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Review Article
 

A Review on the Biochemical and Molecular Mechanisms of Phthalate-Induced Toxicity in Various Organs with a Focus on the Reproductive System



Mohammad Hossein Asghari, Soodabeh Saeidnia and Mohammad Abdollahi
 
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ABSTRACT

Phthalates are a large group of chemicals, used in plasticizers and industrial solvents, to make them flexible and soluble, especially when these materials are applied in the production of toys, medical equipment and drugs coverings. It seems that phthalates induce multi-organ damage through a number of mechanisms such as oxidative stress via generation of Reactive Oxygen Species (ROS), DNA damage, lipid peroxidation, disrupting cell function and also altering the expression and activity of the most important antioxidant enzymes. In this study, we reviewed the recent publications that evaluated the contribution of oxidative stress in phthalate toxicity. Alteration of antioxidant enzymes such as a reduced SOD (Cu/Zn superoxide dismutase) activity as well as an increased CAT (catalase) function normally occur and can be observed particularly with higher doses of phthalates. Moreover, these compounds decrease GPX (glutathione peroxidase) and GST (glutathione S-transferase) activities. Nevertheless, controversy is found in the levels of cellular antioxidants like SOD showing a reduction in many organs like liver, kidney and reproductive system, whereas, its increase has been reported in a few studies. In summary, among various organs, reproductive system seems was affected further by oxidative stress through disruption of spermatogenesis, inducing mitochondrial dysfunction in gonocytes, impairment of cellular redox mechanism and increasing peroxiredoxin 3 and cycloxygenase 2 levels in spermatocytes. The phthalates are being replaced in some countries by other safe plasticizers.

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Mohammad Hossein Asghari, Soodabeh Saeidnia and Mohammad Abdollahi, 2015. A Review on the Biochemical and Molecular Mechanisms of Phthalate-Induced Toxicity in Various Organs with a Focus on the Reproductive System. International Journal of Pharmacology, 11: 95-105.

DOI: 10.3923/ijp.2015.95.105

URL: https://scialert.net/abstract/?doi=ijp.2015.95.105
 
Received: September 08, 2014; Accepted: December 12, 2014; Published: February 16, 2015



INTRODUCTION

Phthalates are one of the most important synthetic chemicals used in various materials such as children’s toys, drug coatings, cosmetics and solvents to give flexibility and solubility to the materials in which they are applied (Saeidnia and Abdollahi, 2013a; Wang et al., 2012a). More than 8.1 million tons of phthalates are released in the world annually (Crinnion, 2010). Phthalates-exposure may happen through oral, dermal and inhalation routes in human (Bahadar et al., 2014). It is also demonstrated that DEHP (Bis(2-ethylhexyl) phthalate) and MEHP (Mono-(2-ethylhexyl) phthalate) are found in the environment and cause adverse health effects in humans (Yang et al., 2012). Phthalates cause multi-organ damage through a number of mechanisms. Some phthalates can cause hepatic toxicity through transactivation of Peroxisome Proliferator-Activated Receptor (PPARs). This activation could cause uncontrolled cell proliferation leading to hepatic tumorogenesis consequences by the alteration of hepatic enzymes activities (Yavasoglu et al., 2014). A recent study showed that DEHP induce necrosis and inflammation in the liver.

Fig. 1:Phthalates induce toxicity via affecting the antioxidant enzymes activity and gene expression

Phthalate-induced oxidative stress results in decreasing the antioxidant capacity, especially in GPX (glutathione peroxidase) and GST (glutathione S-transferase). Moreover, increase in lipid peroxidation, CAT (Catalase) and SOD (Cu/Zn superoxide dismutase) activity were observed. Reactive Oxygen Species (ROS) can affect Peroxisome Proliferator (PP) leading to parenchymal cell proliferation. It is mentioned that Kupffer cells are a potential oxidant in rodent liver (Erkekoglu et al., 2014). Actually, phthalates induce toxicity not only via affecting the antioxidant enzymes activity but also through gene expression (Fig. 1).

