RAPID COMMUNICA TIONS Ultrastructural and Cytochemical Demonstration of Peroxisomes in Cultured Fibroblasts from Patients with Peroxisomal Deficiency Disorders

The oxidation of very long chain fatty acids and synthesis of ether glycerolipids (plasmalogens) occurs mainly in peroxisomes. Zellweger's cerebrohepatorenal syndrome (CHRS) is a rare, inherited metabolic disease characterized by an apparent absence of peroxisomes, an accumulation of very long chain fatty acids, and a decrease of plasmalogens in tissues and cultured fibroblasts from these patients. As peroxisomes are ubiquitous in mammalian cells, we examined normal and CHRScultured fibroblasts for their presence, using an electron microscopic histochemical procedure for the subcellular localization of catalase, a peroxisomal marker enzyme. Small (0.08-0.20/zm) round or slightly oval peroxisomes were seen in both normal and CHRS fibroblasts. The number of peroxisomes was analyzed morphometrically and found to be significantly reduced in all CHRS cell lines. These results are discussed in relation to the underlying defect in peroxisomal function and biogenesis in this disease. Peroxisomes are respiratory organelles that contain H202generating oxidases and catalase (12). They are found in plants, fungi, protozoa, and metazoan organisms, and are ubiquitous in mammalian cells (11). Their unique role in ether glycerolipid (plasmalogen) biosynthesis (7, 8) and very long chain fatty acid oxidation (23) suggests that they are essential to cellular metabolism. Further support for this concept is derived from studies of inborn errors of metabolism where expression of a hereditary abnormality results in defective peroxisomal function and severe multisystem disease (5, 6). Zellweger's cerebrohepatorenal syndrome (CHRS) t is an inherited metabolic disorder characterized by severe nervous system dysfunction, including demyelination and defective neuronal migration, hepatic fibrosis, skeletal abnormalities, and renal cortical cysts (I, 24). Affected infants rarely survive beyond one year. At the subeellular level, the most striking abnormality is an apparent lack ofperoxisomes in hepatocytes (6, 14, 18) and renal proximal tubular epithelia (6). As a disease, CHRS appears to be unique in that the biogenesis of a constitutive organelle is defective. Other syndromes such as neonatal adrenoleukodystrophy share many of the clinical, biochemical, and morphological features of CHRS (1, 4). In f Abbreviation used in this paper. CHRS, cerebrohepatorenal syndrome. contrast to CHRS, reduced numbers of small hepatocellular peroxisomes have been detected in these patients, who are less impaired and survive for as long as six years (4). Biochemical evidence in support of the concept that CHRS represents a profound deficiency in peroxisomes comes from two sets of observations. In the liver,/~-oxidation of very long chain fatty acids (C:22-26) and the initial step in ether glycerolipid (plasmalogen) synthesis occurs mainly, if not exclusively, in peroxisomes (7, 8, 23). Very long chain fatty acids accumulate in tissues and fibroblasts from CHRS patients (1), whereas plasmalogen concentrations and the activity of dihydroxyacetone phosphate acyltransferase, a key enzyme in plasmalogen synthesis, are <10% of normal (3, 9). These data have been interpreted as evidence for the absence of peroxisomes in cultured fibroblasts and tissues from patients with CHRS. The reported absence of peroxisomes in CHRS fibroblasts, which nevertheless grow normally in vitro, and in other tissues is contrary to evidence for an essential role of this organelle in mammalian cell function. Therefore, we conducted an ultrastructural and morphometric study to determine whether peroxisomes are present in normal and CHRS fibroblasts, using a histochemical procedure for the subcellular localization of peroxisomal catalase (15). The results show that small catalase-positive peroxisomes were present in the cytoplasm of normal and CHRS fibroblasts. Morphometric analysis of THE JOURNAL OF CELL BIOLOGY VOLUME 100 MAY 1985 1789-1792 1789 © The Rockefeller University Press. 0021-9525/85/05/1789104 $1.00 on A ril 9, 2017 D ow nladed fom Published May 1, 1985