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. 2025 Sep 10;40(3):e2025020. doi: 10.5620/eaht.2025020

Evaluation of the divergent effects of pyrethroid and D-allethrin-based coils fumes in male Wistar rats

Jelili Abiodun Badmus 1,4, John Olabode Fatoki 2,*, Adebayo Lawrence Adedeji 1, Ikeola Khalilat Mustapha 1, Blessing Chioma Ovili 1, Oyekunle Olufemi Oyedijo 1, Tolulope Fiyinfolu Olabode 1, Kemisola Deborah Joseph 1, Emmanuel Olusola Yawson 3, Donavon Charles Hiss 4
PMCID: PMC12890321  PMID: 41265404

Abstract

Mosquito coil fume is a cheap and commonly used method of reducing malaria incidence in third-world countries. The effects of fumes from pyrethroid and D-allethrin-based mosquito coils available in the Nigerian market were assessed in male Wistar rats. The rats were exposed to the insecticide fumes for 7, 14, and 21 days, while another group served as a control. The experiment consisted of seven randomly divided groups of six weight-matched animals per group. Plasma alanine aminotransferase (ALT), aspartate aminotransferase (AST), lactate dehydrogenase (LDH), cholesterol, phospholipids, high-density lipoprotein cholesterol (HDL-C), and high-density lipoprotein phospholipid (HDL-P) were evaluated. Lung-, liver- and kidney-reduced glutathione (GSH), glutathione peroxide (GPx), glutathione-S-transferase (GST), superoxide dismutase (SOD), malondialdehyde/lipid peroxidation (MDA) and lipid hydroperoxide (LOOH) were also evaluated. Histoimmunochemistry was used to assess lung p53 and Bcl-2 expressions. Pyrethroid and D-allethrin-based fumes induced a significant (p < 0.05) increase in plasma AST, LDH, cholesterol, phospholipids, and HDL-P, with a reduction of HDL-C levels. The fumes significantly and differently dysregulated antioxidant enzymes. The inhalations of the fumes induced significant (p < 0.05) increases in kidney MDA and LOOH levels, liver MDA by pyrethroid fume, and lung MDA by D-allethrin only, but lung LOOH by inhalations of both fumes. The increased expression of lung p53 and repression of Bcl-2 by both fumes were duration-dependent. The fume-induced disproportionate tissue function biomarkers, redox status, and apoptosis-related proteins. These effects are a possible panoply of divergent modes by which exposure to coil fumes can be deleterious to human health.

Keywords: insecticide, pyrethroid, D-allethrin, redox, apoptosis, malondialdehyde

Introduction

Malaria is widespread in tropical and subtropical regions, including sub-Saharan Africa and Asia [1]. The condition is caused by the bite of a female Anopheles mosquito infected with Plasmodium falciparum. In 2020, there were 241 million cases of malaria, and 627 thousand deaths were estimated, the majority of which occurred in sub-Saharan Africa and accounted for the leading cause of mortality in children under the age of five years [2]. The risk of illness can be contained by preventing mosquito bites through nets and mosquito repellents, or other measures such as spraying insecticides or burning insecticide coils and draining stagnant water [3]. The World Health Organization (WHO) recommends long-lasting insecticidal-treated nets free of charge and ensuring equal access for all people at risk of mosquito bites in Nigeria. Households resorted to insecticide coils to control the mosquito bites because of a lack of access to the nets. A similar scenario is obtainable from other parts of African countries that do not officially include the coil as a malaria control program [4]. The coil is the cheapest, easiest-accessible form of driving away mosquitoes while sleeping.

