Metabolic models predict fotemustine and the combination of eflornithine/rifamycin and adapalene/cannabidiol for the treatment of gliomas.

Gliomas are the most common type of malignant brain tumors, with glioblastoma multiforme (GBM) having a median survival of 15 months due to drug resistance and relapse. The treatment of gliomas relies on surgery, radiotherapy and chemotherapy. Only 12 anti-brain tumor chemotherapies (AntiBCs), mostly alkylating agents, have been approved so far. Glioma subtype-specific metabolic models were reconstructed to simulate metabolite exchanges, in silico knockouts and the prediction of drug and drug combinations for all three subtypes. The simulations were confronted with literature, high-throughput screenings (HTSs), xenograft and clinical trial data to validate the workflow and further prioritize the drug candidates. The three subtype models accurately displayed different degrees of dependencies toward glutamine and glutamate. Furthermore, 33 single drugs, mainly antimetabolites and TXNRD1-inhibitors, as well as 17 drug combinations were predicted as potential candidates for gliomas. Half of these drug candidates have been previously tested in HTSs. Half of the tested drug candidates reduce proliferation in cell lines and two-thirds in xenografts. Most combinations were predicted to be efficient for all three glioma types. However, eflornithine/rifamycin and cannabidiol/adapalene were predicted specifically for GBM and low-grade glioma, respectively. Most drug candidates had comparable efficiency in preclinical tests, cerebrospinal fluid bioavailability and mode-of-action to AntiBCs. However, fotemustine and valganciclovir alone and eflornithine and celecoxib in combination with AntiBCs improved the survival compared to AntiBCs in two-arms, phase I/II and higher glioma clinical trials. Our work highlights the potential of metabolic modeling in advancing glioma drug discovery, which accurately predicted metabolic vulnerabilities, repurposable drugs and combinations for the glioma subtypes.

Elevated Na is a dynamic and reversible modulator of mitochondrial metabolism in the heart.

Elevated intracellular sodium Nai adversely affects mitochondrial metabolism and is a common feature of heart failure. The reversibility of acute Na induced metabolic changes is evaluated in Langendorff perfused rat hearts using the Na/K ATPase inhibitor ouabain and the myosin-uncoupler para-aminoblebbistatin to maintain constant energetic demand. Elevated Nai decreases Gibb's free energy of ATP hydrolysis, increases the TCA cycle intermediates succinate and fumarate, decreases ETC activity at Complexes I, II and III, and causes a redox shift of CoQ to CoQH2, which are all reversed on lowering Nai to baseline levels. Pseudo hypoxia and stabilization of HIF-1α is observed despite normal tissue oxygenation. Inhibition of mitochondrial Na/Ca-exchange with CGP-37517 or treatment with the mitochondrial ROS scavenger MitoQ prevents the metabolic alterations during Nai elevation. Elevated Nai plays a reversible role in the metabolic and functional changes and is a novel therapeutic target to correct metabolic dysfunction in heart failure.

Ethylene promotes fruit ripening initiation by downregulating photosynthesis, enhancing abscisic acid and suppressing jasmonic acid in blueberry (Vaccinium ashei).

BACKGROUND: Blueberry fruit exhibit atypical climacteric ripening with a non-auto-catalytic increase in ethylene coincident with initiation of ripening. Further, application of ethephon, an ethylene-releasing plant growth regulator, accelerates ripening by increasing the proportion of ripe (blue) fruit as compared to the control treatment. To investigate the mechanistic role of ethylene in regulating blueberry ripening, we performed transcriptome analysis on fruit treated with ethephon, an ethylene-releasing plant growth regulator. RESULTS: RNA-Sequencing was performed on two sets of rabbiteye blueberry ('Powderblue') fruit: (1) fruit from divergent developmental stages; and (2) fruit treated with ethephon, an ethylene-releasing compound. Differentially expressed genes (DEGs) from divergent developmental stages clustered into nine groups, among which cluster 1 displayed reduction in expression during ripening initiation and was enriched with photosynthesis related genes, while cluster 7 displayed increased expression during ripening and was enriched with aromatic-amino acid family catabolism genes, suggesting stimulation of anthocyanin biosynthesis. More DEGs were apparent at 1 day after ethephon treatment suggesting its early influence during ripening initiation. Overall, a higher number of genes were downregulated in response to ethylene. Many of these overlapped with cluster 1 genes, indicating that ethylene-mediated downregulation of photosynthesis is an important developmental event during the ripening transition. Analyses of DEGs in response to ethylene also indicated interplay among phytohormones. Ethylene positively regulated abscisic acid (ABA), negatively regulated jasmonates (JAs), and influenced auxin (IAA) metabolism and signaling genes. Phytohormone quantification supported these effects of ethylene, indicating coordination of blueberry fruit ripening by ethylene. CONCLUSION: This study provides insights into the role of ethylene in blueberry fruit ripening. Ethylene initiates blueberry ripening by downregulating photosynthesis-related genes. Also, ethylene regulates phytohormone-metabolism and signaling related genes, increases ABA, and decreases JA concentrations. Together, these results indicate that interplay among multiple phytohormones regulates the progression of ripening, and that ethylene is an important coordinator of such interactions during blueberry fruit ripening.

