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Indian Journal of Pharmacology Mar 2024Sildenafil, a common over-the-counter pill often self-administered at high doses for erectile dysfunction, has been reported to rarely cause prothrombotic events and...
Off-target effect of high-dose sildenafil on adenosine 5'- diphosphate and collagen-induced platelet activation through mitogen-activated protein kinase pathway in treated BALB/C mice and in vitro experiments: A preliminary study.
Sildenafil, a common over-the-counter pill often self-administered at high doses for erectile dysfunction, has been reported to rarely cause prothrombotic events and sudden cardiac death in a few case reports. Therefore, we investigated the in vitro and in vivo effect of sildenafil treatment and dosage on platelet activation and mitogen-activated protein kinase (MAPK) phosphorylation. BALB/C mice were segregated into four groups, each having four mice each (control, low [3.25 mg/kg], medium [6.5 mg/kg], and high [13 mg/kg] sildenafil), and after the treatment, blood was drawn from each mouse and washed platelets prepared. Washed platelets were incubated with CD41 PE-Cy7 and Phospho-p38 MAPK PE antibodies and analyzed using a flow cytometer for platelet activation and adenosine 5'- diphosphate (ADP)/collagen-induced MAPK phosphorylation. Washed platelets obtained from the venous blood of 18 human volunteers, were incubated with PAC-1 FITC and Phospho-p38 MAPK PE antibodies, and platelet activation (ADP and collagen), followed by flow cytometry analysis. There was a significant increase in both platelet activation as well as MAPK phosphorylation in the presence of collagen in the high-dose (13 mg/kg) sildenafil group (P = 0.000774). Further, increased platelet activation was observed in samples that were treated with high-dose sildenafil as compared to the untreated samples (P < 0.00001). These studies show the risk of prothrombotic episodes in patients on high-dose sildenafil (100 mg), in those with even mild endothelial dysfunction due to ADP, and collagen-induced platelet activation through MAPK phosphorylation, which was not seen in the low-and intermediate-dose cohorts.
Topics: Animals; Sildenafil Citrate; Platelet Activation; Mice, Inbred BALB C; Male; Humans; Collagen; Mice; Adenosine Diphosphate; Blood Platelets; Phosphorylation; Mitogen-Activated Protein Kinases; MAP Kinase Signaling System; Phosphodiesterase 5 Inhibitors; Dose-Response Relationship, Drug; Adult
PubMed: 38808925
DOI: 10.4103/ijp.ijp_312_23 -
Journal of the American Heart... Jun 2024ADP and ATP are importantly involved in vascular and thrombotic homeostasis, via multiple receptor pathways. Blockade of ADP P2Y receptors inhibits platelet aggregation... (Randomized Controlled Trial)
Randomized Controlled Trial
Safety, Tolerability, and Pharmacodynamics of AZD3366 (Optimized Human CD39L3 Apyrase) Alone and in Combination With Ticagrelor and Acetylsalicylic Acid: A Phase 1, Randomized, Placebo-Controlled Study.
BACKGROUND
ADP and ATP are importantly involved in vascular and thrombotic homeostasis, via multiple receptor pathways. Blockade of ADP P2Y receptors inhibits platelet aggregation and represents an effective cardiovascular disease prevention strategy. AZD3366 (APT102), a long-acting recombinant form of an optimized CD39L3 human apyrase, has effectively reduced ATP, ADP, and platelet aggregation and provided tissue protection in preclinical models, features that could be very beneficial in treating patients with cardiovascular disease.
METHODS AND RESULTS
We conducted this phase 1, first-in-human study of single ascending doses of intravenous AZD3366 or placebo, including doses added to dual antiplatelet therapy with ticagrelor and acetylsalicylic acid. The primary objective was safety and tolerability; secondary and exploratory objectives included pharmacokinetics, pharmacodynamics (measured as inhibition of platelet aggregation), adenosine diphosphatase (ADPase) activity, and ATP/ADP metabolism. In total, 104 participants were randomized. AZD3366 was generally well tolerated, with no major safety concerns observed. ADPase activity increased in a dose-dependent manner with a strong correlation to AZD3366 exposure. Inhibition of ADP-stimulated platelet aggregation was immediate, substantial, and durable. In addition, there was a prompt decrease in systemic ATP concentration and an increase in adenosine monophosphate concentrations, whereas ADP concentration appeared generally unaltered. At higher doses, there was a prolongation of capillary bleeding time without detectable changes in the ex vivo thromboelastometric parameters.
CONCLUSIONS
AZD3366 was well tolerated in healthy participants and demonstrated substantial and durable inhibition of platelet aggregation after single dosing. Higher doses prolonged capillary bleeding time without detectable changes in ex vivo thromboelastometric parameters.
