A low blood lymphocyte count is associated with an expansion of activated cytotoxic lymphocytes in patients with B-cell chronic lymphocytic leukaemia

Abstract: In order to determine the relationships between CD2+ lymphocyte subpopulations and tumour mass, the immunophenotype of natural killer (NK) cells and T lymphocyte subsets was studied in 56 B-chronic lymphocytic leukaemia (B-CLL) patients and 38 healthy subjects. The patients were classified according to their blood lymphocyte count (BLC). Forty patients had BLC<30×10⁹/l (low BLC, less tumour mass) and 16 patients had BLC>30×10⁹/l (high BLC, larger tumour mass). The percentage of CD3^– CD56+ cells, as well as of CD8+, CD8+CD45RO+ and CD3+CD57+ T subsets in low BLC patients, were higher than those found in high BLC patients. Conversely, the percentages of CD3+HLA DR+, CD4+ and CD4+CD45RO+ lymphocytes were higher in high BLC patients than in low BLC patients. The CD4/CD8 ratio was decreased in low BLC patients while it was increased in high BLC patients and a significant positive correlation was found between their CD4/CD8 ratio and their BLC. We conclude that in low BLC B-CLL patients there is a decreased percentage of activated helper lymphocytes and an increased percentage of NK cells and activated cytotoxic T lymphocytes. These results suggest a role for NK cells, and helper and cytotoxic T lymphocytes in the control of tumour burden in B-CLL patients.     

T-cell activation and subset patterns are altered in B-CLL and correlate with the stage of the disease

Two-color FACS analysis was used to study activated and "functional" T and natural killer (NK) cell subsets of circulating lymphocytes in 23 patients with B-type chronic lymphocytic leukemia (B-CLL) and in 30 healthy subjects. As compared with controls, B-CLL patients had increased absolute numbers of phenotypically activated, HLA-DR+ CD4+ and CD8+ cells and T suppressor/effector (CD11b+CD8+) cells. When patients in Rai stages II through IV (n = 11) were compared with cases in Rai stages O through I (n = 12), the former group of patients had higher numbers of activated CD4+ and CD8+ T cells and decreased levels of suppressor/effector T cells. The absolute numbers of T suppressor/inducer (CD45R+CD4+) cells were elevated in patients with stage O through I disease but within normal range in stage II through IV leukemia. We further showed that the absolute numbers of NK-like (CD16+) cells and their activated counterparts (DR+CD16+) are elevated in B-CLL patients as compared with healthy subjects. The comparison of relative T and NK subsets in the blood of patients and controls showed that the proportions of CD4+, CD8+, and CD16+ cells expressing the activation marker HLA-DR were increased in B-CLL. Furthermore, the percentage of T-suppressor/inducer (CD45R+) cells within the CD4+ population was decreased in the patients. The proportion of T- suppressor/effector (CD11b+) cells within the CD8+ subset was reduced in subjects with stage II-IV disease only. When stimulated in vitro with the T-cell-dependent inducer TPA, B-CLL cells from patients in Rai stages II through IV secreted larger amounts of IgM as compared with cells from stage O through I patients. A positive correlation was observed between the degree of phenotypic activation of CD4+ T-helper cells and their functional capacity to augment IgM secretion by autologous B-CLL cells. Our findings indicate a tumor cell-directed regulatory role of T cells (and possibly NK cells as well) in B-CLL. Furthermore, monitoring of phenotypically activated and functional T- cell subsets may be helpful in the prediction of disease progression and timing of therapy in B-CLL.     

