Acetamiprid is a new generation neonicotinoid insecticide, widely used against insect pests as an alternative to other traditional pesticides. However, growing evidence has reported ACMP-exposure mediated severe toxic effects both at cellular and tissue levels in non-targeted organisms (Phogat et al. 2022). Various studies have shown oxidative stress generation during ACMP exposure eliciting organ toxicity by depleting antioxidants level (Chakroun et al. 2016; Erdemli et al. 2020; Khovarnagh and Seyedalipour 2021). The liver has been reported as one of the prime targeted organ for ACMP toxicity due to its involvement in metabolism, detoxification and biotransformation processes (Eftekhari et al. 2020). Nowadays, natural antioxidants with various biological activities have gained much attention and are of significant interest in attenuating oxidative injuries. Extensive investigations have been made to discover the antioxidative and therapeutic potential of natural antioxidants. Berberine is one such natural alkaloid with antioxidative and anti-inflammatory potential. So in the present study, we investigated the ameliorative potential of natural alkaloid; BBR against ACMP-induced oxidative stress, inflammatory changes and histopathological alterations in liver tissue of rats. Findings of the current study suggested that BBR pretreatment effectively ameliorated ACMP-induced oxidative stress in rats via prevention of oxidative injury to biomolecules, regulation of endogenous antioxidants and regulated inflammation and also prevented structural damages.
Oxidative stress-mediated biomolecular damage has been identified as a prime mechanism in ACMP-induced toxicity. Earlier findings have demonstrated increased LPO and protein oxidation in liver and kidney tissues of rats following ACMP exposure. The present study demonstrated that ACMP exposure significantly increased LPO and protein oxidation, suggesting that ACMP exposure could have generated free radicals that attributed to oxidative damage to membranous proteins and lipids, resulting in loss of cellular functioning. Conversely, treatment of BBR diminished LPO and protein oxidation levels in ACMP-intoxicated rats suggesting ameliorative efficacy of BBR against ACMP-mediated oxidative stress. Our results are consistent with earlier findings where supplementation of BBR attenuated hepatic oxidative stress by quenching free radicals and inhibiting the MDA formation in diabetic rats (Zhou and Zhou 2011), ethanol treated mice (Zhang et al. 2014) and acetaminophen-exposed mice (Zhao et al. 2018).
Further, the FTIR findings also confirmed the oxidative injuries to lipids and proteins following ACMP exposure. The shift in peak position at 3546 cm− 1 and 3467 cm− 1 in ACMP-exposed rats indicates the oxidative formation of lipids hydroperoxyl group which can be associated with LPO (Raouf et al. 2012). Amide A (3413 cm− 1 and 3294 cm− 1) and B (3071 cm− 1) peaks arise mainly due to N-H stretching in proteins. The reduced band area of Amide A and B reflects a decrease in the quantity or synthesis of proteins in rat liver administered to ACMP. Asymmetric and symmetric vibrations or stretching of CH3 and CH2 indicate the ‘State-of-order of hydrocarbon tail in lipids’. Reduction in peak area and shift in the position of these areas suggested conformational disorder and oxidative modifications in lipid acyl chain that coincide with LPO and protein oxidation observed in biochemical findings of the present study. The enhanced modification of lipid acyl chain might alter membrane permeability and fluidity, promoting oxidative stress generation that further causes alterations in proteins and lipids. These findings also confirm that oxidative stress is degenerative to cellular biomolecules, which is in relation with earlier findings (Borchman et al. 2000; Rodrigues et al. 2002). The significant decrease in the peak area of Amide I (1642, 1621) and Amide II (1548) after ACMP-intoxication reflects decrease in protein content and alteration in the secondary structure of proteins (Liu and Mantsch 2001), which can be attributed to protein oxidation and depletion of endogenous antioxidants as observed in the biochemical analysis. The marked reduction in peak area and frequency shift around 1452 cm− 1 also indicates lipid peroxidation. FTIR analysis revealed changes in lipids and protein content along with altered membrane fluidity that might attribute to the peroxidation of lipids and proteins due to ACMP-exposure in rat liver. Conversely, supplementation of BBR pronouncedly normalized the peak area and maintained the position of peaks nearly similar to control rats confirming its efficacy in preventing morphological and physiological changes in proteins and lipids. The protective efficacy of BBR, as demonstrated in spectral analysis, is similar to the findings of the previous study where antioxidant supplementation reversed oxidative damage to biomolecules in rat hepatic tissue exposed to organophosphate pesticides (Singh et al. 2021).
