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Introduction: Trypanosoma cruzi is a protozoan parasite that causes the tropical ailment known as Chagas disease, which has its origins in South America. Globally, it has a major impact on health and is transported by insect vector that serves as a parasite. Given the scarcity of vaccines and the limited treatment choices, we conducted a comprehensive investigation of core proteomics to explore a potential reverse vaccine candidate with high antigenicity., Methods: To identify the immunodominant epitopes, T. cruzi core proteomics was initially explored. Consequently, the vaccine sequence was engineered to possess characteristics of non-allergenicity, antigenicity, immunogenicity, and enhanced solubility. After modeling the tertiary structure of the human TLR4 receptor, the binding affinities were assessed employing molecular docking and molecular dynamics simulations (MDS)., Results: Docking of the final vaccine design with TLR4 receptors revealed substantial hydrogen bond interactions. A server-based methodology for immunological simulation was developed to forecast the effectiveness against antibodies (IgM + IgG) and interferons (IFN-g). The MDS analysis revealed notable levels of structural compactness and binding stability with average RMSD of 5.03 Aring;, beta-factor 1.09e+5 Å, Rg is 44.7 Aring; and RMSF of 49.50 Aring;. This is followed by binding free energies calculation. The system stability was compromised by the complexes, as evidenced by their corresponding Gibbs free energies of -54.6 kcal/mol., Discussion: Subtractive proteomics approach was applied to determine the antigenic regions of the T cruzi. Our study utilized computational techniques to identify B- and T-cell epitopes in the T. cruzi core proteome. In current study the developed vaccine candidate exhibits immunodominant features. Our findings suggest that formulating a vaccine targeting the causative agent of Chagas disease should be the initial step in its development., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Albaqami, Altharawi, Althurwi and Alharthy.)

Inflammatory responses and oxidative stress contribute to the pathogenesis of brain ischemia/reperfusion (IR) injury. Naturally occurring bioflavonoids possess antioxidant and anti-inflammatory properties. The phytochemicals of Juniperus sabina L. , known as "Abhal" in Saudi Arabia, have been studied and cupressuflavone (CUP) has been isolated as the major bioflavonoid. This study aimed to investigate the neuroprotective potential of CUP in reducing brain IR damage in rats and to understand probable mechanisms. After 60 min of inducing cerebral ischemia by closing the left common carotid artery (CCA), blood flow was restored to allow reperfusion. The same surgical procedure was performed on sham-operated control rats, excluding cerebral IR. CUP or vehicle was given orally to rats for 3 days prior to ischemia induction and for a further 3 days following reperfusion. Based on the findings of this study, compared to the IR control group, CUP-administered group demonstrated reduced neurological deficits, improved motor coordination, balance, and locomotor activity. Additionally, brain homogenates of IR rats showed a decrease in malondialdehyde (MDA) level, an increase in reduced glutathione (GSH) content, and an increase in catalase (CAT) enzyme activity following CUP treatment. CUP suppressed neuro-inflammation via reducing serum inflammatory cytokine levels, particularly those of tumor necrosis factor alpha (TNF-α), interleukin-6 (IL-6), and interleukin-1 beta (IL-1β) and enhancing the inflammatory cytokine levels, such as Nuclear factor kappa- B (NF-κB), TANK-binding kinase-1 (TBK1), and interferon beta (IFN-β) in brain tissues. Furthermore, CUP ameliorated the histological alterations in the brain tissues of IR rats. CUP significantly suppressed caspase-3 expression and downregulated the Toll-like receptor 4 (TLR4)/NF-κB signaling pathway as a result of suppressing High mobility group box 1 (HMGB1). To our knowledge, this is the first study to document the neuroprotective properties of CUP. Thus, the study findings revealed that CUP ameliorates IR-induced cerebral injury possibly by enhancing brain antioxidant contents, reducing serum inflammatory cytokine levels, potentiating the brain contents of TBK1 and IFN-β and suppressing the HMGB1/TLR-4 signaling pathway. Hence, CUP may serve as a potential preventive and therapeutic alternative for cerebral stroke., Competing Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2023 The Authors.)

The purpose of this study was to evaluate the effectiveness of samarcandin (SMR) in preventing testicular injury caused by ischemia/reperfusion (I/R) in rats. Rats were divided into 4 groups at random: the sham group, the T/D control group (CONT), the T/D group receiving SMR treatment at 10 mg/kg (SMR-10), and the T/D group receiving SMR treatment at 20 mg/kg (SMR-20). When compared to the CONT group, SMR improved the oxidant/antioxidant balance by reducing malondialdehyde (MDA), nitric oxide (NOx), and increasing reduced glutathione (GSH), gluta-thione peroxide (GSH-Px), and superoxide dismutase (SOD). Moreover, SMR increased the levels of the steroid hormones' testosterone (TST), follicle-stimulating hormone (FSH), and luteinizing hormone (LH) in the blood as well as controlled the inflammatory mediators; interleukin-6 (IL6), tumor necrosis factor alpha (TNF-α), and nuclear factor κB (NF-κB). Nevertheless, SMR-treated animals showed a considerable downregulation of the apoptotic marker caspase-3. The T/D-induced histopathological changes were reduced and Proliferating Cell Nuclear Antigen (PCNA) protein expression was enhanced by SMR. These effects are associated with upregulation of testicular (Nuclear factor erythroid 2-related factor 2 (Nrf2), Heme oxygenase-1 (HO-1), and downregulation of NF-κB mRNA expression levels. These findings suggest that SMR may be able to prevent T/D-induced testis damage by mainly regulating the expression of Nrf2 and NF-B, which seems to mediate its promising antioxidant, anti-inflammatory and antiapoptotic effects seen in this study., Competing Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2023 The Author(s).)

