CD45RA expression by CD4 T lymphocytes in tumors invaded by B-cell non-Hodgkin's lymphoma (NHL) or Hodgkin's disease (HD).

Little is known about the role of tumor infiltrating T lymphocytes (TIL-T) in the pathogenesis of malignant diseases and collaboration between normal and malignant cells has not yet been proved. In the present work, we have investigated whether immune T lymphocytes exist in tumors invaded by B-cell non-Hodgkin's lymphoma (NHL) or Hodgkin's disease (HD). Therefore, we have studied the reactivity of the CD45RA monoclonal antibody, which discriminates between naive and memory CD4 T lymphocytes. Our results showed far lower percentages of CD4+ CD45RA+ in malignant lymphoma (30.3 +/- 15.0% in B-cell NHL, and 37.4 +/- 18.6% in HD) than in reactive hyperplasia (54.7 +/- 13.2%), leading to the conclusion of an accumulation of immune cells in tumor microenvironment. A further heterogeneity in the relative proportion of naive and memory TIL-T was also observed within lymphoma (range: 11 to 68% in B-cell NHL, 5 to 69% in HD). In B-cell NHL, it was related to histological features, as documented by the Kiel classification (P = .028), and to a stronger extent to cytological characteristics analysed with the Grenoble classification (P less than .0001): class 1 NHL, which are essentially indolent NHL displayed lower naive cells (22.2 +/- 7.4%) than class 3 NHL, which are more aggressive (40.1 +/- 16.1%). Among the monoclonal antibodies (mAb) defining the B-cell clone phenotype or activation state (CD19, CD20, CD21, CD22, CD23, CD24, CD5, CD10, CD11a, and Ki67), only CD23 (P = .0003) and Ki67 (P = .0007) revealed statistical association with the percentage of naive CD4 lymphocytes. No correlation could be demonstrated with the proportion of whole TIL-T, activated CD3 DR TIL-T, or CD4 subset.     

Poor lymphocyte recovery following CD34-selected autologous peripheral blood stem cell transplantation for non-Hodgkin's lymphoma

Positive selection of CD34+ cells in autologous grafts, designed to deplete tumour cells, also results in T-cell depletion. To assess the reconstitution of the different lymphocyte subsets and of the T-cell repertoire diversity following autologous transplantation of selected CD34+ peripheral blood stem cells (PBSC), we analysed sequential blood samples in eight patients autografted for advanced B-cell non-Hodgkin's lymphoma in a phase I-II pilot study. Although natural killer cell recovery was rapid, T- and B-cell recovery was delayed with a median of 110/microliters CD4+, 175/microliters CD8+ T cells and 45/microliters B cells at 12 months post-transplant. The naive CD45RA+ T-cell compartment was profoundly deficient up to 12 months for both CD4+ and CD8+ subsets. A transient expansion of memory CD8+CD45RO+ T cells consisting of an increased percentage of CD57+CD28- cells occurred within the first 3 months post-transplant, but the memory CD4+CD45RO+ T cells remained far below the normal value. The CD8+CD28+ T-cell subset did not recover. Using multiplex PCR analysis of the T-cell receptor gamma locus, we found that the repertoire diversity improved at 12 months after being poor and oligoclonal during the first 3 months post-transplant. As shown by monoplex PCRgamma analysis of every VJ combination, despite T-cell depletion of the graft, mature T cells were carried over with the selected CD34+ PBSC and contributed to the T-cell recovery after transplantation.     

Effect of didanosine, stavudine, and hydroxyurea therapy on apoptosis in CD45RA+ and CD45RO+ T lymphocyte subpopulations

