World Applied Sciences Journal 16 (3): 376-388, 2012

ISSN 1818-4952
© IDOSI Publications, 2012

Histopathological Alterations in Renal Tubules of

Female Rats Topically Treated with Paraphenylen Diamine

Laila A. Hummdi

Department of Biology, Science Faculty for Girls,

King Abdul-Aziz University, Jeddah, Saudi Arabia

Abstract: During recent years, several publications have reported series cases of severe systemic intoxication
in people who had used para-phenylendiamine (PPD) containing skin paints (Temporary tattoos) or following
the usual exposure through use of permanent hair dyes. The presenter experiment involved 2 treatment groups
and 2 control groups 20 rats each. PPD was topically applied on rats back at doses of 3 or 6 mg/kg of PPD for
30 minutes once daily for 6 months. Increased levels of serum creatinine, urea nitrogen, alkaline phosphatase
and decreased serum Na+ and Ca2+ were observed as significant biochemical parameters in the treated rats.
There was a significant increase in body and kidney relative weights in groups PPD poisoning. Most of the
histo-pathological changes appeared more pronounced on the proximal convoluted tubules (PCTs) than distal
convoluted tubules(DCTs). PCTs at 3 mg/kg of PPD treated group III showed brush border fragmented, cells
necrosis and nuclear pyknosis and karyolysis. Cytoplasmic vacuolation with accumulation of eosinophilic
homogenous material in tubular lumina also seen. Extensive tubular epithelial vacuolation and necrosis, as well
as cell lysis, focal haemorrhagic and cellular infiltrating, interstitial edema and vascular degenerative changes
were evident in all rats treated with 6mg/kg. While,ultra-structurally, PPD induced various degrees damage in
PCTs epithelium in groups III and IV, such as low cytoplasmic electron density, loss cellular organelles,
deformed nuclei, widen basal infolding with deformed, vacuolated, mitochondria. Brush border shrinkage and
vacuolated with fragmented and dilatation of rough ER in 6 mg/kg PPD-treated group. DCTs at 3 mg/kg PPD
revealed intracellular vacuolations, destruction of cytoplasmic organelles, deformed mitochondria and Golgi
bodies with invagination of the nuclei membrane. Additionally, short and damage basal infolding with sever
reduction in mitochondrial number and size were appeared in DCTs of 6mg/kg PPD treated group IV. Also it is
important there was a strong aware for the regulation and restriction of sale of para-phenylenediamine.

Key words: PPD Proximal tubules Distal tubules Histopathology Ultra-structure

INTRODUCTION PPD poisoning is a common health problem in

Paraphenylene diamine (PPD) [C6H4(NH2)2] is an
aromatic amine not found in nature and it is commercially
produced by many industrial companies. It is a derivative
of para nitro analine that is available in the form of white
crystals when pure and rapidly turns to brown when
exposed to air [1]. It is widely used in industrial products
such as textile or fur dyes, dark colored cosmetics,
temporary tattoos, photographic development and
lithography plates, photocopying and printing inks,
black rubber, oils, greases and gasoline [2]. It is also used
to kill wild animals when added to food [3].
the Middle East, it is also common in India, but rare
in the west. Most reported cases were from
adolescents and adults [4,5]. Several acute PPD poisoning
cases had been reported, in particular, from Africa and
Asia whereas it is traditionally used mixed with henna
(leave of Lawsonia alba) tattoos which is traditionally
applied to color the palms of hands and to dye the
hairs[6].
Acute toxicity has been reported following oral,
subcutaneous, intraperitoneal and topical application
in a variety of species. The LD50 following oral
administration was 80-100 mg/kg in the rat, 290 mg/kg in

