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Current Molecular Pharmacology, 2014, 7, 000-000 1

Effect of Environmental Contaminants on Mammalian Testis

Faustin Pascal Tsagué Manfo1*, Edouard Akono Nantia2 and Premendu Prakash Mathur3*

1
Department of Biochemistry and Molecular Biology, Faculty of Science, University of
Buea, PO Box 63 Buea, Cameroon; 2Department of Biochemistry, Faculty of Science,
University of Bamenda, PO Box 39 Bambili, Cameroon; 3KIIT University, Bhu-
baneshwar-751024, Odisha, India

Abstract: Exposure of humans and wildlife to pollutants released in the environment is a centre
of attention nowadays. Many of these chemicals (generally referred to as environmental pollut-
ants) have been shown to interfere with normal hormonal signalling and biological functions,
Faustin Pascal Tsagué Manfo leading to reproductive disorders or infertility, which has been a matter of concern within the Premendu Prakash Mathur
recent decades. The present paper reviews adverse effects of these toxicants on mammalian testes, with emphasis on al-
teration of steroidogenesis, spermatogenesis, and histopathological effects. From the publications reviewed, it appears
that environmental toxicants, especially heavy metals and organic chemicals of synthetic and microbiological origins, dis-
rupt hormone production and action in the mammalian testes. Endocrine disruption leads to disorders of testicular function
and thereby compromises the normal p henotypic development of male sexual characteristics, initiation and maintenance
of spermatogenesis. The toxicants also induce impairment of testicular cells function, testicular histology, and sperm cells
function directly. The release of the toxicants in the environment is still ongoing, despite alarming quantities that already
exist in the atmosphere. If appropriate measures are not taken, their impact on the male reproductive function and espe-
cially on testicular function will be more serious.
Keywords: Hormone, mammal, spermatogenesis, testis, toxicity.

I. INTRODUCTION due to prolonged exposure to environmental risks/pollution.

Environmental factors are estimated to contribute for 24% of
Chemicals are used in a wide variety of products and the global disease burden and 23% of all deaths [5]. Numer-
processes while they are major contributors to national and ous studies have implicated these toxicants inducing dys-
world economies. Nowadays, there is hardly any industry functions such as respiratory difficulties, cancer, neurode-
where chemicals are not used and there is no single eco- generative disorders, immune disorders, hepatic damage,
nomic sector where chemicals do not play an important role. reproductive disorders (e.g., hypospadia, cryptorchidism),
Chemicals have thus been produced in huge quantities, and etc. [1, 6, 7]. Despite these adverse effects on human health,
have become an integral part of daily life in today’s world. the pollutants are being produced and released in the envi-
These chemicals include organic chemicals, metals and their ronment continuously; and this could explain the decline in
complexes, and radionuclides [1-3]. Organic chemicals in- male fertility, as well as the high impact of environmental
clude products from petrochemical industry (products de- factors on human health [5]. Moreover, a 50% world-wide
rived from crude oil and natural gas), plastics, detergents, decline in sperm density among humans (from 1940 to 1990)
synthetic fibres, pesticides used in agriculture (herbicides, has been reported by Carlsen et al. [8]. The frequency of
insecticides, fungicides), substances of microbiological (bac- pathophysiological conditions associated to male infertility
terial and fungal) origin (e.g., aflatoxins from fungi), etc., are thus increasing, and several studies have shown that ex-
while metals include cadmium, zinc, copper, iron, etc. [2, 3]. posure to environmental toxicants (e.g., solvents, pesticides)
These substances are being released into the environment in represents one of the major triggering factors [9, 10]. Envi-
the course of their production, use, and disposal [1]. The ronmental toxicants affect the male reproductive system by
environmental contaminants have been detected in crops, directly interfering with the process of spermatogenesis in
water, air, ground, etc. [2]. testis, or through dysregulation of the hypothalamic-pituitary-
testicular (HPT) axis [Box 1]. The testis is among the most
Environmental pollutants are in contact with humans and sensitive organs to xenobiotics within the male reproductive
wildlife, and are thus capable of altering their physiology [4]. system. This observation is sustained by several analyses of
The World Health Organization (WHO) estimates that about existing reproductive toxicity study databases [11-13]. The
a quarter of the diseases mankind is facing today occur high sensitivity of testis to xenobiotics makes it a good start-
ing point for determination of the no observable adverse ef-
*Address correspondence to these authors at the Department of Biochemistry fect level (NOAEL) of toxicants in risk assessment studies.
and Molecular Biology, Faculty of Science, University of Buea, PO Box 63 To facilitate such studies, the present review paper gathers
Buea, Cameroon, Tel: +237 675 546 276, E-mail: faustinpascal@yahoo.fr
KIIT University, Bhubaneshwar-751024, Odisha, India; information on the range of endpoints such as systemic and
E-mail: ppmathur@gmail.com developmental toxicity of selected groups of environmental

