Synergism between phorbol myristate acetate and calcium ionophore in inducing proliferation of in vitro γ-irradiated murine lymphocytes.

The objective of this study was to analyze the in vitro effects of γ-irradiation (0-5 Gy) on lymphocyte proliferation in animals sensitive to radiation as BALB/c mice. Lymphocytes were irradiated and underwent different treatments: quiescent cells were cultured with calcium ionophore A23187 (5 min or 48 h) with or without phorbol myristate acetate (PMA); lymphocytes (control cells or incubated with A23187 and PMA) were also cultured with four mitogens that are specific to the different subpopulations to determine the degree of inhibition of the response to radiation. Results obtained indicated that in quiescent cells, A23187 and PMA treatment had a mitogenic effect, which peaked with long A23187 treatment (48 h); synergism was further demonstrated between both drugs and was enhanced with higher ionizing radiation doses. However, in both irradiated and non-irradiated mitogen-stimulated cells, A23187 (48 h) and PMA had a strong inhibitory effect on cell proliferation. In conclusion these results indicate that irradiated BALB/c mice lymphocytes respond to treatment with A23187 and PMA more actively than controls. Inhibition of the post-exposure mitogen-induced proliferative response and the synergic effect between A23187 and PMA also suggest altered PKC activation mechanisms in cell membranes. Comparing with previous studies with in vivo irradiated mice, the effects of IR in vitro were less intense.

Immediate and short-, mid- and long-term effects of in vivo ionizing radiation exposure in BALB/c mice: II. activation of phorbol myristate acetate and/or calcium ionophore on lymphocyte proliferation.

AIM: To study the effects of whole-body irradiation (WBI) on lymphocyte proliferation at different times (0, 7, 15, 30, 90 and 180 days) in an animal sensitive to radiation, BALB/c-mice. MATERIALS AND METHODS: Mice were irradiated (4 Gy) and euthanized at different times, and lymphocytes underwent different treatments: quiescent cells were cultured with calcium ionophore (5 min or 48 h) with/without phorbol myristate acetate (PMA). Lymphocytes were cultured with mitogens and underwent the same treatment. Cell proliferation was measured by a tritiated thymidine assay. RESULTS: The results obtained varied at different time points: at 15 days post-irradiation, quiescent cells and PMA-treated cells showed a significantly decreased proliferation, but increased at 90 days; moreover, when cells were treated with ionophore, a significant stimulation was noted at all times. When cells were exposed to mitogens, they behaved according to its nature: thus, concanavalin A (ConA) and phytohaemaglutinin A (PHA) behaved differently with PMA, while lipopolysaccharide (LPS) had an inhibitory effect at 30 days post-irradiation, and pokeweed (PWM) stimulated proliferation at both 90 and 180 days. Accordingly, there were very few variations in the test results when mitogen concanavalin A (ConA) and calcium ionophore with/without PMA were used. CONCLUSION: Our model is based on BALB/c mice. Cells induced to proliferate by the PKC enzyme and calcium ionophore are more resistant to irradiation than the same cells treated with specific T- and B-cells mitogen.

Immediate and short-, mid- and long-term effects of in vivo ionizing radiation exposure in BALB/c mice: I. Activation of lymphocytes and subpopulations.

BACKGROUND: The main aim of this work was to study the effects of single whole-body irradiation (WBI) on the lymphoproliferative response in radiation-sensitive BALB/c mice. MATERIALS AND METHODS: Mice were irradiated (0-5 Gy) and euthanized immediately afterward; other animals were subjected to WBI (4 Gy) and were analyzed after periods of 0-180 days. Splenic cell number, lymphoproliferative response and lymphocyte subpopulations were studied. RESULTS: This study shows that immediately after exposure, an inhibition of the basal mitogen lymphoproliferative response was produced; furthermore, B-cells appear to be more radiosensitive than T-cells. However, up to 90 day's post-irradiation, mice spleens clearly show low cell numbers, and subpopulations and T-cell mitogens did not return to normal, while the basal response and B-cell mitogens peaked on day 15 post-WBI. CONCLUSION: In our model, B-cells regenerate earlier than T-cells, while Th lymphocytes regenerate faster than Tc lymphocytes.