|
Journal of Applied Sciences and Environmental Management
World Bank assisted National Agricultural Research Project (NARP) - University of Port Harcourt
ISSN: 1119-8362
Vol. 8, Num. 1, 2004, pp. 29-32
|
Journal of Applied Sciences & Environmental Management,
Vol. 8, No. 1, June, 2004, pp. 29-32
Effects of Oral Cadmium Exposure on Renal
Glomerular and Tubular Functions in the Rat
1*ASAGBA, S O; 2OBI,
F O
1Department of Biochemistry, Faculty
of Science, Delta State University, Abraka, Nigeria
2Department of Biochemistry, Faculty of Science, University
of Benin, Benin City, Nigeria
ABSTRACT:
The
effects of orally consumed cadmium on the functions of the kidney have been
investigated in rats based on the reported level of the toxicant in Warri
River. Relative
to the corresponding controls
there were significant (P < 0.05) increases in the amount of cadmium in the
kidneys of rats in all the test groups. Biochemical analysis revealed significant
(P < 0.05) changes in plasma creatinine after 2 months (control 1.20 ± 0.20 ´ 10-2;
test 0.92 ± 0.26 ´ 10-2 mg/ml)
and
glucose after 1-month (control 91.67 ± 3.39; test 102.75 ± 5.99 mg/dL) exposure. Twenty-four
hours urine volume
were significantly decreased (P < 0.05) in rats exposed to cadmium for 1 and
2 months. Also in the Cd-exposed rats urine protein was significantly elevated
in those exposed for 2 and 3 months but their urine glucose was demonstratively
elevated only in those exposed for 2 months (control 33.00 ± 7.80; test 43.00 ± 9.80
mg/dL). Urine creatinine was not significantly
altered in any of the test groups. Consistently there were significant (P < 0.05)
decreases in total ATPase and Mg2+ - ATPase activities at the end
of the 2 and 3 months exposure when compared to the controls @JASEM
The kidneys and liver are among
the major target organs of cadmium (Cd) accumulation and intoxication (WHO
report, 1992). The presence of cadmium in all links of the food chain, poor
excretability, propensity for bioaccumulation and persistence make it a source
of considerable
risk to human health (Cabrera et al., 1998) vis-à-vis the functional integrity
of the two organs indicated above and others. Cd-induced injury to both organs
has been attributed to its ability to enhance free radical
formation in vivo (Gupta et al., 1991; Bagchi et al.,
1996). Some characteristic biochemical features associated with Cd-induced kidney
damage are urinary excretion of protein, N-acetyl b D glucosaminidase,
glucose and amino
acids. Others are marked increase of plasma creatinine or blood urea nitrogen
levels (Horiguchi et al., 1996) as well as altered plasma glucose level
(Chapatwala et al., 1980). Consumption of contaminated water is the major
way by which humans are exposed to Cd (WHO report, 1992) and the maximum allowable
level in drinking-water is 0.005 mg/dL (WHO report, 1984). In many countries
including Nigeria contamination of rivers and adjoining seas by Cd and other
heavy metals occurs in a number of ways among which is the discharge of waste
liquid matter from industrial sites. The average level of Cd in Warri River
waters between 1986 and 1991 was 0.3mg/L (Egborge, 1994). This is 60 fold
above the maximum allowable level in drinking water. Naturally this is
worrisome. Therefore this paper focuses on the toxicity of Cd in rats, which
can be related to the Warri River level of the toxicant within the period in
question.
MATERIALS AND METHODS
Animals: Thirty-six adult
albino rats (Wistar strain), 180 190g, bred in the Animal Unit of the School
of Pharmacy, University of Benin were used for this
study.
Chemicals and reagents: Bovine
serum albumin, cadmium sulphate (3CdSO4.8H2O), chloroform
and sodium hydroxide were purchased from May and Baker, Dagenham,
England. Adenosine triphosphate (disodium salt) was purchased from Sigma Chemical
Company, U.S.A. Glucose oxidase kit was obtained from QCA (Spain).
Treatment of animals: The
rats were divided into six experimental groups of six rats each and housed
in standard rat cages. From these, three subgroups made up of two cages of
rat each were produced. One set of rats in a subgroup served as the control
while the other served as the test. The control groups were provided with
distilled water as drinking-water while the test groups were provided with
aqueous solution of CdSO4 containing the equivalent of 0.30mg Cd/L
for one, two and three months respectively. All rats were allowed free access
to chow (BFFM, Ewu, Nigeria).
