对过继免疫治疗中外周血单个核细胞采集术的探讨

目的:探讨在恶性肿瘤患者过继免疫治疗中外周血单个核细胞的采集技术.方法:根据患者外周血单个核细胞计数值,应用CS-3000PLUS血细胞分离机,采集外周血单个核细胞,运用Ficoll法分离纯化,再用贴壁法分离出贴壁细胞进行诱导分化为树突状细胞(DC),未贴壁细胞(主要为淋巴细胞)诱导分化为肿瘤杀伤细胞(CIK)或体外扩增为记忆性淋巴细胞(ALT),最后进行细胞回输.结果:在8例13人次的采集中,处理总循环血量,获得单个核细胞总数,单个核细胞纯度,细胞采集效率,血小板丢失率分别为(6.99±1.74)L,(3.47±1.44)×109/L,(90±6)%、(5L 3±13)%、(43.5±13.1)%.DC加CIK治疗1例,CK治疗3例,化疗后ALT免疫重建治疗4例,均取得了一定的临床疗效.结论:外周血单个核细胞采集术,不良反应发生率低、单个核细胞的采集效率高,用于恶性肿瘤过继免疫治疗的临床效果较好.但采集后血小板的丢失严重,对于恶性肿瘤化疗后血小板减低的患者应引起关注.     

血细胞分离机采集正常献血者外周血单个核细胞前后电解质的变化

目的:比较正常献血者利用血细胞分离机行外周血单个核细胞采集前、采集后及采后2周电解质指标的变化。方法:利用Amicus、com.tec、MCS+、COBE Spectra、Spectra Optia 5种血细胞分离机分别采集12例固定献血者外周血单个核细胞,单采前后15 min采集外周血液标本,检测电解质结果,单采后2周采集血液标本行电解质检测,观察这3次血液电解质的变化情况。结果:单采后献血者血K~+、血Cl~-与单采前比较明显下降(P0.05),而血Na~+在单采前后无明显变化(P0.05);采后2周血K~+、血Cl~-与采集前比较无明显变化(P0.05)。结论:采集单个核细胞后献血者血K~+、血Cl~-有一过性降低,需要在采集前进行适当干预或者在采集后进行治疗。     

糖尿病对骨髓干细胞动员效果的影响

目的 探讨糖尿病(DM)与骨髓干细胞动员效果的相互影响.方法将实验动物新西兰兔分为健康组(A组)、DM组(B组)、未成模组(C组),三组在骨髓干细胞动员前检测外周血单个核细胞(MNC),动员第6天检测外周血及骨髓MNC及粒-单核细胞集落形成单位(CFU-CM)计数.B组在动员前1 d及动员第6天监测血红蛋白(Hb)、血小板(PLT)、空腹血糖(FPG)及血液流变学指标.结果 三组动员前后外周血、骨髓MNC及CFU-GM计数比较均无统计学差异(P均0.05).B组动员前后Hb、PLT、FPC、血浆黏度及红细胞聚集性变化均无统计学差异(P均>0.05),动员后全血黏度显著高于动员前(P<0.05).结论 DM对骨髓干细胞动员效果无影响.用G-CSF行骨髓干细胞动员对DM兔的Hb、PLT、FPG无影响,但对血液流变学有影响,提示临床应予重视并干预.     

直肠癌患者外周血单核细胞诱导为树突细胞

目的 用直肠癌患者外周血单核细胞通过体外培养的方法诱导为成熟树突细胞.方法 CS-3000血细胞分离机采集直肠癌患者外周血单核细胞悬液,Ficoll分离单个核细胞,用含10%FBS的RPMI-1640培养基培养,第0天加入GM-CSF 1 000 U/ml、IL-4 500 U/ml,第4天2/3量换液,补足GM-CSF、白介素(IL)-4,并加入肿瘤坏死因子(TNF)-α 500 U/ml诱导其成熟,以后每隔3天2/3量换液.结果 培养至第11天,收集培养的1×105～2×105个树突细胞经流式细胞仪检测表达CD83+(52.9±5.8)%,CD86+(79.2±8.1)%,HLA-DR(65.8±5.7)%,CD1a(51.3±6.4)%,及CD+14(4.72±2.16)%.结论 直肠癌患者外周血单核细胞经GM-CSF、IL-4及TNF-α联合培养能诱导出成熟树突细胞.为直肠癌的免疫治疗提供了临床应用基础.     