Anticancer Effects of Mitoquinone via Cell Cycle Arrest and Apoptosis in Canine Mammary Gland Tumor Cells.

Treating female canine mammary gland tumors is crucial owing to their propensity for rapid progression and metastasis, significantly impacting the overall health and well-being of dogs. Mitoquinone (MitoQ), an antioxidant, has shown promise in inhibiting the migration, invasion, and clonogenicity of human breast cancer cells. Thus, we investigated MitoQ's potential anticancer properties against canine mammary gland tumor cells, CMT-U27 and CF41.Mg. MitoQ markedly suppressed the proliferation and migration of both CMT-U27 and CF41.Mg cells and induced apoptotic cell death in a dose-dependent manner. Furthermore, treatment with MitoQ led to increased levels of pro-apoptotic proteins, including cleaved-caspase3, BAX, and phospho-p53. Cell cycle analysis revealed that MitoQ hindered cell progression in the G1 and S phases in CMT-U27 and CF41.Mg cells. These findings were supported using western blot analysis, demonstrating elevated levels of cleaved caspase-3, a hallmark of apoptosis, and decreased expression of cyclin-dependent kinase (CDK) 2 and cyclin D4, pivotal regulators of the cell cycle. In conclusion, MitoQ exhibits in vitro antitumor effects by inducing apoptosis and arresting the cell cycle in canine mammary gland tumors, suggesting its potential as a preventive or therapeutic agent against canine mammary cancer.

Treatment of Organophosphorus Poisoning with 6-Alkoxypyridin-3-ol Quinone Methide Precursors: Resurrection of Methylphosphonate-Aged Acetylcholinesterase.

Organophosphorus (OP) nerve agents inhibit acetylcholinesterase (AChE), creating a cholinergic crisis in which death can occur. The phosphylated serine residue spontaneously dealkylates to the OP-aged form, which current therapeutics cannot reverse. Soman's aging half-life is 4.2 min, so immediate recovery (resurrection) of OP-aged AChE is needed. In 2018, we showed pyridin-3-ol-based quinone methide precursors (QMPs) can resurrect OP-aged electric eel AChE in vitro, achieving 2% resurrection after 24 h of incubation (pH 7, 4 mM). We prepared 50 unique 6-alkoxypyridin-3-ol QMPs with 10 alkoxy groups and five amine leaving groups to improve AChE resurrection. These compounds are predicted in silico to cross the blood-brain barrier and treat AChE in the central nervous system. This library resurrected 7.9% activity of OP-aged recombinant human AChE after 24 h at 250 µM, a 4-fold increase from our 2018 report. The best QMP (1b), with a 6-methoxypyridin-3-ol core and a diethylamine leaving group, recovered 20.8% (1 mM), 34% (4 mM), and 42.5% (predicted maximum) of methylphosphonate-aged AChE activity over 24 h. Seven QMPs recovered activity from AChE aged with Soman and a VX degradation product (EA-2192). We hypothesize that QMPs form the quinone methide (QM) to realkylate the phosphylated serine residue as the first step of resurrection. We calculated thermodynamic energetics for QM formation, but there was no trend with the experimental biochemical data. Molecular docking studies revealed that QMP binding to OP-aged AChE is not the determining factor for the observed biochemical trends; thus, QM formation may be enzyme-mediated.

Relationship of Cytotoxic and Antimicrobial Effects of Triphenylphosphonium Conjugates with Various Quinone Derivatives.

