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PubMed1975

Characterization of the lymphocyte surface receptors for Con A and PHA.

Human peripheral blood lymphocytes were iodinated by the lactoperoxidase technique, and the surface receptors for the mitogens concanavalin A (Con A) and phytohemagglutinin (PHA) were analyzed by solubilizing the membrane protein and incubating with mitogen and antimitigen antibody. The resulting precipitates were washed and analyzed by gel electrophoresis. Con A was to bind to about twice as much labeled protein as PHA. Sodium dodecyl sulfate gels showed that multiple surface membrane proteins bind to these mitogens and that the pattern of peaks of Con A receptors in generally similar to the pattern of PHA receptors. Long-term lymphoid tissue culture cells lines also show multiple receptor peaks for mitogens, indicating that the heterogeneity of receptors is not caused entirely by lymphocyte heterogeneity.

Antibodies, Anti-Idiotypic↗
PubMed1979

[Stimulation of proliferation of rat spleen cells, in vitro, by a nonspecific acute inflammatory exudate. Modulation of cell response to phytohemagglutinin (PHA)].

The effect of an acute non-specific inflammatory exudate with mitogenic activity on macrophages in culture has been tested on the spontaneous and PHA-induced DNA synthesis of spleen cells in vitro. Stimulatory effect of this exudate was observed on spontaneous DNA synthesis which was detectable over a range of 1 : 4 to 1 : 4,000 concentrations. After optimal PHA stimulation, an inhibition of mitogen-induced DNA synthesis was observed when the cells were exposed to the highest concentrations (up to 1 : 128) of the exudate. Thereafter, the phenomenon could be reversed and the stimulation was maximal at a concentration of 1 : 2,000. When a sub-optimal dose of PHA was used, the simulatory effect was more pronounced and detected from 1 : 8 up to 1 : 4,000 concentrations.

Acute Disease↗
PubMed1976

Cellular interactions in the proliferative response of human circulating lymphocytes to PHA-analysis of the impaired response in old age.

Peripheral blood lymphocytes (PBL) from old subjects were less responsive to PHA than those from young controls, in both groups the PBL content of macrophages, T and B cells were comparable. Macrophages were identified as peroxydase containing cells, rosette formation with sheep red blood cell was used as a T cell marker and immunoglobulin bearing lymphocytes were considered to be B cells. PBL were separated in two subpopulations of purified T cells (F1 and F2) differing in their responsiveness to PHA and in their ability to cooperate with B cells. Purified B cells had a synergistic effect on the F1 response to PHA whereas they were often suppressive for the corresponding F2 response. Peroxydase containing cells enhanced the response of unfractionated total PBL and of purified T cells in the young subjects. The depressed response of the old age group could be related mainly to a suppressive effect of monocytes and a decreased circulating pool of F1 T cells.

Adult↗
PubMed1975

PHA stimulation of separated human lymphocyte populations.

Lymphocyte preparations from peripheral blood and tonsils were separated into populations enriched with T or B cells by formation of rosettes with SRBC and separation of the rosette-forming and non-rosette-forming populations. T cell-enriched populations were also prepared by nylon column filtration. Using these methods preparations were obtained which comprised 80--95% T or B lymphocytes as determined by E-rosette formation and surface immunoglobulin (Ig) staining. PHA responsiveness, measured by [3H]thymidine incorporation, varied between relatively wide limits and was critically dependent on the degree of separation obtained. Relatively pure B-cell populations (less than 12% T cells) from blood and tonsils gave low PHA responses while preparations from blood still containing 24--38% T cells gave responses equal to or even greater than those of unseparated controls (60--78% T cells). T cell-enriched populations (80--86% T cells) responded to an equal or greater degree than controls but more efficient separation (greater than 90% T cells) resulted in markedly reduced stimulation. There was thus no simple correlation between the degree of phytomitogen-induced transformation and the number of T cells present. It is concluded that the low response of relatively pure T-cell populations may be due to depletion of B cells or non-lymphoid cells (or both) during the separation procedures. These observations have implications for the use of PHA stimulation as a measure of T-cell activity in mixed populations such as those of human peripheral blood leucocytes.

B-Lymphocytes↗
PubMed1992

Inhibition of PHA-stimulated proliferation of peripheral blood mononuclear cells by human seminal plasma inhibin (HSPI).