梅毒与梅毒合并病毒性肝炎患者外周血T淋巴细胞亚群变化*

目的 调查梅毒及其合并病毒性肝炎患者外周血T淋巴细胞亚群的变化。方法 2016年1月～2020年6月我院收治的93例感染苍白螺旋体(TP)梅毒患者中,单纯梅毒感染61例,TP合并CHB患者21例和TP合并CHC患者11例,另选择健康体检者84例。使用流式细胞仪检测外周血T淋巴细胞亚群。结果 梅毒患者外周血CD3⁺、CD4⁺、CD4⁺CD45RO⁺和CD8⁺CD45RA⁺细胞百分比及CD4⁺/CD8⁺细胞比值分别为(52.2±8.5)%、(40.3±5.7)%、(18.1±3.9)%、(12.4±3.7)%和(1.2±0.3),均显著低于健康人[分别为(69.1±7.6)%、(50.7±6.9)%、(20.6±4.7)%、(16.2±4.3)%和(1.9±0.5),P＜0.05],而外周血CD8⁺、CD4⁺CD45RA⁺和CD8⁺CD45RO⁺细胞百分比显著高于健康人[分别为(32.4±7.3)%、(24.7±6.5)%和(8.7±1.5)%对(26.2±5.4)%、(21.8±6.2)%和(5.4±1.1)%,P＜0.05];三组外周血CD8⁺、CD4⁺CD45RA⁺、CD4⁺CD45RO⁺、CD8⁺CD45RA⁺和CD8⁺CD45RO⁺细胞百分比及CD4⁺/CD8⁺细胞比值比较,差异有统计学意义(P＜0.05),TP合并CHB组和TP合并CHC组患者外周血CD8⁺、CD4⁺CD45RA⁺和CD8⁺CD45RO⁺细胞百分比均显著高于TP组(P＜0.05),而TP合并CHB组和TP合并CHC组患者外周血CD4⁺/CD8⁺比值、CD4⁺CD45RO⁺和CD8⁺CD45RA⁺细胞百分比显著低于TP组(P＜0.05),TP合并CHB组与TP合并CHC组患者外周血CD8⁺、CD4⁺CD45RA⁺、CD4⁺CD45RO⁺、CD8⁺CD45RA⁺和CD8⁺CD45RO⁺细胞百分比及CD4⁺/CD8⁺细胞比值比较均无统计学差异(P＞0.05)。结论 梅毒患者存在显著的外周血淋巴细胞亚群变化,合并CHB或合并CHC患者细胞免疫功能变化更明显,其临床意义值得进一步探讨。     

Altered pattern of cytokine production by peripheral blood CD2+ cells from B chronic lymphocytic leukemia patients

To determine if activation-induced cytokine production is altered in CD2+ lymphocytes from B-CLL patients, cytokine levels were determined by ELISA in supernatants of PHA-stimulated cultures of CD2+ cells from 33 B-CLL patients and 22 healthy controls. The production of Interferon γ (IFN-γ) and Tumor Necrosis Factor (TNF-α) by mitogen-activated CD2+ lymphocytes from B-CLL patients was higher than that found in healthy controls, while no differences were found in TNF-β production. IFN-γ and TNF-α levels determined at 72 h in PHA-stimulated CD2+ cell cultures from B-CLL patients statistically correlated with the percentages of CD3+CD45RO+ and CD3−CD56+ lymphocytes, respectively. Although there were differences in the production kinetics of interleukins (ILs) 2 and 4 between B-CLL patients and the healthy controls, no differences were found at the time when the levels of both interleukins peak. The production of both IFN-γ and IL-4 by PHA-stimulated CD2+ lymphocytes from non-smouldering B-CLL patients was significantly higher than that from smouldering B-CLL patients while no significant differences were found in the production of IL-2, TNF-α, and TNF-β between the two B-CLL patient groups. These data suggest that functional alterations in the production of cytokines by CD2+ cells from B-CLL patients could help to explain the expansion of leukemic cells in B-CLL patients. Am. J. Hematol. 57:93–100, 1998. © 1998 Wiley-Liss, Inc.     