The coil burns slowly and emits smoke containing one or more insecticides. Each coil is usually used near the individual requiring mosquito bite protection [5]. General use of these mosquito coils poses a serious public health challenge, especially the chronic inhalation of fumes and consumption of produce that might have been inadvertently laced with chemical constituents of insecticide [6]. Exposure to the fumes has been shown to have a carcinogenic predisposition following degradation of the coil active principle to the potent lung carcinogen bis-(chloromethyl)-esther [7]. The active component of this coil is pyrethroid/pyrethrum, which accounts for 25% of the world insecticide market [8], [9], or allethrin, a synthetic analog of the natural pyrethrum insecticides obtained from the flower of Chrysanthemum cineraria folium [10]. Human exposure to these compounds for at least 8 hours a day can lead to bioaccumulation in all biomembranes of tissues such as blood due to their lipophilic nature [11]. Bioaccumulation is a known inducer of the overwhelming production of reactive oxygen species (ROS), more than the system's antioxidant capability and resultant oxidative damage to macromolecules in mammals [12]. To the best of our knowledge, there is an information gap in the understanding of the potential comparative consequences of inhaling pyrethroid and allethrin-based mosquito coil combustion in the lung, liver, and kidney redox status and p53 and Bcl-2 protein expressions of male Wistar rats. The present study is aimed at bridging this existing gap.

Materials and Methods

Reagents

Trichloroacetic acid (TCA), thiobarbituric acid (TBA), Tris-HCl buffer, KCl, Ellman’s reagent, 2,4-dinitrochlorobenzene, adrenaline, and hydrogen peroxide were obtained from Sigma-Aldrich (St Louis, MO, USA). All other chemicals used were of analytical grade.

Mosquito coils

Mosquito coils were sourced from the local market in Ogbomoso, Oyo State, Nigeria. Each brand of the two coils contained either 0.2% w/w pyrethroid or D-transallethrin as the main active agent and 99% w/w inert ingredients. Each mosquito coil is expected to burn for 8 hours.

Experimental Animals

Experimental protocols were conducted by the Guidelines of the Institutional Animal Care and Use Committee and approved by the Animal Ethics Committee of the Faculty of Basic Sciences, Ladoke Akintola University of Technology, Ogbomoso, Nigeria (LAUTECH/FBMS/18/167). Male Wistar albino rats with an average weight of 150 ± 20 g were purchased from a commercial breeder in Ogbomoso. They were kept for two weeks in the Departmental Animal House to acclimatize. The animals were housed in a cage under normal animal laboratory conditions of a 12-hour dark/light cycle. They were given rat pellets and water ad libitum.

Animal Grouping and Design

Rats were randomly divided into seven groups of six (6) animals each and housed separately according to their assigned group as stated: Group A animals served as control unexposed, Group B was exposed to pyrethroid-based mosquito coil (Swan®) for 7 days while Groups C and D were exposed to the coil for 14 and 21 days, respectively. Groups E, F, and G animals were exposed to D-allethrin-based mosquito coil (Rambo®) for 7, 14, and 21 days, respectively. A coil was administered to each group of animals per night for each period of exposure time. The animal respiratory exposure mimicked that of humans by placing the lit coil in the respective plastic cages, measuring 480 x 350 x 250 mm. The exposure was done at room temperature, which ranged between 25-30 °C. Two net windows measuring 200 x 130 mm, each provided cross ventilation in the cage.

Preparation of Tissue Homogenates

The animals were sacrificed under mild anaesthesia, and blood samples were collected by heart puncture. The lung, heart, kidney, and liver were quickly harvested, weighed, and washed in normal saline (0.9% NaCl) and homogenized in cold homogenizing buffer using a Teflon head homogenizer. The homogenate was centrifuged at 10,000 rpm for 15 min to obtain the post-mitochondrial fraction, and aliquots were kept in a freezer until further use.

Biochemical assay

Plasma alanine aminotransferase (ALT), aspartate aminotransferase (AST), lactate dehydrogenase (LDH), cholesterol, phospholipids, glucose, cholesterol, and HDL-C were assessed using RANDOX (Randox Laboratories, Ltd, 55 Diamond Road, Crumlin, County Antrim, UK.) reagent kits according to the manufacturer’s instructions.

Tissue antioxidant enzyme and lipid peroxidation evaluations

The method described by Beutler et al. [13] was employed in evaluating the level of reduced glutathione in the tissue homogenates. The protocol follows the reaction between Ellman’s (DNTB, 5,5′-dithiobis (2-nitrobenzoic acid)) reagent and free sulfhydryl group (thiols) of protein to generate yellow product (2-nitro-5-thiobenzoate (TNB)) that absorbs maximally at 412 nm. The level of GSH is equivalent to absorbance reading using a molar extinction value of 14,150 M-1 cm-1.

