Immunoregulatory subpopulations of T-lymphocytes and their distribution in the peripheral blood, lymph nodes and spleen of patients with lymphogranulomatosis

Monoclonal antibodies OKT 3, OKT 4 and OKT 8 were used in 20 patients with Hodgkin's disease in order to characterize lymphocyte subpopulations in the peripheral blood, lymph nodes and spleen. As compared to the normal donors' peripheral blood and other parameters, the content of the OKT 3+, OKT 4+ cells in the involved lymph nodes and in the splenocyte suspension turned out to be high.     

Lymphocyte subsets in human peripheral blood after splenectomy and autotransplantation of splenic tissue

Leucocyte and lymphocyte subpopulations and serum immunoglobulins were studied in 14 patients who underwent splenectomy for trauma, nine patients with an autologous replant of splenic tissue after traumatic rupture of the spleen, and 23 age- and sex-matched healthy individuals. Buffy coat cells were stained by direct or indirect immunofluorescence with goat antihuman F(ab')2 immunoglobulin (B cells), Leu-1 (T cells), Leu-2a (T suppressor cells), and Leu-3a (T helper cells). Leucocytes and lymphocyte subsets were analyzed in a cytofluorograf . After splenectomy, patients showed a marked increase in both monocytes (135%) and lymphocytes (137%), with a fourfold rise in B cells, a nearly twofold rise in T cells, a reduced T/B cell ratio, and an unchanged T helper/T suppressor cell ratio. The number of B cells was significantly related to the serum immunoglobulin M level (r = 0.807, p less than 0.001). Patients with an autologous replant had moderate monocytosis (35%) and lymphocytosis (42%), with a selective rise of B cells (60%) and T suppressor cells (75%), which led to a 50% reduction of the T helper/T suppressor cell ratio. The changes observed after splenectomy indicate the major role of the spleen in cellular homing. In patients with an autologous replant , there is a tendency for normal storage of most cells, but an altered contribution of T suppressor cells to the circulating pool.     

Antibody formation by single cells from lymph nodes and efferent lymph of sheep

The antibody-forming cells which appear in the popliteal lymph node and efferent lymph of the sheep following immunization with boiled Salmonella have been studied by light and electron microscopy. Cells were incubated in monolayers with target erythrocytes sensitized with bacterial lipopolysaccharide. Three types of interaction between a proportion of the lymph cells and the erythrocytes surrounding them have been shown to indicate antibody formation: plaque-formation, immuno-cyto-adherence, and localized agglutination. At the peak of the response, 4 days after antigenic stimulation approximately 1 cell in every 200 from lymph node suspensions produces detectable specific antibody, while up to 1 cell in 20 in the lymph is active. For light microscope examination, individual antibody-forming cells were smeared in serum and stained with Leishman''s stain. For electron microscopy, a number of active cells were clumped with antiserum to form a specimen of convenient size, then sectioned. Most of the active cells from efferent lymph are large and basophilic, while a small proportion are blastlike. These cells contain abundant free ribosomes and very little endoplasmic reticulum. In the node only, an additional class of antibody-forming plasma cells is found which have considerable amounts of endoplasmic reticulum in their cytoplasm.     

Distribution of T cell subsets in human lymph nodes

A series of T cell-specific monoclonal antibodies was used to determine the location of T lymphocyte subpopulations in frozen sections of human lymph nodes by means of an immunoperoxidase technique. The majority of cells in the paracortical regions were reactive with anti-T1 and anti-T3 antibodies, which define all mature peripheral T cells. In contrast, the majority of cells within primary follicles were unreactive with anti-T1 and anti-T3 antibodies, but were reactive with anti-Ia and anti-IgM antibodies. In addition, a substantial number of T1+, T3+ cells were found in the germinal centers of secondary follicles on the capsular side. The vast majority of T1+, T3+ cells in the paracortex and the follicles were reactive with anti-T4 antibody, which defines inducer/helper T cells. Only a minority of cells in these areas were reactive with anti-T5 and anti-T8 antibodies, which define cytotoxic/suppressor cells. No lymphocytes were stained with anti-T6 antibody, which reacts with a majority of thymocytes but not with peripheral T cells. Scattered cells in the paracortex showed staining for Ia antigen in an irregular dendritic pattern. The findings demonstrate that the major T cell population found within human lymph node bears the mature T1+, T3+, T4+ phenotype characteristic of inducer T cells. Moreover, the location of this population indicates that they play a role in the induction of B cell differentiation in vivo.     

