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Biogenesis of Peroxisomes in Fetal Liver
of Anatomy, Embryology and Histology, Section of Anatomy and Embryology, University of Gent, B-9000 Gent, Belgium
of Cellular and Developmental Biology, University of Rome ‘‘La Sapienza,’’ 00185 Rome, Italy
peroxisome; biogenesis; differentiation; liver; congenital disease; oocyte; inheritance
Peroxisomes are single membrane-limited cell organelles that are involved in
numerous metabolic functions. Peroxisomes do not contain DNA; the matrix and membrane
proteins are encoded by the nuclear genome. It is assumed that new peroxisomes are formed by
division of existing organelles. The present article gives an overview of microscopic studies and
recent unpublished results dealing with peroxisome biogenesis in mammalian fetal liver and
presents data on peroxisomes in oocytes.
Cytochemical (catalase and D-aminoacid oxidase activity) and immunocytochemical data in rat
and human liver (antigens of catalase, the three peroxisomal b-oxidation enzymes, alanine:
glyoxylate aminotransferase, peroxisomal membrane proteins with molecular weights of 42 and 70
kDa) indicate that during embryonic and fetal development the peroxisomal population undergoes a
differentiation with respect to the composition of the matrix and to the size and number of the
organelles. In the youngest stages, rare and small peroxisomes are present, into which the matrix
components are imported in a sequential way. The import seems asynchronous in peroxisomes of the
same hepatocyte. The size and number of the peroxisomes increase during liver development. In rat
and human liver, no morphological or immunocytochemical evidence for an elaborate network of
interconnected peroxisomes (‘‘reticulum’’) was found. Instead, peroxisomes presented as individual
organelles, which occasionally show membrane extensions.
The importance of the metabolic functions of peroxisomes in human liver is emphasized by the
peroxisomal disorders. In the liver of affected fetuses, the microscopic features associated with the
defect can already be recognized; i.e., either catalase containing peroxisomes are absent and
catalase is localized in the cytoplasm (in fetuses affected with Zellweger syndrome or with infantile
Refsum disease) or peroxisomes are present but they are abnormally enlarged (e.g., a fetus affected
with acyl-CoA oxidase deficiency).
In the quail ovary, numerous peroxisomes are observed in the oocyte and in the granulosa cells
during follicle maturation, but not in the full-grown egg. Thus, the mechanism of peroxisome
inheritance remains unresolved. Microsc. Res. Tech. 39:453–466, 1997. r 1997 Wiley-Liss, Inc.
Peroxisomes are single membrane-bound organelles
that are present in all eukaryotic cells. They belong to
the family of microbodies, which includes peroxisomes,
glycosomes, and glyoxysomes. The common feature of
microbodies is that they are limited by a single membrane and that they share a fatty acid b-oxidation
system, which is different from that in mitochondria.
Peroxisomes occur in yeasts, protozoans, plant leaves,
and animal cells. They do not contain DNA; their
proteins are encoded by nuclear genes. The peroxisomal
proteins are synthetized on free ribosomes; they are
incorporated without posttranslational modifications
into existing peroxisomes (except for 3-ketoacylCoA
thiolase and sterol-carrier protein, which are synthesized as a precursor containing an aminoterminal
presequence, and acyl-CoA oxidase, which is cleaved
into two subunits; for reviews, see Borst, 1989; Lazarow
and Fujiki, 1985; Van den Bosch et al., 1992).
The first description of peroxisomes was given by
Rhodin (1954) who designated spherical organelles in
the proximal tubules of mouse kidney as microbodies,
and Rouiller and Bernhard (1956) described microbodies in the liver of the rat. By cell fractionation, De Duve
and Baudhuin (1966) demonstrated that rat liver peroxisomes contain several hydrogen-peroxide-producing
oxidases and catalase; the latter decomposes hydrogen
peroxide in either a catalatic or a peroxidatic way. In
mammalian peroxisomes, more than 40 distinct enzymes have been identified. The enzyme content of
peroxisomes is tissue dependent; the conversion of a
given substrate may be catalyzed by different enzymes
Contract grant sponsor: Nationaal fonds voor Wetenschappelikj Onderzoek;
Contract grant sponsor: Nationale Loterij; Contract grant number: 9.0027.88;
Contract grant sponsor: Flemish Government, Verkennend Europees Onderzoek;
Contract grant sponsor: European Biomed Concerted Action ‘‘Peroxisomal Leukodystrophy’’; Contract grant number: BMH4 CT96-1621 DGXII; Contract grant
sponsor: University of Ghent, Bijzonder Ondersoeksfonds; Contract grant sponsor: Ministero dell’Università e della Ricerca Scientifica e Tecnologica.
*Correspondence to: Marc Espeel, Department of Anatomy, Embryology and
Histology, Section of Anatomy and Embryology, University of Gent, Godshuizenlaan 4, B-9000 Gent, Belgium. E-mail:
Received 18 April 1995; Accepted in revised form 13 October 1995.
in different species. Human liver peroxisomes are involved a.o. in the biosynthesis of plasmalogens, bile
acids, cholesterol, and dolichol and in the degradation
of very-long-chain fatty acids, phytanic acid, and the
transamination of glyoxylate to glycin (for review, see
Mannaerts and Van Veldhoven, 1993; Van den Bosch et
al., 1992; Wanders et al., 1993).
The size and shape of mammalian peroxisomes varies between cell types and between different species. In
human and rat liver, the organelles have more or less a
spherical shape; the matrix is finely granular (Sternlieb
and Quintana, 1977; Wedel and Berger, 1975). Slender,
vermiform peroxisomes cooccurring with spherical organelles are present in human duodenal epithelium
(Roels et al., 1991a). Elongated peroxisomes are also
common in the proximal tubules of human kidney
(Roels and Goldfischer, 1979).
In several cases, the peroxisomal matrix is divided
into subcompartments. In rat liver peroxisomes, urate
oxidase is present in the crystalline core (Völkl et al.,
1988), and D-aminoacid oxidase is confined to an electron-lucent area of the matrix (Usuda et al., 1991a). In
rat renal tubules, the peroxisomal matrix is divided
into a peripheral electron-dense margin and an electronlucent central area. Some enzymes are predominantly
present in the outer dense matrix (e.g., the peroxisomal
b-oxidation enzymes), whereas others are exclusively
localized in the central area (e.g., D-aminoacid oxidase)
(Yokota et al., 1987).
In normal human hepatic peroxisomes, there are no
detectable subcompartments of the matrix. In pathological conditions, however, corelike inclusions have been
reported: in patients with colorectal cancer (Denetto et
al. (1991), in a specific subgroup of patients with
primary hyperoxaluria type 1 (Danpure et al., 1993), in
hepatic peroxisomes of a girl with a NALD-like syndrome (Roels et al., 1988), and in the hepatic and renal
peroxisomes in a male neonate with a peroxisomal
b-oxidation defect (Espeel et al., 1991). Rare hepatic
peroxisomes with a marginal plate were present in a
girl with a NALD-like syndrome (Roels et al., 1988).
Peroxisomelike organelles with an electron-lucent central area have been reported in the liver of a patient
with infantile Refsum’s disease (Roels et al., 1986,
Cytochemical investigations have demonstrated that,
within the same tissue and even within the same cell,
the organelles display heterogeneity regarding their
enzyme activity: catalase activity differs between individual organelles in human liver (Roels, 1991; Roels
and Cornelis, 1989) and staining heterogeneity for
D-aminoacid oxidase activity has been shown in rat
liver and kidney (Angermüller and Fahimi, 1988; Stefanini et al., 1985; also see Fig. 13 of the present article;
for overviews and discussion, see Roels, 1991; Van den
Munckhof (1996).
In addition to the differences in the composition of the
peroxisomal matrix among tissues, Usuda et al. (1991b)
showed that the relative abundance of the membrane
proteins differs among different rat organs.
A remarkable feature of peroxisomes is that, in the
liver of several mammalian species, their proliferation
can be induced by administration of various chemical
substances. The first observations of peroxisome proliferation induced by xenobiotics were made in rats (Hess
et al., 1965). The molecular mechanism of the induction
and the response mechanism have been intensively
investigated. The proliferative response is species and
gender dependent. Human liver peroxisomes do not
respond to induction by these xenobiotics (for review,
see Gibson and Lake, 1993), but proliferation in patients under several conditions has been described (De
Craemer et al. 1991). Proliferation of the peroxisome
population is also observed in regenerating rat liver
after partial hepatectomy (Rigatuso et al., 1970). Peroxisome proliferation induced by drugs or by partial
hepatectomy in rat liver has been used as a model for
the study of mammalian peroxisome biogenesis (for
review, see Fahimi et al., 1993; following section).