Corresponding Author: Laila A. Hummdi, Department of Biology, Science Faculty for Girls,
King Abdul-Aziz University, Jeddah, Saudi Arabia.
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 World Appl. Sci. J., 16 (3): 376-388, 2012
mice, 250 mg/kg in rabbit and 100 mg/kg in cats [3,7]. The
lethal dose for human was estimated to be 10 gram of pure
PPD [3,8].
Few studies investigated the dermal absorption
potential of PPD in humans and animals[9]. Under the
intended-use conditions, dermal absorption of 0.54 to
2.7% in volunteers [5,10-12] and 2.7% in monkeys
[10,11,13] has been reported. Dermal absorption of 2.7%
has been noted in human cadaver skin [14]. The degree of
dermal absorption was reported as 0.93% in excised pig
skin[11]. Hueber-Becker et al. [12] reported similar dermal
absorption values, of 2.44 and 3.39% in vitro, in human
and pig skin, respectively.
Hence, the henna tattoo industry is not regulated,
people are still getting black henna tattoos and exposing
themselves to serious toxicity problems. In addition,
tattoos are very popular in Arabian Gulf, especially in
Saudi Arabia and there is no report has been available
regarding chronic PPD dermal applied and subsequent
nephrotoxicity. Therefore, the main objective of the
present work was to investigate the effect of repeated
topical application of PPD on the histological picture of
the kidney in female rats.
MATERIALS AND METHODS
Chemical: The tested chemical P-Phenylene diamine
(PPD), CAS No: 106-50-3). Batch: 99E483. Molecular
weight (MW): 108 and 98% purity was purchased from
commercial markets in Jeddah of Saudi Arabia.
Chemical Formula: C6H8N2 (free base),C6H8N2. 2HCl
(dihydrochloride),C6H8N2. H2SO4(sulfate) [9].
Structural Formula
Physical Form: White to light purple powder[9].
Animal Treatment: According to OECD [15,16], on the
design and conduct of chronic toxicity and the Council of
Europe Convention Appendix A [17] recommendations on
rodents housing, groups of total 80 female rats were
obtained from the Animal House of the King Fahd Center
for Medical Research, King Abdul Aziz University in
377
Jeddah of Saudi Arabia, weighing 282.67±7.10 g. Rats
were kept on a 12 h/12 h light/dark schedule with a free
access to standard laboratory food and water at room
temperature. Rats were divided into four groups 20 rats
each. Group III and IV were painted on shaved skin of
approximately 10% of the total body surface area on the
back(interscapular region) with 3and 6mg/kg respectively,
using a 1:2 mixture of PPD and degassed purified water for
30 minutes, daily for a period of 6 months according to
guideline for chemical testing OECD[18,19] and [20,21].
While, groups I and II received topical application of 3
and 6mg/kg of degassed purified water, respectively and
served as controls. The rats were observed for overt signs
of toxicity and for mortality, the individual body weights
were recorded weekly. Animals were killed by cervical
dislocation, kidneys absolute and relative weights were
recorded at necropsy. Blood biochemical studies were
done after the end of the experiment (day 180 of study).
Biochemical Assays: At the end of the trail-24 hours after
the experiment, blood samples were collected and
analyzed for serum urea nitrogen (UN), creatinine (Cre),
alkaline phosphatase (ALP),Na+ and Ca2+ were performed
by standard methods at the Central Laboratory of King
Fahd Center for Medical Research, KAU in Jeddah of
Saudi Arabia.
Histological Study: Kidneys were excised and fixed in
10% buffered formalin, dehydrated in graded ethanol and
embedded in paraffin by standard methods. Two ìm thick
transversal sections of the kidney were mounted on
positively charged glass slides. Subsequently, the
sections were stained with hematoxilin/eosin (H&E),
according to standard methods [22].
Transmission Electron Microscopy: The kidneys were
treated according to the standard procedure [23]. kidney
blocks were trimmed to 1×1×1-mm size and washed,
quenched, post-fixed with ethanol, dehydrated into
propylene oxide and embedded in epoxy resin for
transmission electron microscopy(TEM)studies. Thin
sections were cut on Reichert-Jung Ultra-cut E, then
stained with uranyl acetate and lead citrate and finally
examined with a Philips EM 100.
Statistical Analysis: All data were presented as means
±SEM. Multiple comparisons were performed using
student's t-test. A value p < 0.05 were taken into
 World Appl. Sci. J., 16 (3): 376-388, 2012