Quinone derivatives of triphenylphosphonium have proven themselves to be effective geroprotectors and antioxidants that prevent oxidation of cell components with participation of active free radicals - peroxide (RO2·), alkoxy (RO·), and alkyl (R·) radicals, as well as reactive oxygen species (superoxide anion, singlet oxygen). Their most studied representatives are derivatives of plastoquinone (SkQ1) and ubiquinone (MitoQ), which in addition to antioxidant properties also have a strong antibacterial effect. In this study, we investigated antibacterial properties of other quinone derivatives based on decyltriphenylphosphonium (SkQ3, SkQT, and SkQThy). We have shown that they, just like SkQ1, inhibit growth of various Gram-positive bacteria at micromolar concentrations, while being less effective against Gram-negative bacteria, which is associated with recognition of the triphenylphosphonium derivatives by the main multidrug resistance (MDR) pump of Gram-negative bacteria, AcrAB-TolC. Antibacterial action of SkQ1 itself was found to be dependent on the number of bacterial cells. It is important to note that the cytotoxic effect of SkQ1 on mammalian cells was observed at higher concentrations than the antibacterial action, which can be explained by (i) the presence of a large number of membrane organelles, (ii) lower membrane potential, (iii) spatial separation of the processes of energy generation and transport, and (iv) differences in the composition of MDR pumps. Differences in the cytotoxic effects on different types of eukaryotic cells may be associated with the degree of membrane organelle development, energy status of the cell, and level of the MDR pump expression.

A highly bioactive THPC-crosslinked recombinant collagen hydrogel implant for aging skin rejuvenation.

Skin aging, a complex physiological progression marked by collagen degradation, poses substantial challenges in dermatology. Recombinant collagen emerges as a potential option for skin revitalization, yet its application is constrained by difficulties in forming hydrogels. We have for the first time developed a highly bioactive Tetrakis(hydroxymethyl) phosphonium chloride (THPC)-crosslinked recombinant collagen hydrogel implant for aging skin rejuvenation. THPC demonstrated superior crosslinking efficiency compared to traditional agents such as EDC/NHS and BDDE, achieving complete recombinant collagen crosslinking at minimal concentrations and effectively inducing hydrogel formation. THPC's four reactive hydroxymethyl groups facilitate robust crosslinking with triple helical recombinant collagen, producing hydrogels with enhanced mechanical strength, excellent injectability, increased stability, and greater durability. Moreover, the hydrogel exhibited remarkable biocompatibility and bioactivity, significantly promoting the proliferation, adhesion, and migration of human foreskin fibroblast-1. In photoaged mice skin models, the THPC-crosslinked collagen hydrogel implant notably improved dermal density, skin elasticity, and reduced transepidermal water loss, creating a conducive environment for fibroblast activity and healthy collagen regeneration. Additionally, it elevated superoxide dismutase (SOD) activity and displayed substantial anti-calcification properties. The THPC-crosslinked recombinant collagen hydrogel implant presents an innovative methodology in combating skin aging, offering significant promise in dermatology and tissue engineering.

Vanadium(V) complexes derived from triphenylphosphonium and hydrazides: cytotoxicity evaluation and interaction with biomolecules.

The development of coordination compounds with antineoplastic therapeutic properties is currently focused on non-covalent interactions with deoxyribonucleic acid (DNA). Additionally, the interaction profiles of these compounds with globular plasma proteins, particularly serum albumin, warrant thorough evaluation. In this study, we report on the interactions between biomolecules and complexes featuring hydrazone-type imine ligands coordinated with vanadium. The potential to enhance the therapeutic efficiency of these compounds through mitochondrial targeting is explored. This targeting is facilitated by the derivatization of ligands with triphenylphosphonium groups. Thus, this work presents the synthesis, characterization, interactions, and cytotoxicity of dioxidovanadium(V) complexes (C1-C5) with a triphenylphosphonium moiety. These VV-species are coordinated to hydrazone-type iminic ligands derived from (3-formyl-4-hydroxybenzyl)triphenylphosphonium chloride ([AH]Cl) and aromatic hydrazides ([H2L1]Cl-[H2L5]Cl). The structures of the five complexes were elucidated through single-crystal X-ray diffraction and vibrational spectroscopies, confirming the presence of dioxidovanadium(V) species in various geometries with degrees of distortion (τ = 0.03-0.50) and highlighting their zwitterionic characteristics. The molecular structural stability of C1-C5 in solution was ascertained using 1H, 19F, 31P, and 51V-nuclear magnetic resonance. Moreover, their interactions with biomolecules were evaluated using diverse spectroscopic methodologies and molecular docking, indicating moderate interactions (Kb ≈ 104 M-1) with calf thymus DNA in the minor groove and with human serum albumin, predominantly in the superficial IB subdomain. Lastly, the cytotoxic potentials of these complexes were assessed in keratinocytes of the HaCaT lineage, revealing that C1-C5 induce a reduction in metabolic activity and cell viability through apoptotic pathways.