自体外周血干细胞回输出现过敏性休克一例

患者 ,男性 ,5 0岁。 2 0 0 1年 7月确诊为 :多发性骨髓瘤 (κ轻链型 )ⅢA期。MOD方案化疗一个疗程后达完全缓解。后予MOD方案巩固 2个疗程 ,骨髓象及外周血象持续缓解。 2 0 0 1年 12月应用环磷酰胺行外周血干细胞动员 (CTX2 g/m2 × 2d) ,2 0 0 2年 1月应用美国MCS +Plans血细胞分离机采集。处理全血 92 0 2ml ,PBSC 70ml。采集单个核细胞数 1.43 7× 10 9/L ,CD3 4 7.81% ,CD3 85 2 .9% ,CD3 4 +/CD3 8 0 .96% ,CD3 4 +/CD3 8+5 1.9%。采集过程顺利 ,病人无不适。采集物应用 5 %二甲亚砜 +6%羟乙基淀粉 +人血清蛋白 (上海中…     

伴海洋性贫血供者外周血造血干细胞采集效果分析

目的观察海洋性贫血供者外周血造血干细胞(PBSC)采集的效果。方法将2008年1月至2010年1月在中山大学附属第一医院血液内科接受PBSC采集的23例HLA全相合亲缘供者分为对照组(不伴海洋性贫血)15例,观察组(伴海洋性贫血)8例。比较两组供者PBSC动员后、采集前的血常规以及单次采集产品中血常规指标以及单个核细胞(MNC)计数和CD34+细胞计数,并观察两组在PBSC回输后造血重建情况。结果两组供者动员后、采集前的血常规中,WBC平均计数和MNC平均计数,比较差异均无统计学意义(P0.05);但在单次采集后,观察组RBC计数和HCT分别为(1.19±0.40)×1012/L和(0.084±0.032),明显高于对照组(0.79±0.17)×1012/L和(0.059±0.016)(P0.05),而WBC计数为(125.2±53.1)×109/L、MNC计数为(3.12±1.72)×108/kg和CD34+细胞平均计数为(1.25±0.69)×106/kg,则分别低于对照组的(220.4±25.1)×109/L、(5.87±1.36)×108/kg和3.52±0.82×106/kg(P0.05);观察组共行采集术13次、对照组供者各行1次采集。观察组MNC计数及CD34+细胞分别为(5.36±0.80)×108/kg和(2.14±0.32)×106/kg,与对照组比较差异均无统计学意义[(6.31±1.37)×108/kg和(3.79±0.82)×106/kg];PBSC回输后,观察组受者中性粒细胞和血小板的造血重建时间与对照组差异无统计学意义(P0.05)。结论海洋性贫血供者PBSC动员虽不受影响,但单次采集的PBSC产品中红细胞污染较重、MNC和CD34+细胞收获率降低,但适当增加采集次数后,其PBSC数量仍可达到移植要求,且不影响受者的造血重建。     

血细胞分析仪实验数据比对分析

目的 评价现用不同型号的血细胞分析仪在日常工作中检测结果的准确性及一致性.方法 以本科室Gen-s System2血细胞分析仪为比对仪器,选择高、中、低值三个档次的新鲜全血标本40份,每台仪器平行测定2次,取平均值进行多个参数分析,以Gen-s Systern2评价其他血细胞分析仪检测结果(WBC、RBC、HGB、PLT、HCT).结果 (1)4台血细胞分析仪的精密度好;(2)WBC、RBC、HGB、PLT、HCT五个指标分别进行配对t检验,均P＞0.05,差异无统计学意义;(3)各台仪器与比对仪器参数的相关系数r＞0.975,相关性好;(4)由回归曲线与回归方程可主要了解各台仪器系统误差的大小.结论 (1)应用新鲜全血对血细胞分析仪进行比对试验.能及时发现仪器间的系统误差;(2)同一实验室用新鲜全血对血细胞分析仪进行定期对比实验及校正,能提高和保证同一实验室血细胞分析仪检测结果的准确度和一致性.     