The aim of the present article is to give an overview of
microscopic studies on peroxisomes during mammalian
liver organogenesis and in patients with peroxisomal
disorder and to discuss the data with regard to peroxisome biogenesis. We also deal with the topic of peroxisome inheritance.
Two main processes are involved in the formation of a
functional peroxisome: (1) the formation of the organelle as a membranous vesicle and (2) the incorporation
of the matrix and the membrane proteins into this
organelle. We use the term ‘‘biogenesis’’ to denote the
process of organelle formation. The term ‘‘peroxisome
assembly’’ is considered to be the result of processes (1)
and (2).
Organelle Formation
For many years, the concept prevailed that peroxisomes were formed by budding from the endoplasmic
reticulum (‘‘budding theory’’). In previous theories,
peroxisomes were considered as precursors of mitochondria (Roullier and Bernhard, 1956) and of lysosomes
(Bruni and Porter, 1965; for a discussion and overview,
see de la Iglesia, 1969).
The budding theory is based on early observations in
rat liver that peroxisomes and endoplasmic reticulum
often occur in close association. The existence of luminal continuities between both compartments was suggested in several electron micrographs prior to the use
of cytochemical markers (Novikoff and Shin, 1964).
Subsequent cytochemical and immunocytochemical
studies, however, have demonstrated that both compartments are strictly separate entities (Shio and Lazarow,
1981; Yokota and Fahimi, 1980).
Based on multiple evidence (see Lazarow and Fujiki,
1985), it is assumed that new peroxisomes originate
from existing ones through division of a mother organelle.
The ultrastructural basis of the formation of new
peroxisomes has been studied in detail in the model of
peroxisome proliferation by drug induction or by partial
hepatectomy in rat liver (for review, see Fahimi et al.,
1993). From these studies, the events that lead to the
formation of a new organelle can be summarized as
follows. The initial step is the formation of extensions of
the peroxisomal membrane. They are attached to the
‘‘mother’’ peroxisome under the form of myelinlike
figures that develop into looplike double-membrane
structures. Upon import of the matrix proteins, the
loops develop into small peroxisomes. They may remain
TABLE 1. Evolution of size and form-ellipse factor of peroxisomes in fetal human liver
Fetal age (weeks)
d-circle (µm)
Mean uncorrected
Form-ellipse factor
briefly attached to the mother organelle but eventually
separate to become ‘‘new’’ individual peroxisomes. The
membrane extensions are strongly immunoreactive for
the 70-kDa peroxisomal membrane protein and are
considered as a reservoir ready for the import of the
matrix proteins (Baumgart et al., 1989). The 70-kDa
peroxisomal membrane protein is an ATP-binding protein that, through its homology with ABC-transporter
proteins, is likely to be involved in the translocation of
molecules across the peroxisomal membrane (Kamijo et
al., 1990; Valle and Gärtner, 1993). In the same model,
Lüers et al. (1990) demonstrated that the biosynthesis
of the peroxisomal membrane proteins precedes that of
the matrix proteins.
Incorporation of the Matrix Proteins
The peroxisomal matrix and membrane proteins are
synthesized on free ribosomes. Two distinct targeting
sequences that direct the matrix proteins to the peroxisomes have been identified so far in mammalian cells: a
carboxy-terminal S-K-L (Ser-Lys-Leu) tripeptide is necessary and sufficient to direct a protein into the peroxisome (Gould et al., 1988). Limited variations of the
SKL-tripeptide remain functional in peroxisomal targeting: serine may be replaced by alanine or cysteine,
lysine may be substituted by arginine or histidine, but
alternatives for the terminal leucine are not allowed.
The minimal consensus sequence is thus S/A/C-K/R/
H-L; it is also called the S-K-L motif or PTS-1 (peroxisomal targeting signal). The S-K-L motif has been
highly conserved during evolution (Gould et al., 1989).
Swinkels et al. (1991) and Osumi et al. (1991) demonstrated that the aminoterminal cleavable extension of
3-ketoacylCoA thiolase contains a peroxisomal targeting sequence consisting of at least 11 amino acids,
called PTS-2. The receptor for both the PTS-1 and
PTS-2 targeting signal have been recently identified in
man (Braverman et al., 1997; Dodt et al., 1995).
The peroxisomal membrane proteins are targeted via
different signals that have been identified in two proteins (McNew and Goodman, 1996; Wiemer et al.,
1996). For reviews on peroxisomal protein sorting and
import, we refer to Subramani (1993) and to Purdue
and Lazarow (1994).
Incorporation of newly synthesized peroxisomal proteins is taking place continuously in adult liver. Although biochemical studies have suggested that in
adult liver peroxisomes as a whole have the same
turnover rate as some of their proteins, there is little
microscopic evidence for this. More likely, peroxisomes
in healthy adult organs are relatively stable structures,
but some of their components are continuously renewed, i.e., imported into existing structures. For a
discussion on these subjects, see Roels (1991).
Fig. 1. Visualization of peroxisomes in fetal human liver (menstrual age of 8 weeks) as tiny, dark granules after staining for catalase
activity with diaminobenzidine. The organelles are abundant in the
parenchymal cells. In addition, nucleated erythroblasts with black
cytoplasm are seen. Semithin Epon section, phase contrast microscopy. 31,000. Bar, 10 µm.
The role of peroxisomes in human metabolism is
emphasized by the peroxisomal disorders, a group of
rare inherited diseases in which one or more peroxisomal enzymes are deficient. Most often, it concerns
very severe diseases with profound neurological impairment.
Figs. 2–9.
The peroxisomal disorders can be classified, on a
biochemical basis, as three groups: the generalized
peroxisomal disorders or peroxisome biogenesis disorders (e.g., Zellweger syndrome, neonatal adrenoleukodystrophy, infantile Refsum disease) with an impairment of most peroxisomal functions; the multiple
peroxisomal enzyme deficiencies (e.g., rhizomelic
chondodysplasia punctata); and the single peroxisomal
enzyme deficiencies (e.g., X-linked adrenoleukodystrophy, primary hyperoxaluria type I, acyl-CoA oxidase
deficiency) (Wanders et al., 1993). In group I, catalase
containing peroxisomes are absent. In spite of the term
‘‘generalized’’ disorder, some peroxisomal functions may
be unaffected. In patients with a peroxisomal disorder
of group II and group III, hepatic peroxisomes are
present. Except for X-linked adrenoleukodystrophy,
they show diverse alterations (size, shape, number,
appearance and electron density of the matrix after
diaminobenzidine incubation). In addition, in all groups,
other typical histopathological features can be present
(for reviews, see Dimmick and Applegarth, 1993; Roels,
1991; Roels et al., 1991b, 1993b).
The most severe clinical condition is the Zellweger
syndrome or cerebro-hepato-renal syndrome; it is the
prototype of the generalized peroxisomal disorder. Patients with Zellweger syndrome show craniofacial and
skeletal malformations and severe neurologic impairments associated with neuronal migration defects; their
life expectancy is less than 1 year (for review, see
Fournier et al., 1994; Lazarow and Moser, 1995). Cytochemistry has shown that morphologically identifiable
peroxisomes are absent in the liver and kidney of these
patients (Goldfischer et al., 1973). At the time of this
microscopic observation, the metabolic functions of
peroxisomes were largely unknown; the relationship
between the absence of peroxisomes and the biochemical profiles of these patients (accumulation of very-longchain fatty acids, phytanic acid, pipecolic acid, presence
Fig. 2. Normal human fetal liver (14 weeks gestation). Staining
intensity after diaminobenzidine incubation for catalase activity
obviously differs among individual organelles. Usually larger organelles are less electron dense, indicating a lower catalase activity
(Roels, 1991; Roels and Cornelis, 1989). 321,600. Bar, 0.5 µm.
Fig. 3.
Human fetal liver (18 weeks of gestation); cluster of
peroxisomes with strong differences in catalase staining intensity.
Many peroxisomes are seen in close association with cisternae of the
endoplasmic reticulum (arrowheads). 326,400. Bar, 0.5 µm.
Figs. 4, 5. Human fetal liver, 8 (Fig. 4) and 10 (Fig. 5) weeks of
gestation. Tailed and elongated peroxisomes are sometimes seen; the
image is reminiscent of peroxisome proliferation in adult regenerating
rat liver (Yamamoto and Fahimi, 1987). Fig. 4: 356,000; bar, 0.25 µm.
Fig. 5: 324,000; bar, 0.5 µm.
Fig. 6. Human fetal liver (23 weeks of gestation). Peroxisomes with
a somewhat angular shape exhibit different catalase staining. L,
lipofuscin granule with heterogeneous contents. Numerous glycogen
rosettes are seen. 325,000. Bar, 0.5 µm.
