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PubMed1993

Distinctive increases in HLA-DR+ and CD8+57+ lymphocyte subsets in Wegener's granulomatosis.

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.

Adult↗
PubMed1997

Interrelations between Immunogenetic Factors (HLA antigens) and Lymphocyte Subset Quantitative Rearrangements in Hodgkin's Disease Patients.

In this report we analyzed interrelations between cell quantity in lymphocyte subsets and lymphocyte immunogenetic (HLA) markers in 30 untreated patients with Hodgkin's disease (HD). We defined percentage and the absolute number of peripheral blood lymphocyte subsets expressing CD3, CD4, CD8, CD16, CD25 and CD72 markers for HD patients referring to their HLA phenotype (A, B, Cw loci and DRB1). HD patients had decreased absolute number of all the lymphocyte subsets. This was apparently associated with reduced number of peripheral blood lymphocytes, since their subset shares remained similar to those of healthy volunteers. HD patients had different HLA repertoire: some displayed simultaneous exertion of four antigens in A and B loci ("full house" patients) and others - reduced HLA repertoire ("non-full house" patients). Simultaneous analysis of lymphocyte subset rearrangements and HLA expression revealed that "full house" patients had no significant rearrangements in lymphocyte subsets, except reduced CD4(+) and increased CD25(+) lymphocytes. The reduction of expressed HLA alleles interrelated with reliable decrease of CD3(+), CD4(+), CD16(+) cells. Expression of such HLA alleles as A1, A2, B13, B16, B17, B21, B27, DR2 and DR4 also interrelated with significant decrease in some lymphocyte subsets, of which CD4(+) cells prevailed. The most distinct reduction of lymphocyte subsets interrelated with expression of B17 and DR2 HLA loci. Hence, in HD patients pathological rearrangements of lymphocyte subsets are strictly associated with their HLA expression.

Journal Article↗
PubMed1984

Distribution of acid alpha-naphthyl acetate esterase among human T lymphocyte subsets.

Human T lymphocyte subsets, identified by means of OKT3, 4 and 8 monoclonal antibodies, were isolated by a fluorescence activated cell sorter (FACS IV) and analyzed for distribution of alpha-naphthyl acetate esterase (ANAE) activity. As compared to OKT8+ lymphocytes a higher proportion of OKT4+ lymphocytes was ANAE-positive exhibiting a spot or dot-like pattern in the cytoplasm. OKT8 and 4 positive subsets showed a similar ANAE distribution in diffuse granular form. Although OKT4 and OKT8 populations presented a different ANAE dot-like reactivity, this marker did not allow as clear a distinction between them as that reported for TG and TM lymphocytes.

Cell Separation↗
PubMed1999

Lymphocyte subsets and mitogen stimulation of blood lymphocytes in preeclampsia.

PROBLEM: The question of whether there are differences in systemic immune reactivity in severe preeclampsia compared with normal pregnancy was addressed. METHOD OF STUDY: During the third trimester, blood samples were taken from 12 pregnant women with severe preeclampsia. Five of the preeclamptic pregnancies were analyzed separately because they were treated with dexamethasone before the blood samples were taken. The seven dexamethasone-treated preeclamptic pregnant women were analyzed and compared with six uncomplicated pregnancies. A control group consisted of 15 nonpregnant females. Lymphocyte subsets were identified by flow cytometry. The function of peripheral blood mononuclear cells (PBMCs) was studied as proliferative responses to mitogens alone and in combination with immunomodulating drugs. RESULTS: An increased number of B lymphocytes (CD19+) (P < 0.05) and natural killer (NK) cells (P < 0.05) was noticed in severe preeclampsia compared with normal pregnancy. The proliferative response of PBMCs in phytohemagglutinin (PHA)-stimulated cultures in autologous serum from patients with severe preeclampsia was reduced (P < 0.05) compared with normal pregnancy. The addition of indomethacin and cimetidine significantly stimulated (P < 0.05) the proliferative responses. The enhancing effect of cimetidine was not found in dexamethasone-treated preeclamptic patients. CONCLUSIONS: The presence of systemic immunosuppression in severe preeclampsia is demonstrated as a reduced proliferative response of PBMCs to PHA, which could be partly restituted by indomethacin or cimetidine, indicating immunosuppressor activity that is mediated by prostaglandin and histamine. Increased levels of B lymphocytes and NK cells were also noticed.