In another study, Li et al. (2013) showed the immunotoxicity of DBP (Dibutyl Phthalate). The cytotoxicity in Peritoneal Exudate Macrophages (PEM) was observed. The PEM are used to express CD36 and CD80. Furthermore, DBP reduce phagocytosis on apoptotic cells. It also decreases cytokine production, immunogenicity and antigen presentation (Li et al., 2013).

In another study, neurotoxicity of phthalate was assessed. Impairment in thyroid homeostasis and activation of PPAR were reported as the most important mechanisms. Phthalates are able to change transcriptional activity of the sodium/iodine symporter leading to disruption of iodine uptake into the thyroid gland. It is shown that maternal hypothyroxinemia especially in a low level of free T4 is associated with belated cognitive and neuromotor development as well as reduced IQ. On the other hands, DEHP may overexpress PPAR-γ, resulting in apoptosis of undifferentiated neurons (Miodovnik et al., 2014).

The cardiotoxicity of phthalates has been also reported. A significant decrease in conduction velocity was noted 24 h after DEHP exposure. Moreover, human embryonic stem cell viability and cardiac differentiation were reduced following MEHP exposure (Posnack, 2014). In the kidney, DEHP exposure could decrease the number of nephrons in offspring at the weaning. Likewise, DEHP induce glomerulosclerosis and interstitial fibrosis in adulthood. The renin and angiotensin ll expression were decreased at birthday and enhanced at weaning (Wei et al., 2012).

The preliminary studies revealed that the reproductive system is more susceptible to phthalates rather than other organs (Martino Andrade and Chahoud, 2010). The mechanisms of reproductive toxicity have not been exactly elucidated so far. It is reported that these effects are correlated with anti-androgenic effect (Noriega et al., 2009) and activation of PPARα which can alter testosterone biosynthesis through inducing variation in the gene expression of the related enzymes. This results in down-regulation of nuclear receptors which are important in testis development (Gazouli et al., 2002; Ward et al., 1998), altering the expression and activity of antioxidant enzymes which may result in DNA damage by producing ROS (Wang et al., 2012b) and impairment in the function of Leydig and Sertoli cell (Martino Andrade and Chahoud, 2010).

In another study, Davis et al. (1994) showed that MEHP decreased the estradiol level in cultured rat granulosa cells via inhibition of aromatase activity. In the mentioned study, MEHP also decreased aromatase transcript and protein level. Literature revealed that MEHP can act through activation of PPAR-α and PPAR-γ to reduce the level of estradiol and related genes expression (Lovekamp-Swan and Davis, 2003). In the same way, activation of PPAR-γ disrupted the growth time and follicle differentiation (Lovekamp-Swan et al., 2003). Many of the toxic compounds can act through oxidative stress causing genotoxicity and various chronic diseases (Mostafalou and Abdollahi, 2012, 2013; Saeidnia and Abdollahi, 2013b).

One of the most important mechanisms for phthalates toxicity is defined regarding oxidative stress. The most common ROS are superoxide and H2O2 that can be converted into H2O and O2 by antioxidant enzymes such as SOD, GPX and CAT (Abdollahi et al., 2014). Phthalates are demonstrated to alter the expression and activity of these enzymes leading to disruption of the cell function (Wang et al., 2012a). Although there are several reports demonstrating the role of oxidative stress in this regard, the presence of a clear relationship between phthalate toxicity in some organs and oxidative stress as the main cause is still under question. Therefore, the main purpose of the present study is to systematically review the related reports and papers published in the valid and credible journals to confirm whether there is a correlation between phthalate toxicity and oxidative stress as a main mechanism of toxicity for these compounds.

APPROACH TO A SYSTEMATIC REVIEW

Electronic searches were performed on Science Finder, PubMed, Science Direct and Google Scholar. Key words included: Phthalate, oxidative stress, antioxidant enzymes, liver toxicity and reproductive toxicity. In vitro and in vivo animal and human studies have been evaluated and included in this review. Also, oxidative stress as one of the mechanisms for phthalate toxicity has been discussed. The results of those selected reports are categorized into four sections including histopathology, hormone analysis, oxidative stress parameters and gene expression.