This report deals with the effect of purified HSPI (10.7 kDa single chain prostatic peptide of 94 amino acids) on PHA-stimulated proliferation of peripheral blood mononuclear cells (PBMNC). HSPI was found to inhibit the incorporation of 3H-thymidine by PHA-stimulated PBMNC in a dose-dependent manner, albeit with the requisite of pretreatment of cells with HSPI. A maximum inhibition was observed on pretreatment for 30 min. These studies thus indicate that HSPI may be interfering with the very early events of PHA-induced signal transduction process in PBMNC.

Cell Division↗
PubMedDate not supplied

[Chediak-Higashi syndrome. Study of lymphocyte transformation stimulated by phytohemaglutinin (PHA)].

The reactivity to PHA stimulation of peripheral blood lymphocytes from a child with a Chediak-Higashi syndrome was studied. Lymphocyte transformation was assayed by tritiated-thymidine incorporation. Patient lymphocytes reacted to much lower concentrations of PHA than normal controls. With an optimal dose of PHA the peak of thymidine uptake occurred on day 2 of incubation instead of day 4 as observed in normals.

Chediak-Higashi Syndrome↗
PubMed1976

[Responsiveness of lymphocytes to stimulation by pha and autologous blasts in acute lymphatic leukemias during remission (author's transl)].

The paper deals with the question how far the lymphocytes of acute lymphatic leukemia patients in remission can be stimulated by PHA and autologous blasts in the MLC-Test. The lymphocyte stimulation by PHA showed considerable variation dependent on the phase of therapy. A significant reduction of the responsiveness to PHA stimulation was found during the prophylactic reinduction treatment with Vincristin and Prednisolon compared with the results during maintenance therapy. The examination of lymphocyte stimulation by conserved autologous blasts in the mixed lymphocyte culture-test yielded positive results in one of three children at the beginning of remission and in 2 of 3 children during the consolidated remission period.

Child↗
PubMed1991

Molecular analysis of cell cycle-related gene expression in anti-HLA class I monoclonal antibody (01.65) treated PHA-activated human T-lymphocytes.

HLA class I antigens seem to be involved in the proliferative response of PHA-activated human T-lymphocytes. We have previously reported that the treatment of PHA-activated peripheral blood mononuclear cells (PBMC) with an anti-HLA class I monoclonal antibody, 01.65, (i) inhibits the tritiated thymidine incorporation, (ii) inactivates cytosolic protein kinase C (PKC) and (iii) causes an increase in the duration of the cell cycle. Northern Blot kinetic analysis of c-fos, c-myc, cdc2, IL-2R, c-myb, ODC, TK and H3, from 10 minutes to 120 hours, was performed in MAb 01.65 treated cultures. We found that the expression of four genes (c-myc, IL-2R, cdc2 and TK) was depressed 24 hours after PHA stimulation.

Antibodies, Monoclonal↗
PubMed1991

Influence of growth hormone-releasing hormone (GHRH) on phytohemagglutinin-induced lymphocyte activation: comparison of two synthetic forms. GHRH and PHA-induced lymphocyte activation.

The in vitro effect of synthetic human growth hormone-releasing hormone (GHRH) on mitogen-induced lymphocyte proliferation and lymphokine secretion was investigated. Peripheral blood mononuclear cells (PBMC) of healthy adults were incubated in the presence and absence of increasing concentrations (from 0.006 to 50 micrograms/ml) of two forms of GHRH differing in amino-acid sequence (GHRH 1-44 and GHRH 1-29) or of increasing concentrations (from 0.0012 to 20 U/ml) of recombinant human insulin (rh-insulin). Low concentrations of GHRH 1-29 increased phytoemoagglutinin (PHA)-induced lymphoproliferation, while high concentrations inhibited lymphocyte response, interleukin-2 (IL-2) secretion and IL-2 receptor expression on activated cells. A toxic effect was excluded since no differences in cell viability were observed between cells cultured with and without hormone. GHRH 1-44 did not affect PHA-induced lymphoproliferation, IL-2 production and IL-2 receptor expression. Low concentrations of rh-insulin increased PHA-elicited lymphoproliferation, while high concentrations did not decrease lymphocyte response. The present study suggests that GHRH modulates in vitro human T lymphocyte functions.

Adult↗
PubMed1991

[The specificity of plague antigen FI in PHA].