Glutathione peroxidase activity was evaluated using the method described by Rotruck and co-workers [14]. The method evaluates the ability of the sample’s GPx to protect the oxidation of GSH incubated with H2O2 for 10 min. The remaining GSH is determined using Ellman’s reagent (DNTB, 5,5′-dithiobis-2-nitrobenzoic acid).

The activity of glutathione-s-transferase (GST) was monitored spectrophotometrically using conjugation of 1-chloro-2,4-dinitrobenzene (CDNB) with glutathione at 340 nm wavelength [15]. The method is based on the conjugation reaction between GST and 1-chloro-2,4-dinitrobenzene to produce reduced glutathione. The activity of GST was expressed as nmoles of CDNB conjugates formed/min/mg protein using a molar extinction coefficient of 9.6 × 103 M-1 cm-1.

An indirect method of adrenaline autooxidation to adrenochrome was employed to evaluate SOD activity [16]. This follows the ability of the sample’s SOD to inhibit the autoxidation of adrenaline to adrenochrome in alkaline carbonate buffer (pH 10.2) measured at 340 nm. Inhibition of the autoxidation by 50% is equivalent to 1 unit value of SOD activity. Lipid peroxidation was evaluated by measuring thiobarbituric reactive oxygen species (TBARS) in the form of MDA using the method described by Varshney and Kale [17]. The procedure is based on the formation of an adduct between TBA and malondialdehyde as an end product of lipid peroxidation when heated in an acidic medium. The pink complex product is absorbed maximally at 532 nm and is extractable into an organic solvent (butanol).

Lipid hydroperoxide was evaluated using ferrous ion xylenol orange (FOX) assay as described by Jiang et al. [18]. Lipid hydroperoxide was evaluated by mixing the sample with FOX reagent (Ammonium ferrous sulfate, H2SO4, and Xylenol). The absorbance of the reacting mixture using acetone as blank was read at 560 nm on a UV-visible spectrophotometer

Immunohistochemistry evaluation

Immunohistochemistry evaluation was done following the standard procedure of immuno-stain formalin-fixed, paraffin-embedded lung tissue sections. The paraffin-embedded tissues were de-paraffinised and incubated with a primary antibody, followed by a secondary antibody (diluted biotinylated + streptavidin HRP). DAB (3,3’-Diaminobenzidine) was used to develop the colour of antibody staining

Statistical analysis

Results are expressed as mean ± SD. One-way analysis of variance (ANOVA) followed by Tukey’s post-hoc test was used to analyse the results, with p < 0.05 considered significant.

Results

The group exposed to pyrethroid fume for 21 days consecutively had significantly (p < 0.05) lower plasma glucose levels compared with the unexposed control group. The D-allentrin group of 14-day consecutive exposure showed significantly (p < 0.05) higher plasma glucose levels when compared with the unexposed control group (Figure 1D). Plasma glucose levels of 14- and 21-day pyrethroid fume-exposed groups were significantly (p < 0.05) lower when compared with D-allethrin-exposed groups at the same time points. Plasma protein levels were not significantly altered in groups exposed to both pyrethroid and D-allethrin during the entire time durations (p > 0.05) (Figure 1E).

Figure 1.

Figure 1.

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Plasma ALT (A), AST (B), LDH (C), glucose (D), and protein (E) levels of rats exposed to fumes of pyrethroid and D-allethrin-based mosquito insecticide coils for 7, 14, and 21 days. Data represent means ± SD of 6 animals per group. Where indicated,* denotes the p < 0.05 significance level of exposed groups compared to the control group. ALT: alanine aminotransferase; AST: aspartate aminotransferase; LDH: lactate dehydrogenase.

Pyrethroid and D-allethrin fumes did not significantly affect the plasma ALT levels compared with the control group during the entire exposure period. In contrast to plasma ALT, AST levels were significantly (p < 0.05) elevated in the groups exposed to pyrethroid for 14 and 21 days when compared with the control group (Figures 1A and 1B). However, D-allethrin induced a significant (p < 0.05) increase in AST levels at 7-, 14- and 21-day exposure compared with the control. No significant changes were observed in the plasma AST of rats exposed to pyrethroid compared with the D-allethrin-exposed group in each corresponding period. Pyrethroid- and D-allentrin-exposed groups showed significant increases in plasma LDH levels only at 21 days of exposure. At 21 days, plasma LDH levels of the D-allentrin-exposed group were significantly higher (p < 0.05) than those of the pyrethroid-exposed group (Figure 1C).

