nM-1 μM Research Articles

Ratiometric fluorescence for ultrasensitive detection of chlortetracycline in milk matrix based on its blue alkaline degradation product and red-emitting cyclodextrin stabilized gold nanocluster

As an emerging fluorescent nanomaterial, gold nanoclusters are of increasing interest in sensing and imaging because of their stable luminescence properties, large Stokes shift and low toxicity. Here, red-emitting gold nanoclusters stabilized by 6-Deoxy-6-mercapto-β-cyclodextrin (6-SH-CD@Au NCs) were successfully synthesized based on photo-reduction method for the detection of chlortetracycline. Under alkaline conditions, chlortetracycline transforms into iso-chlortetracycline (iso-CTC), which has a distinct blue fluorescence at 415 nm. The fluorescence intensity at 415 nm changed with increasing concentration of chlortetracycline and inner filter effect (IFE) of 6-SH-CD@Au NCs, while the fluorescence of the gold nanocluster at 670 nm decreased due to the IFE of iso-CTC. This fluorescent system detected chlortetracycline with good linearity in the concentration range of 5 nM-1 μM, and the limit of detection is 2.7 nM. The developed fluorescence sensor and test paper assay had been successfully applied for the detection of chlortetracycline in milk samples. The results demonstrated that the ratiometric sensor has advantages of simplicity, high sensitivity and specificity, cost-effectiveness and ease of operation.

Graphene-anchored conductive polymer aerogel composite for the electrocatalytic detection of hydrogen peroxide and bisphenol A

An aerogel composite of reduced graphene oxide (rGO) anchored on amine groups of conductive polyterthiophene (p(APT-rGO) aerogel is synthesized for the first time, using a hydrothermal method. The electrocatalytic performance of the composite electrode was explored for bisphenol A (BPA) oxidation and hydrogen peroxide reduction. The surface of catalyst was found to possess numerous active sites owing to exposed edges and large surface area of a three-dimensional aerogel. Additionally, incorporation of rGO into poly(aminopyrimidyl terthiophene) (pAPT) is found to enhance the catalytic performance and stability. The p(APT-rGO) aerogel coated screen printed carbon electrode (SPCE) displayed a superior catalytic performance for the BPA detection with two dynamic ranges of 5–500 nM and 1–200 µM with the detection limit of 4.28 ± 0.75 nM. Otherwise, the dynamic ranges of H2O2 detection were 100 nM-100 µM and 200 µM-3 mM with the detection limit of 9.12 ± 1.03 nM. The catalytic active site of the composite aerogel was investigated for BPA oxidation and H2O2 reduction. The proposed sensor demonstrated an excellent analytical performance for the determination of BPA extracted from commercial polycarbonate bottles and trace amounts of H2O2 released from living cells by drug stimulation.

The Small Molecule PPARγ Agonist GL516 Induces Feeding-Stimulatory Effects in Hypothalamus Cells Hypo-E22 and Isolated Hypothalami.

PPARγ agonists are implicated in the regulation of diabetes and metabolic syndrome and have therapeutic potential in brain disorders. PPARγ modulates appetite through its central effects, especially on the hypothalamic arcuate nucleus (ARC). Previous studies demonstrated that the small molecule GL516 is a PPARγ agonist able to reduce oxidative stress and apoptosis with a potential neuroprotective role. Herein, we investigated the effects of GL516, in vitro and ex vivo, on the levels of hypothalamic dopamine (DA) and serotonin (5-HT). The gene expressions of neuropeptide Y, CART, AgRP, and POMC, which play master roles in the neuroendocrine regulation of feeding behavior and energy balance, were also evaluated. HypoE22 cells were treated with H2O2 (300 μM) for 2 h e 30’ and with different concentrations of GL516 (1 nM-100 µM). The cell viability was evaluated after 24 and 48 h of culturing using the MTT test. DA and 5-HT levels in the HypoE22 cell supernatants were analyzed through HPLC; an ex vivo study on isolated hypothalamic specimens challenged with scalar concentrations of GL516 (1–100 µM) and with pioglitazone (10 µM) was carried out. The gene expressions of CART, NPY, AgRP, and POMC were also determined by a quantitative real-time PCR. The results obtained showed that GL516 was able to reduce DA and 5-HT turnover; moreover, it was effective in stimulating NPY and AgRP gene expressions with a concomitant reduction in CART and POMC gene expressions. These results highlight the capability of GL516 to modulate neuropeptide pathways deeply involved in appetite control suggesting an orexigenic effect. These findings emphasize the potential use of GL516 as a promising candidate for therapeutical applications in neurodegenerative diseases associated with the reduction in food intake and stimulation of catabolic pathways.

