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Protein–calixarenes binding plays an increasingly central role in many applications, spanning from molecular recognition to drug delivery strategies and protein inhibition. These ligands obey a specific bio-supramolecular chemistry, which can be revealed by computational approaches, such as molecular dynamics simulations. In this paper, we rely on all-atom, explicit-solvent molecular dynamics simulations to capture the electrostatically driven association of a phosphonated calix-[4]-arene with cytochome-C, which critically relies on surface-exposed paired lysines. Beyond two binding sites identified in direct agreement with the x-ray structure, the association has a larger structural impact on the protein dynamics. Then, our simulations allow a direct comparison to analogous calixarenes, namely, sulfonato, similarly reported as "molecular glue." Our work can contribute to a robust in silico predictive tool to assess binding sites for any given protein of interest for crystallization, with the specificity of a macromolecular cage whose endo/exo orientation plays a role in the binding. [ABSTRACT FROM AUTHOR]

Summary: In this work, deferoxamine (DFO) was injected into Tibetan sheep meat (distilled water as control) to reveal the effects of lysosomal Fe2+ on mitochondrial apoptosis and muscle tenderness. The reactive oxygen species (ROS) content, lysosomal membrane stability, cathepsins activity, mitochondrial membrane permeability transition pore (MPTP), mitochondrial membrane potential (MMP), cytochrome c (Cyt‐c) reduction level, caspase activity, myofibril fragmentation index (MFI) and shear force of Oula Tibetan sheep meat were measured at different ageing times. The results indicated that the ROS content, cathepsins activity, MPTP opening degree, lysosomal membrane stability, caspase‐3/9 activity and MFI of Tibetan sheep meat were significantly decreased after DFO treatment, whilst MMP, shear force and Cyt‐c reduction levels were noticeably increased. Furthermore, correlation analysis and principal component analysis showed that mitochondrial damage was positively correlated with meat tenderness. In sum, the study suggests that lysosomal Fe2+ may promote mitochondrial apoptosis and improve the tenderness of Tibetan sheep meat. [ABSTRACT FROM AUTHOR]

The Calotes genus presents a challenge due to the complexity of its species. However, research on the cryptic species complex within the Indonesian Calotes genus is still lacking. This study aims to determine the extent of genetic relationships and assess the potential existence of a species complex within the Indonesian genus Calotes (Daudin, 1802) (Squamata, Agamidae) using the partial Cytochrome c Oxidase Subunit 1 (COI) gene sequence as a molecular marker. Samples of the Indonesian Calotes genus in this study were collected from South Lampung (Lampung), Bogor (West Java), and Langkat (North Sumatra). By aligning 582 bp sequence similarities with reference sequences in GenBank, we confirmed that seven out of eight samples analyzed belonged to Calotes vultuosus, while one sample was identified as Calotes versicolor. The identity values ranged from 96 to 100%. The C. vultuosus samples in this study displayed lower genetic distances, ranging from 0 to 3%, with the reference C. vultuosus sequence from Indonesia compared to the reference sequence from India, which ranged from 6 to 9%. Phylogenetic tree reconstruction, utilizing both maximum likelihood with IQ-Tree and Bayesian Inference with BEAST methods, further supports these findings. It reveals distinct groupings between C. vultuosus samples from Indonesia and India. These results suggest the potential occurrence of a species complex within the Indonesian genus Calotes. Furthermore, the inclusion of eight COI gene sequences from two Calotes species in the GenBank database has the potential to confirm the existence of previously undocumented species in Indonesia. [ABSTRACT FROM AUTHOR]

Metal complexes based on N‐heterocyclic carbene (NHC) ligands have emerged as promising broad‐spectrum antitumor agents in bioorganometallic medicinal chemistry. In recent decades, studies on cytotoxic metal–NHC complexes have yielded numerous compounds exhibiting superior cytotoxicity compared to cisplatin. Although the molecular mechanisms of these anticancer complexes are not fully understood, some potential targets and modes of action have been identified. However, a comprehensive review of their biological mechanisms is currently absent. In general, apoptosis caused by metal–NHCs is common in tumor cells. They can cause a series of changes after entering cells, such as mitochondrial membrane potential (MMP) variation, reactive oxygen species (ROS) generation, cytochrome c (cyt c) release, endoplasmic reticulum (ER) stress, lysosome damage, and caspase activation, ultimately leading to apoptosis. Therefore, a detailed understanding of the influence of metal–NHCs on cancer cell apoptosis is crucial. In this review, we provide a comprehensive summary of recent advances in metal–NHC complexes that trigger apoptotic cell death via different apoptosis‐related targets or signaling pathways, including B‐cell lymphoma 2 (Bcl‐2 family), p53, cyt c, ER stress, lysosome damage, thioredoxin reductase (TrxR) inhibition, and so forth. We also discuss the challenges, limitations, and future directions of metal–NHC complexes to elucidate their emerging application in medicinal chemistry. [ABSTRACT FROM AUTHOR]

Summary The CYCS gene is highly evolutionarily conserved, with only a few pathogenic variants that cause thrombocytopenia‐4 (THC4). Here, we report a novel CYCS variant NM_018947.6: c.59C>T [NP_061820.1:p.(Thr20Ile)] segregating with thrombocytopenia in three generations of a Czech family. The phenotype of the patients corresponds to THC4 with platelets of normal size and morphology and dominant inheritance. Intriguingly, a gradual decline in platelet counts was observed across generations. CRISPR/Cas9‐mediated gene editing was used to introduce the new CYCS gene variant into a megakaryoblast cell line (MEG‐01). Subsequently, the adhesion, shape, size, ploidy, viability, mitochondrial respiration, cytochrome c protein (CYCS) expression, cell surface antigen expression and caspase activity were analysed in cells carrying the studied variant. Interestingly, the variant decreases the expression of CYCS while increasing mitochondrial respiration and the expression of CD9 cell surface antigen. Surprisingly, the variant abates caspase activation, contrasting with previously known effects of other CYCS variants. Some reports indicate that caspases may be involved in thrombopoiesis; thus, the observed dysregulation of caspase activity might contribute to thrombocytopenia. The findings significantly enhance our understanding of the molecular mechanisms underlying inherited thrombocytopenia and may have implications for diagnosis, prognosis and future targeted therapies. [ABSTRACT FROM AUTHOR]

Cytochrome c (CytC) is conjugated with a small molecule TG6 to give TG6–CytC, which is directly delivered into cytosol, triggering the release of endogenous CytC from mitochondria, and inducing a caspase-3-dependent apoptosis with an IC50 down to 2.4 nM. This work shows an efficient strategy for intracellular protein delivery. [ABSTRACT FROM AUTHOR]

Retinal neurodegeneration, characterized by retinal ganglion cell (RGC) death, is a leading cause of vision impairment and loss in blind diseases, such as glaucoma. Müller cells play crucial roles in maintaining retinal homeostasis. Thus, dysfunction of Müller cells has been implicated as one of the causes of retinal diseases. Yes-associated protein 1 (YAP), a nuclear effector of the Hippo pathway, regulates mammalian cell survival. In this study, we investigated the role of YAP in Müller cells during N-methyl-D-aspartic acid (NMDA)-induced excitotoxic RGC injury in rats. We found that YAP expression increased and was activated in Müller cells after NMDA-induced RGC injury. This YAP response was partly due to an increase in Yap mRNA levels, although it may be independent of the Hippo pathway and ß-TrCP-mediated YAP degradation. Morphological analysis revealed that verteporfin, a selective YAP inhibitor, exacerbated NMDA-induced RGC degeneration, suggesting that YAP activation in Müller cells contributes to RGC survival in NMDA-treated retinas. Studies in the rat Müller cell line (rMC-1) demonstrated that overexpression of YAP increased the levels of Bcl-xL, while verteporfin decreased the levels of Bcl-xL and cell viability and increased the levels of cytochrome c released from mitochondria and cleaved caspase-3. Finally, we found that Bcl-xL expression increased slightly in NMDA-treated retinas, whereas intravitreal injection of verteporfin suppressed this increase. Our findings suggest that activated YAP in Müller cells protects against NMDAinduced RGC injury by upregulating Bcl-xL expression. [ABSTRACT FROM AUTHOR]