Plasma cholesterol levels, as shown in Figure 2A, were found to be significantly (p < 0.05) lower in 7-day and 14-day pyrethroid fume-exposed groups, while no significant change was observed in the 21-day exposed group. The group exposed to D-allethrin for 14 days showed significantly (p > 0.05) higher cholesterol levels. The increase observed in 7- and 21-day D-allentrin was not significant compared with the control group. Plasma cholesterol levels of the 7- and 14-day pyrethroid-exposed groups were significantly (p < 0.05) lower than D-allethrin-exposed groups of the same period. Pyrethroid increased the plasma phospholipid concentration with significant increases occurring at 14- and 21-day exposures (Figure 2B), while significant increases (p < 0.05) in D-allentrin-exposed groups (7-, 14- and 21-day exposures) were not duration-dependent. The plasma phospholipid increase of the 7-day pyrethroid exposure group was significantly (p < 0.05) higher than 7-day D-allentrin-exposed group. By contrast, phospholipid levels measured in the 14- and 21-day D-allethrin-exposed groups were significantly (p < 0.05) higher than pyrethroid-exposed groups at the same time points. Figure 2C shows that plasma HDL-C levels were found to be significantly (p < 0.05) lower due to exposure to both fumes, except at 7 days in the pyrethroid-exposed group, with a non-significant reduction. On the other hand, HDL-P levels were elevated significantly by both fumes at all the time intervals (Figure 2D). While marked increases in plasma HDL-P were observed for the pyrethroid-fume-exposed group, the D-allentrin-fume-exposed group showed a significant surge at 14 days.

Figure 2.

Figure 2.

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Plasma cholesterol (A), phospholipids (B), HDL-C (C), and HDL-P (D) levels of rats exposed to fumes of pyrethroid and D-allethrin-based mosquito insecticide coils for 7, 14, and 21 days. Data represent means ± SD of 6 animals per group. Where indicated,* denotes the p < 0.05 significance level of exposed groups compared to the control group. HDL-C: high-density lipoprotein cholesterol; HDL-P: high-density lipoprotein phospholipid.

The D-allenthrin-exposed group showed no significant (p > 0.05) change in liver GSH level at 7-day exposure, while a significant (p < 0.05) increase was observed at 14- and 21-day exposures. Liver GSH was not affected by pyrethroid fume at the entire exposure time (Figure 3A). The two fumes significantly (p < 0.05) lowered kidney (Figure 3B) and lung (Figure 3C) GSH levels. However, the reduction observed in the kidney GSH level was not duration-dependent (Figure 3A), while lung GSH level reduction was duration-dependent when compared with the control (Figure 3C).

Figure 3.

Figure 3.

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GSH levels of the liver (A), kidney (B), and lung (C) of rats exposed to fumes of pyrethroid, and Dallethrin-based mosquito insecticide coils for 7, 14, and 21 days. Data represent means ± SD of 6 animals per group. Where indicated,* denotes p < 0.05 significance level of exposed groups compared to the control group. GSH: glutathione.

Both fumes induced a significant (p < 0.05) increase in the activities of both liver (Figure 4A) and kidney (Figure 4B) GST at each period of exposure, while a significant (p < 0.05) increase in lung GST was observed only at the 7-day exposure (Figure 4C). The induced increase in kidney GST was duration-dependent (Figure 4B), in contrast to liver GST, which was not duration-dependent (Figure 4A).

Figure 4.

Figure 4.

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GST activities in the liver (A), kidney (B), and lung (C) of rats exposed to fumes of pyrethroid and Dallethrin-based mosquito insecticide coils for 7, 14, and 21 days. Data represent means ± SD of 6 animals per group. Where indicated, * denotes p < 0.05 significance level of exposed groups compared to the control group. GST: glutathione-S-transferase.