The mitochondria-targeted antioxidant MitoQ ameliorates inorganic arsenic-induced DCs/Th1/Th2/Th17/Treg differentiation partially by activating PINK1-mediated mitophagy in murine liver.

Inorganic arsenic is a well-established environmental toxicant linked to acute liver injury, fibrosis, and cancer. While oxidative stress, pyroptosis, and ferroptosis are known contributors, the role of PTEN-induced kinase 1 (PINK1)-mediated mitophagy in arsenic-induced hepatic immunotoxicity remains underexplored. Our study revealed that acute arsenic exposure prompts differentiation of hepatic dendritic cells (DCs) and T helper (Th) 1, Th2, Th17, and regulatory T (Treg) cells, alongside increased transcription factors and cytokines. Inorganic arsenic triggered liver redox imbalance, leading to elevated alanine transaminase (ALT), hydrogen peroxide (H2O2), malondialdehyde (MDA), and activation of nuclear factor erythroid 2-related factor (Nrf2)/heme oxygenase-1 (HO-1) pathway. PINK1-mediated mitophagy was initiated, and its inhibition exacerbates H2O2 accumulation while promoting DCs/Th1/Th2/Treg differentiation in the liver of arsenic-exposed mice. Mitoquinone (MitoQ) pretreatment relieved arsenic-induced acute liver injury and immune imbalance by activating Nrf2/HO-1 and PINK1-mediated mitophagy. To our knowledge, this is the first report identifying PINK1-mediated mitophagy as a protective factor against inorganic arsenic-induced hepatic DCs/Th1/Th2 differentiation. This study has provided new insights on the immunotoxicity of inorganic arsenic and established a foundation for exploring preventive and therapeutic strategies targeting PINK1-mediated mitophagy in acute liver injury. Consequently, the application of mitochondrial antioxidant MitoQ may offer a promising treatment for the metalloid-induced acute liver injury.

Linking triphenylphosphonium cation to a bicyclic hydroquinone improves their antiplatelet effect via the regulation of mitochondrial function.

Platelets are the critical target for preventing and treating pathological thrombus formation. However, despite current antiplatelet therapy, cardiovascular mortality remains high, and cardiovascular events continue in prescribed patients. In this study, first results were obtained with ortho-carbonyl hydroquinones as antiplatelet agents; we found that linking triphenylphosphonium cation to a bicyclic ortho-carbonyl hydroquinone moiety by a short alkyl chain significantly improved their antiplatelet effect by affecting the mitochondrial functioning. The mechanism of action involves uncoupling OXPHOS, which leads to an increase in mitochondrial ROS production and a decrease in the mitochondrial membrane potential and OCR. This alteration disrupts the energy production by mitochondrial function necessary for the platelet activation process. These effects are responsive to the complete structure of the compounds and not to isolated parts of the compounds tested. The results obtained in this research can be used as the basis for developing new antiplatelet agents that target mitochondria.

4-Amidophenol Quinone Methide Precursors: Effective and Broad-Scope Nonoxime Reactivators of Organophosphorus-Inhibited Cholinesterases and Resurrectors of Organophosphorus-Aged Acetylcholinesterase.

Acetylcholinesterase (AChE) inhibition by organophosphorus (OP) compounds poses a serious health risk to humans. While many therapeutics have been tested for treatment after OP exposure, there is still a need for efficient reactivation against all kinds of OP compounds, and current oxime therapeutics have poor blood-brain barrier penetration into the central nervous system, while offering no recovery in activity from the OP-aged forms of AChE. Herein, we report a novel library of 4-amidophenol quinone methide precursors (QMP) that provide effective reactivation against multiple OP-inhibited forms of AChE in addition to resurrecting the aged form of AChE after exposure to a pesticide or some phosphoramidates. Furthermore, these QMP compounds also reactivate OP-inhibited butyrylcholinesterase (BChE) which is an in vivo, endogenous scavenger of OP compounds. The in vitro efficacies of these QMP compounds were tested for reactivation and resurrection of soluble forms of human AChE and BChE and for reactivation of cholinesterases within human blood as well as blood and brain samples from a humanized mouse model. We identify compound 10c as a lead candidate due to its broad-scope efficacy against multiple OP compounds as well as both cholinesterases. With methylphosphonates, compound 10c (250 µM, 1 h) shows >60% recovered activity from OEt-inhibited AChE in human blood as well as mouse blood and brain, thus highlighting its potential for future in vivo analysis. For 10c, the effective concentration (EC50) is less than 25 µM for reactivation of three different methylphosphonate-inhibited forms of AChE, with a maximum reactivation yield above 80%. Similarly, for OP-inhibited BChE, 10c has EC50 values that are less than 150 µM for two different methylphosphonate compounds. Furthermore, an in vitro kinetic analysis show that 10c has a 2.2- and 92.1-fold superior reactivation efficiency against OEt-inhibited and OiBu-inhibited AChE, respectively, when compared to an oxime control. In addition to 10c being a potent reactivator of AChE and BChE, we also show that 10c is capable of resurrecting (ethyl paraoxon)-aged AChE, which is another current limitation of oximes.