consideration for determining significance. All
Statistical procedures were computed using SPSS 18.0
software.
RESULTS AND DISCUSSION
Clinical Manifestations and Mortality: There was a high
percentage of mortalities and clinical signs related to the
test concentration of PPD in both treated groups III and
IV as compared to controls (Table1). A number of earlier
reports suggested that a fraction of topically applied PPD
alone or in combination with an oxidizing agent reach
systemic circulation after percutaneous absorption
[11,24]. The lethal dose of PPD is not known; estimates
vary from 7-10 grams [24]. From the present data, it was
evident that the treatment of PPD caused a swelling face
and neck, dark discoloration of urine, ataxia, dermatitis
and mortality in rats of treated groups III&IV, all these
clinical sings were matched with previous studies
[3,4,25-28] and correspond with our study in female rats at
dose 0.5 and 1mg/kg of PPD [29]. Moreover, Prasad
et al. [30] showed that PPD produces local toxic effects in
the form of skin irritation, contact dermatitis, with 5-10ml.
In contrast, groups of 5 female rabbits received 5 or 10%
PPD applied twice weekly for 85 weeks, no abnormalities
were found in the blood or urine of the rabbits and no
treatment-related local changes were observed [31].
Mortalities were observed in a previous study at dose of
0.2 in 9 male rats and in 1 female rats a 12-week oral
toxicity with PPD on F344 rats administered
concentrations of 0.05, 0.1, 0.2 and 0.4 % /day [32]. Ayoub
et al. [26] reported that mortality in male rats treated with
PPD is 21.1% and 22% in poisoning from hair dye [8],and
10% -35 in females Wistar rats [29] and 41.9% in PPD
poisoning cases [24]. However, the mortality of rats in the
resent study was more 40 and 55% of groups III and IV,
respectively, probably because of prolong PPD exposure
period and PPD concentrations. PPD has also been shown
to produce myocarditis and arrhythmias leading to
sudden death [33]. Death is usually caused by
angioneurotic edema or arrhythmias due to direct
cardiotoxicity of PPD cases of poisoning of PPD which
develops renal failure require dialysis. The cause of renal
injury is probably direct nephrotoxicity of compound [24].
Body and Kidney Weights: Based on data in Table 2, a
significant increase in body weight in both treated groups
as compared to controls, as well as increased absolute
and relative kidney weights were noted. Previous study
showed both absolute and relative kidney weight of rats
exhibits a dose response increase after PPD treatment [34].
Wakelin et al. [35] revealed that mean absolute and
relative liver and kidney weights raised in males wistar
rats topically given 5 mg/kg/day or greater. Likewise, in

Table 1: Clinical symptoms and the mortality recorded in groups 3and4 of PDD treatment rats after 24 hr- 6 months.
No. of Experimental animals =20 No. of Experimental animals =20
Clinical symptoms N0.of individual % N0.of individual %
Swelling face and neck 17 85% 19 95%
Dark discoloration of urine 19 95% 19 95%
Ataxia 11 55% 16 80%
Skin changes(dermatitis) 9 45% 14 70%
Mortality 8 40% 11 55%

Table 2: Body weight and kidney weight(absolute and relative) of experimental rats after 6 month topical application of PPD.
Experimental Groups
Weights Control (G1) Control (G2) Treated (G3) Treated (G4)
Body Weigh M 282.6667 283.5664 292.0270 301.31233
±SEM ±7.10712 ±7.00732 ±4.76599 ±7.15936
P - - .003* .000**
Absolute Kidney weight M .7200 .7190 .8903 .9854
±SEM ±.02000 ±.02100 ±.56879 ±.35444
P - - .0530* .0310*
Relative Kidney weight M .3780 .3696 .4042 .4623
±SEM ±.01158 ±.01178 ±.03323 .04765±
P - - .0011* .000**
* P < 0.05,** P < 0.005