HISTOPATHOLOGICAL ALTERATIONS

Reproductive toxicity: Recently, the reproductive toxicity of phthalate has been evaluated by Wang et al. (2012b). In that study the antral follicles isolated from CD-1 mice which were received DEHP and MEHP at doses: 1, 10 and 100 μg mL‾1 during 96 h. The results showed that the antral follicle growth decreased using all doses of DEHP and MEHP. In another study, Erkekoglu et al. (2010) examined the influence of DEHP and MEHP on MA-10 Leydig cell in mice. Resulted data showed that about 60-80% of the cells survived at high dose of DEHP but there is no cell survival at the dose 10 μM of MEHP.

Furthermore, Zhou et al. (2010) investigated the effects of DBP (dibutyl phthalate) in rats. Animals were given DBP at the doses of 100, 250 and 500 mg/kg/day orally during two weeks. Sperm count and motility significantly decreased at 250 and 500 mg kg‾1. Moreover, seminiferous tubules atrophy was observed in 500 mg kg‾1. In addition, decreased testis weight was observed in several reports (Kasahara et al., 2002; Shono and Taguchi, 2014). Moreover, Botelho et al. (2009) investigated the DEHP toxicity on the reproductive system. The results revealed that the number of multinucleated gonocytes was increased.

Liver toxicity: Seo et al. (2004) assessed the effects of DEHP (Bis(2-ethylhexyl) phthalate), DBP (dibutyl phthalate) and BBP (benzyl butyl phthalate) at the doses of 50, 200 and 1000 mg kg‾1 orally during 14 days in the rat. Hepatomegaly was observed by the DEHP administration at all doses while it happened at a dose of 1000 mg kg‾1 for BBP and DBP. Moreover, Yavasoglu et al. (2014) investigated the effects of BCP (butyl cyclohexyl phthalate) toxicity in mouse liver. The animals received BCP orally at three doses of 100, 200 and 400 mg kg‾1 for 20 days. The results showed congestions in vena centralis, an enlargement of the sinusoids, degeneration in hepatocytes, vacuole formations and presence of lipid droplets in hepatocytes in all groups (Yavasoglu et al., 2014). Additionally, Zhou et al. (2011) evaluated the effects of DBP at three doses of 100, 250 and 500 orally during 2 weeks. A decrease in epididymal weight, an atrophy of epididymal tubules and hyperemia of interstitial vascular were also noted.

Neurotoxicity: There are a few articles on neurotoxicity of phthalates through the oxidative stress. In a study, Tseng et al. (2013) evaluated the influence of DEHP (2 and 20 ppm), DBP (500 and 1000 ppm) and DIBP (100 and 1000 ppm) on nematode. The results showed that intracellular ROS was increased at all doses. In the recent study, Zuo et al. (2014) assessed the properties of DBP on antioxidant parameters. Mice were given DBP at doses of 0.45 and 45 mg kg‾1 orally during 32 days. A significant increase of ROS, GSH and MDA (malondialdehyde) level was observed in the brain.

SEXUAL HORMONE ALTERATIONS

Botelho et al. (2009) investigated the effects of DEHP on sexual hormones at a dose of 500 mg kg‾1 from gestational day (seventh) to lactation day (second) in rats. The results showed that DEHP had no effect on testosterone in offspring. In another study, Lee et al. (2007) assessed DBP exposure on rat. The animals received DBP in drinking water at a single dose of 750 mg/kg/day for 30 days. Statistical analysis identified a significant decrease in testosterone level but there was no effect on thyroid hormones.