Figs. 7–9. Human fetal liver (17 weeks of gestation). Peroxisomes
are in close association with cisternae of granular (Fig. 7) and smooth
endoplasmic reticulum. Although several peroxisomes are nearly in
contact with each other, membrane continuities are not found. In
Figure 8, adjacent peroxisomes have different concentrations of
catalase reaction product, which argues against a continuous exchange of matrix components. In Figure 9, a cisterna of the endoplasmic reticulum is situated in between two peroxisomes (arrow). Fig. 7:
333,600; Fig. 8: 356,000; Fig. 9: 344,000. Bar, 0.5 µm.
of bile acid intermediates, deficiency of plasmalogens)
became established later (Heymans et al., 1983; Schutgens et al., 1986). The absence of hepatic peroxisomes
has also been demonstrated in patients with related,
milder syndromes, e.g., infantile Refsum disease (Roels
et al., 1986).
The absence of peroxisomes was considered to result
from a defect in organelle formation. Subsequent investigations have demonstrated that in cultured fibroblasts from patients with Zellweger syndrome ‘‘peroxisomal membrane ghosts’’ are present; these are enlarged
and seemingly empty organelles, recognized as peroxisomes through the immunoreactivity of their membrane for peroxisomal membrane proteins. The membrane ghosts were considered to represent peroxisomes,
which are unable to import the matrix proteins into the
organelle (Santos et al., 1988a,b; Wiemer et al., 1989).
The existence of ghosts in the liver of patients (Espeel
et al., 1995a) is discussed below.
Complementation studies with cultured skin fibroblasts from patients with generalized peroxisomal disorders have shown that these diseases are genetically
heterogeneous. Eleven complementation groups in peroxisome biognesis disorders have been identified so far,
suggesting that in man at least 11 different genes are
involved in the formation of import competent organelles (Moser et al., 1995; Poulos et al., 1995; Shimozawa
et al., 1993; Yajima et al., 1992). For some complementation groups, the involved gene and gene product have
been identified: a 35-kDa peroxisomal membrane protein (PMP), called PAF-1 (peroxisome assembly factor1). Its precise role in peroxisome assembly is unknown;
transfection of cultured skin fibroblasts from the patients with wild type PAF-1-cDNA restored peroxisome
assembly and functioning. An experimental model for
this complementation group is available: in a peroxisome deficient mutant line of Chinese hamster ovary
cells (CHO-Z65), the defect is in the PAF-1 encoding
gene (Shimozawa et al., 1992a; Tsukamoto, et al.,
1991). Another complementation group is associated
with the PXR1 gene, which encodes the PTS-1 receptor
protein (Dodt et al., 1995). In addition, a defect in the
gene encoding the 70-kDa PMP provokes a peroxisome
biogenesis disorder (Gärtner et al., 1992).
In addition to the CHO cell mutants (e.g. Shimozawa
et al., 1992b; Thieringer and Raetz, 1993), various
mutants of diverse yeast species are currently under
intensive study to elucidate peroxisome biogenesis and
assembly at the molecular level. The studies in yeast
fall outside the scope of the present text (for a review,
see Purdue and Lazarow, 1994). A unified nomenclature
to designate the homologous genes in various organisms and their products (‘‘peroxins’’) responsible for
peroxisome assembly has been introduced (Distel et al.,
Normal Fetal Human Liver1 (Overview)
In a series of 15 fetal liver samples, with an in utero
age of 8–23 weeks, Kerckaert (1990) described catalasecontaining peroxisomes in the parenchymal cells of all
observations refer to peroxisomes in the parenchymal cells.
Fig. 10. Fetal liver of 7 weeks (menstrual age). Diaminobenzidine
incubation revealed the cooccurrence of catalase-containing (arrowhead) and catalase-unreactive (star) peroxisomes. The arrow indicates
an image of budding in a catalase-unreactive peroxisome. The inset
shows three catalase-negative peroxisomes (star) present in a 6-week
specimen (menstrual age). The matrix of the catalase-unreactive
organelles is reticular in both samples. 327,600. Inset: 325,500. Bar,
0.2 µm.
TABLE 2. Morphometry of peroxisomes in human liver
Normal liver
6 weeks1 7 weeks1 17 weeks2 20 weeks2 6 weeks2 4 months2 Adult3
Number measured
d-circle (µm)
Mean of profiles
Mean corrected5
Maximum measured
Volume density (%)
Numerical density (µm23)
Surface density (µm21)
Form ellipse
‘‘Empty’’-looking peroxisomes
in generalized peroxisomal disorders4
et al. (1993a).
et al. (1991b).
samples; De Craemer et al. (1993).
4Two patients; Espeel et al. (1995a).
5Corrected for sectioning effect according to formula 9 from Abe et al. (1983).
6Not determined.
samples. The size of the organelles (see Table 1) and
their electron density after staining for catalase activity showed an increase during development; it reached
a plateau value at the 17th week. Kerckaert (1990) also
noticed the presence of irregularly shaped organelles,
organelles with membrane extensions (‘‘tailed’’ peroxi-
somes), clusters of peroxisomes, and a heterogeneity of
staining for catalase activity between individual organelles. In the clusters of adjacent peroxisomes, cisternae
of endoplasmic reticulum were present in between the
organelles (Figs. 1–9). The presence of elongated peroxisomes was reflected in the low minimal value of the
Fig. 11. Liver (18th week menstrual age) from a fetus affected with
Zellweger syndrome. Immunostaining for catalase protein results in a
diffuse reaction in the cytoplasm of the parenchymal cells; in many
parenchymal cells, catalase immunoreactivity is also present in the
form-ellipse factor (i.e., the ratio between the long and
the short axis of the object; see Table 1).
Roels (1991) reported that in human fetal liver
parenchyma images of budding (dividing) peroxisomes
are rarely observed in these rapidly proliferating cells
and that, in the clusters of peroxisomes, adjacent
organelles display different amounts of diaminobenzidine reaction product. He concluded that if new organelles are formed by division of a mother organelle, the
process of fission must proceed rapidly and that the
separated mother and daughter organelles must have
acquired their own individual enzyme content soon
after the division.
In a 6-week fetus (menstrual age), catalase-containing peroxisomes were not found after diaminobenzidine
(DAB) cytochemistry; instead, small bodies with a
reticular matrix and probably corresponding to a primitive stage of peroxisome development were present
(Roels, 1991) (inset of Figure 10).
In a 7-week specimen (menstrual age), similar catalase-negative organelles were present. They cooccurred
in the same cell with catalase-positive organelles, the
latter representing one-third of the population (Fig.
Fig. 12. Catalase immunostaining in infant control liver shows a
granular reaction pattern, reflecting a peroxisomal localization of
catalase; the nuclei are unreactive. 3750. Bar, 10 µm in Figs. 11, 12.
Fig. 13. D-aminoacid oxidase cytochemistry in the liver of 15-day
old fetal rat. Peroxisomes of different staining degrees are recognizable (arrowheads) in the hepatocyte cytoplasm; the star marks an
unreactive peroxisome. 335,100. Bar, 0.2 µm.
Fig. 14. Small peroxisome with diaminobenzidine reactive matrix
in a presumptive hepatocyte of rat embryonic liver at 11.5 days
postcoitum. Organelles are very rare at this stage. 372,000. Bar,
0.2 µm.
Fig. 15. Small peroxisome with immunoreactive matrix for acylCoA oxidase in a presumptive hepatocyte of rat embryonic liver at 12.5
days postcoitum. 372,000. Bar, 0.20 µm.
10). In these young stages, the peroxisomes were distinctly smaller than in older fetuses and in postnatal
liver (see Table 2). There were no significant size
differences between the catalase-containing and the
catalase-negative organelles. The catalase-positive organelles showed an extreme staining heterogeneity for
catalase activity. This activity was considered, with the
presence of the catalase-negative organelles, to reflect
the onset of catalase import into the peroxisomes. In
addition, we have suggested that in human embryonic
liver catalase is imported in an asynchronous way (see
Fig. 10) into individual, preexisting organelles (Espeel
et al., 1993a).
Only later during development do the three peroxisomal b-oxidation enzymes become detectable by immunocytochemistry: acyl-CoA oxidase and bifunctional
enzyme in the 10th week and 3-ketoacylCoA thiolase in
the 9th week (menstrual age). The immunoreactive
signal for thiolase in simultaneously treated sections of
the different stages showed a distinct increase of staining intensity with fetal age (Espeel et al., 1990). An
increase of the intensity of catalase staining during
development was also noticed (Kerckaert, 1990). Both
observations indicate a progressive and continuous
import of the peroxisomal matrix proteins during development.
Our studies could not confirm the observation by de
la Iglesia (1969) that in human fetal liver in the 6th
week of gestation a nucleoid is present in the peroxisomal matrix.
sent in the unprocessed form (Wanders et al., 1995) and
that bifunctional enzyme is degraded (Tager et al.,
1985; Wanders et al., 1995).