Antibiotic resistance is a major public health concern that has resulted in high healthcare costs, increased mortality, and the emergence of novel bacterial diseases. Cardiobacterium valvarum , an antibiotic-resistant bacterium, is one of the leading causes of heart disease. Currently, there is no licensed vaccination against C. valvarum . In this research, an in silico -based vaccine was designed against C. valvarum using reverse vaccinology, bioinformatics, and immunoinformatics techniques. 4206 core proteins, 2027 nonredundant proteins, and 2179 redundant proteins were predicted. Among nonredundant proteins, 23 proteins were predicted in an extracellular membrane, 30 in the outer membrane, and 62 in the periplasmic membrane region. After applying several subtractive proteomics filters, two proteins, TonB-dependent siderophore receptor and hypothetical protein, were chosen for epitope prediction. In the epitope selection phase, B and T-cellepitopes were analyzed and shortlisted for vaccine design. The vaccine model was designed by linking selected epitopes with GPGPG linkers to avoid flexibility. Furthermore, the vaccine model was linked to cholera toxin B adjuvant to induce a proper immune response. The docking approach was utilized to analyze binding affinity to immune cell receptors. Molecular docking results predicted 12.75 kcal/mol for a Vaccine with MHC-I, 6.89 for a vaccine with MHC-II, and 19.51 vaccine with TLR-4. The MMGBSA estimated -94, -78, and -76 kcal/mol for TLR-4 and vaccine, MHC-I and vaccine, and MHC-II and vaccine, while the MMPBSA analysis estimated -97, -61, and -72 kcal/mol for TLR-4 with the vaccine, MHC-I with vaccine, and MHC-II with a vaccine. Molecular dynamic simulation analysis revealed that the designed vaccine construct has proper stability with immune cell receptors as it is essential for inducing an immune response. In conclusion, we observed that the model vaccine candidate has the potency to induce an immune response in the host. However, the study is designed purely on a computational basis; hence, experimental validation is strongly recommended., Competing Interests: The author declares no conflict of interest., (Copyright © 2023 Faisal F. Albaqami et al.)

Bone graft techniques are used to compensate for bone loss in areas with deficient regeneration. However, matrix metalloproteases (MMPs) can limit bone formation by degrading extracellular matrices, which are required for bone regrowth. Noteworthily, rutin is a natural flavonoid compound that inhibits the genetic expression of various MMPs. Therefore, rutin may serve as an inexpensive and stable alternative to the growth factors used to accelerate dental bone graft healing. This study aimed to evaluate the potential of mixing rutin gel with allograft bone to accelerate the healing of bone defects in an in vivo rabbit model. Bone defects were surgically induced in New Zealand rabbits ( n = 3 per group) and subsequently treated with bone grafts along with rutin or control gel. Overall, treatment with rutin significantly prevented the expression of several MMPs and increased type III collagen in the gingiva around the surgical site. Additionally, rutin-treated animals showed enhanced bone formation with higher bone marrow content in the jawbone defect area compared with the control group. Taken together, these findings demonstrate that rutin gel, when added to bone grafts, quickly enhances bone formation and may serve as a suitable alternative to expensive growth factors for the same purpose.

A hemorrhagic fever caused by the Marburg virus (MARV) belongs to the Filoviridae family and has been classified as a risk group 4 pathogen. To this day, there are no approved effective vaccinations or medications available to prevent or treat MARV infections. Reverse vaccinology-based approach was formulated to prioritize B and T cell epitopes utilizing a numerous immunoinformatics tools. Potential epitopes were systematically screened based on various parameters needed for an ideal vaccine such as allergenicity, solubility, and toxicity. The most suitable epitopes capable of inducing immune response were shortlisted. Epitopes with population coverage of 100% and fulfilling set parameters were selected for docking with human leukocyte antigen molecules, and binding affinity of each peptide was analyzed. Finally, 4 CTL and HTL each while 6 B cell 16-mers were used for designing multiepitope subunit (MSV) and mRNA vaccine joined via suitable linkers. Immune simulations were used to validate the constructed vaccine's capacity to induce a robust immune response whereas molecular dynamics simulations were used to confirm epitope-HLA complex stability. Based on these parameter's studies, both the vaccines constructed in this study offer a promising choice against MARV but require further experimental verification. This study provides a rationale point to begin with the development of an efficient vaccine against Marburg virus; however, the findings need further experimental validation to confirm the computational finding of this study., Competing Interests: The authors declare no conflict of interest., (Copyright © 2023 Faisal F. Albaqami et al.)