CHANGING THE OXIDATIVE STRESS PARAMETERS

In a recent study, Wang et al. (2012a) investigated the effect of DEHP on the activity of antioxidant enzymes. At the beginning, SOD activity significantly increased during 24 h by DEHP (10 μg mL‾1) but its level decreased at 72 and 96 h. GPX and CAT activities exhibited no alteration. Furthermore, Wang et al. (2012b) assessed MEHP properties on antioxidant enzymes activities at 0.1, 1, 10 and 100 μg mL‾1. It was observed that GPX activity decreased at 1, 10 and 100 μg mL‾1 and also at 0.1 μg mL‾1 after 96 h. CAT activity increased at two doses of 0.1 and 1 μg mL‾1. Moreover, SOD activity decreased at a single dose of 100 μg mL‾1, whereas, SOD activity increased in a single dose of 10 μg mL‾1. Moreover, Erkekoglu et al. (2010) examined the influence of DEHP and MEHP on MA-10 Leydig cell and measured the antioxidant enzymes activity. Reduction of the activity of GPX, TrxR and GST was observed with both DEHP and MEHP. Moreover, total GSH levels decreased. On the other side, Botelho et al. (2009) assessed the DEHP toxicity in rats. The animals were given DEHP in a dose of 500 mg kg‾1 from gestational day (seventh day) to lactation day (second day) and the antioxidant enzymes activity was measured in offspring. Increase of CAT activity and a decrease in GST activity were noted. In another study, Zhou et al. (2010) evaluated the effect of DBP on antioxidant enzymes activity. The findings showed that SOD, GSH-Px and GSH activity decreased at two doses of 250 and 500 mg kg‾1 and MDA (malondialdehyde) level was increased at the same doses in the testes. In a previous study, Lee et al. (2007) examined DBP properties at a dose of 750 mg kg‾1 orally during 30 days and measured the antioxidant enzymes activity. A significant increase was noted in MDA and 8-OHdG (8-hydroxy-2-deoxyguanosine) levels. Additionally, SOD, CAT and GPx activity increased. Bibliography revealed that Kasahara et al. (2002) evaluated the influence of DEHP in rat. The animals were treated with DEHP at 1 and 2 g/kg/day in drinking water during seven days. Reduction of free thiol, GSH and ascorbic acid, were reported at all doses. In addition, CAT and GPX activity increased at all doses too.

Erkekoglu et al. (2011) examined the oral administration of DEHP at a single dose of 1000 mg kg‾1 in rat testis during 10 days. The results showed a decrease in both Cu.Zn-SOD activity and GSH level. Also, an increase of TBARS level was noted. In a previous study, Ambruosi et al. (2011) investigated the influence of DEHP on Equine COCs (cumulus-oocyte complexes) at three doses of 0.12, 12 and 1200 μM during 1 h. An increase in CC apoptosis was observed at all doses. At the lowest dose, oocyte maturation was inhibited and ROS level was decreased, whereas, at other doses, there were no changes in oocyte maturation or ROS levels.

Moreover, Zhou et al. (2011) evaluated the effects of DBP in rat testis. The animals received DBP orally at doses of 100, 250 and 500 mg kg‾1 during 2 weeks. The results showed a decrease of SOD activity and an increase of MDA level in all groups. A further decrease of GSH-Px was observed at a dose of 500 mg kg‾1. In another study, Seo et al. (2004) investigated the influence of DEHP, DBP and BBP at three doses of 50, 200 and 1000 mg kg‾1 in rat liver. The MDA level increased significantly with all compounds at doses of 200 and 1000 mg kg‾1. It is also increased in a dose of 50 mg kg‾1 with DEHP. Moreover, enhancement of 8-OHdG was observed at a dose of 1000 mg kg‾1 with DEHP. Kang et al. (2010) examined the effects of DEP by three consecutive daily intraperitoneal (i.p.) injections at doses of 100, 300 and 900 mg kg‾1 in fish liver. The results showed an increase in LPO levels in all groups and decreasing the activity of CAT, AST and A LT at a dose of 900 mg kg‾1.

In a recent study, Yavasoglu et al. (2014) assessed the effects of BCP orally for 20 days at doses of 100, 200 and 400 mg kg‾1 in mouse liver. A significant decrease was observed in all groups. In addition decrease of SOD activity and increase of TBARS level were noted at doses of 200 and 400 mg kg‾1.

Recently, phthalate toxicity has been evaluated by Yang et al. (2012). In that study, the HepG2 cells were received MEHP at doses of 6.25, 12.5, 25, 50 and 100 μM during 36 h. The results showed an increase of MDA level and a decrease of GPx activity at all doses. It was also noted an increase of 8-OHdG at doses of 25, 50 and 100 μM. In another study, Erkekoglu et al. (2010) investigated the effects of DEHP (0.01-10 mM) and MEHP (3-30 μM) on LNCaP cells during 72 h. Furthermore, decrease of GPx activity and increase of GR activity (glutathione reductase) as well as DNA damage were all observed.