In the liver of a 16-week fetus, in which previous
microscopic and biochemical examinations of the chorionic villi demonstrated that it was affected with Zellweger syndrome (Roels et al., 1993a), catalase was
immunolocalized in the cytoplasm of the parenchymal
cells. Immunoreactivity for catalase was also present in
at least a part of the nuclei (Fig. 11). This type of
catalase immunolocalization is typically found in postnatal liver of patients with a generalized peroxisomal
disorder but not in pre- or postnatal control liver (Fig.
12). Peroxisomes were also absent in the liver of a fetus
with infantile Refsum disease (Poll-The et al., 1987).
Enlarged peroxisomes have been reported in a fetus
affected with acyl-CoA oxidase deficiency (De Craemer
et al., 1991) and in a fetus with rhizomelic chondrodysplasia punctata (Roels et al., 1991b). Thus, the typical
feature of enlarged peroxisomes in the postnatal liver
associated with these syndromes (De Craemer et al.,
1991; Heymans et al., 1986; Hughes et al., 1992;
Poll-The et al., 1988) is already observed in the fetal
liver. For a discussion on enlarged hepatic peroxisomes
in peroxisomal disorder patients, see Roels (1991).
Otherwise, in the liver of a stillborn fetus with X-linked
recessive chondrodysplasia punctata (peroxisomal functions are normal with this disease; the fetus was
delivered in the 26th postmenstrual week), no alterations of the peroxisomes were found (Van Maldergem
et al., 1991).
Fetal Liver Affected by Peroxisomal Disorders
In the group I peroxisomal disorders (e.g., Zellweger
syndrome), the peroxisomal matrix proteins are not
imported into peroxisomes, and they remain localized
in the cytoplasm. Despite this mislocalization, several
enzymes remain stable and active (e.g., catalase, alanine/glyoxylate aminotransferase; Roels, 1991; Wanders et al., 1984, 1987). Immunoblotting studies have
shown that in Zellweger liver the b-oxidation enzymes
acyl-CoA oxidase and 3-keto-acylCoA thiolase are pre-
Fetal Rat Liver (Overview)
Without the use of cytochemical markers, Tsukada et
al. (1968) described microbodylike organelles in fetal
rat liver from 15 days onward (the youngest stage
examined) and reported an evident increase of their
number between the 15th and 18th days. Activity
measurements showed that catalase, urate oxidase,
and D-amino acid oxidase became detectable sequentially. Stefanini et al. (1985) found that catalase-
Fig. 16. Granulosa cell of a primordial (prelampbrush) quail
follicle after diaminobenzidine incubation for catalase activity. Five
peroxisomes are recognized due to the precipitated diaminobenzidine;
their membranes are clearly visible. Cytoplasm also contains mitochondria, rough endoplasmic reticulum, and free ribosomes. BM, basement
membrane of the granulosal epithelium. 337,500. Bar, 0.5 µm.
Fig. 17. Primordial follicle (prelampbrush stage) after diaminobenzidine incubation for catalase activity. G, monolayered granulosa with
groups of rounded peroxisomes; O, oocyte with wormlike peroxisomes
(arrow); N, granulosal nuclei. Phase contrast image of semithin (2 µm)
Epon section. 31,750. Bar, 10 µm.
Fig. 18. Oocytal peroxisome with a tubular form in primordial
follicle after diaminobenzidine incubation for catalase; membrane is
clearly visible. A cisterna of smooth endoplasmic reticulum is in close
proximity to the peroxisome. 360,000. Bar, 250 nm.
Fig. 19. Catalase immunoreactivity in a prelampbrush oocyte; the
gold particles are concentrated in a wormlike structure (cf. Fig. 19);
the mitochondrial (M) profiles are devoid of gold particles. 336,000.
Bar, 0.5 µm.
containing organelles were present at the 14th day of
intrauterine life and that D-amino acid oxidase activity
was detectable from the 16th day. The import of the
latter enzyme does not occur simultaneously in all
peroxisomes because organelles with strong and weak
reaction products coexist in the same cell with unreactive organelles (Fig. 13).
Recent Studies in Human and Rat Liver
Collaborative studies by our group and Stefanini’s
group in young embryonic rat liver (10.5–15.5 days
postcoitum) have provided new data on the differentiation of the peroxisome population during liver development. Already at 10.5 days, peroxisomes were visualized by their catalase activity in presumptive
hepatocytes; they were very rare and small. At 11.5
days, catalase protein and the 43-kDa membrane protein were detectable in the peroxisomal matrix and
membrane, respectively, and at 12.5 days immunoreactivity for the three peroxisomal b-oxidation enzymes
was demonstrated in the matrix (Figs. 14, 15). In none
of these stages was a nucleoid found; it was seen at 18
days. The size of the organelles increased from 13.5
days onward and at 15.5 days (d-circle 5 0.27 µm), it
was almost twofold the value of the initial stages
(10.5–12.5 days; d-circle 5 0.16 µm) but only two-thirds
of that in adult rat liver. These are the youngest stages
examined so far in rat liver.
In human embryonic liver staged at 38 days postfertilization, peroxisomes contained immunoreactive catalase, AGT, acyl-CoA oxidase, 3-ketoacylCoA thiolase,
and the peroxisomal membrane immunoreacted for the
43- and 70-kDa membrane proteins (Depreter et al.,
In the liver of patients with a generalized peroxisomal disorder, rare and empty-looking organelles,
identified as abnormal peroxisomes by the reactivity of
their membrane for a 43-kDa peroxisomal membrane
protein, were demonstrated. Two different types were
identified, and they are considered to be the homologue
of the ‘‘peroxisomal membrane ghosts’’ in the fibroblasts
of Zellweger patients, i.e., organelles uncapable to
properly import their matrix components (Espeel et al.,
1995a). The size of these organelles is obviously smaller
than that of the peroxisomes in postnatal control liver
(Table 2).
In other patients, a mosaic distribution of parenchymal cells with and without peroxisomes was found
(Espeel et al., 1995b; Giros et al., 1995; Mandel et al.,
1994; Roels et al., 1996a). Several hypotheses have
been proposed to explain the mosaic distribution, for
example, that the microenvironment (extracellular matrix) influences the differentiation of the peroxisome
population. This hypothesis was inferred from the facts
that (1) hepatocyte-specific features are determined by
tissue interactions early during development (e.g. Caron,
1990; Cascio and Zaret, 1991) and (2) the number, size,
shape, and catalase activity of peroxisomes in cultured
hepatocytes depend on the culture conditions (Furukawa et al., 1988; Mitaka et al., 1993; Roels et al.,
1996a). Thus far, studies of early embryonic rat liver
have not confirmed a relationship between peroxisome
maturation and the presence of extracellular matrix
proteins laminin and collagen IV (Depreter et al., 1997).
Another explanation for the mosaic distribution is
derived from the observation that new peroxisomes
originate by division of existing organelles. This notion
implies that each eukaryotic cell should contain at least
one peroxisome throughout its life. At the moment of
mitotic division, the peroxisomes should be present in a
number that is sufficient to guarantee that the organelles segregate to both daughter cells. If a daughter cell
without peroxisomes were to originate, then all of its
offspring should remain peroxisome deficient. Warren
and Wickner (1996) distinguished a stochastic and an
ordered inheritance of cell organelles. Recent studies
have shown that in different cell types peroxisomes are
associated with microtubules and that the organelles
are capable of making directional movements along the
microtubules (Rapp et al., 1996; Schrader et al., 1996;
Wiemer et al., 1997). According to Wiemer et al.,
peroxisomes in CV1 cells (green monkey kidney cells)
distribute themselves in a stochastic rather than an
ordered way to the daughter cells at the time of mitosis,
the majority of peroxisomes not being associated with
microtubules of the mitotic spindle.
In line with the concept of fission of existing peroxisomes to form new organelles, peroxisomes must be
present in germ cells to be passed to the next generation. Goldfischer (1987) assumed, as it is the case for
mitochondria, that peroxisomes are delivered to the
zygote by the oocyte. Therefore, the identification of
peroxisomes in oocytes is an intriguing question. We
have examined by catalase cytochemistry and immunocytochemistry the presence of peroxisomes in follicles
from the Japanese quail (Coturnix coturnix japonica)
(Roels et al., 1996b).
Numerous diaminobenzidine reactive peroxisomes
with a tubular shape are present in the oocytes during
the small previtellogenic stages (follicle diameter 5 50–
500 µm; Fig. 17). The tubular organelles often show a
constricted middle part and they are frequently lie in
the vicinity of smooth endoplasmic reticulum cisternae
(Fig. 18). After immunostaining against catalase, the
label is concentrated over organelles with a similar size
and shape as the DAB-reactive peroxisomes (Fig. 19).
