1. Introduction and Problem at Hand
Glomerulonephritis is a collective word to describe inflammation
of the glomerulus. The glomerulus is an arterial net, surrounded
by an epithelial capsule, known as the Bowman capsule. The central
unit of the glomerulus is the glomerate membrane with its typical
three-tiered layers, consisting of endotheliod capillary cells,
basal cells and poduroid epithelia.
The cause of the glomerulonephritis often remains unknown.
However, reactions within the immune system do play a major role
in the development of the inflammation. The filter system of
the glomerulus is designed to regulate the internal environment
of tissues. Due to the large number of glomeruli and their capacity
to conserve energy, a large reserve capacity exists which lets
the kidneys function normally even after the disease has progressed.
Hence, azoturia (excess of nitrogenous matter in the urine),
which is an indicator of a functional breakdown , does not occur
until three quarters of the glomeruli have ceased to function.
Consequently, it is essential that the veterinarian be able to
diagnose
glomerulonephritis before the reserve is exhausted.
In the dog, glomerulonephritis is characterized by persistent
proteinuria in unspecific urine sedimentation tests. Unfortunately,
it is only after the disease has reached advanced stages and tissue
changes have occurred that the dog's owner will see symptoms.
Generally, only these symptoms will prompt the owner to seek
help from a veterinarian. Unfortunately, at this point in time,
the disease has, in most cases, caused irreversible, chronic
and progres-sive kidney insufficiency of the highest order. Therefore,
treatment is of necessity limited to treating the symptoms.
At the Veterinary Hospital, University of Zurich, faculty and
clinicians made a subjective observation in recent years that
the occurrence of glomerulonephritis in Bernese Mountain Dogs
seemed to have increased. Therefore, it was the object of this
study to
determine, based on past and future examinations, whether in
fact an increase in glomerulonephritis in Bernese Mountain Dogs
has occurred. The basis of this study consists of patient records
from the Small Animal Clinic of the Veterinary Hospital and of
the Institute of Veterinay Pathology, both of which are part of
the University of Zurich. In investigating the cause of the disease,
particular emphasis was placed on the possibility of genetic predisposition
in Bernese Mountain Dogs. Also, improved non-invasive methods
for early diagnosis were sought.
2. Filtration Characteristics of the Glomeruli
The glomerulus filters particles based on molecular weight.
For example, the endothelium prevents blood cells from passing,
the Basal membrane retains particles of 200k Dalton and above
and the poduridae normally retain particles of 65k Dalton plus.
(Note: Albumin weighs 69k Dalton) The Basal membrane and the
endothelium are rich in sialinic acid containing proteoglycogen
and glycoproteins, therefore they behave like porous membranes
with negative wall loads. Bohrer et al, who researched the filtration
characteristics of anionic, neutral and cationic dextra molecules
in the glomeruli, were able to show that
polycatonic molecules pass much easier through the filter membranes
than neutral or polyanionic molecules of same size. Serum albumin
has a physiologic pH of 7.4 and therefore is classified as polyanion.
Unlike neutral and cationic molecules of same size they are retained
in the plasma and only traces of them pass through the filter
system.
3. Development of Glomerulonephritis
The following mechanisms may lead to inflammatory but non-infectious
impairment of the glomeruli. It is possible that circulating
soluable antigen antibody complexes get trapped and settle in
the filter system of the glomerular capillaries. Such antigen
antibody complexes form typically as a result of antigen excesses.
The Basal membrane has a high
concentration of organ specific antigen. It is possible that
as a result of an immune system reaction kidney specific antibodies
are released and targeted at the Basal membrane. This results
in an in-situ immune complex formation. Another possible in-situ
immune complex formation may result when antibodies containing
cationic
proteins attach themselves to the anionic Basal membrane.
The formation of resultant lesions is influenced not only by the
location within the glomeruli where antigen antibody deposits
occur, but also by the inflammatory mediators involved. Because
of loss in the negative wall load, the deposit of antibodies alone
can contribute to increased permeability of the glomeruli. Complement
fragments (C3a, C5a) can, after direct or indirect activation,
attract neutrophilic granulocytes and especially macrophages.
Release of lysosomatic enzymes and oxygen radicals results.