1874-4672/14 $58.00+.00 © 2014 Bentham Science Publishers

2 Current Molecular Pharmacology, 2014, Vol. 7, No. 2 Manfo et al.

toxicants that result in impairment of testicular function in of the percentage of tubules producing differentiating germ
mammals, with heavy reliance on animal data. cells in DBCP –exposed male rats [19, 27].
The pesticide DDT has been banned in western countries
II. PESTICIDES
because of its non target effects and its environmental persis-
The need to produce more has made the agricultural tence. Most of the endocrine effects induced by DDT and its
chemistry more important with the rapid growth of the global major metabolite Dichlorodiphenoxydichloroethylene (DDE)
population. This has led to extensive use of agrochemical result from the ability to mimic 17-
-estradiol. In experimen-
products (pesticides) such as herbicides, insecticides, fungi- tal animals, DDT and its metabolites trigger their deleterious
cides and fertilizers, to maximize yields per hectare, through effect on spermatogenesis through direct action on Sertoli
elimination of diseases and sustainment of plant growth. cells. DDE decrease the level of FSH binding sites, modulate
Pesticides can contaminate human directly during farming expression of androgen binding protein (ABP), and induce
activities when personal protective equipments are not ap- mitochondria-mediated apoptosis in Sertoli cells [28-30].
propriately used [14, 15]. In addition to lack of protective The intracellular mechanism of o,p’-DDT is mediated
equipment, occupational exposure to agropesticides is sus- through the estrogen receptor (ER) pathway [31]. 3-
tained by increased time spent for cropping or spraying the Methylsulfonyl-DDE (Me-DDE), which also results from
pesticide, frequency and quantity of pesticide sprayed [3, DDT metabolism, down-regulates steroidogenic genes in
16]. These chemicals can also get into human system indi- Leydig cells [32].The deleterious effect of DDE on testicular
rectly through contaminated food stuffs or water [14, 15]. function in humans was demonstrated in an epidemiological
Some pesticides have not been produced and used for few study conduced in malaria endemic-areas in Chiapas (Mex-
years only, but are very persistent and tend to accumulate in ico), where the parent compound DDT was sprayed until
soil and sediments (e.g., dichloro-diphenyl-trichloroethane 2000. A regression analysis revealed a correlation between
(DDT) and related metabolites). Once pesticides are intro- reduction of motile sperm cells percentage and increase of
duced into the environment, they may travel beyond their plasma p,p'-DDE concentrations in humans [18].
point of application or discharge. Some pesticides including Dichlorvos (2, 2-dichlorovinyl phosphate, DDVP) and
triazine group, dichlorophenoxyacetic acid and glyphosate, Chlorpyrifos are organophosphorus insecticides used world-
are used for road and rail weed treatment, gardens, parks and wide for pest control in agriculture [33, 34]. Subcutaneous
urban woodland areas [17]. Pesticides and related metabo- injection of DDVP (1 and 4 mg/kg body weight (bwt), 6
lites have been detected in biological samples (blood, urine) days a week) to Wistar rats for 9 weeks induced a decrease
of exposed men [16, 18]. Their adverse effects on testicular in sperm motility [33]. In another study, DDVP (1.6
function have been illustrated in studies involving men or mg/kg/day) altered sperm morphology, and induced a de-
experimental rodents. crease in testis weights and serum testosterone levels in ex-
The nematocide dibromochloropropane (DBCP) is one posed rats after 4 and 7 weeks treatment. DDVP also induced
of the best known pesticides with proven testicular toxicity necrosis, oedema and cellular damage in testicular tissues of
in men [19, 20]. DBCP was widely used in the United the animals [34]. The DDVP–induced effects are likely to
States and other countries such as Costa Rica [19, 21, 22]. occur in male humans. A study conducted in indoor pesticide
The pesticide induced oligozospermia and azoospermia in sprayers who were often using DDVP and other organo-
men involved in its manufacture in the factory, and in phoshate (fenitrothion, chlorpyrifos/chlorpyrifos-methyl) and
farmers who were using it on crops [20, 22, 23]. The dele- pyrethroid (permethrin) insecticides, revealed reduction in
terious effect was positively correlated with the duration of testicular volume and impairment of sperm quality (reduction
exposure [10, 23]. DBCP was demonstrated to induce ele- of sperm motility) in the pesticide sprayers [35]. Chlorpyrifos
vation of serum gonadotropins (FSH) [23, 24] and DNA reduced serum testosterone levels and decreased testicular
damage [21], and histological analyses in exposed men sperm counts in male rats [36]. Administration of chlorpyrifos
revealed testicular necrosis [22, 25]. The DBCP –induced (5 and 10 mg/kg bwt) to rats for 30 days induced shrinkage of
seminiferous tubules and drastic changes in germ cells, along
adverse effects were confirmed and further clarified in ex-
with plasma levels of hormones that regulate spermatogenesis
perimental rats. Four daily injections of DBCP produced
(testosterone, FSH and LH). Chlorpyrifos induced oxidative
prolonged oligospermia in rats. Six and 20 weeks after
stress in the testis (increased levels of testicular lipid peroxida-
DBCP treatment, about 10% seminiferous tubules were
tion, and decreased activities of superoxide dismutase (SOD)
occluded or had morphologic Sertoli cells alterations in the
catalase (CAT), glutathione peroxidise (GPx)). Chlorpyrifos –
exposed rats, while up to 70% of the tubules showed an inhibitory effect on testosterone production involves inhibition
epithelium with no differentiating germ cells [26]. The lat- of 3
-hydroxysteroid dehydrogenase (3
-HSD) and 17
-
ter tubules without differentiating germ cells contained hydroxysteroid dehydrogenase (17
-HSD) enzymes [37]. The
actively proliferating type A spermatogonia, which under- deleterious effect of organophosphate pesticides on sperm
went apoptosis instead of differentiation; indicating that morphology was also reported in humans. Urinary concentra-
DBCP act on mammalian testis by inducing loss of stem tions of organophosphate metabolites (Diethyldithiophosphate
cells’ ability to undergo differentiation. The proposed ac- and diethylthiophosphate) were associated to reduction of se-
tion mechanism was comforted by elevated levels of repro- men volume and an increased semen pH in pesticide applica-
ductive hormones (FSH levels and intratesticular testoster- tors [16, 38].
one concentrations) in the exposed rats [26]. The spermato-
genic effects of DBCP are usually irreversible, though sup- Testosterone levels were also suppressed in rats by the
pression of intratesticular testosterone and serum FSH lev- insecticide Malathion (60 days treatment; 50 - 250
els with the GnRH agonist Lupron could induce increment mg/kg/bwt/day) [39]. Parathion is metabolised into p-
Effect of Environmental Contaminants on Mammalian Testis Current Molecular Pharmacology, 2014, Vol. 7, No. 2 3

Nitrophenol (PNP) [40], which is an endocrine disruptor mRNA levels of StAR protein and P450-17
in the testes of
commonly found in water and soil, and capable of estrogenic 6-week old male offspring [55]. Decreased StAR and serum
and anti-androgenic activities [41, 42]. PNP (10 mg/kg/day testosterone level were also observed in rats following oral
for 4 weeks) inhibits expression of testicular estrogen recep- administration of deltamethrin for 4 weeks (5 mg/kg bwt/day).
tor
(ER
) and androgen receptor (AR), reduced ER ex- Moreover, deltamethrin increased testicular total oxidant
pression, and induced Leydig cells hyperplasia in immature capacity (TOC), poly (ADP-ribose) polymerase (PARP),
male rats [42]. lactate dehydrogenase (LDH) and DNA damage, illustrating
significant peroxidation and cellular disruptive action of the
Pesticides accumulate in testes, and this may potentiate
pesticide. These adverse effects of the pesticide were how-
their deleterious effect on the organ. Examples of such pesti-
ever alleviated by the co-administration of a carotenoid de-
cides include the organochlorine compound Lindane, which
accumulates in rat testes, where it causes degenerative rivative antioxidant, lycopene [56-58].
changes in germinal epithelium. Lindane induced Sertoli cell A benzimidazole carbamate, carbendazim (methyl-2-
fragmentation and impaired sperm parameters (decreased benzimidazole), is a fungicide used in agriculture. Carben-
sperm counts and increased sperm abnormalities) [43-45], dazin decreased testis weight, reduced Leydig cell activities
and this was partly attributed to its ability to disrupt andro- of the antioxidant enzymes (SOD, CAT, GPx) and non-
gen synthesis in Leydig cells [44]. Disruption of Leydig cell enzymatic antioxidants (GSH, vitamins E, C and A), when
steroidogenesis was also reported following exposure to orally administered to male rats at 25 mg/kg bwt/day for 48
methoxychlor. Methoxychlor inhibited cholesterol side-chain days [59]. A decrease in the activities of testicular SOD and
cleavage enzyme (cytochrome P450scc) in the cells, reduced CAT was also observed, together with increased lipid per-
steroidogenic acute regulatory (StAR) protein expression and oxidation levels in the rats when orally administered at a
decreased 3-hydroxysteroid dehydrogenase (3-HSD) and single dose of the pesticide (200 mg/kg bwt) [60], and results
17-HSD activities in rat testes [46, 47]. Methoxychlor is corroborate the endocrine-disrupting effect of the fungicide.
metabolised into 2,2-bis-(p-hydroxyphenyl)-1,1,1-trichloroe- Indeed, carbendazim decreased serum testosterone and estra-
thane (HPTE), which also reduced testosterone production in diol in male rats, through reduction of the activities of steroi-
rat Leydig cells, through inhibition of several steroidogenic dogenic enzymes (3-HSD, 17-HSD) in Leydig cells [59],
enzymes (3-HSD, 17-HSD type 3, 11-hydroxysteroid as well as reduction of StAR protein and ABP expressions
dehydrogenase, and 17
-hydroxylase/17,20-lyase (CYP17A1)) [60]. The fungicide maneb also prevents release of testoster-
[48-50]. Interestingly, HPTE –induced inhibition of CYP17A1 one precursor (pregnenolone) in Leydig cells of rats after 9 -
was more efficient in human testicular microsomes, com- 18 days exposure (4 mg/kg), leading to the reduction of
pared to those of rats (with the IC50 values of 1.13±0.10 and plasma testosterone and decreased fertility in the animals
6.87±0.13 μM, for human and rat tests microsomes, respec- [61, 62a]. These results indicate that maneb might contribute
tively); suggesting a high sensitivity of humans to the to impairment of testicular Leydig cell steroidogenesis in
chemical compared to rats [50]. male farmers in Djutitsa (West Cameroon). Indeed, the
farmers were using maneb most frequently, and showed im-
Synthetic pyrethroid pesticides also induced serious balances of steroid hormones (increased serum andros-
damage on mammalian testis. For instance, exposure of male tenedione and decreased testosterone levels in blood), which
mice to fenvalerate during lactation (60 mg/kg bwt from are mainly produced by testes [62b].
postnatal day (PND) 0 to PND 21) decreased the weights of
Carbendazim increased androgen receptor (AR) concentra-
testes, induced apoptosis and spermatogenic failure in the
tions in rat testis, and replaced binding of [3H]-5 alpha-
animals. Fenvalerate exposure resulted in decreased mRNA
dihydrotestosterone to AR [63]. The insecticides carbaryl in-
and protein levels of testicular cytochrome P450scc, leading
duced accumulation of cellular mass in the lumen of
to reduction in serum and testicular testosterone levels [51].
seminiferous tubules and loss of sperms in albino rats [64].
This suggests impairment of spermatogenesis, and the latter Shalaby et al. [65] also reported alteration of spermatogenesis
pesticide has even been associated with poor semen quality (reduction in sperm count and sperm motility) in rats by the
in human beings who were exposed to 21.55 x 10-4 mg fen- insecticide methomyl, which consequently led to decreased
valerate /m3 ambient air at the work place [52]. Administra- fertility index in the exposed animals. Likewise, Carbofuran
tion of the insecticide cypermethrin to mature male mice (10 induced Sertoli and germ cells alterations, testicular edema
and 20 mg/kg bwt/day, 35 days) induced decrease in testicu- and accumulation of cellular debris in the lumen of the
lar weight and altered sperm development in seminiferous seminiferous tubules, leading to decreased sperm count and
tubules. Cypermethrin inhibited serum testosterone concen- morphology [66]. Oral administration of Mancozeb to Wistar
tration through alteration of steroidogenic Leydig cells (de- albino rats (500 mg/kg for 30 days) resulted in the reduction of
generation of mitochondria and the smooth endoplasmic testicular sperm counts and decrease of the weight of testis
reticulum of Leydig cells) [53]. Cypermethrin exposure (25 [67]. Oyewopo et al. [68] also reported a decreased testicular
mg/kg bwt/day by gavage, from PND35 - PND70) also re- weight and volume, as well as induction of testicular oxidative
sulted in inhibition of mRNA level of testicular StAR and stress in adult male rats exposed to Propoxur (2-isopropoxy-
decrease of testosterone concentrations in serum of male phenyl-N-methylcarbamate; 10 mg/kg bwt per day) orally for
mice [54]. Bifenthrin (2-methylbiphenyl-3-ylmethyl-(Z)- 30 days. This contrasted with observations of Ngoula et al.,
(1RS)-cis-3-(2-chloro-3,3,3-trifluoroprop-1-enyl)-2,2-dime- [69], who reported increase of testicular weight following ex-
thylcyclopropane carboxylate) is one of the most used syn- posure of rats to Propoxur (5.2 mg/kg bwt) for 90 successive
thetic pyrethroids for field and household pest control [51]. days. However, the pesticide induced modulation of testicular
Administration of 15 mg/kg of Bifethrin for 3 weeks to fe- weight, leading to impairment of its function, as evidenced by
male mice during pregnancy significantly reduced the spermatogenesis impairment in the 2 studies [68, 69].
4 Current Molecular Pharmacology, 2014, Vol. 7, No. 2 Manfo et al.