In the larger previtellogenic follicles (lampbrush stage
II: 0.5–1.5 mm), the organelles are smaller and much
fewer; their shape is spherical. In all vitellogenic stages
(postlampbrush oocytes: 1.5–19 mm) examined so far,
DAB-reactive peroxisomes were no longer observed.
Granulosa Cells
The granulosa cell layer in previtellogenic stages
contains numerous and large round-to-ovoid peroxisomes; they sometimes have an angular outline and
they occur in groups (Fig. 16). In vitellogenic follicles
with diameters of 1.5–10 mm, peroxisomes are present,
but their size (100–200 nm) is smaller than that in the
previtellogenic stages (300–500 nm). Remarkably, in
the three largest preovulatory follicles, i.e., yellow
follicles with diameters of 10–19 mm, DAB-containing
peroxisomes are no longer found.
The observations are interpreted as follows. The
image of elongated peroxisomes connected with a DABunreactive stalk in the small previtellogenic follicles
represents morphological evidence for the formation of
new organelles (cf. Baumgart et al., 1989). At this stage
of oocyte development, new peroxisomes must be formed
(biogenesis) at a high rate to match the large volume
expansion of the oocyte. Indeed, peroxisomes are still
detected in lampbrush II stage. The microscopic disappearance of peroxisomes in the vitellogenic oocytes is
difficult to interpret. Volume expansion is enormous (a
factor of 2,000 for growth from 1.5 to 19 mm oocyte) and
could easily result in a ‘‘dilution effect.’’ A second
explanation could be a stop in catalase synthesis during
the last weeks of oocyte maturation: the normal enzyme
turnover will result in organelles devoid of catalase.
The absence of catalase-containing peroxisomes in the
granulosa cells of mature follicles does not result from a
volume expansion of the cells. It appears that these
cells, which in birds die after ovulation, do not sustain a
differentiated peroxisome population until the end of
their life span.
The observations in fetal human and rat liver indicate that even in the youngest stages examined peroxisomes are detectable; they are very rare and small.
During development, the matrix seems to obtain its
complete composition in a stepwise way: e.g., in rat
liver catalase (antigen and activity) is detectable prior
to the three b-oxidation enzymes and the nucleoid in
the rat liver peroxisomes appears still later. This
finding suggests a sequential import, which seems to be
asynchronous for individual organelles, given the interperoxisomal staining heterogeneity. The sequential and
asynchronous import reflects a differentiation process
(‘‘maturation’’) of individual peroxisomes, which is also
associated with an increase of the size of the organelles.
The factors governing this differentiation are unknown, and it is an open question as to whether these
factors are specific for a given tissue. A parallel study of
the peroxisome population of, e.g., the primitive gut
endoderm outside the liver bud or of neuroepithelial
cells could be relevant to this question. According to the
observations by Depreter et al. (1997), there is no direct
role for laminin or collagen IV in the differentiation of
peroxisomes. With respect to the question of organelle
inheritance, it is sufficient that peroxisomes are present in the ‘‘immature’’ form, provided their number is
high enough to ensure segregation to the daughter
cells. The necessity for a sufficient number of organelles
implies that these immature organelles should be capable to divide. The empty-looking membrane ghosts in
the postnatal liver of patients with a generalized peroxisomal disorder (into which import and incorporation of
functional proteins have failed) could be considered as
homologous to the immature peroxisomes, and they
both share the feature of a small size.
In view of the suggested differentiation process, the
observations on peroxisomes during kidney embryonic
development should be mentioned: whereas, in murine
(Pipan and Psenicnik, 1975) and human fetuses (Bri-
ère, 1986), DAB-reactive peroxisomes are found as
early as the 14th day and the 10th week of gestation,
respectively, in rat proximal tubules the appearance of
catalase-containing peroxisomes occurs as an abrupt
phenomenon at the end of the prenatal period; in the
preceding stages catalase-devoid organelles could not
be detected (Stefanini et al., 1994). The dramatic
increases of peroxisomal number and enzymatic content, found near birth in mouse and rat kidney proximal tubules, suggest a specific role for these organelles
during the maturation of this part of the nephron
(Goeckermann and Vigil, 1975; Pipan and Psenicnik,
1975; Stefanini et al., 1995). The existence of the
membranous compartment (immature peroxisomes) in
rat kidney at younger stages can be usefully investigated by immunocytochemistry for the peroxisomal
membrane proteins.
The cytochemical demonstration of peroxisomes in
oocytes is a recently explored domain. During growth of
the quail oocyte, peroxisomes apparently proliferate. In
mature eggs, however, they can no longer be detected by
catalase cytochemistry, which may be a result of the
sensitivity of our methods or the matrix of the peroxisomes transiently may not contain catalase. The question remains open as to how peroxisomes are inherited
by the zygote.
We thank Dr. E. Jauniaux (Université Libre de
Bruxelles, Belgium), Dr. N. Brière (Université de Sherbrooke, Québec, Canada), and Prof. P. Thorogood and
Dr. C.T.J. Chan (Institute of Child Health, MRC Human Embryonic Tissue Bank, London, UK) for providing the fetal human liver samples. The studies were
supported by grants from the Nationaal Fonds voor
Wetenschappelijk Onderzoek (Krediet aan Navorsers
to K.D., M.E., and F.R.), the Nationale Loterij
(9.0027.88.) to F.R., the Flemish Government (Verkennend Europees Onderzoek) to F.R., the European
Biomed Concerted Action ‘‘Peroxisomal Leukodystrophy’’ (BMH4 CT96-1621 DGXII), the University of
Ghent (Bijzonder Onderzoeksfonds), and the Ministero
dell’Università e della Ricerca Scientifica e Tecnologica
Abe, K., Matsushima, S., and Mia, M. (1983) Estimation of the mean
size of spheres from the profiles in ultrathin sections: A stereological
method. J. Electron Microsc., 32:57–60.
Angermüller, S., and Fahimi, H.D. (1988) Heterogenous staining of
D-amino acid oxidase in peroxisomes of rat liver and kidney.
Histochemistry, 88:277–285.
Baumgart, E., Völkl, A., Hashimoto, T., and Fahimi, H.D. (1989)
Biogenesis of peroxisomes: Immunocytochemical investigation of
peroxisomal membrane proteins in proliferating rat liver peroxisomes and in catalase-negative membrane loops. J. Cell Biol.,
Borst, P. (1989) Peroxisome biogenesis revisited. Biochim. Biophys.
Acta, 1008:1–13.
Braverman, N., Steel, G., Obie, C., Moser, A., Moser, H., Gould, S.J.,
and Valle, D. (1997) Human PEX7 encodes the peroxisomal PTS2
receptor and is responsible for rhizomelic chondrodysplasia punctata. Nature Genet., 15:369–376.
Brière, N. (1986) Peroxisomes in human foetal kidney: Variations in
size and number during development. Anat. Embryol. 174:235–242.
Bruni, C., and Porter, K.R. (1965) The fine structure of the parenchymal cells of the normal rat liver. I. General observations. Am. J.
Pathol., 46:691–755.
Caron, J.M. (1990) Induction of albumin gene transcription in hepato-
cytes by extracellular matrix proteins. Mol. Cell Biol., 10:1239–
Cascio, S., and Zaret, K.S. (1991) Hepatocyte differentiation initiates
during endodermal-mesenchymal interactions prior to liver formation. Development, 113:217–225.
Danpure, C.J., Purdue, P.E., Fryer, P., Griffiths, S., Allsop, J., Lumb,
M.J., Guttridge, K.M., Jennings, P.R., Scheinman, J.I., Mauer, S.M.,
and Davidson, N.O. (1993) Enzymological and mutational analysis
of a complex primary hyperoxaluria type I phenotype involving
alanine:glyoxylate aminotransferase peroxisome-to-mitochondrion
mistargeting and intraperoxisomal aggregation. Am. J. Hum. Genet.,
De Craemer, D., Zweens, M.J., Lyonnet, S., Poll-The, B.T., Schutgens,
R.B.H., Wanders, R.J.A., Waelkens, J.J.J., Saudubray, J.M., and
Roels, F. (1991) Very large peroxisomes in peroxisomal deficiency
disorders (rhizomelic chondrodysplasia punctata and acyl-CoA oxidase deficiency): Novel data. Virchows Arch. A Pathol. Anat., 419:523–
De Craemer, D., Kerckaert, I., and Roels, F. (1991) Hepatocellular
Peroxisomes in Human Alcoholic and Drug-induced Hepatitis: A
Quantitative Study. Hepatology, 14:811–817.
De Craemer, D., Pauwels, M., Hautekeete, M., and Roels, F. (1993)
Alterations of hepatocellular peroxisomes in patients with cancer.
Cancer, 71:3851–3858.
De Duve, C., and Baudhuin, P. (1966) Peroxisomes (microbodies and
related particles). Physiol. Rev., 46:323–357.
de la Iglesia, F.A. (1969). Comparative analysis of hepatic microbodies
(a review). Acta Hepato-Splenol., 16:141–160.