Free interconversion of cytochrome C (CytC) between native ferrous (Cyt‐FeII) and oxidized ferric (CytC‐FeIII) states is necessary to maintain the respiratory function of mitochondria. Disturbances in CytC‐FeIII to total CytC ratio may indicate mitochondrial dysfunction and apoptosis. Thus, tracking CytC oxidation state delivers important information about cellular physiology. In this work, we propose a novel methodology based on resonance Raman (rR) imaging optimized uniquely to track and qualitatively analyze the transition of Cyt‐FeII to CytC‐FeIII within live cells without affecting their morphology. None of the commonly used excitation lines allows such clear‐cut differentiation, contrary to the 405 nm applied in this work. The presented methodology provides a novel pathway in the label‐free detection of ferrous and ferric heme proteins. [ABSTRACT FROM AUTHOR]

We designed a tris-catecholate-based siderophore mimic, H6-T-CATL, to selectively chelate iron(III) from mitochondrial cytochromes and other iron-containing proteins within cellular matrices. This strategic sequestration aims to trigger apoptosis or ferroptosis in cancer cells through the glutathione (GSH)-dependent release of reduced iron and subsequent ROS-mediated cytotoxicity. Synthesis of H6-T-CATL involved precise peptide coupling reactions. Using the Fe(III)-porphyrin model (Fe-TPP-Cl), akin to cytochrome c, we studied H6-T-CATL's ability to extract iron(III), yielding a binding constant (Krel) of 1014 for the resulting iron(III) complex (FeIII-T-CATL)3−. This complex readily underwent GSH-mediated reduction to release bioavailable iron(II), which catalyzed Fenton-like reactions generating hydroxyl radicals (˙OH), confirmed by spectroscopic analyses. Our research underscores the potential of H6-T-CATL to induce cancer cell death by depleting iron(III) from cellular metalloproteins, releasing pro-apoptotic iron(II). Evaluation across various cancer types, including normal cells, demonstrated H6-T-CATL's cytotoxicity through ROS production, mitochondrial dysfunction, and activation of ferroptosis and DNA damage pathways. These findings propose a novel mechanism for cancer therapy, leveraging endogenous iron stores within cells. H6-T-CATL emerges as a promising next-generation anticancer agent, exploiting iron metabolism vulnerabilities to induce selective cancer cell death through ferroptosis induction. [ABSTRACT FROM AUTHOR]

Triple‐negative breast cancer (TNBC) has short survival rates. This study aimed to prepare a novel formula of sorafenib, carbon nanotubes (CNTs), and folic acid to be tested as a drug delivery system targeting versus TNBC compared with free sorafenib and to evaluate the formula stability, in vitro pharmacodynamic, and in vivo pharmacokinetic properties. The formula preparation was done by the synthesis of polyethylene glycol bis amine linker, CNT PEGylation, folic acid attachment, and sorafenib loading. The prepared formula has been characterized using X‐ray diffraction, Flourier‐transform infrared, 1HNMR, UV, high resolution–transmission electron microscope, field emission scanning electron microscopy, and Zeta potential. In vitro studies included drug release determination, MTT assay, flow cytometry to determine the apoptotic stage with percent, cell cycle analysis, and apoptotic marker assays for caspase‐3, 8, 9, cytochrome c, and BCL‐2. The in vivo study was performed to determine bioavailability and half‐life in rats. The in vitro MTT antiproliferative assay revealed that the formula was threefold more cytotoxic toward TNBC cells than free sorafenib, and the flow cytometry showed a significant increase in apoptosis and necrosis. The formula has a greater inhibitory effect on BCL‐2 and a lessening effect on cytochrome c and caspases 3, 8, and 9 than free sorafenib. In vivo experiments proved that our novel formula was superior to free sorafenib by increasing bioavailability by eight times and prolonging the half‐life by three times. These results confirmed the successful preparation of the desired formula with better pharmacodynamic and pharmacokinetic properties. These promising results may show a novel therapeutic strategy for TNBC patients. [ABSTRACT FROM AUTHOR]

Background and Aim: Hydronidone (HDD) is a novel pirfenidone derivative developed initially to reduce hepatotoxicity. Our previous studies in animals and humans have demonstrated that HDD treatment effectively attenuates liver fibrosis, yet the underlying mechanism remains unclear. This study aimed to investigate whether HDD exerts its anti‐fibrotic effect by inducing apoptosis in activated hepatic stellate cells (aHSCs) through the endoplasmic reticulum stress (ERS)‐associated mitochondrial apoptotic pathway. Methods: The carbon tetrachloride (CCl4)‐ and 3,5‐diethoxycarbonyl‐1,4‐dihydrocollidine (DDC)‐induced liver fibrosis models were used for in vivo studies. In vitro studies were conducted using the human hepatic stellate cell line LX‐2. The apoptotic effect of HDD on aHSCs was examined using TUNEL and flow cytometry assays. The small interfering RNA (siRNA) technique was employed to downregulate the expression of interest genes. Results: HDD treatment significantly promoted apoptosis in aHSCs in both the CCl4‐ and DDC‐induced liver fibrosis in mice and LX‐2 cells. Mechanistic studies revealed that HDD triggered ERS and subsequently activated the IRE1α‐ASK1‐JNK pathway. Furthermore, the influx of cytochrome c from the mitochondria into the cytoplasm was increased, leading to mitochondrial dysfunction and ultimately triggering apoptosis in aHSCs. Notably, inhibition of IRE1α or ASK1 by siRNA partially abrogated the pro‐apoptotic effect of HDD in aHSCs. Conclusions: The findings of both in vivo and in vitro studies suggest that HDD induces apoptosis in aHSCs via the ERS‐associated mitochondrial apoptotic pathway, potentially contributing to the amelioration of liver fibrosis. [ABSTRACT FROM AUTHOR]

Cytochrome c (CytC), a one-electron carrier, transfers electrons from complex bc1 to cytochrome c oxidase (CcO) in the electron-transport chain. Electrostatic interaction with the partners, complex bc1 and CcO, is ensured by a lysine cluster near the heme forming the Universal Binding Site (UBS). We constructed three mutant variants of mitochondrial CytC with one (2Mut), four (5Mut), and five (8Mut) Lys->Glu substitutions in the UBS and some compensating Glu->Lys substitutions at the periphery of the UBS for charge compensation. All mutants showed a 4–6 times increased peroxidase activity and accelerated binding of cyanide to the ferric heme of CytC. In contrast, decomposition of the cyanide complex with ferrous CytC, as monitored by magnetic circular dichroism spectroscopy, was slower in mutants compared to WT. Molecular dynamic simulations revealed the increase in the fluctuations of Cα atoms of individual residues of mutant CytC compared to WT, especially in the Ω-loop (70–85), which can cause destabilization of the Fe...S(Met80) coordination link, facilitation of the binding of exogenous ligands cyanide and peroxide, and an increase in peroxidase activity. It was found that only one substitution K72E is enough to induce all these changes, indicating the significance of K72 and the Ω-loop (70–85) for the structure and physiology of mitochondrial CytC. In this work, we also propose using a ferro-ferricyanide buffer as a substrate to monitor the peroxidase activity of CytC. This new approach allows us to determine the rate of peroxidase activity at moderate (200 µM) concentrations of H2O2 and avoid complications of radical formation during the reaction. [ABSTRACT FROM AUTHOR]

The versatility of cytochrome P450 reductase (CPR) in transferring electrons to P450s from other closely related species has been extensively exploited, e.g., by using An. gambiae CPR (AgCPR), as a homologous surrogate, to validate the role of An. funestus P450s in insecticide resistance. However, genomic variation between the AgCPR and An. funestus CPR (AfCPR) suggests that the full metabolism spectrum of An. funestus P450s might be missed when using AgCPR. To test this hypothesis, we expressed AgCPR and AfCPR side-by-side with CYP6P9a and CYP6P9b and functionally validated their role in the detoxification of insecticides from five different classes. Major variations were observed within the FAD- and NADP-binding domains of AgCPR and AfCPR, e.g., the coordinates of the second FAD stacking residue AfCPR-Y456 differ from that of AgCPR-His456. While no significant differences were observed in the cytochrome c reductase activities, when co-expressed with their endogenous AfCPR, the P450s significantly metabolized higher amounts of permethrin and deltamethrin, with CYP6P9b-AfCPR membrane metabolizing α-cypermethrin as well. Only the CYP6P9a-AfCPR membrane significantly metabolized DDT (producing dicofol), bendiocarb, clothianidin, and chlorfenapyr (bioactivation into tralopyril). This demonstrates the broad substrate specificity of An. funestus CYP6P9a/-b, capturing their role in conferring cross-resistance towards unrelated insecticide classes, which can complicate resistance management. [ABSTRACT FROM AUTHOR]