Animal sacrifice and tissue
preparation: After the specified period of exposure rats in each subgroup
were transferred to metabolic cages equipped with accessory for collecting
urine. The 24 hours urine samples were collected after which each rat was
anaesthetized with chloroform. While under anaesthesia blood samples were
obtained via heart puncture and transferred to heparinized tubes standing
on ice. The kidneys were excised. Plasma was obtained by centrifugation
at 3000 rpm for 10 minutes. The kidneys were homogenized as described by
Adam Vizi and Seregi (1982).
Biochemical assays: The activities of ATPases in the kidney homogenates
were assayed as described by Adam Vizi and Seregi (1982). Protein content
of the homogenate and urine was measured by the Lowry method as described previously
(Obi et al., 1998). Plasma and urine glucose estimation was based on
the glucose oxidase assay procedure described in the
QCA kit instruction leaflet. Plasma and urine creatinine levels were determined
by the Jaffe reaction in which a coloured product is formed from creatinine and
picric acid in alkaline solution.
Cadmium analysis: Kidney
cadmium content was estimated with atomic absorption spectrophotometer (Varian
AA 1475) after wet digestion of the
tissues. For the digestion, 20ml HNO3 HCLO4 mixture
(4:1) was introduced into a beaker containing 1g of a given kidney sample followed
by heating at 100ºC until the sample was completely dissolved. Each digest was
thereafter diluted to 100ml with distilled deionized water.
Statistical analysis: The
data are presented as means ± SD. The mean values of the control and test
groups were compared using Students t-test. The significant level was set
at P < 0.05.
RESULTS AND
DISCUSSION
In this study we provided water
and a solution of CdSO4 (º0.30mg Cd/L) ad
libitum to rats for
oral consumption for 1, 2 and 3 months. At the end of each period of exposure
the state of various plasma and urine parameters normally used as indicators
of the proper functioning of the kidney were assessed. Changes in rat kidney
Cd
load caused by this treatment are presented in table 1.
Table 1. Cadmium concentration
in Kidney and Cd-exposed rats
(mean ± sd); n = 6
Duration of exposure (month)
|
Concentration of Cd in kidney (μg x 10 3)
|
|
Control (- Cd)
|
Test (+ Cd)
|
1
|
2.1±0.07
|
6.70±0.20*
|
2
|
6.80±0.60
|
28.00±5.00*
|
3
|
11.00±6.00
|
56.00±8.21*
|
*Values statistically
significant from control (P < 0.05)
The data in table 2 show the results
of the analysis of plasma parameters while that in table 3 represent the values
for urine
parameters. Table 4 shows the results of total ATPase, Mg2+ ATPase
and Na+ K+ ATPase assay. The results presented in table
1 for kidney Cd load reveals that the control rats were not Cd-free. At the
end of the first month of exposure kidneys from the control rats had 0.021 mg
Cd/g. By the end of 2 and 3 months the kidney load has increased to 0.068 and
0.11 mg/g respectively. The second and third month
values represent an increase of 223.8 and 423.8 percent over and above the 1
month
value. The presence of Cd in the kidney of the control rats and the progressive
manner in which it increased are indications that their water and/or feed were
tainted with Cd. This is not surprising in view of the wide distribution of
Cd in the general environment today. It is conceivable though that the result
of our investigation is likely to be affected by the presence of Cd in the kidney
of control rats. While this is not unlikely it is pertinent to note that reproducible
results were obtained by Horiguchi et al.
(1996) who had the same problem. In their study they administered supposedly
Cd-free saline solution to control rats, subcutaneously, once a week and detected
0.041 ± 0.016 and 0.065 ± 0.009 mg Cd/g kidney at
the end of the 6 and 9 months
exposure period respectively. Evidently the sodium chloride and/or water (the
basic components of the saline) and/or the feed on which the control rats were
maintained must have been
tainted with Cd. Again our test rats were provided with CdSO4 solution
prepared with the same distilled water that was provided as drinking water for
the control rats. Both group of rats were also maintained on the
same rat chow. Evidently the control and the test rats likely had identical
baseline Cd level and effects or the same incremented build up of Cd from any
other source upon which the load and effects due to CdSO4 consumption
will be distinctly superimposed.
Table 2. Volumes and biochemical
parameters of urine from control and Cd-exposed rats (mean ± sd)
Duration of exposure
(month)
|
A
|
B
|
C
|
D
|
X
|
Z
|
X
|
Z
|
X
|
Z
|
X
|
Z
|
1
|
14.0±2.7
|
10.3±1.8*
|
5.74±0.13
|
5.06±0.9
|
36.0±1.7
|
35.0±1.20
|
68.0±21.0
|
80.0±23.4
|
2
|
13.1±1.7
|
8.1±2.5*
|
8.1±0.22
|
10.2±1.1*
|
33.0±7.8
|
43.0±9.8*
|
32.0±6.9
|
25.3±7.5
|
3.
|
8.2±3.7
|
6.2±1.1
|
5.6±0.15
|
9.64±0.06*
|
14.0±7.1
|
11.1±8.0
|
94.0±14.2
|
81.3±29.0
|
* Values statistically significant different from control
(p < 0.005).A = 24h urine volume (ml), n = 5; B = urine protein conc. (mg/day),
n = 5: C = urine glucose conc. (mg/dL), n = 6; D = urine creatine conc.
