Mobile phones are the devices known for the release of EMF into the atmosphere can impact the cellular metabolic process and wield several biological effects of thermal or non-thermal mechanisms (Balawender and Orkisz 2020). Head is the most exposed region of the body to the radiation during the conversation. EMF in the frequency range of 900–2200 MHz can penetrate the cranium and reach the deep brain, altering brain function and behaviour (Ntzouni et al. 2011, 2013; Aldad et al. 2012; Li et al. 2012; Saikhedkar et al. 2014; Narayanan et al. 2015, 2018; Ahmadi et al. 2018; Singh et al. 2020). Several other studies reported stress and associated problems such as headache, tiredness, impairment of behavioural and cognitive functions due to the exposure of mobile phone radiation (Behari 2010; Kivrak et al. 2017; Narayanan et al. 2019). It was observed form previous studies that, the exposure to mobile phone radiation can lead to ROS generation (Singh et al. 2020), impairment of antioxidant system (Kesari et al. 2011; Singh et al. 2020), genotoxicity leading to apoptosis (Motawi et al. 2014; Hussein et al. 2016) in the regions of brain. The slow release of EMF from the electronic devices especially from mobile phones accumulate in the living organism and may disrupt the functionality of systems. There were no protective drugs over radiation effects, but numerous synthetic and natural substances were studied for their ability to reverse the effects of mobile phone radiation. (Ahmed et al. 2017; Altun et al. 2017; Asl et al. 2020).
Because of its structural similarities to curcumin, HIS, a natural polyphenolic derived from several mushrooms, piqued the interest of researchers to investigate for its therapeutic actions and is also considered a curcumin derivative (Ravindran et al. 2010; Amalraj et al. 2017; Chethna et al. 2018b) and has been reported for various pharmacological activities such as anti-bacterial, anti-oxidant, anti-tumour, antiviral, analgesic and anti-inflammatory, hepatoprotective, immunomodulatory, cerebroprotective and anti-diabetic (Ali et al. 2003; Chang et al. 2007; Ravindran et al. 2010; Huang et al. 2011; Chen et al. 2013; Gröndemann et al. 2016; Hsin et al. 2017; Prasanth et al. 2021). This made the researchers to synthesise the new derivatives to explore the spectrum of pharmacological activities (Shaikh et al.; Balaji et al. 2015, 2017). Considering the reported antioxidant activity of the HIS and its derivatives such as HP, HME, HMEP were studied for their protective role in primary cortical neuronal culture cells against the impact caused by the mobile phone radiation.
In the present study, HIS and its derivatives were assessed for their protection against radiation induced cytotoxicity. Mobile phone radiation exposure lead to cell death in various cell types including primary cultured neuronal cells (Cotgreave 2005; Xu et al. 2010; Nageshwar Rao and Satish Rao 2010a; Tohidi et al. 2021). The results of the present study are in line with the previous studies, exhibited the cell death in the primary cortical neuronal cells when exposed to the mobile phone for 2 hours. Among the treated groups, HP and HMEP administered groups showed significant decrease in cytotoxicity indicating protection when compared to the cells exposed to radiation alone which were in line the previous study attributed to the pyrazole groups present in diketo region of HIS and HME (Chethna et al. 2018b). The cell cycle analysis revealed significant increase in pre G1 peak indicating apoptosis in all treated groups. HP and HMEP significantly augmented pre G1 peak when compared to the radiation alone exposed group of cells. To understand the mechanism of protection of the pyrazole derivatives of HIS and HME, ROS levels were assessed hypothesizing that HP and HMEP alleviate the oxidative stress concentrated the neurons due to EMF from mobile phone radiation. Radiations emitted from mobile phones were reported develop oxidative stress by elevating the levels of ROS intra cellularly (Motawi et al. 2014). Nerve cells were more sensitive to ROS because of their weak antioxidant system, high metabolic rate, and decreased cellular turnover, ROS accumulates in neuronal cells (Lin and Beal 2006). Moreover, EMF also inhibit mitochondrial respiratory chain and prolong the half-life of ROS in the cells may lead to the cellular damage (Köylü et al. 2006). It can be observed from the results of the current study that, ROS levels were elevated in the primary cortical neuronal cells which are in line with the previous studies (Zhao et al. 2007; Nageshwar Rao and Satish Rao 2010b; Motawi et al. 2014). However there are many studies claimed no effect of mobile phone radiation on the development of oxidative stress (Hook et al. 2004; Zeni et al. 2007a). This incongruities between the studies may be due to the different systems of radio frequency (RF) generators, exposure protocols, and cell types (Zeni et al. 2007b; Simko 2007; Schwarz et al. 2008). HP, HME, and HMEP significantly reduced the levels of ROS in primary cortical neuronal cells when treated along with the radiation. They could be able to reverse the oxidative stress due to radiation as they were the derivatives of a bioactive polyphenolic compound HIS and are in line with the previous studies reported of their ROS scavenge in cell free systems(Shaikh et al.).