Administration of atrazine (a tirazine pesticide) to rats for with occurrence of testicular dysfunction and infertility in
60 days (60 and 120 mg/kg twice a week), induced degenera- living organisms, including humans [92-94].
tive changes in seminiferous tubules in the testes [70], and
It is well known that cadmium is a major environmental
this corroborate with decreased sperm quality observed in
toxicant that causes adverse effects on male reproductive
exposed rats [70] and male human workers [71]. Atrazine
organs [6, 89, 91]. For assessment its action mechanism on
(25 and 50 μg/mL) induced cell cytotoxicity and decreased
testicular toxicity, adult male rats were administered single
the levels of mRNA expression of StAR, and 3-HSD in rat
dose of cadmium chloride [1 - 1.2 mg ⁄ kg bwt; intraperito-
Leydig cells [72]. Atrazine also increased cAMP levels in rat
Leydig cells [73]. The carbamate fungicide Benomyl caused neal injection (i.p)], and followed up for 7 or 56 days. The
occlusion of the efferent ducts, preventing the passage of the lower dose caused slight alterations, while the higher dose
sperm from testis into the epididymis [74]. caused significant reduction in testis weight, gonadosomatic
index and length and diameter of seminiferous tubule after
Dichlorophenoxyacetic acid induced germ cells and 56 days [91]. The tubule lumens were filled with degener-
sperm head abnormalities when administered to male rats ated germ cells and multinucleated spermatid aggregates
(3.3 mg/kg/day, oral) for 3 and 5 consecutive days [75]. The after 7 days. Vacuolization of the seminiferous epithelium
organotin pesticides fentin hydroxide and fenbutatin oxide was also observed, and became more pronounced after 56
(25 g/kg bwt, i.p) also induced impairment of sperm cells days, with vacuolated seminiferous tubule consisting only of
and function in mice, at least in part, through inhibition of Sertoli cells [91]. Cadmium induced degeneration in testes
testosterone synthesis (inhibition of testicular steroidogenic and seminiferous tubules, and decreased testicular weights of
enzyme activities (3-HSD and 17-HSD)), which is re- mice/rats [94, 95]. A cDNA microarray and quantitative real-
quired for normal spermatogenesis [76]. time RT-PCR analysis in the testes of CD-1 mice injected
Alteration of testicular cells by pesticides may be through with CdCl2 (5 micromol/kg, subcutaneous injection) revealed
induction of free radicals production in testes and/or inhibition alteration in the expression of some genes that are likely to
of the antioxidant enzymes SOD, CAT, GPx, glutathione re- be important in toxicity responses. Expression of cell cycle-
ductase (GR), and glutathione S-transferase (GST) [77-79]. regulated cyclin B1 and CDC2 proteins was observed, sug-
Examples include methoxychlor, which was introduced as a gesting G2/M arrest and disturbance of meiosis in the testis
replacement for the banned pesticide, DDT [50]. A glypho- following cadmium exposure [96]. Cadmium also induced
sate-based herbicide Roundup (36 ppm) decreased glutathione disruption of cell junctions in the testis, including Sertoli cell
and increased Thiobarbituric acid reactive substances blood-testis barrier (BTB), through acceleration of endocyto-
(TBARS) levels in pubertal rat testis, leading to Sertoli cell sis of BTB components occludin and N-cadherin [97]. Per-
death [80]. Roundup also damaged Leydig cells and decreased turbation of BTB disrupts the microenvironment needed in
testosterone release in the animals after 24 h exposure [81]. seminiferous tubules for normal spermatogenesis [Box 1].
Administration of methoxichlor to male rats induced an in- Lower doses of Cd (
5 mol/kg) inhibit spermiation [98], and
creased generation of hydrogen peroxide, and reduced the induced spermatogenic cells apoptosis [99, 100]. Cadmium
activity of testicular SOD, CAT, GR, and GPx. In addition to exposure (0.4 mg/kg) also caused lipid peroxidation and al-
oxidative stress, methoxychlor decreased testes weight in the tered the antioxidant defense system (decreased activity of
rats [82]. The reactive oxygen species (ROS) -mediated effect SOD, CAT, GSH-Px, GR and GST) in rat testes. However,
of environmental toxicants is most likely to be detrimental to these defects on the antioxidant system were reversed by the
sperm cells, which are particularly vulnerable. Sperm cells antioxidants coenzyme Q and vitamin E [93].
contain high levels of polyunsaturated fatty acids that may be
subjected to peroxidation by ROS (lipid peroixdation). Alteration of testicular steroidogenesis by Cd was also
Moreover, sperm cells have a limited store of antioxidant reported, as well as imbalances in the levels of gonadotropins
molecules/enzymes, which could protect against oxidative that regulate spermatogenesis. In utero exposure of male mice
stress [79, 83]. foetuses to Cd (0.5 mg/kg/day, i.p, from gestational day (GD)
-13 to GD-17) down regulated expression of steroidogenic
Prochoraz is an immidazole fungicide, which inhibits enzymes/proteins (StAR protein, cytochrome P450scc and
aromatase, and act as an estrogen and androgen receptor an- 17-HSD) in testes, leading to decreased serum testosterone
tagonist in vitro [84]. The fungicide Vinclozolin acts as anti- levels in the animals [94]. A decreased serum testosterone
androgen by inhibiting androgen receptor (AR) activity [85], levels was also observed on male rats and rabbits exposed to
and induced differential gene expression in developing F1 cadmium [101, 102]. Moreover, the cadmium -exposed rats
generation rat testis [86]. showed increased FSH level, and reduction of LH levels [101].