The effect of aggressive antiretroviral therapy on spontaneous apoptosis (AP) in CD4+ and CD8+ lymphocytes expressing CD45RO (memory cells) and CD45RA (naive cells) and their relationship to cellular activation and viral load were examined. Ten patients receiving simultaneous treatment with d4T, ddI, and HU were evaluated. Flow cytometric analysis showed significant levels of AP (measured by TUNEL assay) among memory and naive T cells and an enhanced expression of CD38 and HLA-DR activation markers. The percentage of apoptotic CD4+CD45RO+ and CD4+CD45RA+ cells decreased, respectively, from 34 +/- 3.3 and 29 +/- 3.6 prior to treatment to 20.5 +/- 4 and 22 +/- 3.8 at week 8 into therapy. The percentage of apoptotic CD8+CD45RO+ and CD8+CD45RA+ cells similarly decreased, respectively, from 20 +/- 2.5 and 24 +/- 3 prior to treatment to 14.5 +/- 2.7 and 16 +/- 3 at week 8 into treatment. The percentage of CD4+ cells expressing the activation markers CD38 and HLA-DR decreased from 27 +/- 6 to 13 +/- 2 and from 26 +/- 4 to 13.5 +/- 3, respectively. The percentage of CD8+ cells expressing either CD38 or HLA-DR fell from 22 +/- 3 to 10 +/- 2 for the former and from 39 +/- 5 to 22 +/- 4 for the latter. This was associated with a significant decrease in viral load (mean, 1.4 log10), and a decline in circulating plasma TNF-alpha and sIL-2R levels from 50.5 +/- 10 to 21 +/- 6 and 92.5 +/- 11 to 68 +/- 9, respectively. These data indicate that short-term therapy with ddI, d4T, and HU in combination diminished AP, immune activation, and partially restored naive and memory T cell subpopulations.     

Ex vivo apoptosis,CD95 and CD28 expression in T cells of children with juvenile idiopathic arthritis

We hypothesise that T-cell apoptosis and the percentage of T cells expressing molecules involved in apoptosis modulation (CD95, CD28) are altered at the inflammation site and in peripheral blood (PB) of children with juvenile idiopathic arthritis (JIA). Paired JIA samples of synovial fluid (SF) and PB ( n=7) and PB samples from age-matched normal children ( n=23) were analysed immediately ex vivo. Apoptosis was measured by detection of phophatidylserine (PS) externalisation on T cells. CD95 or CD28 was detected by FACS, and soluble CD95 and CD95 ligand levels were detected by enzyme-linked immunosorbent assay (ELISA). In SF, the mean percentage of apoptotic T cells was somewhat higher than in PB. However, the percentages of T cells expressing CD95 and soluble CD95 levels were markedly higher in SF (CD4 cells 96+/-2%, CD8 91+/-6%, soluble CD95 6,420+/-2,571 pg/ml) than in PB (CD4 32+/-10%, CD8 36+/-9%, soluble CD95 4,296+/-2,142 pg/ml). Peripheral blood T-cell apoptosis in JIA (CD4 20+/-8%, CD8 42+/-19%) was higher than in the control group (CD4 5+/-2%, CD8 9+/-6%). Interestingly, the percentage of PB CD4 cells expressing CD28 was lower in JIA than controls. In conclusion, systemic T-cell apoptosis was higher in JIA while a substantial number of SF T cells survived locally, despite the fact that almost all cells express CD95.     

Epstein-barr virus-associated composite lymphoma composed of peripheral t-cell lymphoma and an anaplastic variant of a diffuse large b-cell type of non-hodgkin’s lymphoma and strongly expressing P53 protein

We report a case of composite lymphoma consisting of peripheral T-cell lymphoma and an anaplastic variant of diffuse large B-cell lymphoma (DLBCL) and associated with Epstein-Barr virus (EBV) infection and strong p53 expression. A 65-year-old Japanese woman developed fever and generalized lymphadenopathy. A biopsy of the cervical node revealed the morphology of malignant lymphoma with 2 kinds of lymphoma coexisting in 1 lymph node. One lymphoma type consisted of immunoblastic large cells with the T-cell marker phenotype CD3+, CD45RO/UCHL-1+, CD20/L26-, CD79-, CD10-, CD30-, and CD15-; the other type consisted of large cells with abundant cytoplasm and pleomorphic nuclei with the marker phenotype CD79+, CD20/L26+, CD45RO/UCHL-1-, CD3-, CD10-, CD30+, NPM/ALK-, and CD15-. Therefore, the diagnosis was composite lymphoma of peripheral T-cell lymphoma and an anaplastic variant of DLBCL, stage IVB, because the patient had bone marrow involvement with peripheral T-cell lymphoma. The biopsy led to findings of latent type II EBV-associated lymphoma in both the peripheral T-cell lymphoma and the anaplastic variant of DLBCL as the result of positive signals for EBV small RNAs by in situ hybridization, positive immunostaining results for EBV latent membrane protein 1 antibody, and negative immunostaining results for EBV nuclear antigen 2. Immunostaining of the mass with p53 antibody also yielded positive results for both types of lymphoma cells. This case suggests that the immunocompromised state of this patient with EBV-related peripheral T-cell lymphoma allowed the emergence of an EBV-related anaplastic variant of DLBCL and suggests a close relationship between p53 expression and latent EBV infection.     