Adult↗
PubMed2001

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.

Adult↗
PubMed1993

Reference ranges for lymphocyte subsets in pediatric patients.

Peripheral blood lymphocyte subset reference ranges were examined in a large group (N = 130) of healthy pediatric patients ranging in age from 1 month to 17 years. All samples were stained with monoclonal antibodies, processed with a whole blood lysis technique, and analyzed on a flow cytometer. Data analysis demonstrated statistically significant changes in most lymphocyte subsets at age 3 years. The relative and absolute numbers of total lymphocytes, CD2, CD4, and CD19 cells; absolute numbers of CD3 and CD8 cells; and CD4/CD8 ratios were high at birth, decreased during early childhood, and closely approximated adult reference values after age 3 years. The relative numbers of CD8 lymphocytes were low in early childhood and then rose to adult values after 3 years of age. The relative percentage of CD3 cells remained stable over all ages studied. Although "adult" lymphocyte subset reference ranges may be similar to those in children older than 3 years, age-adjusted reference ranges should be used for the early childhood period.

Adolescent↗
PubMedDate not supplied

Age-related changes in blood lymphocyte subsets of south Indian children.

BACKGROUND: Enumeration of lymphocyte subsets has been widely used for the diagnosis and monitoring of several haematological and immunological disorders. Various studies have demonstrated age, sex and racial differences in lymphocyte subset expression. Reference values are not available for Indian children and there is a need for this information to replace commonly used, but inappropriate, adult lymphocyte subset ranges. METHODS: One hundred thirty-eight healthy children between 3 and 15 years of age, attending a local government school in Chennai, South India were Included in the study. Haemoglobin levels, and total and differential cell counts were determined using an automated counter and lymphocyte subsets were analysed by flowcytometry. RESULTS: The mean (SD) absolute lymphocyte count declined with age from 4338 (1031) at 3 years to reach a plateau of 3096 (914) at 11-13 years (p < 0.05). A significant decline was also observed in the absolute numbers of CD3+, CD4+, CD8+ and CD19+ cells. However, the percentage values of CD3+, CD4+, CD8+, CD16/56+ cells and the CD4/CD8 ratio remained fairly stable across the age range. CONCLUSION: Our data would prove useful in interpreting disease-related changes in lymphocyte subsets in Indian children of different age groups. Age-related decrease in the absolute lymphocyte count as well as numbers of CD4 and CD8 cells was found to occur between the ages of 3 and 11 years. A normogram relating age to CD4 count has been developed.

Adolescent↗
PubMed1986

Analysis of responding lymphocyte subsets in allogeneic mixed lymphocyte cultures in the neonates and adults.

An analysis of proliferating cell subsets in the allogeneic mixed lymphocyte culture (MLC) in adults (n = 19) and newborns (n = 12) was performed by a method combining immunoperoxidase staining and autoradiography. The main cell type responding in adult cells were OKT8-positive (38%), whereas in neonates OKT4-positive cells (31%) predominated. Furthermore, in the neonatal allogeneic MLC there were significantly more 3H-thymidine incorporating Leu 12-positive B cells than in adults (12% vs 4%).

Adult↗
PubMed1995

Selective recruitment of lymphocyte subsets to the inflamed appendix.