III. HEAVY METALS The use of uranium in industrial or military activities has
led to its spread in the environment, raising concerns about
Heavy metals are elements commonly used in industry its toxicity. Histopathological examination of the testes in
and generally toxic to animals and to aerobic and anaerobic rats exposed to uranium revealed interstitial alterations, such
processes. Heavy metals include cadmium, chromium, cop- as focal atrophy, and binucleated cells [103]. Assessment of
per, mercury, lead, among others [87-89]. Heavy metals can its reproductive toxicity was conducted using human testes,
occur naturally in the earth crust at low levels, but mainly obtained from 7-12 weeks of gestation foetuses. Incubation
from industrial extraction [89, 90]. The general population is of the foetal testes with 0.1 mM uranyl acetate leads to re-
exposed to heavy metals via contaminants found in air, duced germ cells number, and the effect was more pro-
drinking water, food, and during occupational exposition nounced when compared to mouse germ cells treated in the
[91, 92]. Several studies have correlated their concentrations same experimental conditions [104]. This indicated that ad-
Effect of Environmental Contaminants on Mammalian Testis Current Molecular Pharmacology, 2014, Vol. 7, No. 2 5

verse effects of the heavy metals may impair the develop- to the cytoplasm of Sertoli cells, suggesting alteration of BTB
ment of human testis and suggest extrapolation of results dynamics.
from mice/rodents to humans.
IV. PHTHALATES
Mercury has been recognized as an environmental pollut-
ant that adversely affects male reproductive systems of ani- Diesters of 1,2-benzenedicarboxylic acid, or phthalates,
mals [92]. Mercuric chloride is one of the most toxic forms have been highly produced to be mainly used in a large range
of mercury because it easily forms organomercury com- of everyday life products, such as construction materials,
plexes with proteins [105]. When administered to sexually adhesives, paints, inks, cosmetics, toys, food packaging,
mature male rats, mercuric chloride (1mg/kg bwt) impaired clothes, medical materials, and drugs [115]. Phthalates also
the antioxidant system (i.e., increase in the TBARS level and serve as antifoaming agent in paper production, as an emulsi-
a decrease in the SOD, CAT, GPx activities) and histopa- fier for cosmetics, in perfumes and pesticides [116]. Some of
thological changes on the testes. Mercuric chloride induced these compounds, such as Mono-n- pentyl phthalate (MPP),
increased testicular absolute and relative weight in rats [105]. mono-2-ethylhexyl phthalate (MEPH), dibutyl phthalate
The histopathological changes induced by mercuric chloride (DBP), diethylhexyl phthalate (DEHP) and diisononyl phtha-
were also observed in mice testes following subchronic ad- late (DiNP), have been shown to induce reproductive dys-
ministration of the chemical (1-2!mg/kg/day, oral, for 30-60 function [115, 117-119]. Several studies have reported the in
days), and this was concomitant with alteration of sperm utero deleterious effect of phthalates on testicular function in
parameters and fertility, as well as inhibition of testicular mammals.
testosterone levels at the work place [106, 107]. Alteration of When pregnant Wistar rats were exposed to DBP (60
testicular function was reported in a 25-year-old infertile mg/kg bwt/day) from GD-7 to GD-20/21, examination of
(severe oligoasthenoteratospermia) man exposed to mercury testes in male offspring at GD-19 or GD-20/21 revealed
for 5 years. The man had high (considerably above levels of Leydig cell hyperplasia, Sertoli cell hyperplasia and presence
unexposed controls) mercury concentrations in hair, blood, of multinuclear gonocytes. The exposed rats also showed
and urine samples, and mercury grains in the interstitial Ley- reduction in expression of steroigogenic enzymes (cyto-
dig cells. Also, testicular biopsies showed interstitial lym- chrome P450scc and StAR proteins) in Leydig cells. Down-
phatic infiltration, Sertoli-cell-only syndrome and tubular regulation of StAR and cytochrome P450scc mRNA expres-
atrophy [108]. However, the histopathological changes and sion was also induced by DBP in fetal Leydig cells [120],
oxidative stress induced in rodents by mercury were allevi- and the alteration of steroidogenic proteins/enzymes cor-
ated when the animals were co-treated with the antioxidants roborated reduction in testicular testosterone production
sodium selenite and/or vitamin E [92]. Da Silva et al. [109] [115] and insl3 gene expression in the foetal testis [121].
also illustrated protective effect of Solanum sessiliflorum Moreover, DEHP induced severe spermatogenic disturbance
Dunal fruit (a source of antioxidants such as zinc and citric in mice [122]. Exposure of female rats to DBP (500 mg/kg)
acid) on testicular toxicity (decreased testis weights, reduced at GD-11/21 induced cryptorchidism and focal dysgenesis of
sperm production, and increased histopathological abnor- testicular somatic (Sertoli cells, Leydig cells) and germ
malities) induced by methylmercury in Wistar rats. (gonocytes) cells in male foetuses [123], and suppressed
Lead, aluminium and chromium also affect testicular cells. (approximately 90%) testicular testosterone levels [124].
When male rats were exposed to lead, a degeneration of sper- Suppression of ABP by Sertoli cells by Di-n-pentyl phthalate
matogenic and Leydig cells was noticed, as well as testicular (DPP) was also observed in immature rats [125]. DPP sup-
atrophy [110, 111]. Further histological examination of the pressed secretion of seminiferous tubule fluid (STF) by Ser-
testes revealed perturbation of spermatogenesis, characterized toli cells, while DEHP induced seminiferous tubular atrophy
by atrophy of seminiferous tubules, decrease in a number of in rats [125]. MPP and MEPH could penetrate BTB, and this
spermatids and spermatozoa, and alteration of Sertoli cells suggests that phthalate esters may act initially to cause injury
[110, 112]. Administration of lead (10 mg and 15 mg /kg in Sertoli cells, that will subsequently induce germ cells loss
bwt/day for 20 days) to adult male rats induced a decrease of [125]. In rat foetal Leydig cells, MEPH inhibited steroi-
serum testosterone levels with deformities in testicular mor- dogenesis by blocking CYP17A1 activity, which converts
phology, including gross damage within seminiferous tubules 17
-hydroxyprogesterone (17
-OHP) to androstenedione
[113]. A decrease in serum testosterone levels was also ob- [126].
served after exposure of animals to lead acetate, with con- Detachment of germ cells from the seminiferous epithe-
comitant reduction in sperm concentration [110]. Aluminium lium following phthalate exposure was reported in vivo [11,
chloride given orally to male rats at 100 and 200 mg/kg bwt 127]. Mono-2-ethylhexyl phthalate produced specific altera-
daily for 48 days also altered plasma membrane enzymes tions of gene expression profile in the testes [128], detach-
(adenosine triphosphatases, alkaline phosphatase, and gamma ment and sloughing of germ cells, and alteration of cyto-
glutamyl transferase) in testes, which may eventually have an plasmatic intermediate filament vimentin in Sertoli cells
impact on the fertility of the animals [114]. By using the Du- [129] in rodents (mice or rats). This well-known endocrine
rand's validated seminiferous tubule culture model, which disruptor has been shown to reduce germ cells number by
mimics the in vivo situation, Carette et al. [88] recently re- increasing apoptosis without change in basal and LH-
ported that hexavalent chromium increase the number of germ stimulated testosterone production in human foetal testes in
cell cytogenetic abnormalities. Exposure to chromium also vitro [130]. Exposure of humans to phthalates may thus in-
increased the transepithelial resistance and strongly delocal- duce a deleterious effect on testicular steroidogenesis and
ized the gap junction protein connexin 43 from the membrane spermatogenesis (Table 1).
6 Current Molecular Pharmacology, 2014, Vol. 7, No. 2 Manfo et al.