It were inoculate a total of the 15 type and 13 group Leptospira strains to the Rattus flavipetus and Rattus lose 137 rates in Lei Zhou peninsula. As the antisera acquired, it has been examined the lyso-agglutinative test in Leptospira strains. Up to standard serum it was to test for PHA of plaque, employing the sensitized blood cell by FI from 4 antiplague institutes. All of tests are negative. In addition to test for 3 immune sera, Hemorrhagic Fever antisera (Immunized animals Apodemus agraius, Rattus norvegicus, Mouse and Rabbit), anthrax antiserum, there were be negative by PHA of plague test, at same time by RIA test also be negative. Therefore all of the other antisera would not be interfering with specificity of the plague FI antiserum in PHA or RIA test.

Animals↗
PubMed1991

[The cancer immunotherapy controlled by the number of PHA-blastogenic cells].

Old BRMs have been conventionally evaluated by comparative studies between the cancer patients treated with or without BRMs. This comparison should be done under the assumption that effectiveness of BRMs is dose-dependent. However BRMs can not be effective without enhancing cellular immunity of the patients. We have demonstrated that PHA blastogenic reaction was not necessarily augmented in vitro in a dose-related manner by MRMs. Nor did the dose-dependent immuno-enhancement always occur by clinical use of BMRs, if the blastogenic ratio was converted to the number of blastogenic cells per mm3 of peripheral blood to monitor the patient's cellular immunity more precisely. If BRMs should be reviewed in terms of the immuno-enhancing activity by the number of PHA blastogenic cells, the clinical needs of BRMs must be increased. It is sure that the patient's survival could be improved if immunotherapy should be done with varying doses or schedule individually by using the adequate immuno-parameters such as PHA blastogenic reaction.

Female↗
PubMed1990

Suppression of delayed-type hypersensitivity to PPD and PHA in elderly HTLV-I carriers.

In a previous study on immune responsiveness among asymptomatic human T-cell leukemia virus type I (HTLV-I) carriers, we found that carriers had significantly reduced delayed-type hypersensitivity (DTH) response to purified protein derivative (PPD) skin testing. The association was strongest among persons at least 60 years of age. In order to evaluate this finding further, we evaluated the response to both PPD and phytohemagglutinin (PHA) in an elderly population. Fifty-six consecutive hospitalized patients with nonimmunosuppressive diseases were examined. None had a history of tuberculosis nor evidence of the known HTLV-I-associated diseases. The subjects' ages ranged from 62-93 years (median = 75 years); 43 were women and 13 were men. Twenty-two of the subjects were HTLV-I antibody positive. Among the carriers, there was an increased level of nonreactivity to PPD, the relative risk adjusted for age (RR) being 1.9 (95% confidence interval, 0.62-5.8), as well as to PHA of RR = 2.3 (0.60-9.0). When subjects were cross-classified for response to both skin tests, 15 of 17 carriers were nonreactive to either or both antigens compared to 15 of 25 noncarriers [RR = 5.1 (0.99-25.9) (p value, one-sided = 0.026)]. The decline in reactivity to both antigens increased with age, but was consistently lower among the carriers. Among subjects with positive reactions to PPD, the degree of reaction as measured by the size of erythema was reduced among the carriers; however, for PHA responders, the response in carriers appeared to be normal. Among the HTLV-I antibody negative subjects, the size of erythema for both antigens was strongly correlated (p = 0.01).(ABSTRACT TRUNCATED AT 250 WORDS)

Aged↗
PubMed1989

PHA stimulation of peripheral blood lymphocytes in oral lichen planus. Abnormality localized between interleukin-2 receptor ligand formation and gamma-interferon secretion.

Peripheral blood T cell function in five oral lichen planus (OLP) patients and five healthy controls was assessed using different activation parameters. Staining with monoclonal antibodies against interleukin-2 receptor and MHC locus II coded Ia antigen, 3H-thymidine incorporation and gamma-interferon secretion were determined in phytohaemagglutinin (PHA) stimulated peripheral blood mononuclear cell cultures at days 0, 1, 3 and 5. The peripheral blood T cell subsets and spontaneous MHC locus II antigen expression were similar in OLP patients and in controls whereas the spontaneous lymphocyte proliferation was lower in OLP patients than in controls (p less than 0.01). This may reflect a slight in vivo preactivation and its effect on lymphocyte recirculation. The PHA-induced expression of IL-2 receptor and T cell proliferation were similar in both groups whereas gamma-interferon secretion and MHC locus II antigen expression were low in OLP patients compared with controls (p less than 0.01). The results suggest a defect in OLP T cell activation disclosed by in vitro PHA stimulation and localized between IL-2 receptor ligand binding and gamma-interferon secretion.