Elevated intracellular Ca2+ functions downstream of mitodysfunction to induce Wallerian-like degeneration and necroptosis in organophosphorus-induced delayed neuropathy.

Neurotoxic organophosphorus compounds can induce a type of delayed neuropathy in humans and sensitive animals, known as organophosphorus-induced delayed neuropathy (OPIDN). OPIDN is characterized by axonal degeneration akin to Wallerian-like degeneration, which is thought to be caused by increased intra-axonal Ca2+ concentrations. This study was designed to investigate that deregulated cytosolic Ca2+ may function downstream of mitodysfunction in activating Wallerian-like degeneration and necroptosis in OPIDN. Adult hens were administrated a single dosage of 750 mg/kg tri-ortho-cresyl phosphate (TOCP), and then sacrificed at 1 day, 5 day, 10 day and 21 day post-exposure, respectively. Sciatic nerves and spinal cords were examined for pathological changes and proteins expression related to Wallerian-like degeneration and necroptosis. In vitro experiments using differentiated neuro-2a (N2a) cells were conducted to investigate the relationship among mitochondrial dysfunction, Ca2+ influx, axonal degeneration, and necroptosis. The cells were co-administered with the Ca2+-chelator BAPTA-AM, the TRPA1 channel inhibitor HC030031, the RIPK1 inhibitor Necrostatin-1, and the mitochondrial-targeted antioxidant MitoQ along with TOCP. Results demonstrated an increase in cytosolic calcium concentration and key proteins associated with Wallerian degeneration and necroptosis in both in vivo and in vitro models after TOCP exposure. Moreover, co-administration with BATPA-AM or HC030031 significantly attenuated the loss of NMNAT2 and STMN2 in N2a cells, as well as the upregulation of SARM1, RIPK1 and p-MLKL. In contrast, Necrostatin-1 treatment only inhibited the TOCP-induced elevation of p-MLKL. Notably, pharmacological protection of mitochondrial function with MitoQ effectively alleviated the increase in intracellular Ca2+ following TOCP and mitigated axonal degeneration and necroptosis in N2a cells, supporting mitochondrial dysfunction as an upstream event of the intracellular Ca2+ imbalance and neuronal damage in OPIDN. These findings suggest that mitochondrial dysfunction post-TOCP intoxication leads to an elevated intracellular Ca2+ concentration, which plays a pivotal role in the initiation and development of OPIDN through inducing SARM1-mediated axonal degeneration and activating the necroptotic signaling pathway.

Triphenylphosphonium-functionalized dimeric BODIPY-based nanoparticles for mitochondria-targeting photodynamic therapy.

The dimerization of boron dipyrromethene (BODIPY) moieties is an appealing molecular design approach for developing heavy-atom-free triplet photosensitizers (PSs). However, BODIPY dimer-based PSs generally lack target specificity, which limits their clinical use for photodynamic therapy. This study reports the synthesis of two mitochondria-targeting triphenylphosphonium (TPP)-functionalized meso-ß directly linked BODIPY dimers (BTPP and BeTPP). Both BODIPY dimers exhibited solvent-polarity-dependent singlet oxygen (1O2) quantum yields, with maximum values of 0.84 and 0.55 for BTPP and BeTPP, respectively, in tetrahydrofuran. The compact orthogonal geometry of the BODIPY dimers facilitated the generation of triplet excited states via photoinduced charge separation (CS) and subsequent spin-orbit charge-transfer intersystem crossing (SOCT-ISC) processes and their rates were dependent on the energetic configuration between the frontier molecular orbitals of the two BODIPY subunits. The as-synthesized compounds were amphiphilic and hence formed stable nanoparticles (∼36 nm in diameter) in aqueous solutions, with a zeta potential of ∼33 mV beneficial for mitochondrial targeting. In vitro experiments with MCF-7 and HeLa cancer cells indicated the effective localization of BTPP and BeTPP within cancer-cell mitochondria. Under light irradiation, BTPP and BeTPP exhibited robust photo-induced therapeutic effects in both cell lines, with half-maximal inhibitory concentration (IC50) values of ∼30 and ∼55 nM, respectively.