Analysis of peripheral blood T-cell subsets in active thyroid-associated ophthalmopathy: absence of effect of octreotide-LAR on T-cell subsets in patients with thyroid-associated ophthalmopathy.

Thyroid-associated ophthalmopathy (TAO) is thought to be a T-cell-mediated autoimmune disorder. We sought to characterize abnormalities in the peripheral blood T-cell subsets in patients with TAO, and examine whether the long-acting somatostatin analogue, octreotide-LAR, treatment affects these cells. We analyzed peripheral blood T-cell subsets by flow cytometry in 26 euthyroid patients with moderately severe active TAO and 24 controls. Twenty-five of the patients with TAO were enrolled in a randomized trial to receive either 30 mg of octreotide-LAR (n = 11) or placebo (n = 14) every 4 weeks for 16 weeks; all 25 patients subsequently received octreotide-LAR 30 mg every 4 weeks from week 16 to 32. T-cell subsets were analysed at baseline, week 16, and week 32. At baseline, the relative percentage of CD4+ helper T-cells (p = 0.0003) and the CD4+/CD8+ ratio (p = 0.008) were significantly higher in patients with TAO compared to controls. Patients with TAO had higher na?ve active T cells (CD45RA+, CD45RA+ CD4+) and lower memory T cells (CD45RO+, CD45RO+ CD4+) than controls. At weeks 16 and 32, there were no significant differences in any T-cell subsets between the octreotide-LAR-treated and placebo groups. These results support a role of T cell in the pathogenesis of TAO, and show that octreotide-LAR has no effect on T-cell subsets during 32-weeks of treatment.     

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.     

The naive T-lymphocyte compartment is well preserved in patients with chronic myelogenous leukaemia in chronic phase

In chronic myelogenous leukaemia (CML), clonal change occurs in all myeloid and B-cell lineages, but very rarely T-cell lineages. A detailed three-colour cytometric analysis of peripheral lymphocytes was performed in 22 patients with chronic-phase CML (CP-CML). CD45 gating analysis was used to discriminate between lymphocytes and basophils. The peripheral lymphocyte pool was comprised of a significant proportion of naive CD4 cells, defined by a CD4+45RA+ phenotype [47.0 +/- 19.6% (mean +/- SD) of the total CD4+ cells], and naive CD8 cells, defined by a CD8+CD45RA+CD28+ phenotype (35.1 +/- 19.7% of total CD8+ cells), even in patients with long disease duration. The percentage of CD8 naive T cells showed inverse correlation with age, whereas no correlation was observed with disease duration. Possible explanations for the preservation of naive lymphocytes include (1) that the naive T cells differentiated from co-existing normal stem cells or (2) that long-lived naive T cells persisted from the CML onset and expanded peripherally (thymus independent). Either mechanism or a combination of both mechanisms might contribute to maintaining the naive compartment size.     

Asthma severity is associated with an increase in both blood CXCR3+ and CCR4+ T cells

OBJECTIVES: A predominance of type 2 helper T cells (Th2) in the bronchoalveolar space and peripheral blood is a well-accepted feature of bronchial asthma. However, the relationship between peripheral blood Th2 cells and asthma severity has not been thoroughly investigated. METHODS: As Th1 cells predominantly express the chemokine receptor CXCR3 and Th2 cells express CCR4, we assessed the distribution of peripheral blood CXCR3+ and CCR4+ lymphocytes using flow cytometry in 186 patients with asthma and 75 normal subjects. RESULTS: The proportion of CXCR3+/CD45RO+ cells in CD4+ T cells increased as the severity of asthma increased. The percentage of CCR4+/CD45RO+ cells in CD4+ T cells were elevated in mild to severe asthma patients compared with controls. However, there was no significant difference in CCR4+/CD45RO+ cells between the mild to severe asthma patients. There was no relationship between the patient's age and the numbers of CXCR3+ or CCR4+ T cells. The percentage of CCR4+ cells in CD45RO+/CD4+ T cells correlated with the levels of total serum IgE (r = 0.630, P < 0.0001). CONCLUSIONS: The proportion of CCR4+ cells in blood memory helper T cells may be increased in patients with asthma and is associated with the level of serum IgE, but severity of asthma is also associated with the increase of blood CXCR3+ cells in memory helper T cells.     

