Augmentation of Human Thymocyte Proliferative Responses by Retinoic Acid

Sidell, Neil; Famatiga, Estela; Golub, Sidney H.

doi:10.1159/000163828

Cited by 18 publications

(11 citation statements)

References 13 publications

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“…NK cells were separated in the LGL fraction of human peripheral blood mononuclear cells as described (3,14). Following separation by discontinuous Percoll density centrifugation, cells were further depleted of T lymphocytes by sheep erythrocyte-rosette formation, and this highly enriched population (>90% LGL) (9) was used for electrical recording.…”

Section: Methodsmentioning

confidence: 99%

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Potassium channels mediate killing by human natural killer cells.

Schlichter¹,

Sidell²,

Hagiwara³

1986

Proc. Natl. Acad. Sci. U.S.A.

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Human natural killer (NK) cells in peripheral blood spontaneously recognize and kill a wide variety of target cells. It has been suggested that ion channels are involved in the killing process because there is a Ca-dependent stage and because killing by presensitized cytotoxic T lymphocytes, which in many respects resembles NK killing, is associated with changes in K and Na transport in the target cell. However, no direct evidence exists for ion channels in NK cells or in their target cells. Using the whole-cell variation of the patch-clamp technique, we found a voltage-dependent potassium (K+) current in NK cells. The K+ current was reduced in a dose-dependent manner by the K-channel blockers 4-aminopyridine and quinidine and by the traditional Ca-channel blockers verapamil and Cd2 . We tested the effects of ionchannel blockers on killing of two commonly used target cell lines: K562, which is derived from a human myeloid leukemia, and U937, which is derived from a human histiocytic leukemia.Killing of K562 target cells, determined in a standard 51Cr-release assay, was inhibited in a dose-dependent manner by verapamil, quinidine, Cd2+, and 4-aminopyridine at concentrations comparable to those that blocked the K+ current in NK cells. In K562 target cells only a voltage-dependent Na' current was found and it was blocked by concentrations of tetrodotoxin that had no effect on killing. Killing of U937 target cells was also inhibited by the two ion-channel blockers tested, quinidine and verapamil. In this cell line only a small K+ current was found that was similar to the one in NK cells. We could not rind any evidence of a Ca21 current in target cells or in NK cells; therefore, our results cannot explain the Ca dependence of killing. Our findings show that there are K channels in NK cells and that these channels play a necessary role in the killing process. In contrast, the endogenous channel type in the target cell is probably not a factor in determining target cell sensitivity to natural killing.

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Section: Methodsmentioning

confidence: 99%

“…For assessing natural killing we used the standard 51Cr-release assay (14). Percent specific lysis was defined as the percent specific 51Cr released, or [(experimental release -spontaneous release)/(maximal releasespontaneous release)] x 100.…”

Section: Methodsmentioning

confidence: 99%

Potassium channels mediate killing by human natural killer cells.

Schlichter¹,

Sidell²,

Hagiwara³

1986

Proc. Natl. Acad. Sci. U.S.A.

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“…Phytohemagglutinin (PHA) was freshly mixed from stock solutions to a final concentration of 10 ,ug/ml in standard recording solution (see below). The mitogenic activity of the PHA was confirmed by its ability to induce blastogenesis of peripheral blood lymphocytes in a standard 3-day assay as described (11). The standard bathing solution contained (in mM) 150 NaCl, 5 KCl, 2.5 CaCl2, 1 MgCl2, 10 D-glucose, 10 Hepes, and 4 NaOH (pH 7.4).…”

Section: Methodsmentioning

confidence: 99%

K channels are expressed early in human T-cell development.

Schlichter¹,

Sidell²,

Hagiwara³

1986

Proc. Natl. Acad. Sci. U.S.A.

