HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS		QUICK SEARCH:   [advanced] Author: Keyword(s): Year:  Vol:  Page:

This Article Full Text (PDF) All Versions of this Article: 16/11/1474 02-0216fjev1    most recent Alert me when this article is cited Alert me if a correction is posted Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by SRIRAM, K. Articles by O’CALLAGHAN, J. P. Search for Related Content PubMed PubMed Citation Articles by SRIRAM, K. Articles by O’CALLAGHAN, J. P.

Mice deficient in TNF receptors are protected against dopaminergic neurotoxicity: implications for Parkinson’s disease¹

Centers for Disease Control and Prevention (CDC)-NIOSH, Morgantown, West Virginia, USA

²Correspondence: HELD/TMBB, CDC-NIOSH, Mailstop L-3014, 1095 Willowdale Road, Morgantown, WV 26505, USA. E-mail : jdo5{at}cdc.gov

SPECIFIC AIMS

The specific aim of this study was to determine the potential role of the proinflammatory cytokine, tumor necrosis factor (TNF-), in the pathogenesis of Parkinson’s disease (PD). We examined the effects of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP), a dopaminergic neurotoxin that causes PD-like features, using mice lacking TNF receptors.

PRINCIPAL FINDINGS

1. MPTP causes a time-dependent increase in striatal TNF- mRNA that precedes the induction of glial fibrillary acidic protein (GFAP) mRNA
To determine the involvement of TNF- in the dopaminergic neurotoxicity of MPTP, we administered MPTP (12.5 mg/kg, s.c.) to male C57BL/6J mice and measured the striatal expression of TNF- and GFAP mRNA. TNF- is a key proinflammatory cytokine implicated in a broad array of diseases states, including PD. The enhanced expression of GFAP, a marker of injury-induced gliosis, is one of the earliest responses to dopaminergic nerve terminal damage in the MPTP model of PD. TNF- mRNA was detectable as early as 3 h after MPTP, but was undetectable in saline-treated controls. Southern blot analysis of the PCR amplimer using a digoxigenin-labeled human cDNA probe to TNF- confirmed expression of TNF- mRNA in the striatum. TNF- mRNA expression preceded the expression of GFAP mRNA, which was first observed by RT-PCR 12 h after MPTP treatment. These findings suggest that a proinflammatory response, as manifested by induction of TNF-, serves as an early event in the dopaminergic neurotoxic effects of MPTP.

2. Mice lacking both TNF receptors are protected against the decrease in striatal dopamine and tyrosine hydroxylase (TH) caused by MPTP
To determine whether modulation of TNF- action could alter dopaminergic neurotoxicity in the MPTP model of PD, we administered MPTP to mice lacking type 1 TNF receptors (TNFR1-/-), type 2 TNF receptors (TNFR2-/-), or both (TNFR-DKO). MPTP caused a 70%, 63%, and 41% loss of striatal dopamine in wild-type+/+, TNFR1-/-, and TNFR2-/- mice, respectively. However, mice deficient in both receptors (TNFR-DKO) were protected against the dopamine decrease caused by MPTP (Fig. 1 A). Similarly, marked decreases in the levels of dihydroxyphenylacetic acid and homovanillic acid were observed in wild-type+/+, TNFR1-/-, and TNFR2-/- mice, but not among TNFR-DKO mice (data not shown). Concurrent with the loss of striatal dopamine, MPTP also decreases the levels of striatal TH, a marker of dopaminergic nerve terminals. Administration of MPTP to wild-type+/+, TNFR1-/-, or TNFR2-/- mice resulted in a 50% decrease in the levels of TH within 48 h of dosing as measured by a fluorescence-based ELISA. However, TNFR-DKO mice were completely protected against the decrease in TH caused by MPTP (Fig. 1B ). In agreement with the results from TH ELISA, immunoblot analysis of TH revealed similar differences between wild-type+/+ and TNFR-DKO mice (data not shown).

View larger version (21K): [in this window] [in a new window]   Figure 1. MPTP-mediated loss of striatal dopaminergic markers and induction of astrogliosis are abolished in TNFR-DKO mice. Wild-type+/+, TNFR1-/-, TNFR2-/-, and TNFR-DKO mice were administered either saline or MPTP (12.5 mg/kg, s.c) and killed 12 or 48 h after treatment. A) Striatal levels of dopamine were quantified by HPLC-EC at 12 h postdosing (n=5 per group) B) Striatal TH protein levels 48 h postdosing was quantified by ELISA (n=8–10 per group). C) Striatal GFAP mRNA expression was analyzed by RT-PCR 48 h post-MPTP (n=4 per group). D) Striatal GFAP protein levels 48 h after treatment was quantified by ELISA (n=8–10 per group). Values are mean ± SE. *Significantly different from respective saline-treated controls (P<0.01). Newman Keuls pairwise comparisons were used for post hoc statistical analysis.