外周血单个核细胞miR-214与急性心肌梗死患者免疫趋化因子表达的相关性

目的探讨急性心肌梗死(AMI)患者外周血单个核细胞miR-214水平变化以及与免疫趋化因子表达的相关性。方法前瞻性选择2019年1月至2019年12月因急性胸闷胸痛于宜昌市第一人民医院(三峡大学人民医院)心血管内科就诊患者174例,其中排除冠状动脉粥样硬化性心脏病(冠心病)者45例(CAG正常组)和AMI患者129例(AMI组)。分别采用实时定量荧光PCR法、ELISA法和Western blot法检测两组外周血单个核细胞mi R-214、血浆单核细胞趋化蛋白-1(MCP-1)和外周血单个核细胞趋化因子功能性受体2(CCR2)水平,并采用Pearson法分析其相关性。结果 AMI组患者外周血单个核细胞miR-214表达量低于CAG正常组[(1.00±0.13)vs.(0.81±0.28),P0.05],同时血浆MCP-1浓度[(97.85±23.86)pg/ml vs.(133.90±47.48)pg/ml]和外周血单个核细胞CCR2蛋白水平[(0.48±0.16)vs.(0.67±0.29)]高于CAG正常组(P0.05)。经Pearson相关性分析,AMI组患者外周血单个核细胞miR-214水平和血浆MCP-1浓度、外周血单个核细胞CCR2蛋白水平均呈负相关性(r=-0.590,-0.433,P0.05)。随着冠状动脉SYNTAX评分增加,患者外周血单个核细胞miR-214水平降低,同时外周血单个核细胞CCR2蛋白水平和血浆MCP-1浓度逐渐升高(P0.05);经多重线性回归分析,miR-214、MCP-1及CCR2与SYNTAX评分均存在线性回归关系(P0.05)。结论 AMI患者外周单个核细胞miR-214表达量降低,且与MCP-1/CCR2传导通路的激活存在负向调控关系,可能是加重AMI疾病进展的重要机制之一。     

伴全血细胞减少的巨幼细胞贫血临床特点分析

目的 探讨伴全血细胞减少的巨幼细胞贫血（MA）患者的临床特点.方法 分析28例伴全血细胞减少的MA患者的临床特点并与34例单纯贫血者比较.观察28例伴全血细胞减少者叶酸及维生素B12治疗期间网织红细胞比率（Ret）、中性粒细胞（NEUT）及血小板计数（PLT）变化.结果 与单纯贫血比较,伴全血细胞减少的MA患者Hb水平更低,病程及住院时间更长（P均＜0.05）.伴全血细胞减少者治疗5d时Ret升高达峰值,9d时明显下降,但仍高于治疗前.NEUT及PLT恢复时间分别为（9.27±2.00）d、（7.82±1.16）d,差异无统计学意义.结论 伴全血细胞减少的MA病情相对更重；Hb为评价MA病情的主要指标,治疗期间NEUT与PLT恢复时间差异无统计学意义.     