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Mature human T lymphocytes proliferate in response to the mitogen phytohemagglutinin (PHA), but immature thymocytes lacking the T3 receptor (T3-thymocytes) do not. Because functioning K channels are required for proliferation of mature T cells, we asked whether immunoincompetent T3-thymocytes lack normal K channels. We report that T3-thymocytes have a K+ current similar to that of mature peripheral T cells-that is, similar voltage dependence, activation and inactivation kinetics, and pharmacology. Moreover, the maximal specific K+ conductance is the same for both cell types, implying a similar density of activable channels in each cell. In assessing the functional responses of the channels to PHA, we found that the K+ current ofimmature and mature cells responds similarly to the mitogen. Responses near the threshold voltage for activating the K+ current were variable; the K+ conductance and rate of activation were increased, decreased, or unchanged after PHA treatment. For several cells, the voltage dependence of the conductance and activation kinetics was shifted in opposite directions. At more positive voltages, PHA consistently caused a 10-20% suppression of conductance that was not due to the addition of an inward current, to changes in the time course of activation or inactivation, or to changes in the steady-state level of inactivation. The effects of PHA on the K+ current cannot be explained by a simple shift in surface potential, as has been hypothesized to be involved in its triggering of T-cell proliferation. Taken together, our findings show (i) K channels are expressed very early in T-cell differentiation, possibly before thymic processing, (i) differential responses of the K+ current to PHA do not account for the failure of T3-thymocytes to proliferate, and (iii) changes in surface potential are probably not a necessary early event in activation of T cells by PHA.

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“…r3H]thymidine incorporation L'HIThymidine incorporation of treated and control LA-N-5 cells, as an index of DNA synthesis, was performed as previously described (Sidell et al, 1981(Sidell et al, , 1983. Briefly, cells (10' cells/well) were plated in quadruplicate wells of flat-bottomed microtest plates (M.A.…”

Section: Measurement Of Cytosolic Free Ca2+ Concentrationmentioning

confidence: 99%

Menthol blocks dihydropyridine‐insensitive Ca²⁺ channels and induces neurite outgrowth in human neuroblastoma cells

Sidell

Verity

Nord³

1990

Journal Cellular Physiology

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Voltage-gated Ca2+ channels were identified in LA-N-5 human neuroblastoma cells using the Ca2+ sensitive fluorescent probe, fura-2. Using a variety of "classical" Ca2+ channel blockers, we have demonstrated the presence of both dihydropyridine (DHP)-sensitive and -insensitive channel types that can be activated by depolarization of the cells with either high K+ or gramicidin in the bathing solution. Brief exposure of LA-N-5 cells to menthol blunted the depolarization-induced Ca2+ influx though both DHP-sensitive and DHP-insensitive channels. This effect is concentration dependent (50% maximal blocking effect with 0.25 mM menthol), rapid in onset, and readily reversible. The specificity of the Ca2(+)-channel blocking effect of menthol was demonstrated in parallel studies using compounds with similar structures: menthone blocked Ca2+ channels with about half the potency of menthol, while cyclohexanol was without effect. Addition of either menthol or menthone to LA-N-5 cultures induced neurite outgrowth, cellular clustering, and reduction of cell growth in a dose-dependent fashion that correlated with the ability of these compounds to inhibit the DHP-insensitive Ca2+ influx. Cyclohexanol had no biologic activity. Taken together, the parallel potency for blockade of DHP-insensitive Ca2+ influx with the biologic activity of menthol suggests a role for certain types of Ca2+ channels in triggering growth and morphologic changes in LA-N-5 cells.

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Augmentation of Human Thymocyte Proliferative Responses by Retinoic Acid

Cited by 18 publications

References 13 publications

Potassium channels mediate killing by human natural killer cells.

Potassium channels mediate killing by human natural killer cells.

K channels are expressed early in human T-cell development.

Menthol blocks dihydropyridine‐insensitive Ca²⁺ channels and induces neurite outgrowth in human neuroblastoma cells

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Augmentation of Human Thymocyte Proliferative Responses by Retinoic Acid

Cited by 18 publications

References 13 publications

Potassium channels mediate killing by human natural killer cells.

Potassium channels mediate killing by human natural killer cells.

K channels are expressed early in human T-cell development.

Menthol blocks dihydropyridine‐insensitive Ca2+ channels and induces neurite outgrowth in human neuroblastoma cells

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Product

Resources

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Menthol blocks dihydropyridine‐insensitive Ca²⁺ channels and induces neurite outgrowth in human neuroblastoma cells