Denetto, L.A., Tappia, P.S., Malik, Z.A., Wood, A.J., Mann, V.M., Jones,
C.J., and Burdett, K. (1991) Human hepatic peroxisomes with
crystalloid cores associated with urate oxidase activity. Adv. Exp.
Med. Biol., 309A:373–376.
Depreter, M., Nardacci, R., Tytgat, T., Espeel, M., Stefanini, S., and
Roels, F. (1997) Extracellular matrix deposition precedes completion
of peroxisome development in liver. Biol. Cell (submitted)
Dimmick, J.E., and Applegarth, D.E. (1993) Pathology of peroxisomal
disorders. Perspect. Pediatr. Pathol., 17:45–98.
Distel, B., Erdmann, R., Gould, S.J., Blobel, G., Crane, D.I., Cregg,
J.M., Dodt, G., Fujiki, Y., Goodman, J.M., Just, W.W., Kiel, J.A.K.W.,
Kunau, W-H., Lazarow, P.B., Mannaerts, G.P., Moser, H.W., Osumi,
T., Rachubinski, R.A., Roscher, A., Subramani, S., Tabak, H.F.,
Tsukamoto, T., Valle, D., van der Klei, I., van Veldhoven, P.P., and
Veenhuis, M. (1996) A unified nomenclature for peroxisome biogenesis factors. J. Cell Biol., 135:1–3.
Dodt, G., Braverman, N., Wong, C., Moser, A., Moser, H., Watkins, P.,
Valle, D., and Gould, S.J. (1995) Mutations in the PTS-1 receptor
gene, PXR1, define complementation group 2 of the peroxisome
biogenesis disorders. Nature Genet., 9:115–125.
Espeel, M., Jauniaux, E., Hashimoto, T., and Roels, F. (1990) Immunocytochemical localization of peroxisomal b-oxidation enzymes in
human fetal liver. Prenat. Diagn., 10:349–357.
Espeel, M., Roels, F., De Craemer, D., Dacremont, G., Wanders, R.J.A.,
Van Maldergem, L., and Hashimoto, T. (1991) Peroxisomal localization of the immunoreactive b-oxidation enzymes in a neonate with a
b-oxidation defect. Pathological observations in liver, adrenal cortex
and kidney. Virchows Archiv A Pathol. Anat., 419:301–308.
Espeel, M., Brière, N., De Cramer, D., Jauniaux, E., and Roels, F.
(1993a) Catalase-negative peroxisomes in human embryonic liver.
Cell Tissue Res., 272:89–92.
Espeel, M., Heikoop, J.C., Smeitink, J.A.M., Beemer, F.A., De Craemer, D., Van den Berg, M., Hashimoto, T., Wanders, R.J.A., Schutgens, R.B.H., Poll-The, B.T., and Roels, F. (1993b) Cytoplasmic
catalase and ghost-like peroxisomes in the liver from a child with
atypical chondrodysplasia punctata. Ultrastruct. Pathol., 17:623–
Espeel, M., Roels, F., Giros, M., Mandel, H., Peltier, A., Poggi, F.,
Poll-The, B.T., Smeitink, J.A.M., Van Maldegem, L., and Santos,
M.J. (1995a) Immunolocalization of a 43kDa peroxisomal membrane protein in the liver of patients with generalized peroxisomal
disorders. Eur. J. Cell Biol., 67:319–327.
Espeel, M., Mandel, H., Poggi, F., Smeitink, J.A.M., Wanders, R.J.A.,
Kerckaert, I., Schutgens, R.B.H., Saudubray, J.M., Poll-The, B.T.,
and Roels, F. (1995b) Hepatic mosaicism in peroxisomal deficiency
patients. Hepatology, 22:497–504.
Fahimi, H.D., Baumgart, E., and Völkl, A. (1993) Ultrastructural
aspects of the biogenesis of peroxisomes in rat liver. Biochimie,
Fournier, B., Smeitink, J.A.M., Dorland, L., Berger, R., Saudubray,
J.M., and Poll-The, B.T. (1994) Peroxisomal disorders: A review. J.
Inher. Metab. Dis., 17:470–486.
Furukawa, K., Mochizuki, Y., Sawada, N., Gotoh, M. and Tsukada, H.
(1988) Morphometric and cytochemical evaluation of clofibrateinduced peroxisomal proliferation in adult rat hepatocytes cultured
on floating collagen gels. Virchows Arch. B (Cell Pathol.), 55:279–
Gärtner, J., Moser, H., and Valle, D. (1992) Mutations in the 70K
peroxisomal membrane protein gene in Zellweger syndrome. Nature
Genet., 1:16–23.
Gibson, G., and Lake, B. (1993) Peroxisomes: Biology and Importance
in Toxicology and Medicine. Taylor & Francis, Washington, DC.
Giros, M., Roels, F., Ruiz, M., Ribes, A., Prats, J., Espeel, M., Wanders,
R.J.A., Schutgens, R.B.H., and Pampols, T. (1996) Long survival in a
case of peroxisomal biogenesis disorder with peroxisome mosaicism
in the liver. Ann. NY Acad. Sci., 80:747–749.
Goeckermann, J.A., and Vigil, E.L. (1975) Peroxisome development in
the metanephric kidney of the mouse. J. Histochem. Cytochem.,
Goldfischer, S.L. (1987) Pathogenesis of Zellweger’s cerebro-hepatorenal syndrome and related peroxisome deficiency disorders. In:
Peroxisomes in Biology and Medicine. H.D. Fahimi and H. Sies, eds.
Springer Verlag, Berlin, pp. 323–334.
Goldfischer, S., Moore, C.L., Johnson, A.B., Spiro, A.J., Valsamis, M.P.,
Wisniewski, H.K., Ritch, R.H., Norton, W.T., Rapin, I., and Gartner,
L.M. (1973) Peroxisomal and mitochondrial defects in the cerebrohepato-renal syndrome. Science, 182:62–64.
Gould, S.J., Keller, G., and Subramani, S. (1988) Identification of
peroxisomal targeting signals located at the carboxy terminus of
four peroxisomal proteins. J. Cell Biol., 107:897–905.
Gould, S.J., Keller, G., Hosken, N., Wilkinson, J., and Subramani, S.
(1989) A conserved tripeptide sorts proteins to peroxisomes. J. Cell
Biol., 108:1657–1664.
Hess, R., Stäubli, W., and Riess, W. (1965) Nature of the hepatomegalic
effect produced by ethyl-chlorophenoxy-isobutyrate in the rat. Nature, 208:856–858.
Heymans, H.S.A., Schutgens, R.B.H., Tan, R., Van Den Bosch, H., and
Borst, P. (1983) Severe plasmalogen deficiency in tissues of infants
without peroxisomes (Zellweger syndrome). Nature, 306:69–70.
Heymans, H.S.A., Oorthuys, J.W.E., Nelck, G., Wanders, R.J.A.,
Dingemans, K.P., and Schutgens, R.B.H. (1986) Peroxisomal abnormalities in rhizomelic chondrodysplasia punctata. J. Inher. Metab.
Dis., 9:29–31.
Hughes, J.L., Poulos, A., Crane, D.I., Chow, C.W., Sheffield, L.J., and
Silence, D. (1992) Ultrastructure and immunocytochemistry of
hepatic peroxisomes in rhizomelic chondrodysplasia punctata. Eur.
J. Pediatr., 151:829–836.
Kamijo, K., Taketani, S., Yokota, S., Osumi, T., and Hashimoto, T.
(1990) The 70-kDa peroxisomal membrane protein is a member of
the Mdr (P-glycoprotein)-related ATP-binding protein superfamily.
J. Biol. Chem., 265:4534–4540.
Kerckaert, I. (1990) Peroxisomes: Experimental and Human Studies.
Thesis, Vrije Universiteit Brussel.
Lazarow, P.B., and Fujiki, Y. (1985) Biogenesis of peroxisomes. Ann.
Rev. Cell Biol., 1:489–530.
Lazarow, P.B., and Moser, H.W. (1995) Disorders of peroxisome
biogenesis. In: The Metabolic and Molecular Bases of Inherited
Disease, 7th ed. C.R. Scriver, A.L. Beaudet, W.S. Sly, and D. Valle,
eds. McGraw-Hill, New York, pp. 2287–2324.
Lüers, G., Beier, K., Hashimoto, T., Fahimi, H.D., and Völkl, A. (1990)
Biogenesis of peroxisomes: Sequential biosynthesis of the membrane and matrix proteins in the course of hepatic regeneration.
Eur. J. Cell Biol., 52:175–184.