造血系统恶性肿瘤自体外周血干细胞动员采集和冻存研究

目的:探讨造血系统恶性肿瘤自体外周血造血干细胞移植(APBSCT)后造血系统重建与采集和冻存自体的外周血干细胞(APBSC)数量及质量的关系。方法:用大剂量化疗加粒细胞集落刺激因子(G-CSF)方法对拟行APBSCT的18例造血系统恶性肿瘤进行动员,CS-3000PLUS血细胞分离机采集APBSC,经程序降温仪处理保存于-196℃液氮中,37~40℃水浴解冻后迅速回输,检测冻存前及解冻后APBSC的锥虫蓝染色拒染率、单个核细胞(MNC)计数、粒-单集落形成单位(CFU-GM)集落数及CD34 细胞百分率。结果:APBSC的采集时间为化疗后12.6(9~19)d,采集次数2.4(1~3)次,MNC采集率为(138.6±52.32)%。解冻后MNC、CD34 及CFU-GM的回收率分别为(91.96±1.37)%,(85.94±0.64)%,(87.69±4.53)%。冻存前及解冻后APBSC的锥虫蓝染色拒染率分别为(96.26±1.33)%,(92.75±2.04)%,二者差别无统计学意义(P>0.05)。骨髓瘤患者CFU-GM集落数、MNC采集率及CD34 细胞百分率较低;化疗疗程>10次的5例患者MNC采集率较低及CFU-GM生长不良,其中3例出现APBSCT后造血重建延迟。结论:rhG-CSF与大剂量化疗联合的动员方案可缩短采集时间,提高MNC采集率;APBSC的质量和数量有显著的个体差异,与疾病的类型和化疗方案有密切关系,移植前化疗次数增多可影响APBSC的数量和质量,导致APBSCT后造血重建延迟。     

Association of periodontitis with increased white blood cell count and blood pressure

This study was aimed to examine the association of periodontitis with white blood cell (WBC) count and blood pressure (BP). In 2002, 424 subjects (manufacturing workers) were investigated for periodontitis by a general dentist. All were Japanese. Among them, 364 subjects (269 men and 95 women) who also attended the next year's (2003) screening were enrolled for this study. Of the 364 subjects, 55 (15.1%) had periodontitis. We also measured the BP and WBC count in periodontitis and non-periodontitis subjects at baseline and 1-year later follow-up. The WBC count higher in subjects with periodontitis than in subjects without periodontitis, both at baseline [mean +/- standard error (SE) 6.6 x 10(3) +/- 0.2 x 10(3)/ml vs 5.8 +/- 0.3 x 10(3)/ml; p < 0.001] and follow-up (7.0 +/- 0.3(3)/ml vs 6.5 +/- 0.1(3)/ml; p = 0.003). The systolic BP was higher in subjects with periodontitis than in subjects without periodontitis, both at the baseline (128 +/- 2.1 mmHg vs 120.8 +/- 0.8 mmHg; p < 0.001) and follow-up (129.2 +/- 2.3 mmHg vs 123.0 +/- 0.8 mmHg; p = 0.011), and so was the diastolic BP both at baseline (76 +/- 1.5 mmHg vs 71.2 +/- 0.6 mmHg; p = 0.003) and follow-up (80.5 +/- 1.7 mmHg vs 75.4 +/- 0.7 mmHg; p = 0.004). Periodontitis is associated with increased BP and WBC count. This finding may provide one underlying pathway linking periodontitis and cardiovascular disease.     

Changes in the blood cell counts with aging

We analyzed the blood cell counts and serum levels of total protein (TP), total cholesterol (TC) and triglyceride (TG) of 2,231 healthy subjects (1,295 men and 936 women) between age 20 and 99 years in order to clarify the following two subjects. (1) In the approximately 10 years since the report of Shirakura et al in 1978, eating habits have improved and the average life expectancy has extended in Japan. Is there any effect of such betterment on blood cell counts of the aged? (2) It has been pointed out that quality of everyday life, such as staying at home but not in an old-age home, working, traveling, and so forth, had an influence on the blood cell counts of aged. Is there any difference between the blood cell counts of people under 60 years and those of people older than 60 years who have a good quality of life as mentioned above? The hemoglobin concentration, red blood cell count, and hematocrit value began to decrease in men in their sixth decade and in women in their seventh decade and the change was more prominent with advancing age, especially in men. The white blood cell count and platelet count tended to decrease with advancing age. The serum levels of TP, TC, and TG also declined with age in those over 60 years of age. These results confirmed that the hemoglobin concentration, red blood cell count, and hematocrit value decrease in the elderly subjects as they grow older and it may be considered that reduced ingestion of protein is one of the causes of the phenomenon.     

