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At least 649 records · Page 36Linked to original sources
PubMed1980

Conditions for measuring DNA synthesis in PHA stimulated human lymphocytes in 20 microliters hanging drops with various cell concentrations and periods of culture.

We have studied conditions for measuring the uptake of [3H]thymidine ([3H]Tdr) by human lymphocytes in inverted microcultures, varying cell concentrations and periods in culture. Analysis of variance of the log values for [3H]Tdr uptake may be used to separate effects of variables and their interactions. A pulse time of 2 h, a total thymidine concentration of about 1 microgram/ml and a specific activity of [3H]Tdr of 2 Ci/mmole were optimal. Variables such as cell concentration, period of culture, type of serum and dose of PHA were shown to interact, suggesting that these variables should be examined together especially when comparing different samples of lymphocytes. Conditions for doing this simply in small volumes are now available for culturing, thymidine pulsing, harvesting and analysis of data.

Cell Division↗
PubMed1984

A new ultra-microculture system. I. Stimulation of human T lymphocytes by phytohemagglutinin (PHA).

We have developed an ultra-microtechnique for culturing lymphocytes in glass capillary tubes at a final culture volume of 1 microliter or 2 microliter. The advantage of the method is that a substantially lower number of cells and minute amounts of culture medium are required. The cultures are premixed in microtubes, sucked into glass capillary tubes and incubated for an appropriate culture period. For determination of [3H]thymidine ([3H]TdR) incorporation, the cells are transferred into the wells of microtiter plates. Some special accessories have been developed which allow routine use of this system for large numbers of cultures. Optimal culture conditions for stimulation of human T lymphocytes by PHA are described.

Blood Physiological Phenomena↗
PubMed1985

Blotting methods applied to investigations of the mitogenic activity of phytohaemagglutinin (PHA).

PHA polypeptides were separated by SDS-polyacrylamide gel electrophoresis and transferred to nitrocellulose filters which were then cut into identical discs and used to perform blastogenic stimulation on cultured human peripheral blood lymphocytes. This method offers the following advantages: the high resolution of SDS-polyacrylamide gel electrophoresis, the sensitive immunochemical identification of the polypeptides involved in the mitogenic responses of lymphocytes, the determination in the same experiment of the molecular weight, antigenicity and biological activities of the proteins.

Collodion↗
PubMed1988

Typing of HLA class II and class I antigens using PHA-activated, IL-2-propagated T lymphocytes.

We describe here a simple procedure, by which HLA class II antigens can be accurately and reliably identified in those patients where there is minimal or absent expression of HLA-DR,DQw antigens on B cells, or when the total number of leukocytes recovered from the patients do not permit reliable typing. Ficoll-Hypaque-separated peripheral blood mononuclear leukocytes, fresh or cryopreserved, were activated by PHA and then propagated in IL-2-containing medium until enough cells for typing were obtained (usually 7-14 days). At this stage, the cultured cells were shown to be primarily T cells (greater than 90% CD3+). Since the activated T cells propagate in the presence of IL-2, even a small number (10(4] of fresh or cryopreserved patients' cells suffice for this protocol. To date we have been able to successfully HLA-DR,DQw type 34/34 bone marrow transplantation candidates and 12/12 long-term dialysis patients, who were untypable using fresh cells. HLA-DR,DQw antigens on activated T cells from normal individuals were identical to those found on their uncultured B cells. In addition, class I antigens that were undetectable on the uncultured cells of one patient could be identified on activated T cells. The HLA antigens identified on the patients' activated T cells were confirmed by phenotypic analysis of cells from family members.

B-Lymphocytes↗
PubMed1976

Lack of synergistic effect between X-ray and UV irradiation on the frequency of chromosome aberrations in PHA-stimulated human lymphocytes in the G1 stage.

PHA-stimulated human lymphocytes in the G1 stage were irradiated with UV radiation and X-rays, and the cells were analyzed for chromosomal aberrations in the first mitotic division. The frequency of dicentric chromosomes after single X-irradiation in the G1 stage was about twice the yield in the G0 stage. No increase in the yield of dicentrics was observed after combined irradiation with UV and X-rays. This is contrary to the finding for G0 lymphocytes, where a 2-fold increase of chromosome aberrations was observed. UV irradiation of G1 lymphocytes induced chromatid-type aberrations whereas no significant yield of dicentric chromosomes was observed. This is in agreement with previous findings in Chinese hamster cells in the G1 stage [7]. Irradiation of G0 lymphocytes with UV radiation induce a low frequency of dicentric chromosomes. Thus, the present data indicate that the ratio between chromosome-type and chromatid-type aberrations is different in the G1 and G0 stages in human lymphocytes irradiated with UV radiation.