Calligonum comosum is a perennial shrub growing and widely used in traditional medicinal system in Saudi Arabia. The total phenolic content and in vitro antioxidant activity were compared between the water extract (WE) and methanol extract (ME). The protective potential against acetic acid (AA) induced ulcerative colitis (UC) was also evaluated in rats. The obtained results showed that the total phenolic content of the WE and ME were 8.378 ± 0.738 and 33.819 ± 0.488 µg/mL. The antioxidant properties of the two extracts were directly influenced by their total phenolic contents. The ME with higher phenolic contents and stronger antioxidant power was more effective than the WE in protection against AA-induced colitis. Phytochemical study of the ME led to the identification of three flavonoid derivatives: (-)- epi -catechin, quercetin-3-O- α -l-arabinofuranoside (Avicularin) and quercetin-3-O- β -d-glucuronide-6″-methyl ester by various spectroscopic methods. (-)- Epi -catechin was the major component while the other two compounds were obtained in minute quantities. The anti-ulcerative colitis effect of the ME can be explained by the presence of the antioxidant flavonoids since AA-induced colitis featured by imbalance between oxidant and antioxidant substances. Further support of such explanation was provided by HPLC quantification of (-)- epi -catechin in the ME and WE. The percentage in ME was higher than the WE but the difference was higher in term of Total Phenolic Content (TPC). These results support the traditional use of C. comosum as anti-ulcerative colitis., Competing Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2023 Published by Elsevier B.V.)

Research has shown that chloroquine (CQ) can effectively help control COVID-19 infection. B 24 N 24 nanocage is a drug delivery system. Thus, through density functional theory, the present study analyzed pristine nanocage-CQ interaction and CQ interaction with Si- and Al -doped nanocage. The findings revealed that nanocage doping, particularly with Si and Al, yields more satisfactory drug delivery for CQ due to their greater electronic and energetic characteristics with CQ., Competing Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2023 Elsevier B.V. All rights reserved.)

Background: Parkinson's disease (PD) is a common age-related and slowly progressive neurodegenerative disease that affects approximately 1% of the elderly population. In recent years, phytocomponents have aroused considerable interest in the research for PD treatment as they provide a plethora of active compounds including antioxidant and anti-inflammatory compounds. Herein, we aimed to investigate the anti-Parkinson's effect of barbigerone, a natural pyranoisoflavone possessing antioxidant activity in a rotenone-induced rat model of PD., Methods: To evaluate antioxidant activity, a 0.5 mg/kg dose of rotenone was injected subcutaneously into rats. Barbigerone (10 and 20 mg/kg) was administered to rats for 28 days 1 h prior to rotenone. All behavioral parameters were assessed before sacrificing the rats. On the 29th day, all of the rats were humanely killed and assessed for biochemical changes in antioxidant enzymes (superoxide dismutase, glutathione, malondialdehyde, and catalase), neurotransmitter levels (dopamine, 5-hydroxyindoleacetic acid, serotonin, dihydroxyphenylacetic acid, and homovanillic acid levels), and neuroinflammatory cytokines [interleukin (IL)-1β, tumor necrosis factor-α, nuclear factor kappa B, and IL-6]., Results: The data presented in this study has shown that barbigerone attenuated rotenone-induced motor deficits including the rotarod test, catalepsy, akinesia, and open-field test. Additionally, barbigerone has shown improvements in the biochemical and neuroinflammatory parameters in the rotenone-induced rat model of PD., Conclusion: The results demonstrated that barbigerone exhibits antioxidant and anti-inflammatory actions via reducing oxidative stress and inflammatory cytokines. Altogether, these findings suggest that barbigerone could potentially be utilized as a therapeutic agent against PD., Competing Interests: The authors declare no competing financial interest., (© 2023 The Authors. Published by American Chemical Society.)

The current investigation assessed the effect of the eudesmanolid, Vulgarin (VGN), obtained from Artemisia judaica ( A. judaica ), on the antidiabetic potential of glibenclamide (GLB) using streptozotocin (STZ) to induce diabetes. Seven groups of rats were used in the study; the first group received the vehicle and served as normal control. The diabetic rats of the second to the fifth groups were treated with the vehicle (negative control), GLB at 5 mg/kg (positive control), VGN at 10 mg/kg (VGN-10) and VGN at 20 mg/kg (VGN-20), respectively. The diabetic rats of the sixth and seventh groups were administered combinations of GLB plus VGN-10 and GLB plus VGN-20, respectively. The diabetic rats treated with GLB plus VGN-20 combination showed marked improvement in the fasting blood glucose (FBG), insulin and glycated hemoglobin (HbA1c), as well as the lipid profile, compared with those treated with GLB alone. Further, the pancreatic tissues of the diabetic rats that received the GLB+VGN-20 combination showed superior improvements in lipid peroxidation and antioxidant parameters than those of GLB monotherapy. The insulin content of the β-cells was restored in all treatments, while the levels of glucagon and somatostatin of the α- and δ-endocrine cells were reduced in the pancreatic islets. In addition, the concurrent administration of GLB+VGN-20 was the most effective in restoring PEPCK and G6Pase mRNA expression in the liver. In conclusion, the results demonstrated that the GLB+VGN-20 combination led to greater glycemic improvement in diabetic rats compared with GLB monotherapy through its antioxidant effect and capability to modulate PEPCK and G6Pase gene expression in their livers.

