Introduction
Radiofrequency-electromagnetic radiation (RF-EMR) is a general term that describes periodic changes between positive and negative, and the speed or the number of changes per second, called the frequency, with Hertz as units (Beraldi et al., Reference Beraldi, Sciamanna, Mangiacasale, Lorenzini and Spadafora2003). RF-EMR generated from mobile phones has frequency radiation between 900–1800 MHz (Safari et al., Reference Safari, Mortazavi, Aghazade and Mousavi2020). Specific absorption rate (SAR) is the amount of RF energy absorbed by a person’s body during a phone call and usually is recorded in units of watts per kilogram (W/kg) or in milliwatts per gram (mW/g) (Gautam et al., Reference Gautam, Singh, Nirala, Murmu, Meena and Rajamani2019). Most phones have SAR ranging from 0.1 to 1.2 W/kg (Safian et al., Reference Safian, Khalili, Khoradmehr, Anbari, Soltani and Halvaei2016). In assisted reproductive technology (ART), embryos are generated and cultured under in vitro conditions. Therefore, environmental disruptions, such as RF-EMR exposure, may affect their survival and developmental capacities, therefore concerns about risk of malformations after being subjected to cell phone radiation have been increasing (Beraldi et al., Reference Beraldi, Sciamanna, Mangiacasale, Lorenzini and Spadafora2003).
There have been some in vitro and in vivo animal studies that assessed the effects on embryogenesis and newborns of RF-EMR propagated from cell phones. Some experiments revealed that RF-EMR from cell phones did not induce significant changes in the rates of fertilization and embryogenesis (Suzuki et al., Reference Suzuki, Okutsu, Suganuma, Komiya, Nakatani-Enomoto, Kobayashi, Ugawa, Tateno and Fujimori2017), in addition to histological or ultrastructural changes in embryos (Klug et al., Reference Klug, Hetscher, Giles, Kohlsmann and Kramer1997). In contrast, other studies have indicated that this radiation may cause decrease in embryo quality and blastulation rates and an increase in the number of embryo deaths (Fatehi et al., Reference Fatehi, Anjomshoa, Mohammadi, Seify and Rostamzadeh2018). From these experiments, it may be concluded that RF-EMR can be considered an environmental stress factor. Exposure of cells to stressful states may be associated with increased synthesis of stress proteins, including heat-shock proteins (Hsp), and increased levels of reactive oxygen species (ROS) (Weisbrot et al., Reference Weisbrot, Lin, Ye, Blank and Goodman2003; Safian et al., Reference Safian, Khalili, Khoradmehr, Anbari, Soltani and Halvaei2016). It has been confirmed that increased levels of ROS have detrimental effects on the embryo and fetus (Lampiao, Reference Lampiao2012; Shafiei et al., Reference Shafiei, Almasi, Nikzad, Miyan, Mahabadi and Moshkdanian2020). RF-EMR could induce numerous anomalies in offspring (El-Sayed et al., Reference El-Sayed, Badr, Yahia, Salem and Kandil2011; Celikozlu et al., Reference Celikozlu, Ozyurt, Cimbiz, Yardimoglu, Cayci and Ozay2012; Haghani et al., Reference Haghani, Shabani and Moazzami2013; Ye et al., Reference Ye, Wang, Zhang, Fang, Zhao and Wang2016), even though these effects may be temporary (Fragopoulou et al., Reference Fragopoulou, Koussoulakos and Margaritis2010).
As other mammals have many similarities to humans in terms of anatomy, physiology and genetics, the aim of this article was to summarize the existing literature that assessed the influences of RF-EMR exposure from cell phone on fertilization and subsequent embryonic and fetus development in other mammals in vitro and in vivo.
Materials and methods
We searched MEDLINE–PubMed (http://www.ncbi.nlm.nih.gov/PubMed), Google Scholar (https://scholar.google.com/), Scopus (https://www.scopus.com) and ISI Web of Science (http://apps.lib.wosg.ir/WOS) databases, from the earliest available online indexing year until April 2021 using the following Medical Subject Headings (MeSH) and non-MeSH keywords: ‘cell phone’, ‘embryo development’, ‘mammals’, and ‘fetal’. The searches were limited to original articles published in English and examined the in vitro and in vitro effects of RF-EMR emitted from cell phones on mammalian embryonic and fetal development. Other animal studies were excluded from this review.