Total lymphocyte counts and the distribution of lymphocyte subsets were determined in peripheral venous blood and appendiceal mononuclear cells from 60 patients who underwent appendicectomy for the clinical diagnosis of appendicitis. A significant peripheral lymphopenia was observed in the 46 patients with histologically confirmed acute appendicitis which was accompanied by an increase in the appendiceal lymphocyte concentration. There was an even greater depletion of CD45RO+ (memory) T lymphocytes in peripheral blood and an increase in the inflamed appendix. Reciprocal changes were observed in the CD45RA+ (naive) T lymphocyte subset. These changes were reflected in the local arterial and venous CD45RA and CD45RO T lymphocyte subsets. Proliferation studies showed an expanded functional repertoire of T lymphocytes in the inflamed appendix. Selective recruitment of memory T lymphocytes from the peripheral blood to the inflamed appendix was demonstrated.

Acute Disease↗
PubMed1994

[Analysis of lymphocyte subsets in human lingual tonsils].

Lymphocytes in the lingual tonsil were analyzed using monoclonal antibodies to cell surface markers. Immunohistologic staining of lingual tonsillar tissue showed localization of the various lymphocyte subsets in the germinal center, mantle zone, and inter-follicular area. Populations of lymphocyte subsets, i.e., T cells, B cells, CD4 positive cells, CD8 positive cells, alpha beta T-cells, gamma delta T-cells, and NK cells were assessed by laser flow cytometry analysis, and we compared the following tissues: peripheral blood, lingual tonsils, palatine tonsils, and pharyngeal tonsils (adenoids).

Adult↗
PubMed1983

Patients with abnormal proportions of T-lymphocyte subsets have reduced in vitro cellular immunity.

Monoclonal antibodies which identify helper/inducer (OKT4) and cytotoxic/suppressor (OKT8) subsets of human T lymphocytes have recently been used to investigate immunoregulation in isolated cell populations, as well as in human disease states. However, the relationship between relative proportions of OKT4- and OKT8-positive blood lymphocytes and in vitro cellular immune function in patients with immunodeficiencies has not been studied extensively. We enumerated T-lymphocyte subsets with OKT4 and OKT8 antibodies, and measured proliferative responses to allogeneic cells in mixed lymphocyte culture (MLC) and to phytohemagglutinin (PHA), in peripheral blood mononuclear cells (PBMCs) from 60 patients with varying degrees of immunodeficiency and 20 healthy controls. Controls had 56.0 +/- 5.3% (mean +/- 1SD) OKT4-positive lymphocytes, 28.6 +/- 5.9% OKT8-positive lymphocytes, and an OKT4/8 ratio of 2.05 +/- 0.55. We defined as abnormal values of less than 40% OKT4-positive or greater than 45% OKT8-positive lymphocytes (3 SD below and above mean values, respectively), or an OKT4/8 ratio of less than 1.2. Patients with these abnormal percentages of T-lymphocyte subsets had significantly lower mean MLC and PHA responses (P less than 0.001), and higher incidences of abnormal MLC and PHA responses (P less than 0.01). Abnormal proportions of T-lymphocyte subsets correlated with low MLC and PHA responses in most immunodeficient patients, although some patients with low MLC and PHA responses had normal subset distributions. T-Cell subset proportions were heterogeneous among patients with the same diagnosis.

Adult↗
PubMed1991

Detection of lymphocyte subsets using three-color/single-laser flow cytometry and the fluorescent dye peridinin chlorophyll-alpha protein.

The fluorescent dye, Peridinin chlorophyll A protein (PerCP) derived from dinoflagellate organisms (Glenodinium sp.) can be excited by a 488 nm laser and emits light with a large Stokes shift and no major spectral overlap with commonly used chromophores such as fluorescein isothiocyanate (FITC) and R-phycoerythrin (PE). PerCP was conjugated directly to various mouse monoclonal antibodies (mAb) specific for human leukocyte markers or to avidin for use with biotinylated-mAb, and used to perform three color single-laser flow cytometry. The efficacy of this method was demonstrated by analyzing the heterogeneity of thymus T lineage subsets and B lymphocyte subsets in blood. CD4-CD8-, CD4+CD8+ and CD4+CD8- or CD4-CD8+ subsets differ in their expression of cell-cell interaction markers including CD18, CD28, CD44 and Leu 8, and activation/subset markers CD45RO, CD45RA and CD26. Some CD5+ peripheral blood B cells, unlike CD5-B cells, expressed CD45RO or high levels of CD54 (ICAM-1) suggesting the CD5+ B cell population contains activated lymphocytes. The availability of such an accessible method for three color analysis will make it possible to do routine three color monitoring of immunologic diseases such as AIDS, and autoimmune or periodontal diseases.

