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Interleukin-1 beta-induced up-regulation of opioid receptors in the untreated and morphine-desensitized U87 MG human astrocytoma cells.

Byrne LS, Peng J, Sarkar S, Chang SL - J Neuroinflammation (2012)

Bottom Line: DOR expression was also elevated, although not significantly.Co-treatment with IL-1β and interleukin-1 receptor antagonist protein (IL-1RAP) resulted in a significant decrease in IL-1β-mediated MOR up-regulation.Our results indicate that the pro-inflammatory cytokine, IL-1β, affects opiate-dependent pathways by up-regulating the expression of the MOR in both untreated and morphine-desensitized U87 MG.

View Article: PubMed Central - HTML - PubMed

Affiliation: Institute of NeuroImmune Pharmacology, Seton Hall University, 400 South Orange Ave, South Orange, NJ 07079, USA.

ABSTRACT

Background: Interleukin-1beta (IL-1β) is a pro-inflammatory cytokine that can be produced in the central nervous system during inflammatory conditions. We have previously shown that IL-1β expression is altered in the rat brain during a morphine tolerant state, indicating that this cytokine may serve as a convergent point between the immune challenge and opiate mediated biological pathways. We hypothesized that IL-1β up-regulates opioid receptors in human astrocytes in both untreated and morphine-desensitized states.

Methods: To test this hypothesis, we compared the basal expression of the mu (MOR), delta (DOR), and kappa (KOR) opioid receptors in the human U87 MG astrocytic cell line to SH-SY5Y neuronal and HL-60 immune cells using absolute quantitative real time RT-PCR (AQ-rt-RT-PCR). To demonstrate that IL-1β induced up-regulation of the MOR, DOR and KOR, U87 MG cells (2 x 105 cells/well) were treated with IL-1β (20 ng/mL or 40 ng/mL), followed by co-treatment with interleukin-1 receptor antagonist protein (IL-1RAP) (400 ng/mL or 400 ng/mL). The above experiment was repeated in the cells desensitized with morphine, where U87 MG cells were pre-treated with 100 nM morphine. The functionality of the MOR in U87 MG cells was then demonstrated using morphine inhibition of forksolin-induced intracellular cAMP, as determined by radioimmunoassay.

Results: U87 MG cells treated with IL-1β for 12 h showed a significant up-regulation of MOR and KOR. DOR expression was also elevated, although not significantly. Treatment with IL-1β also showed a significant up-regulation of the MOR in U87 MG cells desensitized with morphine. Co-treatment with IL-1β and interleukin-1 receptor antagonist protein (IL-1RAP) resulted in a significant decrease in IL-1β-mediated MOR up-regulation.

Conclusion: Our results indicate that the pro-inflammatory cytokine, IL-1β, affects opiate-dependent pathways by up-regulating the expression of the MOR in both untreated and morphine-desensitized U87 MG.

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The effects of IL-1RAP on IL-1β-induced up-regulation of the MOR in U87 MG cells. U87 MG cells were treated with medium (control), IL-1β (20 ng/mL), IL-1RAP (400 ng/mL) + vehicle, IL-1RAP (400 ng/mL) + IL-1β (20 ng/mL), IL-1RAP (4,000 ng/mL) + vehicle, or IL-1RAP (4,000 ng/mL) + IL-1β (20 ng/mL) for 12 h. Real time RT-PCR was used to determine the levels of the MOR; GAPDH was used to normalize the MOR levels. Data are the mean ± SE. A Student t-test was used to determine significance. *P <0.05 compared to control; ^P <0.001 compared to IL-1β (alone); #P <0.01 compared to IL-1β (alone).
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Figure 5: The effects of IL-1RAP on IL-1β-induced up-regulation of the MOR in U87 MG cells. U87 MG cells were treated with medium (control), IL-1β (20 ng/mL), IL-1RAP (400 ng/mL) + vehicle, IL-1RAP (400 ng/mL) + IL-1β (20 ng/mL), IL-1RAP (4,000 ng/mL) + vehicle, or IL-1RAP (4,000 ng/mL) + IL-1β (20 ng/mL) for 12 h. Real time RT-PCR was used to determine the levels of the MOR; GAPDH was used to normalize the MOR levels. Data are the mean ± SE. A Student t-test was used to determine significance. *P <0.05 compared to control; ^P <0.001 compared to IL-1β (alone); #P <0.01 compared to IL-1β (alone).

Mentions: IL-1β exerts its biological effects by binding to interleukin-1 receptor 1 (IL-1R1). An antagonist to this receptor, interleukin-1 receptor antagonist protein (IL-1RAP), has been shown to decrease IL-1β’s effects [28]. We used IL-1RAP to examine whether IL-1β’s up-regulation of the MOR is mediated through the IL-1R1. U87 MG cells treated with IL-1β (20 ng/mL) demonstrated an increase in the MOR (3.8 ± 0.05) compared to the U87 MG cells treated with control (Figure 5). U87 MG cells treated with IL-1RAP (400 ng/mL) + vehicle showed a significant decrease in MOR expression (0.16 ± 0.64) compared to IL-1β treatment alone or control. Co-treatment with IL-1RAP (400 ng/mL) + IL-1β (20 ng/mL) also resulted in a significant decrease of the MOR (0.36 ± 0.27) compared to the vehicle treated control and to IL-1β alone.