Results
Effects of RF-EMR on embryo development
There have been few studies with different protocols that assessed the consequences of cell phone radiation on embryo development (Table 1). Klug et al. (Reference Klug, Hetscher, Giles, Kohlsmann and Kramer1997) exposed 11.5-day-old rat embryos (two or three somites) to RF-EMR. After 48 h, the rates of embryo growth and differentiation were evaluated using a scoring system. Both histological examination and protein content of the embryos were determined. They suggested that exposure to electric or magnetic fields did not significantly influence normal growth and differentiation of rat embryos in vitro (Klug et al., Reference Klug, Hetscher, Giles, Kohlsmann and Kramer1997). Also, Safian and co-workers (2016) evaluated the effects of cell phone radiation on in vitro development of mouse embryos. In their study, two-cell embryos were exposed to RF-EMR for 30 min/day for 4 days consecutively. The quality of embryos was documented daily and fluorescence staining was used for identification of viable blastocysts. Based on their results, both embryo survival and blastocyst formation rates were similar in both groups. However, death of embryos at the 2-cell stage and loss of cell viability in blastocysts were significantly higher in the experimental group (Safian et al., Reference Safian, Khalili, Khoradmehr, Anbari, Soltani and Halvaei2016). In a study administered by Suzuki and associates, both spermatozoa and oocytes were separately exposed to RF-EMR for 60 min. Then, in vitro fertilization (IVF) and intracytoplasmic sperm injection (ICSI) were performed using combinations of exposed and non-exposed oocytes plus spermatozoa. Interestingly, their results showed that waves propagated from cell phones had no significant detrimental effects on the rates of chromosomal abnormality, as well as IVF and ICSI fertilization rates, even though the amount of exposure in their experiment was 100 times greater than regular exposure to human spermatozoa and oocytes (Suzuki et al., Reference Suzuki, Okutsu, Suganuma, Komiya, Nakatani-Enomoto, Kobayashi, Ugawa, Tateno and Fujimori2017).
Effects of RF-EMR emitted from cell phone on embryo development in mammals
In a recent study, ovulating mice were irradiated at three RF-EMR intensities low intensity (mean intensity 150 μW/cm2), mid intensity (mean intensity 570 μW/cm2), and high intensity (mean intensity 1400 μW/cm2) for 4 or 2 h/day for 3 consecutive days. After that, the ova were collected for the IVF protocol. Their results indicated a decline in the IVF rate in the treatment groups compared with the control (Chen et al., Reference Chen, Qu and Liu2017). Also, Fatehi and colleagues (Reference Fatehi, Anjomshoa, Mohammadi, Seify and Rostamzadeh2018) separated male mice into five groups as control (I) and experimental groups (II–IV). Groups II, III and IV were exposed to cell phones on ‘Standby mode’ for 1, 5 and 10 h/day, respectively. Group V, however, was exposed to a cell phone on ‘Active mode’ 1 h/day. At 30 days after treatment, the mice were sacrificed and IVF was performed subsequently. Based on their studies, RF-EMR for 1 h on Active mode had more damaging effects compared with the 1 h Standby mode. Also, increased exposure time from 1 h to 5 h and 10 h, resulted in a higher decrease in the numbers of two-cell embryos and blastocyst rates (Fatehi et al., Reference Fatehi, Anjomshoa, Mohammadi, Seify and Rostamzadeh2018).
Effects of RF-EMR on offspring
Some studies focused on offspring following daily exposure of pregnant animals near to the RF-EMR emitted by a mobile phone (Table 2). Ogawa and her teammates subjected 7–17-day-old gestational rats to RF-EMR. In their study, the prenatal rats were allocated into four groups, with five pregnant rats in each group. Group 1 (control): animals were kept in conventional cages. Rats in Groups 2–4 were positioned in the exposure chamber for 90 min/day in the morning: Group 2 was considered as sham. Groups 3 and 4 were subjected to RF-EMR at low (average SAR: 0.67 W/kg) and high (average SAR: 2.0 W/kg) levels, respectively. Based on their results, exposure of the animal’s head to weak mobile phone-like signals during gestation induced no significant developmental changes in the offspring (Ogawa et al., Reference Ogawa, Nabae, Wang, Wake, Watanabe, Kawabe, Fujiwara, Takahashi, Ichihara, Tamano and Shirai2009). Furthermore, Fragopoulou and co-operators exposed 5-day pre-pregnancy female mice to RF-EMR. The 5-day period before pregnancy exposure was proposed to permit stimulation of the intrinsic defence system. After positive mating (vaginal smear), pregnant mice were exposed again until term (12–14 days).