Table 1. Adverse effects of selected environmental pollutants on testicular function in humans and rodents

Class Chemical Studied species Toxicity and testicular cell(s) affected Reference(s)

Proliferating type A spermatogonia, resulting in low sperm count;

Dibromo-chloropropane
Human, rat Hormone levels imbalances (elevation of intratesticular testosterone [21, 23, 26, 27]
(DBCP)
concentrations)

Dichlorodiphenoxy-
Rat, mouse Apoptosis of Sertoli cells [30]
dichloroethylene (DDE)

Hormone alteration
Dichlorvos Rat [34]
Decrease of testis weight

Chlorpyrifos Rat Reduction in Sperm count [36]

Lindane Rat Degeneration of germinal epithelium, Sertoli cell Apoptosis [43]

Cell apoptosis
Fenvalerate Mouse [51]
Decrease of organ weights
Pesticides
Inhibition of serum testosterone through alteration of testicular
Cypermethrin Rat /Leydig cells steroidogenesis; decrease in testicular weight and [53, 54]
alteration of sperm development in seminiferous tubules

Disruption of antioxidant system, decrease of Leydig cell steroi-

Carbendazim Rat [60]
dogenesis

Inhibition of steroid production by Leydig cells, leading to de-

Maneb Rat, rabbit [62a, 62b, 222]
creased sperm number and reduced fertility

Carbofuran Rat Sertoli and germ cells alterations [66]

4-Nitrophenol
Rat Hyperplasia of testicular Leydig cells [42]
(PNP)

Roundup Rat Disruption of antioxidant system and inhibition of testosterone synthesis [80, 81]

Methoxichlor Rat Disruption of the testicular antioxidant system [82]

Reduction testis weights

degeneration in testes

disruption of cell junctions [91, 93, 95,

Cadmium Rat
spermatogenic cells apoptosis 152]

Disruption of antioxidant system,

Heavy hormone levels and steroidogenesis
metals
Chromium Rat Alteration of BTB [88]

Oxidative stress and alteration of sperm parameters in rats; Severe

Mercury Rat, human oligoasthenoteratospermia, interstitial lymphatic infiltration in testis, [105, 108]
and Sertoli-cell-only syndrome in humans

Rat, mouse, and Testicular atrophy in rats, reduced germ cells number in mouse and
Uranium [103, 104]
human human fetal testis
Effect of Environmental Contaminants on Mammalian Testis Current Molecular Pharmacology, 2014, Vol. 7, No. 2 7

Table (1) contd….

Class Chemical Studied species Toxicity and testicular cell(s) affected Reference(s)

Mono-2-ethylhexyl

phthalate Rat Inhibition of CYP17A1 activity [126]

(MEPH)

hyperplasia of Leydig cells and Sertoli cells

inhibition of testicular testosterone production and Please

aromatase,include "Mouse"
Dibutyl phthalate
Rat [121, 125, 120]
(DBP) here
reduction in expression of cytochrome P450scc, StAR protein and
Phthalates
insl3 genes in testis or Leydig cells

Diethylhexyl phthalate
Oxidative stress in testis, and inhibition of spermatogenesis [122]
(DEHP)

Mono-2-ethylhexyl Rodents (mice,

Alterations of gene expression profile in rodent testes, and increase
rats), human [128-130]
phthalate of apoptosis in germ cells in human foetal testes in vitro
foetal testis

Zearalenone Rat Decreased serum testosterone [161]

Induction of Leydig cells hyperplasia, increased the plasma level of

Mycotoxins Patulin Rat testosterone levels, and disorganization of seminiferous tubule [162, 163]
epithelium

Tetrachlorodibenzo-p-dioxin
Dioxins Rat Inhibition of steroidogenesis and induction of oxidative stress [131, 138, 139]
(TCDD)

Alteration of Ca2+ homeostasis, inhibition of hCG-induced testos-

Nonylphenol (NP) Rat [140, 142]
Sulfactants terone release in Leydig cells

Perfluorododecanoic acid Rat Inhibtion of steroidogenesis and testosterone level [150, 151]

2-methoxyethanol Rat spermatocytes necrosis/degeneration [168-170]

Organic 2, 5-hexanedione
solvents Testicular atrophy, germ cell loss (detachment from the seminifer- [127, 171, 174,
(n-hexane and methyl n-butyl Rat
ous epithelium) 175]
ketone metabolite)

Decrease in sperm number and degeneration of germ cells in rats;

2,4,6-trinitrotoluene (TNT) Rat, human decreased serum testosterone levels and impairment of sperm pa- [165, 166]
rameters
Other xeno-
biotics Germ cell apoptosis (increased of Fas and FasL, Bax, caspase-9)
[12, 126, 183,
Bisphenol A Rat disruption of BTB, inhibition of steroidogenesis and decrease of 184, 188]
testosterone level

Legend: numbers are references as indicated in references section.

electrolysis), from a de-novo synthesis from thermal sources

V. DIOXINS
(waste incineration plants and incomplete combustion proc-
Dioxins are a mixture of 219 different polychlorinated esses in landfills, combustion plants, iron smelting, sinter
dibenzo-p-dioxins and furans. The most well-known and plants, non-ferrous smelting and recycling plants), and from
hazardous dioxin is tetrachlorodibenzo-p-dioxin (TCDD), an incomplete combustions processes involving chlorine [132-
endocrine-disrupting environmental pollutant [131]. Dioxins 135]. For example, TCDD is generated during incineration
can arise as unwanted by-products from the production or of medical wastes, chlorine bleaching of paper and manufac-
use of many organochlorine compounds (such as chlorine ture of pesticides. It is also generated in exhaust from cars
bleaching of cellulose in paper production and chlorine alkali and cigarette smoke [133, 136, 137]. The compound TCDD
8 Current Molecular Pharmacology, 2014, Vol. 7, No. 2 Manfo et al.