6-Nitrodopamine is an endogenous selective dopamine receptor antagonist in Chelonoidis carbonaria aorta

Chelonoidis carbonaria aortic rings present endothelium-derived release of dopamine, noradrenaline, adrenaline and 6-nitrodopamine (6-ND). Here it was investigated whether 6-ND release is coupled to nitric oxide (NO) synthesis and its action on the vascular smooth muscle reactivity.Basal release of 6-ND from aortic rings in the absence and presence of the NO synthesis inhibitor L-NAME was quantified by LC-MS-MS. Aortic rings were suspended vertically between two metal hooks in 10-mL organ baths containing Krebs-Henseleit's solution and attached to isometric transducers. The tissues were allowed to equilibrate for 1 h before starting the experiments.The release of 6-ND was significantly reduced by previous incubation with L-NAME. 6-ND (up to 300 μM) had no contractile activity in the aortic rings. 6-ND (1, 3 and 10 μM) produced significant rightward shifts of the concentration-response curves to dopamine in endothelium-intact (pA2 6.09) and L-NAME pre-treated endothelium-intact (pA2 7.06) aortic rings. Contractions induced by noradrenaline and adrenaline were not affected by pre-incubation with 6-ND. The EFS (16 Hz)-induced aortic contractions were significantly inhibited by incubation with 6-ND (10 μM).In the thromboxane A2 mimetic U-46619 (30 nM) pre-contracted endothelium intact aortic rings, 6-ND (1 nM–1 μM) and the dopamine D2-receptor antagonist haloperidol (1 nM-1 μM) induced concentration-dependent relaxations. The relaxations were not present in endothelium-removed aortic rings but they were not affected by incubation with L-NAME in endothelium-intact aortic rings.The results indicate that the synthesis of this novel catecholamine in Chelonoidis carbonaria aortic rings is coupled to NO release and that 6-ND acts as a highly selective dopamine D2-like receptor antagonist.

Preparation of sulphuric acid-mediated N,S-codoped red emissive carbon dots: Applications in food dyes detection, solid-state luminescence and cell imaging

As diseases such as cardiovascular disease and cancer caused by food problems are more and more frequent, food safety has received great attention. Among them, the safety problem caused by food dyes is more prominent. Thus, it is of great value to develop sensitive detection methods for food dyes. In present study, sulphuric acid-mediated N,S-codoped red emissive carbon dots (namely as R-CDs) had been manufactured by using N-phenyl-o-phenylenediamine as precursor, sulfuric acid as additive for the first time. The structural and fluorescence properties of R-CDs had been systematically studied. The results demonstrated that R-CDs showed uniform spherical morphology and had a graphite-like structure, for which the average diameters size was 5.05 nm. Due to the various functional groups such as hydroxyl, pyridinic N, pyrrolic N and -C-SO4, R-CDs emitted bright red fluorescence. Importantly, because of the interactions between the functional groups of R-CDs with the selected food dyes, three dyes including amaranth, brilliant blue FCF and methylene blue can sensitively quench the fluorescence of R-CDs through IFE and static quenching effects. The linearity ranges of them were separately detected as 0.20 μM −20 μM, 10 nM-1 μM and 60 nM-8 μM. The limits of detection (LODs) of them were 70 nM, 4 nM and 20 nM, respectively. Further, R-CDs was successfully applied to the sensitive detection of three dyes from various food samples. To maximize the fluorescence properties of R-CDs, a R-CDs/PVA composite gel was fabricated to make R-CDs fluoresce in solid state condition. The potential of R-CDs/PVA composite gel for preliminary visualization analysis of three dyes was studied. Finally, ascribed to the low toxicity and good biocompatibility, the potential of R-CDs as probe for cell imaging was explored preliminarily.

Abstract 3267: ARID1A mutated bladder cancer cells are sensitized to BET protein inhibition with OTX-015