Rifampicin is a strong inducer of cytochrome P450 (CYP3A4) and P-glycoprotein (P-gp/ABCB1), leading to profound drug–drug interactions. In contrast, the chemically related rifabutin does not show such pronounced induction properties in vivo. The aim of our study was to conduct a comprehensive analysis of the different induction potentials of rifampicin and rifabutin in primary human hepatocytes and to analyze the mechanism of potential differences. Therefore, we evaluated CYP3A4/ABCB1 mRNA expression (polymerase chain reaction), CYP3A4/P-gp protein expression (immunoaffinity–liquid chromatography–mass spectrometry, IA-LC-MS/MS), CYP3A4 activity (testosterone hydroxylation), and considered intracellular drug uptake after treatment with increasing rifamycin concentrations (0.01–10 µM). Furthermore, rifamycin effects on the protein levels of CYP2C8, CYP2C9, and CYP2C19 were analyzed (IA-LC-MS/MS). Mechanistic analysis included the evaluation of possible suicide CYP3A4 inhibition (IC50 shift assay) and drug impact on translational efficiency (cell-free luminescence assays). Rifabutin accumulated 6- to 15-fold higher in hepatocytes than rifampicin, but induced CYP3A4 mRNA comparably to rifampicin (e. g. rifampicin 61-fold vs. rifabutin 44-fold, 72 h). While rifampicin for example enhanced protein (10 µM: 21-fold) and activity levels considerably (53-fold), rifabutin only slightly increased CYP3A4 protein expression (10 µM: 3.3-fold) or activity (11-fold) compared to rifampicin after 72 h. Both rifamycins similarly influenced expression of other eliminating proteins. A potential CYP3A4 suicide inhibition by a specific rifabutin metabolite or disruption of ribosome function were excluded experimentally. In conclusion, the lack of protein enhancement, could explain rifabutin's weaker induction-related drug–drug interaction risk in vivo. [ABSTRACT FROM AUTHOR]

This article presents a study on the distribution of cytochrome P450 (CYP450) alleles in the Danish population. CYP450 enzymes are important for drug metabolism, and genetic variations in the genes encoding these enzymes can impact drug response. The study compared genotype data from different genotyping platforms and found that the CYP2C19*17 and CYP2C9*2 alleles were more common than the CYP2C19*2 and CYP2C9*3 alleles. The results provide valuable insights into the genetic variability of these enzymes in the Danish population and suggest that the Illumina GSA platform could be useful for identifying drug-gene interactions. [Extracted from the article]

The complex III of the mitochondrial respiration chain is an effective fungicidal target in plant pathogenic fungi. Especially the QoIs (FRAC Group 11 and 11A) fungicides cover a broad pathogen spectrum, that includes ascomycetes, basidiomycetes and oomycetes, by inhibiting the complex III at the outer binding site. Many of the target pathogens have developed resistance mediated by the target site mutation G143A. All fungicides that belong to Group 11 are affected by this mutation. Metyltetraprole has been recently developed, and it also binds at the Qo-site but is not affected by the G143A mutation and has therefore been classified differently (FRAC Group 11A). In experiments on solid medium with sequential exposure to complex III inhibitors, laboratory mutants of Zymoseptoria tritici with a resistance to metyltetraprole could be selected. Resistance to metyltetraprole was mediated by the target site mutation in the cytochrome B gene leading to the aa change Y132C. Metyltetraprole resistant isolates could only be generated from wildtype parents, but not from G143A parents. Further experiments used serial batch cultivations in liquid media with increasing fungicide concentrations. These were aimed to select double mutants that harboured both G143A and Y132C. Several different parental strains were treated with increasing concentrations of metyltetraprole, but no Y132C mutants were found when G143A parents were used. When Y132C mutants were used as the initial inoculum, and these were exposed to increasing concentrations of pyraclostrobin, the parental mutation was replaced by G143A within a few cycles; this could suggest that both mutations are incompatible. [ABSTRACT FROM AUTHOR]

Carbon black nanoparticles (CBNPs) are widely distributed in the environment and are increasingly recognized as a contributor in the development of cardiovascular disease. A variety of cardiac injuries and diseases result from structural and functional damage to cardiomyocytes. This study explored the mechanisms of CBNPs-mediated myocardial toxicity. CBNPs were given to mice through intra-tracheal instillation and it was demonstrated that the particles can be taken up into the cardiac tissue. Exposure to CBNPs induced cardiomyocyte inflammation and apoptosis. In combination with in vitro experiments, we showed that CBNPs increased the ROS and induced mitochondria fragmentation. Functionally, CBNPs-exposed cardiomyocyte exhibited depolarization of the mitochondrial membrane potential, release of cytochrome c , and activation of pro-apoptotic BAX, thereby initiating programmed cell death. On the other hand, CBNPs impaired autophagy, leading to the inadequate removal of dysfunctional mitochondria. The excess accumulation of damaged mitochondria further stimulated NF-κB activation and triggered the NLRP3 inflammasome pathway. Both the antioxidant N -acetylcysteine and the autophagy activator rapamycin were effective to attenuate the damage of CBNPs on cardiomyocytes. Taken together, this study elucidated the potential mechanism underlying CBNPs-induced myocardial injury and provided a scientific reference for the evaluation and prevention of the CBNPs-related heart risk. [Display omitted] • CBNPs can be taken up into cardiomyocytes through intra-tracheal instillation and induce myocardial toxicity. • ROS mediates myocardial cell damage through disruption of mitochondrial quality control. • Inhibition of mitophagy triggers the inflammation and mitochondrial-dependent apoptosis in myocardial cell. [ABSTRACT FROM AUTHOR]

Oxidative stress is a hallmark of secondary injury of spinal cord injuries. Controlling oxidative stress is crucial for mitigating secondary injury and promoting functional recovery after spinal cord injuries. Calycosin is an O‐methylated isoflavone with antioxidant activity. To evaluate the effect of calycosin on spinal cord neurons under oxidative stress and clarify the molecular mechanism underlying the effect, we tested the neuroprotective activity of calycosin in a primary spinal cord neuron culture model. We found that calycosin protected neurons from H2O2‐induced neuronal death in a dose‐dependent manner. Further experiments revealed that calycosin decreased H2O2‐induced mitochondrial fragmentation and mitochondrial membrane potential loss, and subsequently reduced H2O2‐triggered release of mitochondrial cytochrome c into the cytoplasm. In addition, calycosin inhibited H2O2‐induced reactive oxygen species generation and activation of NF‐κB signaling in spinal cord neurons. Furthermore, the expression of several antioxidant enzymes such as HO‐1, NQO1, GCLC, GCLM, TrxR1, and Trx1 was significantly promoted by calycosin. More importantly, we revealed that the Nrf2/Keap1 signal is crucial for the effect of calycosin, because calycosin increased the amount of nuclear Nrf2 while decreasing the amount of cytoplasmic Nrf2. Nrf2 knockdown with siRNA transfection abolished the neuroprotective effect of calycosin. Taken together, this study disclosed a novel mechanism by which calycosin combats oxidative stress. Our study thus sheds light on the potential clinical application of calycosin in SCI treatment. [ABSTRACT FROM AUTHOR]

The bioreduction characteristics and mechanisms of Cr(VI) onto Bacillus cereus RCr enhanced by ferric citrate were investigated. The optimum conditions were initial pH 9, temperature 40 °C, inoculation amount 4%, and glucose 3 g/L, respectively. The addition of 1.5 g/L ferric citrate increased the average reduction rate from 120.43 to 220.61 mg/(L∙h) compared with the control (without ferric citrate). The binding capacity of Cr(III) on the cell surface increased to 21%, in which the precipitates were mainly CrO(OH), Cr2O3, and FeCr2O4. Cell membrane was the main site of reduction, related important functional groups: − COOH, C-H, − NH2, C = C, and P-O. Fe(III) increased the yield of NADH and cytochrome c by approximately 48.51% and 68.63%, which significantly facilitated the electron generation and electron transfer, thus increasing the amount of electrons in the bioreduction of heavy metals by an average of 110%. Among the electrons obtained by Cr(VI), the proportion of indirect reduction mediated by Fe(III)/Fe(II) shuttle was 62% on average, whereas direct reduction mediated by reductase was 38%. These results may provide insights into the bioreduction process by bacteria enhanced by Fe(III) for detoxification of heavy metals with multiple valences, as an important step towards improving microbial remediation. [ABSTRACT FROM AUTHOR]

