Value of fluorescence in situ hybridization for detecting the bcr/abl gene fusion in interphase cells of routine bone marrow specimens

A patient presented with a severe nephrotic syndrome and a renal biopsy consistent with early focal segmental glomerulosclerosis (FSGS). After a year of intensive immunosuppressive therapy proteinuria was unabated and renal function began to deteriorate. Treatment with weekly plasmapheresis combined with moderate doses of prednisone and azathioprine produced a dramatic decrease in proteinuria and serum creatinine. A marked fall in the B-cell population occurred during treatment, as well as an increase both in T-lymphocytes of the mature CD4+ helper/suppressor phenotype and in the immature/cytotoxic CD8+ phenotype. Activation of the immune system during treatment was demonstrated by an increase in the rate of spontaneous proliferation of peripheral blood mononuclear cells and an increase in T-cell expression of the interleukin-2 receptor.     

Abnormal thymocyte subset distribution and differential reduction of CD4+ and CD8+ T cell subsets during peripheral maturation in diabetes-prone BioBreeding rats

In this study we quantified CD8+ and CD4+ T cells in T lymphocytopenic BB rats as compared with control rats at given stages along the maturational pathway from immature thymocytes to mature peripheral T cells. Our results show that BB rats exhibit abnormal thymocyte subset distribution. Numbers of mature TCRhigh/CD4-8+ thymocytes, and also their TCRhigh/CD4+8+ precursors were decreased, as were levels of CD8 expression on all thymocyte subsets investigated. By analogy with mouse thymocyte development, these findings suggest a decreased efficiency for positive selection of CD8 precursors in BB rats. Furthermore, as related to the number of available mature TCRhigh single positive thymocytes, numbers of CD4+ and CD8+ T cells most recently migrated from the thymus were severely decreased in BB blood, indicating either reduced thymic output or rapid cell death after migration. Subsequently, in peripheral blood and cervical lymph nodes, a 95% decrease of CD8+ and a 50 to 80% decrease of CD4+ T cells were demonstrated upon maturation from recent thymic migrants to mature peripheral T cells, leaving the BB rat with a severely reduced T cell population, consisting of CD4+ T cells and a minute population of CD8+ T cells. The vast majority of the latter was found to have an immature peripheral phenotype. Possible consequences of our findings for the generation of autoreactive CD8+ T cells are discussed.     

B-lymphocyte suppression in multiple myeloma is a reversible phenomenon specific to normal B-cell progenitors and plasma cell precursors

The reduced levels of normal immunoglobulin in patients with myeloma may be due to suppression of normal B-cell differentiation. However, reports on the numbers of B cells vary, with some finding decreases consistent with immunoparesis, and others reporting expansions of phenotypically aberrant cells. We have therefore assessed the phenotype and levels of B lymphocytes in patients at presentation (n = 23), in plateau or complete remission (PB n = 42, BM n = 18), and in relapse (PB n = 17, BM n = 14), in comparison to normal individuals (n = 10). Phenotypic analysis was performed using five-parameter flow cytometry, with CD14 used to exclude monocytes where necessary. We found no evidence of a phenotypically distinctive blood or marrow B-cell population in patients with myeloma, nor of an increase in the levels of any B-cell subset. Numbers of blood CD19+ 38+ normal plasma cell precursors were significantly reduced in presentation/relapse patients, but not in patients in plateau/remission. Total CD19+ cells were significantly reduced only in patients with circulating myeloma cells, detected by IgH-PCR. In the marrow, CD19+ B cells expressing CD5, CD10, CD34, CD38, CD45(low) and Syndecan-1 were significantly decreased at presentation/relapse, but not in patients in plateau/remission. The majority of these antigens are expressed by normal B-cell progenitors, indicating that myeloma also affects the early stages of B-cell development. The suppression of progenitor cells was not restricted to B-lymphoid differentiation, as total CD34+ cells were also significantly reduced in the marrow of myeloma patients at presentation. These results indicate that, if neoplastic B cells are present in myeloma, they are low in number and have a phenotype similar to their normal counterparts. Furthermore, there is a reversible suppression of CD19+ B lymphocytes that correlates inversely with disease stage, and specifically affects the early and late stages of normal B-cell differentiation.     