Endogenous antioxidants are protective machinery for eliminating free radicals formed during biotransformation of pesticides and oxidative injuries to tissue. Depletion in enzymatic and non-enzymatic antioxidants disrupts balance between ROS production and their elimination during stress conditions. GSH is a prime non-enzymatic antioxidant involved in the elimination and detoxification of pesticide metabolites inside the liver. While SOD and catalase are enzymatic antioxidants that eliminate superoxides radicals and decompose H2O2 to oxygen and water, respectively, to detoxify free radicals. So, in the current study, the decreased activities of SOD and catalase and decline in GSH content following ACMP exposure indicate insufficient detoxification and cellular oxidative damage to rat liver tissue. Remarkably, treatment of BBR restored GSH content and maintained antioxidants enzymes activities in co-treated rats suggesting the antioxidative potential of BBR, possibly by scavenging free radicals. Our results are in agreement with some earlier in vivo and in vitro studies suggesting that administration of BBR assists the antioxidant defense system during pathology of diabetes (Zhou and Zhou 2011; Zhang et al. 2016), xenobiotics exposure (Eftekhari et al. 2020), heavy metals contamination (Othman et al. 2014; Hasanein et al. 2016), drug abuse (Mahmoud et al. 2017) and chemical exposure (Li et al. 2014; Eissa et al. 2018). In addition, the results are in accordance with previous observation showing that BBR modulates Nrf2 signaling pathway, which decreases ROS production and subsequently improves endogenous antioxidants level and imparts hepatoprotection in rats (Mahmoud et al. 2017; Deng et al. 2019)
Mounting evidences have suggested that oxidative stress and inflammatory response are closely associated. Studies have reported that ROS activates the translocation of transcription factor NF-κB from the cytosol to the nucleus, promoting the expression of other inflammatory cytokines (Xu et al. 2016). Exposure to ACMP is associated with the augmentation of inflammatory response via activation of proinflammatory mediators (NF-κB and TNF-α) in kidney tissue (Alhusaini et al. 2019; Erdemli et al. 2020). In agreement with earlier reports, the current study illustrated that ACMP exposure pronouncedly increased the levels of NF-κB, IL-1β, IL-6, IL-12 and TNF-α. BBR treatment to ACMP exposed rats significantly normalized the expression of NF-κB and associated proinflammatory cytokines suggesting the potential anti-inflammatory and modulatory effects of BBR. An earlier study has also observed similar effects of BBR on NF-κB and TNF-α following cisplatin exposure in mice kidney (Domitrović et al. 2013). Consistent with our observations, other studies have reported that berberine chloride downregulated the expression of proinflammatory cytokines, possibly either via blocking IL-6/STAT3/NF-κB signaling pathway (Zhu et al. 2019) or through up-regulating Nrf2/HO-1 signaling (Zhang et al. 2016; Mahmoud et al. 2017). In addition, BBR has alleviated thioacetamide-mediated hepatic inflammation in male rats through scavenging ROS and stimulating anti-inflammatory cytokines production (Eissa et al. 2018).
Histopathological examination of liver sections exposed to ACMP showed abnormalities as evident from the congestion of portal vein, leucocytes infiltration, dilation of central vein, vacuolization and broadening of sinusoidal space, which agrees with previous studies (Chakroun et al. 2016; Doltade et al. 2019; Karaca et al. 2019). The observed histopathological changes are direct shreds of oxidative stress and inflammation and support the biochemical and FTIR results of the present study. Meanwhile, in co-treated rats, BBR treatment significantly normalized the histopathological changes and preserved liver integrity, almost similar to control rats. Our findings are in concordance with earlier studies where BBR showed protection against drug-induced histopathological alterations in the liver of mice (Zhao et al. 2012) and rats (Mahmoud et al. 2017). The findings suggest that BBR imparts antioxidative potential against structural and molecular damage in liver tissue.