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Drug-induced liver injury (DILI) represents a major issue for pharmaceutical companies, being a potential cause of black-box warnings on marketed pharmaceuticals, or drug withdrawal from the market. Lipid accumulation in the liver also referred to as steatosis, may be secondary to impaired mitochondrial fatty acid oxidation (mtFAO). However, an overall causal relationship between drug-induced mtFAO inhibition and the occurrence of steatosis in patients has not yet been established with a high number of pharmaceuticals. Hence, 32 steatogenic and 13 nonsteatogenic drugs were tested for their ability to inhibit mtFAO in isolated mouse liver mitochondria. To this end, mitochondrial respiration was measured with palmitoyl-l-carnitine, palmitoyl-CoA + l-carnitine, or octanoyl- l-carnitine. This mtFAO tri-parametric assay was able to predict the occurrence of steatosis in patients with a sensitivity and positive predictive value above 88%. To get further information regarding the mechanism of drug-induced mtFAO impairment, mitochondrial respiration was also measured with malate/glutamate or succinate. Drugs such as diclofenac, methotrexate, and troglitazone could inhibit mtFAO secondary to an impairment of the mitochondrial respiratory chain, whereas dexamethasone, olanzapine, and zidovudine appeared to impair mtFAO directly. Mitochondrial swelling, transmembrane potential, and production of reactive oxygen species were also assessed for all compounds. Only the steatogenic drugs amiodarone, ketoconazole, lovastatin, and toremifene altered all these 3 mitochondrial parameters. In conclusion, our tri-parametric mtFAO assay could be useful in predicting the occurrence of steatosis in patients. The combination of this assay with other mitochondrial parameters could also help to better understand the mechanism of drug-induced mtFAO inhibition., (© The Author(s) 2024. Published by Oxford University Press on behalf of the Society of Toxicology. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

The aim of the current study was to investigate the potential of in ovo-fed amino acids (AA) to reduce the effects of heat stress on finishing broiler chickens. To achieve this, a total of 1,400 fertile hatching eggs were randomly distributed into 5 groups (n = 280/group) and injected with one of the following in ovo treatments on embryonic day 18: 52 µL of sterile diluent/egg (CTRL), CTRL + 1.0 mg of L-Leucine (T1), CTRL + 0.45 mg of leucine + 1.15 mg of methionine (T2), CTRL + 3.0 mg of methionine + 2.0 mg of cysteine (T3), and CTRL + 0.40 mg of leucine + 1.60 mg of methionine + 1.60 mg of cysteine (T4). After hatch, chicks were allocated according to a complete randomized block design comprising 2 thermal conditions: thermoneutral (24°C, 45% RH) and heat stress (34°C, 55-60% RH) with 5 pens/group/condition. The cyclical heat stress regimen (10 h/d) was then applied from d 29 to d 34. Compared to the CTRL group, T3 and T4 exhibited a higher BW during the starter phase (P < 0.001). T4 also had a lower feed conversion ratio (FCR) than CTRL during this same phase (P = 0.03). During the grower phase, males of all treatment groups consistently exhibited higher BW compared to the CTRL group, which was not observed among female birds (P Sex × TRT = 0.005). During the finisher phase, the in ovo treatment effect on performance was not significant. However, heat-stressed birds from treatment group T3 and T4 exhibited lower facial temperatures (P day × TRT < 0.001) as well as lower plasma (P condition x TRT = 0.039) and liver (P condition x TRT < 0.001) malonaldehyde concentrations compared to the CTRL group. In conclusion, in ovo-fed AA have the potential to modulate the effects of heat stress on finishing broiler chickens by limiting its detrimental consequences, including increased body temperature and oxidative damage., Competing Interests: DISCLOSURES The authors declare no conflicts of interest., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