Comparative study of transfer factor and acyclovir in the treatment of herpes zoster

Reactivation of varicella herpes virus (VHV), latent in individuals who have previously suffered varicella, gives rise to herpes zoster and in some cases leads to a sequela of post herpetic neuritis with severe pain which is refractory to analgesics. Many different antiviral agents have been tried without achieving satisfactory results. Of all the antiviral agents employed, acyclovir has been the most successful in reducing post herpetic pain. However acyclovir has not been as reliable as interferon alpha (IFN-alpha). We have previously looked into the use of transfer factor (TF) as a modulator of the immune system, specifically with respect to its effectiveness in the treatment of herpes zoster. In this work findings from a comparative clinical evaluation are presented. A double blind clinical trial of TF vs acyclovir was carried out in which 28 patients, presenting acute stage herpes zoster, were randomly assigned to either treatment group. Treatment was administered for seven days and the patients were subsequently submitted to daily clinical observation for an additional 14 days. An analogue visual scale was implemented in order to record pain and thereby served as the clinical parameter for scoring results. The group treated with TF was found to have a more favorable clinical course, P < or = 0.015. Laboratory tests to assess the immune profile of the patients were performed two days prior and 14 days after initial treatment. The results of these tests showed an increase in IFN-gamma levels, augmentation in the CD4+ cell population but not the percentage of T rosettes in the TF treated group. These parameters were however insignificantly modified in patients receiving acyclovir. Although TF treated patients showed an increase in CD4+ counts these cells remained below the levels for healthy individuals. The fact that IFN-gamma levels as well as the counts for CD4+ cells rose in the TF treated group and not in the acyclovir one is very significant and confirms the immunomodulating properties of TF.     

The functional phenotype of the primitive plasma cell in patients with multiple myeloma correlates with the clinical state

The malignant plasma cells from patients with multiple myeloma display considerable phenotypic heterogeneity. All plasma cells express high intensity CD38 (CD38++), cytoplasmic immunoglobulin and either kappa or lambda light chains. Subpopulations of mature (CD45-), immature (CD45+) and primitive (CD45++, CD19+) plasma cells can be defined but little is known about the functional differences and clinical significance of these subpopulations. Three colour flow cytometry and permeabilisation was used to determine the expression of functionally important antigens in plasma cell subpopulations. These antigens included the labelling index (LI, bromodeoxyuridine), number of nucleoside transporter per cell, p-glycoprotein (JSB-1), and oncoprotein expression (c-myc, c-fos, c-neu, bcl-2, p-ras, p53m, p-53w, and Rb). In progressive disease there was an increase in the absolute number but not the percentage of CD45++ plasma cells. There was a significant difference in the mean LI of the CD38++, CD45++ population in progressive disease compared with stable disease (9.2% vs 2.2%; z = 19.9, p < 0.001). The LI of CD45++ cells ranged up to 45% and provided a better correlation with disease status than the LI of the total cell population. Any increase in nucleoside transporters or p-glycoprotein expression was almost entirely attributable to an increase in the primitive plasma cell population. In 96% (n = 28) of samples from patients in progressive disease there was at least one abnormality in the functional phenotype of the primitive plasma cells. This is in contrast with 44% of samples from patients in stable disease (n = 58). These studies suggest that the functional phenotype of the primitive plasma cell determines the clinical phenotype of patients with myeloma.     

Herpes zoster, immunological deterioration and disease progression in HIV-1 infection

OBJECTIVE: To study the incidence of herpes zoster, the relationship between herpes zoster and immunological markers, and the prognostic value of herpes zoster for progression of HIV disease. DESIGN AND METHODS: A total of 966 homosexual participants in The Amsterdam Cohort Study were studied. Herpes zoster was defined by its characteristic clinical presentation. Incidence was calculated using Poisson regression, cumulative incidence by the Kaplan-Meier product-limit method and the prognostic value was evaluated using Cox proportional hazards model. RESULTS: The incidence of first episodes of herpes zoster was 3.31 per 1000 person-years (PY) in HIV-seronegatives and 51.51 per 1000 PY in HIV-1-seropositive individuals. Recurrences only occurred in HIV-1-positive patients (25.6%). Cumulative incidences of first episodes increased linearly with the duration of follow-up. In HIV-1-seropositives the incidence was 31.2 per 1000 PY at CD4+ cells > or = 500 x 10(6)/l, 47.2 per 1000 PY [relative risk (RR), 1.51; 95% confidence interval (CI), 0.78-2.94] at CD4+ cells 200-499 x 10(6)/l and 97.5 per 1000 PY (RR, 3.13; 95% CI, 1.54-6.32) at CD4+ cells < 200 x 10(6)/l. Besides CD4+ cell counts, CD3 monoclonal antibodies and phytohaemagglutinin-induced T-cell reactivity were independent predictors for herpes zoster. The hazard ratio for AIDS after herpes zoster was 1.6 (95% CI, 1.1-2.4) and for death 1.7 (95% CI, 1.1-2.5), but these were not independent from CD4+ cell counts. CONCLUSION: In HIV-1 infection the incidence of herpes zoster increases with the decrease of CD4+ cell counts and T-cell reactivity, but herpes zoster is not an independent predictor for disease progression.     

