Ketamine, as adjuvant analgesics for patients with refractory cancer pain, does affect IL-2/IFN-γ expression of T cells in vitro?: A prospective, randomized, double-blind study

Abstract

Background: Ketamine has been used as an analgesic adjuvant with morphine in the treatment of refractory cancer pain recently. But both morphine and ketamine have been reported to produce a number of immunomodulatory effects. The current study was performed to assess whether the concentration of ketamine, as adjuvant analgesics for patient with refractory cancer pain, was related to its effect on T cells interleukin-2 (IL-2)/interferon-γ (IFN-γ) expression in vitro.

Methods: Peripheral blood mononuclear cells (PBMCs) were isolated from venous blood of patients with refractory cancer pain over a Ficoll-Hypaque density gradient. T cells were isolated from by positive selection using anti-CD3 beads. T cells were then treated with vehicle (C group), morphine (200 ng/mL, M group), morphine (200 ng/mL), and different dose of ketamine (100, 200, 1000 ng/mL; MK1, MK5, MK10 group) for 24 hours before stimulation with anti-CD3 and anti-CD28. Then supernatant IL-2 and IFN-γ protein analysis, quantitative reverse transcription polymerase chain reaction (RT-PCR) for IL-2 and IFN-γ were done.

Results: There were no significant difference of supernatant IL-2 and IFN-γ among C group, M group, and MK1 group, but the mRNA of M group and MK1 group were decreased compared with C group (P < .05). Compared with C group, both of the supernatant protein and the mRNA of MK5 group and MK10 group were all significantly decreased (P < .01). Compared with M group, both of the supernatant protein and the mRNA of MK5 group and MK10 group were all decreased (P < .05), while supernatant IL-2 and the mRNA of MK10 group were significantly decreased (P < .01).

Conclusion: In conclusion, we confirmed that just as morphine, ketamine dose-dependently suppressed IL-2 and IFN-γ of activated T lymphocyte of patients with refractory cancer pain in vitro, but the inhibitory action of low dose ketamine could be neglected.

Figure 1

Effects of morphine and ketamine on T cells of patients with refractory cancer pain. PBMCs were isolated over a Ficoll-Hypaque density gradient. T cells were isolated from PBMCs by positive selection using anti-CD3 beads. T cells were then treated with vehicle, morphine (200 ng/mL), and different dose of ketamine (100, 500, 1000 ng/mL) for 24 hours before stimulation with anti-CD3 and anti-CD28. Then supernatant IL-2 and IFN-γ immunoreactive protein concentrations were measured using cytokine-specific enzyme-linked immunosorbent assay kits. ^∗/^∗∗, significant at level P < .05/.01 compared with the C group; ^#/^##, significant at level P < .05/.01 compared with the M group. IFN = interferon, IL = interleukin, PBMCs = peripheral blood mononuclear cells.

Figure 2

Effects of morphine and ketamine on T cells of patients with refractory cancer pain. PBMCs were isolated over a Ficoll-Hypaque density gradient. T cells were isolated from PBMCs by positive selection using anti-CD3 beads. T cells were then treated with vehicle, morphine (200 ng/mL), and different dose of ketamine (100, 500, 1000 ng/mL) for 24 hours before stimulation with anti-CD3 and anti-CD28. Then total RNA of T cells was extracted by cell lysis in guanidinium isothiocyanate, followed by phenol acid extraction. And amplification was performed with the oligonucleotide primers specific for human IL-2, IFN-γ. Quantitative real-time RT-PCR was done using a LightCycler-Fast Start DNA Master SYBR Green I kit (IL-2) or Revert Aid TM First Strand cDNA Synthesis Kit (IFN-γ). ^∗/^∗∗, significant at level P < .05/.01 compared with the C group; ^#/^##, significant at level P < .05/.01 compared with the M group. IFN = interferon, IL = interleukin, PBMCs = peripheral blood mononuclear cells, RT-PCR = reverse transcription polymerase chain reaction.