Oxidative stress and inflammatory reaction play critical roles in ischemia/reperfusion (I/R) injury in the brain. β-carotene (βCAR) is a naturally occurring pigment present in fruits and vegetables that expresses antioxidant and anti-inflammatory activities. This study was conducted to investigate the involvement of Bcl2/Bax and NF-κB signaling pathways in the potential protective role of βCAR against brain injury in a middle cerebral artery occlusion (MCAO) rat model. A focal brain ischemia model was created for 2 h, followed by reperfusion. Rats were given 10 and 20 mg/kg of βCAR for 7 days orally before induction of ischemia, at the start of reperfusion, and 3 days after ischemia. Scores of neurological deficit were rated 24 h after induction of ischemia. Motor coordination and spontaneous coordinate activities were assessed using rotarod and activity cage, respectively. After 2 h of the last dose, the animals were killed and their brains were extracted for further examinations. The results of the study show that βCAR diminished the score of neurological deficits and ameliorated motor coordination, balance, and locomotor activity in the I/R control group. Further, βCAR resulted in diminution of malondialdehyde (MDA) and augmentation of reduced glutathione (GSH) contents, as well as the elevation of superoxide dismutase (SOD), glutathione peroxidase (GPx), and catalase (CAT) enzyme activities in the brain homogenates of I/R rats. βCAR treatment significantly reduced nuclear factor kappa B (NF-κB) brain content and myeloperoxidase (MPO) activity and ameliorated the histological alterations in the brain tissues. βCAR significantly suppressed Bcl-2-associated X protein (Bax) and caspase-3 expression, as well as upregulated B-cell lymphoma-2 (Bcl-2) expression, suggesting a neuroprotective potential via downregulating NF-kB and protecting the rat brain against the I/R-associated apoptotic injury.

Background: Europinidin is a derivative of delphinidin obtained from the plants Plumbago Europea and Ceratostigma plumbaginoides. This herb has wide medicinal applications in treating various diseases but there are very few studies available on this bioactive compound. Considering this background, the present study is designed for the evaluation of Europinidin against Parkinson’s disease. Aim: The investigation aims to assess the effect of Europinidin in the rotenone-activated Parkinson’s paradigm. Methods: To evaluate neuroprotective activity, rotenone (1.5 mg/kg s.c) and europinidin (10 mg/kg and 20 mg/kg) was administered in rats for 21 days. The behavioural parameters were performed before sacrificing the rats. On the 22nd day, all the rats were assessed for biochemical markers (SOD, GSH, MDA, Catalase), neurotransmitter levels (Dopamine, 5-HIAA, DOPAC, and HVA levels), and neuroinflammatory markers (IL-6, IL-1β and TNF-α). Results: It was found that rotenone produced significant (p < 0.001) oxidative damage, a cholinergic deficit, dopaminergic loss, and a rise in neuroinflammatory markers in rats. Conclusion: The study concludes that europinidin possesses anti-oxidant and anti-inflammatory properties. The results suggest the therapeutic role of europinidin against rotenone-activated behavioural, biochemical, and neuroinflammatory alterations in rats.

Epstein-Barr Virus (EBV) is a human pathogen that has a morbidity rate of 90% in adults worldwide. Infectious mononucleosis is caused by EBV replication in B cells and epithelial cells of the host. EBV has also been related to autoimmune illnesses, including multiple sclerosis and cancers like nasopharyngeal carcinomas and Burkitt's lymphoma. Currently, no effective medications or vaccinations are available to treat or prevent EBV infection. Thus, the current study focuses on a bioinformatics approach to design an mRNA-based multi-epitope (MEV) vaccine to prevent EBV infections. For this purpose, we selected six antigenic proteins from the EBV proteome based on their role in pathogenicity to predict, extract, and analyze T and B cell epitopes using immunoinformatics tools. The epitopes were directed through filtering parameters including allergenicity, toxicity, antigenicity, solubility, and immunogenicity assessment, and finally, the most potent epitopes able to induce T and B cell immune response were selected. In silico molecular docking of prioritized T cell peptides with respective Human Leukocytes Antigens molecules, were carried out to evaluate the individual peptide's binding affinity. Six CTL, four HTL, and ten linear B cell epitopes fulfilled the set parameters and were selected for MEV-based mRNA vaccine. The prioritized epitopes were joined using suitable linkers to improve epitope presentation. The immune simulation results affirmed the designed vaccine's capacity to elicit a proper immune response. The MEV-based mRNA vaccine constructed in this study offers a promising choice for a potent vaccine against EBV.

More than 90% of diabetic patients suffer from sexual dysfunction, including diminished sperm count, sperm motility, and sperm viability, and low testosterone levels. The effects of Momordica charantia (MC) were studied by estimating the blood levels of insulin, glucose, glycosylated hemoglobin (HbA1c), testosterone (TST), follicle-stimulating hormone (FSH), and luteinizing hormone (LH) in diabetic rats treated with 250 and 500 mg/kg b.w. of the total extract. Testicular antioxidants, epididymal sperm characteristics, testicular histopathology, and lesion scoring were also investigated. Testicular mRNA expression of apoptosis-related markers such as antiapoptotic B-cell lymphoma-2 (Bcl-2) and proapoptotic Bcl-2-associated X protein (Bax) were evaluated by real-time PCR. Furthermore, caspase-3 protein expression was evaluated by immunohistochemistry. MC administration resulted in a significant reduction in blood glucose and HbA1c and marked elevation of serum levels of insulin, TST, and gonadotropins in diabetic rats. It induced a significant recovery of testicular antioxidant enzymes, improved histopathological changes of the testes, and decreased spermatogenic and Sertoli cell apoptosis. MC effectively inhibited testicular apoptosis, as evidenced by upregulation of Bcl-2 and downregulation of Bax and caspase-3. Moreover, reduction in apoptotic potential in MC-treated groups was confirmed by reduction in the Bax/Bcl-2 mRNA expression ratio.