Abstract Background: Bladder cancer (BC) is a common and deadly disease. Inactivating ARID1A mutations occur in up to 30% of metastatic BC tumors, making it the most commonly mutated epigenetic gene and the 4th most commonly mutated gene overall in BC. ARID1A mutations are associated with decreased response to therapy and poor prognosis; thus, there is a need to develop therapies specifically targeting ARID1A mutant tumors. Using the UNC EpiG Diamond compound library, a set of well-annotated small molecule probes targeting chromatin regulatory proteins, we determined that inhibitors of bromodomain and extraterminal (BET) proteins potently inhibit the viability of BC cells. Here, we tested the hypothesis that ARID1Amut cells are particularly sensitized to the pan-BET inhibitor OTX-015. Methods: AR1D1A-competent (ARID1AWT) 5637 and ARID1A-mutated (ARID1Amut) HT1197 cells were treated with eight ascending concentrations of OTX-015 (0.1 nM-100 µM), and cell viability was measured using CellTiter-Glo™ after 72-120-hour incubations. IC50 values were calculated using a four-parameter non-linear regression model. Gene expression of BET inhibitor target genes, MYC, ARID1B, and RAD51, were evaluated by RT-PCR after a 48-hour incubation, normalized to an SDHA housekeeper and compared to a 0.1% DMSO control. CellTiter-Glo™ and RT-PCR experiments were conducted in technical and biologic triplicates. Western blotting evaluated OTX-015 effects on c-MYC, BAF250B (encoded by ARID1B), and RAD51 protein expression after 72-hour incubation. Results: OTX-015 was 8-times more potent in ARID1Amut HT1197 cells at 120 h than in ARID1AWT 5637 cells (IC50: 0.12 μM vs. 1.0 μM). OTX-015 treatment (1 µM) significantly reduced ARID1B and RAD51 mRNA expression versus 0.1% DMSO control (83% and 86% reduction, respectively, both P<0.0001). At the same concentration, OTX-015 did not significantly reduce MYC mRNA expression HT1197 cells (25% reduction, P=0.31), but did significantly reduce MYC expression in 5637 cells (57% reduction, P<0.0001) at 48 h. When comparing HT1197 to 5637 cells, OTX-015 treatment (1 µM) caused a greater reduction in ARID1B mRNA expression (83% vs. 62% reduction, P=0.02) and RAD51 mRNA expression (86% vs. 57%, P=0.001) at 48 h. Finally, OTX-015 treatment (1 µM) also resulted in more dramatically reduced c-MYC, ARID1B and RAD51 protein expression in HT1197 cells at 72 h, when compared to 5637 cells. Conclusions: These preliminary results support future lines of inquiry into the molecular mechanisms that underlie sensitization of ARID1Amut cells to BET protein inhibition. Citation Format: Ryan M. Kemper, Heemaja Mewada, Jeffrey S. Damrauer, Brian Hardy, Stephen F. Frye, Kenneth H. Pearce, Jesse Raab, William Y. Kim, Daniel James Crona. ARID1A mutated bladder cancer cells are sensitized to BET protein inhibition with OTX-015 [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3267.

Prostaglandin E2 Exerts Biphasic Dose Response on the PreBötzinger Complex Respiratory-Related Rhythm.

Inflammation in infants can cause respiratory dysfunction and is potentially life-threatening. Prostaglandin E2 (PGE2) is released during inflammatory events and perturbs breathing behavior in vivo. Here we study the effects of PGE2 on inspiratory motor rhythm generated by the preBötzinger complex (preBötC). We measured the concentration dependence of PGE2 (1 nM-1 μM) on inspiratory-related motor output in rhythmic medullary slice preparations. Low concentrations (1–10 nM) of PGE2 increased the duration of the inspiratory burst period, while higher concentrations (1 μM) decreased the burst period duration. Using specific pharmacology for prostanoid receptors (EP1-4R, FPR, and DP2R), we determined that coactivation of both EP2R and EP3R is necessary for PGE2 to modulate the inspiratory burst period. Additionally, biased activation of EP3 receptors lengthened the duration of the inspiratory burst period, while biased activation of EP2 receptors shortened the burst period. To help delineate which cell populations are affected by exposure to PGE2, we analyzed single-cell RNA-Seq data derived from preBötC cells. Transcripts encoding for EP2R (Ptger2) were differentially expressed in a cluster of excitatory neurons putatively located in the preBötC. A separate cluster of mixed inhibitory neurons differentially expressed EP3R (Ptger3). Our data provide evidence that EP2 and EP3 receptors increase the duration of the inspiratory burst period at 1–10 nM PGE2 and decrease the burst period duration at 1 μM. Further, the biphasic dose response likely results from differences in receptor binding affinity among prostanoid receptors.

Sex Difference in Renal Artery Contractility in a Novel CRISPR/Cas9‐Generated P2X7 Knockout Rat

