Elsevier

Neurobiology of Disease

Volume 71, November 2014, Pages 280-291
Neurobiology of Disease

Dynamic changes in pro- and anti-inflammatory cytokines in microglia after PPAR-γ agonist neuroprotective treatment in the MPTPp mouse model of progressive Parkinson's disease

https://doi.org/10.1016/j.nbd.2014.08.011Get rights and content

Highlights

  • We used a mouse model of progressive PD induced by chronic MPTP/probenecid exposure.

  • We assessed changes in pro/anti-inflammatory cytokines and neurodegeneration.

  • MPTPp induced pro-inflammatory cytokines in microglia and neurodegeneration.

  • Neuroprotective rosiglitazone induced anti and inhibited pro-inflammatory cytokines.

  • Switching microglia polarization to M2 may mediate PPAR-γ agonist neuroprotection.

Abstract

Neuroinflammatory changes play a pivotal role in the progression of Parkinson's disease (PD) pathogenesis. Recent findings have suggested that activated microglia may polarize similarly to peripheral macrophages in the central nervous system (CNS), assuming a pro-inflammatory M1 phenotype or the alternative anti-inflammatory M2 phenotype via cytokine production. A skewed M1 activation over M2 has been related to disease progression in Alzheimer disease, and modulation of microglia polarization may be a therapeutic target for neuroprotection. By using the 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine–probenecid (MPTPp) mouse model of progressive PD, we investigated dynamic changes in the production of pro-inflammatory cytokines, such as tumor necrosis factor (TNF)-α and interleukin (IL)-1β, and anti-inflammatory cytokines, such as transforming growth factor (TGF)-β and IL-10, within Iba-1-positive cells in the substantia nigra compacta (SNc). In addition, to further characterize changes in the M2 phenotype, we measured CD206 in microglia. Moreover, in order to target microglia polarization, we evaluated the effect of the peroxisome-proliferator-activated receptor (PPAR)-γ agonist rosiglitazone, which has been shown to exert neuroprotective effects on nigral dopaminergic neurons in PD models, and acts as a modulator of cytokine production and phenotype in peripheral macrophages.

Chronic treatment with MPTPp induced a progressive degeneration of SNc neurons. The neurotoxin treatment was associated with a gradual increase in both TNF-α and IL-1β colocalization with Iba-1-positive cells, suggesting an increase in pro-inflammatory microglia. In contrast, TGF-β colocalization was reduced by the neurotoxin treatment, while IL-10 was mostly unchanged. Administration of rosiglitazone during the full duration of MPTPp treatment reverted both TNF-α and IL-1β colocalization with Iba-1 to control levels. Moreover, rosiglitazone induced an increase in TGF-β and IL-10 colocalization compared with the MPTPp treatment. CD206 was gradually reduced by the chronic MPTPp treatment, while rosiglitazone restored control levels, suggesting that M2 anti-inflammatory microglia were stimulated and inflammatory microglia were inhibited by the neuroprotective treatment. The results show that the dopaminergic degeneration was associated with a gradual microglia polarization to the inflammatory over the anti-inflammatory phenotype in a chronic mouse model of PD. Neuroprotective treatment with rosiglitazone modulated microglia polarization, boosting the M2 over the pro-inflammatory phenotype. PPAR-γ agonists may offer a novel approach to neuroprotection, acting as disease-modifying drugs through an immunomodulatory action in the CNS.

Section snippets

Drugs

MPTP-HCl (Sigma, Italy) was dissolved in saline; probenecid (Sigma, Italy) was dissolved in 5% NaHCO3. Rosiglitazone (Santa Cruz Biotechnology) was suspended in 0.5% methylcellulose.

Treatment

Three-month-old male C57BL/6J mice (Charles River, Italy) were divided into seven groups (n = 5–10 for each group). The control group received saline as a vehicle, groups M1, M3, M7, and M10 received 1, 3, 7, and 10 doses of MPTP (25 mg/kg i.p.) respectively, plus probenecid (100 mg/kg i.p.) (MPTPp). MPTPp was injected

Evaluation of MPTP-induced nigral degeneration

MPTPp chronic treatment reduced both the density and the number of TH-positive neurons as well as Nissl-stained cells in the SNc by about 40% (Fig. 1). Neurodegeneration was gradual along the chronic neurotoxin treatment, with a slight reduction in the number of TH-positive neurons after three MPTP injections (M3), and a larger significant neuronal loss after subsequent injections, in agreement with the previous characterization of this MPTPp protocol (Schintu et al., 2009b). The daily

Discussion

We report that the progressive degeneration of nigral neurons and their rescue by rosiglitazone in the chronic MPTPp treatment were associated with dynamic changes in cytokine production by microglia/macrophages in the SNc. Although fluorescent immunohistochemistry only provides a semi-quantitative measurement of protein levels, it allows to assess the extent of proteins colocalization, and to use this value for comparison among experimental groups. Moreover this technique, by evaluating

Conclusions

Progression of neurodegeneration in a chronic model of PD was associated with a gradual increase of neurotoxic pro-inflammatory microglia over the anti-inflammatory phenotype. Moreover, pharmacological treatment with a PPAR-γ agonist switched microglia polarization to anti-inflammatory, turning microglia activation into a beneficial event in the diseased brain. Therefore, although microgliosis is generally regarded as a toxic event in PD pathogenesis, results suggest that microglia might

Acknowledgments

We thank Prof. Giacomo Diaz, PhD, Dept. of Biomedical Sciences, Univ. of Cagliari, for assistance in statistical analysis. We thank Susan Bos of Medical Edit for language revision of the manuscript.

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