Immunomodulatory effects of nicotine on interleukin 1β activated human astrocytes and the role of cyclooxygenase 2 in the underlying mechanism.
BACKGROUND
The cholinergic anti-inflammatory pathway (CAP) primarily functions through acetylcholine (ACh)-alpha7 nicotinic acetylcholine receptor (α7nAChR) interaction on macrophages to control peripheral inflammation. Interestingly, ACh can also bind α7nAChRs on microglia resulting in neuroprotective effects. However, ACh effects on astrocytes remain elusive. Here, we investigated the effects of nicotine, an ACh receptor agonist, on the cytokine and cholinesterase production of immunocompetent human astrocytes stimulated with interleukin 1β (IL-1β) in vitro. In addition, the potential involvement of prostaglandins as mediators of nicotine was studied using cyclooxygenase 2 (COX-2) inhibition.


METHODS
Cultured human fetal astrocytes were stimulated with human recombinant IL-1β and treated simultaneously with nicotine at different concentrations (1, 10, and 100 μM). Cell supernatants were collected for cytokine and cholinesterase profiling using ELISA and MesoScale multiplex assay. α7nAChR expression on activated human astrocytes was studied using immunofluorescence. For the COX-2 inhibition studies, enzyme activity was inhibited using NS-398. One-way ANOVA was used to perform statistical analyses.


RESULTS
Nicotine treatment dose dependently limits the production of critical proinflammatory cytokines such as IL-6 (60.5 ± 3.3, %inhibition), IL-1β (42.4 ± 1.7, %inhibition), and TNF-α (68.9 ± 7.7, %inhibition) by activated human astrocytes. Interestingly, it also inhibits IL-8 chemokine (31.4 ± 8.5, %inhibition), IL-13 (34.243 ± 4.9, %inhibition), and butyrylcholinesterase (20.8 ± 2.8, %inhibition) production at 100 μM. Expression of α7nAChR was detected on the activated human astrocytes. Importantly, nicotine's inhibitory effect on IL-6 production was reversed with the specific COX-2 inhibitor NS-398.


CONCLUSIONS
Activation of the cholinergic system through α7nAChR agonists has been known to suppress inflammation both in the CNS and periphery. In the CNS, earlier experimental data shows that cholinergic activation through nicotine inhibits microglial activation and proinflammatory cytokine release. Here, we report similar anti-inflammatory effects of cholinergic activation on human astrocytes, at least partly mediated through the COX-2 pathway. These results confirm the potential for cholinergic neuroprotection, which is looked upon as a promising therapy for neuroinflammation as well as neurodegenerative diseases and stroke. Our data implicates an important role for the prostaglandin system in cholinergic regulatory effects.
Department of Medicine, Unit of Rheumatology, Center for Molecular Medicine (CMM), Karolinska Institute, Karolinska University Hospital, Stockholm, Sweden. Priya.Revathikumar@ki.se.
Priya
http://orcid.org/0000-0001-9761-4014
P
Revathikumar
Department of Medicine, Unit of Rheumatology, Center for Molecular Medicine (CMM), Karolinska Institute, Karolinska University Hospital, Stockholm, Sweden.
Filip
F
Bergqvist
Department of Medicine, Unit of Rheumatology, Center for Molecular Medicine (CMM), Karolinska Institute, Karolinska University Hospital, Stockholm, Sweden.
Srividya
S
Gopalakrishnan
Department of Medicine, Unit of Rheumatology, Center for Molecular Medicine (CMM), Karolinska Institute, Karolinska University Hospital, Stockholm, Sweden.
Marina
M
Korotkova
Department of Medicine, Unit of Rheumatology, Center for Molecular Medicine (CMM), Karolinska Institute, Karolinska University Hospital, Stockholm, Sweden.
Per-Johan
PJ
Jakobsson
Department of Medicine, Unit of Rheumatology, Center for Molecular Medicine (CMM), Karolinska Institute, Karolinska University Hospital, Stockholm, Sweden.
Jon
J
Lampa
Department of Medicine, Unit of Rheumatology, Center for Molecular Medicine (CMM), Karolinska Institute, Karolinska University Hospital, Stockholm, Sweden.
Erwan
E
Le Maître
Alpha7 nicotinic acetylcholine receptor; Astrocytes; Cholinergic immune regulation; Cyclooxygenase 2; Neuroinflammation; Nicotine; Prostaglandins
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