Purinergic signaling regulates numerous physiological processes, including renal hemodynamics. The P2X7 receptor (P2X7) is a non‐selective cation channel activated by extra‐cellular adenosine triphosphate (ATP) which plays an important role in immune cell response and inflammation. P2X7 is also expressed in endothelial cells. In the kidney, acute pharmacological blockade of P2X7 increases renal perfusion in healthy rats (Kidney Int. 88(5): 1079–1087, 2015). Here, we generated a novel P2X7 knockout (KO) rat and examined ex vivo contractility of the renal artery in male and female rats.A P2rx7 global KO rat was generated on a F344 IcoCrl background using CRISPR/Cas9 technology. KO was achieved via a two‐adenine insertion predicted to introduce a premature stop codon in exon 2. RNA and protein were extracted from KO and wild‐type (WT) kidneys to detect P2X7 by RTqPCR (n=4‐5/group) and western blotting (n=2/group). Bone marrow‐derived macrophages (BMDM) from KO and WT rats were used to assess P2X7 function by measuring interleukin (IL)‐1β release (ELISA) after stimulation with lipopolysaccharide (LPS) (1μg/mL, 4h) and ATP (3mM, 1h) (n=5‐6/group). RNA from BMDM was extracted after LPS to detect P2rx7, Il1b, Tnf and Il6 mRNA by RTqPCR (n=3/group), and Griess reaction was used to assess BMDM nitric oxide (NO) production (n=5‐6/group). One main renal artery from 2.5‐to‐4.5‐month‐old male and female KO and WT rats (n=8‐11/group) was mounted on a wire myograph to assess the response to 125 mM potassium physiological solution (KPSS) and to cumulative doses (1 nM‐30 μM) of phenylephrine (PE), acetylcholine (ACh) and sodium nitroprusside (SNP).The P2rx7 mutation in exon 2 was confirmed by genome sequencing (Sanger). P2rx7 mRNA abundance was decreased by ~60% in kidney and ~53% in BMDM from KO vs WT. P2X7 protein was detected in kidneys from WT but not from KO rats. P2X7 KO did not alter BMDM polarization after LPS stimulation. The induction of NO production was not different between genotypes (2.14±0.26 vs 2.31±0.43 μM, NS) and mRNA expression of M1 markers Il1b, Tnf and Il6 was similarly increased. In BMDM from male WT rats, ATP stimulation induced IL‐1β release; this response was markedly suppressed in KO BMDM (2577±70 vs 413±19 pg/mL, p<0.0001). In the renal artery, the constriction induced by KPSS depolarization or by PE was not different between genotypes for either sex. In male rats, the maximum vasodilation induced by ACh was significantly decreased in KO compared to WT (IC50=0.25 vs 0.20 μM, NS; %Imax= 24±3 vs 10±2%, p=0.0015); this was not observed in females (IC50=0.11 vs 8.28 μM, NS; %Imax= 19±4 vs 12±4%, NS). The dilation induced by the direct NO donor, SNP was not affected by either genotype or sex.Global genetic deletion of P2X7 impaired endothelial‐dependent vasodilation in the renal artery of male rats that may reflect diminished NO production in response to ACh. Thus, the increased renal perfusion induced by P2X7 inhibitor may arise from off‐target effects. The potential sexual dimorphism in vascular expression and/or function of P2X7 requires further investigation.

Soluble Protein Oligomers Induce Endoplasmic Reticulum Stress and Exacerbated Vasodilation via Nitric Oxide Release in Resistance Arteries from Male and Female Mice

Amyloid b (Ab) protein which gives rise to small, toxic, and soluble Ab oligomers (SPOs) is capable of interfering with synaptic function and integrity, thus memory loss in Alzhemier’s disease (AD). Further their accumulation induces endoplasmic reticulum (ER) stress, leading to cell death in several cardiovascular diseases. Our previous data showed for the first time that in arteries from both male and female mice, SPOs exert their vasoactive effects by inducing ER stress in acute conditions, resulting in pathologically exacerbated endothelium‐dependent vasodilation due to loss of vascular autoregulation, edema formation and chronic, irreversible vascular collapse. ER stress was also confirmed with anti‐KDEL antibody staining, specific for ER resident chaperones Grp78/94 and visualized with multiphoton fluorescent confocal microscopy. Additionally, we showed that ER stress induced further release of SPOs in arteries from female mice, showing a positive feedforward mechanism. Based on these prior research, we hypothetized that SPOs lead to exacerbated and deleterious vasodilation via nitric oxide synthase (NOS) and nicotinamide adenine dinucleotide phosphate oxidase (NADPH oxidase) activation. Mesenteric resistance arteries (MRA) were isolated from male and female C57BL/6J mice (~14 weeks old) (n=4‐7). The arteries were pre‐incubated for 30 minutes in the presence or absence of specific hydrogen peroxide (H2O2) inhibitor, catalase (500u/ml); NOS inhibitor, L‐NAME (100µM); cyclooxygenase (COX) inhibitor, Indomethacin (10µM); NADPH oxidase‐1 (Nox1) inhibitor (10 µM) and Nox 1/4 inhibitor (GKT136901,10μM), followed by incubation with SPOs (0.1 μM) for 45 minutes. Subsequently, concentration response curves to acetylcholine (1 nM‐30 µM) were performed after contraction with phenylephrine (10 µM). No differences were observed in the presence of catalase, indomethacin or GKT136901. However, L‐NAME was able to eliminate the SPOs ability to increase vasodilation in arteries form both male and female mice (Fig 1. A and B, Two‐way ANOVA). We therefore concluded that the mechanism that SPOs use to pathologically exacerbate endothelium‐dependent vasodilation in the resistance arteries is NO dependent, and was observed in arteries from both male and female mice. This could be one of the mechanisms that AD leads to loss of myogenic tone and edema in the microvasculature.