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Purpose: We aimed to investigate the urinary caspase-3 and cytochrome c levels in patients with unilateral antenatal hydronephrosis and to determine whether changes in urinary biomarker levels could be useful for both predicting the need for surgical intervention due to ureteropelvic junction obstruction (UPJO) and postoperative surgical success. Methods: Sixty-five children with a history of unilateral antenatal hydronephrosis and postnatal anteroposterior diameter ≥ 10 mm were included in this prospective case–control study between January 2013 and December 2021. The obstruction group consisted of 33 patients (28 boys, 84.8%) who underwent open dismembered pyeloplasty due to UPJO. The non-obstructive dilatation (NOD) group consisted of 32 patients (27 boys, 84.4%) with stable or improving hydronephrosis and no significant reduction in ipsilateral split renal function during follow-up, whereas 34 healthy children were enrolled in the study as a control group. Urinary urinary caspase-3 and cytochrome c levels using ELISA were measured. Results: The median preoperative urinary caspase-3 level was significantly higher in the obstruction group when compared to the NOD group (4.82 ng/mgCr vs. 2.61 ng/mgCr, p = 0.013) as well as the control group (4.82 ng/mgCr vs. 1.72 ng/mgCr, p = 0.002). In the postoperative period, urinary caspase-3 levels significantly decreased compared to preoperative measurements (4.82 ng/mgCr vs. 2.51 ng/mgCr, p = 0.006) and became similar to the control group (2.51 ng/mgCr vs. 1.72 ng/mgCr, p = 0.422). On the other hand, no significant differences were observed in urinary cytochrome c levels between the groups. All patients who underwent pyeloplasty achieved postoperative resolution in hydronephrosis and improved drainage on MAG-3, so none of the patients required re-do pyeloplasty. Postoperative decrease in caspase-3 level was found to be compatible with adequate urine drainage on MAG-3 scan. The cut-off value of urinary caspase-3 to predict patients requiring pyeloplasty was found to be 3.31 ng/mg creatinine with 63.6% sensitivity, 62.5% specificity (AUC = 0.679). In the multivariable analysis, urinary caspase-3 level (OR: 1.653, p = 0.019), anteroposterior pelvic diameter (OR: 1.401, p = 0.001), and split renal function on MAG-3 (OR: 1.277, p = 0.011) were found to be independent factors in determining patients who require surgery. Conclusion: Based on our preliminary findings, urinary caspase-3 levels could be a useful biomarker not only for predicting the need for surgical intervention but also for determining the postoperative surgical success in children with UPJO. [ABSTRACT FROM AUTHOR]

Background: The anti-diabetic drug, empagliflozin (EMPA), has many pleiotropic actions and is challenged recently to possess renoprotective properties. This renoprotective potential is proposed to be mediated via the activation of AMP-activated protein kinase (AMPK)/nuclear factor erythroid 2-related factor 2 (Nrf2) signaling pathways. This research investigated the renoprotective potential and the mechanistic pathway of EMPA against methotrexate (MTX)-induced nephrotoxicity and evaluated the role of AMPK by utilizing an AMPK inhibitor, dorsomorphin (Dorso). Methods: Thirty male Wistar rats, weighing 180–200 g, were divided equally into five groups. Group I represented the control group. Nephrotoxicity was induced in the remaining rats through the administration of a single intraperitoneal injection of MTX (20 mg/kg). Rats were then randomly assigned to: Group 2 (received MTX injection only); Group 3 (received MTX and EMPA 30 mg/kg/day); Group 4 (received MTX and Dorso 0.2 mg/kg/day), Group 5 (received MTX, Dorso, EMPA). After one week, blood samples were collected, the rats were euthanized, and renal tissues were harvested for biochemical and histomorphometric assessments. Results: MTX produced a significant rise in serum creatinine and tissue MDA levels; an increase in BAX, p53, cytochrome-c expression; a reduction in Bcl2 level; and disruption of renal microarchitecture. In contrast, EMPA therapy in group 3, resulted in a significant improvement of all these parameters, correlated with significant increase in AMPK phosphorylation and Nrf2 expression. Importantly, the co-administration of Dorso, in group 5, prevented EMPA's beneficial effects. Conclusion: EMPA has a potential protective effect against MTX-induced toxicity through the activation of the AMPK/Nrf2 signaling pathway. [ABSTRACT FROM AUTHOR]

Ciprofol is a novel short-acting intravenous anaesthetic developed in China that is mainly metabolized by cytochrome P450 2B6 (CYP2B6) and uridine diphosphate glucuronosyltransferase 1A9 (UGT1A9). Currently, insufficient evidence is available to support drug‒drug interactions between ciprofol and CYP2B6 inactivators. Here, we established a high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) method to assess the concentration of ciprofol and investigated the effects of psoralen and clopidogrel on the metabolism of ciprofol in liver microsomes and rats. In rat and human liver microsomes, the median inhibitory concentration (IC50) values of psoralen were 63.31 μmol·L-1 and 34.05 μmol·L-1, respectively, showing mild inhibitory effects on ciprofol metabolism, whereas the IC50 values of clopidogrel were 6.380 μmol·L-1 and 2.565 μmol·L-1, respectively, with moderate inhibitory effects. SD rats were randomly divided into three groups: psoralen (27 mg·kg-1), clopidogrel (7.5 mg·kg-1), and the same volume of 0.5% carboxy methyl cellulose. After 7 days, all rats were injected with 2.4 mg·kg-1 ciprofol. Compared with the control group, the AUC and MRT values of ciprofol in the psoralen and clopidogrel groups were significantly greater, whereas the CL values were significantly lower. In addition, the durations of loss of righting reflex (LORR) in the psoralen and clopidogrel groups were 16.1% and 23.0% longer than that in the control group, respectively. In conclusion, psoralen and clopidogrel inhibit ciprofol metabolism to different degrees and prolong the duration of LORR in rats. [ABSTRACT FROM AUTHOR]

Oxetane synthase (TmCYP1), a novel cytochrome P450 enzyme from Taxus×media cell cultures, has been functionally characterized to efficiently catalyse the formation of the oxetane ring in tetracyclic taxoids. Transient expression of TmCYP1 in Nicotiana benthamiana using 2α,5α,7β,9α,10β,13α‐hexaacetoxytaxa‐4(20),11(12)‐diene (1) as a substrate led to the production of a major oxetane derivative, 1β‐dehydroxybaccatin IV (1 a), and a minor 4β,20‐epoxide derivative, baccatin I (1 b). However, feeding the substrate decinnamoyltaxinine J (2), a 5‐deacetylated derivative of 1, yielded only 5α‐deacetylbaccatin I (2 b), a 4β,20‐epoxide. A possible reaction mechanism was proposed on the basis of substrate‐feeding, 2H and 18O isotope labelling experiments, and density functional theory calculations. This reaction could be an intramolecular oxidation‐acetoxyl rearrangement and the construction of the oxetane ring may occur through a concerted process; however, the 4β,20‐epoxide might be a shunt product. In this process, the C5‐O‐acetyl group in substrate is crucial for the oxetane ring formation but not for the 4(20)‐epoxy ring formation by TmCYP1. These findings provide a better understanding of the enzymatic formation of the oxetane ring in paclitaxel biosynthesis. [ABSTRACT FROM AUTHOR]

Two new pyrazolidine-3,5-dione derivatives namely (Z)-4-(2-(4-bromophenyl)hydrazono)-1-phenylpyrazolidine-3,5-dione (2) and (E)-1-phenyl-4-((pyridin-2-ylamino)methylene)pyrazolidine-3,5-dione (3) were synthesized and their structures were elucidated by single-crystal X-ray analysis along with spectroscopic techniques. Density functional theory (DFT) studies were carried out to examine the distribution of charge, and natural bond orbital (NBO) analysis and frontier molecular orbital (FMO) analysis were carried out. Hirshfeld surface analysis was carried out to examine the intermolecular interactions, revealing a prevalence of H⋯H interactions in the crystals of both molecules. NBO analysis was carried out to investigate the stabilization energy, while MEP analysis was conducted to explore regions susceptible to nucleophilic and electrophilic attacks. The molecules were screened for their medicinal potential based on physiochemical and pharmacokinetics including (gastrointestinal) GI absorption, blood brain barrier (BBB) permeation, skin permeation capability, Caco-2 permeability, Madin–Darby canine kidney (MDCK) permeability, drug metabolism by the cytochrome P450 (CYP) family of liver enzymes, and toxicity evaluations. The expected metabolic reactions in the real cell system and the resulting metabolites along with their polarities for the determination of the probability of excretion from the body and drug-likeness were studied. To investigate the anticancer potency of 2 and 3, both were docked with three enzymes as drug targets for anticancer studies including HER2 (PDB ID: 3WSQ), EGFR (PDB ID: 5WB7), and the extracellular domain of the Tdp enzyme (PDB ID: 6N0D). From the calculated binding energies and inhibition constants, 2 is a better inhibitor of the target substrates than 3. [ABSTRACT FROM AUTHOR]