3. Mice lacking both TNF receptors are protected against the increase in striatal GFAP caused by MPTP
GFAP mRNA and protein expression were assayed as indices of the astroglial response to MPTP-induced dopaminergic nerve terminal damage. Administration of MPTP to wild-type+/+, TNFR1, or TNFR2 mutant mice resulted in a twofold increase in the expression of striatal GFAP mRNA (Fig. 1C ) and a fourfold increase in striatal GFAP protein content (Fig. 1D ). In TNFR-DKO mice, the MPTP-induced expression of GFAP mRNA and protein was almost completely blocked (Fig. 1C, D ). This absence of a glial response to neuronal injury was anticipated, since the dopaminergic neurotoxicity in these mice was abolished.

4. Mice lacking both TNF receptors are protected against the loss of dopaminergic nerve terminals and the associated reactive gliosis caused by MPTP
Immunohistochemical analysis confirmed that mice deficient in both the TNF receptors are protected against MPTP-mediated loss of striatal TH immunoreactivity and associated reactive gliosis. In wild-type+/+ mice, MPTP treatment resulted in loss of TH immunoreactive nerve terminals (Fig. 2 A), left panels) and a concomitant astrogliosis (Fig. 2B , left panels). In TNFR-DKO mice, however, the loss of TH immunoreactive nerve terminals and the induction of astrogliosis by MPTP were abolished (Fig. 2A , right panels; Fig. 2B , right panels, respectively). Taken together, these results confirm involvement of TNF- in the dopaminergic neurotoxicity caused by MPTP.

View larger version (67K): [in this window] [in a new window]   Figure 2. MPTP-mediated loss of TH immunoreactivity and increase in GFAP immunoreactivity are abolished in TNFR-DKO mice. Wild-type+/+, TNFR1-/-, TNFR2-/-, and TNFR-DKO mice were administered saline or MPTP (12.5 mg/kg, s.c) and killed 48 h later. Two sets of coronal or sagittal serial sections were independently stained in each analysis for TH (polyclonal, 1:1000) and GFAP (polyclonal, 1: 5000). A) TH immunoreactivity in striatum showing decreased staining in MPTP-treated wild-type+/+ mice and complete protection against MPTP neurotoxicity in TNFR-DKO mice. B) Concomitant increase in GFAP immunoreactivity (reactive astrogliosis) seen in the same field in wild-type+/+ mice is abolished in TNFR-DKO mice. [Scale bar = 140 µm].

CONCLUSIONS AND SIGNIFICANCE

We have demonstrated that enhanced expression of TNF- is associated with the earliest stages of damage in the MPTP model of dopaminergic neurotoxicity. Moreover, using mice lacking receptors for TNF, we showed complete protection against MPTP-induced neurotoxicity. The early onset of TNF- expression after MPTP and the neuroprotective effect afforded to dopaminergic neurons by TNF receptor deficiency implicate this proinflammatory cytokine as an early effector in the neurodegenerative processes underlying PD.

These data suggest that neuroprotection requires the absence or deficiency of both TNF receptor subtypes. Traditionally, the TNFR1 is considered the essential subtype for TNF-induced modulation of gene expression and cytotoxicity. Nevertheless, TNFR2 alone is capable of signaling cell death. Although each TNF- receptor was thought to mediate distinct cellular responses, there is increasing evidence accumulating to demonstrate receptor cooperation in mediating the cytocidal effect of TNF-. Thus, inhibition or deficiency of either of the receptors alone may not prevent detrimental aspects of TNF--mediated events. Our observations are in concordance with these findings. Within the central nervous system, TNF- is expressed in both microglia and astrocytes. In the substantia nigra of patients with PD, TNF- appears to be localized to activated microglia. Because the time course for expression of TNF- after MPTP parallels the time course for microglial activation, it would appear that activated microglia are a likely source for this cytokine in the MPTP model, as they seem to be in PD. Besides their presence on astrocytes and microglia, TNF receptors have been found on nigrostriatal dopaminergic neurons in PD. Their localization on the very cell type selectively vulnerable to damage in PD is consistent with the region-specific nature of the detrimental effects of TNF-. For example, whereas enhanced expression of TNF- appears to be associated with damage to dopaminergic neurons in the basal ganglia, the lack of TNF receptors or inhibition of TNF- appears to exacerbate toxic injuries of the hippocampus. Whether the effects of TNF- in the MPTP model or, potentially, in PD are mediated through microglial, astroglial, or neuronal-localized receptors remains to be determined. The molecular mechanisms downstream of TNF receptor activation remain to be elucidated as well.