Effects of RF-EMR emitted from cell phone on offspring in mammals
All the mice in both control and experiment groups were weighed regularly, and observed daily throughout pregnancy for overall appearance and locomotor activity by cage-side observation. At 5 h after birth, most pups were fixed for histological assessment and others remained for structural assessment. Their data indicated that mild exposure to RF-EMR from cell phones adversely influenced mice fetal development at the ossification level (Fragopoulou et al., Reference Fragopoulou, Koussoulakos and Margaritis2010). Also, El-Sayed and teammates exposed pregnant rats to RF-EMR from the 1st to 20th and 7th to 16th days of pregnancy. At the 20th day, the fetuses were detached from the uterus and evaluated. Liver of pregnant rats and their cubs were used for extraction of total RNA for quantification of Msx1 and Cx43 genes using real-time PCR. They noticed that subjecting to the rats to cell phones throughout the gestational period changed gene expression as well as some morphological and physiological factors in prenatal rats and their fetuses (El-Sayed et al., Reference El-Sayed, Badr, Yahia, Salem and Kandil2011).
In a study conducted by Jing and colleagues, pregnant rats were exposed to different intensities of RF-EMR. Experimental groups were irradiated three times a day from the 1st day of gestation for consecutively 20 days. On the 21st day, the fetuses were removed and the levels of superoxide dismutase (SOD), glutathione peroxidase (GSH-Px), malondialdehyde (MDA), noradrenaline (NE), dopamine (DA), and 5-hydroxyindole acetic acid (5-HT) in the brain were examined. The distance between the antenna and pregnant rat’s ear completely mimicked human methods of phone calling. Also, throughout the call, the antenna of the phone was kept near to the ear. They agreed that RF-EMR from cellular phones during the prenatal period had adverse effects on rat fetal brains (Jing et al., Reference Jing, Yuhua, Xiao-qian, Rongping, Dong-mei and Xi2012). In the same year, Celikozlu and colleagues evaluated the influence of magnetic fields from cell phones on some blood parameters and neurons in the brain of rats. The cell phones were located on the wall of the cage and were in standby mode for an entire day and in talk mode for 30 min per day. Rats were exposed to RF-EMR during prenatal and postnatal periods until they were 80 days old. Their findings proved that exposure to cell phones influenced some biochemical factors, particularly in the cortex region of the brain (Celikozlu et al., Reference Celikozlu, Ozyurt, Cimbiz, Yardimoglu, Cayci and Ozay2012).
In 2013, Haghani and co-workers exposed female rats to RF-EMR throughout the pregnancy period. Then, the newborns were evaluated for behavioural and electrophysiological status. Their outcomes demonstrated that prenatal RF-EMR exposure led to altered electrophysiological properties in Purkinje neurons. These modifications, however, were not serious enough to modify cerebellum-dependent functional properties (Haghani et al., Reference Haghani, Shabani and Moazzami2013). In another research study, rat cubs were exposed to RF-EMR from the beginning of pregnancy until 6 weeks after births. During the 4th to 6th weeks of the research, kidney and testis of offspring were removed and the levels of lipid peroxidation (LPO), copper, zinc, GSH, GSH-Px and total antioxidant status (TAS) were assessed. The data showed that waves from mobile phones led to precocious puberty and oxidative damage to the kidneys and testes of the rats (Özorak et al., Reference Özorak, Nazıroğlu, Çelik, Yüksel, Özçelik, Özkaya, Çetin, Kahya and Kose2013).
In addition, Bedir and colleagues irradiated rats with cell phone on ‘talk mode’ during first 20 days of the gestational period for 24 h per day. They evaluated histopathological variations and apoptosis in the kidneys, together with the levels of urea, creatinine and electrolytes in serum. The histological samples were assessed using immunohistochemistry. Their outcome indicated that RF-EMR emitted from cell phones had damaging effects on the renal development of pregnant rats (Bedir et al., Reference Bedir, Tumkaya, Şehitoğlu İ Kalkan, Yilmaz and Şahin2015).
Yüksel and team workers subjected pregnant rats (60 min/day for 5 days a week) for four generations. At the beginning of the pregnancy, female adults were irradiated for an additional 3 weeks. After specifying the sex of the newborns, the female progenies were exposed to additional radiation up to 6 weeks of age. They showed that mobile phones induced changes in progesterone, oestrogen and prolactin concentrations, oxidative stress (OS) levels, and endocrine functions in growing and maternal rats. Also, EMR could be a source of uterine injury in growing rats (Yüksel et al., Reference Yüksel, Nazıroğlu and Özkaya2016).