ALTERATIONS IN GENE EXPRESSION OF THE ANTIOXIDANT ENZYMES

Literature revealed that Wang et al. (2012a) assessed the effect of DEHP on gene expression of antioxidant enzymes. A decrease in SOD1 expression was noted. In another study, Wang et al. (2012b) evaluated the effects of MEHP on the gene expression. The results revealed that SOD1 and GPX expression decreased at 100 μg mL‾1. Decrease of anti-apoptotic factor (Bcl-2) expression and increase of pro-apoptotic factor (Bax) expression were noted at three doses of 1, 10 and 100 μg mL‾1. Moreover, expression of cell cycle genes decreased at the same doses.

Erkekoglu et al. (2010) evaluated the DEHP and MEHP toxicity on gene expression too. In that study, p53 expression increased by MEHP (IC50 = 3 mM). In addition, DNA damage was observed by increased tail intensity and tail moment. Also, these authors (Erkekoglu et al., 2011) investigated the effects of MEHP at a dose of 3 μM on LNCaP cell line. The results showed an increase in expression of p53 and p21. Additionally, in a previous study by Lee et al. (2007), the influence of DBP was evaluated on gene expression in rat. A significant increase of PPAR-r protein expression and PXRr (Retinoid X Receptors) expression were noted. All the mentioned data are exhibited in Table 1-5. Moreover, the diagrammatic representation designed for collecting data from the studies is shown in Fig. 1.

DISCUSSION

Oxidative stress and reproductive system: After conducting a vast systematic literature review regarding phthalate toxicity in reproductive organs and OS, it was pointed out that administration of the phthalate alters reproductive function by increasing of ROS (Akingbemi et al., 2004). First, germ cells showed high sensitivity to phthalates (Kasahara et al., 2002) and also MEHP increased Fas ligand expression in Sertoli cells which induced apoptosis in spermatocytes (Lee et al., 1997, 1999). On the other side, it was reported that DEHP can induce apoptosis in pachytene spermatocytes. The differences observed were related to the GSH levels and related enzymes in the cells (Bauche et al., 1994; Yoganathan et al., 1989). Obviously, phthalates can produce ROS through some pathways such as activation of PPARα that is assumed to induce reproductive disorder (Rusyn et al., 2006). Moreover, it is reported that phthalates can disrupt spermatogenesis and induce mitochondrial dysfunction in gonocytes (Suna et al., 2007).

Table 1: Toxic effects of phthalate on the reproductive system
DBP: Dibutyl phthalate, MBP: Mono-n-butyl phthalate, DEHP: Di-(2-ethylhexyl) phthalate, MEHP: Mono-(2-ethylhexyl) phthalate, ROS: Reactive oxygen species, CAT: Catalase, GPX: Glutathione peroxidase, SOD1: Cu/Zn superoxide dismutase, TrxR: Thioredoxin reductase, GST: Glutathione S-transferase, Trα-1: Thyroid hormone receptor α-1, PPAR-r: Peroxisome proliferator-activated receptor-r, 8-OHdG: 8-hydroxy-2-deoxyguanosine, MDA: Malondialdehyde, RXRs: Retinoid X receptors, GSH: Glutathione, COCs: Cumulus-oocyte complexes

Table 2: Toxic effects of phthalate in human
DEHP: Di-(2-ethylhexyl) phthalate, MEHP: Mono-(2-ethylhexyl) phthalate, ROS: Reactive oxygen species, GPX: Glutathione peroxidase, SOD1: Cu/Zn superoxide dismutase, 8-OHdG: 8-hydroxy-2-deoxyguanosine, MDA: Malondialdehyde, GR: Glutathione reductase