Anticancer photodynamic activities of triphenylphosphine-labelled phthalocyanines and their bovine serum albumin-gold nanoparticles- complexes on melanoma A375 cell lines in vitro.

This work reports on the synthesis of triphenylphosphine-labelled cationic phthalocyanines (Pc) complexed with bovine serum albumin (BSA) and gold nanoparticles (Au NPs). This nano-complex (Pc-BSA-Au) is studied for its photodynamic therapy (PDT) activity compared to the non-complexed Pc counterpart. The photochemical properties and in vitro PDT efficacies of the Pc and the nano-complex were determined and are compared herein. The singlet oxygen (1O2) yields of the Pcs were determined and are reported in DMF. A singlet oxygen quantum yield of 0.47 was obtained for the Pcs. The PDT efficacies of the complexes were thereafter determined using malignant melanoma A375 cancer cell line in vitro. An increase in the cell toxicity was observed for cells treated with Pc-BSA-Au compared to those treated with the Pc alone. The cell survival percentages were 23.1% for cells treated with Pc-BSA-Au and 48.7% for those treated with Pc alone under PDT treatments.

Acute effects of MitoQ on vascular endothelial function are influenced by cardiorespiratory fitness and baseline FMD in middle-aged and older adults.

A key mechanism promoting vascular endothelial dysfunction is mitochondrial-derived reactive oxygen species (mtROS). Aerobic exercise preserves endothelial function in preclinical models by lowering mtROS. However, the effects of mtROS on endothelial function in exercising and non-exercising adults is limited. In a double-blind, randomized, placebo-controlled crossover study design 23 (10 M/13 F, age 62.1 ± 11.5 years) middle-aged and older (MA/O, ≥45 years) adults were divided into two groups: exercisers (EX, n = 11) and non-exercisers (NEX, n = 12). All participants had endothelial function (brachial artery flow-mediated dilatation, FMDBA) measured before and ∼1 h after mitoquinone mesylate (MitoQ) (single dose, 80 mg) and placebo supplementation. A two-way repeated measures ANOVA was used to determine the effects of MitoQ and placebo on FMDBA. Pearson correlations assessed the association between the change in FMDBA with MitoQ and baseline FMDBA and cardiorespiratory fitness (CRF). Compared with placebo, MitoQ increased FMDBA in NEX by + 2.1% (MitoQ pre: 4.9 ± 0.4 vs. post: 7.0 ± 0.4 %, P = 0.004, interaction) but not in EX (P = 0.695, interaction). MitoQ also increased endothelial function in adults with a FMDBA <6% (P < 0.0001, interaction) but not >6% (P = 0.855, interaction). Baseline FMDBA and CRF were correlated (r = 0.44, P = 0.037), whereas the change in FMDBA with MitoQ was inversely correlated with CRF (r = -0.66, P < 0.001) and baseline FMDBA (r = -0.73, P < 0.0001). The relationship between the change in FMDBA and baseline FMDBA remained correlated after adjusting for CRF (r = -0.55, P = 0.007). These data demonstrate that MitoQ acutely improves FMDBA in NEX and EX adults who have a baseline FMDBA <6%. KEY POINTS: A key age-related change contributing to increased cardiovascular disease (CVD) risk is vascular endothelial dysfunction due to increased mitochondrial-derived reactive oxygen species (mtROS). Aerobic exercise preserves endothelial function via suppression of mtROS in preclinical models but the evidence in humans is limited. In the present study, a single dose of the mitochondria-targeted antioxidant, mitoquinone mesylate (MitoQ), increases endothelial function in non-exercisers with lower cardiorespiratory fitness (CRF) but not in exercisers with higher CRF. The acute effects of MitoQ on endothelial function in middle-aged and older adults (MA/O) are influenced by baseline endothelial function independent of CRF. These data provide initial evidence that the acute MitoQ-enhancing effects on endothelial function in MA/O adults are influenced, in part, via CRF and baseline endothelial function.

The role of T-type calcium channels in elderly human vascular function: A pilot randomized controlled trial.