DNA array analysis for red blood cell antigens facilitates the transfusion support with antigen-matched blood in patients with sickle cell disease

Background  Blood samples from patients with sickle cell disease (SCD) present to transfusion service with numerous antibodies, making the searching for compatible red blood cells (RBC) a challenge. To overcome this problem we developed an effective strategy to meet needs of supplying RBC-compatible units to SCD patients using DNA arrays.
Methods  We selected DNA samples from 144 SCD patients with multiple (receiving > 5 units) transfusions previously phenotyped for ABO, Rh(D, C, c, E, e), K1, Fy^a and Jk^a. We also selected DNA samples from 948 Brazilian blood donors whose ABO/RhD phenotype matched that of the patients. All samples were analysed by DNA array analysis (HEA Beadchip^TM, Bioarray Solutions) to determine polymorphisms associated with antigen expression for 11 blood group systems (Rh, Kell, Kidd, Duffy, MNS, Dombrock, Lutheran, Landsteiner-Wiener, Diego, Colton, Scianna); and one mutation associated with haemoglobinopathies.
Results  Based on genotype results we were able to predict phenotype-compatible donors needed in order to provide compatible units to this group of patients. Based on their ABO/Rh phenotype we were able to find in this pool of donors compatible units for 134 SCD patients.
Conclusion  Blood group genotyping by DNA array contributes to the management of transfusions in SCD patients by facilitating the transfusion support with antigen-matched blood. It has the potential to improve the life of thousands of SCD-transfused patients by reducing mortality due to transfusion reactions and immunization.     

Association of metabolic syndrome with white blood cell subtype and red blood cells

Inflammation and thrombogenesis have been suggested as possible causes for cardiovascular events in patients suffering from metabolic syndrome (MS). The primary objective of this study was to determine the relationship between red blood cell (RBC) or white blood cell (WBC) subtypes and MS. The secondary objective was to reveal any gender differences inherent to this association. Body mass index (BMI), blood pressure, serum high-density lipoprotein cholesterol, triglycerides, and glucose were measured. The numbers of WBC subtypes and RBCs were determined in healthy adults. In male subjects, the numbers of total leukocytes, neutrophils, and lymphocytes was elevated in the MS patients (P<0.05). In the male subjects, the numbers of total leukocytes, neutrophils, and lymphocytes were elevated in accordance with the metabolic component count (P<0.05). RBC, monocyte, eosinophil, and basophil counts did not differ in accordance with metabolic component counts (r = 0.406, r = 0.304, r = 0.366; P<0.05). In the female subjects, we determined there to be no differences in the numbers of RBC and WBC subtypes in the MS patients, in accordance with metabolic component counts. The numbers of total leukocytes, neutrophils, and lymphocytes were elevated in the male MS subjects in this study, and these counts increased in accordance with the metabolic component counts. In the female subjects in this study, we determined there to be no association between RBC and WBC subtype counts with MS.     

Trial of ABO and Rh blood typing with an automated blood cell counter

We designed a method for ABO and Rh typing using a blood cell counter. Blood specimens were diluted with physiological saline, mixed with an antiserum, and incubated. The cell count and size distribution curves were obtained by an automated blood cell counter. The samples with agglutination had a decreased cell count and a characteristic size distribution curve with a population of doublet cells on the right side. The results were correlated with the degree of agglutination observed by the manual method, used as a reference. It should be possible to adapt blood cell counters for use in blood typing.     