(ug/mL), n = 6; X =
control (- Cd); Z = test (+ Cd)
Twenty-four hour urine
volume, urinary protein, glucose, amino acid and N-acetyl b D glucosaminidase
are usually studied as indices of the functional integrity of renal proximal
tubules while proper glomerular function is assessed by examining the plasma
creatinine or blood urea nitrogen
level (Friberg et al., 1986). In the present study we examined urine
volume, urinary protein and glucose as well as plasma creatinine. We found that
24 h urine volumes were significantly (P < 0.05) decreased in the test rats
at the end of 1 and 2 months respectively (Table 2). Urine proteins were, however,
significantly (P < 0.05) increased in the test rats at the end of 2 and 3
months respectively. Significant (p < 0.05) increase in glucose was demonstrated
in the urine of the test rats exposed to Cd for 2 months. Increased urine protein
and glucose levels were observed at the end of 2 months
of oral Cd exposure. Proteinuria has been observed in man and animals and shown
to be due to renal tubular dysfunction (Friberg et al., 1986; Horiguchi et
al., 1996). Evidently bioaccumulation of cadmium in the kidney of the test
rats appreciated to a point (by the end of the second month in particular) where
it caused proteinuria and glucosuria, two conditions which in combination with
others are evidences of renal tubular dysfunction.
Table 3. Plasma creatine and
glucose in control and Cd-exposed rats (mean ± sd); n = 6.
Duration of exposure (month)
|
Creatine conc. ug/ml x 10-2
|
Glucose conc. (mg/dL0
|
Control (-Cd)
|
Test (+Cd)
|
Control (-Cd)
|
Test (+Cd)
|
1
|
6.6±1.2
|
7.6±1.5
|
91.7±3.4
|
102.8±6.0*
|
2
|
1.2±0.2
|
0.9±0.3*
|
106.0±9.5
|
100.0±1.8
|
3
|
6.8±1.4
|
6.6±0.7
|
91.9±3.2
|
91.4±1.96
|
*Values statistically significantly
different from control (p<0.05)
In our study 24 h urine volume
did not increase but
decreased in the test rats. Urinary protein was increased in the test rats only
at the end of 2 and 3 months. Urinary glucose was significantly increased in
the test rat only at the end of 2 months exposures. Relative to the controls
Horiguchi et al. (1996) observed significant increase in 24 h urine volume,
urinary protein and glucose at the end of their 6 and 9 months
exposure periods. The absence of increased urine volume and lack of consistency
in the increase of protein and glucose in the urine as reported here are likely
attributable to the low level of cadmium in the kidney of our
test rats. As is evident in table 1, the 2 and 3 months test kidney Cd load
were 28.00 ± 5.00 ´ 10-2 and 56.00 ± 8.21 ´ 10-2 mg/g
respectively whereas the 6 and 9 month test kidney Cd load reported by Horiguchi et
al. (1996) were 163.2 ± 5.0 and
134.3 ± 3.8 mg/g respectively.
Plasma creatinine or blood urea nitrogen is used for assessing renal glomerular
function. In this study we assessed the
level of creatinine for this purpose. Our results show no difference in plasma
creatinine level between the control and test groups at the end of the first
and third months of exposure. At the end of 2 months of exposure plasma creatinine
was significantly (P < 0.05) decreased relative to the control. Absence of
increased plasma creatinine indicates that renal glomerular function was not
compromised in this study. Our finding is apparently in agreement with that
of
Horiguchi et al. (1996). Their treatment failed to produce pronounced
difference in plasma creatinine between the control and test at the end of their
exposure periods. Interestingly at the end of their first exposure period plasma
creatinine of the Cd-exposed rats was lower than that of the
control rats. This is very much like our finding at the end of 2 months of exposure
although the decrease in our own case was significant. Taken together, therefore,
our results provide partial evidence of tubular rather
than glomerular dysfunction occasioned by oral consumption of water containing
0.3mg Cd/L.