The effect of hemodialysis on the virgin CD45RA+, CD45RO- and memory CD45RO+, CD45RA- lymphocyte count in patients with chronic renal failure

The hallmark of immunological memory is a quick and effective response to a repeated antigen exposure. Virgin lymphocytes, with their surface receptors CD45RA+, CD45RO- are produced in primary lymphatic organs, then migrating to secondary lymphatic structures. Memory lymphocytes CD45RO+, CD45RA- produced in these organs migrate to non-lymphatic organs--a possible location of inflammatory process, thus enabling the immunological system to eliminate effectively the same antigen, when repeatedly present. The aim of the study was 1) to test the influence of hemodialysis on the number of virgin lymphocytes and/or memory lymphocytes; 2) whether such impact (if any) depends on the type of dialysis membrane used (cuprophan or polysulphon), 3) if the effect is different in patients with or without diabetes. Overall number of virgin T helper lymphocytes CD45RA+CD4+ was significantly lower in patients with end-stage renal disease, while the number of total CD45RO+, CD45RA- memory lymphocytes was significantly greater among patients with diabetic nephropathy, compared to normal control subjects. After 15 minutes of hemodialysis, number of virgin lymphocytes CD45RA+, CD45RO- (p < 0.001, p < 0.01) and their subclasses, as well as memory lymphocytes CD45RO+, CD45RA- were significantly decreased. After 15 minutes of hemodialysis with polysulphon membrane, the decrease in T virgin cytotoxic, B virgin CD45RA+CD4-, T memory cytotoxic as well as B memory CD45RO+CD4- lymphocytes was significantly lower, when compared with cuprophan membrane (p < 0.02). Among patients treated with cuprophan hemodialysis, the decrease of T helper memory CD45RO+CD4+ lymphocytes was significantly lower in patients with diabetic nephropathy, than in non-diabetic patients. CONCLUSIONS: In all patients with end-stage renal disease, the impact of hemodialysis on the number of memory lymphocytes CD45RO+, CD45RA-, as well as virgin lymphocytes CD45RA+, CD45RO- was shown, but the effect was less profound during hemodialysis with polysulphon membrane, compared to cuprophan. The presence of diabetic nephropathy effects the hemodialysis-induced changes in the number of T memory helper CD45RO+ CD45RO+CD4+ lymphocytes, with no impact on other subclasses of the examined cells.     

中国乙型肝炎患者外周血淋巴细胞亚群频率参考值范围

目的 调查中国乙型肝炎患者外周血淋巴细胞亚群频率参考值范围.方法 利用流式细胞术检测2846例乙型肝炎患者和117例健康人群外周血淋巴细胞亚群数值,调查我国健康人群和乙型肝炎人群的参考值范围.结果 调查了16～60岁健康人群和HBV感染相关的急性肝炎、慢性肝炎、重型肝炎和肝硬化人群外周血CD3+T淋巴细胞、CD3+CD...     

Phenotypic and clonal analysis of T lymphocytes in childhood immune thrombocytopenic purpura

In an attempt to identify and characterize T-lymphocyte immunoregulatory abnormalities in immune thrombocytopenic purpura (ITP), we have performed phenotypic and clonal analysis on peripheral T lymphocytes from 23 children with ITP. Quantitation of lymphocyte subpopulations showed that children with acute ITP had higher numbers of CD45RA+ and lower numbers of CD45RO+ T cells than children with chronic ITP or controls, but these differences may be age related. Analysis of T-cell receptor variable beta gene usage identified 2 boys with chronic ITP and elevated numbers of V beta 8+ T cells. Eight T- cell clones were established (6 CD4+, 4B4+ helper-inducer lines and 2 CD8+ lines) that showed in vitro proliferation against allogeneic platelets. The addition of autologous antigen-presenting cells enhanced the proliferation of six clones, but not for two clones that coexpressed natural killer (NK) markers. Four of seven positive clones also had measurable interleukin (IL)-2 secretion following platelet stimulation, providing further evidence for T-cell reactivity. Our results provide the first evidence that patients with ITP may have platelet-reactive T lymphocytes identifiable at the clonal level, supporting the hypothesis that autoreactive peripheral T lymphocytes may mediate or participate in the pathogenesis of this disorder.     

