FcgammaR and CR, except for CD16 on monocytes (20% positive). Lymphocytes expressed CD16 and CD32 but not CD64. PD, HD and CRF children had lower percentages of CD16(+) and CD32(+) lymphocytes compared with HC. The percentage of CD11b(+) lymphocytes was lower only in PD and the percentage of CD35(+) lymphocytes was lower in HD and CRF compared with HC. The median CD32 mean fluorescense intensity (MFI) on lymphocytes, monocytes and neutrophils was lower in PD, HD and CRF compared with HC. On the other hand, CD11b MFI on lymphocytes, monocytes and neutrophils was higher in PD, HD and CRF children compared with HC. CD16 and CD64 MFI were not different among the groups and CD35 MFI was only lower on lymphocytes from PD, HD and CRF compared with HC. CONCLUSIONS: In children with chronic renal failure, whether dialysed or not, FcgammaRII expression on lymphocytes, monocytes and neutrophils was reduced and CR3 expression was increased. Furthermore, CR1 expression on lymphocytes, important for the humoral response, was lower in children with renal failure. Age and uraemia are associated with these abnormalities and might contribute to impaired immune function in children with chronic renal failure.     

慢性乙型肝炎病毒感染者外周血T细胞亚群及NK细胞的特点及意义

目的探讨慢性乙型肝炎病毒（hepatitis B virus,HBV）感染者外周血T细胞亚群及NK细胞的特点及临床意义。方法采用流式细胞术检测各研究组,包括慢性乙型肝炎（chronic hepatitis B,CHB）组33例、乙型肝炎失代偿期肝硬化（1iver cirrhosis,LC）组20例、慢性乙型重型肝炎（chronic severe hepatitis B,CSH）组17例及健康对照组（Control）20例的外周血T细胞亚群及NK细胞相对计数,并检测肝功能、HBVDNA含量及HBV血清标志物。结果按Control、CHB、LC、CSH顺序,CD3^＋T细胞、CD8^＋T细胞百分比依次升高,而CIM^＋T细胞、CD4^＋／CD8^＋比值及NK细胞百分比依次降低,且CHB、LC、CSH组与Control组及CHB组与CSH组相比,差异均有统计学意义（P〈0．05或P〈0．008）。CHB患者的CD3^＋T细胞与血清总胆红素（total bilirubin,TB）、HBVDNA含量（log_10）呈正相关（P〈0．001;P〈0．001）;CD8^＋T细胞与HBVDNA含量（log_10）呈正相关（P=0．007）,NK细胞与HBVDNA含量（log_10）（P=0．001）呈负相关。CHB组乙型肝炎e抗原（HBeAg）阳性者的CD4^＋T细胞及CD4^＋／CD8^＋比值低于HBeAg阴性者（P=0．018;P〈0．001）,而HBVDNA含量（log_10）和CD8^＋T细胞高于HBeAg阴性者（P=0．012;P=0．019）。结论慢性HBV感染者外周血T细胞亚群及NK细胞相对值紊乱,且与临床类型、病情、血清HBVDNA水平及HBeAg相关。     

Proto-oncogene expression in peripheral blood mononuclear cells in IgA nephropathy.

We have investigated the expression of proto-oncogenes in mononuclear cells obtained from patients with IgA nephropathy using a RNA hybridization technique. Patients with IgA nephropathy expressed more c-myc, c-raf, c-fos, and c-jun proto-oncogene RNA than did normal controls. However, no significant expression of c-N-ras, c-mos or c-myb genes was found in the mononuclear cells of these patients. When the amount of urinary protein excretion was used as an indicator of disease activity (greater than 1 g/day), a positive correlation was found between c-myc, c-raf, c-fos, and c-jun expression and urinary protein excretion (P less than 0.01). The expression of these genes correlated also with the serum IgA concentration (P less than 0.01), IgA immune complex (P less than 0.01), and histopathological changes in renal tissues obtained from patients with IgA nephropathy (P less than 0.01). The results of this survey suggest that abnormally regulated proto-oncogene expression in mononuclear cells may play an important role in the progression of IgA nephropathy and may be useful as an indicator of disease activity and/or prognosis.     

尿毒症患者外周血单个核细胞超保守区域转录表达水平的分析

目的 尿毒症是由于肾衰竭致使代谢产物和其他有毒物质在体内蓄积而引起的一种自身中毒综合征,超保守区域转录子(transcribed ultra-conserved regions,T-UCRs)是lncRNA重要组成部分,转录自人类基因组中481个极度保守区域,希望分析尿毒症外周血单个核细胞的超保守区域转录表达水平,从而发现尿毒症潜在的诊断和预后的生物标志物.方法 本研究采用T-UCRs芯片技术分析了15例尿毒症患者和15例健康对照组的外周血单个核细胞的超保守区域转录表达水平,并且用实时荧光定量PCR技术验证了尿毒症患者与健康对照组表达差异最显著的基因的表达水平.结果 对比尿毒症患者与健康对照组,通过芯片筛选出T UCRs发现21个T-UCRs表达水平上调,49个T-UCRs表达水平下调,在表达上调和下调的T-UCRs中,对应的超保守区域uc.458-表达水平最显著(P-2.47524E-24;FC=3118.5361),其相关基因为RBFOX2.结论 通过GO分析以及验证后发现uc.458-可能是尿毒症潜在的生物标志物,RBFOX2可能是尿毒症潜在的关键基因.     