Antibodies, Monoclonal↗
PubMed1995

Alterations in lymphocyte subsets as prognosticators of postoperative infections.

OBJECTIVE: To evaluate changes in lymphocyte subsets after major abdominal and thoracic operations, and to correlate changes with the development of clinically relevant infections postoperatively. DESIGN: Open study. SETTING: University hospital, Italy. SUBJECTS: 33 patients who were to undergo major abdominal or thoracic operations. INTERVENTIONS: Lymphocyte subsets were measured by cytofluorimetry before operation and 1, 3, 5, and 7 days postoperatively. MAIN OUTCOME MEASURES: Correlation between changes in the number of lymphocyte subsets and development of infection. RESULTS: Lymphocyte subsets were within the reference range in all patients before operation. 10/33 Patients developed infections (pneumonia, bacteraemia, or wound or urinary tract infections) between the second and the ninth days postoperatively (30%). On day 1 the numbers of all lymphocyte subsets had decreased significantly compared with the preoperative measurements in all patients (CD3 p < 0.01, CD4 p < 0.001, and CD8 p < 0.05). The reduction in CD3 was significantly greater in the group that developed infections (p < 0.001). Among patients who did not develop infections the numbers of lymphocyte subsets had returned to the reference range within a week of operation whereas among patients who developed infections they remained depressed (p < 0.05). CONCLUSION: The synchronous reduction in numbers of all lymphocyte subsets on the first day postoperatively to below 50% of the reference range (CD3 to < 600/microliters, CD4 to < 400/microliters, and CD8 to < 250/microliters) predicted the development of infection postoperatively with an accuracy of 89%, a sensitivity of 80%, and a specificity of 96%.

Adult↗
PubMed1995

Characterization of distribution of T lymphocyte subsets and activated T lymphocytes infiltrating into sarcoid lesions.

We studied the relationship between various T lymphocyte functions and granuloma formation in 5 lung tissue and 4 lymph node tissue samples from patients with sarcoidosis by immunohistological methods. In the lesion of sarcoidosis, T cells were positive for alpha beta TCR, but gamma delta TCR-positive T cells were rarely observed. The results of analysis of functional subsets showed that T cells in the internal area of granuloma were predominantly helper/inducer subset (CD4+CD45RA-). On the other hand, cytotoxic T cells (CD8+CD45RA-CD11b-) were present in abundance in the outer boundaries of granuloma. In addition, suppressor-inducer T cells (CD4+CD45RA+) were present in the surrounding areas. However, T cells of various subsets were present sporadically outside the granulomas. We also studied the distribution of T cells expressing activation-related antigens. The results showed that T lymphocytes in the internal area of granulomas more frequently had these antigens than did T lymphocytes in the external area. These findings suggested that T cells infiltrating into the sarcoidosis lesion demonstrated a layer-like distribution based on functional subsets. These findings also confirm that activated T cells were more abundantly distributed in the internal area of sarcoid granuloma than in the external area.

Antibodies, Monoclonal↗
PubMed1995

Regulation of T lymphocyte subsets.