Atorvastatin Modulates the Function and Trafficking of Dopamine Transporters

IntroductionDopaminergic signaling is involved in learning, memory, reward, movement, and emotion. The dopamine transporter (DAT) is the primary mechanism for terminating this signaling via DA uptake. A few lines of evidence indicate that DAT function is regulated by membrane cholesterol. Cholesterol is a critical component of plasma membranes and plays an essential role in regulating membrane fluidity, protein compartmentalization, and function. Statins are cholesterol‐lowering medications that have been linked to mild cognitive impairment. The goal of this project was to examine the effects of atorvastatin‐mediated reductions in cholesterol content on DAT function and trafficking.MethodsCultured cells & cholesterol measurement: HA‐tagged human DAT (hDAT) was stably expressed in neuroblastoma 2a (N2A) cells using G418 as selection. Cells were treated with atorvastatin (0.1, 0.3, 1.0, or 10 μM) and membrane cholesterol content was measured using Amplex Red.[3H]DA Uptake: After 24hr atorvastatin or vehicle treatment, cells were incubated with 15 nM [3H]DA in the presence of various concentrations of non‐labeled DA and [3H]DA uptake was measured to determine Vmax and Km.Cocaine inhibition of DA uptake & cholesterol replenishment: Cells were treated for 24hr with atorvastatin (1 μM) or vehicle and incubated with 15 nM [3H]DA and increasing concentrations of cocaine (100 nM‐10 μM). To determine if there was a causal effect of reduced cholesterol content and cocaine inhibition of DA uptake, atorvastatin‐treated cells were replenished with water soluble cholesterol (~40 mg per gram solid; MβCD) prior to cocaine inhibition.Amphetamine‐induced, DAT‐mediated DA efflux: Cells treated for 24hrs with either atorvastatin (1 μM) or vehicle were preloaded with 30 nM of [3H]DA. Then, cells were incubated with amphetamine (1 or 10 μM). AMPH‐induced DA efflux was determined by measuring DA content in the medium across time.Surface DAT Biotinylation: Basal surface levels of DAT in atorvastatin and vehicle treated cells, as well as DA‐induced DAT internalization, were measured by surface biotinylation using membrane non‐permeable sulfo‐NHS‐SS biotin.ImmunocytochemistryWe measured levels of F‐actin and G‐actin to determine the effects of atorvastatin treatment on actin dynamics using immunocytochemistry.ResultsWe found that atorvastatin dose dependently reduced membrane cholesterol content. This reduction was accompanied by decreases in the maximal rate of DA uptake and by decreases in the potency of cocaine to inhibit [3H]DA uptake. In contrast, atorvastatin treatment did not significantly alter AMPH‐induced DA efflux. Importantly, the reduced ability of cocaine to inhibit DA uptake was reversed by replenishing cholesterol in atorvastatin‐treated cells, suggesting a direct causal effect of reduced cholesterol content and cocaine inhibition. Lastly, we found that atorvastatin disrupted actin dynamics and abolished DA‐induced DAT internalization. The present study provides evidence for the first time that atorvastatin dysregulates DAT function and trafficking. Furthermore, this dysregulation may be associated with the cognitive deficits observed in patients receiving atorvastatin treatment.

Switching on-off-on colorimetric sensor based on Fe-N/S-C single-atom nanozyme for ultrasensitive and multimodal detection of Hg2+

Single-atom nanozymes (SAzymes) as a new class of efficient nanozymes have attracted extensive research interest due to their high catalytic activity and specificity. However, it is challenging to develop a novel nanoenzyme with high activity, good stability and reproducibility. In this paper, the nitrogen and sulfur coordinated Fe-N/S-C SAzymes were synthesized using peanuts shells as carbon, nitrogen and sulfur source. It shows high oxidase-like activities due to the doping of S induced geometric and electronic effects, which is further confirmed by density functional theory calculations. The prepared Fe-N/S-C SAzymes with the remarkable oxidase-mimicking activity could oxidize TMB to blue oxTMB, but the GSH can inhibit the oxidation of TMB resulting in blue fading. However, when Hg2+ is added into above system, Hg2+-SH complexes are generated attributed to a high affinity between GSH and Hg2+, ultimately leading to blue recovery. Based on this phenomenon, we constructed a novel “on-off-on” colorimetric sensor for the simultaneous detection of GSH (off) and Hg2+ (on), and the signal is acquired by various modes such as naked eye, UV–Vis spectrometer and smartphone. The colorimetric detection mode based on a smartphone showed a good linear response from 10 to 80 μM for GSH with a detection limit of 3.92 μM, and for Hg2+ with a linear range of 1 nM-10 μM and LOD of 0.17 nM, which is more suitable for routine laboratory applications. More importantly, the proposed colorimetric sensor has been successfully applied to the detection of GSH and Hg2+ in real samples with good analytical performance. This work not only provides a simple and cost-effective method to detect GSH and Hg2+ but also makes a certain contribution to environmental protection.