Analysis of bone marrow and peripheral blood immunoregulatory lymphocytes in patients with myelodysplastic syndrome

The cell surface phenotype of immunoregulatory lymphocytes in bone marrow (BM) and peripheral blood (PB) in myelodysplastic syndrome (MDS), a stem cell disorder, was analyzed. Mononuclear cells from 25 patients with refractory anemia (RA) and nine with RA with an excess of blasts (RAEB) were characterized by two-color flow cytometry using various monoclonal antibodies. No significant change of CD3+, CD4+, and CD8+ cells in PB, but a decrease of the percent of positive cells for CD8+ among the total lymphocyte (%CD8+ +) was noticed in RA patients. On the other hand, in BM of RA patients, a decrease in the number of CD4+cells, but not CD8+ +cells, was noted. In RAEB patients, the absolute numbers of CD3+, CD4+, CD8+, and CD8+ +cells in BM were decreased; however, the ratio of these lymphocytes was not changed. No change was observed among the CD4 + subsets in PB of RA or RAEB patients. In BM, a decrease in percentage of CD4+ CD45RA+ (% CD4+ CD45RA+; naive cell) and increases in CD4+ CD45RO+ (% CD4+ CD45RO+; memory cell) and CD4+ CD29+ (%CD4+ CD29+; helper/inducer) among CD4+ cells were found in both RA and RAEB patients. Analysis of the CD8+ + subset showed an increased number of CD8+ + CD11a+ cells (activated CTL) in both BM and PB of RA patients, but not of RAEB patients. Furthermore, increments in CD56+ and CD16+ cells among CD3- cells (natural killer; NK cells) were seen in RA patients but not in RAEB patients. It remains unclear whether lymphocytes in MDS patients were involved in the abnormal (MDS) clones, but our results regarding the increments of CD8+ + CD11a+ and NK cells in RA patients suggest that the mechanism of immune surveillance against the abnormal MDS clones was activated in these RA patients, but not in RAEB patients. Further investigation is required to clarify the functions of these immunoregulatory lymphocytes in MDS patients.     

Flow cytometry of blood and bone marrow cells from patients with hairy cell leukemia: phenotype of hairy cells and lymphocyte subsets after treatment with 2-chlorodeoxyadenosine

By flow cytometry and an extensive set of markers, we characterized leukemic cells from the blood and bone marrow of 68 symptomatic patients with hairy cell leukemia (HCL). Hairy cells identified in the large cell gate always expressed CD19, CD20, HLA-DR, CD45RA, and B-ly 7. Other markers were occasionally expressed, such as CD38, CD45RO, CD23, CD15, CD4, CD5, and CD10 (expressed on more than 20% of the hairy cells in 44%, 25%, 21%, 18%, 12%, 10%, and 5% of evaluated cases, respectively). During treatment with 2-chlorodeoxyadenosine (CdA), the median lymphocyte counts decreased from 2,000/microL to 300/microL. Flow cytometry was repeated at the nadir (n = 24) of lymphocyte counts, at 3 months (n = 46), at 6 months (n = 50), at 1 year (n = 39), and at 2 years (n = 12) after treatment. The initial decrease of CD8+ and CD20+ cells was greater than that of CD4+ and natural killer (NK) cells, leading to an increasing CD4/CD8 ratio. Median nadir values of CD4+, CD8+, CD20+, and NK cells were 128/microL, 78/microL, 10/microL, and 13/microL, respectively. The subsequent recovery was quicker for CD8+ and NK cells, leading to a normalization within 3 months, whereas CD20+ and CD4+ cells required 1 or 2 years to enter the normal range. The CD4/CD8 ratio thus decreased after the nadir and remained less than 1. CD45RA+ CD4 cells and CD45RA+/CD45RO+ double-positive cells were less affected by CdA. Activated T cells, ie, HLA-DR+ cells, rarely decreased below the normal range and often recovered with an overshoot. CD10+ cells increased in the bone marrow posttreatment as an indication of normal B-cell regeneration in 16 of 36 (44%) patients. The quick regeneration of certain lymphoid subsets might explain the lack of late infections in CdA-treated HCL patients.     