Specimen identification at the species level is a critical challenge for understanding community structure and conserving biodiversity. The use of mitochondrial DNA barcodes, in addition to morphology, has proven to be a useful tool for earthworm identification, but it has also raised difficulties. Thus, approaches to delineating molecular operational taxonomic units (MOTUs) from cytochrome c oxidase subunit 1 (COI) or 16S rDNA sequence data often reveal more mitochondrial lineages than morphospecies, raising the question of whether these MOTUs should be used as taxonomic entities in community structure studies. Here, we used a newly acquired dataset of 576 COI barcodes of earthworms from Metropolitan France that were clustered in 36 MOTUS, corresponding to 21 morphospecies. We also incorporated data gathered from the literature to investigate this question. In order to match our MOTUs with already-described mitochondrial lineages, we downloaded reference sequences from the GenBank and BOLD platforms. In light of the difficulties encountered in recovering these sequences, we recommend that any new mitochondrial lineage described in a study be named consistently with previous works. Next, we analyzed the biological, ecological, and molecular data available in the literature on the different mitochondrial lineages that matched our MOTUs in order to determine if there was a consensus for species delimitation. Although the study specimens mainly belong to the Lumbricidae, which is one of the most studied families of earthworms, the data are often missing to determine if the MOTUs correspond to different species. Pending revision of the taxonomy, MOTUs for which mitochondrial divergence has been confirmed by morphological differences should be considered distinct taxonomic entities in community structure studies. In the absence of morphological differences and pending more data, we propose to distinguish for these analyses, within sexually reproducing morphospecies, the MOTUs for which mitochondrial divergence has been confirmed by multilocus nuclear data, while in the case of reproduction by parthenogenesis, ecological differences between MOTUs are necessary to consider them as different taxonomic entities. [ABSTRACT FROM AUTHOR]

Background: The marbling trait of cattle muscles, being a key indicator, played an important role in evaluating beef quality. Two breeds of cattle, namely a high-marbling (Angus) and a low-marbling (Nanyang) one, with their cattle muscles selected as our samples for transcriptome sequencing, were aimed to identify differentially expressed long non-coding RNAs (lncRNAs) and their targets associated with the marbling trait. Results: Transcriptome sequencing identified 487 and 283 differentially expressed mRNAs and lncRNAs respectively between the high-marbling (Angus) and low-marbling (Nanyang) cattle muscles. Twenty-seven pairs of differentially expressed lncRNAs-mRNAs, including eighteen lncRNAs and eleven target genes, were found to be involved in fat deposition and lipid metabolism. We established a positive correlation between fourteen up-regulated (NONBTAT000849.2, MSTRG.9591.1, NONBTAT031089.1, MSTRG.3720.1, NONBTAT029718.1, NONBTAT004228.2, NONBTAT007494.2, NONBTAT011094.2, NONBTAT015080.2, NONBTAT030943.1, NONBTAT021005.2, NONBTAT021004.2, NONBTAT025985.2, and NONBTAT023845.2) and four down-regulated (NONBTAT000850.2, MSTRG.22188.3, MSTRG.22188.4, and MSTRG.22188.5) lncRNAs and eleven genes related to adiponectin family protein (ADIPOQ), cytochrome P450 family (CYP4V2), 3-hydroxyacyl-CoA dehydratase family (HACD4), kinesin family (KIF5C), lipin family (LPIN2), perilipin family (PLIN1), prostaglandin family (PTGIS), solute carrier family (SLC16A7, SLC2213, and SLCO4C1), and containing a transmembrane domain protein family (VSTM1). Conclusions: These candidate genes and lncRNAs can be regarded as being responsible for regulating the marbling trait of cattle. lncRNAs along with the variations in intramuscular fat marbling established a foundation for elucidating the genetic basis of high marbling in cattle. [ABSTRACT FROM AUTHOR]

The kidney undergoes structural and physiological changes with age, predominantly studied in glomeruli and proximal tubules. However, limited knowledge is available about the impact of aging and anti‐aging interventions on distal tubules. In this study, we investigated the effects of cytochrome b5 reductase 3 (CYB5R3) overexpression and/or dietary nicotinamide riboside (NR) supplementation on distal tubule mitochondria. Initially, transcriptomic data were analyzed to evaluate key genes related with distal tubules, CYB5R3, and NAD+ metabolism, showing significant differences between males and females in adult and old mice. Subsequently, our emphasis focused on assessing how these interventions, that have demonstrated the anti‐aging potential, influenced structural parameters of distal tubule mitochondria, such as morphology and mass, as well as abundance, distance, and length of mitochondria‐endoplasmic reticulum contact sites, employing an electron microscopy approach. Our findings indicate that both interventions have differential effects depending on the age and sex of the mice. Aging resulted in an increase in mitochondrial size and a decrease in mitochondrial abundance in males, while a reduction in abundance, size, and mitochondrial mass was observed in old females when compared with their adult counterparts. Combining both the interventions, CYB5R3 overexpression and dietary NR supplementation mitigated age‐related changes; however, these effects were mainly accounted by NR in males and by transgenesis in females. In conclusion, the influence of CYB5R3 overexpression and dietary NR supplementation on distal tubule mitochondria depends on sex, genotype, and diet. This underscores the importance of incorporating these variables in subsequent studies to comprehensively address the multifaceted aspects of aging. [ABSTRACT FROM AUTHOR]

Introduction: Campylobacter spp. are a public health concern, yet there is still no effective vaccine or medicine available. Methods: Here, we developed a Campylobacter jejuni-specific antibody and found that it targeted a menaquinol cytochrome c reductase complex QcrC. Results: The antibody was specifically reactive to multiple C. jejuni strains including clinical isolates from patients with acute enteritis and was found to inhibit the energy metabolism and growth of C. jejuni. Different culture conditions produced different expression levels of QcrC in C. jejuni, and these levels were closely related not only to the energy metabolism of C. jejuni but also its pathogenicity. Furthermore, immunization of mice with recombinant QcrC induced protective immunity against C. jejuni infection. Discussion: Taken together, our present findings highlight a possible antibody- or vaccination-based strategy to prevent or control Campylobacter infection by targeting the QcrC-mediated metabolic pathway. [ABSTRACT FROM AUTHOR]

The structure of the sidechain crosslinked Tyr‐Leu‐Trp peptide produced by the biarylitide crosslinking cytochrome P450Blt from Micromonospora sp. MW‐13 has been reanalysed by a series of NMR, computational and isotope labelling experiments and shown to contain a C−N rather than a C−O bond. Additional in vivo experiments using such a modified peptide show there is a general tolerance of biarylitide crosslinking P450 enzymes for histidine to tryptophan mutations within their minimal peptide substrate sequences despite the lack of such residues noted in natural biarylitide gene clusters. This work further highlights the impressive ability of P450s from biarylitide biosynthesis pathways to act as biocatalysts for the formation of a range of sidechain crosslinked tripeptides. [ABSTRACT FROM AUTHOR]

The structure of the sidechain crosslinked Tyr‐Leu‐Trp peptide produced by the biarylitide crosslinking cytochrome P450Blt from Micromonospora sp. MW‐13 has been reanalysed by a series of NMR, computational and isotope labelling experiments and shown to contain a C−N rather than a C−O bond. Additional in vivo experiments using such a modified peptide show there is a general tolerance of biarylitide crosslinking P450 enzymes for histidine to tryptophan mutations within their minimal peptide substrate sequences despite the lack of such residues noted in natural biarylitide gene clusters. This work further highlights the impressive ability of P450s from biarylitide biosynthesis pathways to act as biocatalysts for the formation of a range of sidechain crosslinked tripeptides. [ABSTRACT FROM AUTHOR]

