Pharmacological induction of heat-shock proteins alleviates polyglutamine-mediated motor neuron disease - PubMed (original) (raw)

Pharmacological induction of heat-shock proteins alleviates polyglutamine-mediated motor neuron disease

Masahisa Katsuno et al. Proc Natl Acad Sci U S A. 2005.

Abstract

Spinal and bulbar muscular atrophy (SBMA) is an adult-onset motor neuron disease caused by the expansion of a trinucleotide CAG repeat encoding the polyglutamine tract in the first exon of the androgen receptor gene (AR). The pathogenic, polyglutamine-expanded AR protein accumulates in the cell nucleus in a ligand-dependent manner and inhibits transcription by interfering with transcriptional factors and coactivators. Heat-shock proteins (HSPs) are stress-induced chaperones that facilitate the refolding and, thus, the degradation of abnormal proteins. Geranylgeranylacetone (GGA), a nontoxic antiulcer drug, has been shown to potently induce HSP expression in various tissues, including the central nervous system. In a cell model of SBMA, GGA increased the levels of Hsp70, Hsp90, and Hsp105 and inhibited cell death and the accumulation of pathogenic AR. Oral administration of GGA also up-regulated the expression of HSPs in the central nervous system of SBMA-transgenic mice and suppressed nuclear accumulation of the pathogenic AR protein, resulting in amelioration of polyglutamine-dependent neuromuscular phenotypes. These observations suggest that, although a high dose appears to be needed for clinical effects, oral GGA administration is a safe and promising therapeutic candidate for polyglutamine-mediated neurodegenerative diseases, including SBMA.

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Figures

Fig. 1.

Fig. 1.

Effects of GGA on polyglutamine toxicity in cultured cell. (A) Punctuated aggregates visualized with GFP (arrowhead) are formed in SHSY-5Y cells infected with an adenovirus vector containing truncated AR with 97 CAGs (tAR97Q-GFP) but not in those bearing tAR24Q. (B) Frequency of cell death 6 days after infection as detected by propidium iodine staining (*, P < 0.05 compared with untreated tAR97Q cells). (C) Suppression of cell death by GGA. (D) Frequency of cells bearing aggregates. (E) Anti-AR analysis of Western blots of extracts from cells infected with tAR97Q. Error bars indicate SD.

Fig. 2.

Fig. 2.

GGA-mediated HSP induction in cultured cell. (A) Anti-HSP analysis of Western blots from cells infected with tAR97Q and treated with GGA. (B) Quantification of the levels of HSPs from tAR97Q-infected cells after 2 days of GGA treatment. (C) Anti-Hsp70 analysis of Western blots from tAR97Q cells treated with or without cycloheximide. (D) Frequency of cell death 2 days after infection as detected by propidium iodine staining (**, P < 0.05 compared with tAR97Q cells treated with GGA but not with cycloheximide). (E and F) Anti-HSF-1 analysis of Western blots of the cellular nuclear fraction (E) and that of total cell lysate (F). Upper bands correspond to the hyperphosphorylated, active form of HSF-1. (G) Immunocytochemistry for HSF-1. Error bars indicate SD.

Fig. 3.

Fig. 3.

Effect of GGA on neurological phenotypes of AR-97Q mice. (A) Muscle atrophy of 13-week AR-97Q mice. (B) Footprints of 13-week AR-97Q mice. Front paws are shown in red, and hind paws are shown in blue. (C) Stride distance of 13-week AR-97Q mice (n = 3 for each group). (_D_-F) Rotarod task (D), body weight (E), and cumulative survival (F) of male AR-97Q mice treated with GGA (n = 12 for each group) and untreated counterparts (n = 15). Rotarod performance significantly improved after GGA at doses of 0.5% and 1.0% (P < 0.0001 at both doses compared with nontreated mice at 20 weeks), and body weight increased significantly at a dose of 0.5% (P < 0.005 at 0.5% and P < 0.05 at 1.0%, at 14 weeks). Error bars indicate SD.

Fig. 4.

Fig. 4.

GGA-mediated HSP induction in AR-97Q mice. (A) Western blotting for various HSPs in the spinal cord of 14-week, wild-type (Wt), AR-24Q and AR-97Q mice. (B) Western blotting for various HSPs in skeletal muscle of 14-week wild-type, AR-24Q, and AR-97Q mice. (C) Immunohistochemistry for Hsp70 in 14-week wild-type, AR-24Q, and AR-97Q mice. (D) Western blotting of nuclear fraction from spinal cord and that from muscle using anti-HSF-1 antibody. Upper bands correspond to the hyperphosphorylated active form of HSF-1.

Fig. 5.

Fig. 5.

Effect of GGA on accumulation of abnormal AR. (A) Immunohistochemistry of 14-week wild-type, AR-24Q, and AR-97Q mice using 1C2 antibody. (B) Quantification of 1C2-positive cells in spinal cord and muscle of AR-97Q mice treated with or without GGA. (C) Western blotting for AR of 14-week AR-97Q mice and quantification of the high-molecular-weight, abnormal AR complex indicated by a smear from the top of the gel. Error bars indicate SD.

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