Nodal lymph flow quantified with afferent vessel input function allows differentiation between normal and cancer-bearing nodes

Morbidity and complexity involved in lymph node staging via surgical resection and biopsy could ideally be improved using node assay techniques that are non-invasive. While visible blue dyes are often used to locate the sentinel lymph nodes from draining lymphatic vessels near a tumor, they do not provide an in situ metric to evaluate presence of cancer. In this study, the transport kinetics of methylene blue were analyzed to determine the potential for better in situ information about metastatic involvement in the nodes. A rat model with cancer cells in the axillary lymph nodes was used, with methylene blue injection to image the fluorescence kinetics. The lymphatic flow from injection sites to nodes was imaged and the relative kinetics from feeding lymphatic ducts relative to lymph nodes was quantified. Large variability existed in raw fluorescence and transport patterns within each cohort resulting in no systematic difference between average nodal uptake in normal, sham control and cancer-bearing nodes. However, when the signal from the afferent lymph vessel fluorescence was used to normalize the signal of the lymph nodes, the high signal heterogeneity was reduced. Using a model, the lymph flow through the nodes (F_LN) was estimated to be 1.49 ± 0.64 ml/g/min in normal nodes, 1.53 ± 0.45 ml/g/min in sham control nodes, and reduced to 0.50 ± 0.24 ml/g/min in cancer-cell injected nodes. This summarizes the significant difference (p = 0.0002) between cancer-free and cancer-bearing nodes in normalized flow. This process of normalized flow imaging could be used as an in situ tool to detect metastatic involvement in nodes.OCIS codes: (170.0170) Medical optics and biotechnology, (170.2655) Functional monitoring and imaging     

Lymphocyte subsets and mitogen induced proliferation in adjuvant arthritis. Part II. Changes in function and numbers of lymph nodes T lymphocytes.

To evaluate the effects of lymphocytes on the pathogenesis of adjuvant arthritis (AA), the lymphocytic responses to phytohemagglutinin (PHA) and concanavalin A (Con A) and the percentages of T cell subpopulations were investigated in the inguinal lymph nodes of Long-Evans (LE) rats and Sprague-Dawley (SD) rats after a single injection of complete Freund's adjuvant. The response of LE to PHA increased to its maximum on the 14th day and correlated with the severity of AA. The responses of LE and SD rats to Con A were decreased on the 7th, 11th, and 14th days. Additionally, percentage of suppressor T cells of LE rats was lower than that of SD rats. Significant increases of percentage of helper T cells and of helper/suppressor T cell ratio were observed in LE rats, indicating a decrease or defect in the suppressor T cells in LE rats. In LE, we postulated that helper T cells are involved in the induction of AA, and a defect in the suppressor T cell function or numbers would subject LE to the development of AA. SD rats, on the other hand, with higher suppressor T cell numbers, were less susceptible to AA.     

Immunoreactivity of lymphocytes from draining lymph nodes, peripheral blood and tumor infiltrates from oral cancer patients

Lymphocytes from metastatic (met) and nonmetastatic (non-met) regional lymph nodes, LNL peripheral blood (PBL) and tumor infiltrating lymphocytes (TIL) of patients with squamous cell carcinoma of the oral cavity and healthy donors were investigated for CD3, CD4, CD8 and HNK-1 phenotypes, Natural Killer (NK) cell Activity, Antibody Dependent Cellular Cytotoxicity (ADCC) and proliferative response to mitogen (PHA). Modulation of NK cytotoxicity with recombinant Interferon alpha (IFN-alpha) was also investigated in some cases. Lymphocytes from met and non-met lymph nodes showed no variation in the percentages of CD3+, CD4+ and CD8+ cells, when compared with each other and with PBL of oral cancer patients. TIL showed significantly less proportion of CD3+ and CD4+ cells. The percentage of HNK-1+ cells was significantly lower in LNL and TIL when compared to PBL of oral cancer patients. The mitogen responses of met and non-met LNL were comparable to each other and better than that of PBL from the same patients, while, TIL showed significant impairment in mitogen responses. The NK cytotoxicity and ADCC of PBL from oral cancer patients were comparable to healthy donors which could be augmented by rIFN alpha. LNL and TIL showed almost negligible NK and ADCC activities and NK activity could not be modulated by rIFN alpha. The results thus demonstrate that in oral cancer patients, lymphocytes from three compartments viz. PBL, LNL and TIL showed differential effector functions. The metastatic status of LN did not affect the immunoreactivity of LNL.     