Osteoclasts and a small population of peripheral blood cells share common surface antigens

Summary Several methods have been tried to identify mononuclear osteoclast precursors. We used a panel of 13 osteoclast-recognizing monoclonal antibodies (mabs) for the identification of osteoclast precursor cells from the bone, bone marrow, and peripheral blood of egg laying hens. Almost all mabs stained some mononuclear cells in the bone. Seven mabs recognized few mononuclear cells in the bone marrow and five mabs gave the positive immunofluorescence reaction in the white blood cell fraction. Possible immediate osteoclast precursor cells differing from osteoclasts in their densities were identified in the bone. Three mabs (K38, K52, and K70) stained the same amount of mononuclear cells (2.6–3.4%) enriched in Percoll density centrifugation. Of the monoclonal antibodies that recognized few cells in blood, K41 stained only osteoclasts. K47 and K52 also recognized some mononuclear cells in the bone marrow. Other monoclonal antibodies K51 and K70 were more unspecific, since they stained cells derived from other tissues. Blood cells detected with these different monoclonal antibodies were negative for tartrate-resistant acid phosphatase (TRAP). On the basis of our results, we suggest that there is in the blood a specific TRAP-negative cell population, which is a good candidate for osteoclast precursor.     

肝癌组织、外周血细胞磷脂酰肌醇蛋白聚糖3 mRNA表达的意义

目的探讨磷脂酰肌醇蛋白聚糖3(GPC3)mRNA表达对肝细胞肝癌(HCC)及其血液微转移的诊断价值。方法参照甲胎蛋白(AFP)mRNA,半定量与巢式逆转录聚合酶链反应(RT-PCR)检测41例HCC、41例癌旁组织、52例非瘤肝组织(41例远离癌灶组织与11例正常肝组织)及67例外周血的GPC3表达。结果半定量PCR显示组织与外周血细胞全部存在GPC3 mRNA表达,HCC、癌旁、非瘤肝组织的表达量分别为78.9±35.5、30.6±21.6、23.8±15.5;而AFP mRNA的表达量依次为61.2±32.6、31.5±23.6、21.2±15.9。各基因在HCC与其他组织中的表达量差异具有统计学意义(P<0.01),以各基因在非瘤肝的相对表达量上限(x-+1.96s)为界,HCC中GPC3 mRNA和AFP mRNA的高表达分别占80.5%与63.4%;至少一个基因高表达HCC达总数的92.7%,与AFPmRNA单独检测差异有统计学意义(P<0.01)。临床病理特征分析发现AFP mRNA与HCC分级及血清AFP浓度有关,GPC3 mRNA表达与HCC分级及侵袭性有关。67例外周血GPC3 mRNA的表达量为15.9±9.0,各类组织中GPC3 mRNA表达均强于相应的外周血表达(P<0.01)。外周血GPC3mRNA表达量在HCC组、非HCC组分别为16.1±8.3、15.6±10.2,组间差异无统计学意义;在HCC转移组、HCC无转移组的表达量各为16.0±9.0、16.3±7.7,组间差异无统计学意义。巢式PCR显示,AFP mRNA表达比率在HCC组与非HCC组各为56.1%、23.1%,组间差异有统计学意义(P=0.011);HCC转移组与无明显转移组各为80.9%、30.0%,组间差异有统计学意义(P=0.002)。结论GPC3 mRNA是潜在的HCC组织诊断指标,与AFP mRNA有明显的互补作用,两者联合检测可提高HCC的阳性检出率;外周血存在GPC3 mRNA的广泛表达,但其与HCC的转移复发无关,不能用于HCC微转移的检测。     

布鲁菌病患者外周血NK细胞和T淋巴细胞亚群的变化

目的了解布鲁菌病患者性别、年龄、职业分布特征及并发症,外周血NK细胞和T淋巴细胞亚群的变化及意义。方法对101例布鲁菌病患者采用流行病学调查法进行调查,采用流式细胞仪抗体双表法检测38例布鲁菌病和非布鲁菌病发热患者外周血NK细胞（CD16^＋CD56^＋）、淋巴细胞绝对值（CD45^＋）、T淋巴细胞绝对值（CD3^＋）、T辅助细胞绝对值（CD3^＋CD4^＋）、T抑制/细胞毒细胞绝对值（CD3^＋CD8^＋）、B淋巴细胞绝对值（CD19^＋）、CD4/CD8比值,并进行分析。结果布鲁菌病患者以男性居多,中老年患者居多,发病人群主要为农牧民及从事防疫工作者,并发症以肝功能损害最多;布鲁菌病患者外周血NK绝对值显著低于非布鲁菌病发热患者组,具有统计学意义（t=-2.58,P〈0.05）。结论布鲁菌病以男性中老年患者居多,以农牧民和防疫员为主,布鲁菌病患者外周血NK细胞常受损伤。     

