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ChemSusChem Jul 2024The common synthesis approach of reduced graphene oxide (rGO) using toxic reducing agents poses a threat to environmental pollution. This study used banana peel extract...
The common synthesis approach of reduced graphene oxide (rGO) using toxic reducing agents poses a threat to environmental pollution. This study used banana peel extract as a green reducing agent for the synthesis of rGO. Ultrasonication was assimilated to expedite the synthesis process. For comparison, rGO was also produced by reducing GO with hydrazine treatment under conventional stirring. Both morphological (SEM) and physicochemical (FTIR and XRD) studies have revealed that banana peel extract can reduce GO to rGO, although its reducing effect is much weaker compared to hydrazine. Despite this, the rGO produced using banana peel extract with the assimilation of ultrasonication technique has a greater interlayer spacing than that formed under the conventional stirring method. In terms of electrical properties, the electrical conductance of hydrazine-produced rGO (5.69 × 10-6 S) is higher than the banana peel extract-produced rGO (3.55 × 10-6 S - 4.56 × 10-6 S). Interestingly, it was found that most of the rGO produced by banana peel extract under ultrasound assistance has higher or comparable electrical conductance compared to the rGO produced by banana peel extract under stirring method. This implies the feasibility of using short-period ultrasound to replace conventional stirring in rGO synthesis.
PubMed: 38948933
DOI: 10.1002/cssc.202400845 -
Synlett : Accounts and Rapid... Nov 2023Potassium trifluoroborates have gained significant utility as coupling partners in organic synthesis, particularly in the Suzuki-Miyaura coupling reaction. Recently,...
Potassium trifluoroborates have gained significant utility as coupling partners in organic synthesis, particularly in the Suzuki-Miyaura coupling reaction. Recently, they have also been used as radical precursors under oxidative conditions to generate carbon-centered radicals. These versatile reagents have found new applications in photoredox catalysis, including radical substitution, conjugate addition reactions, and transition metal dual catalysis. In addition, this photomediated redox neutral process has enabled radical-radical coupling with persistent radicals in the absence of a metal, and this process remains to be fully explored. In this study, we report the radical-radical coupling of benzylic potassium trifluoroborate salts with isolated acyl azolium triflates, which are persistent radical precursors. The reaction is catalyzed by an organic photocatalyst and forms isolable tertiary alcohol species. These compounds can be transformed into a range of substituted ketone products by simple treatment with a mild base.
PubMed: 38948905
DOI: 10.1055/s-0041-1738448 -
BioRxiv : the Preprint Server For... Jun 2024Under stress conditions, cells reprogram their molecular machineries to mitigate damage and promote survival. Ubiquitin signaling is globally increased during oxidative...
Under stress conditions, cells reprogram their molecular machineries to mitigate damage and promote survival. Ubiquitin signaling is globally increased during oxidative stress, controlling protein fate and supporting stress defenses at several subcellular compartments. However, the rules driving subcellular ubiquitin localization to promote these concerted response mechanisms remain understudied. Here, we show that K63-linked ubiquitin chains, known to promote proteasome-independent pathways, accumulate primarily in non-cytosolic compartments during oxidative stress induced by sodium arsenite in mammalian cells. Our subcellular ubiquitin proteomic analyses of non-cytosolic compartments expanded 10-fold the pool of proteins known to be ubiquitinated during arsenite stress (2,046) and revealed their involvement in pathways related to immune signaling and translation control. Moreover, subcellular proteome analyses revealed proteins that are recruited to non-cytosolic compartments under stress, including a significant enrichment of helper ubiquitin-binding adaptors of the ATPase VCP that processes ubiquitinated substrates for downstream signaling. We further show that VCP recruitment to non-cytosolic compartments under arsenite stress occurs in a ubiquitin-dependent manner mediated by its adaptor NPLOC4. Additionally, we show that VCP and NPLOC4 activities are critical to sustain low levels of non-cytosolic K63-linked ubiquitin chains, supporting a cyclical model of ubiquitin conjugation and removal that is disrupted by cellular exposure to reactive oxygen species. This work deepens our understanding of the role of localized ubiquitin and VCP signaling in the basic mechanisms of stress response and highlights new pathways and molecular players that are essential to reshape the composition and function of the human subcellular proteome under dynamic environments.
PubMed: 38948861
DOI: 10.1101/2024.06.20.598218 -
BioRxiv : the Preprint Server For... Jun 2024Oxidative stress is implicated in the pathogenesis and progression of abdominal aortic aneurysm (AAA). Antioxidant delivery as a therapeutic for AAA is of substantial...