induced reproductive toxicity in males. TCDD decreased fluorooctanoate is rapidly absorbed and bioaccumulated in
activities of testicular steroidogenic enzymes (3-HSD and humans (half- life thought to be in the range of 2.3-3.8 years)
17-HSD) when administered to rats (100 ng/kg bwt/ day for [147, 155], and this may potentiate its testicular toxicity,
15 days) [131], leading to the reduction of serum testoster- regarding the aforementioned adverse effects in other mam-
one level [138]. TCDD increased oxidative stress and sup- malian species.
pressed the activities of testicular antioxidant enzymes [138,
139]. TCDD also modulated the gene expression levels of VII. MYCOTOXINS
arylhydrocarbon receptor (AhR) and its repressor AhRR in
Mycotoxins are toxic secondary metabolites produced by
Leydig cells in vitro [135].
toxigenic fungi species. Mycotoxins commonly contaminate
animal feed and human food commodities [156]. Examples
VI. SURFACTANTS
of mycotoxins that may be important in connection with
Nonylphenol (NP) is a widely distributed environment foods are aflatoxins, ochratoxin A, patulin, and trichothe-
contaminant and has been documented to disrupt testicular cenes [156-158]. The administration of mycotoxins (afla-
development and decrease male fertility [140]. It is a de- toxin, ochratoxins and zearalenone) to male albino rats at the
graded product of alkylphenol polyethoxylates (APEOs), doses of 0.5, 1.0 and 2.5 ppm in feed, respectively), for 6
which are non- ionic surfactants used as components of months significantly reduced testosterone, LH and FSH lev-
cleaners, detergents and emulsifiers [141]. Amongst possible els [159]. Ochratoxin A in particular is commonly found as
targets of NP are testicular Sertoli cells, which play a crucial contaminant in foods such as coffee, grapes, wine, grains,
role in supporting and nourishing sperm cells. Nonylphenol and mainly produced by various species of the Aspergillus
alters Ca2+ homeostasis, causing dramatic morphological genus and one species of Penicillium, Penicillium verrucos
changes and decreased viability of Sertoli cells, as well as [160]. Treatment of H295R cells with Ochratoxin A in-
activation of the endoplasmic reticulum -stress signal path- creased aromatase protein, and leads to enhanced production
way in the cells [140]. NP inhibited hCG-induced testoster- of estradiol [158]. The effect of this mycotoxin on aromatase
one release both in vitro (rat Leydig cells) and in vivo (in may comprise the testicular balance in steroid enzymes
rats) [142]. NP and octylphenol caused downregulation of thereby negatively affect male reproductive function.
testicular 3-HSD mRNA when orally administered to 4- Zearalenone is a non-steroidal mycotoxin produced by sev-
week-old Sprague-Dawley rats (10-250 mg/kg bwt/day) for 3 eral species of Fusarium fungi. Zearalenone is frequently
weeks [143]. Downregulation of testicular 3-HSD mRNA found in cereals used for human food stuffs and animal
was also observed in rats exposed to octylphenol (OP; 10- feeds, and capable of altering gene expression profile in the
250 mg/kg bwt/day for 3 weeks), which is another degrada- developing testes of mice [128]. Zearalenone decreased se-
tion product of APEOs [143]. OP decreased viability and rum testosterone in rats after 3 days (i.p, 20 mg/kg bwt). This
increased apoptosis of Sertoli cells, induced up-regulation of negative effect of Zearalenone was however alleviated by the
Bax, and down-regulated the expression of procaspase-3 and simultaneous administration of vitamin E to animals [161].
Bcl-2 [144]. NP was detected in 51% urine samples (concen- The mycotoxin Patulin produced in many species (including
tration
0.1 g/L) collected from humans [145], and may Aspergillus sp., Penicillium sp., Paecilomyces sp. and Bys-
constitute a potential triggering factor for testicular dycfunc- sochlamys sp.) is found in a variety of food commodities. It
tion and infertility in men. increased the plasma level of testosterone, induced hyperpla-
sia of Leydig cells and disorganization of seminiferous tu-
Perfluoroalkyl acids, including perfluorododecanoic acid,
bule epithelium [162, 163].
are used as surfactants in industrial and consumer products,
and have been detected in environmental matrices, in wild-
VIII. OTHER XENOBIOTICS
life and in human samples (blood, tissues) in various coun-
tries [146, 147]. Perfluorooctanoate has been used in the The compound 2,4,6-trinitrotoluene (TNT) is a com-
production of various fluoropolymers, and is also an end monly used explosive. Several studies in both humans and
product of many fluorochemical compounds in the environ- animals have showed its ability to induce testicular toxicity.
ment [148, 149]. Chronic administration of Perfluorodode- Toxic effects following administration of TNT to Fischer
canoic acid to male rats (0.5mg/kg/day, 110 days) resulted in 344 male rats (125 and 300 mg /kg/day for 13 weeks in the
decreased protein levels of StAR, cytochrome P450scc, diet) included testicular atrophy with degeneration of the
along with reduced mRNA levels of insulin-like growth fac- seminiferous tubular epithelium [164]. In another study,
tor I (IGF-I), IGF-I receptor, and interleukin 1 in rat testes TNT administration to rats induced germ cell degeneration
[150]. Oral administration of ammonium perfluorooctanoate and disappearance of spermatozoa in seminiferous tubules,
(1.0-5.0 mg/kg bwt/day) to mice for 6 weeks resulted in re- resulting in a significant decrease in sperm number in testis.
duced plasma testosterone concentrations in the animals. TNT also increased the formation of 8-oxo-7,8-dihydro-2'-
Moreover, perfluorooctanoate decreased testosterone levels deoxyguanosine in sperm without affecting plasma testoster-
in rat testis [151], and the decrease was associated with re- one levels in the animals; suggesting a direct action of the
duced expression of mitochondrial cytochrome P450scc and toxicant on sperm cells [165]. Reproductive toxicity of TNT
StAR protein, which are all involved in the steroidogenesis was assessed in male humans occupationally exposed to the
process [152]. Inhibition of 3-HSD and 17-HSD activities chemical in China. The TNT concentrations in the workplace
and induction of cell adenomas in rat Leydig cells exposed to air exceeded maximal allowable concentration (i.e. > 1
perfluorooctanoate were also reported [153, 154]. Per- mg/m3) [166, 167]. The exposed male workers showed sig-
Effect of Environmental Contaminants on Mammalian Testis Current Molecular Pharmacology, 2014, Vol. 7, No. 2 9