Alchalabi and teammates also exposed pregnant rats to RF-EMR from cell phones in the 1st to 3rd weeks of pregnancy. They saw uterine congestion, haemorrhage, and dead and reabsorbed fetuses during the 2nd and 3rd weeks of pregnancy and a meaningful decrease in the implantation sites and embryos during the 1st and 2nd weeks (Alchalabi et al., Reference Alchalabi, Aklilu, Aziz, Malek, Ronald and Khan2016). The effects of RF-EMR exposure to mobile phones during the gestational period on some biomarkers of OS were assessed in tissues, such as heart, liver, kidney, and the cerebellum of offspring mice. Based on the results, increased levels of MDA and decreased levels of total thiol groups (TTG), SOD, and catalase (CAT) were observed in all tissues from both mothers and offspring (Bahreyni Toossi et al., Reference Bahreyni Toossi, Sadeghnia, Mohammad Mahdizadeh Feyzabadi, Hosseini, Hedayati, Mosallanejad, Beheshti and Alizadeh Rahvar2018).
More recently, Haghani and colleagues exposed prenatal rats to cell phone radiation during pregnancy for 6 h/day. Their results indicated that exposure to cell phone radiation did not induce any significant alternations in body temperature, litter size, length of gravidity, body weight of newborns and mortality rate between control and experimental groups. However, it damaged the regular current flow of the cerebellar Purkinje neurons (Haghani et al., Reference Haghani, Pouladvand, Mortazavi, Razavinasab, Bayat and Shabani2020).
Discussion and limitations of studies
Cell phones are an indispensable method of communication in our daily lives. However, there are worries about the healthcare effects from RF-EMR emitted from cell phones. Epidemiological studies have suggested that RF-EMR from cellular phones may increase the risk of brain tumours (Sato et al., Reference Sato, Akiba, Kubo and Yamaguchi2011), diabetes (Sakurai et al., Reference Sakurai, Satake, Sumi, Inoue and Miyakoshi2004), sleep and personality disorders (Abdel-Rassoul et al., Reference Abdel-Rassoul, El-Fateh, Salem, Michael, Farahat, El-Batanouny and Salem2007). In addition, some studies have demonstrated an association between RF-EMR exposure and human reproductive diseases. In reproduction technology, there has been a focus on the effects of RF-EMR exposure on sperm parameters (Gutschi et al., Reference Gutschi, Mohamad Al-Ali, Shamloul, Pummer and Trummer2011; Okechukwu, Reference Okechukwu2020), fibroblasts and granulosa cells (Diem et al., Reference Diem, Schwarz, Adlkofer, Jahn and Rüdiger2005), ovarian follicles (Gul et al., Reference Gul, Celebi and Uğraş2009; Türedi et al., Reference Türedi, Hancı, Çolakoğlu, Kaya and Odacı2016; Koohestani et al., Reference Koohestani, Zavareh, Lashkarbolouki and Azimipour2019; Azimipour et al., Reference Azimipour, Zavareh and Lashkarbolouki2020) fertilization and embryo development (Batellier et al., Reference Batellier, Couty, Picard and Brillard2008).
This review summarized available published studies that assessed the effects of exposing in vitro and in vivo produced mammalian embryos to cell phone radiation on subsequent embryo survival. Inconsistency of results might be due to different experimental settings, animal models or various effects of RF-EMR in animals of different sizes (Santini et al., Reference Santini, Cordone, Falone, Mijit, Tatone, Amicarelli and Di Emidio2018). It appears that numerous factors could affect the outcome of RF-EMR experiments such as exposure conditions, cell phone types, frequency of the mobile phone, and duration of exposure (Safian et al., Reference Safian, Khalili, Khoradmehr, Anbari, Soltani and Halvaei2016). Also, RF-EMR from cellular phones always varies according to the surrounding environment and signal conditions. In addition, the amount of exposure to RF-EMR depends on the size of the exposed target, whether the target is stationary or moving, or whether the target is in air or liquid and, if in liquid, the properties of the liquid should be considered (Suzuki et al., Reference Suzuki, Okutsu, Suganuma, Komiya, Nakatani-Enomoto, Kobayashi, Ugawa, Tateno and Fujimori2017).