Cardiac hypertrophy is an independent risk factor of many cardiovascular diseases. Several cardiovascular protective properties of Cymbopogon proximus have been reported. However, no reports investigating the direct effect of C. proximus essential oil on the heart are available. The goal of this study was to explore the cardioprotective effect of C. proximus on cardiac hypertrophy and fibrosis. Male albino rats were administered C. proximus essential oil in the presence or absence of hypertrophic agonist isoproterenol. Cardiac hypertrophy and fibrosis were assessed using real-time polymerase chain reaction (PCR) and histological examination. Pre- treatment of rats with C. proximus decreased the ratio of heart weight to body weight and gene expression of hypertrophy markers atrial natriuretic peptide (ANP), brain natriuretic peptide (BNP), and β-myosin heavy chain (β-MHC), which were induced by isoproterenol. Moreover, C. proximus prevented the increase in gene expression of fibrosis markers procollagen I and procollagen III and alleviated the collagen volume fraction caused by isoproterenol. The pre- treatment with C. proximus essential oil conferred cardio-protection against isoproterenol- induced cardiac hypertrophy and fibrosis.

Several studies have demonstrated the role of cytochrome P450 (CYP) and its associated arachidonic acid (AA) metabolites in the anthracyclines-induced cardiac toxicity. However, the ability of daunorubicin (DNR) to induce cardiotoxicity through the modulation of CYP and its associated AA metabolites has not been investigated yet. Therefore, we hypothesized that DNR-induced cardiotoxicity is mediated through the induction of cardiotoxic hydroxyeicosatetraenoic acids and/or the inhibition of cardioprotctive epoxyeicosatrienoic acids (EETs). To test our hypothesis, Sprague-Dawley rats were treated with DNR (5 mg/kg i.p.) for 24 h, whereas human ventricular cardiomyocytes RL-14 cells were exposed to DNR in the presence and absence of 4-[[trans-4-[[(tricyclo[3.3.1.13,7]dec-1-ylamino)carbonyl]amino]cyclohexyl]oxy]-benzoic acid (tAUCB), a soluble epoxide hydrolase (sEH) inhibitor. Thereafter, real-time PCR, Western blot analysis and liquid chromatography-electron spray ionization mass spectroscopy were used to determine the level of gene expression, protein expression and AA metabolites, respectively. Our results showed that DNR-induced cardiotoxicity in vivo and in vitro as evidenced by the induction of hypertrophic and fibrotic markers. Moreover, the DNR-induced cardiotoxicity was associated with a dramatic increase in the formation of cardiac DHET/EET metabolites both in vivo and in RL-14 cells suggesting a sEH enzyme dependent mechanism. Interestingly, inhibition of sEH using tAUCB, a selective sEH inhibitor, significantly protects against DNR-induced cardiotoxicity. Mechanistically, the protective effect tAUCB was mediated through the induction of P50 nuclear factor-κB and the inhibition of phosphorylated p38. In conclusion, our study provides the first evidence that DNR induces cardiotoxicity through a sEH-mediated EETs degradation-dependent mechanism.

1. Dimethylarsinic acid (DMA(V)) is the major metabolite of inorganic arsenic in human body. Thus we investigated the effect of DMA(V) on the alteration of phase I (typified by Cyp1a) and phase II (typified by Nqo1) AhR-regulated genes in vivo. C57BL/6 mice received DMA(V) (13.3 mg/kg, i.p.) with or without TCDD (15 μg/kg, i.p.), thereafter the liver, lung, and kidney were harvested at 6 and 24 h post-treatment. 2. Results demonstrated that DMA(V) has no significant effect on Cyp1a mRNA and protein expression or catalytic activity in the liver. On the other hand, DMA(V) significantly potentiated the TCDD-mediated induction of Cyp1a mRNA and protein expression, with a subsequent potentiation of catalytic activity in the lung. Moreover, DMA(V) significantly inhibited the TCDD-mediated induction of Cyp1a mRNA and protein expression with subsequent inhibition of catalytic activity in the kidney. 3. Regarding to phase II AhR-regulated genes, DMA(V) has no significant effect on Nqo1 mRNA and protein expression, or activity neither in the liver, lung, or kidney. 4. In conclusion, the present work demonstrates for the first time that DMA(V) modulates AhR-regulated genes in a tissue- and enzyme-specific manner. This modulation may play a crucial role in arsenic-induced toxicity and carcinogenicity.