Mannaerts, G.P., and Van Veldhoven, P.P. (1993) Metabolic role of
mammalian peroxisomes. In: Peroxisomes. Biology and Importance
in Toxicology and Medicine. G. Gibson and B. Lake, eds. Taylor &
Francis, London. pp. 19–62.
Mandel, H., Espeel, M., Roels, F., Sofer, N., Luder, A., Iancu, T., Aizin,
A., Berant, M., Wanders, R.J.A., and Schutgens, R.B.H. (1994) A
new type of peroxisomal disorder with variable expression in liver
and fibroblasts. J. Pediatr., 125:549–555.
McNew, J.A., and Goodman, J.M. (1996) The targeting and assembly
of peroxisomal proteins: Some old rules do not apply. Trends
Biochem. Sci., 21:54–58.
Mitaka, T., Norioka, K., and Mochizuki, Y. (1993) Redifferentiation of
proliferated rat hepatocytes cultured in L15 medium supplemented
with EGF and DMSO. In Vitro Cell Dev. Biol., 29A:714–722.
Moser, A., Rasmussen, M., Naidu, S., Watkins, P.A., McGuinness, M.,
Haijra, A.K., Chen, G., Raymond, G., Liu, A., Gordon, D., Gamaas,
K., Walton, D.S., Skjeldal, O.H., Guggenheim, M.A., Jackson, L.G.,
Elias, E.R., and Moser, H.W. (1995) Phenotype of patients with
peroxisomal disorders subdivided into sixteen complementation
groups. J. Pediatr., 127:13–22.
Novikoff, A.B., and Shin W.-Y. (1964) The endoplasmic reticulum in the
Golgi zone and its relations to microbodies, Golgi apparatus and
autophagic vacuoles in rat liver cells. J. Microsc., 3:187–206.
Osumi, T., Tsukamoto, T., Hata, S., Yokota, S., Miura, S., Fujiki, Y.,
Hijikata, M., Miyazawa, S., and Hashimoto, T. (1991) Aminoterminal presequence of the precursor of peroxisomal 3-ketoacylCoA thiolase is a cleavable signal peptide for peroxisomal targeting.
Biochem. Biophys. Res. Commun., 181:947–954.
Pipan, N., and Psenicnik, M. (1975) The development of microperoxisomes in the cells of the proximal tubules of the kidney and
epithelium of small intestine during the embryonic development
and postnatal period. Histochemistry, 44:12–21.
Poll-The, B.T., Saudubray, J.M., Rocchioccioli, F., Scotto, J., Roels, F.,
Boue, J., Ogier, H., Dumez, Y., Wanders, R.J.A., Schutgens, R.B.H.,
Schram, A.W., and Tager, J.M. (1987) Prenatal diagnosis and
confirmation of infantile Refsum’s disease. J. Inher. Metab. Dis.
Suppl. 2, 10:229–232.
Poll-The, B.T., Roels, F., Ogier, H., Scotto, J., Vamecq, J., Schutgens,
R.B.H., Wanders, R.J.A., Van Roermund, C.W.T., Van Wyland,
M.J.A., Schram, A.W., Tager, J.M., and Saudubray, J.M. (1988) A
new peroxisomal disorder with enlarged peroxisomes and a specific
deficiency of acyl-CoA oxidase (Pseudo-neonatal adrenoleukodystrophy). Am. J. Hum. Genet., 42:422–434.
Poulos, A., Christodoulou, J., Chow, C.W., Goldblatt, J., Paton, B.C.,
Orii, T., Suzuki, Y., and Shimozawa, N. (1995) Peroxisomal assembly
defects: Clinical, pathologic and biochemical findings in two patients
in a newly identified complementation group. J. Pediatr., 127:596–
Purdue, P.E., and Lazarow, P.B. (1994) Peroxisomal biogenesis: Multiple pathways of protein import. J. Biol. Chem., 269:30065–30068.
Rapp, S., Saffrich, R., Anton, M., Jäkle, U., Ansorge, W., Gorgas, K.,
and Just, W. (1996) Microtubule-based peroxisome movement. J.
Cell Sci., 109:837–849.
Rhodin, J. (1954) Correlation of Ultrastructural Organization and
Function in Normal and Experimentally Changed Proximal Convoluted Tubule Cells of the Mouse Kidney. Ph.D. thesis. Aktiebolaget
Godvil, Stockholm.
Rigatuso, J.L., Legg, P.G., and Wood, R.I. (1970) Microbody formation
in regenrating rat liver. J. Histochem. Cytochem., 18:893–900.
Roels, F. (1991) Peroxisomes. A Personal Account. VUB Press, Brussels.
Roels, F., and Cornelis, A. (1989) Heterogeneity of catalase staining in
human hepatocellular peroxisomes. J. Histochem. Cytochem., 37:
Roels, F., and Goldfischer, S. (1979) Cytochemistry of human catalase.
The demonstration of hepatic and renal peroxisomes by a high
temperature procedure. J. Histochem. Cytochem., 27:1471–1477.
Roels, F., Pauwels, M., Cornelis, A., Kerckaert, I., Van Der Spek, P.,
Goovaerts, G., Versieck, J., and Goldfischer, S. (1983) Peroxisomes
(microbodies) in human liver. Cytochemical and quantitative studies in 85 biopsies. J. Histochem. Cytochem., 31:235–237.
Roels, F., Cornelis, A., Poll-The, B.T., Aubourg, P., Ogier, H., Scotto, J.,
and Saudubray, J.M. (1986) Hepatic peroxisomes are deficient in
infantile Refsum disease: A cytochemical study of 4 cases. Am. J.
Med. Genet., 25:257–271.
Roels, F., Pauwels, M., Poll-The, B.T., Scotto, J., Ogier, H., Aubourg, P.,
and Saudubray, J.M. (1988) Hepatic peroxisomes in adrenoleukodystrophy and related syndromes. Cytochemical and morphometric
data. Virchows Arch. A Pathol. Anat., 413:275–285.
Roels, F., Espeel, M., Pauwels, M., De Craemer, D., Egberts, H.J.A.,
and Van der Spek, P. (1991a) Different types of peroxisomes in
human duodenal epithelium. Gut, 32:858–865.
Roels, F., Espeel, M., and De Craemer, D. (1991b) Liver pathology and
immunocytochemistry in peroxisomal disorders: A review. J. Inher.
Metab. Dis., 14:853–875.
Roels, F., Espeel, M., Poggi, F., Mandel, H., Van Maldergem, L., and
Saudubray, J.M. (1993a) Human liver pathology in peroxisomal
diseases: A review including novel data. Biochimie, 75:281–292.
Roels, F., Espeel, M., Lissens, W., Schutgens, R.B.H., Wanders, R.J.A.,
Pauwels, M., Foulon, W., and Liebaers, I. (1993b) Fast prenatal
diagnosis of Zellweger cerebro-hepato-renal syndrome by microscopy of first trimester chorionic villi. In: Abstracts of the 31st SSIEM
Annual Symposium. Kluwer Academic Publishers, UK.
Roels, F., Tytgat, T., Beken, S., Giros, M., Espeel, M., De Prest, B.,
Kerckaert, I., Pampols, T., and Rogiers, V. (1996a) Peroxisome
mosaics in the liver of patients and the regulation of peroxisome
expression in rat hepatocyte culture. Ann. N.Y. Acad. Sci., 804:502–
Roels, F., D’Herde, K., Vamecq, J., Kerckaert, I., Dacremont, G., and
Espeel, M. (1996b) Peroxisomes in oocytes. Acta Histochem. Cytochem. Suppl., 29:222–223.
Rouiller, C., and Bernhard, W. (1956) ‘‘Microbodies’’ and the problem of
mitochondrial regeneration in rat liver cells. J. Biophys. Biochem.
Cytol., 2:355–358.
Santos, M.J., Imanaka, T., Shio, H., Small, G.M., and Lazarow, P.B.
(1988a) Peroxisomal membrane ghosts in Zellweger syndrome—
Aberrant organelle assembly. Science, 239:1536–1538.
Santos, M.J., Imanaka, T., Shio, H., and Lazarow, P.B. (1988b)
Peroxisomal integral membrane proteins in control and zellweger
fibroblasts. J. Biol. Chem., 263:10502–10509.
Schrader, M., Burkhardt, J.K., Baumgart, E., Lüers, G., Spring, H.,
Völkl, A., and Fahimi, H.D. (1996) Interaction of microtubules with
peroxisomes. Tubular and spherical peroxisomes in HepG2 cells and
their alterations induced by microtubule-active drugs. Eur. J. Cell
Biol., 69:24–35.
Schutgens, R.B.H., Heymans, H.S.A., Wanders, R.J.A., Van Den
Bosch, H., and Tager, J.M. (1986) Peroxisomal disorders: A newly
recognized group of genetic diseases. Eur. J. Pediatr., 144:430–440.