Table 3: Toxic effects of phthalate on liver
DBP: Dibutyl phthalate, BBP: n-butylbenzyl phthalate, BCP: Butyl cyclohexyl phthalate, DEHP: Di-(2-ethylhexyl) phthalate, CAT: Catalase, SOD: Superoxide dismutase, 8-OHdG: 8-hydroxy-2-deoxyguanosine, MDA: Malondialdehyde, GSH: Glutathione, LPO: Lipid peroxide, TBARS: Thiobarbituric acid reactive substances

Table 4: Tixuc effects of phthalate on kidney
DEP: Diethyl phthalate, LPO: Lipid peroxide, GST: Glutathione S-transferase and GR: Glutathione reductase

Table 5: Neurotoxicity of phthalate
DEHP: Di (2-ethylhexyl) phthalate, DIBPL: Diisobutyl phthalate, DBP: Dibutyl phthalate, OVA: Ovalbumin-immunized, MDA: Malondialdehyde, GSH: Glutathione

Onorato et al. (2008) indicated that the impairment of cellular redox mechanism increased peroxiredoxin 3 and cyclooxygenase 2 levels in spermatocytes. Thus, increase of intracellular ROS is undoubtedly one of the main mechanisms of phthalate-induced reproductive toxicity (Erkekoglu et al., 2010).

Hormonal evaluations in the reproductive system demonstrated that DEHP exposure in prenatal period increased MNGs in the rats possessed testosterone insufficiently (Botelho et al., 2009; Andrade et al., 2006; Mahood et al., 2007). In addition, administration of DEHP or MEHP was able to decrease the testosterone level in mice. Although DEHP reduced testosterone, the injection of testosterone had no effect on the prevention of testicular atrophy. Thus, it is possible that testosterone has no effect on the pathogenesis of testicular atrophy (Kasahara et al., 2002). On the other hand, superoxide radical and H2O2 that are able to increase the ROS levels might play more important roles, where MEHP (but not DEHP) increased the release of cytochrome c (a key factor for inducing apoptosis) in testis mitochondria in low concentration (Lee et al., 1999). DEHP and MEHP can also affect Sertoli cells and induce vacuolization (Kasahara et al., 2002).

A literature review for other kinds of phthalates showed that even DBP increased ROS by DNA hunting (increase of 8-OHdG) and lipids as well (increase of MDA) in testis which prevented cell growth and decreased testosterone level (Lee et al., 2007). Moreover, it was shown that prepubertal exposure to DBP can cause a significant abnormality in male reproductive track (Mylchreest et al., 2002). In addition, DBP is reported to induce atrophy in testis and hyperplasia in Leydig cells in neonatal rats (Kim et al., 2004). Some pathways for DBP toxicity have been suggested. These are including the decrease of testosterone level (Parks et al., 2000), anti-androgenic activity (Mylchreest et al., 1998) and zinc depletion (Fukuoka et al., 1995).

More investigations on DBP demonstrated that this compound was able to block Sertoli cell maturation and consequently AR expression level decreased in testis (Lee et al., 2007). Also, it is observed that DBP decreased the number of sperms by affecting Sertoli cells (Kleymenova et al., 2005). Likewise, DBP (750 mg kg‾1) enhanced TRα-1 protein level that decreased testosterone synthesis and thus testicular size reduced. In order to explain these alterations, it should be noted that there are peroxisomes in the testis (Corton and Lapinskas, 2005) and peroxisome proliferators can cause the protection of antioxidant enzymes against ROS (Mehrotra et al., 1999). CAT is one of the most important enzymes located in peroxisome in testis, nevertheless previous studies revealed that DEHP enhanced GPx and CAT activity while decreased GSH level in testis (Kasahara et al., 2002).

Although DEHP is known to cause alterations in the activity and expression of the antioxidant enzymes (Erkekoglu et al., 2010; Botelho et al., 2009), it is reported that DEHP inhibited antral follicle growth and reduced SOD1 activity and expression via an in vitro study (Wang et al., 2012a). SOD is an antioxidant enzyme converting superoxide into H2O2 while GPX and CAT convert this product into H2O and O2. Further, there are two major enzymes in rodent ovarian cells, including copper/zinc superoxide dismutase (SOD1) and manganese superoxide dismutase (SOD2). SOD1 and SOD2 are located in the cytoplasm and mitochondria, respectively (Matzuk et al., 1998; Tilly et al., 1995).