Analysis of the in vitro cytokine production by liver-infiltrating T cells of patients with autoimmune hepatitis

The pathogenic mechanisms underlying the development of autoimmune hepatitis (AIH) are still unclear. Since AIH is associated with the presence of various autoantibodies and certain HLA subtypes, it is likely that T and B cells play a major role in this disease. In this study we have determined the functional capacities of in vivo preactivated liver-infiltrating T cells (LTC) from patients with AIH. As controls we used LTC from patients with non-autoimmune hepatitis (non-AIH). Our results show that preactivated LTC from patients with AIH predominantly (190/255 clones) reside in the CD4+ population, whereas LTC in non-AIH are dominated by the CD8+ phenotype (148/254 clones). In view of this finding we have investigated the cytokine secretion patterns of 102 randomly chosen CD4+ T cell clones from six patients with AIH. As controls we have used 58 CD4+ LTC from 11 patients with non-AIH. All clones were stimulated by lectin and irradiated accessory cells and subsequent cytokine production was evaluated. LTC from patients with AIH have a lower interferon-gamma (IFN-gamma)/IL-4 ratio compared with LTC from non-AIH. Although clones from some patients with AIH produced very high amounts of IL-4 in vitro, this was not a constant finding. These results show that in vivo preactivated LTC from patients with AIH are mostly CD4+ T cells that produce more IL-4 than IFN-gamma. In contrast, LTC from patients with non-AIH are dominated by CD8+ and CD4+ T cells that produce significantly less IL-4 than IFN-gamma. Thus, liver-infiltrating T cells from patients with AIH and non-AIH belong to different functional T cell subsets. This may have implications for the regulation of humoral and cellular immune responses in inflammatory liver disease.     

Thiazolidinediones--tools for the research of metabolic syndrome X

Recent studies have demonstrated that the expression of Fas by peripheral T cells from HIV-1+ patients is deregulated and increases the susceptibility of these cells to undergo apoptosis. Here, we show that secretion of Fas-ligand (L), the complementary agonist of Fas, is abnormally upregulated in CD4+ cells from HIV-1-infected individuals, particularly during the non-lymphopenic stages of the disease. An increase of soluble Fas-L occurred in T cell cultures from 26 patients with a number of CD4+ cells higher than 400/microliter, whereas it was almost undetectable in cultures from 21 severely lymphopenic patients (CD4+ < 200/microliter). The MTT test, cytofluorimetric analysis of cellular DNA, cytotoxicity, and proliferative assays using the Fas-transfected WC8 mouse lymphoma confirmed the cytocidal capability of T cell supernatants from non-lymphopenic patients. Double-fluorescence analysis revealed that the majority of CD4+ cells (approximately 90%) in these cultures secreted Fas-L in the presence of high intracellular gamma-interferon and low Bcl-2. In contrast, the CD8+/Fas-L+ population was comparably decreased (approximately 55%). Molecular cloning of Fas-L revealed a substantial expression of Fas-L mRNA in cells from non-lymphopenic patients compared with patients with advanced disease and healthy controls. Since CD4+ cells of Th1 phenotype are impaired during HIV-1 infection and show high cellular expression of Fas-L, it is conceivable that excess Fas-L during the early or non-lymphopenic phase of the disease increases the extent of apoptosis in these cells by the Fas/Fas-L pathway. The defective expression of the ligand in severely lymphopenic stages could be explained by exhaustion of this mechanism as the disease progresses.     