Shimozawa, N., Tsukamoto, T., Suzuki, Y., Orii, T., Shirayoshi, Y.,
Mori, T., and Fujiki, Y. (1992a) A human gene responsible for
Zellweger syndrome that affects peroxisome assembly. Science,
Shimozawa, N., Tsukamoto, T., Suzuki, Y., Orii, T., and Fujiki, Y.
(1992b) Animal cell mutants represent two complementation groups
of peroxisome-defective Zellweger syndrome. J. Clin. Invest., 90:
Shimozawa, N., Suzuki, Y., Orii, T., Moser, A., Moser, H.W., and
Wanders, R.J.A. (1993) Standardization of complementation grouping of peroxisome-deficient disorders and the second Zellweger
patient with peroxisomal assembly factor-1-(PAF-1) defect. Am. J.
Hum. Genet., 52:843–844.
Shio, H., and Lazarow, P.B. (1981) Relationships between peroxisomes
and endoplasmic reticulum investigated by combined catalase and
glucose-6-phosphate cytochemistry. J. Histochem. Cytochem., 29:
Stefanini, S., Farrace, M.G., and Ceru Argento, M.P. (1985) Differentiation of liver peroxisomes in the foetal and newborn rat. Cytochemistry of catalase and D-amino acid oxidase. J. Embryol. Exp. Morphol.,
Stefanini, S., Serafini, B., Cimini, A., and Sartori, C. (1994) Differentiation of kidney cortex peroxisomes in fetal and newborn rats. Biol.
Cell 80:185–193.
Sternlieb, I., and Quintana, N. (1977) The peroxisomes of human
hepatocytes. Lab. Invest., 36:140–149.
Subramani, S. (1993) Import of proteins into peroxisomes and biogenesis of the organelle. Annu. Rev. Cell Biol., 9:447–478.
Swinkels, B.W., Gould, S.J., Bodnar, A.G., Rachubinski, R.A., and
Subramani, S. (1991) A novel cleavable peroxisomal targeting signal
at the amino-terminus of the rat 3-ketoacylCoA thiolase. EMBO J.,
Tager, J.M., Ten Harmsen Van De Beek, W.A., Wanders, R.J.A.,
Hashimoto, T., Heymans, H.S.A., Van Den Bosch, H., Schutgens,
R.B.H., and Schram, A.W. (1985) Peroxisomal b-oxidation enzyme
proteins in the Zellweger syndrome. Biochem. Biophys. Res. Commun., 126:1269–1275.
Thieringer, R., and Raetz, C.R.H. (1993) Peroxisome-deficient chinese
hamster ovary cells with point mutations in peroxisome assembly
factor-1. J. Biol. Chem., 268:12631–12636.
Tsukada, H., Mochizuki, Y., and Konishi, T. (1968) Morphogenesis and
development of microbodies of hepatocytes of rats during pre- and
postnatal growth. J. Cell Biol., 37:231–243.
Tsukamoto, T., Miura, S., and Fujiki, Y. (1991) Restoration by a 35K
membrane protein of peroxisome assembly in a peroxisome-deficient
mammalian cell mutant. Nature, 350:77–81.
Usuda, N., Yokota, S., Ichikawa, R., Hashimoto, T., and Nagata, T.
(1991a) Immunoelectron microscopic study of a new D-amino acid
oxidase-immunoreactive subcompartment in rat liver peroxisomes.
J. Histochem. Cytochem., 39:95–102.
Usuda, N., Kuwabara, T., Ichikawa, R., Hashimoto, T., and Nagata, T.
(1991b) Immunoelectron microscopic evidence for organ differences
in the composition of peroxisome-specific membrane polypeptides
among three rat organs: liver, kidney, and small intestine. J.
Histochem. Cytochem., 39:1357–1366.
Valle, D., and Gärtner, J. (1993) Penetrating the peroxisome. Nature,
Van den Bosch, H., Schutgens, R.B.H., Wanders, R.J.A., and Tager,
J.M. (1992) Biochemistry of peroxisomes. Annu. Rev. Biochem.,
Van den Munckhof, R.J.M. (1996) In situ heterogeneity of peroxisomal
oxidase activities: An update. Histochem. J., 28:401–429.
Van Maldergem, L., Espeel, M., Roels, F., Petit, C., Dacremont, G.,
Wanders, R.J.A., Verloes, A., and Gillerot, Y. (1991) X-linked reces-
sive chondrodysplasia punctata with XY translocation in a stillborn
fetus. Hum. Genet. 87:661–664.
Völkl, A., Baumgart, E., and Fahimi, H.D. (1988) Localization of urate
oxidase in the crystalline cores of rat liver peroxisomes by immunocytochemistry and immunoblotting. J. Histochem. Cytochem., 36:
Wanders, R.J.A., Kos, M., Roest, B., Meijer, A.J., Schrakamp, G.,
Heymans, H.S.A., Tegelaers, W.H.H., Van Den Bosch, H., Schutgens, R.B.H., and Tager, J.M. (1984) Activity of peroxisomal enzymes and intracellular distribution of catalase in Zellweger syndrome. Biochem. Biophys. Res. Commun. 123:1054–1061.
Wanders, R.J.A., Van Roermund, C.W.T., Westra, R., Schutgens,
R.B.H., Van Der Ende, M.A., Tager, J.M., Monnens, L.A.H., Baadenhuysen, H., Govaerts, L., Przyrembel, H., Wolff, E.D., Blom, W.,
Huijmans, J.G.M., and Van Laerhoven, F.G.M. (1987) Alanine
glyoxylate aminotransferase and the urinary excretion of oxalate
and glycollate in hyperoxaluria type I and the Zellweger syndrome.
Clin. Chim. Acta, 165:311–319.
Wanders, R.J.A., Barth, P.G., Schutgens, R.B.H., and Tager, J.M.
(1993) Peroxisomal disorders. In: Peroxisomes. Biology and Importance in Toxicology and Medicine. G. Gibson and B. Lake, eds. Taylor
& Francis, London. pp. 63–98.
Wanders, R.J.A., Dekker, C., Ofman, R., Schutgens, R.B.H., and
Mooijer, P. (1995) Immunoblot analysis of peroxisomal proteins in
liver and fibroblasts from patients. In: Diagnosis of Peroxisomal
Disorders. A Handbook. F. Roels, S. De Bie, and R.B.H. Schutgens,
eds. Kluwer, Dordrecht, pp. 101–112.
Warren, G., and Wickner, W. (1996) Organelle inheritance. Cell,
Wedel, F.P., and Berger, E.R. (1975) On the quantitative stereomorphology of microbodies in rat hepatocytes. J. Ultrastr. Res.,
Wiemer, E.C., Brul, S., Just, W.W., Van Driel, R., Brouwer-Kelder, E.,
Van Den Berg, M., Weijers, P.J., Schutgens, R.B.H., Van Den Bosch,
H., Schram, A., Wanders, R.J.A., and Tager, J.M. (1989) Presence of
peroxisomal membrane proteins in liver and fibroblasts from patients with the Zellweger syndrome and related disorders: evidence
for the existence of peroxisomal ghosts. Eur. J. Cell Biol., 50:407–
Wiemer, E.A.C., Lüers, G., Faber, K.N., Wenzel, T., Veenhuis, M., and
Subramani, S. (1996) Isolation and characterisation of Pas2p, a
peroxisomal membrane protein essential for peroxisome biogenesis
in the methylotrophic yeast Pichia pastoris. J. Biol. Chem., 271:
Wiemer, E.A.C., Wenzel, T., Deerinck, T.J., Ellisma, M.H., Subramani,
S. (1997) Visualization of the peroxisomal compartment in living
mammalian cells: Dynamic behavior and association with microtubules. J. Cell Biol., 136:71–80.
Yajima, S., Suzuki, Y., Shimozowa, N., Yamaguchi, S., Orii, T., Fujiki,
Y., Osumi, T., Hashimoto, T., and Moser, H.W. (1992) Complementation study of peroxisome-deficient disorders by immunofluorescence
staining and characterization of fused cells. Hum. Genet., 88:491–
Yamamoto, K., and Fahimi, H.D. (1987) Three-dimensional reconstruction of a peroxisomal reticulum in regenerating rat liver: Evidence of
interconnections between heterogenous segments. J. Cell Biol.,
Yokota, S., and Fahimi, H.D. (1980) Immunocytochemical localization
of catalase in rat liver. J. Histochem. Cytochem., 29:805–812.
Yokota, S., Völkl, A., Hashimoto, T., and Fahimi, H.D. (1987) Immunoelectron microscopy of peroxisomal enzymles; their substructural
association and compartmentalization in rat kidney peroxisomes.
In: Peroxisomes in Biology and Medicine. H.D. Fahimi and H. Sies,
eds. Springer Verlag, Berlin, pp. 115–127.
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