OKT8 Cells Research Articles

Phenotypic characteristics of peripheral blood T cells regulating colony growth by nontransformed human B lymphocytes.

The phenotypic characteristics of peripheral blood T cell subpopulations regulating human B cell colony growth stimulated by Staph protein A were investigated. Colony growth was facilitated by OKT4 cells, and T cells expressing DR antigens were found to be partially responsible for colony facilitation. A linear increase in the magnitude of colony growth was observed with greater T cell numbers, and maximal colony enhancement occurred when T cells were present during the early stages of colony formation. OKT8 cells did not enhance colony growth and also inhibited the facilitation of colony formation by OKT4 cells. Other experiments showed that the functional activities of OKT4 and OKT8 cells differed in their requirements for DNA synthesis. Although active T cell DNA synthesis was absolutely required for the facilitation of colony growth at all concentrations tested, DNA synthesis was not needed for OKT8 inhibition of OKT4 promotion of colony formation. Thus, distinct T cell subsets whose functional properties differ in their requirements for DNA synthesis regulate human colony growth.

Lymphocyte subpopulations in patients with multiple sclerosis.

Using monoclonal antibodies, peripheral blood helper/inducer (OKT4) and cytotoxic/suppressor (OKT8) lymphocytes were measured in 14 normal controls, 36 patients with multiple sclerosis at different stages of the disease and 15 patients with isolated optic neuritis. Thirty-four of these individuals were studied on two or more occasions at intervals up to 340 days. Patients with multiple sclerosis in relapse had low levels of OKT8 cells (14.07% +/- 3.79) compared with controls (29.42% +/- 4.69) and this abnormality returned to normal within approximately one month of the onset of new symptoms. Further changes occurred with new relapses. Low OKT8 cells were also found in patients with isolated optic neuritis (18.76 +/- 3.71) or progressive multiple sclerosis (19.91% +/- 7.96); the same pattern of recovery was seen in these two groups as in patients with multiple sclerosis in relapse. In 25% patients studied on two or more occasions after an episode of demyelination abnormalities of lymphocyte subpopulations occurred which were not accompanied by new clinical symptoms or signs. Fluctuations of this kind did not occur in controls. The findings have implications for the pathogenesis and management of patients with multiple sclerosis.

Lymphocyte subsets in Chediak-Higashi patients

Peripheral blood lymphocyte subsets were studied in 6 Chediak-Higashi patients and 12 family members. The lymphocyte subsets were characterized by monoclonal antibody reagents and fluorescence flow-cytometry. An increase in OKT8 (suppressor/ cytotoxic) and a decrease in OKT4 (helper) cell populations was observed in all patients studied. No correlation was seen between the clinical status (presence or absence of the lymphoproliferative phase) and the percentage of the lymphocyte subsets. The patient's mothers also had an increased percentage of OKT8-positive cells. The significance of these findings is discussed regarding the patients clinical course.

Active T rosettes, human autologous T rosettes, OKT8 and OKT4 cells, Con A-induced suppressive activity, and autoantibodies: Clinical correlations

Correlations between various T-cell subsets: OKT4, OKT8, TE h (human autologous T rosettes), and TE a (active T rosettes) cells and the concanavalin A-induced suppressor function assessed on Con A proliferation have been analyzed in 46 patients with various secondary immunological disorders. The different T subsets and the suppressive activity were also compared in patients with and without autoantibodies. Significant positive correlations were found between T-cell markers TE a-OKT8 and TE h-OKT4. Significant inverse correlations were also found between Con A-induced suppressive activity- OKT4 OKT8 ratios, and TE a- OKT4 OKT8 ratios suggesting that the subpopulations identified by the active T rosettes are probably involved in T immunoregulatory mechanisms. There was no association between the Con A-induced suppressive function or the T markers TE a, OKT4, and OKT8, and the presence of autoantibodies. Only subjects with autoantibodies had a lower percentage of T lymphocytes forming autologous rosettes. These observations emphasize the fact that lack of correlation may exist between markers or function and immune status in some patients.

Identification of subpopulations of T lymphocytes and Ia positive B lymphocytes using a rosette assay based upon the biotin-avidin interaction

Bovine erythrocytes were coated with avidin using a chromic chloride coupling technique and used successfully in an indirect rosette assay to identify and quantitate: (a) T-lymphocyte helper and suppressor subpopulations, and (b) Ia positive B-lymphocyte populations. Using mouse monoclonal antibodies to the T-lymphocyte markers OKT3, OKT4, and OKT8 it was shown that the proportion of OKT4 and OKT8 positive cells were respectively 60.9 and 39.2% of the total OKT-3 positive T lymphocytes. In a similar way, using a monoclonal anti-human Ia antibody, the percentage Ia positive cells in B-lymphocyte enriched preparations was shown to vary between 18 and 55% for normal peripheral blood and 31 and 58% for chronic lymphocytic leukaemia derived peripheral blood.