REGISTRATION
URL: https://www.clinicaltrials.gov; Unique Identifier: NCT04588727.
Topics: Humans; Male; Ticagrelor; Female; Apyrase; Platelet Aggregation; Aspirin; Platelet Aggregation Inhibitors; Middle Aged; Adult; Double-Blind Method; Dual Anti-Platelet Therapy; Drug Therapy, Combination; Young Adult; Adenosine Diphosphate; Blood Platelets; Dose-Response Relationship, Drug; Treatment Outcome; Recombinant Proteins; Purinergic P2Y Receptor Antagonists
PubMed: 38804212
DOI: 10.1161/JAHA.123.033985 -
Nature Communications May 2024Actin nucleotide-dependent actin remodeling is essential to orchestrate signal transduction and cell adaptation. Rapid energy starvation requires accurate and timely...
Actin nucleotide-dependent actin remodeling is essential to orchestrate signal transduction and cell adaptation. Rapid energy starvation requires accurate and timely reorganization of the actin network. Despite distinct treadmilling mechanisms of ADP- and ATP-actin filaments, their filament structures are nearly identical. How other actin-binding proteins regulate ADP-actin filament assembly is unclear. Here, we show that Spa2 which is the polarisome scaffold protein specifically remodels ADP-actin upon energy starvation in budding yeast. Spa2 triggers ADP-actin monomer nucleation rapidly through a dimeric core of Spa2 (aa 281-535). Concurrently, the intrinsically disordered region (IDR, aa 1-281) guides Spa2 undergoing phase separation and wetting on the surface of ADP-G-actin-derived F-actin and bundles the filaments. Both ADP-actin-specific nucleation and bundling activities of Spa2 are actin D-loop dependent. The IDR and nucleation core of Spa2 are evolutionarily conserved by coexistence in the fungus kingdom, suggesting a universal adaptation mechanism in the fungal kingdom in response to glucose starvation, regulating ADP-G-actin and ADP-F-actin with high nucleotide homogeneity.
Topics: Actin Cytoskeleton; Actins; Adenosine Diphosphate; Glucose; Microfilament Proteins; Saccharomyces cerevisiae; Saccharomyces cerevisiae Proteins
PubMed: 38802374
DOI: 10.1038/s41467-024-48863-4 -
Journal of the American Chemical Society Jun 2024Liquid-jet photoemission spectroscopy (LJ-PES) allows for a direct probing of electronic structure in aqueous solutions. We show the applicability of the approach to...
Liquid-jet photoemission spectroscopy (LJ-PES) allows for a direct probing of electronic structure in aqueous solutions. We show the applicability of the approach to biomolecules in a complex environment, exploring site-specific information on the interaction of adenosine triphosphate in the aqueous phase (ATP) with magnesium (Mg), highlighting the synergy brought about by the simultaneous analysis of different regions in the photoelectron spectrum. In particular, we demonstrate intermolecular Coulombic decay (ICD) spectroscopy as a new and powerful addition to the arsenal of techniques for biomolecular structure investigation. We apply LJ-PES assisted by electronic-structure calculations to study ATP solutions with and without dissolved Mg. Valence photoelectron data reveal spectral changes in the phosphate and adenine features of ATP due to interactions with the divalent cation. Chemical shifts in Mg 2p, Mg 2s, P 2p, and P 2s core-level spectra as a function of the Mg/ATP concentration ratio are correlated to the formation of [Mg(ATP) ], [MgATP], and [MgATP] complexes, demonstrating the element sensitivity of the technique to Mg-phosphate interactions. The most direct probe of the intermolecular interactions between ATP and Mg is delivered by the emerging ICD electrons following ionization of Mg 1s electrons. ICD spectra are shown to sensitively probe ligand exchange in the Mg-ATP coordination environment. In addition, we report and compare P 2s data from ATP and adenosine mono- and diphosphate (AMP and ADP, respectively) solutions, probing the electronic structure of the phosphate chain and the local environment of individual phosphate units in ATP. Our results provide a comprehensive view of the electronic structure of ATP and Mg-ATP complexes relevant to phosphorylation and dephosphorylation reactions that are central to bioenergetics in living organisms.
Topics: Magnesium; Adenosine Triphosphate; Photoelectron Spectroscopy
PubMed: 38802319
DOI: 10.1021/jacs.4c03174 -
Research (Washington, D.C.) 2024Poly (adenosine 5'-diphosphate-ribose) polymerase inhibitors (PARPi) are increasingly important in the treatment of ovarian cancer. However, more than 40% of deficient...