BACKGROUND
Oxidative stress is implicated in the pathogenesis and progression of abdominal aortic aneurysm (AAA). Antioxidant delivery as a therapeutic for AAA is of substantial interest although clinical translation of antioxidant therapy has met with significant challenges due to limitations in achieving sufficient antioxidant levels at the site of AAA. We posit that nanoparticle-based approaches hold promise to overcome challenges associated with systemic administration of antioxidants.
METHODS
We employed a peptide-based nanoplatform to overexpress a key modulator of oxidative stress, superoxide dismutase 2 (SOD2). The efficacy of systemic delivery of SOD2 mRNA as a nanotherapeutic agent was studied in two different murine AAA models. Unbiased mass spectrometry-enabled proteomics and high-dimensional bioinformatics were used to examine pathways modulated by SOD2 overexpression.
RESULTS
The murine SOD2 mRNA sequence was mixed with p5RHH, an amphipathic peptide capable of delivering nucleic acids to form self-assembled nanoparticles of ∼55 nm in diameter. We further demonstrated that the nanoparticle was stable and functional up to four weeks following self-assembly when coated with hyaluronic acid. Delivery of SOD2 mRNA mitigated the expansion of small AAA and largely prevented rupture. Mitigation of AAA was accompanied by enhanced SOD2 protein expression in aortic wall tissue. Concomitant suppression of nitric oxide, inducible nitric oxide synthase expression, and cell death was observed. Proteomic profiling of AAA tissues suggests that SOD2 overexpression augments levels of microRNAs that regulate vascular inflammation and cell apoptosis, inhibits platelet activation/aggregation, and downregulates mitogen-activated protein kinase signaling. Gene set enrichment analysis shows that SOD2 mRNA delivery is associated with activation of oxidative phosphorylation, lipid metabolism, respiratory electron transportation, and tricarboxylic acid cycle pathways.
CONCLUSIONS
These results confirm that SOD2 is key modulator of oxidative stress in AAA. This nanotherapeutic mRNA delivery approach may find translational application in the medical management of small AAA and the prevention of AAA rupture.
PubMed: 38948794
DOI: 10.1101/2024.06.17.599454 -
BioRxiv : the Preprint Server For... Apr 2024The renin-angiotensin system involves many more enzymes, receptors and biologically active peptides than originally thought. With this study, we investigated whether...
BACKGROUND
The renin-angiotensin system involves many more enzymes, receptors and biologically active peptides than originally thought. With this study, we investigated whether angiotensin-(1-5) [Ang-(1-5)], a 5-amino acid fragment of angiotensin II, has biological activity, and through which receptor it elicits effects.
METHODS
The effect of Ang-(1-5) (1µM) on nitric oxide release was measured by DAF-FM staining in human aortic endothelial cells (HAEC), or Chinese Hamster Ovary (CHO) cells stably transfected with the angiotensin AT -receptor (AT R) or the receptor Mas. A potential vasodilatory effect of Ang-(1-5) was tested in mouse mesenteric and human renal arteries by wire myography; the effect on blood pressure was evaluated in normotensive C57BL/6 mice by Millar catheter. These experiments were performed in the presence or absence of a range of antagonists or inhibitors or in AT R-knockout mice. Binding of Ang-(1-5) to the AT R was confirmed and the preferred conformations determined by docking simulations. The signaling network of Ang-(1-5) was mapped by quantitative phosphoproteomics.
RESULTS
Key findings included: (1) Ang-(1-5) induced activation of eNOS by changes in phosphorylation at eNOS and eNOS and thereby (2) increased NO release from HAEC and AT R-transfected CHO cells, but not from Mas-transfected or non-transfected CHO cells. (3) Ang-(1-5) induced relaxation of preconstricted mouse mesenteric and human renal arteries and (4) lowered blood pressure in normotensive mice - effects which were respectively absent in arteries from AT R-KO or in PD123319-treated mice and which were more potent than effects of the established AT R-agonist C21. (5) According to modelling, Ang-(1-5) binds to the AT R in two preferred conformations, one differing substantially from where the first five amino acids within angiotensin II bind to the AT R. (6) Ang-(1-5) modifies signaling pathways in a protective RAS-typical way and with relevance for endothelial cell physiology and disease.
CONCLUSIONS
Ang-(1-5) is a potent, endogenous AT R-agonist.
PubMed: 38948791
DOI: 10.1101/2024.04.05.588367 -
BioRxiv : the Preprint Server For... Jun 2024Sickle cell disease is caused by a mutation in the beta subunit of hemoglobin (HbSS) that drives Hb fiber formation when the protein is in the deoxygenated (tense, T)...