Characterization of T-lymphocyte subsets in hairy-cell leukaemia (HCL) by monoclonal antibodies: comparison with Fc gamma, Fc mu receptors and correlation with disease activity.

T lymphocytes from 22 patients with hairy-cell leukaemia (HCL) were assessed on the basis of their ability to bind the Fc receptors for IgM (T mu) or IgG (T gamma), and by the capacity to react with OKT monoclonal antibodies. T-cell subsets defined by the presence of Fc receptors for IgM or IgG showed an overall increase in the proportion of T gamma cells and a non-significant decrease of T mu cells, regardless of the clinical state of the disease. Results with monoclonal antibodies showed that in patients with HCL in clinical remission T-cell subsets were normally balanced, while in patients with active disease the distribution and absolute number of T-cell subpopulations appeared markedly impaired, with a significant increase of OKT8 positive cells (suppressor/cytotoxic) and a significant reduction of OKT4 positive cells (helper/inducer) compared both with active disease patients and with normal controls. The OKT4+/OKT8+ ratio was also significantly reduced in patients with active disease compared with those in clinical remission and with controls (0.96 v 1.63 and v 1.94, respectively). Our findings confirm the heterogeneity of T-cell subset positivity defined by monoclonal antibodies and by Fc mu and Fc gamma receptors and suggest that in patients with HCL the distribution of OKT4 and OKT8 positive cells is closely correlated to the clinical state of the disease.

The correlation between inducer/suppressor ratios, generation of concanavalin A-activated suppressor cells, and mitogen-stimulated proliferation of peripheral blood lymphocytes in patients with alopecia areata and normal controls

Suppressor T-cell function activated by preincubation of peripheral blood mononuclear cells with concanavalin A (Con A) is mediated by the human lymphocyte subset which reacts with monoclonal antibody OKT5 8 (E. L. Reinherz, P. C. Kung, G. Goldsteing, and S. F. Schlossman, J. Immunol. 124, 1243, 1980; E. L. Reinherz and S. F. Schlossman, Cell 19, 821, 1980). Proliferative responses stimulated by mitogenic lectins involve T cells of both the OKT4 and OKT8 phenotypes (E. L. Reinherz and S. F. Schlossman, N. Engl. J. Med. 303, 370, 1980). As part of a study of immunologic parameters in patients with alopecia areata and concurrent normal controls we analyzed the results of functional responses of the subjects' T cells (mitogen proliferation and ability to generate Con A-activated suppression) according to the proportions of OKT4 (helper/inducer) and OKT8 (suppressor/cytotoxic) cells quantitated at the time of study ( OKT4 OKT8 ratios ranging from 0.53 to 3.55). Absolute total T-cell counts and lectin-stimulated proliferative responses (phytohemagglutinin, Con A) were similar for patient and control groups. Patients with alopecia areata, and particularly those who demonstrated regrowth of hair at the time that immunologic studies were performed, generated increased Con A-activated suppression in conjugation with an increase in the proportions of peripheral OKT8 cells. For all subjects studied, OKT4 OKT8 ≤ 1.5 correlated with Con A-activated suppression > 50% ( P = 0.0006), and relatively decreased PHA and Con A proliferative responses ( P < 0.01, P = 0.01, respectively). These findings suggest that functional responses of peripheral lymphocytes reflect the proportional distribution of peripheral immunoregulatory subsets among healthy individuals as well as patients with alopecia areata.

Reduction of the Fraction of Circulating Helper-Induced T Cells Identified by Monoclonal Antibodies in Psoriatic Patients Treated with Long-term Psoralen/Ultraviolet-A Radiation (PUVA)

Ultraviolet radiation has been found to alter the distribution and function of human lymphocytes. To determine whether photochemotherapy (PUVA) alters circulating levels of T cell subset marker-bearing lymphocytes, cells from 9 patients with psoriasis undergoing PUVA therapy for several years (mean 4.6 +/- 1.4 yr), 17 patients with active untreated psoriasis, and 20 healthy volunteers were reacted with monoclonal antibodies to T cell surface markers, including OKT3 (all peripheral blood T cells), OKT4 (helper/inducer T cells), OKT6 (common thymocytes), and OKT8 (suppressor/cytotoxic T cells), and analyzed by flow cytometry. There were no differences in the distribution of T cell subsets between healthy volunteers and patients with active psoriasis. In contrast, the percentages of lymphocytes reacting with OKT3 and OKT4 were lower (by 16% and 12% percent respectively, p less than 0.0025) in the PUVA-treated patients compared to healthy volunteers or patients with active psoriasis that had not received PUVA therapy. There was no difference in the percentage of OKT8 and OKT6 bearing cells. Squamous cell carcinoma of the skin subsequently developed in 2 of 3 PUVA-treated patients with the lowest percentages of T4-bearing cells. These findings indicate that long-term PUVA therapy is associated with a reduction in circulating helper/inducer T cells. This reduction may have a role in the altered immune function reported in PUVA-treated patients.