nificant sexual disorders (impotence, loss of libido and hy- male infertility [12, 179, 180]. BPA showed a mitogenic
poesthesia), with increased sperm malformation, reduced effect on Leydig cells in the prepubertal period in rats, lead-
semen volume and percentage of motile spermatozoa. The ing to increased Leydig cell numbers in the testes of adult
serum testosterone content in the workers was also signifi- males [180, 181]. BPA induced germ cell apoptosis through
cantly decreased [166]. TNT exposure induced a decrease of the Fas/FasL signalling pathway and the mitochondrial apop-
Cu, Zn, Na, Mg, and Se concentrations in the semen of the totic pathway [181]. BPA exposure also evoked the activa-
workers. Moreover, TNT exposure extended the liquefying tion of caspase-8 and caspase-3 in testes, induced cyto-
time of semen, increased the incidence of sperm malforma- chrome c translocation from mitochondria into cytosol, and
tion, and reduced the percentage of motile spermatozoa upregulated the level of testicular Bax and active caspase-9
[167]. [181]. Prenatal or neonatal exposure to BPA disrupts BTB in
mice, through down-regulation of genes associated with Ser-
Organic solvents are also implicated in testicular dys-
toli cell function (Msi1h, Ncoa1, Nid1, Hspb2, and Gata6),
function. Histological alterations were observed in testes of
and modulation in the expression of BTB proteins (increases
rats following administration of the glycol ether solvent 2-
in the expression of N-cadherin and Zona Occludin-1, and
methoxyethanol (150 mg/kg bwt/day for 3 days). The in-
reduction in the expression of Connexin 43) [182, 183].
duced histopathological changes consisted of degeneration or
necrosis of spermatocytes [168]. Spermatocytes degeneration Considering the importance of BTB for normal spermato-
was also observed in rats treated with a single oral dose of 2- genesis [Box 1], the BTB destruction may explain the de-
cline in total sperm counts in rodents following BPA expo-
methoxyethanol [169, 170]. The compound 2,5-hexanedione
sure [182-184].
is a toxic metabolite of industrial solvents such as n-hexane
and methyl n-butyl ketone [171], and has been shown to de- Acrylonitrile (CH‚CH–CN, ACN) have been widely used
crease the transport rate of the luminal oil droplets in in various synthetic chemical industries, including produc-
seminiferous tubules of rats [172]. This suggests the ability tion of plastics. When acrylonitrile was administered daily to
of 2,5-hexanedione to reduce production of STF in Sertoli male albino rats at a dose of 30 mg/kg bwt for 5 days, a re-
cells, a key factor in maintenance of spermatogenesis and duction of glutathione content and GST activity was ob-
transport sperm from the seminiferous tubule through the served in the testes of the animals. Acrylonitrile also induced
rete testis [125, 173]. Exposure to 2, 5-hexanedione (1%, in maturation arrest, depletion and abnormalities (altered cyto-
drinking water) for 3-5 weeks resulted in testicular atrophy, plasmic organelles, and deformed spermatids) in spermato-
with decreased seminiferous tubule fluid secretion and de- genic cells. Moreover, testosterone –producing Leydig cells
cline in germ cell number [174, 175]. exhibited mitochondrial disruption and reduced amount of
smooth endoplasmic reticulum [185], and this corroborated
Bisphenol A (BPA) is a well known environmental es- the reduction of testosterone concentrations in sera of young
trogen that is most commonly used to produce polycarbonate and adult male humans exposed to acrylonitrile (vinylcyanid)
plastic [12, 79]. BPA has been one of the highest volume in a chemical factory [186].
chemicals produced worldwide, with over 6 billion pounds
produced each year and over 100 tons released into the at- Massive use of Brominated flame retardants (BFRs) in
mosphere by yearly production [176]. BPA can leach into order to reduce the risk of fire accidents, have led to its oc-
drinking water from polycarbonate bottles including poly- currence in almost all the environmental compartments, in-
carbonate baby bottles and reusable water bottles. BPA have cluding humans and wildlife organisms. BFRs are endocrine
been detected in serum and urine samples from humans. A disrupting chemicals, and affect mammalian testicular func-
study conducted on a reference human population of the tion [187]. These include Hexabromocyclododecane, 1,2,5,6-
tetrabromocyclooctane and bis-(2-ethylhexyl) tetrabro-
United States revealed that BPA concentration was
0.1
mophthalate. Hexabromocyclododecane induced a decreased
g/L in
95% urine samples collected from the participants
weight of the testis in F1 male pups in a one-generation re-
[145]. BPA was also detected in serum samples from men,
production assay in Wistar rats [188]. Hexabromocyclodode-
with higher concentrations and greater detection rates re-
cane inhibited steroidogenesis in peripubertal rat Leydig cell
corded in males occupationally exposed than those not ex-
cultures [189]. The chemical 1,2,5,6-tetrabromocyclooctane
posed to the chemical. Interestingly, BPA concentrations
modulated CYP17A1, leading to stimulation of testosterone
were associated with low androstenedione and high sex hor-
and estradiol production in primary porcine testicular cells
mone-binding globuline levels [177], suggesting impairment
[190].
of testicular function. The BPA-induced reproductive dys-
function in experimental animals is well documented [12, 13, Environmental toxicants induced impairment on repro-
178]. When administered to rats by oral route at doses of ductive function in males in general, and particularly on
0.005 - 500 g/kg bwt/day for 45 days, BPA dose- mammalian testis, and this is illustrated by correlations be-
dependently decreased the activities of testicular antioxidant tween pollution level and defects in semen samples. For ex-
enzymes, steroidogenic enzymes (3--HSD, and 17--HSD), ample, a study conducted in Chongqing, China, revealed that
and reduced StAR expression; leading to decrease in testos- air pollution level was more pronounced in urban area com-
terone secretion by Leydig cells [12]. BPA also decreased pared to rural area, and was associated with male semen
LH receptor (LHCGR) in Leydig cells. Moreover, investiga- quality. Concentrations of particulate matter, sulphur diox-
tions on BPA revealed impairment of insulin signalling and ide, and nitrous dioxide were negatively associated with
glucose transport in testes of exposed rats, and this dysfunc- normal sperm morphology percentage and sperm kinetic
tion could lead to impairment of testicular functions and parameters [96]. These observations were in line with the
10 Current Molecular Pharmacology, 2014, Vol. 7, No. 2 Manfo et al.

earlier report on the evidence that sulphur dioxide, and ni- plants [201, 204-206]. Because of rapid increase in the num-
trous dioxide are toxins for the reproductive system of ber of automobiles and industrial production, metabolites of
mammals [191]. The alteration of sperm parameters may PAHs were detected in 100% of test candidates in a recent
also occur indirectly through disruption of hormonal regula- study in China, and higher levels were associated with male
tion of the testicular function. infertility [207]. It is also well established from both human
Polychlorinated biphenyls (PCBs) are either manufactured and animal studies that exogenous administration of oestro-
directly by chlorination of biphenyls, or generated during gens results in increased incidence of male reproductive dis-
combustion processes (e.g., from waste incineration plants, orders, such as cryptorchidism [208]. The effect of radionu-
fossil fuel burning). Manufactured PCBs were used as lubri- cleoides and temperature on testicular function was also re-
cants, impregnating agents for wood and paper, flame protec- ported. A transient elevation in serum FSH and decreased
tive substances, in transformers and condensers, as carrier inhibin B levels was observed in adult men after exposure to
substances for insecticides, etc. [192, 193]. Although banned radioiodine for thyroid cancer treatment. This reflected se-
for several decades, PCBs continue to exist in the environment vere impairment of spermatogenesis by the radionucleoide
because of their long half-life, continued presence in items [209, 210]. Although the FSH levels normalized within 9-18
produced before the ban, and poor disposal practices [194]. months from the last radioiodine administration, this result
PCBs are implicated in testicular dysfunction. The PCB Aro- indicate the potential adverse effect of radonucleoides in
clor 1254 decreased testicular weights and reproductive suc- spermatogenesis (or HPT axis [Box 1]) in the exposed pa-
cess of male pups in adulthood when given to male rats tients, and particularly those requiring multiple administra-
through lactation (8, 32, or 64 mg/kg to dams) [195, 196]. tions of radionucleoides for persistent cancer treatment [209,
Treatment of Leydig cells with the PCB Aroclor 1254, re- 210]. Amara et al. [211] also exposed rats to static magnetic
sulted in a significant reduction of serum testosterone and es- field for 128 mT intensity, 1 h/d for 30 days, and observed a
tradiol. In addition, the Leydig cell surface LH receptors, ac- reduction in testosterone concentration with increased oxida-
tivities of the steroidogenic enzymes such as cytochrome tive stress in the animals [199].
P450scc, 3-HSD, 17-HSD, antioxidant enzymes SOD,
CAT, GPX, GR, -GT, GST and non-enzymatic antioxidants
such as vitamin C and E were significantly diminished, Box 1: Sertoli cells in spermatogenesis
whereas LPO and ROS were markedly elevated. However, the The overall objective of the reproductive process for the
simultaneous administration of vitamin C and E in Aroclor adult male is the production of gametes capable of fertilization and
1254 exposed rats resulted in a significant restoration of all the the production of viable offspring [212]. The male gamete results
above-mentioned parameters to the control level [197]. from spermatogenesis, an elaborate process of germ cell prolifera-
tion and differentiation that leads to the production and release of
A nitroaromatic intermediate 1,3-Dinitrobenzene, used in
spermatozoa from the testis [213]. Spermatogenesis involves mito-
manufacture of dyes, plastics and explosives, induced tes-
sis, meiosis, and cellular differentiation that transform spermatogo-
ticular atrophy in male rats [198, 199]. Daily administration nia into mature spermatozoa [77]. The testis consists of seminifer-
of 1,3-Dinitrobenzene (4mg/kg/day) to rats induced testicular ous tubules that comprise of Sertoli cells, and the interstitial com-
injury, particularly to Sertoli cells, and induced apoptosis in partment that comprises testosterone-producing Leydig cells [89,
the surrounding germinal cells. Plasma testosterone and tes- 212, 214, 215]. Testicular spermatogenesis and steroidogenesis
ticular steroidogenesis-related gene expression were affected processes are regulated by the HPT axis which involves the pitui-
in the animals, suggesting an interference with the endocrine tary gonadotropins LH and FSH. In addition to FSH and LH, a
system [200]. number of hormones and growth factors, including androgens, insu-
Environmental toxicants also include diesel exhaust par- lin-like growth factor, oxytocin, and transforming growth factor