UNLABELLED
Sickle cell disease is caused by a mutation in the beta subunit of hemoglobin (HbSS) that drives Hb fiber formation when the protein is in the deoxygenated (tense, T) state. The drug voxelotor was recently approved to treat sickle cell disease by preventing HbSS fiber formation. Voxelotor acts as an allosteric inhibitor of polymerization by maintaining the HbSS protein in the relaxed (R) conformation, limiting polymerization of T-state fibers. Normal blood cells contain small amounts of natural Hb fibers and a few percent of the Fe ferric form, metHb, incapable of binding oxygen. Although the drug Voxelotor is now in use, the effect of the drug on the oxidized metHb state has not been reported. Here we assessed the influence of voxelotor on normal human metHb. We compared the aggregation of metHb at two pH values (5.5 and 7.1). MetHb is known to form organized fiber structures at or below pH 5.5. We find that voxelotor significantly enhances fiber formation of metHb R-state at pH 5.5, consistent with the mode of action for this drug in maintaining the Hb R conformation. The opposite effect is observed at physiological pH values. Voxelotor significantly decreases the rate of metHb aggregate formation at pH 7.1 but did not affect protein stability. Notably, drug binding drives metHb into novel spherical particles with a morphology never seen before for Hb. The formation of these particles should be considered in patients being treated for sickle cell disease with voxelotor.
WHY IT MATTERS
Voxelotor is an FDA-approved drug for sickle cell anemia, known to prevent hemoglobin fiber formation. Here, we investigate its effect on methemoglobin, the form of hemoglobin in which iron takes on the ferric Fe state. Our study examines voxelotor's impact on methemoglobin aggregation and stability. At pH 7.1, we found voxelotor to have an effect on methemoglobin solubility as evidenced by the formation of novel methemoglobin spherical structures. We observe that voxelotor significantly increases methemoglobin fiber formation at pH 5.5 but, notably, reduces methemoglobin aggregation at physiological pH levels. Minimal impact on methemoglobin thermodynamic stability is noted. These findings suggest voxelotor's potential therapeutic efficacy for various hemoglobinopathies, including conditions characterized by Heinz body formation.
PubMed: 38948767
DOI: 10.1101/2024.06.16.599216 -
BioRxiv : the Preprint Server For... Jun 2024Beckwith-Wiedemann Syndrome (BWS) is an epigenetic overgrowth syndrome caused by methylation changes in the human 11p15 chromosomal locus. Patients with BWS exhibit...
Beckwith-Wiedemann Syndrome (BWS) is an epigenetic overgrowth syndrome caused by methylation changes in the human 11p15 chromosomal locus. Patients with BWS exhibit tissue overgrowth, as well as an increased risk of childhood neoplasms in the liver and kidney. To understand the impact of these 11p15 changes, specifically in the liver, we performed single-nucleus RNA sequencing (snRNA-seq) and single-nucleus assay for transposase-accessible chromatin with sequencing (snATAC-seq) to generate paired, cell-type-specific transcriptional and chromatin accessibility profiles of both BWS-liver and nonBWS-liver nontumorous tissue. Our integrated RNA+ATACseq multiomic approach uncovered hepatocyte-specific enrichment and activation of the peroxisome proliferator-activated receptor α (PPARA) - a liver metabolic regulator. To confirm our findings, we utilized a BWS-induced pluripotent stem cell (iPSC) model, where cells were differentiated into hepatocytes. Our data demonstrates the dysregulation of lipid metabolism in BWS-liver, which coincided with observed upregulation of PPARA during hepatocyte differentiation. BWS liver cells exhibited decreased neutral lipids and increased fatty acid β-oxidation, relative to controls. We also observed increased reactive oxygen species (ROS) byproducts in the form of peroxidated lipids in BWS hepatocytes, which coincided with increased oxidative DNA damage. This study proposes a putative mechanism for overgrowth and cancer predisposition in BWS liver due to perturbed metabolism.
PubMed: 38948745
DOI: 10.1101/2024.06.14.599077 -
Several common methods of making vesicles (except an emulsion method) capture intended lipid ratios.BioRxiv : the Preprint Server For... Jun 2024Researchers choose different methods of making giant unilamellar vesicles in order to satisfy different constraints of their experimental designs. A challenge of using a...