In summary, our results implicate TNF- as an obligatory component of dopaminergic neurotoxicity caused by MPTP and suggest that microglia may serve as cellular effectors of TNF--mediated neurodegeneration. The broad implication of these findings is that pharmacological modulation of the TNF receptors or intermediates in its signaling cascade may provide novel and perhaps a mechanistically based approach for the treatment of PD.

View larger version (15K): [in this window] [in a new window]   Figure 3. Schematic diagram depicting the involvement of TNF- in the striatal dopaminergic neurotoxicity of MPTP and the neuroprotection afforded in mice lacking TNF receptors. MPTP treatment results in the activation of microglia, which then release TNF-. TNF- may possibly trigger degeneration of the dopaminergic nerve terminals by signaling via its receptors localized on the dopaminergic neurons. Deficiency of both the TNF receptors blocks the downstream signaling and thereby affords neuroprotection.

FOOTNOTES

¹ To read the full text of this article, go to http://www.fasebj.org/cgi/doi/10.1096/fj.02-0216fje; to cite this article, use FASEB J. (July 18, 2002) 10.1096/fj.02-0216fje

This article has been cited by other articles:

				J.-K. Lee, M. K. McCoy, A. S. Harms, K. A. Ruhn, S. J. Gold, and M. G. Tansey Regulator of G-Protein Signaling 10 Promotes Dopaminergic Neuron Survival via Regulation of the Microglial Inflammatory Response J. Neurosci., August 20, 2008; 28(34): 8517 - 8528. [Abstract] [Full Text] [PDF]

				L. Qian, S.-J. Wei, D. Zhang, X. Hu, Z. Xu, B. Wilson, J. El-Benna, J.-S. Hong, and P. M. Flood Potent Anti-Inflammatory and Neuroprotective Effects of TGF-{beta}1 Are Mediated through the Inhibition of ERK and p47phox-Ser345 Phosphorylation and Translocation in Microglia J. Immunol., July 1, 2008; 181(1): 660 - 668. [Abstract] [Full Text] [PDF]

				A. Ghosh, A. Roy, X. Liu, J. H. Kordower, E. J. Mufson, D. M. Hartley, S. Ghosh, R. L. Mosley, H. E. Gendelman, and K. Pahan Selective inhibition of NF-{kappa}B activation prevents dopaminergic neuronal loss in a mouse model of Parkinson's disease PNAS, November 20, 2007; 104(47): 18754 - 18759. [Abstract] [Full Text] [PDF]

				M.-J. Wang, S.-Z. Lin, J.-S. Kuo, H.-Y. Huang, S.-F. Tzeng, C.-H. Liao, D.-C. Chen, and W.-F. Chen Urocortin Modulates Inflammatory Response and Neurotoxicity Induced by Microglial Activation J. Immunol., November 1, 2007; 179(9): 6204 - 6214. [Abstract] [Full Text] [PDF]

				L. Qian, K. S. Tan, S.-J. Wei, H.-M. Wu, Z. Xu, B. Wilson, R.-B. Lu, J.-S. Hong, and P. M. Flood Microglia-Mediated Neurotoxicity Is Inhibited by Morphine through an Opioid Receptor-Independent Reduction of NADPH Oxidase Activity J. Immunol., July 15, 2007; 179(2): 1198 - 1209. [Abstract] [Full Text] [PDF]

				A. D. Wahner, J. S. Sinsheimer, Jeff. M. Bronstein, and B. Ritz Inflammatory Cytokine Gene Polymorphisms and Increased Risk of Parkinson Disease Arch Neurol, June 1, 2007; 64(6): 836 - 840. [Abstract] [Full Text] [PDF]

				M. Timmer, K. Cesnulevicius, C. Winkler, J. Kolb, E. Lipokatic-Takacs, J. Jungnickel, and C. Grothe Fibroblast Growth Factor (FGF)-2 and FGF Receptor 3 Are Required for the Development of the Substantia Nigra, and FGF-2 Plays a Crucial Role for the Rescue of Dopaminergic Neurons after 6-Hydroxydopamine Lesion J. Neurosci., January 17, 2007; 27(3): 459 - 471. [Abstract] [Full Text] [PDF]

				W. Zhang, E.-J. Shin, T. Wang, P. H. Lee, H. Pang, M.-B. Wie, W.-K. Kim, S.-J. Kim, W.-H. Huang, Y. Wang, et al. 3-Hydroxymorphinan, a metabolite of dextromethorphan, protects nigrostriatal pathway against MPTP-elicited damage both in vivo and in vitro FASEB J, December 1, 2006; 20(14): 2496 - 2511. [Abstract] [Full Text] [PDF]