ticles (DEPs) and polycyclic aromatic hydrocarbons (PAHs) and estrogens, are also proposed to regulate testicular function [124,
[201, 202]. DEPs are air pollutants resulting from increased 216-218]. In addition to hormonal stimulation, spermatogenesis
depends upon dynamic interactions between the Sertoli cells and the
use of vehicles, especially diesel engine- powered cars and
germ cells of the seminiferous epithelium [213]. Sertoli cells secrete
trucks. The DEPs may induce hazardous effects on human
nutritive factors into the adluminal compartment that creates a spe-
health, including endocrine disruption [202, 203]. Exposure
cialized microenvironment suitable for development and viability of
to nanoparticle-rich diesel exhaust increased serum and testis
germ cells [213]. Indeed, the BTB formed by tight junctions be-
testosterone levels in male rats [203], through increased tween adjacent Sertoli cells prevents large molecules from passing
StAR protein and cytochrome P450scc mRNA and their directly from the blood into the lumen of the seminiferous tubule
protein expressions in the testis of the exposed rats. Interest- and separates spermatogonia in the basal compartment from sper-
ingly, the exposed rats also showed significant increase in matocytes and spermatids in the adluminal compartment [11]. Ser-
growth hormone receptor expression and increased insulin- toli cells also form sites of attachment to germ cells that provide
like growth factor I-mRNA levels in the testis [202]. PAHs efficient paracrine signaling mechanisms between these cells as
are formed during incomplete combustion of organic matter well as physical support to developing germ cells [213, 219]. Sertoli
such as wood, carbon or mineral oil. An example of PAHs cells express androgen receptor and FSH receptor, to which testos-
source includes car exhaust particles, road runoff, and prod- terone FSH bind, respectively, to inhibit death signals sent to germ
ucts originating from incomplete combustion processes in cells [220, 221]. Exposure to Sertoli cell toxicants is likely to in-
urban landfills, forest and house fires, heat and energy power duce detachment of germ cells from the seminiferous epithelium
stations, vehicle traffic, waste incineration and industrial [11, 127].
Effect of Environmental Contaminants on Mammalian Testis Current Molecular Pharmacology, 2014, Vol. 7, No. 2 11

Box 1. contd…. rats and mice have been so far the optimal experimental sub-
The BTB comprises ectoplasmic specialization (ES) (in addi-
jects as mammalian species, probably due to their ease of
tion to tight junction (TJ), desmosome, and gap junction) which handling and high productivity in short term. Direct effect on
anchors developing spermatids in the seminiferous epithelium until human testis have been illustrated also by few studies. Ma-
they are fully developed, prior to release into the seminiferous jority of the toxicants reviewed affect the male reproductive
tubule lumen. Thus, disruption of the apical ES by xenobiotics system by direct action on testis. This action results in reduc-
causes the premature release of spermatids that are structurally tion testis weights in general, or decrease of seminiferous
defective and unable to fertilize the ovum [184]. Sertoli cells pro- epithelial supportive Sertoli cells and interstitial Leydig cells
duce STF, which is essential to maintain the appropriate nutritional (Fig. 1). The toxicants also induce reduction of sperm pa-
and hormonal environment for spermatogenesis and to transport rameters (counts, motility, viability and density, and increase
sperm from the seminiferous tubule through the rete testis and into in sperm DNA damage and deformity, among others), which
the epididymis. Changes in fluid production and lack of resorption represents a key factor for successful fecundation and fertil-
of fluid in the epididymis can create abnormal conditions in the ity in males. The available toxicological data on mammalian
seminiferous tubules, leading to disrupted spermatogenesis [125, testis are chiefly based on reduction of testicular size and
173]. Maintenance of Sertoli cell population is also a critical fac- alteration of sperm cells within the seminiferous epithelium.
tor. Several reports have elucidated the effect of various environ- Induction of oxidative stress is emphasized as another com-
mental toxicants on depletion of Sertoli cell population, alteration mon response after exposure to environmental toxicants, and
of their secretary function, induction of germ cells distachment several reports have suggested the alleviating effect of well-
from seminiferous epithelium (Sertoli cells), as well as impairment known antioxidant molecules. Moreover, the reviewed data
of the BTB [89, 184, 222]. indicated that xenobiotic –induced alteration of the testicular
function may occur from the in utero period throughout eld-
erly life. The environmental toxicants are usually present in
CONCLUSION
the environment as complex mixtures of chemicals that can
This review paper presents updated data on testicular interact to cause increased effects. The data hereby reviewed
toxicity in mammals. The reviewed literature revealed that are somehow silent on the effect of chemical cocktails on

Fig. (1). Cellular target of different environmental pollutants within mammalian testis. Actions of environmental toxicants include in-
duction of oxidative stress in the testis, inhibition of steroidogenesis in Leydig cells (LC), inhibition of Sertoli cells (SC) function, direct
inhibition of spermatogenesis, and alteration of sperm cells. PTC: peritubular cells, SPG: spermatogonia, SPT1: spermatocyte 1, SPT2:
spermatocyte 2, RS: round spermatid, ES: elongated spermatid, SPZ: spermatozoa.
12 Current Molecular Pharmacology, 2014, Vol. 7, No. 2 Manfo et al.

testicular function. The deleterious effects reported should [16] Yucra, S.; Gasco, M.; Rubio, J.; Gonzales, G.F. Semen quality in
thus be considered with stringency, in order to contribute Peruvian pesticide applicators: association between urinary or-
ganophosphate metabolites and semen parameters. Environ.
alleviating the burden of xenobiotics around us. This is a Health, 2008, 7, 59. doi: 10.1186/1476-069X-7-59.
prerequisite for preservation of the mammalian specie, as [17] Revitt, D.; Ellis, J.B.; Llewellyn, N,R. Herbicide behaviour in the
spermatogenesis represents the milestone in reproduction. runoff from an urban catchment. 8th ICUSD, pp 96-104, Sydney,
Australia, 1999.
[18] De Jager, C.; Farias, P.; Barraza-Villarreal, A.; Avila, M.H.; Ayotte,
CONFLICT OF INTEREST P.; Dewailly, E.; Dombrowski, C.; Rousseau, F.; Sanchez, V.D.; Bai-
ley, J.L. Reduced seminal parameters associated with environmental
The authors confirm that this article content has no con- DDT exposure and p,p'-DDE concentrations in men in Chiapas, Mex-
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Received: August 10, 2014 Revised: November 3, 2014 Accepted: December 19, 2014