Chromosome Aberrations↗
PubMed1983

Studies on endoreduplication. III. Endoreduplication induced by mitomycin C in PHA-stimulated tonsillar lymphocyte cultures.

The activity of mitomycin C (MMC) to induce endoreduplication was examined in PHA-stimulated tonsillar lymphocyte cultures. Cellular kinetics of both diploid and endoreduplicated cells was studied by the combined techniques of thymidine and BrdU incorporation. The kinetic data showed that the increase of endoreduplicated cells induced by MMC correlated with the increase of the second-generation diploid cells. Also, it showed that the majority of endoreduplicated cells found at the time when the maximal number of endoreduplicated cells was obtained had incorporated BrdU only once into their diplochromosomes. This suggests that MMC-induced endoreduplicated cells originate from MMC-induced G2-arrested cells.

Cells, Cultured↗
PubMed1978

Cytochemical study of human lymphocytes stimulated by PHA in function of donor age.

The 3H-AM binding reflects the structural changes involved in the cellular differentiation. This parameter was studied during blastic transformation of human lymphocytes, in relation to the age of the donor. Although they are individual variations, the 3H-AM binding is higher in the young group than in the aged subjects, as well as the blast transformation score. These results indicate that the weak lymphocyte response to the mitogen lectine (PHA) stimulation could be related to some age-induced structural alterations of the chromatin, resulting in an irreversible blockage in G1, at least in some of the T lymphocytes.

Adult↗
PubMed1984

PHA-ICC, ADCC and NK in patients with ANLL in CR: human fibroblastic interferon fails to increase NK-active cell frequency.

PHA-ICC, ADCC and NK activity of PBL were studied in ten patients with ANLL in CR and in eighteen normal controls in the presence and absence of HFIF. No statistically significant differences were recorded among the two groups with regard to basic lymphocyte functions. Although the parameters of lymphocyte function remained analogous for those tested, the analysis at the single cell level revealed that HFIF stimulation increases the number of NK active cells and target binding cells among normals, but not in leukemic patients.

Adult↗
PubMed1994

PHA induces IL-2 receptors on B-CLL cells and is a potential biological response modifier for the LIL-2-diphtheria toxin, DAB486IL-2.

DAB486IL-2 is an IL-2-diphtheria toxin conjugate which was developed to be specifically cytotoxic to cells bearing high affinity IL-2 receptors. The high affinity IL-2 receptor is a heterodimer comprising p55 and p75 subunits. While the p75 subunit appears to be ubiquitously expressed among the common North American leukemias and lymphomas, the p55 subunit is more restricted in its expression. To broaden the therapeutic relevance of the DAB486IL-2 we have sought physiologically feasible inducers of the p55 IL-2 receptor subunit. This report describes that PHA, in vitro, induces the p55 IL-2 receptor subunit on initially p55-negative B-CLL cells and converts toxin-insensitive leukemia cells to a toxin-sensitive state.

Diphtheria Toxin↗
PubMed1979

Increased development of PHA-stimulated murine colonies in vitro subsequent to adriamycin or daunorubicin treatment of spleen donor mice.

Spleen cell populations from mice treated with Adriamycin or daunorubicin were found to develop a greater PHA-stimulated clonal response than spleen cells from untreated or cyclophosphamide-treated animals. Daunorubicin, in addition, stimulated an unusual monocytic macrophage-like colony. The results are consistent with the possibility that the concentration of nonspecific T-lymphocyte progenitor and/or accessory cells in the spleen are increased consequent to drug treatment.

Animals↗
PubMed1979

In vitro studies in reproductive immunology. - 2. Demonstration of the inhibitory effect of male genital tract constituents on PHA-stimulated mitogenesis and E-rosette formation of human lymphocytes.

The effect of male genital tract components (human spermatozoa, intact and chromatographed seminal plasma fractions) on in vitro cell-mediated immune reactions was examined. Their addition to PHA-containing lymphocyte cultures resulted in a marked degree of inhibition of DNA synthesis, the response varying with the fractions employed. The interference by such components with a terminal cell event during blast transformation was suggested by the failure of stimulated cultures to incorporate thymidine irrespective of the time during the incubation period at which the constituents were added. Seminal plasma and sperm extracts were also shown to inhibit T-cell associated E-rosette formation. The inhibitory properties of genital fractions remained unaltered after repetitive freeze-thaw cycling when tested in the transformation and E-rosette assays. Heating to 100 degrees C modified the response with some of the fractions.

Humans↗
PubMed1991

Immunoregulatory effects of ovine trophoblastin protein (oTP): all five isoforms suppress PHA-induced lymphocyte proliferation.