Even without
complement activation, it is possible that macrophages attach
themselves to Fc particles of immune globulin, thereby starting
the inflammatory process. Due to histamine degranula-tion, the
permeability of capillaries is increased. As intercapillary currents
change, platelets can accumulate in the impaired endothelium.
As a result, vasoactive amines may be released as well as cationic
proteins. These in turn may stimulate an immune complex formation
and
act as chemotaxis. It is also possible that the mesangial cells
in the glomeruli themselves are participating in the release of
inflammation mediators.
4. Functional Consequences of Inflammation in the Glomeruli
The filtration capacity is seriously disturbed as a result
of load shifts in the endothelium and the Basal membrane. The
increased glomerular permeability allows primarily the passing
of plasma proteins, especially albumin. Proteinuria results.
A prolonged loss of protein leads to hypoalbuminemia, to reduction
in oncotic pressure n the blood and the formation of edema. Hypoalbuminosis
causes a decrease in plasma volume which in turn reduces renal
plasma flow and finally results in the activation of the renin-angiotonic
system. This causes water and natrium retention. Hence, the
possibility of edema formation and effusion is increased. The
hypoalbuminosis activates the synthesis of lipoproteins in an
effort to maintain the oncotic pressure within physiological limits.
As a result hyperlipoproteinosis occurs. Additional side effects
associated with the loss of proteins are losses of highly molecular
carrier proteins for hormones, vitamins and trace elements such
as Transferrin. An increased susceptibility for infections is
also observed. The loss of the heparin cofactor Antithrombin
III, a protein molecule of 65k Dalton and a strong inhibitor of
peptidose thrombin of the coagulation system, increases the risk
of thromboembolism. This tendency is further intensified by an
increased protein synthesis through the liver, inclusive of various
highly molecular coagulation factors.
5. Etiology of Glomerulonephritis
The follwing is a summary of causative factors for glomerulonephritis.
Infections: contagious canine hepatitis (Wright et al., 1974;
Morrison et al., 1975); Ehrlichiasis (Troy et al., 1980); Brucellosis
(Green and George, 1984); Leischmaniasis (Chapman and Hanson,
1984); bacterial endocardities (Highman et al., 1959); pyometra
(Asheim, 1965); dirofilariasis (Simpson et al., 1974; Casey and
Splitter, 1975; Aikawa et al., 1981); borreliasis (Grauer et al.,
1988).Organ and autoimmune diseases: pancreatitis (Lewis, 1976;
Slausen and Lewis, 1979); systemic lupos erythematosus (Lewis,
1972; Lewis et al., 1973; Osborne et al., 1973).Neoplasia: mastocytoma
(Hattendorf and Nielson, 1968); lymphosarcoma (Slauson and Lewis,
1979).Genetic diseases: collagen malformation in Samoyeds and
Doberman Pinschers (Jansen et al., 1987; Thorner et al., 1987)
and Picut and Lewis, 1986), respectively.
Idiopathic disease: in many cases glomerulonephritis could not
be attributed to any of the above or other causes (Jergens, 1987).
6. Therapy for Glomerulonephritis
First the therapy should be directed at a possible primary
illness which may have caused the glomerulonephritis, such as
pyometra, for example. Since patients are often brought in only
after kidney insufficiency has become chronic, therapy is limited
to treating uremic symptoms and to delay progression of the illness.
It is possible to influence the renal hemodynamic with limited
intake of protein. When large amounts of protein are consumed,
the renal flow and hence the glomerular filtration ratio is increased.
By limiting
protein intake, this process can be slowed down. The need for
protein in uremic dogs is not known. Polzin and Osborn recommend,
therefore an intake of 2.0 to 2.2 grams per kilo of body weight
per day of a biologically high-grade protein. The protein loss
in the urine must be added to this.
Secondary hyperparathyroidism and the lack of vitamin D hormones
disturb the calcium-phosphorus metabolism. Hence calcium deposits
may occur in the kidney parenchyma and lead to hyperphosphatemia.
Should the hyperphosphotemia remain despite diet protein and
phosphate restrictions, the oral administration of phosphorus
binders must be
considered. The reduction of phosphorus levels is aimed at the
following: avoidance of calcium associated cell damage, the reduction
of cellular energy expenditure and the minimization of immunologic
response reactions. The composition of lipids provided in the
diet seems to influence blood pressure, platelet aggregation,
blood viscosity, the immune system and the fibrinolitic activity.