Expression of the activation antigen CD27 in rheumatoid arthritis

Differentiation of CD4+ T-cells is reflected by the change from the CD45RA+CD27+ phenotype via CD45RO+CD27+ to the CD45RO+CD27- phenotype. To provide insight into the migration and activation of T-cells at the site of inflammation in rheumatoid arthritis (RA), CD27 expression by T-cells in peripheral blood (PB), synovial fluid (SF), and synovial tissue (ST) as well as the levels of the soluble form of CD27 (sCD27) in plasma and SF were studied in patients with RA. Since CD4+CD27+ T-cells are involved in providing helper activity for B-cells, we also investigated the levels of rheumatoid factors in serum and SF in relation to CD27 expression. The mean level of sCD27, which is produced by CD27+ cells, and the mean percentage of CD27 T-cells within the CD4+CD45RA- subset were higher in SF than in PB. SF sCD27 levels were higher in the patients with RA than in the patients with osteoarthritis, who served as controls. In ST infiltration by CD4+CD45RO+CD27+ T-cells, could be demonstrated in the rheumatoid perivascular lymphocytic aggregates with a relative increase in the percentage of CD27- T-cells in the diffuse lymphocytic infiltrate. The sCD27 levels and the percentages of CD4+CD27+ cells in SF correlated positively with the levels of rheumatoid factors in serum and SF. The findings presented in this study suggest a continuous influx of preactivated CD4+CD45RO+CD27+ cells from the PB into the rheumatoid ST and further activation and differentiation to CD4+CD45RO+CD27- cells in situ, followed by migration to the SF. These activated T-cells are likely to play a role in synovial inflammation.     

Increase in adhesion molecules on CD4+ cells and CD4+ cell subsets in synovial fluid from patients with rheumatoid arthritis

OBJECTIVE: To elucidate the role of adhesion molecules in the pathogenesis of rheumatoid arthritis (RA). METHODS: We evaluated their expression and that of an activation marker on CD4+ cell populations and CD4+ cell subsets in specimens of peripheral blood (PB) and synovial fluid (SF) obtained from 10 patients with RA and 7 with osteoarthritis (OA). A 2 or 3-color immunofluorescent method was used for analysis. RESULTS: The SF from both groups of patients showed a greater density of adhesion molecules including LFA-1 alpha, LFA-1 beta, CD2, VLA-4 alpha and VLA-5 alpha on CD4+ cells, and a higher percentage of CD4+HLA-DR+ cells compared with their PB. IN PB-CD4+ cell subsets from the arthritic and healthy subjects, the CD4+CD45RO+ cell population showed an increased expression of adhesion molecules compared with CD4+CD45RA+ cell population. The expression of adhesion molecules on circulating CD4+ cell population and CD4+ cell subsets from the patients with RA and OA was comparable to that from healthy subjects. SF from both groups of patients showed a higher percentage of CD4+CD45RO+ cells and a lower percentage of CD4+CD45RA+ cells. In SF-CD4+ cell subsets from patients with RA, the CD4+CD45RO+ cell population had an increased expression of VLA-4 alpha compared to the CD4+CD45RA+ cell population; however, there was no significant difference in other adhesion molecule expression and the percentage of HLA-DR+ cells between the 2 cell subsets. Furthermore, the expression of VLA-4 alpha on the CD4+CD45RO+ cell population in SF from patients with RA was significantly higher than that in matched PB. In CD4+CD45RA+ cell population from both groups of patients, SF showed an enhanced expression of adhesion molecules and an increased percentage of HLA-DR+ cells compared with matched PB. CONCLUSION: Our results suggest that increased expression of adhesion molecules and increased percentage of HLA-DR+ cells on CD4+ cells in SF may be responsible for cellular interactions between these cells and synovial cells or extracellular matrix.     

Induction of type 2 cytokines by a staphylococcal enterotoxin superantigen

Persistent intramammary infections of dairy cows with Staphylococcus aureus may involve immunosuppression mediated by bacterial toxins such as enterotoxins and other super-antigens (SAgs). Previously we found that stimulation of bovine PBMC with staphylococcal enterotoxin C (SEC) induced a unique phenotype of activated CD8+ T cells expressing a newly identified activation molecule, ACT3. In the present study we found that SEC induced the expression of interleukin (IL)-4 and IL-10 mRNAs, two cytokines associated with type 2 responses. Elevated levels of IL-4 and IL-10, observed between day 0 and day 4 of culture, were associated with temporary inhibition of proliferative responses of T cells, evidenced by a decrease in numbers of CD4+ T cells and a small increase in numbers of CD8+ T cells. Vigorous proliferation of T cells occurred between days 4 and 7 of culture and with a bias towards CD8+ T cells. Acquisition of the ACT3+ phenotype by CD8+ T cells was preceded by induction of IL-4 mRNA. Thus, in the bovine system, SAgs may hinder protective responses by inducing type 2 cytokines, which interfere with immune clearance of many microbial pathogens. The results of the study are consistent with the hypothesis that SAgs are involved in immunosuppression, and suggest possible immunomodulatory mechanisms.     