Red blood cell function and blood storage

Red blood cells are ideal vehicles for delivering oxygen to tissues, but their functions deteriorate during liquid preservation. In this article, we review the role of red blood cells in oxygen delivery and methods to evaluate the effectiveness of red blood cell transfusion. Quantitative estimation of transfusion effects could avoid unnecessary transfusion and reduce the risk of transfusion-associated disorders. We also describe the benefits of transfusion of red blood cells having a higher oxygen-delivering capacity. Phosphoenolpyruvate is a promising component to prepare red blood cells having a higher oxygen-delivering capacity.     

Current issues with blood transfusions in sickle cell disease

With increased recognition of the profound morbidity of sickle cell disease and with growing evidence of the efficacy of transfusion therapy in prevention and treatment of sickle cell complications, most patients now receive intermittent transfusion therapy. The purpose of this report is to review blood component therapy and its risks for sickle cell patients. Packed red cells are the preferred blood component. Leukocyte-reduced units should be standard because of their beneficial effects in reducing alloimmunization, transfusion reactions, platelet refractoriness, and infection transmission. The use of washed, frozen, or irradiated units is limited to specific problems. Sickle trait-positive units function normally, but because of difficulties with calculating hemoglobin S percentages and leukocyte filters, they are not routinely used. Transfusion-acquired infections have shown a marked decrease but still present a major risk. Viral hepatitis transmission is currently low, but at least 10% of adult sickle cell patients are hepatitis C positive, and they often have liver damage. Although bacterial infections are rare, they account for 16% of transfusion-related fatalities. Patients who are iron overloaded are particularly vulnerable to Yersina enterocolitica. Red cell alloimmunization is a serious problem that could potentially affect 50% of transfused patients. However, preventive phenotypic matching for common antigens can minimize alloimmunization; limited matching for at least E, C, and K has become the standard of care. Recently, more patients are being identified who have developed red cell autoantibodies, which can mask alloantibodies and occasionally are hemolytic. Careful laboratory evaluation of all cases is essential. Transfusions also may trigger sickle cell events, including pain crises, stroke, and acute pulmonary deterioration. In part, these are induced by blood viscosity and increased blood pressure. Diuretic therapy and close monitoring of transfusion volume and vital signs can minimize these events. In summary, transfusion therapy carries risks, but the routine use of leukocyte-reduced, phenotypically matched units in conjunction with close monitoring of patients can make transfusion therapy safer.     

Rapid bedside rejuvenation of red blood cell with an autologous cell salvage device

Background and Objectives

During storage, red blood cells (RBCs) undergo physicochemical changes which affect the quality, function, and in vivo survival of transfused packed RBCs (pRBC). Changes include decreased 2,3‐diphosphoglycerate (2,3‐DPG) levels, decreased ATP, changes in mechanical properties and oxidative injury. RBC rejuvenation is a method used to increase levels of 2,3‐DPG and ATP in pRBCs. This process requires incubating the pRBCs with a rejuvenation solution and subsequent washing. Standard blood bank protocols using the COBE 2991 Cell Processor require several hours of preparation. The objective of this study was to verify if a bedside protocol for rejuvenating pRBC and washing with the Sorin Xtra autologous cell salvage system could be used.

Materials and Methods

Outdated pRBC units were obtained and rejuvenated in a model operating suite using a dry air incubator for 1 h at 37°C. Six units of pRBCs were pre‐diluted with saline (1000 ml) and six units were not pre‐diluted with saline. All units were washed with normal saline (1000 ml) using an apheresis‐design cell salvage device in manual mode and wash volume set to 3000 ml. Samples were collected and analyzed for standard RBC quality parameters at baseline and post‐wash.

Results

Total pRBC wash efficiency was 94% ± 12% at a final hematocrit of 67.7 ± 5.9% while maintaining post‐wash hemolysis 0.24 ± 0.12 %. Pre‐dilution prior to washing did not confer statistically significant differences in final RBC quality parameters with the notable exceptions of calculated hemolysis and supernatant potassium levels (P < 0.05). The washing process can be completed within 10 min. The post‐wash RBC parameters are appropriate for immediate transfusion to patients.