Sodium Ligninsulfonate-assisted Synthesis of Lithium Bismuthate/bismuth Oxide Microspheres and Solar Light Photocatalytic Performance

Background: Great attention has been paid to the environmental pollution by organic dyes, which are difficult to be degraded in the natural environment and have been an unavoidable and urgent global problem. It is essential to develop green wastewater treatment technology with high removal efficiency and low-cost for protecting the surrounding and human health. Objective: The aim of the research is to synthesize lithium bismuthate/bismuth oxide microspheres with good photocatalytic performance for the removal of gentian violet (GV). Methods: Lithium bismuthate/bismuth oxide microspheres were successfully prepared by a sodium ligninsulfonate-assisted hydrothermal synthesis route. The lithium bismuthate/bismuth oxide microspheres were investigated by X-ray diffraction (XRD), scanning electron microscopy (SEM), Fourier Transform infrared spectroscopy (FTIR), and solid UV-vis diffuse reflectance spectrum. Results: XRD pattern and SEM observation show that the lithium bismuthate/bismuth oxide microspheres are composed of cubic LiBi12O18.50 and monoclinic Bi2O3 with a diameter of 250 nm-1 μm. Irregular microscale and nanoscale particles are formed under low hydrothermal temperature, low sodium ligninsulfonate concentration, and short duration time. By increasing the sodium ligninsulfonate concentration, hydrothermal temperature, and duration time, irregular particles are transferred into microspheres. Lithium bismuthate/bismuth oxide microspheres possess a band gap energy of 1.85 eV, suggesting good visible light absorption ability. The photocatalytic removal ability for GV is enhanced by prolonging light irradiation time and microspheres dosage. GV solution with the concentration of 10 mg•L-1 is able to be totally degraded by 10 mg lithium bismuthate/bismuth oxide microspheres in 10 mL GV solution under solar light irradiation for 6 h. Conclusion: The lithium bismuthate/bismuth oxide microspheres show good photocatalytic removal ability toward GV in wastewater under solar light irradiation.

Stem cell-derived extracellular vesicle therapy for acute brain insults and neurodegenerative diseases

Stem cell-based therapy is a promising approach for treating a variety of disorders, including acute brain insults and neurodegenerative diseases. Stem cells such as mesenchymal stem cells (MSCs) secrete extracellular vesicles (EVs), circular membrane fragments (30 nm−1 μm) that are shed from the cell surface, carrying several therapeutic molecules such as proteins and microRNAs. Because EV-based therapy is superior to cell therapy in terms of scalable production, biodistribution, and safety profiles, it can be used to treat brain diseases as an alternative to stem cell therapy. This review presents evidences evaluating the role of stem cell-derived EVs in stroke, traumatic brain injury, and degenerative brain diseases, such as Alzheimer’s disease and Parkinson’ disease. In addition, stem cell-derived EVs have better profiles in biocompatibility, immunogenicity, and safety than those of small chemical and macromolecules. The advantages and disadvantages of EVs compared with other strategies are discussed. Even though EVs obtained from native stem cells have potential in the treatment of brain diseases, the successful clinical application is limited by the short half-life, limited targeting, rapid clearance after application, and insufficient payload. We discuss the strategies to enhance the efficacy of EV therapeutics. Finally, EV therapies have yet to be approved by the regulatory authorities. Major issues are discussed together with relevant advances in the clinical application of EV therapeutics.

Detection of picric acid in industrial effluents using multifunctional green fluorescent B/N-carbon quantum dots

Carbon quantum dots have recently gained widespread attention due to their excellent physicochemical features. The rapid escalation in the dumping of hazardous chemicals into water, spurred demand for developing efficient and selective sensors for toxic chemicals. Herein, we have developed a novel fluorescence sensor for picric acid which is a major pollutant in industrial effluents. The new strategy exploits the development of a fluorescence sensor based on N-doped carbon quantum dots (N-CQDs) followed by boron functionalization. The N-CQDs were synthesized in a rapid single-step microwave technique by employing L-serine and citric acid. Subsequent boron functionalization of N-CQDs was carried out using boric acid for the synthesis of Boron-nitrogen carbon quantum dots (B/N-CQDs). The B/N-CQDs were found to exhibit high quantum yield (24%), good water solubility, outstanding photostability features, and bright green fluorescence under UV light. The morphology of B/N-CQDs is spherical, with scattered particle sizes ranging from 2 to 8 nanometers. Furthermore, B/N-CQDs were found to be an effective fluorescence probe for the selective and sensitive detection of picric acid, with a good linear range of 37 nM-30 µM and a detection limit of 1.8 nM. The Photoluminescence (PL) intensity of B/N-CQDs was selectively quenched by picric acid. The quenching mechanism was conclusively established using fluorescence lifetime decay studies. Moreover, the synthesized B/N-CQDs was successfully employed for the analysis of picric acid from industrial effluents and cell imaging with Hela cells to showcase the utility of the developed fluorescent probe.