Direct ex vivo flow cytometric analysis of human microglial cell CD4 expression: examination of central nervous system biopsy specimens from HIV-seropositive patients and patients with other neurological disease

OBJECTIVE: To define a clear ex vivo flow cytometric phenotype for adult human microglia that would distinguish it from all other macrophage lineage cells in the central nervous system (CNS) or blood, and to utilize this phenotype to examine the activation state and CD4 expression of microglia freshly derived from CNS tissue of HIV-positive patients and those with other neurological diseases. DESIGN: Fresh human CNS tissue from both HIV-uninfected and HIV-infected individuals was obtained by biopsy or resection, and cells isolated immediately, labelled for flow cytometry and analysed. METHODS: A Percoll density gradient isolation technique and phenotypic characteristics used for rodent microglia were applied and modified. RESULTS: Resident microglia could clearly be defined by the flow cytometric phenotype CD45low CD4- CD11b+ CD11chigh major histocompatibility complex (MHC) class II+ CD26- CD14-. Assuming normally low-level MHC class II expression in the healthy CNS, it was likely that MHC class II positivity reflected underlying pathology necessitating biopsy or resection and appeared to be a 'leaky' activation marker. Microglia activation was observed in specimens from only six (35%) out of 17 HIV-uninfected but all four (100%) HIV-infected patients, defined strictly as any level of upregulation of CD4 expression, to produce the phenotype CD45low/medium CD4low CD11b+ CD1.1Chigh MHC class II+/+2 CD26- CD14-. Where examined by immunohistology, CD68 was also upregulated in these cases. CONCLUSIONS: When activated in situ, microglia express low levels of CD4 and this is always seen in tissue from HIV-infected patients. Using the flow cytometric phenotype established here, microglia from HIV-infected tissue can now be isolated in pure form and studied directly ex vivo.     

Age-related increase in the fraction of CD27-CD4+ T cells and IL-4 production as a feature of CD4+ T cell differentiation in vivo

The influence of ageing on phenotype and function of CD4+ T cells was studied by comparing young (19-28 years of age) and aged (75-84 years of age) donors that were selected using the SENIEUR protocol to exclude underlying disease. An age-related increase was observed in the relative number of memory cells, not only on the basis of a decreased CD45RA and increased CD45RO expression, but also on the basis of a decrease in the fraction of CD27+CD4+ T cells. Our observation that the absolute number of CD45RO+CD4+ T cells was increased, while absolute numbers of CD27-CD4+ T cells remained unchanged in aged donors, indicates that the latter subset does not merely reflect the size of the CD45RO+CD4+ T cell pool. The increased fraction of memory cells in the aged was functionally reflected in an increased IL-4 production and T cell proliferation, when cells were activated with the combination of anti-CD2 and anti-CD28, whereas IL-2 production was comparable between both groups. No differences were observed with respect to proliferative T cell responses or IL-2 production using plate-bound anti-CD3 or phytohaemagglutinin (PHA). The observation that IL-4 production correlated with the fraction of memory cells in young donors but not in aged donors suggests different functional characteristics of this subset in aged donors.     

The CD7- T cell subset represents the majority of IL-5-secreting cells within CD4+CD45RA- T cells

Absence of CD7 is a stable phenotype in a subset of normal human T cells. Most circulating CD7- T cells express the CD4CD45RO+CD45RA- memory phenotype. We analysed CD4+CD45RA- peripheral blood lymphocytes that were separated into CD7+ and CD7- for their in vitro cytokine secretion in response to different stimuli. The CD4+CD7- subpopulation was found to secrete significantly higher levels of IL-5 compared with the CD4+CD7- subset upon stimulation with ionomycin/phorbol myristate acetate (PMA) plus anti-CD28 MoAbs. In contrast to IL-5 secretion, IL-4 and interferon-gamma (IFN-gamma) secretion was not significantly different in CD7+ and CD7- T cells upon stimulation in vitro. The data indicate that the CD4+CD7- T cell represents the majority of IL-5-secreting cells within the population of CD4+CD45RA- memory T cells. Since CD4+CD7- T cells were found to be enriched in various skin lesions associated with eosinophilic infiltration, the results of our study support the hypothesis that skin-infiltrating CD7- T cells are one of the major sources of IL-5 responsible for the development of eosinophilic inflammation in certain skin diseases.     

