Vol. 9, No 1, 2002 pp. 64 - 65
UC 616.61-002.828
EXPERIMENTAL MYCOTOXIC NEPHROPATHIES
AND BALKAN ENDEMIC NEPHROPATHY
Peter G. Mantle
Department of Environmental Science and Technology, Imperial College
of Science, Technology and Medicine, London, UK
Nearly half a century after its first recognition in Bulgaria, the cause
of Balkan endemic nephropathy (BEN) and its associated higher incidence
of urinary tract tumours remains obscure. Even 20 years ago some current
ideas concerning viruses, bacterial pathogens, heavy metals from geological
sources, and nephrotoxic medicinal plants such as Aristolochia, were discounted
in a comprehensive review (1) as not fitting the mosaic of disease incidence.
However, as part of the present new review of BEN it may be helpful to
reflect on the topic of nephrotoxic mycotoxins, which has long been a popular
source of putative aetiological agents. The present author does so, following
20 years of active research interest both in BEN 'in the field' in Croatia
and Bulgaria, and in experimental science of some nephrotoxic mycotoxins
of possible relevance.
Ochratoxin A has been the main focus of international research concerning
BEN because of its role as a cause of the Danish porcine nephropathy that
became of economic concern in the bacon industry in the mid 20th century.
In that case, the fungus (now known as Penicillium verrucosum, naturally
producing the toxin in northern European latitudes) was clearly demonstrated
as a common spoilage microorganism of poorly-stored barley. Its occurrence
was correlated with the presence of its key metabolite ochratoxin A and
experiments in pigs with the pure toxin and experimentally spoiled grain
confirmed that ochratoxin A was clinically toxic at concentrations commonly
occurring in years of poor harvest weather. Thus a satisfactory principal
cause and effect correlation was proved for ochratoxin A concerning that
porcine nephropathy.
It became reasonable, therefore to suggest extrapolation from Danish
mycotoxic porcine nephropathy to the mysterious and idiopathic BEN in the
early 1970s (2). However, it may have slanted some investigations of the
human disease away from a search for its cause and towards an attempt to
show that ochratoxin A also fits as a cause of BEN. Further, the claim
at that time of similar renal histopathology between porcine nephropathy
and BEN has since received rather little direct challenge. Of course there
are features in common. But it is usually ignored that nephropathy in growing
pigs is a renal hypertrophy, often quite marked and correlated with impaired
live weight gain, while BEN is a very silent renal atrophy with accompanying
fibrosis in peasant farmers who do not seem to have a history of chronic
poor health. Interestingly, there does not seem to be nephropathy in the
pigs that the BEN subjects almost all keep and which are likely to eat
a diet of inferior quality.
The situation is the more complicated by the frequent cooccurrence
of urinary tract tumours with the renal atrophy of BEN, and the clear role
of ochratoxin A in causing renal tumours in rats. Ochratoxin A thus seems
to have an array of toxic attributes ideally suited to connection with
many aspects of the Balkan disease syndrome. Therefore it has been important
to study the toxicology of ochratoxin A and other nephrotoxic metabolites
of food/feed-spoilage moulds in experimental animals with respect to both
renal tubular morbidity and tumourigenesis.
Of course, ochratoxin A is a very toxic substance. Perhaps it is too
toxic and broady toxic to cause the specific and silent BEN. Direct renal
insults as tubule epithelial cell loss appear to be mainly necrotic. Claims
of evidence for loss by apoptosis in vivo seem weak since they are not
based on specific staining. Indeed, apoptosis is not readily compatible
with carcinogenicity. Further, our recent studies have shown that administration
to rats by oral gavage causes much greater damage than when ingested via
feed (3). Some experimentation may therefore have given exaggerated effects.
Similarly, a recent 1-year experiment in pigs fed continuously on a diet
containing 800ppb ochratoxin A revealed a slight effect on growth and a
rather mild renal histopathology. It was judged that the animals would
not have been recognised by routine meat inspection at slaughter as subjects
with ochratoxicosis (4). In contrast a similar dose given to pigs over
a shorter time, but augmented with the common mycotoxin penicillic acid
in the diet, showed much more severe effects (5). Experimentation that
can be meaningful in terms of risk assessment for humans and applicability
to the BEN/ urinary tract tu-mour complex should therefore include diets
which have a heterogeneous mycotoxin composition relevant to the range
of natural circumstances. It is also questioned whether experimentation
in rats dosed with ochratoxin A on an increasing body-weight basis during
life also exaggerates response relative to normal dietary intake and body
conformation in humans, especially concerning effects on excretory organs.
Another experimental mycotoxic nephropathy is that caused in rats by
crude water extract of the food-spoil-age mould, now known as Penicillium
polonicum, which is common in the Balkans. This causes extensive apoptosis
in proximal tubule epithelia (6). It is also potently mitogenic, resulting
in huge and persistent karyomegaly (7). The clear demonstration of tissue-specific
rat renal tubule apoptosis by a common mould stimulated the idea that silent
apoptosis is an attractive mechanism by which BEN might arise and progress.
P. polonicum nephrotoxin might only be a model for a similar effect from
an otherwise unexpected source (6). Very early stages in BEN are difficult
to specify. The author is not aware of any confident description of such;
so-called early stages in a kidney with atrophy may actually be early end-stage
in the whole 'lifetime' aetiology of BEN. Strangely, the same mould has
been shown to alter the debrisoquine-metabolising capability in rats (8),
so as to give a scientific base for questioning the fidelity of a statistical
finding of nephropathy patients tending also to be faster metabolisers
of debrisoquine.
Current focus in the author's laboratory is collaborative research
with others in Western Europe towards fundamental understanding of aspects
of ochratoxin toxicity. This involves broad study of rat gene expression
changes using 1000 gene DNA microarrays. We would be pleased to include
analogous study of small amounts of suitable ultrafresh renal tissue from
BEN subjects that may become available. This might help in comparing the
human disease with some known effects of ochratoxin A to see the extent
of fit. It could also provide a valuable knowledge base concerning the
gene expression status of human tissue captured during the development
of BEN.
Always it is necessary to remember that the incidence of BEN is highly
mosaic both within and between endemic villages. Any putative cause must
fit this, as also the strong familial character of the disease. Also, aetiological
study should strive for rigorous proof analogous to that prescribed in
the late 19th century by Robert Koch for establishing the cause of infectious
disease. In the Balkans ochratoxin A has been found in cereal commodities
in small amounts from time to time, much as it has in some other regions
of the world, and sometimes much emphasis is placed on the occasional higher
than usual concentration. However, unlike in northern European latitudes,
P. verrucosum seems not to be a toxigenic mould. Aspergillus ochraceus,
the source of the original discovery of ochratoxin A in South Africa in
the early 1960s, is not common and rather few isolates have been found
to produce significant amounts of the toxin. Some other ochratoxigenic
Penicillium spp. have been found, but generally the source of ochratoxin
A in human food in the Balkans is obscure (9). This illustrates the neglect
of rigorous fo-rensic study of BEN for far too long and does not compel
confidence in convicting nephrotoxic mycotoxins of a causative role in
BEN beyond reasonable doubt. However, even if, for whatever reason, the
incidence of BEN can be shown, fortunately, to be in decline it is still
important to find the cause as a model for addressing other apparently
intractable aetiological questions in human medicine.