二期梅毒患者外周血单个核细胞基因表达谱研究

目的:分析二期梅毒患者外周血单个核细胞基因表达谱特征,探索机体抗梅毒免疫的分子机制。方法:采用全基因组高通量的Illumina测序技术,对4例二期梅毒患者和4例健康对照者外周血单个核细胞行数字基因转录组分析。后行实时PCR对其结果进行验证。结果:与健康对照者相比较,二期梅毒患者外周血单个核细胞共发现差异表达基因78个,其中有16个免疫应答相关基因。肿瘤坏死因子超家族成员17(TNFRSF17)、IL-17C、IL-21、IL-31受体A(IL-31RA)、C-X-C配体10(CXCL10)、趋化因子配体1(CCL1)等炎症因子和相关受体、CD4+T细胞激活标志CD38、Fc类吞噬性受体(FcγR1A、FcγR3B)以及补体(C2、SERPING1)等均显著上调。结论:二期梅毒患者多种天然免疫和适应性免疫分子参与机体系统性抗梅毒应答。     

脐血CD34+细胞表面粘附因子受体和趋化因子受体表达的检测

目的　检测脐血CD34 细胞表面粘附因子受体和趋化因子受体 (CXCR)表达情况。方法　取 15份新鲜和冻存脐血 ,采用流式细胞术检测脐血中CD34 细胞上CD6 2L、CD2 9及CXCR4等受体的表达情况。结果　 15份脐血CD34 细胞中CD6 2L表达率为 (74.0 7± 2 3.42 ) % ,CD2 9表达率为 (93.45± 10 .35 ) % ,CXCR4表达率为 (5 5 .73± 17.73) % ;三者平均表达强度分别为为 178.78±76 .5 9,15 6 .17± 6 4.6 1,5 6 .82± 2 6 .90。结论　与其它文献报道比较 ,脐血与骨髓、外周血CD34 细胞上述受体表达存在显著差异 ,这可能是导致脐血移植造血恢复慢的原因     

肝衰竭患者外周血单核细胞Toll样受体表达与炎性细胞因子关系的研究

目的探讨肝衰竭患者外周血单核细胞（PBMC）Toll样受体2（TLR2）和TLR4表达的变化及其在肝衰竭发病机制中的作用。方法采用RT—PCR法检测健康对照组（20名）、慢性乙型肝炎组（CHB,20例）、肝衰竭早期组（18例）和肝衰竭中晚期组（14例）患者PBMC中TLR2 mRNA和TLR4 mRNA的表达。ELISA法检测患者血中内毒素、TNF-α和IL-6的水平。结果与健康对照组比较,CHB组、肝衰竭早期组和中晚期组患者PBMC中的TLR2 mRNA、TLR4 mRNA,以及血中内毒素、TNF-α和IL-6水平有所上升,差异具有统计学意义（F值分别为32．997、37．476、23．951、57．265和38．403,P值均〈0．01）。TLR2 mRNA的表达在肝衰竭中晚期组中高于早期组,而TLR4 mRNA的表达则相反。在肝衰竭早期和中晚期,TLR2 mRNA的表达与TNF-α、IL-6均有明显相关性（FTNF为0．865和0．921,rIL-6为0．762和0．851,P值均〈0．01）,TLR4 mRNA在肝衰竭早期与TNF—α和IL-6具有相关性（r值分别为0．917和0．788,P值均〈0．01）。结论TLR2和TLR4介导的炎性反应可能参与了肝衰竭的发病机制。     

维生素D对复发性流产患者外周血自然杀伤细胞的调节作用

目的探讨维生素D对复发性流产(recurrent miscarriage,RM)患者外周血自然杀伤细胞(natural killer cells,NK)的调节作用。方法收集99例RM患者黄体中期的外周血,流式细胞术检测外周血NK细胞的数量和细胞毒性。根据患者体内维生素D水平将其分为三组:维生素D正常组(vitamin D normal,VDN)、维生素D不足组(vitamin D insufficiency,VDI)和维生素D缺乏组(vitamin D deficiency,VDD)。对维生素D缺乏和不足的RM患者补充1,25(OH)2D,分析维生素D补充对RM患者的NK细胞的调节作用。结果 CD3~-CD56~+NK细胞比例在三组间无显著差异(P0.05)。VDI组中NK细胞毒性在效靶比(effector-to-target ratio,E:T)为50∶1(51.6±8.8 vs.45.3±12.4)(P0.05)、25∶1(43.2±9.1 vs.35.8±11.8)(P0.01)和12.5∶1(30.2±10.3 vs.23.0±10.6)(P0.01)这三个条件下均显著高于VDN组。补充维生素D后,可显著降低E:T为50∶1(41.6±14.0 vs.51.0±10.5)(P0.01)、25∶1(34.9±14.3 vs.43.1±10.8)(P0.01)和12.5∶1(22.6±11.4 vs.29.9±10.7)(P0.01)的NK细胞毒性。结论低水平维生素D的RM患者外周血NK细胞毒性增加,而补充1,25(OH)2D可在一定程度上降调NK细胞毒性。