Endothelial dysfunction develops with age and may precede cardiovascular disease. Animal data suggest that T-type calcium channels play an important role in endothelial function, but data from humans are lacking. This study included 15 healthy, sedentary, elderly males for a double blinded, randomized controlled trial. For 8 weeks, they were given 40 mg/day of either efonidipine (L- and T-type calcium channel blocker (CCB)) or nifedipine (L-type CCB). Vascular function was evaluated by graded femoral arterial infusions of acetylcholine (ACh; endothelium-dependent vasodilator) and sodium nitroprusside (endothelium-independent vasodilator) both with and without co-infusion of N-acetylcysteine (NAC; antioxidant). We measured leg blood flow and mean arterial pressure and calculated leg vascular conductance to evaluate the leg vascular responses. Despite no significant change in blood pressure in either group, we observed higher leg blood flow responses (Δ 0.43 ± 0.45 l/min, P = 0.006) and leg vascular conductance (Δ 5.38 ± 5.67 ml/min/mmHg, P = 0.005) to intra-arterial ACh after efonidipine, whereas there was no change in the nifedipine group, and no differences between groups. We found no upregulation of endothelial nitric oxide synthase in vastus lateralis muscle biopsies within or between groups. Smooth muscle cell responsiveness was unaltered by efonidipine or nifedipine. Intravenous co-infusion of NAC did not affect endothelium-dependent vasodilatation in either of the CCB groups. These results suggest that 8 weeks' inhibition of T- and L-type calcium channels augments endothelium-dependent vasodilatory function in healthy elderly males. Further studies are required to elucidate if T-type calcium channel inhibition can counteract endothelial dysfunction.

MitoQ protects against carbon tetrachloride-induced hepatocyte ferroptosis and acute liver injury by suppressing mtROS-mediated ACSL4 upregulation.

Ferroptosis has been shown to be involved in carbon tetrachloride (CCl4)-induced acute liver injury (ALI). The mitochondrion-targeted antioxidant MitoQ can eliminate the production of mitochondrial reactive oxygen species (mtROS). This study investigated the role of MitoQ in CCl4-induced hepatocytic ferroptosis and ALI. MDA and 4HNE were elevated in CCl4-induced mice. In vitro, CCl4 exposure elevated the levels of oxidized lipids in HepG2 cells. Alterations in the mitochondrial ultrastructure of hepatocytes were observed in the livers of CCl4-evoked mice. Ferrostatin-1 (Fer-1) attenuated CCl4-induced hepatic lipid peroxidation, mitochondrial ultrastructure alterations and ALI. Mechanistically, acyl-CoA synthetase long-chain family member 4 (ACSL4) was upregulated in CCl4-exposed human hepatocytes and mouse livers. The ACSL4 inhibitor rosiglitazone alleviated CCl4-induced hepatic lipid peroxidation and ALI. ACSL4 knockdown inhibited oxidized lipids in CCl4-exposed human hepatocytes. Moreover, CCl4 exposure decreased the mitochondrial membrane potential and OXPHOS subunit levels and increased the mtROS level in HepG2 cells. Correspondingly, MitoQ pretreatment inhibited the upregulation of ACSL4 in CCl4-evoked mouse livers and HepG2 cells. MitoQ attenuated lipid peroxidation in vivo and in vitro after CCl4 exposure. Finally, MitoQ pretreatment alleviated CCl4-induced hepatocytic ferroptosis and ALI. These findings suggest that MitoQ protects against hepatocyte ferroptosis in CCl4-induced ALI via the mtROS-ACSL4 pathway.

Two natural compounds as potential inhibitors against the Helicobacter pylori and Acinetobacter baumannii IspD enzymes.

In a vast majority of bacteria, protozoa and plants, the methylerythritol phosphate (MEP) pathway is utilized for the synthesis of isopentenyl diphosphate (IDP) and dimethylallyl diphosphate (DMADP), which are precursors for isoprenoids. Isoprenoids, such as cholesterol and coenzyme Q, play a variety of crucial roles in physiological activities, including cell-membrane formation, protein degradation, cell apoptosis, and transcription regulation. In contrast, humans employ the mevalonate (MVA) pathway for the production of IDP and DMADP, rendering proteins in the MEP pathway appealing targets for antimicrobial agents. This pathway consists of seven consecutive enzymatic reactions, of which 4-diphosphocytidyl-2C-methyl-D-erythritol synthase (IspD) and 2C-methyl-D-erythritol 2,4-cyclodiphosphate synthase (IspF) catalyze the third and fifth steps, respectively. In this study, we characterized the enzymatic activities and protein structures of Helicobacter pylori IspDF and Acinetobacter baumannii IspD. Then, using the direct interaction-based thermal shift assay, we conducted a compound screening of an approved drug library and identified 27 hit compounds potentially binding to AbIspD. Among them, two natural products, rosmarinic acid and tanshinone IIA sodium sulfonate, exhibited inhibitory activities against HpIspDF and AbIspD, by competing with one of the substrates, MEP. Moreover, tanshinone IIA sodium sulfonate also demonstrated certain antibacterial effects against H. pylori. In summary, we identified two IspD inhibitors from approved ingredients, broadening the scope for antibiotic discovery targeting the MEP pathway.