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Table 3: Serum creatinine and serum urea of experimental animals kidney after 6 month topical application of PPD.
Experimental Groups
Kidney bioassay Control (G1) Control (G2) Treated (G3) Treated (G4)
S. creatinine mg/dl M .5300 .5310 .9543 1.96320
±SEM ±.04637 ±.12627 ±.08380 ±.05337
P - - . 000** .000**
Blood urea mg/dl M 5.1800 5.1900 22.2780 25.0340
±SEM ±.32772 ±.34726 ±.56925 ±.49989
P - - .000** .000**
Alkaline phosphatase (U/L) M 48.326 50.221 116.195 198.545
±SEM ±.7200 ±.8400 ±.11076 ±.9513
P - - .000** .000**
Na+ (mmol/L) M 14.023 14.322 8.7722 8.2856
±SEM ±.2330 ±.2371 ±.3200 ±.5041
P - - .000** .000**
Ca2+ (mmol/L) M 13.0430 15.1009 8.7391 8.5911
±SEM ±.392 ±.2899 ±.372 ±.5008
P - - .001** .001**
* P < 0.05,** P < 0.005

females given 0.05 and 1 mg/kg of PPD were reported in
our previous study [29]. In addition, the body-weight-
related kidney weights were significantly increased of
females given 8 mg/kg/day and 16 mg/kg/day at 13 weeks
treatment [36]. However, that PPD administered to Wistar
rats with 0.05ml once a week for 18 months, resulted in a
slight decrease in the bodyweights of the males after 30
weeks of exposure and no such effects were found among
the females [37]. Imaida et al. [32] noted that at 0.02% of
PPD, a 50 % reduction in body weight in both sexes of
rats as, as well as increased relative liver and kidney
weights.
Effects of PPD on Renal Function: From the
previous studies and the present results (Table3),
it could be concluded that the urinary parameters
measured showed significant (P<0.005) rising in UN and
Cre levels correspondingly, ALP activity was
increased in two treated animals groups. The serum Na2+
and Ca2+ concentrations in dosed animals were markedly
lower than in the control animals. These data indicate
reduced ability of the kidney to eliminate the toxic
metabolic substances and reabsorb the metal and non-
metal ions [3]. Moreover, there are some reports which
have been pointed out the changes in the biochemical
profile in patients who have consumed the hair dye
containing PPD [2,38-40]. Other findings showed that
chronic frequent PPD exposure is deleterious to kidney
structure and function of female rats[29]and in part, due
to the presence of quinine-diamine which is a potentially
nephrotoxic substance [2,3].
Histological Results: Histological examination restricted
significant findings in renal tubules at the cortical portion
of kidney in treated groups III and IV. Most of the
pathological changes appeared more pronounced on the
proximal convoluted tubules (PCTs) than distal
convoluted tubules (DCTs). In group III treated with 3
mg/kg PPD consisted of brush border fragmented, cells
necrosis with dilated intercellular spaces and epithelium
detachment, nuclear pyknosis and karyolysis (Figs.1a-c).
In addition, cytoplasmic vacuolation, dilated with
accumulation of eosinophilic homogenous material in
tubular lumina (Fig.1d).
These findings are consistent with Sampath and
Yesudas study [33] who reported that renal structure
shows acute tubular necrosis in PPD toxicity Chugh et al.
[1] described the histological change of acute tubular
necrosis in PPD poisoning as vacuolar degeneration,
nuclear pyknosis and cytoplasmic vacuolations brush
border damage. The most pronounced histological
changes that observed in glomerular cells in our previous
experiment were similar to those described in tubular cells
in present experiment such as vacuolization of endothelial
cells with necrotic changes of many mesangial cells and
podocytes [29]. Tubular obstruction may be caused by
release of cytoplasmic fragments mixed with intratubular
proteins. This can lead to increased intratubular pressure,
which again may result in sufficient back pressure to alter
the transglomerular hydrostatic pressure. Autopsy of PPD
poisoning patients revealed renal tubular occlusion due
to myoglobin casts with histological evidence of acute
tubular necrosis [3,24,40]. Myoglobin, which was released