Interleukin-1 beta-induced up-regulation of opioid receptors in the untreated and morphine-desensitized U87 MG human astrocytoma cells.

Byrne LS, Peng J, Sarkar S, Chang SL - J Neuroinflammation (2012)

The effects of IL-1RAP on IL-1β-induced up-regulation of the MOR in U87 MG cells. U87 MG cells were treated with medium (control), IL-1β (20 ng/mL), IL-1RAP (400 ng/mL) + vehicle, IL-1RAP (400 ng/mL) + IL-1β (20 ng/mL), IL-1RAP (4,000 ng/mL) + vehicle, or IL-1RAP (4,000 ng/mL) + IL-1β (20 ng/mL) for 12 h. Real time RT-PCR was used to determine the levels of the MOR; GAPDH was used to normalize the MOR levels. Data are the mean ± SE. A Student t-test was used to determine significance. *P <0.05 compared to control; ^P <0.001 compared to IL-1β (alone); #P <0.01 compared to IL-1β (alone).
© Copyright Policy - open-access
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC3526549&req=5

Figure 5: The effects of IL-1RAP on IL-1β-induced up-regulation of the MOR in U87 MG cells. U87 MG cells were treated with medium (control), IL-1β (20 ng/mL), IL-1RAP (400 ng/mL) + vehicle, IL-1RAP (400 ng/mL) + IL-1β (20 ng/mL), IL-1RAP (4,000 ng/mL) + vehicle, or IL-1RAP (4,000 ng/mL) + IL-1β (20 ng/mL) for 12 h. Real time RT-PCR was used to determine the levels of the MOR; GAPDH was used to normalize the MOR levels. Data are the mean ± SE. A Student t-test was used to determine significance. *P <0.05 compared to control; ^P <0.001 compared to IL-1β (alone); #P <0.01 compared to IL-1β (alone).
Mentions: IL-1β exerts its biological effects by binding to interleukin-1 receptor 1 (IL-1R1). An antagonist to this receptor, interleukin-1 receptor antagonist protein (IL-1RAP), has been shown to decrease IL-1β’s effects [28]. We used IL-1RAP to examine whether IL-1β’s up-regulation of the MOR is mediated through the IL-1R1. U87 MG cells treated with IL-1β (20 ng/mL) demonstrated an increase in the MOR (3.8 ± 0.05) compared to the U87 MG cells treated with control (Figure 5). U87 MG cells treated with IL-1RAP (400 ng/mL) + vehicle showed a significant decrease in MOR expression (0.16 ± 0.64) compared to IL-1β treatment alone or control. Co-treatment with IL-1RAP (400 ng/mL) + IL-1β (20 ng/mL) also resulted in a significant decrease of the MOR (0.36 ± 0.27) compared to the vehicle treated control and to IL-1β alone.

Bottom Line: DOR expression was also elevated, although not significantly.Co-treatment with IL-1β and interleukin-1 receptor antagonist protein (IL-1RAP) resulted in a significant decrease in IL-1β-mediated MOR up-regulation.Our results indicate that the pro-inflammatory cytokine, IL-1β, affects opiate-dependent pathways by up-regulating the expression of the MOR in both untreated and morphine-desensitized U87 MG.

View Article: PubMed Central - HTML - PubMed

Affiliation: Institute of NeuroImmune Pharmacology, Seton Hall University, 400 South Orange Ave, South Orange, NJ 07079, USA.

ABSTRACT

Background: Interleukin-1beta (IL-1β) is a pro-inflammatory cytokine that can be produced in the central nervous system during inflammatory conditions. We have previously shown that IL-1β expression is altered in the rat brain during a morphine tolerant state, indicating that this cytokine may serve as a convergent point between the immune challenge and opiate mediated biological pathways. We hypothesized that IL-1β up-regulates opioid receptors in human astrocytes in both untreated and morphine-desensitized states.

Methods: To test this hypothesis, we compared the basal expression of the mu (MOR), delta (DOR), and kappa (KOR) opioid receptors in the human U87 MG astrocytic cell line to SH-SY5Y neuronal and HL-60 immune cells using absolute quantitative real time RT-PCR (AQ-rt-RT-PCR). To demonstrate that IL-1β induced up-regulation of the MOR, DOR and KOR, U87 MG cells (2 x 105 cells/well) were treated with IL-1β (20 ng/mL or 40 ng/mL), followed by co-treatment with interleukin-1 receptor antagonist protein (IL-1RAP) (400 ng/mL or 400 ng/mL). The above experiment was repeated in the cells desensitized with morphine, where U87 MG cells were pre-treated with 100 nM morphine. The functionality of the MOR in U87 MG cells was then demonstrated using morphine inhibition of forksolin-induced intracellular cAMP, as determined by radioimmunoassay.

Results: U87 MG cells treated with IL-1β for 12 h showed a significant up-regulation of MOR and KOR. DOR expression was also elevated, although not significantly. Treatment with IL-1β also showed a significant up-regulation of the MOR in U87 MG cells desensitized with morphine. Co-treatment with IL-1β and interleukin-1 receptor antagonist protein (IL-1RAP) resulted in a significant decrease in IL-1β-mediated MOR up-regulation.

Conclusion: Our results indicate that the pro-inflammatory cytokine, IL-1β, affects opiate-dependent pathways by up-regulating the expression of the MOR in both untreated and morphine-desensitized U87 MG.

Show MeSH
Related in: MedlinePlus