				S. Yang, J. Yang, Z. Yang, P. Chen, A. Fraser, W. Zhang, H. Pang, X. Gao, B. Wilson, J.-S. Hong, et al. Pituitary Adenylate Cyclase-Activating Polypeptide (PACAP) 38 and PACAP4-6 Are Neuroprotective through Inhibition of NADPH Oxidase: Potent Regulators of Microglia-Mediated Oxidative Stress J. Pharmacol. Exp. Ther., November 1, 2006; 319(2): 595 - 603. [Abstract] [Full Text] [PDF]

				L. Qian, M. L. Block, S.-J. Wei, C.-f. Lin, J. Reece, H. Pang, B. Wilson, J.-S. Hong, and P. M. Flood Interleukin-10 Protects Lipopolysaccharide-Induced Neurotoxicity in Primary Midbrain Cultures by Inhibiting the Function of NADPH Oxidase J. Pharmacol. Exp. Ther., October 1, 2006; 319(1): 44 - 52. [Abstract] [Full Text] [PDF]

				M. K. McCoy, T. N. Martinez, K. A. Ruhn, D. E. Szymkowski, C. G. Smith, B. R. Botterman, K. E. Tansey, and M. G. Tansey Blocking soluble tumor necrosis factor signaling with dominant-negative tumor necrosis factor inhibitor attenuates loss of dopaminergic neurons in models of Parkinson's disease. J. Neurosci., September 13, 2006; 26(37): 9365 - 9375. [Abstract] [Full Text] [PDF]

				K. Sriram, J. M. Matheson, S. A. Benkovic, D. B. Miller, M. I. Luster, and J. P. O'Callaghan Deficiency of TNF receptors suppresses microglial activation and alters the susceptibility of brain regions to MPTP-induced neurotoxicity: role of TNF-{alpha} FASEB J, April 1, 2006; 20(6): 670 - 682. [Abstract] [Full Text] [PDF]

				Y. S. Kim, S. S. Kim, J. J. Cho, D. H. Choi, O. Hwang, D. H. Shin, H. S. Chun, M. F. Beal, and T. H. Joh Matrix Metalloproteinase-3: A Novel Signaling Proteinase from Apoptotic Neuronal Cells That Activates Microglia J. Neurosci., April 6, 2005; 25(14): 3701 - 3711. [Abstract] [Full Text] [PDF]

				N. M. Filipov, R. F. Seegal, and D. A. Lawrence Manganese Potentiates In Vitro Production of Proinflammatory Cytokines and Nitric Oxide by Microglia Through a Nuclear Factor kappa B-Dependent Mechanism Toxicol. Sci., March 1, 2005; 84(1): 139 - 148. [Abstract] [Full Text] [PDF]

				M. A. Christie and S. M. Hersch Demonstration of nondeclarative sequence learning in mice: Development of an animal analog of the human serial reaction time task Learn. Mem., November 1, 2004; 11(6): 720 - 723. [Abstract] [Full Text] [PDF]

				K. Sriram, S. A. Benkovic, M. A. Hebert, D. B. Miller, and J. P. O'Callaghan Induction of gp130-related Cytokines and Activation of JAK2/STAT3 Pathway in Astrocytes Precedes Up-regulation of Glial Fibrillary Acidic Protein in the 1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine Model of Neurodegeneration: KEY SIGNALING PATHWAY FOR ASTROGLIOSIS IN VIVO? J. Biol. Chem., May 7, 2004; 279(19): 19936 - 19947. [Abstract] [Full Text] [PDF]

				L. Qin, Y. Liu, T. Wang, S.-J. Wei, M. L. Block, B. Wilson, B. Liu, and J.-S. Hong NADPH Oxidase Mediates Lipopolysaccharide-induced Neurotoxicity and Proinflammatory Gene Expression in Activated Microglia J. Biol. Chem., January 9, 2004; 279(2): 1415 - 1421. [Abstract] [Full Text] [PDF]

				Q. Ma, K. Kinneer, J. Ye, and B. J. Chen Inhibition of Nuclear Factor {kappa}B by Phenolic Antioxidants: Interplay between Antioxidant Signaling and Inflammatory Cytokine Expression Mol. Pharmacol., August 1, 2003; 64(2): 211 - 219. [Abstract] [Full Text] [PDF]

This Article Full Text (PDF) All Versions of this Article: 16/11/1474 02-0216fjev1    most recent Alert me when this article is cited Alert me if a correction is posted Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by SRIRAM, K. Articles by O’CALLAGHAN, J. P. Search for Related Content PubMed PubMed Citation Articles by SRIRAM, K. Articles by O’CALLAGHAN, J. P.