Fumes from pyrethroid insecticide coils induced a significant (p < 0.05) duration-dependent increase in kidney GPx activity (Figure 5B), while such an increase was only observed at the 7-day exposure in the D-allethrin-exposed group (Figure 5A) relative to the control group. Interestingly, significant (p < 0.05) decreases in liver GPx activities were noted in the D-allethrin-exposed group after the 14- 21-day exposure times compared with the control (Figure 5A). Pyrethroid fume induced a significant (p < 0.05) increase in the activity of kidney GPx while D-allentrin significantly (p < 0.05) increased such activity at 7- and 14-day exposures while no change was observed at the 21-day exposure when compared with the control (Figure 5B). Both fumes induced significant (p < 0.05) decreases in lung GPx activity (Figure 5C).

Figure 5.

Figure 5.

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Figure caption. GPx activities measured in the liver (A), kidney (B), and lung (C) of rats exposed to fumes of pyrethroid and D-allethrin-based mosquito insecticide coils for 7, 14, and 21 days. Data represent means ± SD of 6 animals per group. Where indicated, * denotes p < 0.05 significance level of exposed groups compared to the control group. GPx: glutathione peroxide.

Pyrethroid fume at 14- and 21-day exposures significantly (p < 0.05) increased the liver SOD activities while D-allethrin exposure led to a significant (p < 0.05) decrease at 21-day only as shown in Figure 6A. Both fumes induced significant (p < 0.05) increases in the activities of kidney SOD except for 7-day exposure to D-allethrin, which represented an insignificant (p > 0.05) lower SOD activity (Figure 6B). Lung SOD activity was found to be lowered at 7-day exposure to pyrethroid (Figure 6C), whereas D-allethrin at 7- and 14-day exposure induced a decreased lung SOD activity relative to control (Figure 6C).

Figure 6.

Figure 6.

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SOD activities in the liver (A), kidney (B), and lung (C) of rats exposed to fumes of pyrethroid and Dallethrin-based mosquito insecticide coils for 7, 14, and 21 days. Data represent means ± SD of 6 animals per group. Where indicated, * denotes the p < 0.05 significance level of exposed groups compared to the control group. SOD: superoxide dismutase.

Liver MDA in rats exposed to pyrethroid was significantly (p < 0.05) higher at 21 days, whereas MDA decreased in the D-allethrin-exposed group (Figure 7A). The fume of D-allethrin significantly induced an increase in the kidney MDA levels of the exposed animals at both the 14- and 21-day periods (Figure 7B), whereas an increase in the level of lung MDA occurred at only the 14-day exposure (Figure 7C). No significant effect was found in the liver LOOH level when the animals were exposed to both fumes (Figure 7D). However, exposure to pyrethroid fumes significantly induced kidney LOOH levels at 14- and 21-day exposure, whereas kidney LOOH levels were significantly induced in D-allethrin-exposed animals (Figure 7E). Lung LOOH levels were significantly (p < 0.05) induced by pyrethroid at 14-day exposure, while D-allethrin significantly (p < 0.05) increased LOOH levels at 21-day exposure (Figure 7F).

Figure 7.

Figure 7.

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Levels of lipid peroxidation in terms of MDA and LOOH. liver MDA (A), kidney MDA (B) lung MDA(C), liver LOOH (D), kidney LOOH (E), and lung LOOH (F) levels of rats exposed to fumes of pyrethroid and Dallenthrin based mosquito insecticide coils for 7, 14 and 21 days. Data represent means ± SD of 6 animals per group. Where indicated,* denotes p < 0.05 significance level of exposed groups compared to the control group. MDA: malondialdehyde/lipid peroxidation; LOOH: lipid hydroperoxide.

Immunohistochemistry using Image J densitometric analysis showed that there was a significant increase in p53 expression in the lungs of animals exposed to both fumes relative to weeks of exposure (Figure 8). The increase according to the period of exposure was observed with D-allenthrin, while pyrethroid at 21-day exposure revealed downregulated expression. Conversely, both fumes induced a significant decrease in lung Bcl-2 expression relative to the period of exposure (Figure 9). The summary of possible overall effects of D-allenthrin and pyrethroid based mosquito coil fumes exposure in rats for 21 days is presented in Figure 10.

Figure 8.

Figure 8.