Based on the Safian study, although RF-EMR did not have significant adverse effects on mouse embryo development, they hypothesized that ROS production may have adverse effects on subsequent embryo survival (Safian et al., Reference Safian, Khalili, Khoradmehr, Anbari, Soltani and Halvaei2016). Several other studies have demonstrated that the exposure of different cell types to a 50 Hz RF-EMR led to an increase in the intracellular ROS levels (Ayşe et al., Reference Ayşe, Zafer, Sule, Işil and Kalkan2010; Mannerling et al., Reference Mannerling, Simkó, Mild and Mattsson2010; Patruno et al., Reference Patruno, Tabrez, Pesce, Shakil, Kamal and Reale2015; Calcabrini et al., Reference Calcabrini, Mancini, De Bellis, Diaz, Martinelli, Cucchiarini, Sestili, Stocchi and Potenza2017; Santini et al., Reference Santini, Cordone, Falone, Mijit, Tatone, Amicarelli and Di Emidio2018). Suzuki and colleagues announced that, even though 60 min exposure time did not cause spermatozoa ageing in in vitro condition, in vivo exposure for longer periods of time could affect fertilization, and early embryonic development (Suzuki et al., Reference Suzuki, Okutsu, Suganuma, Komiya, Nakatani-Enomoto, Kobayashi, Ugawa, Tateno and Fujimori2017).
Personal protective measures and future view
Mobile phones are commonly placed on the lap or in pockets (Swerdlow et al., Reference Swerdlow, Feychting, Green, Leeka Kheifets and Savitz2011), therefore exposing the genital area to RF-EMF. Germ cells are more susceptible to RF-EMF, as they are rapidly dividing through meiosis and mitosis. EMF induces modifications in cellular levels such as activation of voltage-gated calcium channels, formation of free radicals, protein misfolding and DNA damage (Altun et al., Reference Altun, Deniz, Yurt, Davis and Kaplan2018). In the general population, pregnant women are at specific risk of exposure to environmental RF-EMF because of their higher oxygen consumption and amniotic fluid-induced ROS production (Çiğ and Nazıroğlu, Reference Çiğ and Nazıroğlu2015) (Fig. 1). Individuals should reduce their rate of exposure to RF-EMF-emitting devices including mobile phones (Okechukwu, Reference Okechukwu2020) as electromagnetic waves enhanced the amount of oxygen free radicals in the body that led to disturbance of spermatogenesis process and variations in spermatozoa membrane, which resulted in changes in capacitation, acrosome reaction and therefore disorder in the process of spermatozoa binding to the oocyte and failure of normal fertilization both in vivo and in vitro (Fatehi et al., Reference Fatehi, Anjomshoa, Mohammadi, Seify and Rostamzadeh2018). In addition, exposure to RF may lead to a decline in ovarian follicle reservoirs at the start of the prepubertal time (Türedi et al., Reference Türedi, Hancı, Çolakoğlu, Kaya and Odacı2016).
Effect of electromagnetic radiation on female reproduction and fetal development. IUGR, intrauterine growth restriction (Agarwal et al., Reference Agarwal, Aponte-Mellado, Premkumar, Shaman and Gupta2012).
To reduce the contact with RF-EMR radiated by cell phones, people should avoid keeping their phones inside their pockets, also use of hand-free materials and gadgets and mobile covers limit direct expose of cell phones to the body (Okechukwu, Reference Okechukwu2020). Also, people do not appreciate that a cell phone is a small base station and usually underrate the importance of distance between the cell phone and the body (Cousin and Siegrist, Reference Cousin and Siegrist2010). In analyzing the relationship between the distance of cell phone antenna from the body and SAR, Hossain and colleagues reported that increasing the distance of cell phone from the body led to diminishing of both electric and magnetic field strength to the body, therefore reducing SAR values (Hossain et al., Reference Hossain, Faruque and Islam2015). Furthermore, it is known that the EMF decreases with distance and magnetic induction as calculated below:
$$B = k.{1 \over {{d^a}}}$$
where k is a constant dependent on the source, d is the distance to the source in metres, and α is commonly between 1 and 3, based on the nature of the source (Staebler, Reference Staebler2017).
Moreover, it is known that vitamins C and E and some natural supplements are helpful in protecting body against RF-EMR effects (Okechukwu, Reference Okechukwu2018, Reference Okechukwu2020). There have been limited studies that assessed the consequence of cellular phone electromagnetic radiation on ART such as IVF or ICSI (Suzuki et al., Reference Suzuki, Okutsu, Suganuma, Komiya, Nakatani-Enomoto, Kobayashi, Ugawa, Tateno and Fujimori2017), therefore additional investigations are required to examine the influence of cell phone usage in ART.
Conclusion
At this time, it is difficult from the available animal studies to document confidently the role of RF-EMR exposure on human embryo development, both in vivo and in vitro. Further investigations with complementary techniques will be necessary to understand the mechanism of action of RF-EMR emitted by cell phones and the consequences on mammals, particularly human beings.
Conflicts of interest
The authors report no conflicts of interest. The authors alone are responsible for the content and writing of this article.