Patterns of cytokine secretion and functional differences distinguish T lymphocyte subsets. T lymphocyte subsets are also regulated differentially. Most established CD8+ lymphocyte clones secrete gamma-interferon (IFN-gamma) but not interleukin 2 (IL-2) or IL-4. Using murine T cells which express a transgenic, antigen-specific alpha/beta T cell receptor (TCR) specific for L(d) class I major histocompatibility complex antigen, we have found that CD8+ lymphocytes can be divided into functional subsets. Freshly isolated CD8+ T cells are not cytolytic, do not proliferate and do not proliferate and do not secrete cytokines. Stimulation of TCR alone does not induce cytokine secretion, but cells become responsive to exogenous IL-2 or IL-4. Stimulation of CD28 together with TCR induces secretion of IL-2 and IFN-gamma, and cells proliferate without exogenous cytokines. Proliferation is necessary for the development of cytolytic activity. If IL-4 is present during initial stimulation, IL-4 is secreted following restimulation. Upon stimulation, some IL-4-producing murine CD8+ T cell clones express CD40 ligand (CD40L), and they potentiate proliferation and immunoglobulin secretion by small resting B cells. Thus, the CD8+ T cell subsets T cytotoxic 1 (Tc1) and Tc2 are analogous to CD4+ T helper 1 (Th1) and Th2. IL-2 production by naive CD8+ cells requires co-stimulation. IL-4 production by CD8+ T cells requires the presence of IL-4 during initial stimulation. Some IL-4-producing CD8+ T cells express CD40L following TCR stimulation and provide help for B cells.

Animals↗
PubMed1996

Modulation of peripheral blood lymphocyte subsets during methylprednisolone pulse therapy.

We determined fluctuations in circulating lymphocyte subsets induced by methylprednisolone pulse therapy (MPT) and the continuous administration of prednisolone (PSL) in 17 patients with autoimmune or systemic rheumatic disease. Two-color flow cytometry, using monoclonal antibodies to various lymphocyte subsets, was performed to identify a possible association between the clinical efficacy of treatment and modulative effects on each subset. Both MPT and continuous oral PSL showed suppressive effects on most of the lymphocyte subsets: CD4+, CD45RA or CD45RA+CD4+, CD8+, CD11b.CD8+, CD5+ B, and CD57+ or CD57 CD16+ cells. Modulation of lymphocyte subsets were more profound with MPT than with continuous oral PSL. The results are relevant to the different degrees of immuno-suppression effected by the two treatment modalities. We found that the number of CD45RA.CD4+ cells after MPT treatment correlated with the clinical efficacy of the treatment: the less CD45RA.CD4+ cell numbers decreased after MPT treatment, the greater was the clinical efficacy of the treatment. The results probably are associated with a rapid recovery of the subset after MPT treatment in the responders. Thus, the sequential monitoring of circulating lymphocyte subsets is useful in predicting the clinical effects of MPT treatment.

Adult↗
PubMed1995

Influence of race, age and sex on the lymphocyte subsets in peripheral blood of healthy Malaysian adults.

The lymphocyte subsets in the peripheral blood of healthy Malaysian adults (212 subjects, age 18-71 years) were analysed using a flow cytometer FACScan in an effort to establish a reference range for the lymphocyte subsets. The lymphocyte subsets studied were T cells (CD3), B cells (CD19), natural killer (NK) cells (CD3- CD16+/CD56+), helper/inducer cells (CD4), cytotoxic/suppressor cells (CD8) and the helper/suppressor ratio (CD4/CD8). The distributions of T cells, CD4 cells and CD8 cells were symmetric about their means while B cells, NK cells and CD4/CD8 ratio followed a skewed distribution. Differences in race were observed for T cells, NK cells, CD4 cells and CD4/CD8 ratio where the Indians were significantly different from the Malays and the Chinese (higher T cells, CD4 cells and CD4/CD8 ratio and lower NK cells). The B cells were significantly lower in the Chinese than the Malays and the Indians. Age differences were seen only in the Chinese where increased CD4 cells and CD4/CD8 ratio, and decreased CD8 cells were observed. A sex difference was observed only in the Chinese where the CD4/CD8 ratio was significantly higher in females than males.

Adolescent↗