Numerous experimental studies have demonstrated the role of cytochrome P450 1B1 (CYP1B1) and its associated mid-chain hydroxyeicosatetraenoic acids (mid-chain HETEs) metabolite in the pathogenesis of cardiac hypertrophy. However, the ability of isoproterenol (ISO) to induce cardiac hypertrophy through mid-chain HETEs has not been investigated yet. Therefore, we hypothesized that ISO induces cardiac hypertrophy through the induction of CYP1B1 and its associated mid-chain HETE metabolites. To test our hypothesis, Sprague-Dawley rats were treated with ISO (5 mg/kg i.p.) for 12 and 72 h whereas, human ventricular cardiomyocytes RL-14 cells were exposed to 100 μM ISO in the presence and absence of 0.5 μM tetramethoxystilbene (TMS) a selective CYP1B1 inhibitor, or 25 nM CYP1B1-siRNA. Moreover, RL-14 cells were transiently transfected with the CRISPR-CYP1B1 plasmid. Thereafter, real-time PCR, western blot analysis, and liquid chromatography-electrospray ionization mass spectroscopy were used to determine the level of gene expression, protein expression, and mid-chain HETEs, respectively. Our results showed that ISO induced CYP1B1 protein expression and the level of cardiac mid-chain HETEs in vivo at pre-hypertrophic and hypertrophic stage. In vitro, inhibition of CYP1B1 using TMS or CYP1B1-siRNA significantly attenuates ISO-induced hypertrophy. Furthermore, overexpression of CYP1B1 significantly induced cellular hypertrophy and mid-chain HETEs metabolite. Mechanistically, the protective effect of TMS against cardiac hypertrophy was mediated through the modulation of superoxide anion, mitogen-activated protein kinases (MAPKs), and nuclear factor-κB (NF-κB). In conclusion, our study provides the first evidence that CYP1B1 and its associated mid-chain HETE metabolites are directly involved in the ISO-induced cardiac hypertrophy.

Doxorubicin (DOX) has been reported to be a very potent and effective anticancer agent. However, clinical treatment with DOX has been greatly limited due to its cardiotoxicity. Furthermore, several studies have suggested a role for cytochrome P450 1B1 (CYP1B1) and mid-chain hydroxyeicosatetraenoic acids (mid-chain HETEs) in DOX-induced cardiac toxicity. Therefore, we hypothesized that DOX induced cardiotoxicity is mediated through the induction of CYP1B1 and its associated mid-chain HETEs metabolite. To test our hypothesis, Sprague-Dawley rats and RL-14 cells were treated with DOX in the presence and absence of 2,3',4,5'-tetramethoxystilbene (TMS), a selective CYP1B1 inhibitor. Thereafter, cardiotoxicity parameters were determined using echocardiography, histopathology, and gene expression. Further, the level of mid-chain HETEs was quantified using liquid chromatography-electron spray ionization-mass spectrometry. Our results showed that DOX induced cardiotoxicity in vivo and in vitro as evidenced by deleterious changes in echocardiography, histopathology, and hypertrophic markers. Importantly, the TMS significantly reversed these changes. Moreover, the DOX-induced cardiotoxicity was associated with a proportional increase in the formation of cardiac mid-chain HETEs both in vivo and in our cell culture model. Interestingly, the inhibition of cardiotoxicity by TMS was associated with a dramatic decrease in the formation of cardiac mid-chain HETEs suggesting a mid-chain HETEs-dependent mechanism. Mechanistically, the protective effect of TMS against DOX-induced cardiotoxicity was mediated through the inhibition of mitogen activated protein kinases (MAPKs) and nuclear factor-κB (NF-κB). In conclusion, our study provides the first evidence that the inhibition of CYP1B1 and mid-chain HETE formation attenuate DOX-induced cardiotoxicity., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

Cytochrome P450 enzymes (P450s), along with their cardioprotective metabolites the epoxyeicosatrienoic acids (EETs) and cardiotoxic metabolite 20-hydroxyeicosatetraeonic acid (20-HETE), were found to be altered in cardiac hypertrophy; however, it is unclear whether these changes are causal or epiphenomenon. Therefore, we hypothesized that P450s and their metabolites play a crucial role in the initiation of cardiac hypertrophy. To test our hypothesis, rats and RL-14 cells were treated with the hypertrophic agonist isoproterenol for different time periods. Thereafter, in vivo heart function and wall thickness were assessed using echocardiography. Moreover, the role of P450 epoxygenases, ω-hydroxylases, and soluble epoxide hydrolase (sEH) were determined at mRNA, protein, and activity levels using real-time polymerase chain reaction, Western blot, and liquid chromatography-mass spectrometry, respectively. Our results show that in vivo and in vitro hypertrophy was initiated after 72 hours and 6 hours of isoproterenol treatment, respectively. Studies performed at the prehypertrophy phase showed a significant decrease in P450 epoxygenases along with a significant induction of sEH activity. Consequently, lower EET and higher dihydroxyeicosatrienoic acid levels were observed during this phase. However, significant increases in P450 ω-hydroxylase along with its associated metabolite, 20-HETE, were detected only in vivo. Interestingly, increasing EET levels by P450 epoxygenase induction, sEH inhibition, or exogenous administration of EET prevented the initiation of cardiac hypertrophy through a nuclear factor-κB-mediated mechanism. Taken together, these findings reveal a crucial role of P450 epoxygenases and EETs in the development of cardiac hypertrophy, which could uncover novel targets for prevention of heart failure at early stages., (Copyright © 2015 by The American Society for Pharmacology and Experimental Therapeutics.)

