A chemical chaperone, sodium 4-phenylbutyric acid, attenuates the pathogenic potency in human α-synuclein A30P + A53T transgenic mice

https://doi.org/10.1016/j.parkreldis.2009.03.002Get rights and content

Abstract

Aggregation and cytotoxicity of misfolded α-synuclein are postulated to be crucial in the disease processes of Parkinson's disease (PD) and other synucleinopathies. Mutations in the α-synuclein gene in some pedigrees of familial PD have been reported. The mutant α-synuclein has been reported to form fibrillar aggregates resulting in biochemical abnormalities that are responsible for the onset of familial PD. Thus, any agent that effectively prevents the development of misfolded and aggregated α-synuclein would be a disease modifying therapeutic candidate. We examined the efficacy of sodium 4-phenylbutyric acid (PBA), one of the chemical chaperons, in transgenic (Tg) mice overexpressing human α-synuclein containing a double mutation (A30P + A53T). To evaluate the therapeutic efficacy, bradykinesia and motor coordination were assessed using a pole test and a rotarod treadmill task, respectively. After PBA treatment, these motor deteriorations gradually improved. In immunohistochemical examinations, both a loss of tyrosine hydroxylase-positive neurons and an increase of phosphorylated α-synuclein in the substantia nigra were inhibited, resulting in no depletion of the striatal dopamine content. These data suggest that PBA might be one of the therapeutic reagents for neurodegenerative disorders.

Introduction

Parkinson's disease (PD) is one of the most common neurodegenerative disorders. The occurrence of PD is largely sporadic, although several families with Mendelian segregation of PD were reported. Thirteen chromosome loci have been linked to familial PD [1]. α-Synuclein (α-Syn) is the major component of Lewy bodies (LBs) and Lewy neuritis which are the hallmark lesions found in the brains with PD and other synucleinopathies [2]. Point mutations of the α-synuclein gene have definite pathological potency. Mutations (A30P, A53T and E46K) promote the oligomer formation and fibrillization of α-Syn [3]. These mutant α-synuclein gene products tend to aggregate resulting in the formation of LBs which has been thought to have neurotoxic effects. However, there are some reports that inclusion bodies like LBs are cytoprotective against neuronal cells [4], [5]. Although the physiological functions of α-Syn still remain unclear, misfolding and aggregation of α-Syn have definite causative effects for enhancing PD onset.

Molecular chaperones such as heat shock proteins (Hsps) are known to protect other proteins from misfoldings, resulting in a reduction in protein–protein aggregate formations and hence, providing cellular protections. Hsp70 can reduce the amount of misfolded and aggregated α-Syn species in vivo and in vitro, and protect neuronal cells from α-Syn-dependent neurotoxicity [6]. Hsp104 reduced the phosphorylation of α-Syn inclusions and prevented the nigrostriatal dopaminergic neurodegeneration induced by mutant α-Syn (A30P) [7].

Chemical chaperones are small molecules with common features that mimic the functions of molecular chaperones. Sodium 4-phenylbutyric acid (PBA), one of the chemical chaperones, is an orally bioavailable short-chain fatty acid used for the treatment of various diseases such as urea metabolism disorders [8], homozygous β-thalassemia [9], spinal muscular atrophy [10] and tumor [11]. It has been shown that PBA could exert significant neuroprotective effects in some mouse models of human diseases [12], [13], [14]. Furthermore, several studies have been reported that PBA is able to reverse the mislocalization and/or aggregation of proteins associated with human diseases and as well as to suppress endoplasmic reticulum (ER) stress [15], [16], [17], [18], [19], [20], [21].

In this study, we observed the clinico-pathological effects of PBA in transgenic mice expressing human α-synuclein with both A30P and A53T mutations (TgαSYN #8713: Tg) [22].

Section snippets

Preparation of sodium 4-phenylbutyric acid

4-Phenylbutyric acid (Sigma–Aldrich, St. Louis, MO) was converted into the water-soluble sodium salt for the purposes of this study. Briefly, sodium ethoxide was obtained by reaction of sodium metal (Nacalai Tesque, Inc., Osaka, Japan) and absolute ethanol. Then, an equal molarity of 4-phenylbutyric acid was added. After the reaction, residual ethanol was removed by a vacuum pump. The lyophilized PBA was stored in a desiccator until use. The purity of synthesized PBA was determined by 1H NMR

Permeability of PBA against the BBB was estimated using an in vitro model of BBB

BBB permeability coefficient of PBA was assessed using an in vitro model of the BBB. The permeability coefficients (cm/min) were estimated as 162.91 ± 21.02 × 10−3 for sodium PBA (n = 4 inserts), and 2.18 ± 0.43 × 10−3 for Na-F (n = 8 inserts) (data not shown).

Oral PBA administration improved bradykinesia and motor coordination

In both the pole test and the rotarod treadmill task, significant differences were observed at the start of the PBA treatment (3 months old) between non-Tg (15 mice) and Tg (30 mice) mice groups. However there was no significant difference between

Discussion

At present, there is no disease modifying therapy for PD. Here we investigated the possibility of PBA treatment as a disease modifying therapy for PD. At first, the permeability of PBA through the BBB was assessed using an in vitro model. The result showed PBA had more than 70 times the permeability of the control (Na-F) [26]. It has been shown that PBA could exert significant neuroprotective effects in mouse models of neurodegenerative diseases sach as Huntington's disease [13] and amyotrophic

Acknowledgements

The authors are grateful to Dr. Kenji Yamagata and Dr. Fumi Okabe for their technical assistance in the preparation of PBA. We are also grateful to Dr. Shinya Dohgu for technical assistance in measuring the BBB permeability of PBA.

References (30)

  • S. Veszelka et al.

    Human serum amyloid P component attenuates the bacterial lipopolysaccharide-induced increase in blood–brain barrier permeability in mice

    Neurosci Lett

    (2003)
  • Y. Mizuno et al.

    Progress in the pathogenesis and genetics of Parkinson's disease

    Philos Trans R Soc Lond B Biol Sci

    (2008)
  • T. Iwatsubo

    Aggregation of α-synuclein in the pathogenesis of Parkinson's disease

    J Neurol

    (2003)
  • L. Chen et al.

    Alpha-synuclein phosphorylation controls neurotoxicity and inclusion formation in a drosophila model of Parkinson's disease

    Nat Neurosci

    (2005)
  • C. Lo Bianco et al.

    Hsp104 antagonizes α-synuclein aggregation and reduces dopaminergic degeneration in a rat model of Parkinson disease

    J Clin Invest

    (2008)
  • Cited by (69)

    View all citing articles on Scopus
    View full text