Antiproliferative effects of δ opioids on highly purified CD4+ and CD8+ murine T cells

N. A. Shahabi, Burt Sharp

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Abstract

Numerous studies have shown that opioids modulate the proliferative response of mixed splenocytes to T cell mitogens. To identify the T cell subpopulations affected by opioids, splenocytes from C57BL/6 and CD1 mice were separated using a fluorescent activated cell sorter (FACS) to obtain 98 to 99% pure populations of either CD4+ or CD8+ T cells. Cells were stimulated to proliferate in serum-free medium by cross-linking the T cell receptor using plate-coated anti-CD3-ε, then 3H-thymidine uptake and cell number were measured at 48 and 72 hr. [D-Ala2]-deltorphin 1 (deltorphin) dose-dependently inhibited the proliferation of C57BL/6 CD4+ T cells by approximately 50%. This effect was maximal when cells were preincubated with deltorphin 60 min before activation, whereas deltorphin was ineffective when added at the time of activation. Similarly, [D-Ala2]-Met-Enkephalinamide (DAME) 10-11 to 10-7 M inhibited CD4+ T cell proliferation. Naltrindole 10-12 M abolished the antiproliferative effect of 10-7 M deltorphin on CD4+ T cells. Proliferation of CD8+ T cells from C57BL/6 mice also was dose-dependently inhibited by deltorphin. At all concentrations of deltorphin, the antiproliferative effects were greater after 48 compared to 72 hr in culture. The effect of deltorphin and DAME on secretion of the T cell growth factor, IL-2, was determined by ELISA analysis of supernatants obtained from CD4+ T cells after 48-hr culture. Deltorphin showed a biphasic effect: 10-11 M enhanced IL-2 secretion, whereas higher concentrations (10-9-10-7 M) were inhibitory. IL-2 secretion was inhibited significantly by all concentrations of DAME (10-13-10-7 M); however, a biphasic response was observed, with greatest inhibition at both the lowest (10-13 M) and highest (10-7 M) concentrations. Using CD4+ T cells obtained from CD1 mice, it was also found that deltorphin, DAME and 2,5 DPDP-enkephalin dose-dependently inhibited proliferation. The antiproliferative effects of deltorphin and DAME (10-7 M) were reduced when CD4+ T cells were stimulated by increasing concentrations of anti-CD3-ε. In summary, δ opiate receptor agonists inhibited proliferation of both CD4+ and CD8+ T cells from two strains of mice. IL-2 secretion from CD4+ T cells also was suppressed by the δ opiate receptor agonists. Inasmuch as these studies used highly purified T cells lacking detectable macrophages, the effects of the δ opiate receptor agonists appear to be mediated by receptors present on quiescent T cells.

Original languageEnglish (US)
Pages (from-to)1105-1113
Number of pages9
JournalJournal of Pharmacology and Experimental Therapeutics
Volume273
Issue number3
StatePublished - Jan 1 1995

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Opioid Analgesics
T-Lymphocytes
Interleukin-2
Opioid Receptors
naltrindole
deltorphin
Enkephalins
Serum-Free Culture Media
T-Cell Antigen Receptor
Inbred C57BL Mouse
Mitogens
Thymidine
Cell Count
Enzyme-Linked Immunosorbent Assay
Macrophages
Cell Proliferation
Met-enkephalinamide

All Science Journal Classification (ASJC) codes

  • Molecular Medicine
  • Pharmacology

Cite this

Antiproliferative effects of δ opioids on highly purified CD4+ and CD8+ murine T cells. / Shahabi, N. A.; Sharp, Burt.

In: Journal of Pharmacology and Experimental Therapeutics, Vol. 273, No. 3, 01.01.1995, p. 1105-1113.