HLA-D Antigens↗
PubMedDate not supplied

Modulating effect of growth hormone (GH) on PHA-induced lymphocyte proliferation.

In order to investigate the ability of growth hormone (GH) to modulate lymphoproliferation, peripheral blood mononuclear cells (PBMC) from healthy adults were stimulated with phytohaemagglutinin (PHA) in the presence of increasing concentrations of pituitary (Crescormon) and biosynthetic (Somatonorm and Genotropin) GH (from 0.425 x 10(3) to 501.5 x 10(3) ng/ml) as well as biosynthetic insulin (rhinsulin, Humulin 1 Eli Lilly: from 0.0012 to 20 U/ml). The results obtained show that low doses of GHs and insulin increase lymphoproliferation. In contrast, high concentrations of GHs (8.5 x 10(3) ng/ml) induce a progressive decrease of PHA-induced lymphoproliferation, while insulin does not. No differences in cell viability were observed with and without GHs. Comparable percentages of lymphocytes bearing interleukin-2 receptor (CD25) and comparable values of interleukin-2 (IL-2) pr oduction were found in all conditions tested. Nevertheless, when exogenous recombinant IL-2 (R IL-2) was added to PHA-induced PBMC cultures with high doses of GH, a reconstitution of lymphoproliferation was observed. Further experiments seem to exclude the presence of suppressor T lymphocyte induction by GH. The present results suggest that hypothesis that high concentrations of GH may interfere with the binding between IL-2 and its membrane receptor on activated lymphocytes.

Adult↗
PubMed1988

Changes in activation markers and cell membrane receptors on human peripheral blood T lymphocytes during cell cycle progression after PHA stimulation.

Phytohaemagglutinin (PHA)-stimulated peripheral blood lymphocytes were examined sequentially for changes in volume, the appearance of cell membrane receptors and nucleic acid synthesis. The kinetics of appearance of activation antigens were compared with the progress of the cell through the separate events of volume growth and nucleic acid syntheses, to determine points at which regulation of receptors may control further progress through the cell cycle. In all samples tested there was a consistent pattern of response in the proportion of cells progressing through the cell cycle. Most of the T cells increased in size (mean 82% at 24 hr), fewer cells entered the Gla/Glb phase with the onset of RNA synthesis (mean 68% at 48 hr) and even fewer entered DNA synthesis (mean 42% at 72 hr). The time-course of appearance and the number of cells expressing IL-2 receptors were almost identical with that of cells responding by RNA synthesis. A similar correlation was observed between expression of the transferrin receptor and DNA synthesis. Addition of anti-Tac antibody temporarily suppressed the onset of RNA synthesis and antibodies to the transferrin receptor suppressed DNA synthesis. These linkages are further evidence that IL-2 and transferrin are the specific signals for cellular RNA and DNA synthesis. With optimal concentrations of PHA, addition of IL-2 did not increase the proportion of cells bearing activation antigens or undergoing nucleic acid synthesis. Suboptimal concentrations of PHA produced a small reduction in the number of cells expressing the IL-2 receptor, but a much greater reduction in the rate of entry into RNA synthesis. There was a consistent increase in all activation parameters tested with the addition of IL-2, but the proportion of cells expressing the transferrin receptor and entering DNA synthesis was consistently lower than that of cells that expressed the IL-2 receptor or entered RNA synthesis. This suggests that regulation of the IL-2 receptor is not responsible for the reduction in the number of cells that proceed to proliferation. The CD2 antigen (T11(1] showed increasing expression in a step-wise fashion after activation, the increases coinciding with the onset of RNA and DNA syntheses.

Cell Cycle↗
PubMed1988

Mechanisms by which oxidative injury inhibits the proliferative response of human lymphocytes to PHA. Effect of the thiol compound 2-mercaptoethanol.