Beside the above mentioned studies, it was previously reported that DEHP (10 and 100 μg mL‾1) decreased the estradiol levels and inhibited the antral follicle growth in vitro (Gupta et al., 2010). In addition, only DEHP-10 (but not DEHP-100) enhanced ROS/RNS levels. Low and high doses of DEHP might act through different pathways. Lower dose of DEHP may work via oxidative stress while high dose of DEHP may affect the follicle growth through decreasing the estradiol levels. Moreover, DEHP-100 decreased gene expression of the so-called cell cycle regulators Ccnd2 (cyclin-D2), Cdk4 (cyclin-depended-kinase 4) and aromatase (Wang et al., 2012a). MEHP is also found to decrease the expression of the cell cycle regulators (Ccnd2, Cdk4 and Ccne1) and anti-apoptotic factor Bcl-2 (B-cell lymphoma 2) while increased the expression of pro-apoptotic factor Bax (Bcl-2-associated X protein) (Wang et al., 2012b).

Probably, the inhibition of GPX activity is one of the main mechanisms of MEHP reproductive toxicity. Accumulation of H2O2 due to inhibition of GPX can activate other antioxidant enzymes. DEHP mainly affects the expression and activity of SOD1 leading to the accumulation of superoxide, whereas MEHP decreased the expression and activity of GPX resulting in accumulation of H2O2 that caused more toxicity in antral follicles.

Oxidative stress and liver/kidney toxicity: Literature reviews demonstrated that PPs can increase H2O2 by induction of enzymes activities. In the same way, H2O2 is able to increase the 8-OHdG levels and carcinogenesis. Also, reduction of the hepatic activity of CYP1A1, 1A2 and 3A4 and selective induction of PPs on CYP4A1 which involved in lipid metabolism, were both demonstrated (Seo et al., 2004). Some studies are summarized in Table 3 indicating the correlation between liver toxicity of phthalates and OS parameters. In fact and in conclusion, by decreasing CAT and SOD, increasing of TBARS and MDA occurred.

An appraisal on Table 3 and 4 shows that LPO level generally increased in liver and kidney. DEP exposure increased detoxifying enzymes and anti-oxidative enzymes such as GSH, GPx and GR to deal with ROS. In contrast, there was no change in GST and CAT activity (Kang et al., 2010). In addition, it is revealed that MEHP decreased the viability of HepG2 cells while increased apoptosis (Yang et al., 2012). A significant increase of 8-OHdG at 24 h and a decrease of 8-OHdG at 36 h suggested that oxidative DNA damage might be repaired after 24 h. MEHP also induces mitochondrial dysfunction and reduces cellular ATP content. Moreover, up-regulation of cytochrome c in cytosol and increase of caspase-9 and caspase-3 were noted as well. DEHP and MEHP are reported to increase the DNA damage and apoptosis while decrease GPx1 activity in LNCaP cells. In that study, MEHP (not DEHP) caused augmentation in TrxR activity. TrxR and GPx1 are two important defense systems (Gromer et al., 2004) that play roles in many biological processes such as apoptosis and DNA repair (Arner and Holmgren, 2000). An increase of H2O2 may lead to a reduction in TrxR activity in small-cell lung carcinoma that has previously been reported (Gandin et al., 2009).

Oxidative stress and neurotoxicity: Previous studies have demonstrated that OS involves in neurological and psychiatric disorders, especially in depression (Ng et al., 2008; Sahin and Gumuşlu, 2004). Likewise, phthalates could induce these disorders as a result of imbalance in antioxidants and ROS levels (Zuo et al., 2014). Moreover, the accumulation of ROS has been related to a variety of neurodegenerative diseases (Bilici et al., 2001). On the other hand, OS cause impairment in learning behavior and reduce motor activity (Kumsta et al., 2001; Murakami and Murakami, 2005). Furthermore, oxidative stress might be considered as a main factor in neurotoxicity of phthalates, a factor by which disruption of neuronal systems may lead to neurobehavioral abnormalities (Tseng et al., 2013).