St. John's Wort preparations are used for the treatment of mild to moderate depression. They are usually well tolerated but can cause adverse reactions including liver toxicity in rare cases. To date, the mechanism(s) underlying the hepatotoxicity of St. John's Wort extracts are poorly investigated. We studied the hepatocellular toxicity of hypericin and hyperforin as the two main ingredients of St. John's Wort extracts in HepG2 and HepaRG cells and compared the effects to citalopram (a synthetic serotonin uptake inhibitor) with a special focus on mitochondrial toxicity and oxidative stress. In HepG2 cells, hypericin was membrane-toxic at 100 µM and depleted ATP at 20 µM. In HepaRG cells, ATP depletion started at 5 µM. In comparison, hyperforin and citalopram were not toxic up to 100 µM. In HepG2 cells, hypericin decreased maximal respiration starting at 2 µM and mitochondrial ATP formation starting at 10 µM but did not affect glycolytic ATP production. Hypericin inhibited the activity of complex I, II and IV of the electron transfer system and caused mitochondrial superoxide accumulation in cells. The protein expression of mitochondrial superoxide dismutase 2 (SOD2) and thioredoxin 2 (TRX2) and total and reduced glutathione decreased in cells exposed to hypericin. Finally, hypericin diminished the mitochondrial DNA copy number and caused cell necrosis but not apoptosis. In conclusion, hypericin, but not hyperforin or citalopram, is a mitochondrial toxicant at low micromolar concentrations. This mechanism may contribute to the hepatotoxicity occasionally observed in susceptible patients treated with St. John's Wort preparations., Competing Interests: Declaration of Competing Interest 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., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Exposure to xenobiotics can adversely affect biochemical reactions, including hepatic bile acid synthesis. Bile acids are essential for dissolving lipophilic compounds in the hydrophilic environment of the gastrointestinal tract. The critical micellar concentration of bile acids depends on the Δ4-reduction stereochemistry, with the 3-oxo-5β-steroid-Δ4-dehydrogenase (AKR1D1) introducing the cis ring A/B conformation. Loss-of-function mutations in AKR1D1 cause hepatic cholestasis, which, if left untreated can progress into steatosis and liver cirrhosis. Furthermore, AKR1D1 is involved in clearing steroids with an A-ring Δ4-double bond. Here, we tested whether anabolic-androgenic steroids (AAS), often taken off-label at high doses, might inhibit AKR1D1, thereby potentially causing hepatotoxicity. A computational molecular model was established and used for virtual screening of the DrugBank database consisting of 2740 molecules, yielding mainly steroidal hits. Fourteen AAS were selected for in vitro evaluation, as such compounds can reach high hepatic concentrations in an abuse situation. Nandrolone, clostebol, methasterone, drostanolone, and methenolone inhibited to various extent the AKR1D1-mediated reduction of testosterone. Molecular modeling suggests that 9 out of 14 investigated AAS are competitive inhibitors. Moreover quantum mechanical calculations show that nadrolone and clostebol are substrates of AKR1D1 with different activation energy barriers for the hydrogen transfer from cofactor to the C5 position affecting their turnover. In this multidisciplinary approach, we established a molecular model of AKR1D1, identified several AAS as inhibitors, and described their binding mode. This approach may be applied to study other classes of inhibitors including non-steroidal compounds., Competing Interests: Declaration of Competing Interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Martin Smiesko reports financial support was provided by Swiss Centre for Applied Human Toxicology (SCAHT)., (Copyright © 2023. Published by Elsevier B.V.)

The heart muscle diseases hypertrophic (HCM) and dilated (DCM) cardiomyopathies are leading causes of sudden death and heart failure in young, otherwise healthy, individuals. We conducted genome-wide association studies and multi-trait analyses in HCM (1,733 cases), DCM (5,521 cases) and nine left ventricular (LV) traits (19,260 UK Biobank participants with structurally normal hearts). We identified 16 loci associated with HCM, 13 with DCM and 23 with LV traits. We show strong genetic correlations between LV traits and cardiomyopathies, with opposing effects in HCM and DCM. Two-sample Mendelian randomization supports a causal association linking increased LV contractility with HCM risk. A polygenic risk score explains a significant portion of phenotypic variability in carriers of HCM-causing rare variants. Our findings thus provide evidence that polygenic risk score may account for variability in Mendelian diseases. More broadly, we provide insights into how genetic pathways may lead to distinct disorders through opposing genetic effects.