Species of the catfish genus Rineloricaria are common in the Paraíba do Sul River basin, in southeastern Brazil; here we present a revision of the taxonomic diversity and geographic distribution of the species of the genus inhabiting the basin, based on novel morphologic and molecular data. Five species delimitation methods based on cytochrome C oxidase subunit 1 nucleotide sequences yielded comparable molecular operational taxonomic units. The automatic barcode gap discovery, assemble species by automatic partitioning, barcode index number, and Bayesian implementation of the Poisson tree process methods supported the recognition of five evolutionary lineages. These taxonomic units were assigned to the previously described Rineloricaria nigricauda, Rineloricaria steindachneri, Rineloricaria zawadzkii, and Rineloricaria nudipectoris, and an additional undescribed species. R. zawadzkii was further divided into two intraspecific geographically structured lineages using the generalized mixed Yule coalescent delimitation method. A maximum likelihood phylogenetic analysis revealed that the five lineages from the Paraíba do Sul have closer relationships to different species from southern and southeastern Brazil (Ribeira de Iguape, Lagoa dos Patos, Uruguay, Paraguay, and Parana river basins) than to each other. Based on the analysis of lectotypes, recently collected material, and specimens from ichthyological collections, the poorly described R. nigricauda and R. steindachneri are redescribed following current descriptive standards. The undescribed species from the middle and upper Paraíba do Sul River basin is formally described. The description of a new species, along with the description of species boundaries in R. nigricauda and R. steindachneri, contributes to the knowledge of the ichthyofauna of the Paraíba do Sul River basin and adjacent coastal drainages of southeastern Brazil. An identification key for the species of Rineloricaria occurring in the Paraíba do Sul River basin is provided. [ABSTRACT FROM AUTHOR]

Summary: Thrombocytopenia 4 (THC4) is an autosomal‐dominant thrombocytopenia caused by mutations in CYCS, the gene encoding cytochrome c (CYCS), a small haeme protein essential for electron transport in mitochondria and cell apoptosis. THC4 is considered an extremely rare condition since only a few patients have been reported so far. These subjects presented mild thrombocytopenia and no or mild bleeding tendency. In this study, we describe six Italian families with five different heterozygous missense CYCS variants: p.Gly42Ser and p.Tyr49His previously associated with THC4, and three novel variants (p.Ala52Thr, p.Arg92Gly, and p.Leu99Val), which have been classified as pathogenic by bioinformatics and segregation analyses. Moreover, we supported functional effects of p.Ala52Thr and p.Arg92Gly on oxidative growth and respiratory activity in a yeast model. The clinical characterization of the 22 affected individuals, the largest series of THC4 patients ever reported, showed that this disorder is characterized by mild‐to‐moderate thrombocytopenia, normal platelet size, and function, low risk of bleeding, and no additional clinical phenotypes associated with reduced platelet count. Finally, we describe a significant correlation between the region of CYCS affected by mutations and the extent of thrombocytopenia, which could reflect different degrees of impairment of CYCS functions caused by different pathogenetic variants. [ABSTRACT FROM AUTHOR]

Cymoxanil (CYM) is a widely used synthetic acetamide fungicide, but its biochemical mode of action remains elusive. Since CYM inhibits cell growth, biomass production, and respiration in Saccharomyces cerevisiae, we used this model to characterize the effect of CYM on mitochondria. We found it inhibits oxygen consumption in both whole cells and isolated mitochondria, specifically inhibiting cytochrome c oxidase (CcO) activity during oxidative phosphorylation. Based on molecular docking, we propose that CYM blocks the interaction of cytochrome c with CcO, hampering electron transfer and inhibiting CcO catalytic activity. Although other targets cannot be excluded, our data offer valuable insights into the mode of action of CYM that will be instrumental in driving informed management of the use of this fungicide. [ABSTRACT FROM AUTHOR]

OBJECTIVE: Develop a cytochrome P450 (CYP) phenotyping cocktail for dogs using specific substrates for hepatic P450 enzymes CYP2B11, CYP2D15, and CYP3A12 and determine whether alternative sampling methods (saliva and urine) or single time point samples could be used instead of multiple blood sampling. ANIMALS: 12 healthy client-owned dogs (8 females and 4 males) from February 2019 to May 2019. METHODS: In a randomized crossover study, dogs received oral administration of the probe drug bupropion (75 mg), dextromethorphan (30 mg), or omeprazole (40 mg) alone or as a 3-drug combination (Program in Individualized Medicine [PrIMe] cocktail) to evaluate simultaneous phenotyping of CYP2B11, CYP2D15, and CYP3A12. Pharmacokinetic profiles for the probe drugs and metabolites were determined using plasma, saliva, and urine. Dogs received probe drugs alone or combined. Pharmacokinetic profiles up to 6 hours postdose for the probe drugs and metabolites were determined using plasma, saliva, and urine. RESULTS: The PrIMe cocktail was well tolerated. There was no statistically significant interaction between the probe drugs when administered together. Single time point plasma metabolic ratios at 4 hours postdose for all probe drugs strongly correlated with the corresponding area under the plasma concentration-versus-time curve (AUC) ratios. Saliva AUC metabolic ratios for CYP3A12 and CYP2D15 and 6-hour urine for CYP2B11 and CYP2D15 were correlated with plasma AUC ratios. CONCLUSIONS: The PrIMe cocktail can be used for simultaneous CYP phenotyping using plasma 4-hour single time point sample metabolic ratios. Saliva and urine sampling are suitable for specific CYPs. CLINICAL RELEVANCE: The PrIMe cocktail has potential as a useful tool in dogs to detect clinically important CYP-mediated drug-drug interactions, identify novel pharmacogenes, determine the drug-metabolizing phenotype of individual dogs, aid in individualized dose selection, and evaluate the effects of various physiological states on drug metabolism. [ABSTRACT FROM AUTHOR]

Pritelivir is a novel viral helicase‐primase inhibitor active against herpes simplex virus. In vitro drug‐drug interaction studies indicated that pritelivir has the potential for clinically relevant interactions on the cytochrome P450 (CYP) enzymes 2C8, 2C9, 3A4, and 2B6, and intestinal uptake transporter organic anion transporting polypeptide (OATP) 2B1 and efflux transporter breast cancer resistance protein (BCRP). This was evaluated in 2 clinical trials. In 1 trial the substrates flurbiprofen (CYP2C9), bupropion (CYP2B6), and midazolam (CYP3A4) were administered simultaneously as part of the Geneva cocktail, while the substrate celiprolol (OAPT2B1) was administered separately. In another trial, the substrates repaglinide (CYP2C8) and rosuvastatin (BCRP) were administered separately. Exposure parameters of the substrates and their metabolites (flurbiprofen and bupropion only) were compared after administration with or without pritelivir under therapeutic concentrations. The results of these trials indicated that pritelivir has no clinically relevant effect on the exposure of substrates for the intestinal uptake transporter OATP2B1 and the CYP enzymes 3A4, 2B6, 2C9, and 2C8, and has a weak inhibitory effect on the intestinal efflux transporter BCRP. In summary, the results suggest that pritelivir has a low drug‐drug interaction potential. [ABSTRACT FROM AUTHOR]

Pancreas ductal adenocarcinoma belongs to the most common cancers, but also to the tumors with the poorest prognosis. Here, we pharmacologically targeted a mitochondrial potassium channel, namely mitochondrial Kv1.3, and investigated the role of sphingolipids and mutated Kirsten Rat Sarcoma Virus (KRAS) in Kv1.3-mediated cell death. We demonstrate that inhibition of Kv1.3 using the Kv1.3-inhibitor PAPTP results in an increase of sphingosine and superoxide in membranes and/or membranes associated with mitochondria, which is enhanced by KRAS mutation. The effect of PAPTP on sphingosine and mitochondrial superoxide formation as well as cell death is prevented by sh-RNA-mediated downregulation of Kv1.3. Induction of sphingosine in human pancreas cancer cells by PAPTP is mediated by activation of sphingosine-1-phosphate phosphatase and prevented by an inhibitor of sphingosine-1-phosphate phosphatase. A rapid depolarization of isolated mitochondria is triggered by binding of sphingosine to cardiolipin, which is neutralized by addition of exogenous cardiolipin. The significance of these findings is indicated by treatment of mutated KRAS-harboring metastasized pancreas cancer with PAPTP in combination with ABC294640, a blocker of sphingosine kinases. This treatment results in increased formation of sphingosine and death of pancreas cancer cells in vitro and, most importantly, prolongs in vivo survival of mice challenged with metastatic pancreas cancer. Key messages: Pancreatic ductal adenocarcinoma (PDAC) is a common tumor with poor prognosis. The mitochondrial Kv1.3 ion channel blocker induced mitochondrial sphingosine. Sphingosine binds to cardiolipin thereby mediating mitochondrial depolarization. Sphingosine is formed by a PAPTP-mediated activation of S1P-Phosphatase. Inhibition of sphingosine-consumption amplifies PAPTP effects on PDAC in vivo. [ABSTRACT FROM AUTHOR]