An increase in the rate of ROS production causes the build-up of ROS-associated damages in DNA, proteins, and lipids, which can lead to progressive cell dysfunction and, as a result, apoptosis, raising the overall probability of pathological conditions in an organism (Tsao and Deng 2004a; Suski et al. 2018). Mitochondria have been implicated in apoptosis regulation (Grebeňová et al. 2003). The membrane permeability of mitochondria is increased in presence of ROS and release its contents which could activate apoptosis (Guo et al. 2013). The results of the current study revealed the same that the ROS developed inside the cell upon mobile phone exposure of primary cortical neuronal cells indicated by green shift in JC-1 assay indicating the raise of mitochondrial membrane permeability might leak the contents of the mitochondria and activate apoptosis (Sivandzade et al. 2019). HIS, HP and HMEP significantly increased red/green ratio in comparison with radiation control and indicating protection of mitochondrial membrane potential under mobile phone radiation. Furthermore, extent of DNA damage and protection of HIS derivatives against the radiation was assessed in the primary cortical neuronal cells. In the brain cells of rats exposed for 2 hours to a 2450 MHz field at 0.6–1.2 W/kg, Lai and Singh found an increase in single and double-strand DNA breakage. They also reported that EMR exposure enhanced apoptosis and generated DNA-protein and DNA-DNA crosslinks in biological samples from rats (Lai and Singh 1995, 1997, 2004; Lai 1996). The current study reveals that there was DNA fragmentation indicating damage when exposed to 2100 MHz mobile phone with 1.6 W/Kg SAR for 2 hours. Pre-treatment with HIS compounds resulted in the significant change in the DNA fragmentation, and the compounds like hispolon monomethyl ether and hispolon showed significant change in the fragmentation indicates the apoptotic nature. The control groups itself showed basal level of DNA fragmentation. Cells has an intrinsic mechanism to detect and repair the breaks in DNA(Chatterjee and Walker 2017). DNA damage leads to the activation of p53, a tumour suppression gene that raises its levels with an increased ability to bind DNA to mediate transcriptional activation, which in turn activates various genes that prompt cell-cycle arrest, apoptosis, or DNA repair (Lakin and Jackson 1999). The results reveal that there was significant difference in the levels of p53 in the radiation control which are in line with the previous studies (Yilmaz et al. 2014). The tumour suppressor protein p53, which induces cell cycle arrest or apoptosis in response to DNA damage, targets both Bcl2 and Bax. Overall, the coordination of these molecules are crucial for directing a cell's life and death (Basu 1998). In the current study, the relative gene expression levels revealed the reason for apoptosis in radiation control group mediated by the upregulating the p53 as well as Bax and downregulating the Bcl2. HIS and its derivatives pre-treatment doesn’t change in the relative expression of apoptotic genes in the neuronal cells. According to Pamela et al. 2020, even short-term exposure to cell phone radiofrequency emissions can up-regulate elements of apoptotic pathways in cells derived from the brain, with neurons appearing to be more sensitive to this effect. The findings were also in par with Pamela et al indicating the upregulation of apoptotic pathways in primary cultured cortical neurons, when exposed to a 2100 MHz mobile phone with 1.6 W/Kg SAR for 2 hours, while pyrazole derivatives significantly altered the effects of mobile phone radiation, indicating their protective effect. At the same time these compounds did not alter gene expression levels opening a gateway to explore other pathways of protection incurred by these compounds.
It was previously recognised that phenolic compounds had antioxidant and cytoprotective properties against radiation-induced oxidative damage. They can act as reducing agents, hydrogen donors, singlet oxygen quenchers, and metal chelating agents due to their high redox potential, giving them intrinsic antioxidant capabilities (Tsao and Deng 2004b; Soobrattee et al. 2005).