Effects of prostaglandin E 2 and preincubation on lectin-stimulated proliferation of human T cell subsets

Our laboratory as well as others has previously noted that sensitivity of lymphocytes to inhibition by prostaglandin (PG)E 2 is lost after overnight preincubation. We now show that the change in sensitivity to PGE 2 with preincubation varies depending upon the T-cell subset in the culture. In phytohemaglutinin (PHA)-stimulated cultures of fresh cells, PGE 2 inhibits proliferation of T cells as well as OKT4 (+) cells and OKT8(+) cells. In PHA-stimulated cultures of preincubated cells, PGE 2 slightly augments proliferation of T cells; however, PGE still inhibits OKT4(+) cells and causes marked augmentation of OKT8(+) cells. Thus the “loss in sensitivity” to PGE 2 with preincubation is actually a result of the balancing effect of PGE 2 inhibiting OKT4(+) cells and stimulating OKT8(+) cells.

Phenotypic heterogeneity of the OKM1-positive lymphocyte population: Reactivity of OKM1 monoclonal antibody with a subset of the suppressor/cytotoxic T-cell population

Upon analysis on a fluorescence-activated cell sorter, about 20% of E-rosette-positive cells from normal donors were found to stain with OKM1, a monoclonal antibody reacting primarily with cells of myelomonocytic origin. These OKM1(+) cells are not monocytes since they have small light scatter and are peroxidase negative. They also stain with OKT11 and antibody 9.6, two monoclonal antibodies against an antigen identical to or related to the E receptor. In two-color immunofluorescence experiments, the existence of an OKM1(+) OKT8(+) cell was demonstrated. This was also confirmed with additive staining experiments. This cell type constitutes ~10% of the E-rosette(+) cell population. OKM1 reacts with approximately 33% of OKT8(+) cells. Conversely, 48% of OKM1(+) E-rosette(+) cells are also OKT8(+). The same overlap was found between Leu 2a and OKM1 while no overlap between OKT4 and OKT8 staining cells, or between OKT4 and OKM1 staining cells was detected. The OKM1(+) lymphocyte population was isolated by removing OKT3(+) cells from nylon wool-nonadherent cells: 43 ± 13% of these cells also reacted with Ab 9.6 and 33 ± 10% with OKT8. At least three subpopulations of OKM1(+) lymphoid cells can therefore be identified: one without any T cell marker, one that is E-receptor positive, and one that has, in addition to the E receptor, a definite T-cell marker (OKT8). In addition, OKT8(+) cells can be separated into OKT8(+) OKT3(+) and OKT8(+) OKT3(−) populations. Only the OKT8(+) OKT3(+) population suppresses Ig synthesis in pokeweed mitogen-stimulated cultures.

Regulatory T cell-subset imbalance in chronic active hepatitis.

Three monoclonal anti-T-cell antibodies, specifically directed against total T cells (OKT3), inducer-helper T cells (OKT4) and suppressor/cytotoxic T cells (OKT8), were used in this study to analyze peripheral T-cell subsets in hepatitis B surface antigen (HBsAg)-positive and -negative chronic active hepatitis (CAH) patients. Results showed that a clear-cut difference exists in the distribution of peripheral T cells of these two groups of subjects. HBsAg-positive CAH patients had a numerical predominance of peripheral T lymphocytes expressing the characteristics of cytotoxic/suppressor T cells. In contrast, patients with "autoimmune" HBsAg-negative CAH exhibit a predominance of OKT4 cells, namely, the helper-inducer T-cell subset. In addition, high numbers of circulating double labeled cells (expressing both the OKT4 and the OKT8 xenoantigens) were detected in some of the HBsAg-positive and HBsAg-negative CAH patients studied.

The measurement of leukocyte subsets in the peripheral blood of cancer patients with solid tumors using monoclonal antibody reagents.