Introduction: RL-14 cells, human fetal ventricular cardiomyocytes, are a commercially available cell line that has been established from non-proliferating primary cultures derived from human fetal heart tissue. However, the expression of different drug metabolizing enzymes (DMEs) in RL-14 cells has not been elucidated yet. Therefore, the main objectives of the current work were to investigate the capacity of RL-14 cells to express different cytochrome P450 (CYP) isoenzymes and correlate this expression to primary cardiomyocytes., Methods: The expression of CYP isoenzymes was determined at mRNA, protein and catalytic activity levels using real time-PCR, Western blot analysis and liquid chromatography-electron spray ionization-mass spectrometry (LC-ESI-MS), respectively., Results: Our results showed that RL-14 cells constitutively express CYP ω-hydroxylases, CYP1A, 1B, 4A and 4F; CYP epoxygenases, CYP2B, 2C and 2J; in addition to soluble epoxide hydrolayse (EPHX2) at mRNA and protein levels. The basal expression of CYP ω-hydroxylases, epoxygenases and EPHX2 was supported by the ability of RL-14 cells to convert arachidonic acid to its biologically active metabolites, 20-hydroxyeicosatetraenoic acids (20-HETEs), 14,15-epoxyeicosatrienoic acids (14,15-EET), 11,12-EET, 8,9-EET, 5,6-EET, 14,15-dihydroxyeicosatrienoic acid (14,15-DHET), 11,12-DHET, 8,9-DHET and 5,6-DHET. Furthermore, RL-14 cells express CYP epoxygenases and ω-hydroxylase at comparable levels to those expressed in adult and fetal human primary cardiomyocytes cells implying the importance of RL-14 cells as a model for studying DMEs in vitro. Lastly, different CYP families were induced in RL-14 cells using 2,3,7,8-tetrachlorodibenzo-p-dioxin and fenofibrate at mRNA and protein levels., Discussion: The current study provides the first evidence that RL-14 cells express CYP isoenzymes at comparable levels to those expressed in the primary cells and thus offers a unique in vitro model to study DMEs in the heart., (Copyright © 2014 Elsevier Inc. All rights reserved.)

It has been previously shown that the cytochrome P450 (P450) modulator, fenofibrate, protects against cardiovascular diseases. P450 and their metabolites, epoxyeicosatrienoic acids (EETs) and 20-hydroxyeicosatetraenoic acid (20-HETE) were found to play an important role in cardiovascular diseases. Therefore, it is important to examine whether fenofibrate would modulate the cardiac P450 and its associated arachidonic acid metabolites and whether this modulation protects against isoproterenol-induced cardiac hypertrophy. For this purpose, male Sprague-Dawley rats were treated with fenofibrate (30 mg·kg·d), isoproterenol (4.2 mg·kg·d), or the combination of both. The expression of hypertrophic markers and different P450s along with their metabolites was determined. Our results showed that fenofibrate significantly induced the cardiac P450 epoxygenases, such as CYP2B1, CYP2B2, CYP2C11, and CYP2C23, whereas it decreased the cardiac ω-hydroxylase, CYP4A3. Moreover, fenofibrate significantly increased the formation of 14,15-EET, 11,12-EET, and 8,9-EET, whereas it decreased the formation of 20-HETE in the heart. Furthermore, fenofibrate significantly decreased the hypertrophic markers and the increase in heart-to-body weight ratio induced by isoproterenol. This study demonstrates that fenofibrate alters the expression of cardiac P450s and their metabolites and partially protects against isoproterenol-induced cardiac hypertrophy, which further confirms the role of P450s, EETs, and 20-HETE in the development of cardiac hypertrophy.

1. Arsenic (As(III)) toxicity has received increasing attention as human exposure to arsenic is associated with pulmonary, hepatic and renal toxicities. Therefore, in the present study, we investigated the effect of acute As(III) treatment on pulmonary, hepatic and renal cytochrome (CYP) P450-mediated arachidonic acid metabolism. 2. Our results demonstrated that acute As(III) treatment (12.5 mg/kg) altered CYP epoxygenases, CYP ω-hydroxylases and EPHX2 mRNA levels that were isozyme and tissue specific. 3. Furthermore, As(III) increased the formation of epoxyeicosatrienoic acids (EETs) in the kidney without affecting their levels in the lung or liver. In addition, acute As(III) treatment increased dihydroxyeicosatrienoic acid (DHETs) formation in the lung, while it did not affect liver DHETs formation and decreased kidney DHETs formation. 4. As(III) also increased total epoxygenases activity in the lung while it decreased its levels in the kidney and had no effect on the liver. Furthermore, As(III) increased 20-hydroxyeicosatetraenoic acid formation in the liver while it decreased its formation in the kidney. 5. Lastly, As(III) increased soluble epoxide hydrolase activity in the lung, while it decreased its levels in the kidney and had no effect on the liver. In conclusion, this is the first demonstration that As(III) alters arachidonic acid metabolism in a tissue specific manner.