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Plate (1a-h): Cross sections of kidney cortex of female wister rats; H&E. (a): kidney section of control rats (G1) showing
the normal histological structure of the proximal convoluted tubules (arrow) & distal convoluted tubules
( ) in the cortical portion;x400.(b): showing a part of renal corpuscle(Rc) and interstitium(It), brush border
(Bb) and large central nuclei in PCTcells;x1000. (c-d): sections of 3 mg/kg PP D treated kidney (G3): (c):
showing brush border fragmented(*), cells necrosis ( ), dilated intercellular spaces( thin arrows ) and
detachment of them from the basement membrane (thick arrows), pyknosis(P)and karyolysis(K) of tubular
cells nuclei; x1000.(d): showing cytoplasmic vacuolations (arrows), accumulation of eosinophilic
homogenous material in dilated tubules lumen ( ) ;x1000. (e-h): sections of 6g/kg PPD treated kidney
(G4):(e): showing massive tubular epithelial cells vacuolations, focal mononuclear leucocytes ( ) and focal
haemorrhagic areas(arrows)a; x1000.(f): inflammatory cells infiltrating and interstitial edema at the
corticomedullary portion;x1000.(g): showing marked architecture degeneration, dilation of the cortical blood
vessel(arrow) with red blood cells stasis ( ); x1000. (h): showing normal structure of the cortical blood
vessel; x1000.

as a result of rhabdomyolysis has been implicated in heme
induced renal damage, mainly by causing oxidative
damage to the renal tubules [41]. However, renal failure
can also occur as a result of other nephrotoxic chemicals
of PPD substance [ 30]. In another observational study of
hair dye poisoning in Hyderabad, India, acute renal failure
developed in 100% of patients due to rhabdomyolysis
since myoglobin is a small molecule with a molecular
weight of 17 kDa, which binds only lightly to the plasma
proteins, it escapes easily in the urine[ 33].

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Plate (2a-c): Electron micrographs(E.M.) of proximal convoluted tubules of kidney rat 3mg/kg PPD treated(G3): (a):
showing brush border disorganized(Bb) cytoplasmic lysis (arrows) irregularly of nuclei membrane with
loose chromatin materials, widen basal membrane infolding ( ), the tubular lumen filled with damage
mitochondria and secretion (*); x2600.(b):showing damage epithelial lining of a PCT cell, note dilated basal
membrane infolding ( ), scattered deformed electron dense mitochondria (arrows) deformed nucleus(N)
coalescence of brush border (*) ; x5800.(c): E.M of a part of PCT cell of a control rat kidney showing a large
rounded central located nucleus(N) with normal chromatin distribution, numerous well organized elongated
mitochondria (arrow) oriented parallel to the cell axis surrounding by basal membrane infolding ( ); x7900.

On the other hand, extensive tubular epithelial cells
vacuolation and necrosis as well as cell lysis, focal
mononuclear leucocytes, haemorrhagic area and
inflammatory cells infiltrating and interstitial edema at the
corticomedullary portion were evident in all rats treated
with 6mg/kg PPD in group IV (Figs. 1e,f). There was an
increase in the incidence of marked vascular degenerative
changes in the lining epithelial cells of the blood vessels
at the cortical portion and distortion their architecture
associated with dilation and red blood cells stasis
(Figs.1g,h). These results were corresponds to those of
Suliman et al. [42] who confirmed that 5mg/kg of PPD
cause acute renal failure within the first week. Similar
study [34] suggested that the common histopathological
finding among all the PPD treated animals at dose 1 mg
and 3mg /kg were severe necrosis and inflammation
characterized by increased infiltration of leucocytic
component accompanied by extensive hemorrhage and
hydropic degeneration in hepatocytes. Acute renal and
hepatic failure with hemorrhage in PPD poisoning [24,29]
and intravascular haemolysis with hair dye poisoning
were showed [43].

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Plate (3a-c): Electron micrographs(E.M.) of proximal convoluted tubules of kidney rat 6g/kg PPD treated(G4): (a):
showing severe impairment of proximal tubule brush border(Bb); shrinkage, loss, swelling and vacuolation,
cytoplasmic lysis(*) and mitochondria vacuolation ( ) fragmented and dilated RER(arrows);x13500.(b,c):
E.M. of apical part of PCT cell of a control rat kidney, (b): showing a large rounded nucleus(N) oval
mitochondria(M) with transverse plate like cristae;x19000.(c):showing brush border (Bb) with closely packed
and elongated microvilli; x13500.