沈阳市某学校教师的亚健康情况及外周血淋巴细胞亚群分析

目的统计沈阳市某学校教师的亚健康情况和分析亚健康状态时外周血淋巴细胞亚群的变化。方法采用现场调查的方法,对参加体检的教师发放亚健康调查表,判断教师健康状况。同时,检测健康教师和亚健康教师的淋巴细胞亚群。同期住院肿瘤患者淋巴细胞亚群结果作为对照。结果共有584名教师问卷合格,其中健康人群300例,患病需治疗的9例,亚健康状态人群275例,亚健康发生率为47.09%,健康教师淋巴细胞亚群分析中,总T细胞、辅助性T细胞(Th)、抑制性T细胞(Ts)、自然杀伤细胞(NK)和B细胞比例分别为(67.19±8.95)%、(37.58±7.23)%、(28.14±7.66)%、(21.16±9.97)%、(9.56±3.80)%。亚健康职工上述细胞比例分别为(65.22±7.75)%、(35.90±7.22)%、(28.87±8.02)%、(24.57±8.33)%、(9.61±4.66)%。同年度肿瘤患者上述细胞比例分别为(60.65±14.39)%、(30.32±15.22)%、(26.36±9.51)%、(30.25±13.36)%、(5.37±4.29%)。亚健康组与健康组比较总T细胞、Th、NK细胞比例差异均有统计学意义(P0.05),Ts及B细胞比例无明显差异。亚健康组与肿瘤组比较总T细胞、Th、Ts、NK细胞及B细胞比例差异均有统计学意义(P0.05),健康组与肿瘤组比较总T细胞、Th、Ts、NK细胞及B细胞比例差异均有统计学意义(P0.05)。结论教师亚健康状态的临床表现多样,其健康状况不容乐观。     

肝病患者的肝组织单细胞悬液与外周血中淋巴细胞亚群的分析比较

目的探讨肝脏组织单细胞悬液的制备方法,了解慢性肝病患者的肝组织和外周血的淋巴细胞亚群差异。方法分别选择慢性乙型肝炎(CHB)14例、自身免疫性肝炎(AIH)4例和原发性肝细胞癌(HCC)3例,在超声引导下经皮作肝脏穿刺活检,用单细胞制备仪或物理研磨方法将肝组织制备成单细胞悬液,肝组织中淋巴细胞经4种特异性荧光抗体标记,在流式细胞仪(FCM)上作四色荧光分析,并与外周血测定结果比较。结果单细胞制备仪和物理研磨方法制备的肝组织单细胞悬液,细胞成活率分别为92.4%和91.5%;检测肝组织单细胞悬液中淋巴细胞亚群,正常对照组和慢性肝病各组间差异均有显著性(P<0.05),AIH组和CHB组间差异有显著性(P<0.05)。而外周血中淋巴细胞亚群检测,HCC组和CHB组间差异有显著性(P<0.05)。肝组织和外周血相应项目比较差异均有显著性(P<0.05)。结论用2种方法制备单细胞悬液能满足FCM检测的特殊需要;肝脏组织中淋巴细胞与外周血的淋巴细胞亚群比例差异有显著性。     

肝病患者的肝组织单细胞悬液与外周血中淋巴细胞亚群的分析比较

目的 探讨肝脏组织单细胞悬液的制备方法,了解慢性肝病患者的肝组织和外周血的淋巴细胞亚群差异.方法 分别选择慢性乙型肝炎（CHB）14例、自身免疫性肝炎（AIH）4例和原发性肝细胞癌（HCC）3例,在超声引导下经皮作肝脏穿刺活检,用单细胞制备仪或物理研磨方法将肝组织制备成单细胞悬液,肝组织中淋巴细胞经4种特异性荧光抗体标记,在流式细胞仪（FCM）上作四色荧光分析,并与外周血测定结果比较.结果 单细胞制备仪和物理研磨方法制备的肝组织单细胞悬液,细胞成活率分别为92.4%和91.5%;检测肝组织单细胞悬液中淋巴细胞亚群,正常对照组和慢性肝病各组间差异均有显著性（P〈0.05）,AIH组和CHB组间差异有显著性（P〈0.05）.而外周血中淋巴细胞亚群检测,HCC组和CHB组间差异有显著性（P〈0.05）.肝组织和外周血相应项目比较差异均有显著性（P〈0.05）.结论 用2种方法制备单细胞悬液能满足FCM检测的特殊需要;肝脏组织中淋巴细胞与外周血的淋巴细胞亚群比例差异有显著性.     

肺癌患者外周血、淋巴结β1,4-N-乙酰氨基半乳糖转移酶mRNA的检测及其临床意义

目的:通过研究分子标志物β1,4-N-乙酰氨基半乳糖转移酶(GalNAc T)mRNA在肺癌患者外周血及淋巴结的表达,探讨肺癌微转移的临床意义。方法:应用巢式RT-PCR技术检测35例Ⅰ、Ⅱ期肺癌患者标本(癌组织、淋巴结及外周血)和12例肺部良性病变患者手术标本及10例来自健康志愿者血液标本中GalNAc T mRNA的表达。结果:GalNAc T mRNA在35例Ⅰ期、Ⅱ期肺癌病人中22例(63%)为阳性,外周血标本中8例(23%)为阳性,43个肺门及纵隔淋巴结中21个(49%)为阳性。而在12例良性肺部肿瘤及10例健康志愿者外周血中均为阴性。结论:GalNAc T mRNA可作为RT-PCR法检测肺癌患者外周血微转移的分子标志物,有助于早期诊断肺癌转移,为制定治疗方案和评估预后提供重要参考依据。