Infinity and the limits of the unconscious

Macrophages play an important role in the intestinal mucosal immune system. However, they are a poorly defined cell population. We therefore determined their phenotype in normal colonic mucosa. Macrophages were isolated from colonic biopsies and surgical specimens by collagenase digestion. Colonic macrophages were positively sorted by anti-CD33 magnetic beads. Flow cytometric triple fluorescence analysis was applied to study CD14, CD16, CD33, CD44, CD11b, CD11c, CD64, HLA-DR, CD80, CD86 and CD3/CD19 expression. CD33 was evaluated as a positive marker for intestinal macrophages. CD33+ cells isolated from normal colonic mucosa showed co-expression of the established intracellular macrophage marker CD68 in FACS analysis. CD33+ cells were capable of phagocytosis. Isolation of this cell population by magnetic anti-CD33 beads and culture resulted in a 4.2-40-fold increase in IL-1beta and 4.5-44-fold increase in tumour necrosis factor-alpha (TNF-alpha) secretion compared with unsorted lamina propria mononuclear cells (LPMC). Of the CD33+ cells, 90.9 +/- 6.9% (mean +/- s.d.) were CD44+. However, macrophages from colonic mucosa showed only a low expression of CD14 (10.5 +/- 3.8%), CD16 (10.1 +/- 3.9%), HLA-DR (27.3 +/- 9.2%), CD11b (17.4 +/- 6.8%), CD11c (17.8 +/- 10.4%). Furthermore, expression of CD80 (9.2 +/- 4.2%) and CD86 (15.1 +/- 7.3%) was low, suggesting a low ability of normal intestinal macrophages to activate T cells and T cell-mediated immune responses. We conclude that CD33 is useful for the isolation and flow cytometric characterization of colonic macrophages. These cells exhibit a single phenotype in normal mucosa (CD33++, CD44++, CD14-, CD16-, CD11b-, CD11c-, HLA-DRlow, CD80-, CD86-) lacking lipopolysaccharide (LPS) receptor and costimulatory molecules.     

CDw60: a marker for human CD8+ T helper cells

The existence of helper cells among the CD8+ T cell subset has been recognized for a long time. However, the phenotype of these cells has remained elusive. In this study, we provide evidence that the expression of the CDw60 antigen on human CD8+ T cell allows one to distinguish between CD8+ T helper cells and CD8+ T cells with cytotoxic and suppressor capacity. CDw60 monoclonal antibodies (mAb) recognize the 9-O-acetylated disialosyl group on ganglioside GD3 expressed on 20-40% of CD8+ cells. By use of the direct and indirect mAb-rosetting technique, we were able to isolate the CDw60+CD8+ and CDw60-CD8+ cells at high purity. The alloantigen-specific cytotoxic activity of CD8+ cells resided entirely in the CDw60- population. Helper and suppressor capacity of both CD8 subsets was assayed by the pokeweed mitogen-induced differentiation of B cells into immunoglobulin-secreting cells. These studies clearly indicate that the CDw60+CD8+ subset provided substantial help to B lymphocytes, whereas the CD8+ cells with the CDw60- phenotype were suppressing B cell differentiation. Both subsets produced similar amounts of interleukin 2 (IL-2) after stimulation with phytohemagglutinin. Activation with phorbol myristate acetate in combination with Ca-ionophore induced IL-4 secretion in both populations, but preferentially in the CDw60+ subset, whereas the vast majority of interferon gamma was produced by the CDw60-CD8+ cells. When used in combination with other markers, CDw60 may prove to be useful in defining CD8+ subsets with reciprocal functional activities.     