We have previously shown that isoproterenol-induced cardiac hypertrophy causes significant changes to cytochromes P450 (CYPs) and soluble epoxide hydrolase (sEH) gene expression. Therefore, in this study, we examined the effect of isoproterenol in H9c2 cells, and the protective effects of 14,15-EET against isoproterenol-induced cellular hypertrophy. Isoproterenol was incubated with H9c2 cells for 24 and 48 h. To determine the protective effects of 14,15-EET, H9c2 cells were incubated with isoproterenol in the absence and presence of 14,15-EET. Thereafter, the expression of hypertrophic markers and different CYP genes were determined by real time-PCR. Our results demonstrated that isoproterenol significantly increased the expression of hypertrophic marker, atrial natriuretic peptide (ANP) and brain natriuretic peptide (BNP), parallel to a significant increase in cell surface area. Also, isoproterenol increased the mRNA expression of CYP1A1, CYP1B1, CYP2J3, CYP4F4 and CYP4F5, as well as the gene encoding sEH, EPHX2. On other hand, 14,15-EET significantly attenuated the isoproterenol-mediated induction of ANP, BNP, CYP1A1, CYP2J3, CYP4F4, CYP4F5 and EPHX2. Moreover 14,15-EET prevented the isoproterenol-mediated increase in cell surface area. Interestingly, 20-hydroxyeicosatetraenoic acid (20-HETE) treatment caused similar effects to that of isoproterenol treatment and induced cellular hypertrophy in H9c2 cells. In conclusion, isoproterenol induces cellular hypertrophy and modulates the expression of CYPs and EPHX2 in H9c2 cells. Furthermore, 14,15-EET exerts a protective effect against isoproterenol-induced cellular hypertrophy whereas, 20-HETE induced cellular hypertrophy in H9c2 cells., (Copyright © 2013 Elsevier Inc. All rights reserved.)

Background and Purpose: We have previously shown that isoprenaline-induced cardiac hypertrophy causes significant changes in the expression of cytochromes P450 (CYP) and soluble epoxide hydrolase (sEH) genes. Therefore, it is important to examine whether the inhibition of sEH by 1-(1-methanesulfonyl-piperidin-4-yl)-3-(4-trifluoromethoxy-phenyl)-urea (TUPS) will protect against isoprenaline-induced cardiac hypertrophy., Experimental Approach: Male Sprague-Dawley rats were treated with TUPS (0.65 mg kg(-1) day(-1), p.o.), isoprenaline (5 mg kg(-1) day(-1), i.p.) or the combination of both. In vitro H9c2 cells were treated with isoprenaline (100 μM) in the presence and absence of either TUPS (1 μM) or 11,12 EET (1 μM). The expression of hypertrophic, fibrotic markers and different CYP genes were determined by real-time PCR., Key Results: Isoprenaline significantly induced the hypertrophic, fibrotic markers as well as the heart to body weight ratio, which was significantly reversed by TUPS. Isoprenaline also caused an induction of CYP1A1, CYP1B1, CYP2B1, CYP2B2, CYP4A3 and CYP4F4 gene expression and TUPS significantly inhibited this isoprenaline-mediated effect. Moreover, isoprenaline significantly reduced 5,6-, 8,9-, 11,12- and 14,15-EET and increased their corresponding 8,9-, 11,12- and 14,15-dihydroxyeicosatrienoic acid (DHET) and the 20-HETE metabolites. TUPS abolished these isoprenaline-mediated changes in arachidonic acid (AA) metabolites. In H9c2 cells, isoprenaline caused a significant induction of ANP, BNP and EPHX2 mRNA levels. Both TUPS and 11,12-EET significantly decreased this isoprenaline-mediated induction of ANP, BNP and EPHX2., Conclusions and Implications: TUPS partially protects against isoprenaline-induced cardiac hypertrophy, which confirms the role of sEH and CYP enzymes in the development of cardiac hypertrophy., (© 2012 The Authors. British Journal of Pharmacology © 2012 The British Pharmacological Society.)

Acute arsenic (As(III)) exposure has been reported to cause cardiac toxicity, however this toxicity was never linked to the disturbance in cytochrome P450 (P450)-mediated arachidonic acid metabolism. Therefore, we investigated the effect of acute As(III) toxicity on the expression of P450 and soluble epoxide hydrolase (sEH) and their associated arachidonic acid metabolism in mice hearts. As(III) toxicity was induced by a single intraperitoneal injection of 12.5 mg/kg of As(III). Our results showed that As(III) treatment caused a significant induction of the cardiac hypertrophic markers in addition to Cyp1b1, Cyp2b, Cyp2c, Cyp4f, and sEH gene expression in mice hearts. Furthermore, As(III) increased sEH protein expression and activity in hearts with a consequent decrease in 11,12-, and 14,15-epoxyeicosatrienoic acids (EETs) formation. Whereas the formation of 8,9-, 11,12-, 14,15-dihydroxyeicosatrienoic acids (DHETs) was significantly increased. As(III) also increased sEH mRNA and protein expression levels in addition to the hypertrophic markers which was reversed by knockdown of sEH in H9c2 cells. In conclusion, acute As(III) toxicity alters the expression of several P450s and sEH enzymes with a consequent decrease in the cardioprotective EETs which may represent a novel mechanism by which As(III) causes progressive cardiotoxicity. Furthermore, inhibiting sEH might represent a novel therapeutic approach to prevent As(III)-induced hypertrophy.