PPD-treated groups III and IV under transmission
electron microscopy, indicated that PPD induced various
degrees of damage on the architecture of proximal tubular
epithelium when compare to controls samples, such as
brush border disorganized and coalescence as well as cell
swelling, cytoplasmic lysis with loss most of the
cytoplasmic electron density and cellular organelles,
irregularly- shaped of nuclei with loose chromatin
materials( karyolysis). In addition to, widening the basal
membrane infolding with scattered deformed electron
dense mitochondria, tubular lumen filled with damage
mitochondria and secretion were observed in groupIII
(Figs. 2a-c). Moreover, PPD caused severe impairment of
proximal tubule brush border, such as shrinkage, loss and
vacuolated with swelling and mitochondrial cristae
vacuolation, fragmented and dilatation of rough
endoplasmic reticulum (RER) in 6mg/kg PPD-treated group
IV(Figs. 3a-c).
The tubular lesions found in rats intoxicated with
PPD show several similarities with the lesions described
for chemical pollutants and different toxins, such as
cytoplasmic vacuolization,low electron density, swollen

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Plate (4a,b): Electron micrographs(E.M.) of distal convoluted tubules cells of kidney rat 3mg/kg PPD treated(G3): (a):
showing intracellular vacuolations ( ), destruction of cytoplasmic organelles(arrows), irregular, dilation
and distortion of basement membrane infolding (*),loose chromatin materials with irregular nucleus
membrane(N);x3400.(b):a part of epithelial lining cell showing loss most of the cytoplasmic electron density
and cellular organelles(*),deformed mitochondria ( ) and Golgi bodies(G), with invagination of the nucleus
membrane (arrow); x19000. (c,d): E.M. of DCTs cells of a control rat kidney;(c):showing apical spheroid
nuclei(N) of DCT, well developed basal infolding ( ) surrounding filamentous elongated
mitochondria(arrows);x2600.(d): high power of previous section showing apical cytoplasmic knobs
(Ck),Golgi apparatus(G), elongated mitochondria(M) marginal heterochromatin in nucleus(N),
desmosomes(arrow);x13500.

nuclei with chromatin margination, brush border
damage,vacuolated and swollen the mitochondria with
cristae devastation [44-49]. The microvilli disappeared
[47,48], increased density and closely packed
mitochondria which probably representing apoptotic cells
were also noted [46.49]. Furthermore,there were several
factors that may have played a role in PPD tubular
injuries, such as decrease in glomerular capillary
permeability, back-leakage of glomerular filtrate through
tubular cell walls [50]. Damage the brush border and
leakage of ALP into urine could have been a result of
toxin binding to the brush border. This enzyme is
associated with the brush border of the renal tubules and
the urinary concentration of ALP in particular has been
used as an early marker of toxic tubular insult [51,52]. The
pathological changes in mitochondria as swollen or

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Plate (5a,b): Electron micrographs(E.M.) of distal convoluted tubules cells of kidney rat6g/kg PPD treated(G4): (a):
showing heterogeneity in epithelial cells damage including necrotic cells with pyknotic nucleus(P) and
hypertrophic cells (*) with low electron dense cytoplasm, loss most cellular organelles and karyolysis
nucleus(N);x5800. (b): high power E.M. from previous section showing short and damage basal infolding
of DCT cell ( ) with sever reduction in number and size of mitochondria ;x19000. (c): basal part of DCT cell
showing well developed basal infolding (arrows)surrounding rod-like shaped elongated mitochondria
(M);x13500.

cavitations are an important indicator of cellular
damage leading to loss of functional efficiency [45].
The mitochondrial cristae Smashed may be due to
disturbance of calcium ions Ca2+ balance into cells, or
block the ATPase synthesis and thus block the
phosphorous oxidation in mitochondria [53,54], or
lack of permeability of mitochondrial membrane [56] or
cellular PH disturbance[56]. However, Prasad et al. [30]
reported that PPD promotes calcium release and the
leakage of calcium ions from the smooth endoplasmic
reticulum, thus causing irreversible change in the cell
structure.