Reduced apoptosis of memory T-cells in the inner airway wall of mild and severe asthma.

Effector memory T-cells (CD45RO+) may provide pro-inflammatory signals that contribute to the persistent airway inflammation that is characteristic of asthma, and reduced apoptosis of these cells may prolong their effects. The present authors compared apoptosis of CD45RO+ T-cells in the inner airway wall in nonfatal asthma (n = 7), fatal asthma (n = 7) and control (n = 8) cases. Apoptotic cells were identified using both the terminal deoxynucleotidyl transferase dNTP nick end-labelling (TUNEL) technique and cell morphology. The percentage of CD45RO+ T-cells that were apoptotic was significantly greater in control cases compared with nonfatal and fatal cases of asthma, respectively, in small (42+/-19, 16+/-9, 7+/-6%), medium (40+/-12, 15+/-11, 12+/-8%) and large airways (42+/-15, 23+/-18, 18+/-12%). The reduction in the percentage of apoptotic CD45RO+ cells in the cases of asthma was observed in both blood vessels and the interstitium in large airways. In conclusion, these data suggest that reduced apoptosis may prolong the active life of effector memory T-cells in the airways. It is possible that survival signals may be received before cells migrate into the interstitium of the inner airway wall.     

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。     

CTLA-4 upregulation during HIV infection: association with anergy and possible target for therapeutic intervention

OBJECTIVE: To study the role of cytotoxic T-lymphocyte-associated antigen 4 (CTLA-4) during HIV infection. METHODS: Intracellular CTLA-4 expression, determined by flow-cytometry, and proliferative responses to HIV antigens, were studied in peripheral blood mononuclear cells (PBMC) from 93 HIV-1-infected [HIV(+)] patients and 40 HIV-1 seronegative controls. RESULTS: The proportions of CTLA-4 expressing CD4+ T cells were: (1) significantly higher in HIV(+) patients, 10.95 +/- 0.66%, than in controls, 6 +/- 0.45% (P < 0.0001); (2) inversely correlated to CD4+ counts (r = -0.67, P < 0.005, n = 16, drug-naive patients; r = -0.57, P < 0.0001, n = 77, HAART-treated patients); and (3) positively correlated to proportion of activated (HLA-DR+CD3+) (r = 0.53, P < 0.0001) and memory (CD45RO+CD4+) T cells (r = 0.46, P < 0.001). CD28 median fluorescence intensity in CTLA-4- cells was twice that in CTLA-4+ cells (140 +/- 5.3 versus 70 +/- 2.28, P < 0.00001), whereas cells low in CD28 and CD4, expressed more CTLA-4 (P < 0.0001). Higher proportion of CTLA-4+CD4+ cells expressed CCR5 and Ki-67, in comparison with CTLA-4-CD4+ cells, (65 +/- 11.9 and 25 +/- 7.5% versus 27 +/- 8.9 and 3.7 +/- 2%, P < 0.0001 and P < 0.01, respectively). Among HAART-treated patients, with viral load below detectable levels, CD4+ cells increase was inversely correlated to %CTLA-4+CD4+ cells (r = -0.5, P = 0.003, n = 39). Proliferation of PBMC to anti-CD3, gp-120 depleted HIV-1 antigen or HIV-1 p24 stimulation was inversely correlated with CTLA-4 levels (r = -0.68, P = 0.0035; r = -0.38,P = 0.04; and r = -0.43, P = 0.028, respectively). CONCLUSIONS: (1) CTLA-4 is upregulated during HIV infection and may therefore account for CD4 T-cell decline and anergy in HIV-1 infection. (2) Increased levels of CTLA-4 may undermine immune responses and in the HAART-treated patient-immune reconstitution. (3) Blocking of CTLA-4 may offer a novel approach for immune-based therapy in HIV infection.     

