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On a 0.1% phytol diet (E),Phyh/mice showed indicators of microsteatosis (small lipid vacuoles within hepatocytes). mouse, high phytanic acid levels cause a peripheral neuropathy and ataxia with loss of Purkinje cells. These findings provide important insights in the pathophysiology of Refsum disease. Keywords:fatty acid oxidation, metabolic disorder, peroxisomes Refsum disease is an autosomal recessive lipid-storage disorder (MIM266500) characterized by the accumulation of the branched-chain fatty acid phytanic acid (3,7,11,15-tetramethylhexadecanoic acid) in tissues and body fluids of patients (1). Phytanic acid levels are elevated because of a deficiency of the enzyme phytanoyl-CoA hydroxylase (PHYH). PHYH catalyzes the first step of the peroxisomal -oxidation pathway, i.e., the conversion of phytanoyl-CoA to 2-hydroxy-phytanoyl-CoA. This -oxidation is required for the degradation of phytanic acid, which contains a methyl group at the third carbon atom of its backbone and therefore cannot be degraded via -oxidation. As a result of -oxidation, the terminal carboxyl-group of phytanic acid is usually released as CO2, producing Harmaline pristanic acid, a 2-methyl branched-chain fatty acid (2,6,10,14-tetramethylpentadecanoic acid), which can be degraded via -oxidation in the peroxisome. The molecular basis of Refsum disease is usually heterogeneous, but the majority of patients have mutations in the gene encoding PHYH (2). In humans, thePHYHgene is usually 21 kb and consists of 9 exons and 8 introns. It encodes a protein with a calculated mass of 38.6 kDa, including a cleavable peroxisomal targeting signal type 2 (PTS2). This PTS2-signal is usually recognized by peroxin 7, which is a receptor required for the import of newly synthesized PTS2-made up of proteins into peroxisomes. Peroxin 7 is usually encoded byPEX7, which is the second gene that can be mutated in patients with Refsum disease (2). The impaired import of PHYH into the peroxisome leads to a deficient PHYH activity in patients with mutations in thePEX7gene. Clinically, Refsum disease is usually characterized by cerebellar ataxia, polyneuropathy, and progressive retinitis pigmentosa, culminating in blindness, (1,3). The age of onset of the symptoms can vary from early childhood to the third or fourth decade of life. No treatment is usually available for patients with Refsum disease, but they benefit from a low phytanic acid diet. Phytanic acid is derived from dietary sources only, specifically from the chlorophyll component phytol. When phytanic acid levels are kept low via dietary reduction, a halt in the progression of the symptoms has been reported (47). Although the clinical symptoms of Refsum disease are well known, only limited postmortem data are available on Refsum patients and the pathogenesis of the disorder is usually poorly understood. Insight herein is usually of crucial importance for the development of potential therapeutic options. Phytanic acid has been claimed to be toxic, and it has been shown to be an activating ligand of the nuclear receptor peroxisome proliferator receptor (PPAR). To study the pathological consequences of phytanic acid accumulation, we generated Harmaline of a mouse model for Refsum disease by targeted disruption of thePhyhgene. == Results == == Disruption of thePhyhGene. == To Harmaline disrupt the mousePhyhgene, part of the gene made up of exons 47 was replaced with the hygromycin B-resistance gene by homologous recombination in ES cells as depicted inFig. 1A. Correct targeting ofPhyhwas confirmed by Southern blot analysis and PCR on genomic DNA (Fig. Rabbit Polyclonal to HOXA1 1BandC). The disruption of thePhyhgene resulted in the absence of detectable mRNA (Fig. 1D), PHYH protein, and a complete loss of PHYH enzyme activity (data not shown). The absence of thePhyhgene did not result in embryonic lethality nor did it affect postnatal viability. When kept under standard laboratory conditions,Phyh/mice developed normally and had no obvious developmental abnormalities. They were fertile, and interbreeding ofPhyh/males Harmaline and females led to viable progeny. == Fig. 1. == Generation of targeting vector andPhyh/mice. (A) Strategy of.