Lymphocyte subset reconstitution after HLA-identical placental blood transplantation (PBT) or PBT plus bone marrow transplantation (BMT) in three children with beta-thalassemia major

The kinetics of circulating lymphoid cells were evaluated in three children suffering from beta-thalassemia major after HLA-identical sibling placental blood transplant (PBT) in one patient and placental blood plus bone marrow transplantation (BMT) in two patients. Recovery of the main lymphocyte subsets, as determined by phenotype analysis of circulating PBMCs, was complete within 2 months after transplant. NK (CD56+) cells were the first to appear in peripheral blood, followed by T (CD3+, CD2+, CD7+) and B (CD19+) cells. Of the T lymphocytes, the CD8+ were the first to reconstitute, but recovery of CD4+ cells was also rapid and within 6 months these T cells reached normal values. The expression of CD57 by NK or T cells was slightly delayed. The evaluation of RA and RO isoform expression of the CD45 molecule showed a prevalence of the CD45RA antigen with a ratio of 2-3:1. In the PBT only patient, T cells expressing the CD45RO antigen prevailed in the early post-transplant period. Severe or chronic GVHD was not observed. This experience demonstrates that reconstitution of lymphocyte subsets is successful in genetic hematological diseases after transplantation of HLA-identical placental blood or placental blood plus bone marrow from healthy or heterozygous siblings. Bone Marrow Transplantation (2000) 26, 743-747.     

Peripheral blood lymphocyte subsets in polymyalgia rheumatica

Summary Peripheral blood lymphocytes from 23 patients with polymyalgia rheumatica (PMR) were characterized using monoclonal antibodies and flow cytometry in a two-year prospective study. There were no significant differences in absolute numbers or relative percentages of lymphocytes or CD3+, CD4+, CD8+ T cells or the CD4+T cell functional subsets, virgin (CD4+CD45RA+) and memory (CD4+CD29+) T cells, in patients before or during corticosteroid treatment compared to controls. Previous reports on decreased levels of CD8+T cells as a characteristic of PMR/giant cell arteritis was not confirmed. The absolute number and relative percentage of lymphocytes with natural killer cell activity, CD16+ CD56+ cells, were significantly lower in patients with active untreated PMR as well as during corticosteriod treatment compared to controls, but at the two-year follow-up the difference was less marked.     

Interleukin-10 expression: is there a neglected contribution of CD8+ T cells in rheumatoid arthritis joints?

OBJECTIVE: To search for RA specific processes among T cell accumulation, T cell activation, or cytokine expression in CD4+ and CD8+ synovial fluid (SF) T cells. METHODS: Flow cytometry of CD4+, CD8+, CD45RA+, CD45RO+, CD69 double or triple stained peripheral blood (PB) and SF T cells. IL-2, IL-10, and IFN-gamma expression was determined in PMA + ionomycin stimulated T cells on the single cell level. Concentrations of secreted IL-2, IL-4, IL-10, and IFN-gamma were quantified in the sera and synovial fluids by enzyme linked immunosorbent assay (ELISA). RESULTS: A preferential recruitment of CD45RO+ memory T cells was found for CD4+ helper T cells, and in similar also for CD8+ suppressor T cells. An elevated CD69 expression was detected in memory, but also in CD45RA+ naive CD4+ and CD8+ SF T cells, whilst IL-2 expression was only demonstrable in a minor proportion of T cells populations. Preferential recruitment of memory T cells, but incomplete activation of naive and memory, CD4+ and CD8+ T cells were in similar found in RA and control patients. In RA but not in the control patients, a relevant proportion of CD4+ and CD8+ PB and SF T cells expressed IL-10 and IFN-gamma. High concentrations of IL-10, that were correlated with the amounts of secreted TNF-alpha, were only detected in RA joints. CONCLUSION: Memory and naive T cell state of CD4+ and CD8+ T cell accumulates in the joints, and early T cell activation occur in similar patterns in RA and control patients. High IL-10 SF concentrations in contrast, and elevated percentages of IFN-gamma and IL-10 expressing CD4+ and CD8+ T cells in the PB and SF were characteristic for RA. Here, CD8+ T cells may contribute to high IL-10 concentrations in RA joints.     