Monocyte and lymphocyte subsets were quantitated in the peripheral blood of normal subjects and patients with solid tumors using the monoclonal antibody reagents OKM1, BRL63D3, OKT3, OKT4, and OKT8. Percentages and numbers of cells reacting with monoclonal reagents were analyzed by indirect immunofluorescence. Correlations between leukocyte subset values and stage of disease or immunocompetence were sought. No differences from normal were seen in the percentage of OKT3, OKT4, and OKT8 cells or in the ratios of OKT4/OKT8 cells in peripheral blood mononuclear cells (PBMC) from all cancer patients, patients with localized disease, or patients with advanced disease. A significant decrease in absolute numbers of lymphocytes, OKT3 cells, OKT4 cells, and OKT8 cells was seen in the peripheral blood of patients with advanced disease reflecting the absolute lymphopenia of these patients. A significant increase was seen in the percentages of PBMC reacting with OKM1 and BRL63D3 from patients with advanced disease compared with normal donors or localized disease patients. A positive correlation was found between PHA responsiveness and absolute numbers of OKT3 and OKT4 cells. A negative correlation was found between PHA responsiveness and percentages of OKM1 cells. These data indicate that malignant disease does not alter T cell subset percentages in patient peripheral blood but may decrease their absolute numbers in association with absolute lymphopenia. On the other hand, percentages of OKM1 and BRL63D3 cells are increased in patients with advanced solid tumors in association with impaired PHA responsiveness.

Interstitial mononuclear cell populations in renal graft rejection. Identification by monoclonal antibodies in tissue sections.

The interstitial mononuclear cell populations of 22 renal grafts with interstitial rejection (IR), 6 grafts with interstitial nephritis without rejection (IN), and 5 kidneys without infiltration (3 donor kidneys, 2 grafts) were identified and quantitated by monoclonal antibodies recognizing T cells (TA-1, OKT3), helper inducer cells (OKT4), cytotoxic/suppressor cells (OKT8), B cells (BA-1), and monocytes and null cells (OKM1). Double-layer fluorochrome enhancement using F(ab')(2) reagents and nuclear counter staining with ethidium bromide enabled quantitation of the number of positive mononuclear cells, interstitial cells, and total cells on each of 30-55 microscopic fields per tissue section. T cells were the most abundant infiltrating cell in tissues with IR (35 +/- 9.8 percent), significantly higher than that seen in IN (21 +/- 16 percent) or in kidneys without infiltration (5.0 +/- 3.9 percent). The percentage of T cells identified by TA-1 or OKT3 was approximately equivalent to the summation of OKT4 plus OKT8. Although no differences were observed in the percentage of OKT4 cells, the percentage of OKT8 was significantly higher in IR (26 +/- 7.7 percent, P {less than} 10(-4)) than in IN (9.3 +/- 6.2 percent) or in kidneys with normal interstitium (3.0 +/- 2.4 percent). The ratio of OKT8/OKT4-positive T cells in 22 graft tissues with IR (3.2 +/- 1.4) was greater (P {less than} 0.0007) than 6 graft tissues with IN without rejection (0.82 +/- 0.39) and the 5 kidney tissues without interstitial infiltration (0.75 +/- 0.25). There was no significant difference between the groups in the relatively low percentage of interstitial cells identified as B cells reacting with BA-1 or containing S(IgD,M). The percentage of interstitial cells recognized by OKM1 was similar in rejection and interstitial nephritis, with both being greater than controls (P {less than} 0.02). The relative numbers of blood mononuclear cells identified by the monoclonal antibodies was generally not predictive of the proportions present in kidney tissue, although OKT4-positive blood cells were less numerous and OKMI+ blood cells were more numerous than in controls (P {less than} 0.002). Quantitative analysis of identifiable interstitial cells in graft rejection reveals that most infiltrating cells are T cells, the greater proportion of which are recognized by OKT8. OKT8-positive cells may play an important role in mediating renal graft rejection.

Use of monoclonal antibodies to T-cell subsets for immunologic monitoring and treatment in recipients of renal allografts.

Using monoclonal antibodies and flow cytometry, wer serially monitored lymphocyte subpopulations in renal-allograft recipients treated with either conventional immunosuppression or a monoclonal antibody. In 29 patients given conventional suppression, highly significant correlations between changes in T-cell subsets and rejection were noted. Normal or elevated ratios of OKT4 (helper/inducer) to OKT8 (suppressor/cytotoxic) cells were associated with rejection unless the donor was HLA identical or the total number of T cells was extremely low. In patients with low ratios, rejection seldom occurred. Two patients treated with OKT3 monoclonal antibody for acute rejection had rapid disappearance of OKT3-reactive cells from the peripheral blood and prompt reversal of rejection. The use of monoclonal antibodies allows the precise determination of changes in T-cell subsets and promises the development of therapeutic protocols that can be designed to manipulate selected lymphocyte populations.