Introduction: Data suggest malfunctioning mitochondria reduce oxidation and adenosine triphosphate (ATP) production, disrupting insulin signalling. Cytochrome c (CC), acylcarnitine (AC) and citrate synthase (CS) are essential components of the mitochondria machinery and can be used as reliable biomarkers of mitochondrial dysfunction. This study aimed to determine whether mitochondrial biomarkers (AC, CS and CC) are altered in individuals with type 2 diabetes mellitus (T2DM) and to examine the association between these biomarkers and insulin resistance. Methodology: A cross‐sectional observational study that recruited 170 participants (88 with T2DM and 82 without DM) was conducted. Blood samples were collected from the recruits and analysed for levels of fasting glucose (FBG), AC, CS, CC, insulin, total cholesterol, triglycerides (TG), glycated haemoglobin (HbA1c) and magnesium. Blood pressure (BP) and anthropometric characteristics of participants were also taken. Appropriate formulas were used to determine %body fat, body mass index (BMI), waist‐to‐hip ratio (WHR), the homeostatic model assessment for insulin resistance (HOMA‐IR) and insulin sensitivity (HOMA‐β). Results: Patients with T2DM had higher levels of CC, %body fat, FBG, TG, HbA1c, BMI and HOMA‐IR than controls (p < 0.05, respectively). Results showed a significant relationship between circulating CC levels versus HOMA‐β (r = −0.40, p = 0.001), CS (r = −0.70, p = 0.001) and AC (r = −0.72, p = 0.001) levels in patients with T2DM. The adjusted odds increased in the T2DM patients for VLDL (OR = 6.66, p = 0.002), HbA1c (OR = 6.50, p = 0.001), FPG (OR = 3.17, p = 0.001), TG (OR = 2.36, p = 0.010), being female (OR = 2.09, p = 0.020) and CC (OR = 1.14, p = 0.016). Conclusion: Overall, alterations in mitochondrial biomarkers, measured by AC, CC and CS, were observed in people with T2DM and showed a direct relationship with insulin resistance. These findings are potentially significant in Africa, although additional confirmation from a larger cohort is necessary. [ABSTRACT FROM AUTHOR]

The coastal marine stingrays (genus Fontitrygon) include two endemic and highly threatened species (Fontitrygon margaritella and Fontitrygon margarita) from the African Gulf of Guinea. However, the lack of robust diagnostic features due to similar external features hinders species identification, thus limiting species‐specific conservation efforts.The present study aims to examine the morphological characteristics and apply a DNA barcoding tool through amplifying mitochondrial cytochrome C oxidase subunit I (mtCOI) to accurately identify F. margaritella and F. margarita from coastal waters in Nigeria, Cameroon, Côte d'Ivoire, Guinea Bissau and Senegal.Morphological evaluation revealed differences in colour pattern, eye shade, mouth shape and pectoral radial count between F. margarita and F. margaritella. We generated 25 mtCOI barcode sequences that included F. margaritella (n = 22) and F. margarita (n = 3). Our query search in the National Center for Biotechnology Information (NCBI) GenBank and Barcode of Life Data system (BOLD) showed a sequence match of our newly collected F. margaritella with other F. margaritella in the global databases, while mtCOI sequences of F. margarita were new to the NCBI GenBank and BOLD databases. The phylogenetic clustering analyses based on the maximum‐likelihood tree grouped morphospecies into highly supported monophyletic units.Our study demonstrates the potential of combined molecular and morphological approaches in the identification of coastal marine stingrays from the Gulf of Guinea. Our study is helpful in future scientific studies and in forming future conservation plans. [ABSTRACT FROM AUTHOR]

BACKGROUND: Spodoptera litura is one of the most harmful lepidoptera pests in China, and is difficult to control due to its strong resistance to the current frequently used insecticide species. The requirement to develop pesticides with novel toxicology mechanisms to control S. litura is urgent. The quassinoid of bruceine D display outstanding systemic properties and strong insecticidal activity against S. litura, which possess notable application potential for integrative management of S. litura, but the mechanism of toxicity remains unclear. RESULTS: In this study, we found that bruceine D exerts potent growth inhibitory activity against S. litura, disrupting the ecdysone and juvenile hormone titers, and causing long‐term adverse effects. Association analysis between transcriptomics and metabolomics suggested that bruceine D affected the digestion and absorption capacity of S. litura larvae by inducing a strong oxidative stress response and cell apoptosis in the intestine. Further analysis demonstrated that bruceine D can inhibit the activities of digestive and antioxidant enzymes and induce malondialdehyde (MDA) and reactive oxygen species (ROS) overaccumulation in the midgut. Moreover, the protein level of Bax, cleavage caspase 3, and cytochrome c expressed in cytoplasm (cyto) were up‐regulated by bruceine D, while Bcl‐2 and cytochrome c expressed in mitochondria (mito) were down‐regulated. In addition, there was a noticeable increase in caspase‐3 protease activity. Histopathological observations revealed that bruceine D damages the structure of midgut epithelial cells and activates lysosomes, which subsequently disrupts the midgut tissue. CONCLUSION: Overall, our findings suggested that bruceine D induced excessive ROS accumulation in midgut epithelial cells. The resulting cell apoptosis disrupted midgut tissue, leading ultimately to reduced nutrient digestion and absorption in the midgut and the inhibition of larval growth. © 2024 Society of Chemical Industry. [ABSTRACT FROM AUTHOR]

Abstract Background The anti-diabetic drug, empagliflozin (EMPA), has many pleiotropic actions and is challenged recently to possess renoprotective properties. This renoprotective potential is proposed to be mediated via the activation of AMP-activated protein kinase (AMPK)/nuclear factor erythroid 2-related factor 2 (Nrf2) signaling pathways. This research investigated the renoprotective potential and the mechanistic pathway of EMPA against methotrexate (MTX)-induced nephrotoxicity and evaluated the role of AMPK by utilizing an AMPK inhibitor, dorsomorphin (Dorso). Methods Thirty male Wistar rats, weighing 180–200 g, were divided equally into five groups. Group I represented the control group. Nephrotoxicity was induced in the remaining rats through the administration of a single intraperitoneal injection of MTX (20 mg/kg). Rats were then randomly assigned to: Group 2 (received MTX injection only); Group 3 (received MTX and EMPA 30 mg/kg/day); Group 4 (received MTX and Dorso 0.2 mg/kg/day), Group 5 (received MTX, Dorso, EMPA). After one week, blood samples were collected, the rats were euthanized, and renal tissues were harvested for biochemical and histomorphometric assessments. Results MTX produced a significant rise in serum creatinine and tissue MDA levels; an increase in BAX, p53, cytochrome-c expression; a reduction in Bcl2 level; and disruption of renal microarchitecture. In contrast, EMPA therapy in group 3, resulted in a significant improvement of all these parameters, correlated with significant increase in AMPK phosphorylation and Nrf2 expression. Importantly, the co-administration of Dorso, in group 5, prevented EMPA’s beneficial effects. Conclusion EMPA has a potential protective effect against MTX-induced toxicity through the activation of the AMPK/Nrf2 signaling pathway.

[Display omitted] • Reducing carbon sources and increasing electron carriers led to "activated" SRB. • "Activated" SRB shows relatively high growth and metabolism. • "Activated" SRB promotes FeS formation compared with normal SRB. • Gauzy FeS induced by "activated" SRB reveals high Cr(VI) removal efficiency. • SRB-induced FeS show high stability, oxidizing resistance and anti-interference. Biosynthetic metal sulfides showed great application prospects in the environmental treatment against high-valence metal pollutants. However, the efficiency of biosynthesis, agglomeration during the reaction process, and the formation of the passivation layer during the reduction process were always the important factors restricting its development. This study explored the composition of the culture medium to promote the growth of highly corrosive sulfate-reducing bacteria (SRB) and its metabolism to produce FeS nanoparticles (NPs). The results showed that reducing the carbon source (CS) and adding electron carriers in the culture medium effectively promoted the production of small, dispersed, and loose FeS NPs in cells. At pH = 7, 24 °C and 10 min reaction time, 0.1 g/L FeS NPs produced by SRB under the conditions of 10 % CS with 10 ppm cytochrome c medium could achieve 100 % removal efficiency of 1 mM hexavalent chromium (Cr(VI)). Under this condition, FeS NPs could be produced by intracellular metabolism in SRB cells, and environmental factors such as pH, metal cations, and Cl− had little effect on the removal of Cr(VI) by this FeS NPs. The surface proteins of FeS NPs significantly enhanced their antioxidant properties. After 7 days of natural environment exposure, the Cr(VI) removal efficiency of FeS NPs was only reduced by 16 % compared with the initial sample. This work provided an in-depth understanding of Cr(VI) removal by SRB biosynthesis of FeS and contributes to the widespread application of FeS in the future. [ABSTRACT FROM AUTHOR]