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Expression of p53 in the lungs of rats exposed to fumes of pyrethroid and D-allethrin-based mosquito insecticide coils for 7, 14, and 21 days. A represents the densitometry of p53 expression obtained from immunohistochemistry plates using the Image J software while B represents plates of immunohistochemistry. The number 1 represents the control group while 2, 3, and 4 are representative of pyrethroid-exposed groups of 7, 14, and 21 days, respectively. The numbers 5, 6, and 7 are representative of D-allethrin exposed groups of 7, 14, and 21 days, respectively.

Figure 9.

Figure 9.

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Expression of Bcl-2 in the lungs of rats exposed to fumes of pyrethroid and D-allethrin-based mosquito insecticide coils for 7, 14, and 21 days. A represents the densitometry of Bcl-2 expression obtained from immunohistochemistry plates using the Image J software while B represents plates of immunohistochemistry. The number 1 represents the control group while 2, 3, and 4 are representative of pyrethroid-exposed groups of 7, 14, and 21 days, respectively. The numbers 5, 6, and 7 are representative of D-allethrin exposed groups of 7, 14, and 21 days, respectively.

Figure 10.

Figure 10.

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Summary of effects of 21 days exposure of D-allenthrin and pyrethroid based mosquito coil fumes in rats. Prolonged exposure could cause apoptosis induction in rat lung with human risk assessment implications. GSH: reduced glutathione, GST: glutathione-S-transferase, SOD: superoxide dismutase, LOOH: lipid hydroperoxide, TBARS: thiobarbituric acid reactive substances, GPx: glutathione peroxide, ALT: alanine amino transferase, LDH: lactate dehydrogenase, AST: aspartate amino transferase, HDL-C: high density lipoprotein cholesterol and HDL-P: high density lipoprotein phospholipid.

Discussion

Evidence-based laboratory and epidemiology studies correlate possible health implications of human exposure to insecticides. Pyrethroids have enjoyed wide and contradictory scientific reportage, while there is a scarcity of information on the assessment of D-allethrin, a product from pyrethroids, which is equally used as an insecticide. Swaminathan [19] linked the occurrence of Type II diabetes to organophosphates, organochlorines, and carbamate exposures [19]. The findings of the current work show that D-allethrin fumes have the propensity to induce diabetic conditions more than pyrethroids. This is due to the observed significant surge of blood sugar level on the 14th day of exposure before the predictable body system reversal of the sugar on the 21st day. By contrast, the work of Narendra et al. [20] showed a significant upsurge in blood glucose levels in human subjects exposed to pyrethroids, allethrin, and prallethrin. High plasma glucose is a product of perturbations of gluconeogenesis and glycogenolysis pathways by the insecticides, causing insulin resistance through pathological tilts towards oxidative stress and pro-inflammatory markers [21]. Similarly, Zhang et al. [22] reported the potential role of D-allethrin and chemical components of insecticides, such as omethoate, in the pathogenesis of insulin resistance.

The increases in plasma AST and LDH levels observed in this study could be linked to damage to tissues other than the hepatic system because of the non-significant increase in plasma ALT levels. The elevated level of plasma ALT is a known specific biomarker for hepatic damage due to exposure to toxicants. Except for the non-significant increase of plasma ALT observed for both fumes, elevated levels of plasma AST and LDH are consistent with the study of Araoud et al. [23] that reported alteration of biochemical and hematological markers in Tunisian agricultural workers exposed to insecticides. Also, Idowu et al. [24] reported elevated levels of these biomarkers in rats exposed to coil emission for an extended period.

Multivariate analysis has linked the increase in ALT to the duration of exposure to pesticides [23]. Pesticides interact with phospholipid constituents of the plasma membrane, leading to increased membrane fluidity and subsequent cellular enzyme leakage into the extracellular lumen and eventual increased activity in the blood [25]. Cholesterol is an essential sterol in animals required as a structural component of membranes and also a precursor to a wide variety of steroid hormones. Cholesterol homeostasis perturbation signals the development and progression of atherosclerotic coronary artery disease (CAD) [26]. Downregulation of plasma cholesterol in animals exposed to pyrethroid fumes for 7 and 14 days consecutively might occur because of impaired cholesterogenesis due to reduced activities of 3-hydroxyl-3-methylglutaryl CoA reductase (HMG-CoA reductase) biosynthesis and mevalonate-5-pyrophosphate carboxylase [27].