抗病毒治疗对HIV/AIDS患者自然杀伤细胞和γδT细胞的重建作用研究

目的 研究高效联合抗逆转录病毒治疗(HAART)对HIV/AIDS患者NK细胞和γδT细胞的重建作用,比较HAART对获得性免疫和固有免疫重建规律的异同.方法 对59例接受规律HAART的HIV/AIDS患者于治疗前、治疗后1、3、6、9、12个月末进行随访,采用分支DNA(bDNA)法检测血浆病毒载量,经流式细胞仪检测患者外周血淋巴细胞亚群.以56例健康献血员作为正常对照.结果 基线时HIV/AIDS患者CD4+T细胞中位数为45(164,13)个/μl,NK细胞和γδT细胞中位数分别为96(135,56)个/μl和30(45,19)个/μl,与正常对照组相比均显著减低(P<0.001).HAART开始后1个月,血浆病毒载量降低了2.33 lg拷贝/ml(P<0.001),CD4+T细胞数增长了111个/μl(P<0.001),主要以记忆CD4+T细胞的快速增长为主,而NK细胞和γδT细胞与基线相比无显著变化.在HAART后3个月NK细胞和.γδT细胞数量出现增长,分别较基线增加26个/μl(P=0.007)和15个/μl(P<0.001).治疗12个月,CD4+T细胞、NK细胞及γδT细胞计数分别达到271(365,134)个/μl、166(281,108)个/μl和46(66,34)个/μl,但仍低于正常对照.结论 HAART可以重建HIV/AIDS患者固有免疫,但NK细胞和γδT细胞数量的增加晚于CD4+ T细胞,HAART对固有免疫的重建作用机制有待进一步研究.     

Phenotypic characteristics of dissociated mononuclear cells from rheumatoid synovial membrane

The phenotypic characteristics of enzymatically dissociated synovial membrane mononuclear cells from 8 patients (14 samples) with rheumatoid arthritis (RA) were assessed by fluorescence activated flow cytometry and compared to peripheral blood (PB) mononuclear cells from 18 patients with RA and 14 normal controls. There was no significant difference between the percentage of CD4+ and CD8+ lymphocytes in synovial membrane compared to RA and normal PB. Double labelling experiments revealed similar percentages of CD4+ CDw29+ (helper-inducer) and CD4+ CD45R+ (suppressor-inducer) cells in RA and normal PB. In contrast, SM CD4+ CDw29+ cells were present in significantly higher proportions than in RA and normal PB (p less than 0.001). Conversely, synovial membrane CD4+ CD45R+ cells were present in significantly lower proportions than in RA and normal PB (p less than 0.001). A similar pattern of CDw29 and CD45R antigen expression was noted on CD8+ lymphocytes reflecting increased killer-effector (p less than 0.001) and decreased suppressor-effector (p less than 0.001) cells, respectively. Other experiments revealed a significant increase in the percentage of synovial membrane CD20+ cells (B lymphocytes) and HLA-DR+ cells compared to RA PB (p less than 0.02 and p less than 0.001) and normal PB (p less than 0.01 and p less than 0.005), and similar proportions of CD14+ cells (monocytes/macrophages). Our results suggest that RA synovial membrane contains populations of T and B lymphocytes that differ quantitatively and qualitatively from those in PB. These may account for some of the abnormalities in intraarticular humoral and cellular immune responses in patients with RA.     

Circulating activated T cell subsets in autoimmune thyroid diseases: differences between untreated and treated patients.

To investigate the relationships between lymphocyte subsets and thyroid function, peripheral blood lymphocytes were analysed with cell surface antigens of activated (HLA-DR+) T, helper T (CD4+ 2H4-, CD4+ 4B4+) and suppressor-inducer T (CD4+ 2H4+, CD4+ 4B4-) cells subsets in 56 patients with Graves' disease, 16 patients with Hashimoto's thyroiditis, 7 patients with typical subacute thyroiditis and 2 patients with the thyrotoxic phase of autoimmune thyroiditis. Both patients with Graves' disease and Hashimoto's thyroiditis had increased percentages of HLA-DR+ T (Ia+ CD3+) cells as well as HLA-DR+ helper-inducer T (Ia+ CD4+) cells, which seemed to be independent of treatments. The percentage of HLA-DR+ suppressor-cytotoxic T (Ia+ CD8+) cells was increased in euthyroid or hypothyroid patients with Graves' disease following treatment, but was normal in hyperthyroid patients. The percentages of Ia+ CD4+ cells and Ia+ CD8+ were also increased in patients with thyroiditis, whereas these abnormal values normalized in the remission phase. These findings suggest that an increase in Ia+ CD4+ cells characteristically occurs during immune system activation in patients with hyperthyroid Graves' disease, Hashimoto's thyroiditis and the thyrotoxic phase of subacute thyroiditis, whereas the activated CD8+ cells in Graves' disease are induced by antithyroidal therapy.     