The steepest-entropy-ascent quantum thermodynamic (SEAQT) framework is used to explore the influence of heating and cooling on polymer chain folding kinetics. The framework predicts how a chain moves from an initial non-equilibrium state to stable equilibrium along a unique thermodynamic path. The thermodynamic state is expressed by occupation probabilities corresponding to the levels of a discrete energy landscape. The landscape is generated using the Replica Exchange Wang–Landau method applied to a polymer chain represented by a sequence of hydrophobic and polar monomers with a simple hydrophobic-polar amino acid model. The chain conformation evolves as energy shifts among the levels of the energy landscape according to the principle of steepest entropy ascent. This principle is implemented via the SEAQT equation of motion. The SEAQT framework has the benefit of providing insight into structural properties under non-equilibrium conditions. Chain conformations during heating and cooling change continuously without sharp transitions in morphology. The changes are more drastic along non-equilibrium paths than along quasi-equilibrium paths. The SEAQT-predicted kinetics are fitted to rates associated with the experimental intensity profiles of cytochrome c protein folding with Rouse dynamics. [ABSTRACT FROM AUTHOR]

Neuroglobin (Ngb) is a cytosolic heme protein that plays an important role in protecting cells from apoptosis through interaction with oxidized cytochrome c (Cyt c) released from mitochondria. The interaction of reduced Ngb and oxidized Cyt c is accompanied by electron transfer between them and the reduction in Cyt c. Despite the growing number of studies on Ngb, the mechanism of interaction between Ngb and Cyt c is still unclear. Using Raman spectroscopy, we studied the effect of charged amino acid substitutions in Ngb and Cyt c on the conformation of their hemes. It has been shown that Ngb mutants E60K, K67E, K95E and E60K/E87K demonstrate changed heme conformations with the lower probability of the heme planar conformation compared to wild-type Ngb. Moreover, oxidized Cyt c mutants K25E, K72E and K25E/K72E demonstrate the decrease in the probability of methyl-radicals vibrations, indicating the higher rigidity of the protein microenvironment. It is possible that these changes can affect electron transfer between Ngb and Cyt c.

Molecular methods are currently some of the best-suited technologies for implementation in insect monitoring. However, the field is developing rapidly and lacks agreement on methodology or community standards. To apply DNA-based methods in large-scale monitoring, and to gain insight across commensurate data, we need easy-to-implement standards that improve data comparability. Here, we provide three recommendations for how to improve and harmonize efforts in biodiversity assessment and monitoring via metabarcoding: (i) we should adopt the use of synthetic spike-ins, which will act as positive controls and internal standards; (ii) we should consider using several markers through a multiplex polymerase chain reaction (PCR) approach; and (iii) we should commit to the publication and transparency of all protocol-associated metadata in a standardized fashion. For (i), we provide a ready-to-use recipe for synthetic cytochrome c oxidase spike-ins, which enable between-sample comparisons. For (ii), we propose two gene regions for the implementation of multiplex PCR approaches, thereby achieving a more comprehensive community description. For (iii), we offer guidelines for transparent and unified reporting of field, wet-laboratory and dry-laboratory procedures, as a key to making comparisons between studies. Together, we feel that these three advances will result in joint quality and calibration standards rather than the current laboratory-specific proof of concepts. This article is part of the theme issue 'Towards a toolkit for global insect biodiversity monitoring'. [ABSTRACT FROM AUTHOR]

Borgs are huge extrachromosomal elements (ECE) of anaerobic methane-consuming "Candidatus Methanoperedens" archaea. Here, we used nanopore sequencing to validate published complete genomes curated from short reads and to reconstruct new genomes. 13 complete and four near-complete linear genomes share 40 genes that define a largely syntenous genome backbone. We use these conserved genes to identify new Borgs from peatland soil and to delineate Borg phylogeny, revealing two major clades. Remarkably, Borg genes encoding nanowire-like electron-transferring cytochromes and cell surface proteins are more highly expressed than those of host Methanoperedens, indicating that Borgs augment the Methanoperedens activity in situ. We reconstructed the first complete 4.00 Mbp genome for a Methanoperedens that is inferred to be a Borg host and predicted its methylation motifs, which differ from pervasive TC and CC methylation motifs of the Borgs. Thus, methylation may enable Methanoperedens to distinguish their genomes from those of Borgs. Very high Borg to Methanoperedens ratios and structural predictions suggest that Borgs may be capable of encapsulation. The findings clearly define Borgs as a distinct class of ECE with shared genomic signatures, establish their diversification from a common ancestor with genetic inheritance, and raise the possibility of periodic existence outside of host cells. Borgs are huge extrachromosomal elements associated with anaerobic methane-consuming archaea. Here, the authors use nanopore sequencing to identify new Borgs from peatland soil and their hosts, uncover their DNA modifications and activities, and elucidate the evolutionary relationships of Borgs. [ABSTRACT FROM AUTHOR]

Excessive reactive oxygen species (ROS) generated during cerebral ischemic reperfusion (CIRI) are crucial for subsequent tissue damage. However, despite the potential benefits of antioxidants reported in clinical applications, few have proven effective in treating CIRI, particularly in the elderly. Epicatechin (EC) is a catechol flavonoid monomer derived from natural tea plants. Multiple phenolic hydroxyl groups give it strong antioxidant properties, which can not only degrade ROS through chemical reactions between hydroxyl and ROS but also enhance the activity of antioxidant enzymes in cells, and it is easy to penetrate the blood–brain barrier. But its antagonistic effect on age‐related CIRI and potential medicinal value are still unknown. Nuclear factor erythroid 2‐related factor 2 (Nrf2) is the most important transcription factor regulating the expression of antioxidant proteins in the body. This study first compared the pathological differences of the Nrf2 system in CIRI between 2‐month‐old and 12‐month‐old Sprague–Dawley (SD) rats. Subsequently, EC was administered to 12‐month‐old rat models of middle cerebral artery occlusion and reperfusion (MCAO/R) and senescent SH‐SY5Y cell models subjected to oxygen glucose deprivation/reoxygenation (OGD/R). EC treatment improved cerebral morphology and function; increased p‐Nrf2, heme oxygenase‐1 (HO‐1), superoxide dismutase (SOD), and glutathione (GSH) expression; reduced infarct volume; and neuronal apoptosis in senescent rats. Moreover, EC enhanced cellular activity and the expression of p‐Nrf2, HO‐1, and quinone oxidoreductase‐1 (NQO‐1) while decreasing ROS and malondialdehyde (MDA) levels and mitigating apoptosis in senescent SH‐SY5Y cells. These effects were reversed upon si‐Nrf2. In sum, we confirm that EC exerts neuroprotective effects by upregulating Nrf2/ARE and reducing oxidative stress, suggesting that EC may be a promising drug for the treatment of senile cerebral apoplexy. This study also provides a scientific basis for the development and selection of new drugs for ischemic stroke in elderly patients. [ABSTRACT FROM AUTHOR]

Photosystem II (PSII) catalyzes water oxidation and plastoquinone reduction by utilizing light energy. It is highly susceptible to photodamage under high-light conditions and the damaged PSII needs to be restored through a process known as the PSII repair cycle. The detailed molecular mechanism underlying the PSII repair process remains mostly elusive. Here, we report biochemical and structural features of a PSII-repair intermediate complex, likely arrested at an early stage of the PSII repair process in the green alga Chlamydomonas reinhardtii. The complex contains three protein factors associated with a damaged PSII core, namely Thylakoid Enriched Factor 14 (TEF14), Photosystem II Repair Factor 1 (PRF1), and Photosystem II Repair Factor 2 (PRF2). TEF14, PRF1 and PRF2 may facilitate the release of the manganese-stabilizing protein PsbO, disassembly of peripheral light-harvesting complexes from PSII and blockage of the QB site, respectively. Moreover, an α-tocopherol quinone molecule is located adjacent to the heme group of cytochrome b559, potentially fulfilling a photoprotective role by preventing the generation of reactive oxygen species. Here the authors show an intermediate state of the photosystem II (PSII) repair cycle, where three protein factors and an α-tocopherol quinone molecule are associated with a damaged PSII at distinct sites, fulfilling their specific functions in the repair process. [ABSTRACT FROM AUTHOR]