An appraisal of the present reports (Table 1-4) on phthalate compounds and oxidative stress shows us there are some controversies in augmentation of oxidative stress parameters after exposure of different organs to the various phthalate derivatives. For instance, although DEHP, DBP and DIBP could increase the levels of ROS, GSH and MDA in brain (Table 5), the level of ROS decreased following exposure to the lowest dose of DEHP 0.12 μM in the reproductive system (Table 1). But the most controversy can be seen in the levels of cellular antioxidants like SOD which generally shows decrease in many organs like liver, kidney and reproductive while in some studies its augmentation has been reported (Table 1). For example, Lee et al. (2007) reported SOD enhancement after exposure to 750 mg kg‾1 of DBP in rat testis. Studies on human cell lines (in vitro) exhibited an increase of MDA, a decrease of GPx activity and enhancement of 8-OHdG, as well as increase in DNA damage and GR activity following exposure to different phthalate derivatives. Nevertheless, extrapolation of the in vitro investigations to animal and human studies may bring some different results due to the lack of intracellular connections and normal physiological states (Shetab-Boushehri and Abdollahi, 2012), a review on the present data shows that, except a few reports almost all the animal studies revealed the critical role of oxidative stress in phthalate toxicity. An explanation for such differences can be found in possibility of compound accumulation in certain organs, tissues or cell types resulting in prolonged or increased exposure of the target tissue and then an in vitro model system may lead to an underestimation of biological activity of the compound under study.

CONCLUSION

Phthalates are among the most important synthetic chemicals playing roles in inducing toxicity in various organs such as liver, kidney and the reproductive system in both human and animals. Their mechanisms of toxicity, especially in reproductive system have not been exactly elucidated so far, however, altering the expression and activity of antioxidant enzymes that result in DNA damage via augmentation of ROS is one of the noteworthy mechanisms involved.

Various kinds of phthalates are able to interfere with antioxidant enzymes in different organs. For instance, DEHP enhances GPX and CAT activity, whereas decreases GSH level in the testis or DBP that acts via increasing ROS by DNA and lipid hunting and increasing MDA in the testis. Moreover, MEHP (but not DEHP) can cause an increase in TrxR activity. TrxR like GPX1 is an important defense system that displays in many biological processes such as apoptosis and DNA repair. An increase of H2O2 may lead to the increase of TrxR activity in a small-cell lung carcinoma cell line which has been previously reported. Since, various phthalate derivatives are able to cause irreversible tissue and organ damages especially in reproductive system via a number of mechanisms such as oxidative stress, recently, these compounds have been replaced in the USA, Canada and European Union by other plasticizers. Considering the U.S. Environmental Protection Agency (EPA), application of the following eight phthalates should be managed carefully: Dibutyl phthalate (DBP), diisobutyl phthalate (DIBP), Butyl Benzyl Phthalate (BBP), di-n-pentyl phthalate (DnPP), di(2-ethylhexyl) phthalate (DEHP), di-n-octyl phthalate (DnOP), diisononyl phthalate (DINP) and diisodecyl phthalate (DIDP), of which, BBP, DEHP and DBP cause the most toxicity to terrestrial organisms, fish and aquatic invertebrates.

ACKNOWLEDGMENT

This invited study is the result of an in-house study and no funding has been received.

ABBREVIATIONS

BBP: Benzyl butyl phthalate
CAT: Catalase
DBP: Dibutyl phthalate
DEHP: Bis(2-ethylhexyl) phthalate
DIBP: Diisobutyl phthalate
DIDP: Diisodecyl phthalate
DINP: Diisononyl phthalate
DnOP: Di-n-octyl phthalate
DnPP: Di-n-pentyl phthalate
EPA: U.S. environmental protection agency
GPXx: Glutathione peroxidase
GST: Glutathione S-transferase
MDA: Malondialdehyde
MEHP: Mono-(2-ethylhexyl) phthalate
PP: Peroxisome proliferator
PPARs: Peroxisome proliferator-activated receptor
ROS: Reactive oxygen species
SOD: Cu/Zn superoxide dismutase
REFERENCES
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