Persistence of replication-competent HIV in both memory and naive CD4 T cell subsets in patients on prolonged and effective HAART

OBJECTIVE: To investigate the phenotypic features of infected lymphocytes in patients on prolonged and effective highly active antiretroviral therapy (HAART). DESIGN: We examined highly purified subsets of memory and naive CD4 T lymphocytes for the presence of replication-competent virus. METHODS: In 11 highly selected HAART-treated patients, we isolated highly purified CD45RO CD45RA CD4 T cells using a magnetic bead-based procedure. In some patients, a subsequent cell separation according to CD62L expression was performed. We quantified total viral DNA in freshly isolated T-cell subsets. To verify whether the virus was replication-competent, HIV RNA was measured in supernatants following cell activation. RESULTS: HIV DNA was detectable in the CD45RO and CD45RA CD4 T-cell subsets in 100% and 90% of the patients tested, respectively. In central memory CD45ROCD62L, effector memory CD45RO+CD62L-, truly naive CD45RACD62L, and CD45RA+CD62L- CD4 T cells, HIV DNA was found in 100%, 55%, 88%, and 50% of the patients tested respectively. HIV DNA was significantly higher in the CD45RO fraction than in the CD45RA subset and in the CD45ROCD62L fraction than in the three other CD45RA/ROCD62L+/- subsets. Detectable HIV RNA was found in the culture supernatants of CD45RO and CD45RA CD4 T-cell subsets in 80% and 66% of the patients tested respectively, and in CD45ROCD62L, CD45RO+CD62L-, CD45RACD62L, and CD45RA+CD62L- CD4 T cells in 100%, 100%, 100% and 50% of the patients tested respectively. CONCLUSIONS: In patients on prolonged and effective HAART, the pool of infected CD4 T lymphocytes consists predominantly of memory cells but also contains naive cells.     

Distinct characteristics of lymphokine-activated killer (LAK) cells derived from patients with B-cell chronic lymphocytic leukemia (B-CLL). A factor in B-CLL serum promotes natural killer cell-like LAK cell growth

We show that lymphokine-activated killer (LAK) cell precursors derived from patients with B-cell chronic lymphocytic leukemia (B-CLL) and cultured in the presence of recombinant interleukin-2 and normal human serum (NHS), develop into primarily NK cell-like (CD 57+) LAK cells, whereas identically prepared LAK cell precursors from normal subjects develop into mainly T cell-like (CD 3+, CD 8+) LAK cells. B-CLL LAK cells exhibited greater proliferative capacity than did normal LAK cells. When normal LAK cells were grown in B-CLL serum instead of NHS, their proliferation increased; NK cell levels also increased to those found in B-CLL LAK cells, suggesting that B-CLL serum contains a factor that promotes NK cell-like growth, LAK cells derived from normal or B-CLL patients demonstrated similar lytic activity toward K562 and Raji cells. Growth in B-CLL serum did not reduce their lytic potential. Thus, the altered phenotype and growth exhibited by B-CLL LAK cells and normal LAK cells grown in B-CLL serum does not lead to abnormalities in their cytolytic functions. We propose instead that the predominance of NK-like cells in B-CLL LAK cell populations and the presence of an NK cell-like growth factor in B-CLL serum reflect abnormalities related to NK cell-mediated B-cell regulation; ie, either inhibition of normal B-cell growth and/or growth stimulation of the leukemic clone in B-CLL.     

