Developmental Neurobiulogy, edited by Philippe Evrard and Alexandra Minkowski. Nestle Nutrition Workshop Series, Vol. 12. Nestec Ltd.. Vevey/Raven Press, Ltd., New York © 1989. Peroxisomes and Central Nervous System Dysgenesis and Dysfunction Sidney L. Goldfischer Department of Pathology, Albert Einstein College of Medicine, The Bronx, New York 10461 Peroxisomes play a key role in a number of genetic diseases. These include disor- ders in which the activity of a peroxisomal enzyme is deficient and an extraordinary group of diseases in which the formation of the organelle itself is defective (1-3) (Table 1). DISORDERS OF PEROXISOMAL BIOGENESIS Zellweger's cerebrohepatorenal syndrome (CHRS) is the first disease in which defective formation of peroxisomes was described (4,5). Infants affected with this rare autosomal recessive disorder usually die within one year. Clinical features of the syndrome include a typical facial appearance, with hypertelorism, a high fore- head, and pursed lips; minor skeletal abnormalities; renal cortical cysts; and severe hepatic fibrosis. Iron storage is frequently seen in the early stage of the disease. The most prominent findings are in the central nervous system and include profound hy- potonia. Cerebral abnormalities include polymicrogyria, pachygyria, olivary dys- plasia, and defective neuronal migration. Gliosis and accumulations of lipid in glia are associated with myelin breakdown, and the disease has been described as a suda- nophilic leukodystrophy (6-9). Hepatocellular peroxisomes have not been detected in children with this disease. This remarkable finding has been confirmed by many laboratories (10-13). It is par- ticularly surprising in view of the fact that there are approximately 1,000 peroxi- somes in a normal human hepatocyte (14). In hepatocytes and proximal renal tubules, where peroxisomes also appear to be absent, their diameter is approxi- mately 0.5 microns and normally they are readily detectable. In addition to the per- oxisomal defects, structural and chemical abnormalities in mitochondria have been described (4,5,10,13). The most common has been the report of diminished oxida- tion of succinate; further on in the respiratory pathway, electron transport is normal. 63 64 PEROXISOMES AND CNS DYSGENESIS TABLE 1. Peroxisomal diseases Defective peroxisomal biogenesis syndromes Zellweger's cerebrohepatorenal syndrome neonatal adrenoleukodystrophy infantile Refsum's disease Deficiency of a peroxisomal enzymatic activity X-linked childhood adrenoleukodystrophy pseudo-Zellweger's syndrome (3-oxoacyl-coA thiolase deficiency) acatalasia cerebrotendinous xanthomatosis (?) A variety of chemical abnormalities are seen in patients with CHRS, and many of these reflect the peroxisomal deficiency. These include markedly increased concen- trations of pipecolic acid (15), very long chain fatty acids (VLCFA) in blood and tis- sues (16,17), and the presence of abnormal bile acid intermediates (18,19). The activity of a key peroxisomal membrane enzyme involved in the synthesis of plas- malogens, dihydroxyacetone phosphate acyltransferase (DHAP-AT), is reduced by approximately 90%, and tissue concentrations of plasmalogens are less than 10% of normal in cells and tissues from patients with CHRS (20-22). Assay of DHAP-AT has been utilized in the antenatal diagnosis of CHRS (23). Neonatal adrenoleukodystrophy (NALD) is another disease in which the forma- tion of hepatocellular peroxisomes is defective (24-26). Although not as severely affected as children with CHRS (affected individuals have survived for as long as 6 years), NALD patients also suffer from severe hypotonia and seizures. Plasma con- centrations of VLCFA are increased six- to ninefold and cultured fibroblasts have a markedly diminished capacity to oxidize these fatty acids. Tissue deposits of VLCFA are widespread. We have examined the hepatocytes in two cases of NALD, and in both instances hepatocellular peroxisomes are markedly reduced in size and number (27). They are less than 0.2 |xm in diameter, and one-tenth as numerous as in the normal hepatocyte. Indeed, they are so small and sparse that several reports have been published stating that they are absent in this disease (28). They can be identified unequivocally by incubation in a cytochemical medium that makes visible a marker peroxisomal enzyme, catalase. Other biochemical findings in this disease are similar to CHRS (27). These include elevated concentrations of pipecolic acid, the presence of abnormal bile acid intermediates, and reduced levels of DHAP-AT, a key enzyme in plasmalogen synthesis (Janna Collins personal communication, 1985). The infantile variant of Refsum's disease is the third disorder in which formation of peroxisomes appears to be defective (29). Evidence for this comes from cyto- chemical and ultrastructural studies of the liver (30) and from biochemical studies of serum and fibroblasts (31,32). The classical form of Refsum's syndrome, which is an autosomal recessive disorder, manifests itself in the first or second decade of life PEROXISOMES AND CNS DYSGENESIS 65 and is characterized by retinitis pigmentosa, peripheral polyneuropathy, and cerebel- lar ataxia. Affected individuals have tissue stores of phytanic acid, a 20-carbon fatty acid derived from plant material. Because the stored material is exogenous in origin, a restricted diet is an effective therapy in this condition. The defect has been attrib- uted to the failure in the first step of catabolism of phytanic acid, that is, the alpha oxidation of the terminal carboxyl group. Recently several cases have been de- scribed of young children with phytanic acid storage disease who manifested symp- toms from birth. Electron microscopic studies by Dr. Frank Roels (30) have demonstrated that hepatocytes do not contain recognizable peroxisomes in this con- dition, and biochemical studies (31,32) have shown increased concentrations of the bile acid intermediate trihydroxycoprostanoic acid (THCA), as well as VLCFA. We are not aware of any studies in which the role of peroxisomes in the oxidation of phytanic acid has been analyzed. However, deficient oxidation of phytanic acid has been described in fibroblasts of such patients. We have recently found that serum phytanic acid is increased in CHRS and NALD, two diseases in which peroxisomes are deficient, but not in diseases associated with a more specific enzymatic defect, such as X-linked adrenoleukodystrophy (ALD) or a multiple peroxisomal oxidative activity deficiency syndrome (pseudo-Zellweger's disease) (33,34). This suggests that phytanic acid accumulation is secondary to a deficiency of peroxisomes, and that peroxisomal oxidative activity plays a critical role in the catabolism of phytanic acid. It is noteworthy that the number of peroxisomes is not diminished in fibro- blasts from patients with the classical form of Refsum's disease (35). It is unclear at the present time whether the CHRS, NALD, and infantile form of Refsum's disease are distinct entities or represent varying degrees of expression of a single disorder. It has recently been demonstrated that the absence of peroxisomes in CHRS is not absolute (36). Fibroblasts from four patients with CHRS or related diseases were examined. Two were classical Zellweger patients who died at the age of 6 months, and two others were longer-lived atypical patients (5 years, and alive at 3 years) who probably had NALD. Peroxisomes were detected, but reduced in number in all of these cell lines. Morphometric analysis demonstrated a reduction of approximately 90% in the first group of classical CHRS patients, and of 60% in the patients with longer survival. It will require much more extensive study to deter- mine whether the severity of disease is a consistent reflection of the number of per- oxisomes. The apparent inconsistency between hepatocytes and fibroblasts may reflect variations in the degree to which the deficiency manifests itself in different tissues. Intestinal peroxisomes do not appear to be reduced in NALD (27). It is note- worthy that in mice with testicular feminization syndrome there is a marked reduc- tion in size and number of peroxisomes in the interstitial cells of the testes (37). It is evident that there is considerable overlap between these syndromes (16). Even apparently distinctive pathologic phenomena, such as normal adrenals in CHRS and adrenal atrophy in NALD, may be a reflection of the shorter life span of these individuals. Recent studies have shown functional adrenal insufficiency in children with Zellweger's CHRS (38). Cerebellar heterotopia and olivary dysplasia occur both in classical CHRS and in NALD, but these phenomena have not been de- 66 PEROXISOMES AND CNS DYSGENESIS scribed in the infantile form of Refsum's disease. Other neurological findings, such as retinitis pigmentosa and demyelination, have been observed in all of these condi- tions, even though the degree of involvement may vary considerably. Comparison of conditions in which peroxisomal biogenesis is defective with several other disor- ders in which there appears to be defective activity of a single peroxisomal enzyme may be instructive in relating specific pathologic events to specific enzymatic activi- ties. It must be stressed that there have been no studies of mitochondrial respiration in NALD and infantile Refsum's disease. PEROXISOMAL DISORDERS AFFECTING SPECIFIC ENZYMATIC ACTIVITIES A second group of peroxisomal disorders resemble the better known deficiency diseases. These are diseases in which the