Hydrogen Bond Organic Frameworks as a Novel Electrochemiluminescence Luminophore: Simple Synthesis and Ultrasensitive Biosensing.

Nowadays, continuous efforts have been devoted to searching highly efficient electrochemiluminescence (ECL) emitters for applications in clinical diagnosis and food safety. In this work, triazinyl-based hydrogen bond organic frameworks (Tr-HOFs) were synthesized by N···H hydrogen bond self-assembly aggregation, where 6,6'-(1,4-phenylene)bis(1,3,5-triazine-2,4-diamine) (phenyDAT) was prepared via the cyclization reaction and behaved as a novel ligand. Impressively, the resulting Tr-HOFs showed strong ECL responses with highly enhanced ECL efficiency (21.3%) relative to the Ru(bpy)32+ standard, while phenyDAT hardly showed any ECL emission in aqueous phase. The Tr-HOFs innovatively worked as a new ECL luminophore to construct a label-free biosensor for assay of kanamycin (Kana). Specifically, the ECL response greatly weakened upon assembly of captured DNA with ferrocene (cDNA-Fc) onto the Tr-HOFs-modified electrode, while the ECL signals were adversely recovered by releasing linked DNA (L-DNA) from double-stranded DNA (dsDNA, hybridization of aptamer DNA (aptDNA) with L-DNA) due to the specific recognition of Kana with the aptDNA combined by the linkage of L-DNA and cDNA-Fc on the electrode. The as-built sensor showed a broadened linear range (1 nM-10 μM) and a limit of detection (LOD) down to 0.28 nM, which also displayed satisfactory results in the analysis of Kana in the milk and diluted human serum samples. This work offers a novel pathway to design an ECL emitter with organic molecules, holding great promise in biomedical analysis and food detection.

A novel covalent organic framework with multiple adsorption sites for removal of Hg2+ and sensitive detection of nitrofural

It is urgent to control serious drug pollution and heavy metal ion pollution. Covalent organic framework (COF) is becoming a promising material for the removal of heavy metal ion and adsorption of organic small molecule because of the designed functional sites, ordered pore channel, high specific surface area and abundant π electrons. Herein, 2,4,6-triformylphloroglucinol (TFP) and benzene-1,3,5-tricarbohydrazide (BTH) was used to design a stable yet efficient beta-ketoenamine COF (COFTFP-BTH) adsorbent, which would provide multiple adsorption sites of Hg2+ by dense acylhydrazine. The maximum adsorption capacity of COFTFP-BTH toward Hg2+ achieved 909 mg g−1, coinciding with Langmuir adsorption and pseudo-second-order adsorption kinetic model, with the h value as high as 71.43 mg g−1 min−1. COFTFP-BTH was also grown on NH2-carbon nanotubes (CNT) to form a novel COFTFP-BTH@NH2-CNT composite for sensitive detection of nitrofural (NF). The NF sensor based on COFTFP-BTH@NH2-CNT showed a low detection limit (3.2 nM) and a wide linear range (9.6 nM-100 μM). The excellent performances were also attributed to the ordered channels of COFTFP-BTH exposing more adsorption sites of Hg2+ and NF, as well as the good stability of beta-ketoenamine COFTFP-BTH.

Label-free Hg(II) electrochemiluminescence sensor based on silica nanoparticles doped with a self-enhanced Ru(bpy)32+-carbon nitride quantum dot luminophore

Herein, a label-free, self-enhanced electrochemiluminescence (ECL) sensing strategy for divalent mercury (Hg(II)) detection was presented. First, a novel self-enhanced ECL luminophore was prepared by combining the ECL reagent tris(2, 2′-bipyridyl) dichlororuthenium(II) hexahydrate (Ru(bpy)32+) and its co-reactant carbon nitride quantum dots (CNQDs) via electrostatic interactions. In contrast to traditional ECL systems where the emitter and its co-reactant underwent an intermolecular reaction, the self-enhanced ECL system exhibited a shortened electron-transfer distance and enhanced luminous efficiency because the electrons transferred from CNQDs to oxidized Ru(bpy)32+ via an intramolecular pathway. Furthermore, the as-prepared self-enhanced ECL material was encapsulated in silica (SiO2) nanoparticles to generate a Ru-QDs@SiO2 luminophore. Based on the different affinity of Ru-QDs@SiO2 nanoparticles for single-stranded DNA (ssDNA) and Hg(II)-triggered double-stranded DNA (dsDNA), a label-free ECL biosensor for Hg(II) detection was developed as follows: in the absence of Hg(II), ssDNA was adsorbed on Ru-QDs@SiO2 surface via hydrogen bond, electrostatic, and hydrophobic interaction. Thus, quenched ECL signal was observed. On the contrary, in the presence of Hg(II), stable dsDNA was formed and carried the ssDNA separating from Ru-QDs@SiO2 surface, resulting in most of Ru-QDs@SiO2 existing in their free state. Therefore, a recovered ECL intensity was obtained. On this basis, Hg(II) was measured by the proposed method in the range of 0.1 nM-10 μM, with a detection limit of 33 pM. Finally, Hg(II) spiked in water samples was measured to evaluate the practicality of the fabricated biosensor.