UNLABELLED
Researchers choose different methods of making giant unilamellar vesicles in order to satisfy different constraints of their experimental designs. A challenge of using a variety of methods is that each may produce vesicles of different lipid compositions, even if all vesicles are made from a common stock mixture. Here, we use mass spectrometry to investigate ratios of lipids in vesicles made by five common methods: electroformation on indium tin oxide slides, electroformation on platinum wires, gentle hydration, emulsion transfer, and extrusion. We made vesicles from either 5-component or binary mixtures of lipids chosen to span a wide range of physical properties: di(18:1)PC, di(16:0)PC, di(18:1)PG, di(12:0)PE, and cholesterol. For a mixture of all five of these lipids, ITO electroformation, Pt electroformation, gentle hydration, and extrusion methods result in only minor shifts (≤ 5 mol%) in lipid ratios of vesicles relative to a common stock solution. In contrast, emulsion transfer results in ∼80% less cholesterol than expected from the stock solution, which is counterbalanced by a surprising overabundance of saturated PC-lipid relative to all other phospholipids. Experiments using binary mixtures of some of the lipids largely support results from the 5-component mixture. Exact values of lipid ratios variations likely depend on the details of each method, so a broader conclusion is that experiments that increment lipid ratios in small steps will be highly sensitive to the method of lipid formation and to sample-to-sample variations, which are low (roughly ±2 mol% in the 5-component mixture and either scale proportionally with increasing mole fraction or remain low). Experiments that increment lipid ratios in larger steps or that seek to explain general trends or new phenomena will be less sensitive to the method used.
SIGNIFICANCE STATEMENT
Small changes to the amounts and types of lipids in membranes can drastically affect the membrane's behavior. Unfortunately, it is unknown whether (or to what extent) different methods of making vesicles alter the ratios of lipids in membranes, even when identical stock solutions are used. This presents challenges for researchers when comparing data with colleagues who use different methods. Here, we measure ratios of lipid types in vesicle membranes produced by five methods. We assess each method's reproducibility and compare resulting vesicle compositions across methods. In doing so, we provide a quantitative basis that the scientific community can use to estimate whether differences between their results can be simply attributed to differences between methods or to sample-to-sample variations.
PubMed: 38948736
DOI: 10.1101/2024.02.21.581444 -
BioRxiv : the Preprint Server For... Jun 2024Oxidative protein folding in the endoplasmic reticulum (ER) is essential for all eukaryotic cells yet generates hydrogen peroxide (H2O2), a reactive oxygen species...
Oxidative protein folding in the endoplasmic reticulum (ER) is essential for all eukaryotic cells yet generates hydrogen peroxide (H2O2), a reactive oxygen species (ROS). The ER-transmembrane protein that provides reducing equivalents to ER and guards the cytosol for antioxidant defense remains unidentified. Here we combine AlphaFold2- based and functional reporter screens in to identify a previously uncharacterized and evolutionarily conserved protein ERGU-1 that fulfills these roles. Deleting ERGU-1 causes excessive H2O2 and transcriptional gene up- regulation through SKN-1, homolog of mammalian antioxidant master regulator NRF2. ERGU-1 deficiency also impairs organismal reproduction and behaviors. Both and human ERGU-1 proteins localize to ER membranes and form network reticulum structures. We name this system ER-GUARD, E ndoplasmic R eticulum Gu ardian A egis of R edox D efense. Human and homologs of ERGU-1 can rescue mutant phenotypes, demonstrating evolutionarily ancient and conserved functions. Together, our results reveal an ER-membrane-specific protein machinery and defense-net system ER-GUARD for peroxide detoxification and suggest a previously unknown but conserved pathway for antioxidant defense in animal cells.
PubMed: 38948723
DOI: 10.1101/2024.06.19.599784 -
BioRxiv : the Preprint Server For... Oct 2023Low nephron endowment at birth is a risk factor for chronic kidney disease. The prevalence of this condition is increasing due to higher survival rates of preterm...
Low nephron endowment at birth is a risk factor for chronic kidney disease. The prevalence of this condition is increasing due to higher survival rates of preterm infants and children with multi- organ birth defect syndromes that affect the kidney and urinary tract. We created a mouse model of congenital low nephron number due to deletion of in nephron progenitor cells. is a core component of the Nucleosome Remodeling and Deacetylase (NuRD) chromatin remodeling complex. These mice developed albuminuria at 4 weeks of age followed by focal segmental glomerulosclerosis (FSGS) at 8 weeks, with progressive kidney injury and fibrosis. Our studies reveal that altered mitochondrial metabolism in the post-natal period leads to accumulation of neutral lipids in glomeruli at 4 weeks of age followed by reduced mitochondrial oxygen consumption. We found that NuRD cooperated with Zbtb7a/7b to regulate a large number of metabolic genes required for fatty acid oxidation and oxidative phosphorylation. Analysis of human kidney tissue also supported a role for reduced mitochondrial lipid metabolism and ZBTB7A/7B in FSGS and CKD. We propose that an inability to meet the physiological and metabolic demands of post-natal somatic growth of the kidney promotes the transition to CKD in the setting of glomerular hypertrophy due to low nephron endowment.
PubMed: 38948707
DOI: 10.1101/2023.10.18.562984