Recovery of lymphocyte and dendritic cell subsets after autologous CD34+ cell transplantation

Following high-dose chemotherapy (HDC) and peripheral blood progenitor cell transplantation (PBPCT), there are profound changes in leukocyte homeostasis and the immune system is compromised. Transplantation of purified CD34+ cells may further compromise immune recovery because the grafts are depleted of mature immune cells. However, a detailed monitoring of immune cell reconstitution has not been done. We monitored blood levels of antigen-presenting cells (APC) and of lymphocytes by multi-color flow cytometry at different times post CD34+ PBPCT. We found a rapid normalization of circulating levels of the antigen-presenting CD11c+ dendritic cells (defined as lineage- HLA-DR+ CD11c+ cells). There was a slight over-representation of lin- DR+ CD11c- cells at day 42 post transplantation suggesting that the composition of the APC population might be affected. Normal levels of total T, B and NK lymphocytes were rapidly achieved but the composition of the T cell population was abnormal. Patients had elevated levels of CD8+ T cells at early times and a persistent reduction in levels of naive CD8+ T cells (CD8+ CD4- CD45RA+ CD27+) and of naive CD4+ T cells (CD4+CD3+ CD8- CD45RA+). Thus, we found a rapid recovery of DC after CD34+ PBPCT but the specific numerical defects in naive T cells are likely to be a major cause of immune dysfunction in the patients. Bone Marrow Transplantation (2000) 25, 1249-1255.     

Lymphocyte subpopulations in peripheral blood in primary sclerosing cholangitis

BACKGROUND/AIMS: Primary sclerosing cholangitis (PSC) is a chronic disease of autoimmunological etiology, leading to inflammation, destruction and atrophy of the bile ducts. The aim of the study was to determine peripheral lymphocyte B, T, and NK cells in PSC. METHODOLOGY: The estimation of peripheral blood lymphocytes in 17 patients (54+/-12 years old) with PSC was carried out; the control group consisted of 27 subjects (38+/-11 years). The following T lymphocyte subpopulations were determined using duo color flow cytometry: CD3+, CD4+, CD8+, CD3++HLA DR+, B cells CD19+, and NK cells CD16+ +CD56+. RESULTS: In PSC we observed doubled increase in activated T lymphocytes of CD3+ +HLA DR+ phenotype as compared to healthy subjects (7.9% vs. 2.7%, p<0.01) and NK cells (12.6% vs. 10.3%, respectively, p<0.05). There were no significant differences in the composition of CD3+, CD4+, and CD8+. In peripheral blood we noted, in patients with PSC, a decrease in B lymphocytes (11.2% vs. 12.3%, p<0.19). CONCLUSIONS: The examinations showed that activated T (HLA DR+) lymphocytes and NK cells played an important role in development of PSC.     

CD7- T cells in rheumatoid arthritis.

OBJECTIVE. Rheumatoid arthritis (RA) is characterized by decreased expression of CD7 in the peripheral blood and in the synovium. The present study was designed to identify the basis for and functional consequences of this decreased expression. METHODS. Peripheral blood lymphocytes from normal controls and from patients with RA or systemic lupus erythematosus (SLE), and T cell lines derived from rheumatoid synovium, were evaluated using 3-color fluorescence-activated cell sorter analysis. RESULTS. Normal subjects and most SLE patients expressed homogeneous, bright CD7 on CD4+, CD45RA+ cells, whereas RA patients demonstrated a significantly increased proportion of CD7- cells. T cell lines derived from rheumatoid synovium demonstrated a striking deficiency of CD7 on CD4+, CD45RA- cells. CD4+, CD45RA+ cells from RA patients changed phenotype after in vitro activation to CD45RA negativity, with up-regulation of CD7. CD7-, CD4+, CD45RA- cells were assessed for their ability to induce pokeweed mitogen-driven IgM and IgM-rheumatoid factor synthesis, and they were found to be potent helper/inducer cells. An increased population of CD7-, CD4+ cells in peripheral blood was found to predict a low response to recall antigens. CONCLUSION. The low expression of CD7 in RA may explain some of the immune abnormalities which may contribute to the pathogenesis of this disease.