The T-type Calcium Channel Cav3.1 in Y79 Retinoblastoma Cells is Regulated by the Epidermal Growth Factor Receptor via the MAPK Signaling Pathway

ABSTRACTPurposeRetinoblastoma is the most frequent intraocular cancer in children. It is also one of the most common causes for enucleation and carries a significant morbidity rate in affected individuals. Hence, studies on its pathophysiological and growth regulatory mechanisms are urgently needed to identify more effective novel therapeutics.MethodsUsing the Y79 retinoblastoma cell line, we investigated the electrophysiological and functional activities of the T-type voltage-gated calcium channel Cav3.1, that is constitutively expressed in these cells. We also analyzed the Akt and MAPK signaling pathways downstream of the epidermal growth factor receptor (EGFR) to understand the mechanism responsible for the inhibition of Cav3.1.ResultsWe demonstrate that the EGFR inhibitor Afatinib significantly reduced cell viability and Cav3.1 mRNA expression and electrophysiological activity. At low concentrations (1 µM), Afatinib reduced the amplitude of Cav3.1 current density, whereas at a high concentration (10 µM), it completely abolished the voltage-gated calcium current. Our results show that inhibition of the MAPK pathway by a specific inhibitor VX-11e affected the Cav3.1 current in a dose-dependent manner. VX-11e (50 nM-1 µM) treatment reduced Cav3.1 current densities in Y79 cells, with complete abolishment of Cav3.1 current at higher concentrations (5 µM). We also demonstrate that the specific inhibition of the Akt kinase (using MK-2206) had no effect on the Cav3.1 currents.ConclusionOur study provides a functional relationship between the MAPK pathway and EGFR signaling and indicates that the MAPK signaling pathway mediates the control of Cav3.1 by EGFR in retinoblastoma.

Biophysical characterization of full-length oleosin in dodecylphosphocholine micelles.

Oleosin is a hydrophobic protein that punctuates the surface of plant seed lipid droplets, which are 20 nm-100 μm entities that serve as reservoirs for high-energy metabolites. Oleosin is purported to stabilize lipid droplets, but its exact mechanism of stabilization has not been established. Probing the structure of oleosin directly in lipid droplets is challenging due to the size of lipid droplets and their high degree of light scattering. Therefore, a medium in which the native structure of oleosin is retained, but is also amenable to spectroscopic studies is needed. Here, we show, using a suite of biophysical techniques, that dodecylphosphocholine micelles appear to support the tertiary structure of the oleosin protein (i.e., hairpin conformation) and render the protein in an oligomeric state that is amenable to more sophisticated biophysical techniques such as NMR.

Examination of the contribution of Nav1.7 to axonal propagation in nociceptors.

Nav1.7 is a promising drug target for the treatment of pain. However, there is a mismatch between the analgesia produced by Nav1.7 loss-of-function and the peripherally restricted Nav1.7 inhibitors, which may reflect a lack of understanding of the function of Nav1.7 in the transmission of nociceptive information. In the periphery, the role of Nav1.7 in transduction at nociceptive peripheral terminals has been comprehensively examined, but its role in axonal propagation in these neurons is less clearly defined. In this study, we examined the contribution of Nav1.7 to axonal propagation in nociceptors using sodium channel blockers in in vivo electrophysiological and calcium imaging recordings in mice. Using the sodium channel blocker tetrodotoxin (TTX) (1-10 µM) to inhibit Nav1.7 and other tetrodotoxin-sensitive sodium channels along the sciatic nerve, we first showed that around two-thirds of nociceptive L4 dorsal root ganglion neurons innervating the skin, but a lower proportion innervating the muscle (45%), are blocked by TTX. By contrast, nearly all large-sized cutaneous afferents (95%-100%) were blocked by axonal TTX. Many cutaneous nociceptors resistant to TTX were polymodal (57%) and capsaicin sensitive (57%). Next, we applied PF-05198007 (300 nM-1 µM) to the sciatic nerve between stimulating and recording sites to selectively block axonal Nav1.7 channels. One hundred to three hundred nanomolar PF-05198007 blocked propagation in 63% of C-fiber sensory neurons, whereas similar concentrations produced minimal block (5%) in rapidly conducting A-fiber neurons. We conclude that Nav1.7 is essential for axonal propagation in around two-thirds of nociceptive cutaneous C-fiber neurons and a lower proportion (≤45%) of nociceptive neurons innervating muscle.