Expression of CD30 in patients with acute graft-versus-host disease

Acute GVHD (aGVHD) remains a major source of morbidity after allogeneic hematopoietic cell transplantation. CD30 is a cell-surface protein expressed on certain activated T cells. We analyzed CD30 expression on peripheral blood T-cell subsets and soluble CD30 levels in 26 patients at the time of presentation of aGVHD, before the initiation of treatment, compared with 27 patients after hematopoietic cell transplantation without aGVHD (NONE). Analysis by flow cytometry showed that patients with aGVHD had a greater percentage of CD30 expressing CD8(+) T cells with the difference especially pronounced in the central memory subset (CD8(+)CD45RO(+)CD62L(+)): GVHD median 12.4% (range, 0.8%-33.4%) versus NONE 2.1% (0.7%, 17.5%), P < .001. There were similar levels of CD30 expression in naive T cells, CD4(+) T cells, and regulatory (CD4(+)CD127(low)CD25(+)) T cells. Plasma levels of soluble CD30 were significantly greater in patients with GVHD: median 61.7 ng/mL (range, 9.8-357.1 ng/mL) versus 17.4 (range, 3.7-142.4 ng/mL) in NONE (P < .001). Immunohistochemical analysis of affected intestinal tissue showed many CD30(+) infiltrating lymphocytes present. These results suggest that CD30 expression on CD8(+) T-cell subsets or plasma levels of soluble CD30 may be a potential biomarker for aGVHD. CD30 may also represent a target for novel therapeutic approaches for aGVHD.     

Changes in the expression of surface receptors on lymphocyte subsets in the elderly: quantitative flow cytometric analysis

The immunophenotype of circulating lymphocytes, including the intensity expression of surface receptors, changes with ageing. Until now, no results of systematic studies on age-dependent changes with respect to the expression of the major lymphocyte surface receptors in healthy elderly subjects have been reported. In order to identify age-related changes in both representation and immunophenotype of lymphocyte populations, we investigated, by means of triple-color whole-blood immunostaining and quantitative flow cytometry, the percent values and the absolute numbers, as well as the levels of surface antigen expression or antigen molecules per cell (ABC values x 10(3)), of different peripheral blood lymphocyte subsets from 23 healthy elderly subjects and 13 young donors. Naive (CD45RA+CD3+) T cells, total B cells, and CD5+ B lymphocytes are decreased (22%, 6%, 0.8% vs. 30%, 12%, 1.4%, respectively), whereas activated (HLA-DR+CD3+) and memory (CD45RO+CD3+) T cells, CD3+CD7- T lymphocytes, and lymphocytes expressing the NK marker CD56 are expanded in the elderly (2%, 53%, 13%, 6% vs. 0.8%, 45%, 8%, 8%, respectively). Moreover, T lymphocytes from elderly individuals express lower CD3 (61 +/- 10) compared to young (69 +/- 10). Considering the different T-cell populations, CD3 antigen is respectively decreased on CD45RO+ T cells (55 +/- 14 vs. 66 +/- 14) and up-regulated on CD56+ T lymphocytes (62 +/- 21 vs. 45 +/- 20). Increased CD8 expression characterizes CD3+CD7- lymphocytes (70 +/- 34 vs. 44 +/- 17) while HLA-DR on activated T cells is lower in old (39 +/- 7) than young (46 +/- 9) donors. CD7 is down-regulated both in T (22 +/- 3 vs. 28 +/- 3) and NK (48 +/- 18 vs. 71 +/- 18) cells, whereas CD2 expression, unchanged on NK cells, is up-regulated on T lymphocytes (54 +/- 10 vs. 41 +/- 8). Age-related changes in B-cell antigen expressions were also found: CD20 is increased (124 +/- 23 vs. 105 +/- 16) whereas, despite the unchanged CD5 expression of T cells, CD5 intensity on the B-cell subset co-expressing this antigen is higher in old (49 +/- 37) than in young (22 +/- 4) people. The observed changes in the expression of functionally important cellular receptors can contribute to the remodeling of immune function characteristic of the elderly. Moreover, since quantitative flow cytometry is becoming widely employed in clinical practice, our results also contribute to the assessment of specific age-dependent antigen expression changes to be considered for diagnostic approaches in the elderly.     