Poly (adenosine 5'-diphosphate-ribose) polymerase inhibitors (PARPi) are increasingly important in the treatment of ovarian cancer. However, more than 40% of deficient patients do not respond to PARPi, and wild-type cases do not show obvious benefit. In this study, we demonstrated that progesterone acted synergistically with niraparib in ovarian cancer cells by enhancing niraparib-mediated DNA damage and death regardless of status. This synergy was validated in an ovarian cancer organoid model and in vivo experiments. Furthermore, we found that progesterone enhances the activity of niraparib in ovarian cancer through inducing ferroptosis by up-regulating palmitoleic acid and causing mitochondrial damage. In clinical cohort, it was observed that progesterone prolonged the survival of patients with ovarian cancer receiving PARPi as second-line maintenance therapy, and high progesterone receptor expression combined with low glutathione peroxidase 4 (GPX4) expression predicted better efficacy of PARPi in patients with ovarian cancer. These findings not only offer new therapeutic strategies for PARPi poor response ovarian cancer but also provide potential molecular markers for predicting the PARPi efficacy.
PubMed: 38798714
DOI: 10.34133/research.0371 -
Food Chemistry Oct 2024Sweet potatoes are rich in cardioprotective phytochemicals with potential anti-platelet aggregation activity, although this benefit may vary among cultivars/genotypes....
Sweet potatoes are rich in cardioprotective phytochemicals with potential anti-platelet aggregation activity, although this benefit may vary among cultivars/genotypes. The phenolic profile [HPLC-ESI-qTOF-MS], cheminformatics (ADMET properties, affinity toward platelet proteins) and anti-PA activity of phenolic-rich hydroalcoholic extracts obtained from orange (OSP) and purple (PSP) sweet potato storage roots, was evaluated. The phenolic richness [Hydroxycinnamic acids> flavonoids> benzoic acids] was PSP > OSP. Their main chlorogenic acids could interact with platelet proteins (integrins/adhesins, kinases/metalloenzymes) but their bioavailability could be poor. Just OSP exhibited a dose-dependent anti-platelet aggregation activity [inductor (IC mg.ml): thrombin receptor activator peptide-6 (0.55) > Adenosine-5'-diphosphate (1.02) > collagen (1.56)] and reduced P-selectin expression (0.75-1.0 mg.ml) but not glycoprotein IIb/IIIa secretion. The explored anti-PA activity of OSP/PSP seems to be inversely related to their phenolic richness. The poor first-pass bioavailability of its chlorogenic acids (documented in silico) may represent a further obstacle for their anti-PA in vivo.
Topics: Ipomoea batatas; Phenols; Platelet Aggregation; Plant Extracts; Platelet Aggregation Inhibitors; Plant Roots; Humans; Cheminformatics; Animals; Blood Platelets
PubMed: 38797094
DOI: 10.1016/j.foodchem.2024.139794 -
European Journal of Cell Biology Jun 2024Intracellular actin networks assemble through the addition of ATP-actin subunits at the growing barbed ends of actin filaments. This is followed by "aging" of the...
Intracellular actin networks assemble through the addition of ATP-actin subunits at the growing barbed ends of actin filaments. This is followed by "aging" of the filament via ATP hydrolysis and subsequent phosphate release. Aged ADP-actin subunits thus "treadmill" through the filament before being released back into the cytoplasmic monomer pool as a result of depolymerization at filament pointed ends. The necessity for aging before filament disassembly is reinforced by preferential binding of cofilin to aged ADP-actin subunits over newly-assembled ADP-P actin subunits in the filament. Consequently, investigations into how cofilin influences pointed-end depolymerization have, thus far, focused exclusively on aged ADP-actin filaments. Using microfluidics-assisted Total Internal Reflection Fluorescence (mf-TIRF) microscopy, we reveal that, similar to their effects on ADP filaments, cofilin and cyclase-associated protein (CAP) also promote pointed-end depolymerization of ADP-P filaments. Interestingly, the maximal rates of ADP-P filament depolymerization by CAP and cofilin together remain approximately 20-40 times lower than for ADP filaments. Further, we find that the promotion of ADP-P pointed-end depolymerization is conserved for all three mammalian cofilin isoforms. Taken together, the mechanisms presented here open the possibility of newly-assembled actin filaments being directly disassembled from their pointed-ends, thus bypassing the slow step of P release in the aging process.
Topics: Actin Cytoskeleton; Animals; Actins; Actin Depolymerizing Factors; Adenosine Diphosphate; Rabbits; Mice; Polymerization; Cofilin 1
PubMed: 38796920
DOI: 10.1016/j.ejcb.2024.151423 -
The Journal of Surgical Research May 2024Previous literature suggests that sphingolipids may impact systemic coagulation and platelet aggregation, thus modulating the risks of thrombotic events. The goal of...
INTRODUCTION
Previous literature suggests that sphingolipids may impact systemic coagulation and platelet aggregation, thus modulating the risks of thrombotic events. The goal of this investigation was to evaluate the role of serum sphingolipids on intrinsic platelet function to assess whether pharmacologic manipulation of sphingolipid metabolites would impact platelet aggregability.