The use of normobaric exposure to O2 as a model for in vitro oxidative injury prevented phytohaemagglutinin (PHA)-stimulated human peripheral blood mononuclear cells (PBMC) from undergoing the G0 to G1 transition, but 5 x 10(-6) M 2-mercaptoethanol (2-ME) almost protected the cells from this blockade. The percentage of cells with IL-2 and transferrin-receptors was reduced by the O2 exposure and, like the cell cycle transition, was protected by 2-ME against oxidative injury. By contrast, IL-2 recovery in the supernatants of O2-exposed PHA-stimulated PBMC was enhanced. This enhancement may be due partly to the reduced IL-2 consumption caused by the decreases in IL-2 receptor expression and in proliferation. On the other hand, IL-2 recovery in the supernatants of O2-treated PBMC was always enhanced compared to the IL-2 control recovery after DNA synthesis was blocked in G1/S by mitomycin c, and the G0/G1 transition was protected by 2-ME. Furthermore, PHA-stimulated monocytes exposed to O2 produced more IL-1 than control cells. This enhanced IL-1 production was not modified by 2-ME. These results suggest that oxidative injury reduces the proliferation of PBMC by interfering with the cellular events that lead to the transition from the G0 to the G1 phase of the cell cycle. The protective effects of 2-ME suggest that thiol compounds have a critical role in the early events of the cell cycle. By contrast, exposure to O2 induced increases in the production of both IL-1 and IL-2 that may not be related to alterations in the thiol status of the cell.

Cell Cycle↗
PubMed1987

Immune dysfunction in primary biliary cirrhosis (PBC): I. Increased sensitivity of PHA stimulated lymphocyte cultures to indomethacin.

A reduced responsiveness of peripheral blood lymphocytes from PBC patients to phytohaemagglutinin (PHA) was observed. Such an alteration may be related either to an intrinsic defect of T cells to proliferate or to a derangement of endogenously secreted mediators with regulatory functions, i.e., prostaglandins (PG). The role of the PG mediated suppressor system in 14 PBC patients and in 12 age and sex matched controls was evaluated. After the addition of indomethacin, the lymphocyte responsiveness to PHA increased in both controls and in PBC patients. However, this increment was statistically significantly only in PBC patients. The addition of PGE1 and PGE2 to PHA stimulated cultures decreased the lymphocyte proliferative activity. No differential sensitivity to PGEs was detectable in lymphocytes from PBC patients. Our data suggest, that, after polyclonal stimulation, mononuclear cells from PBC patient may release increased levels of PGEs which in turn affect the proliferative activity of mitogen activated lymphocytes.

Adult↗
PubMed1985

The decline in murine splenic PHA and LPS responsiveness with age is primarily due to an intrinsic mechanism.

Changes in splenic B and T lymphocyte number and mitogenic activity with age were quantitated in (A X C57BL/6)F1 (AB6F1) hybrid mice. Although both the B and T lymphocyte proliferative reactivity to their respective mitogens, lipopolysaccharide (LPS) and phytohemagglutinin (PHA), declined significantly with age, an earlier and more marked reduction was recorded for the T cell response. The decline in B and T lymphocyte mitogenic activity with age could not be correlated with a corresponding reduction in the percentage of splenic B or T lymphocytes. The main focus of this study was to determine if the reduction in T and B lymphocyte mitogenic activity with age results primarily from a mechanism intrinsic to the lymphoid lineage itself or from adverse extracellular factors that increase with age. Bone marrow cells (BMC) derived from individual young and old donor AB6F1 mice were transplanted into the neutral environment of young, lethally irradiated syngeneic recipients. Number and mitogenic activity of splenic T and B lymphocytes were recorded for the original BMC donors as well as for the recipients of the young and old BMC lines 9 mo after the BMC transplants. A predominance of the donor (male) rather than recipient (female) karyotype within the mitogen-responding populations of recipient mice confirmed a donor BMC take. The PHA and LPS response levels exhibited by the old donors were 30% and 70% of those of the young donors, respectively. These differences in PHA and LPS reactivity recorded between young and old donors were maintained between recipients of young and old donor BMC lines. Thus, even under the influence of a young recipient environment, old BMC were incapable of giving rise to mitogen responding cells with a functional competence equivalent to that of their younger counterparts. This finding would lend further support to the theory that an intrinsic mechanism is responsible for the decline in murine mitogenic activity with age.

Aging↗