T-cell division in human immunodeficiency virus (HIV)-1 infection is mainly due to immune activation: a longitudinal analysis in patients before and during highly active antiretroviral therapy (HAART)

In human immunodeficiency virus (HIV)-1 infection, highly increased T-cell turnover was proposed to cause exhaustion of lymphocyte production and consequently development of AIDS. Here, we investigated cell proliferation, as measured by expression of the Ki-67 nuclear antigen, in peripheral blood CD4(+) and CD8(+) lymphocyte subpopulations before and during highly active antiretroviral therapy (HAART). In untreated HIV-1 infection, both the percentage and number of Ki-67(+) CD4(+) and CD8(+) lymphocytes were significantly increased, compared with values obtained from healthy individuals. A more than 10-fold increase in the percentage of dividing naive CD4(+) T cells in the blood was found when the number of these cells were below 100 per microL. HAART induced an immediate decline in Ki-67 antigen expression, despite often very low CD4(+) T-cell numbers, arguing against increased proliferation being a homeostatic response. After approximately 24 weeks of HAART treatment, a transient increase in the number of proliferating cells was seen, but only in the CD4(+) CD27(+) memory pool. In the CD8(+) T-cell compartment, the number of dividing cells was elevated 20- to 25-fold. This increase was most notable in the CD27(+) CD 45RO(+) and CD27(-) CD45RO(+) memory CD8(+) T-cell pool, corresponding with the degree of expansion of these subsets. Reduction of plasma HIV-RNA load by HAART was accompanied by a decrease in numbers and percentages of dividing cells in all CD8(+) T-cell subsets. Taken together, our results indicate that peripheral T-cell proliferation is a consequence of generalized immune activation. (Blood. 2000;95:249-255)     

Predominance of CD8+ T lymphocytes in psoriatic arthritis.

OBJECTIVE: To characterize the synovial fluid (SF) derived T cell populations in psoriatic arthritis (PsA) and compare with similar populations from rheumatoid arthritis (RA). METHODS: Paired peripheral blood (PB) and SF samples were analyzed by 3 color flow cytometry using monoclonal antibodies to CD3, CD4, CD8, HLA-DR, CD25, CD45RA, and CD45RO. RESULTS: There was a significantly increased CD8+ T cell population in PsA SF compared to RA: PsA mean 61% (range 35-93), RA mean 46% (range 6-72) (p < 0.005). This resulted in a reversal of the CD4:CD8 ratio in PsA SF compared to RA SF (p < 0.001). Patients with oligoarticular PsA had the most pronounced differences in SF derived T cell populations compared to RA (p < 0.0005) but these results were not significantly different from PsA patients with a polyarticular disease pattern. PB PsA T cell populations were not different from controls, in contrast to RA, where the CD4+ T cell population was increased (p < 0.0026), giving an exaggerated PB CD4:CD8 ratio. The majority of PsA SF CD8+ T cells expressed CD45RO, mean 73% (range 58-95), and HLA-DR antigen: mean 72% (range 38-94). Low levels of CD25 were detectable in this population, indicating a nonclassical activation pattern: mean 2% (range 0.3-4.4). CONCLUSION: In PsA, activated (HLA-DR+) and mature (CD45RO+) CD8+ T cells predominate in SE Analysis of this population may uncover clues to pathogenesis in this HLA class I mediated disease.     

Absence of bcl-2 expression by activated CD45RO+ T lymphocytes in acute infectious mononucleosis supporting their susceptibility to programmed cell death

bcl-2 proto-oncogene encodes an inner mitochondrial membrane protein that blocks programmed cell death (apoptosis). There is now increasing evidence that regulation of bcl-2 expression is a determinant of life or death in normal lymphocytes. We have recently described that activated (CD45RO+) CD4+ and CD8+ T cells in acute infectious mononucleosis (IM) undergo apoptotic cell death on culturing, indicating an activation-driven cell death of mature T cells. In this work, we examine bcl-2 expression by activated T cells in acute IM using a flow-cytometric analysis with an anti-bcl-2 monoclonal antibody (MoAb). It was consistently observed that most T cells from acute IM patients displayed only much less bcl-2, while normal T cells expressed bcl-2 relatively strongly. Multicolor analysis showed that bcl-2- lacking T cells in acute IM were restricted to the CD45RO+ (activated) populations of CD4+, as well as CD8+ T cells. In contrast, the relatively intense levels of bcl-2 were expressed in both CD45RO+ and CD45RO- T-cell populations from normal subjects. This marked difference in bcl-2 expression of CD45RO+ T cells between acute IM and normal controls was also confirmed by Western blot analysis. Activated (CD45RO+) T cells with low bcl-2 expression, but not bcl-2-expressing CD45RO- T cells, in acute IM patients were found to die easily when cultured without added growth factors. However, in normal individuals, both CD45RO+ and CD45RO- T cells were relatively stable on culturing. These findings suggest that lack of bcl-2 expression by activated (CD45RO+) T cells in acute IM might be associated with their susceptibility to programmed cell death.     