The immunosuppressive properties of ovine trophoblastin protein (oTP) isoforms purified to homogeneity by DEAE HPLC have been studied within and across species barriers by in vitro assays. It has been demonstrated that not only the classical oTP 1, but in fact all 5 isoforms, are immunosuppressive in a PHA-induced proliferation assay, whilst being ineffective on IL-2 dependent CTL-L2 cell replication. The significance of these findings is discussed.

Animals↗
PubMed1989

The effect of immunomodulators on PHA or gamma-IFN induced release of neopterin from purified macrophages and peripheral blood mononuclear cells.

Immunomodulators cause changes in neopterin-release from purified macrophages or peripheral blood mononuclear cells by affecting the macrophage and T cell subsets activity, the intracellular cGMP/cAMP balance, or the intracellular pteridines-related biochemical pathways. Increased neopterin release was achieved by gamma-IFN or its inducers (PHA, IL-2), by interfering with the biopterin production by increased levels of cGMP or by decreasing the activity of the T suppressor cells. The released neopterin levels decreased due to decreased macrophage and T-helper cell activity or due to increased levels of cAMP. The in vitro effect of the immunomodulators has to be taken into account when assessing the neopterin levels in immunomodulators-treated patients.

Adjuvants, Immunologic↗
PubMed1992

Valid estimation of IL2 secretion by PHA-stimulated T-cell clones absolutely requires the use of anti-CD25 monoclonal antibody to prevent IL2 consumption.

A major problem encountered for quantification of IL2 production by stimulated T cells is its simultaneous consumption by these activated cells. In the present study, 40 T-cell clones (TCC) derived from normal peripheral blood, hyperplastic lymph nodes (LN) or lymph nodes involved by malignant lymphomas, were studied for their ability to produce IL2. When supernatants were generated in the presence of 20% fetal calf serum (FCS), no IL2 could be detected for 22 of the 40 TCC, whereas very low levels were found for the 18 other TCC (mean value 31 pg/ml; range from 10 pg/ml to 114 pg/ml); in contrast, when conditioned media were produced with reduced amounts of FCS (final concentration, 1%) as well as in the presence of an anti-CD25 monoclonal antibody (final concentration, 50 micrograms/ml), all TCC were found to release IL2, and very high quantities of this lymphokine were measured (mean value: 11,387 pg/ml; range, from 250 pg/ml to 37,000 pg/ml). Consequently, inhibition of IL2 consumption by PHA-stimulated TCC seems to be an absolute requirement for estimating the true capacity of T cells to produce this lymphokine.

Animals↗
PubMed1982

A clinical assessment of fluorescence polarisation changes in lymphocytes stimulated by phytohaemagglutinin (PHA) in malignant and benign diseases.

The modified double-zone SCM technique, developed in these laboratories as an in vitro test for cancer, is based on the differential response to PHA of lymphocytes harvested from two regions of a Ficoll-Triosil gradient. Lymphocyte responses are measured by changes in intracellular fluorescein fluorescence polarisation. We report its continued clinical evaluation in patients hospitalised with malignant and non-malignant diseases, including a blind trial of 78 patients with disorders of the gastrointestinal tract, breast and lung. Overall "false' negative and "false' positive rates from 336 blood samples were 1.8 and 3.3% respectively.

Adult↗
PubMed1984

Amygdalostriatal projections in the rat. Topographical organization and fiber morphology shown using the lectin PHA-L as an anterograde tracer.

Amygdalostriatal projections in the rat were studied with the anterograde tracer phaseolus vulgaris leucoagglutinin (PHA-L), which allows direct visualization of axon morphology. The appearance of fibers in the striatum which were labeled from injections into the amygdaloid complex suggests that they form 'en passant' synapses. A topographic organization was found in the projection from the basolateral amygdaloid nucleus: the most caudal part of this nucleus projects to the anteromedial edge of the striatum, including the medial nucleus accumbens, while progressively more rostral parts of the nucleus project to more lateral and caudal portions of the striatum. No amygdaloid fibers were found in the most rostro-dorsolateral part of the caudatoputamen.

Amygdala↗
PubMed1990

Arborization of corticothalamic axons in the auditory thalamus of the cat: a PHA-L tracing study.

As a result of Phaseolus vulgaris leucoagglutinin (PHA-L) injection in the primary auditory cortex of the cat, portions of corticofugal axons could be traced individually in the reticular complex of the thalamus (RE) and in the medial geniculate body (MGB). On their way to the MGB, axons were seen to give off collaterals which terminated in RE. In the MGB, an individual corticofugal axon was fully reconstructed. It ramified at 3 distinct frontal levels in the rostral half of the pars lateralis (LV). Its terminal branches gave rise to boutons en passant and terminaux in a stripe of LV (parallel to the isofrequency contours), overlapping a cluster of retrogradely labeled neurons, supporting at the microscopic level the principle of the reciprocity of thalamocortical and corticothalamic projections.

Animals↗