Statins are compounds that reduce cholesterol synthesis through HMG-CoA reductase inhibition [28]. Pyrethroid may be one of the compounds that mimic the effect of statins on HMG-CoA reductase. The increased phospholipids in both plasma and HDL in pyrethroid-exposed groups are likely the result of significant concentrations of free fatty acids (FFA) deployed for phospholipid synthesis instead of cholesterol synthesis. This might be another plausible factor for the reduced cholesterol observed in rats exposed to pyrethroid fumes. The work of Rader [29] confirmed reduced HDL cholesterol and hypocholesterolemia as consequences of lipoprotein lipase (LPL) reduced activity [29]. It is also not inappropriate to hypothesize that the observed hypocholesterolemia in this study is a result of inhibition of LPL by pyrethroid exposure.

Contrary to pyrethroid exposure, D-allethrin fumes resulted in hypercholesterolemia that could be linked to enhancement of cholesterogenesis in the liver and/or induction of the activity of HMG CoA reductase. Rotimi et al. [30]. linked the activity of HMG-CoA reductase to the quantity of the enzyme available, which could fluctuate due to the effect of toxicants on the enzyme's synthesis and/or degradation and intrinsic catalytic efficiency influenced by the effects of toxicants on the phosphorylation or dephosphorylation of the enzyme [30].

Phospholipidosis is a storage disorder characterised by a disproportionate accumulation of phospholipids in various tissues [31]. Phospholipidosis is a hallmark of exposure to pyrethroid and D-allethrin fumes because of significantly elevated plasma phospholipids. This is consistent with our earlier postulation that the bulk of FFA available is likely to have been routed through phospholipid synthesis. Phospholipidosis initiation time frame ranges between a few days and depends strongly on the agent's affinity for susceptible cells [29]. The decrease in HDL-C and the increased phospholipids in HDL as observed for both pyrethroid and D-allethrin fumes is reflective of lipoprotein abnormalities as earlier stated [30].

Previous studies have implicated lipid and lipoprotein anomalies as independent risk factors in the pathogenesis and progression of CAD [32], [33]. These findings therefore indicate the potential of pyrethroid and D-allenthrin fumes as risk factors in the pathogenesis and progression of CAD. Bao et al. [34] have demonstrated a causal link between exposure to pyrethroid insecticides and cardiovascular-related deaths [34]. A tilt of the equilibrium toward reactive oxygen species (ROS) leads to oxidative stress with deleterious consequences [35]. The reduced kidney and lung GSH levels observed in this study might be due to oxidative stress induced by both fumes in the tissues. The liver GSH contents are not affected like those of the lung and kidney, which shows that the liver is not susceptible to the oxidative effects of the fumes within the period of evaluations. The reduced GSH levels observed in the present study are a corollary of induced oxidative stress in the organs.

This result is consistent with previous work presenting the potential of insecticides to disrupt the synthesis and utilization of GSH [36, 37]. GSH plays pivotal physiological roles by using its thiol group to maintain the systemic balance of reduction and oxidation. The reduction of GSH makes the tissues susceptible to a series of reactive oxygen species that attack with devastating consequences [38].

Aerobic animals, in addition to GSH, are endowed with interrelated cascades of endogenous antioxidant enzymes that protect cells from oxidative stress-induced macromolecular damage [39]. The endogenous antioxidant enzymes are reducing elements such as superoxide dismutase (SOD), glutathione peroxidase (GPx), and glutathione-S-transferase (GST) [40]. They are enzymes that mitigate the effects of ROS build-up during normal metabolic processes [36]. The endogenous antioxidant enzymes continuously keep ROS generated from normal metabolic processes in balance to maintain homeostasis [41]. This study shows that the effects of both fumes on the antioxidant enzymes are organ- and insecticide-specific. The organ-specific effects of the insecticides on all the antioxidants tested may be a result of varying amounts of the toxicants distributed to the respective organs. As expected, both the pyrethroid and D-allethrin will induce more consequences on the organs where they are more concentrated than those where they are less concentrated. Meanwhile, the distribution of drugs among organs is likely to be influenced by the affinity of organs for certain toxicants, tissue perfusion, and blood flow.