Certain lipids appear to facilitate the damage to the glomerular
Basal membrane and mesangial structures. The use of certain drugs
may be
indicated to control hypertension, and thromboembolic complications.
7. Analysis of Pedigrees
Between 1983 - 1987 of 19 BMDs diagnosed with glomerulonephritis
only six pedigreees could be obtained. Between 1988 and 1989
of 52 BMDs with no kidney problems, 12 pedigrees could be obtained.
In studying the pedigrees of the BMDs with kidney disease it
was noted that certain stud dogs where listed in all six pedigrees.
The relationship coefficient established by Wright was employed
in factoring the relationship between descendants of stud dogs
A,B and C. For sire A the coefficient was 0.119, sire B 0.15
and sire C 0.229. For the healthy BMDs the coefficients were
0.03, 0.059 and 0.106 for sires A, B and C, respectively. In
the ancestry of both the healthy and the sick BMDs, sires A,B
and C produced the majority of stud dogs in the composite pedigree.
The stud dogs in this line were represented in the ancestry of
healthy dogs by 83.3% and by 100% in the ancestry of sick dogs.
8. Sex and Age Predispositions
There appears to be no difference in predisposition for glomerulonephritis
between males and females. The average age of BMD's with glomerulonephritis
in test groups I and II was 5.25 years. It is important to note
that dogs with high degrees of glomerulonephritis were diagnosed
at the latest in their 7th year of life. The average age of BMDs
diagnosed with glomerulopathy in test group II was 4.5 years,
with a male-female ratio of 1:1. Patients
suffering from kidneyamyloidosis had a higher average age, namely
7.5 years and the ratio between males and females was 0:4, which
indicates the possibility that bitches are more predisposed to
amyloidosis. Again, the number of dogs involved was too small
to prove this theory. The average age for BMDs in both groups
which were diagnosed with
interstitial nephritis was 5.52 years. The ratio between sexes
was 12:5, which indicates that males may be more predisposed to
this illness. A possible explanation for this may be the fact
that dogs always retain some urine for "marking". This
encourages the deposit of pathogenic bacteria which may then enter
the kidneys. The average age of dogs diagnosed with glomerulopathy
in test group III was 3.83 years and the ratio between sexes was
1:1. In group III the average age of BMDs with interstitial nephritis
was 5.92 years. As in groups I and II, males appear to be more
susceptible to this illness than
females. The ratio between sexes was 9:4.
9. Discussion Regarding Possible Etiology for Glomerulonephritis
in BMDs
Glomerulonephritis in the dog can evolve as a secondary illness,
in other words be part of a multi-system occurrence or it can
be a primary (idiopathic) disease. Murray and Wright (1974) found
that 70% of the dogs they dissected and which represented a multitude
of
breeds, suffered from serious extra renal illnesses in addition
to glomerulonephritis. In 40% of these dogs they found neoplasia
and in 16% they found chronic inflammatory processes. Center
and Smith (1987) found extra renal and chronic illnesses in 88%
of their dogs, representing many breeds. Clucocorticoid was found
in 34%, chronic skin infections in 27%, neoplasia in 17%, rheumatoid
arthritis in 12% and systemic lupus erythematosus in 7%. In comparison,
in the BMD group I, a total of 47% of the dogs suffered
from kidney disease only. Chronic inflammatory illnesses were
found in 35% and neoplasia and rheumatoid arthritis in 12%.
This shows that among BMD's a tendency for idiopathic glomerulonephritis
exists. The only other breeds susceptible to glomerulonephritis
are the Samoyeds and the Doberman Pinschers. However, in these
breeds the problem is a collagen misformation in the
glomeruli caused by a Basal membrane defect. In the Samoyeds,
Thorner et al (1987) were able to identify an x-chromosomal dominant
inheritance factor with manifestation of clinical symptoms during
the first year of a descendants life. In BMDs such a simple correlation
was not possible because (1) BMDs reached a lifespan of up to
seven years despite having glomerulonephritis and (2) only a small
number of pedigrees were available for analysis. However, the
limited sample does indicate that the relationship coefficient
among descendants of a certain stud dog line was higher by 3 for
dogs having been diagnosed with the disease. The genetic (probably
polygene) disposition for glomerulonephritis is a possibility
although additional research is required to prove this point.