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From the ultrastructural observation of samples of the
distal convoluted tubules(DCTs) in PPD-treated groups
showed changes of the DCTs cells after administration of
3 mg/kg of PPD (Figs. 4a,b) when compared to the
controls (Figs. 4c,d).These comprised intracellular
vacuolations, destruction of cytoplasmic organelles,
deformed mitochondria and Golgi bodies, splitting and
irregular of basement membrane with dilation and
distortion of basal infolding. Associated with, lose of
chromatin materials and invagination of the nuclear
membrane. Additionally, the DCTs cells in PPD group
given 6mg/kg appeared heterogeneity in epithelial cells
damage including necrotic cells with pyknotic nuclei or
hypertrophic cells with shorten and damage basal
infolding associated by sever reduction in number and
size of mitochondria (Figs. 5a,b) when compared to the
controls (Figs. 5c). All these degenerative changes are
consistent with the several studies reported renal toxicity
associated with toxic substances leading to DCTs
histological damage [46-49,57].However, Kaloyanides and
Ramsammy [58] reported that the rough ER injury due to
collect of the pollutants and their metabolites products in
inside the ER cisternae. The kidney is an important organ
for maintaining the calcium homeostasis. Ca2+ is
reabsorbed by paracellular diffusion and transcellular
active transport [59]. There has been evidence for
mitochondrial accumulation of Ca2+ in ischemic acute renal
failure and that this calcium uptake impairs renal
mitochondrial phosphorylation and adenosine
triphosphate synthesis [60]. Mitochondrial uptake of
calcium may explain the tendency toward lowering of
serum calcium levels and may have played a role in the
damages observed in the epithelial cells.
On the basis of previous studies,the nephrotoxicity
manifest in this study mainly attributed to the smaller
amount of parent PPD molecules that reach systemic
circulation and after percutaneous absorption.
Furthermore, The plasma clearance rate of PPD is biphasic
and plasma half life is estimated to be 24 min and 43.5 h,
respectively [34]. This may explain the apparent
nephrotoxicity of the PPD molecule, since a little fraction
of PPD absorbed through skin is possibly retained for
a longer period (43.5 h) so as to produce toxicity.
The mechanisms of acute tubular necrosis involved in the
development of the changes in morphologic and
functional parameters are certainly many and complex.
Most of the injuries resulting from hypoxia dehydration,
intravascular haemolysis, Methaemoglobinaemia and
direct toxic actions of chemical or it?s by products on the
renal tubules [24].
385
Other possible mechanisms may involve reactive
intermediates or oxidative stress or both [50]. Biologically
reactive intermediates are electron-deficient compounds
(electrophiles) that bind to cellular electron-rich
compounds (nucleophiles), such as proteins and lipids
[61]. Mixed-function oxidases catalyze the formation of
these toxic metabolites. Reactive intermediates bind
covalently to critical cellular macromolecules and interfere
with normal biologic activity. Oxidative stress is induced
by increased production of reactive oxygen species
(ROS), such as superoxide anion, hydrogen peroxide and
hydroxyl radicals [61]. ROS can induce lipid peroxidation,
inactivate cellular enzymes, depolymerize polysaccharides
and induce deoxyribonucleic acid breaks and chromosome
breakage. Swelling of the mitochondria observed in our
study could have been a result of inhibited mitochondrial
respiration. Many of the common nephrotoxic substances
appear to act by means of alterations in the renal
perfusion [62] and some of our findings were in agreement
with such a mechanism. The dominant tubular lesions
were in the outer cortex. This zone is limited oxygen
supply [62,63] and thus easily get damaged if a
nephrotoxic substance diminishes the renal perfusion.
CONCLUSION
It is concluded that the test substance PPD is an
extremely potent kidney toxicant in female rats, confirming
results from numerous previous studies.PPD intoxication
is a life threatening condition. Health authorities should
call for the prevention of the use and trade of PPD in the
market. Awareness programs about its toxicity should be
implemented at different levels.
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