Apoptosis (the 1992 Frank Rose Memorial Lecture)

Lymphocyte subset abnormalities have been observed in a series of patients diagnosed with Wegener's granulomatosis (WG). Fifteen patients with WG were evaluated for lymphocyte subset abnormalities by two-color immunofluorescence flow cytometry. The WG group showed a decrease in CD4+ cells and an extraordinary increase in HLA-DR+ cells. The average percentages of HLA-DR+ cells of the normal group and WG group were 11.0 +/- 4.0 and 40.5 +/- 13.9% (CD3+HLA-DR+ = 20.3 + 9.4%). Within the CD8 family, CD8+57+, CD8+38+ and CD8+HLA-DR+ cells were markedly elevated. Our findings differ from other series which reported that the lymphocyte subsets in the peripheral blood of patients with WG were normal.     

Coexpression of GD3 ganglioside with CD45RO in resting and activated human T lymphocytes

The ganglioside GD3 is preferentially expressed on the surface of malignant T cell lymphoblasts and on resting T cells which express the memory cell phenotype, CD45RA-CD29+. However, GD3 expression in activated T cells and its potential function in proliferating normal and malignant T cells are unclear. Utilizing three-color immunostaining and flow cytometry, we examined changes in the expression of GD3 in conjunction with the RA and RO isoforms of CD45 during in vitro T cell activation. GD3 was equally expressed in resting CD4 and CD8 cells and was specifically found in the CD45RO+RA population. Activation of T cells with PHA resulted in an increased percentage of GD3+ cells. This increase was evident by the first day and was observed in the CD45RO (naive cell) population; by 2 days, GD3 was expressed heterogeneously in a large population of CD45RO+RA+ cells. Further activation of T cells with PHA or anti-CD3 monoclonal antibody (OKT3) resulted in a further increase in GD3-expressing cells, and the increase in GD3 density correlated with increased CD45RO and loss of CD45RA. In contrast, increases in GD3 and interleukin-2 receptor (CD25) expression in response to PHA or OKT3 occurred independently, indicating that the GD3/ CD45RO coexpression observed was not a general consequence of cell activation. The results provide evidence for specific comodulation of GD3 and CD45RO during T cell mitogenesis, and thus suggest that these molecules may colocalize on the T cell surface.     

Gender differences in use of stress testing and coronary heart disease mortality: a population-based study in Olmsted County, Minnesota

The distribution of CD57+ T and CD56+ T cells in patients with RA was examined. In control osteoarthritis patients, these cells exist as a minor population in the peripheral blood. Our data show that in patients with RA, CD57+ T cell levels are elevated in peripheral blood, knee joint fluid, knee synovial membrane and bone marrow (BM), compared with peripheral blood of controls. CD57+ T cells are especially high in knee joint fluid and joint-adjacent BM, while CD56+ T cells show no such increase. CD57+ T cells contain a major population of CD8+ cells and higher proportions of CD4-8- cells and gammadelta T cells than do CD57- T cells. CD57+ T cells in peripheral blood and joint fluid increase with the duration of disease. Erythrocyte sedimentation rate (ESR) is inversely correlated with the proportion of CD57+ T cells in the joint fluid. Although RA frequently occurs in patients with CD3+57+ cell leukaemia, and some CD57+ T cells are likely to be involved in the onset of RA, we suggest that CD57+ T cells may rather suppress inflammation of RA, and other cellular components (e.g. granulocytes) may govern the severity of the inflammation of RA. These CD57+ T cells are probably generated extrathymically in the adjacent BM or joint space.     

The CD8+ cell phenotype mediating antiviral activity in feline immunodeficiency virus-infected cats is characterized by reduced surface expression of the CD8 beta chain

The acute stage of feline immunodeficiency virus (FIV) infection is characterized by a CD8+ anti-FIV response that parallels the appearance of a CD8+ subpopulation with reduced expression of the beta chain (CD8 alpha + beta lo). The relationship between the CD8 alpha + beta lo phenotype and CD8+ anti-FIV activity was examined. Flow cytometric analysis of peripheral blood mononuclear cells with anti-CD8 beta chain monoclonal antibody 117 revealed that the CD8 alpha + beta lo phenotype expanded throughout the asymptomatic infection, constituting 80%-90% of the CD8 beta + cells in long-term-infected cats. Purified CD8 alpha + beta hi and CD8 alpha + beta lo subpopulations were analyzed for anti-FIV activity in an acute infection assay. Anti-FIV activity resided principally in the CD8 alpha + beta lo population and was demonstrated in acute FIV infections, as well as in long-term asymptomatic infections. These data suggest that a unique CD8 alpha + beta lo anti-FIV phenotype arises early in infection and may play a major role in eliminating virus and maintaining the asymptomatic infection.