Because the illness is latent for many years, research through
breeding seems impractical. Therefore, gene
technology should be utilized to find the answer.
The immunohistologic examinations served to identify immune complex
deposits in the glomeruli. Immune complex deposits were found
not only in dogs with glomerulonephritis but also in those diagnosed
with interstitial nephritis and those with kidney amyloidosis.
It is possible, however, that in these cases it was only a transient
deposit of IgM. Medium to high deposits, however, were only found
in dogs with glomerulonephritis. Especially interesting were
two cases of glomerulonephritis where absolutely no immune complex
deposits could be found. One was a patient with a medium grade
of mesangio-proliferous and the other with chronic sclerose glomerulonephritis.
These cases may serve to illustrate the development of histologic
changes in absence of the trigger noxa and point to the self supporting
character of the illness.
10. Final Observations
With this study we are able to illustrate that kidney disease
in general and glomeruloneph-ritis in particular is found more
often in Bernese Mountain Dogs than in the general population.
To diagnose the disease, lab tests have proven to be effective.
Routine tests
carried out in many labs (serum and urine analysis) serve as
the basis for further testing. The protein/creatinine ratio test
was successfully utilized to diagnose loss of protein through
urine.
Additional tests employed to narrow down the diagnosis were
the creatinine-clearance, radioisotope-clearance and kidney biopsy.
Although the two clearance tests both exhibited the same amount
of sensitivity to glomerulopathy, the radioisotope-clearance was
yielding
more detailed information into the disease process in the glomeruli.
A large advantage over the standard clearance methods was its
accuracy. In all the cases where glomerulopathy was suspected,
additional tests only served to confirm what routine lab tests
had indicated. Which means that routine urine tests combined
with protein/creatinine ratios suffice to establish a diagnosis
of glomerulopathy. Unfortunately, kidney function tests can not
differentiate between kidney amyloidosis and glomerulonephritis.
The only way to differentiate between the two diseases is by
biopsy. Understandably, it is difficult to receive permission
from the dog owner to perform a biopsy in the early stages of
the disease and
before serious symptoms are apparent.
Since it was not possible to perform biopsies on all patients,
it was not possible to assess tissue damage in the glomeruli.
It is our conclusion that the protein/creatinine ratio is the
best method for early detection of glomerulopathy and this test
is available to every
verterinarian. The tissue analysis and a prognostic assessment
however, is dependent on a kidney biopsy. The importance of
early detection is demonstrated in the three patients who after
two years of dietary therapy showed no tendency towards the disease.
It is speculated that the predisposition to glomerulonephritis
is polygenetic and is passed on to offspring. Definite proof
of this point is difficult to come by in light of current breeding
practices (breeding without pedigree). An answer to this complex
question could
probably be obtained through gene technology.
11. Summary
At the Veterinary Clinic of the University of Zurich a study
based on past and prospective examinations was conducted to establish
frequency, cause and diagnostic possibilities of glomerulonephritis
in Bernese Mountain Dogs. The study based on past cases included
the
years 1983 to 1987. The case histories of 261 BMDs were included.
In 46 (17.62%) of the 261 BMDs, kidney disease was diagnosed
and in 19 (7.2%) the diagnosis of glomerulonephritis could be
confirmed through histopathology. In 6 (2.29%) BMDs, damage to
the glomeruli was established through clinical examination although
tissue was not available to specify the cause for the damage.
The study based on prospective examinations included 72 Bernese Mountain Dogs which were seen between January 15, 1988 and March 15, 1989. Based on clinical examinations, lab results, clearance-examinations and tissue evaluation, it was possible to diagnose kidney disease in 20 (26.38%) BMDs. In 6 (8.33%) dogs, the clinical diagnosis of glomerulopathy was established. In two of these cases, tissue evaluation made it was possible to definitely diagnose glomerulonephritis.
There is a definite trend among Bernese Mountain Dogs to suffer
from glomerulonephritis. A genetic predisposition is possible.
Early detection of development of proteinuria is possible through
a simple test measuring the protein/creatinine ratio. The radioisotope-clearance
by means of 3H-Inulin and 14C-Tetraethylammoniumbromid leads to
a quick and accurate diagnosis of glomerular function.