The lung is the only organ in this study that shows the same trend in the effects of both fumes on antioxidant enzymes. This is likely because the organ is directly exposed to the fumes, unlike others that depend on its metabolites. Upregulation or downregulation of these enzymes, as observed in this study, is in strong agreement with previous studies [36, 38, 42, 43]. The increases in these enzymes are likely due to adaptive responses to protect the organ from the overwhelming effects of induced ROS, and the decreased lung SOD and GPx activities could be due to reduced protein synthesis or degradation of the proteins due to oxidative stress [44]. Lipid peroxidation is a resultant effect of the overwhelming influence of oxidative stress on the lipid components of the cell membrane.

The influence is known to cause the oxidation of lipids and subsequent membrane damage and membrane protein inactivation [45]. Lipid peroxidation is assessed through thiobarbituric acid reactive substances (TBARS) in the form of malondialdehyde (MDA) or lipid hydroperoxides (LOOH) [46]. The increases in the levels of MDA and LOOH at different time points in some of the organs confirm the involvement of induced oxidative stress occasioned by exposure to coil fumes. This corresponds with previous work on mosquito repellants in different animal organs [47].

The seemingly D-allethrin reduction of liver MDA and no apparent changes in liver LOOH level in the presence of both fumes show that the time needed for the expected increased lipid peroxidation could be longer. This agrees with the study of Madhubabu and Yenugu [42], who reported a reduction in MDA of some reproductive organs for an exposure period of less than 30 days.

Exposure to pesticides is one of the environmental factors that induces apoptosis through the generation of overwhelming ROS in aerobic organisms. p53 and Bcl-2 play critical roles and tightly regulate the apoptosis process. p53 is a known promoter while Bcl-2 acts as an antagonist of the apoptosis pathway [48].

The increased p53 and reduction of Bcl-2 level observed in this study may be due to an adaptive response to ROS-induced apoptosis. The expression of p53 is negatively associated with Bcl-2 expression as shown in this study, which is consistent with the role of p53 as a transcriptional repressor of Bcl-2 expression for the induction of apoptosis. However, the pattern of the results obtained on the expressions of P53 and Bcl-2 is not different from some agents that induce toxicity through oxidative stress, such as acetamiprid (neonicotinoid pesticide), lead, cadmium, etc. These compounds have been shown to either increase the expression of p53 or reduce the expression of Bcl-2 in different tissue types [49, 50, 51].

Conclusions

This study showed that fumes from pyrethroid- and D-allethrin-based mosquito coils induce different patterns of dysregulation in antioxidant enzymes, liver function biomarkers, and lipid profiles. Both fumes had the same pattern of effects on the expressions of p53 and Bcl-2, which indicates induced cell apoptosis (Figure 10). Therefore, prolonged exposure to fumes from pyrethroid and D-allethrin-based mosquito coils could be a pathway to one of the non-communicable diseases related to oxidative stress.

Footnotes

Acknowledgement

The authors acknowledge the Technical Staff of the Department of Biochemistry, College of Health Sciences, Ladoke Akintola University of Technology, Ogbomoso, Nigeria, for their technical assistance.

The authors confirm that the data used to support this study's findings are included in the manuscript. The raw data are available upon request from the corresponding author.

The author(s) reported no funding associated with the work featured in this article.

Conflict of interest

No potential conflict of interest was reported by the author(s).

CRediT author statement

JAB: Conceptualization, Methodology, Validation, Investigation, Supervision, Formal analysis, Resources, Writing – Original draft preparation, Writing – Review and Editing, Project administration; JOF: Conceptualization, Methodology, Validation, Investigation, Formal analysis, Resources, Writing – Review and Editing; ALA: Conceptualization, Methodology, Investigation, Writing – Review and Editing; DCH: Conceptualization, Methodology, Validation, Investigation, Resources, Writing – Review and Editing, Project administration; EOY: Investigation, Writing – Review and Editing; IKM: Investigation, Writing – Review and Editing; BCO: Investigation, Writing – Review and Editing; OOO: Investigation, Writing – Review and Editing; TFO: Investigation, Writing – Review and Editing; KDJ: Investigation, Writing – Review and Editing.

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