METHODS
C57BL/6J mice were injected with either normal saline, 1 mg/kg FTY720 (synthetic sphingosine-1-phosphate [S1P] receptor analog), or 5 mg/kg SLM6031434 (sphingosine kinase two inhibitor). Mice were sacrificed at 6 h and whole blood (WB) was collected for impedance aggregometry assessing platelet responsiveness to arachidonic acid or adenosine diphosphate. Ex vivo studies utilized WB or platelet-rich plasma that was pretreated with S1P, FTY720, amitriptyline, or d-sphingosine then analyzed by aggregability and flow cytometry for platelet and platelet-derived microvesicle characteristics.
RESULTS
FTY720 and SLM6031434 pretreated induced similar arachidonic acid and adenosine diphosphate-mediated platelet aggregation as controls. Ex vivo WB and platelet-rich plasma treatment with S1P, FTY720, amitriptyline and d-sphingosine did not impact platelet aggregation. The percentages of CD41+, CD62P+ and CD41+/ceramide+, CD62P+/ceramide + platelets, and platelet-derived microvesicle were not significantly different between amitriptyline-treated and normal saline-treated cohorts.
CONCLUSIONS
Sphingolipid modulating agents, such as FTY720, SLM6031434, S1P, amitriptyline, ceramide, and d-sphingosine do not appear to independently impact platelet aggregation in murine models.
PubMed: 38795670
DOI: 10.1016/j.jss.2024.04.063 -
Microorganisms May 2024In cyanobacteria and chloroplasts (in algae and plants), ATP synthase plays a pivotal role as a photosynthetic membrane complex responsible for producing ATP from... (Review)
Review
In cyanobacteria and chloroplasts (in algae and plants), ATP synthase plays a pivotal role as a photosynthetic membrane complex responsible for producing ATP from adenosine diphosphate and inorganic phosphate, utilizing a proton motive force gradient induced by photosynthesis. These two ATP synthases exhibit similarities in gene organization, amino acid sequences of subunits, structure, and functional mechanisms, suggesting that cyanobacterial ATP synthase is probably the evolutionary precursor to chloroplast ATP synthase. In this review, we explore the precise synthesis and assembly of ATP synthase subunits to address the uneven stoichiometry within the complex during transcription, translation, and assembly processes. We also compare the regulatory strategies governing ATP synthase activity to meet varying energy demands in cyanobacteria and chloroplasts amid fluctuating natural environments. Furthermore, we delve into the role of ATP synthase in stress tolerance and photosynthetic carbon fixation efficiency in oxygenic photosynthetic organisms (OPsOs), along with the current researches on modifying ATP synthase to enhance carbon fixation efficiency under stress conditions. This review aims to offer theoretical insights and serve as a reference for understanding the functional mechanisms of ATP synthase, sparking innovative ideas for enhancing photosynthetic carbon fixation efficiency by utilizing ATP synthase as an effective module in OPsOs.
PubMed: 38792770
DOI: 10.3390/microorganisms12050940 -
International Journal of Molecular... May 2024The excessive activation of frog eggs, referred to as overactivation, can be initiated by strong oxidative stress, leading to expedited calcium-dependent non-apoptotic...
The excessive activation of frog eggs, referred to as overactivation, can be initiated by strong oxidative stress, leading to expedited calcium-dependent non-apoptotic cell death. Overactivation also occurs spontaneously, albeit at a low frequency, in natural populations of spawned frog eggs. Currently, the cytological and biochemical events of the spontaneous process have not been characterized. In the present study, we demonstrate that the spontaneous overactivation of frog eggs, similarly to oxidative stress- and mechanical stress-induced overactivation, is characterized by the fast and irreversible contraction of the egg's cortical layer, an increase in egg size, the depletion of intracellular ATP, a drastic increase in the intracellular ADP/ATP ratio, and the degradation of M phase-specific cyclin B2. These events manifest in eggs in the absence of caspase activation within one hour of triggering overactivation. Importantly, substantial amounts of ATP and ADP leak from the overactivated eggs, indicating that plasma membrane integrity is compromised in these cells. The rupture of the plasma membrane and acute depletion of intracellular ATP explicitly define necrotic cell death. Finally, we report that egg overactivation can occur in the frog's genital tract. Our data suggest that mechanical stress may be a key factor promoting egg overactivation during oviposition in frogs.
Topics: Animals; Adenosine Triphosphate; Ovum; Necrosis; Xenopus laevis; Female; Oxidative Stress; Adenosine Diphosphate; Cell Death; Cell Membrane; Stress, Mechanical
PubMed: 38791359
DOI: 10.3390/ijms25105321