Research output: Contribution to journalArticle

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abstract = "Numerous studies have shown that opioids modulate the proliferative response of mixed splenocytes to T cell mitogens. To identify the T cell subpopulations affected by opioids, splenocytes from C57BL/6 and CD1 mice were separated using a fluorescent activated cell sorter (FACS) to obtain 98 to 99{\%} pure populations of either CD4+ or CD8+ T cells. Cells were stimulated to proliferate in serum-free medium by cross-linking the T cell receptor using plate-coated anti-CD3-ε, then 3H-thymidine uptake and cell number were measured at 48 and 72 hr. [D-Ala2]-deltorphin 1 (deltorphin) dose-dependently inhibited the proliferation of C57BL/6 CD4+ T cells by approximately 50{\%}. This effect was maximal when cells were preincubated with deltorphin 60 min before activation, whereas deltorphin was ineffective when added at the time of activation. Similarly, [D-Ala2]-Met-Enkephalinamide (DAME) 10-11 to 10-7 M inhibited CD4+ T cell proliferation. Naltrindole 10-12 M abolished the antiproliferative effect of 10-7 M deltorphin on CD4+ T cells. Proliferation of CD8+ T cells from C57BL/6 mice also was dose-dependently inhibited by deltorphin. At all concentrations of deltorphin, the antiproliferative effects were greater after 48 compared to 72 hr in culture. The effect of deltorphin and DAME on secretion of the T cell growth factor, IL-2, was determined by ELISA analysis of supernatants obtained from CD4+ T cells after 48-hr culture. Deltorphin showed a biphasic effect: 10-11 M enhanced IL-2 secretion, whereas higher concentrations (10-9-10-7 M) were inhibitory. IL-2 secretion was inhibited significantly by all concentrations of DAME (10-13-10-7 M); however, a biphasic response was observed, with greatest inhibition at both the lowest (10-13 M) and highest (10-7 M) concentrations. Using CD4+ T cells obtained from CD1 mice, it was also found that deltorphin, DAME and 2,5 DPDP-enkephalin dose-dependently inhibited proliferation. The antiproliferative effects of deltorphin and DAME (10-7 M) were reduced when CD4+ T cells were stimulated by increasing concentrations of anti-CD3-ε. In summary, δ opiate receptor agonists inhibited proliferation of both CD4+ and CD8+ T cells from two strains of mice. IL-2 secretion from CD4+ T cells also was suppressed by the δ opiate receptor agonists. Inasmuch as these studies used highly purified T cells lacking detectable macrophages, the effects of the δ opiate receptor agonists appear to be mediated by receptors present on quiescent T cells.",
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N2 - Numerous studies have shown that opioids modulate the proliferative response of mixed splenocytes to T cell mitogens. To identify the T cell subpopulations affected by opioids, splenocytes from C57BL/6 and CD1 mice were separated using a fluorescent activated cell sorter (FACS) to obtain 98 to 99% pure populations of either CD4+ or CD8+ T cells. Cells were stimulated to proliferate in serum-free medium by cross-linking the T cell receptor using plate-coated anti-CD3-ε, then 3H-thymidine uptake and cell number were measured at 48 and 72 hr. [D-Ala2]-deltorphin 1 (deltorphin) dose-dependently inhibited the proliferation of C57BL/6 CD4+ T cells by approximately 50%. This effect was maximal when cells were preincubated with deltorphin 60 min before activation, whereas deltorphin was ineffective when added at the time of activation. Similarly, [D-Ala2]-Met-Enkephalinamide (DAME) 10-11 to 10-7 M inhibited CD4+ T cell proliferation. Naltrindole 10-12 M abolished the antiproliferative effect of 10-7 M deltorphin on CD4+ T cells. Proliferation of CD8+ T cells from C57BL/6 mice also was dose-dependently inhibited by deltorphin. At all concentrations of deltorphin, the antiproliferative effects were greater after 48 compared to 72 hr in culture. The effect of deltorphin and DAME on secretion of the T cell growth factor, IL-2, was determined by ELISA analysis of supernatants obtained from CD4+ T cells after 48-hr culture. Deltorphin showed a biphasic effect: 10-11 M enhanced IL-2 secretion, whereas higher concentrations (10-9-10-7 M) were inhibitory. IL-2 secretion was inhibited significantly by all concentrations of DAME (10-13-10-7 M); however, a biphasic response was observed, with greatest inhibition at both the lowest (10-13 M) and highest (10-7 M) concentrations. Using CD4+ T cells obtained from CD1 mice, it was also found that deltorphin, DAME and 2,5 DPDP-enkephalin dose-dependently inhibited proliferation. The antiproliferative effects of deltorphin and DAME (10-7 M) were reduced when CD4+ T cells were stimulated by increasing concentrations of anti-CD3-ε. In summary, δ opiate receptor agonists inhibited proliferation of both CD4+ and CD8+ T cells from two strains of mice. IL-2 secretion from CD4+ T cells also was suppressed by the δ opiate receptor agonists. Inasmuch as these studies used highly purified T cells lacking detectable macrophages, the effects of the δ opiate receptor agonists appear to be mediated by receptors present on quiescent T cells.

AB - Numerous studies have shown that opioids modulate the proliferative response of mixed splenocytes to T cell mitogens. To identify the T cell subpopulations affected by opioids, splenocytes from C57BL/6 and CD1 mice were separated using a fluorescent activated cell sorter (FACS) to obtain 98 to 99% pure populations of either CD4+ or CD8+ T cells. Cells were stimulated to proliferate in serum-free medium by cross-linking the T cell receptor using plate-coated anti-CD3-ε, then 3H-thymidine uptake and cell number were measured at 48 and 72 hr. [D-Ala2]-deltorphin 1 (deltorphin) dose-dependently inhibited the proliferation of C57BL/6 CD4+ T cells by approximately 50%. This effect was maximal when cells were preincubated with deltorphin 60 min before activation, whereas deltorphin was ineffective when added at the time of activation. Similarly, [D-Ala2]-Met-Enkephalinamide (DAME) 10-11 to 10-7 M inhibited CD4+ T cell proliferation. Naltrindole 10-12 M abolished the antiproliferative effect of 10-7 M deltorphin on CD4+ T cells. Proliferation of CD8+ T cells from C57BL/6 mice also was dose-dependently inhibited by deltorphin. At all concentrations of deltorphin, the antiproliferative effects were greater after 48 compared to 72 hr in culture. The effect of deltorphin and DAME on secretion of the T cell growth factor, IL-2, was determined by ELISA analysis of supernatants obtained from CD4+ T cells after 48-hr culture. Deltorphin showed a biphasic effect: 10-11 M enhanced IL-2 secretion, whereas higher concentrations (10-9-10-7 M) were inhibitory. IL-2 secretion was inhibited significantly by all concentrations of DAME (10-13-10-7 M); however, a biphasic response was observed, with greatest inhibition at both the lowest (10-13 M) and highest (10-7 M) concentrations. Using CD4+ T cells obtained from CD1 mice, it was also found that deltorphin, DAME and 2,5 DPDP-enkephalin dose-dependently inhibited proliferation. The antiproliferative effects of deltorphin and DAME (10-7 M) were reduced when CD4+ T cells were stimulated by increasing concentrations of anti-CD3-ε. In summary, δ opiate receptor agonists inhibited proliferation of both CD4+ and CD8+ T cells from two strains of mice. IL-2 secretion from CD4+ T cells also was suppressed by the δ opiate receptor agonists. Inasmuch as these studies used highly purified T cells lacking detectable macrophages, the effects of the δ opiate receptor agonists appear to be mediated by receptors present on quiescent T cells.

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