Table 4. Activity of Renal ATPase in control and Cd-exposed
Rats (Mean ± SD; n = 6)
Duration of exposure (month)
|
mMol
pi/min/mg protein ´ 10-2
|
Total ATPase
|
Mg2+ ATPase
|
Na+ -
K+ATPase
|
X
|
Z
|
X
|
Z
|
X
|
Z
|
1
|
0.41±0.02
|
0.49±0.04
|
0.28±0.004
|
0.35±0.006*
|
0.13±0.03
|
0.14±0.01
|
2
|
0.37±0.10
|
0.29±0.12*
|
0.27±0.09
|
0.21±0.08*
|
0.098±0.034
|
0.082±0.047
|
3
|
0.23±0.07
|
0.18±0.048*
|
0.17±0.04
|
0.11±0.027*
|
0.066±0.027
|
0.068±0.023
|
*Values statistically significantly different from corresponding
control (p<0.05)
X = control (-Cd)
and Z = test (+Cd)
Our 24 h urine volumes at the end of 1 and 2 months exposure
periods were significantly (P < 0.05)
decreased (Table 2). Extracellular fluid (ECF) volume is influenced by the movement
of Na+ into and out of a cell. Increased ECF osmolarity decreases
renal water excretion (Abraham and Schrier, 1997). Water and Na+ movement
across plasma membrane is controlled by a transmembrane enzyme, Na+ -
K+ ATPase, one of a number of pump enzymes (Voet and Voet,
1995). In order to obtain a possible explanation for our Cd-induced reduction
in 24 h urine volume we assayed for the activity of total ATPase, Mg2+ ATPase
and Na+ K+ ATPase in the kidney
homogenates. We found that total ATPase and Mg2+ ATPase activities
were significantly reduced in Cd exposed rats at the end of 2 and 3 months
exposure (Table 4). Na+ K+ ATPase activity was not
altered. It does appear then that the Cd induced low 24 h urine volume is
not brought about by altered Na+ K+ - ATPase
activity. All the same others (Nishiyama et al., 1986) have reported
that Cd exposure increases sodium and water retention, low urine volume. However,
Horiguchi et al. (1996) observed increased 24 h urine volume among Cd-exposed
rats. It is therefore obvious that the factors that influence Cd-induced alteration
in 24 h urine volume in man and animals are yet to be clearly established and
deserves further investigation.The kidney is the critical target organ in man
when subjected to long-term exposure to low or
high doses of cadmium. The results presented here suggest that in rats, consumption
of 0.3mg Cd/L, the level of the toxicant in Warri River between 1986 and 1991
(Egborge 1994) brought about partial tubular dysfunction after 2
months of exposure. Therefore, barring any species difference continuous consumption
of water contaminated with cadmium to the extent reported by Egborge (1994) will
likely cause some degree of renal tubular dysfunction.
REFERENCES
-
Abraham W.T., Schrier
R.W. (1997). Renal sodium
excretion, oedematous disorders and diuretic use. In: Schrier R.W. (ed).
Renal and electrolyte disorders, 5th Edn. Lippincott Raven publishers.
Philadelphia, p 72 130.
-
Adam-Vizi V., Seregi
M. (1982). Receptor dependent
stimulatory effect of noradrenalin on Na+/K+ ATPase in
rat brain homogenate. Role of lipid peroxidation. Biochem. Pharmacol. 31: 2231 2236.
-
Cabrera C., Ortega E.,
Lorenzo M., Lopez M. (1998). Cadmium contamination of vegetable crops,
farmlands, and irrigation waters. Rev. Environ. Contam. Toxicol. 154: 55 81.
-
Chapatwala D.K., Rajanna
B., Desaiah D. (1980).
Cadmium induced changes in gluconeogenic enzymes in rat kidney and liver. Drug
Chem. Toxicol. 3(4): 407 420.
-
Egborge A.B.M. (1994). Water
pollution in Nigeria.
Biodiversity and Chemistry of Warri River. Ben Miller Books, (Nig) Ltd.,
Warri.
-
Horiguchi H., Sato M.,
Konno N., Fukushima M. (1996). Long-term cadmium exposure induces anaemia
in rats through hypoinduction of erytrhopoietin in the kidney. Arch. Toxicol.
71: 11 19.
-
Nishiyama S., Nakamura
K., Konishi Y. (1986). Blood pressure and urinary sodium and potassium excretion
in cadmium treated male
rats. Environ. Res. 40: 357 364.
-
Obi F.O., Usenu, I.A.,
Osayande J.O. (1998). Prevention of carbon tetrachloride-induced hepatotoxicity
in the rat by H. rosasinensis
anthocyanin extract administered in ethanol. Toxicology 131: 93 98.
-
Voet D., Voet J.G.(1995).
Biochemistry, 2nd Edn. John Wiley and Sons, Inc. New York, USA.
-
WHO
Report. (1984). Guidelines for drinking water quality. Health criteria and
other supporting
information. World Health Organisation. Geneva
pp. 84-90
-
WHO report (1992). Environmental
Health Criteria.
134, Cadmium. World Health Organization, Geneva.
Copyright 2004 - Journal of Applied Sciences & Environmental Management
|