Flow cytometric assessment of lymphocyte subsets, lymphoid progenitors, and hematopoietic stem cells in allogeneic stem cell grafts.

Currently, bone marrow (BM), cord blood (CB), and G-CSF-mobilized peripheral blood progenitor cells (PBPCs) are the most commonly used sources for allogeneic stem cell transplantation (SCT). The aim of this study was to assess the yields and distribution of lymphocyte subsets, lymphocyte progenitors and hematopoietic stem cells (HSC) in each type of allograft by three-color flow cytometry. The yields of CD34(+)CD38(-) HSCs did not differ significantly between BM grafts (2.80 +/- 0.74 x 10(6)) and leukapheresis products (LPs) (1.82 +/- 0.64 x 10(6)), and were lowest in CB grafts (0.21 +/- 0.05 x 10(6)). For most lymphocyte subsets yields were lowest in CB grafts and significantly higher in LPs than in BM grafts. BM grafts, however, contained the highest yields of CD34(+)CD19(+)CD20(-) B cell progenitors and CD19(+)CD20(-) B cells. The relative frequencies of the naive CD45RA(+)CD45RO(-) phenotype among CD4(+) and CD8(high) T cells were highest in CB grafts (P < or = 0.001), and higher in LPs than in BM grafts (P < or = 0.02). The latter finding was in accordance with a preferential G-CSF mobilization of naive T cells relative to the total lymphocyte population (P < or = 0.014). CD3(+)CD8(low) and CD3(+)CD8(low)CD4(-) subsets, which facilitate engraftment in murine transplantation models, demonstrated a tendency towards lower frequencies among T cells in CB grafts and LPs compared to BM grafts. This observation coincided with a significantly reduced mobilization of subsets potentially enriched for facilitating cells as compared to the total lymphocyte population (P < or = 0.036). The CD34(+) compartment of CB grafts contained a significantly higher percentage (12.1%) of CD34(+)CD7(+)CD3(-) T cell progenitors than those of BM grafts (5.1%) and LPs (3.6%). In addition, CB lymphocytes contained the highest fraction of CD3(-)CD16/56(+) NK cells (P < or = 0.013) and almost no CD3(+)CD16/56(+) NKT cells (P < 0.001) compared to adult cell sources. In summary, LPs, CB allografts and BM allografts differ widely with respect to the cellular composition of their lymphocyte compartments, which is partially affected by a varying mobilization efficiency of G-CSF for distinct lymphocyte subsets.     

Comparison between sirolimus- and paclitaxel-eluting stent in T-cell subsets redistribution

We sought to investigate the effects of 2 different coronary drug-eluting stents on the distribution of central or effector memory T cells circulating in the coronary sinus of patients with coronary artery disease who underwent percutaneous coronary revascularization. We randomly assigned 43 patients (mean age 65.4 +/- 4.3 years; 34 men) presenting with stable coronary disease and angiographically proved stenosis of the left anterior descending artery to treatment with sirolimus- or paclitaxel-eluting stents. Heparinized blood samples were obtained from the coronary sinus before and 20 minutes after stent implantation. Analysis of surface phenotype was performed by 4-color flow cytometry, and data are expressed as the percentage of positive cells. The percentages of CD8+ and CD4+ effector memory T cells, as defined by the CD3+CD45RO+CD27- phenotype, were significantly reduced in patients who received a sirolimus-eluting stent compared with the basal values. Conversely, the percentages of CD8+, but not CD4+, central memory T cells (CD3+CD45RO+CD27+) were increased in the same treatment group after the revascularization procedure. No changes in the percentages of memory T-cell populations in the paclitaxel-eluting stent group were observed. These findings show that sirolimus-eluting stents rapidly induced a redistribution of memory T lymphocytes, with a significant decrease of proinflammatory effector memory T cells circulating within the coronary sinus.