Mn-Based Artificial Mitochondrial Complex "VI" Acts as an Electron and Free Radical Conversion Factory to Suppress Macrophage Inflammatory Response.

Disturbance in the mitochondrial electron transport chain (ETC) is a key factor in the emerging discovery of immune cell activation in inflammatory diseases, yet specific regulation of ETC homeostasis is extremely challenging. In this paper, a mitochondrial complex biomimetic nanozyme (MCBN), which plays the role of an artificial "VI" complex and acts as an electron and free radical conversion factory to regulate ETC homeostasis is creatively developed. MCBN is composed of bovine serum albumin (BSA), polyethylene glycol (PEG), and triphenylphosphine (TPP) hierarchically encapsulating MnO2 polycrystalline particles. It has nanoscale size and biological properties like natural complexes. In vivo and in vitro experiments confirm that MCBN can target the mitochondrial complexes of inflammatory macrophages, absorb excess electrons in ETC, and convert the electrons to decompose H2O2. By reducing the ROS and ATP bursts and converting existing free radicals, inhibiting NLRP3 inflammatory vesicle activation and NF-κB signaling pathway, MCBN effectively suppresses macrophage M1 activation and inflammatory factor secretion. It also demonstrates good inflammation control and significantly alleviates alveolar bone loss in a mouse model of ligation-induced periodontitis. This is the first nanozyme that mimics the mitochondrial complex and regulates ETC, demonstrating the potential application of MCBN in immune diseases.

Long-Term Neuropsychiatric Developmental Defects after Neonatal Organophosphate Exposure: Mitigation by Synthetic Neurosteroids.

Children are much more susceptible to the neurotoxic effects of organophosphate (OP) pesticides and nerve agents than adults. OP poisoning in children leads to acute seizures and neuropsychiatric sequela, including the development of long-term disabilities and cognitive impairments. Despite these risks, there are few chronic rodent models that use pediatric OP exposure for studying neurodevelopmental consequences and interventions. Here, we investigated the protective effect of the neurosteroid ganaxolone (GX) on the long-term developmental impact of neonatal exposure to the OP compound, diisopropyl-fluorophosphate (DFP). Pediatric postnatal day-28 rats were acutely exposed to DFP, and at 3 and 10 months after exposure, they were evaluated using a series of cognitive and behavioral tests with or without the postexposure treatment of GX. Analysis of the neuropathology was performed after 10 months. DFP-exposed animals displayed significant long-term deficits in mood, anxiety, depression, and aggressive traits. In spatial and nonspatial cognitive tests, they displayed striking impairments in learning and memory. Analysis of brain sections showed significant loss of neuronal nuclei antigen(+) principal neurons, parvalbumin(+) inhibitory interneurons, and neurogenesis, along with increased astrogliosis, microglial neuroinflammation, and mossy fiber sprouting. These detrimental neuropathological changes are consistent with behavioral dysfunctions. In the neurosteroid GX-treated cohort, behavioral and cognitive deficits were significantly reduced and were associated with strong protection against long-term neuroinflammation and neurodegeneration. In conclusion, this pediatric model replicates the salient features of children exposed to OPs, and the protective outcomes from neurosteroid intervention support the viability of developing this strategy for mitigating the long-term effects of acute OP exposure in children. SIGNIFICANCE STATEMENT: An estimated 3 million organophosphate exposures occur annually worldwide, with children comprising over 30% of all victims. Our understanding of the neurodevelopmental consequences in children exposed to organophosphates is limited. Here, we investigated the long-term impact of neonatal exposure to diisopropyl-fluorophosphate in pediatric rats. Neurosteroid treatment protected against major deficits in behavior and memory and was well correlated with neuropathological changes. Overall, this pediatric model is helpful to screen novel therapies to mitigate long-term developmental deficits of organophosphate exposure.