High SERS performance of functionalized carbon dots in the detection of dye contaminants.

INTRODUCTION: The long-term overuse of malachite green (MG) has potential carcinogenic, teratogenic, and mutagenic effects. The functional nanocomposite is novel and challenging to construct and implement through surface enhanced Raman scattering (SERS) strategy to reveal the contributions in application. OBJECTIVES: The novel Ag-CDs (carbon dots)-PBA (phenyl boric acid) nanocomposite was constructed by a facile route to detect toxic MG molecule with high SERS sensitivity and good uniformity. METHODS: The enhanced substrate used for the detection of MG has been successfully constructed using PBA modulated Ag-CDs on a structured surface with rich binding sites. RESULTS: The fabricated Ag-CDs-PBA substrate can be used to analyze various probe molecules exhibiting high sensitivity, good signal reproducibility, and excellent stability. The mechanism between components has been proved by calculations originating from the plasmonic Ag and active electronic transmission among the bridging CDs and PBA via the close spatial π-π effect. In addition, the accelerated separation of electron-hole pairs was triggered to further improve the SERS activity of the hybrid via a bidirectional charge transfer (CT) process. Significantly, the Ag-CDs-PBA system shows distinctive selectivity, in which PBA can hinder the interference of other species without specific hydroxyl groups. CONCLUSION: Based on this deeper insight on plasmon-mediated mechanism, the SERS substrate was successfully practiced for quantitative determination in real water and fish samples. The strategy developed promises to be a new sensor technology and has great potential for environmental and food safety applications.

p53-dependent HIF-1α /autophagy mediated glycolysis to support Cr(VI)-induced cell growth and cell migration.

Cr(VI) is known to be seriously toxic and carcinogenic. Hypoxia-inducible factor-1α (HIF-1α) is a crucial regulator to promote tumor development. In this study, we found that Cr(VI) significantly increased the expression of HIF-1α in A549 cells and in lung of BALB/c mice but not in HELF cells. Treatment with Lificiguat (YC-1), HIF-1α inhibitor, or CoCl2, HIF-1α inducer, could alter Cr(VI)-induced autophagy, glycolysis, and cell growth in A549 cells but not in HELF cells, validating the involvement of HIF-1α in these effects of Cr(VI) in A549 cells. Co-treatments of pcATG4B with YC-1, or siATG4B with CoCl2 demonstrated the role of HIF-1α / autophagy axis in inducing glycolysis and cell growth in A549 cells. In HELF cells, however, only autophagy but not HIF-1α played a role in inducing glycolysis. The protein level of p53 was significantly lower in A549 cells than in HELF cells. RITA, a p53 inducer, attenuated Cr(VI)-induced HIF-1α and LC3-II in A549 cells, suggesting that p53 might be the mechanism underlying the different effects of Cr(VI) on HIF-1α in A549 and HELF cells. Thus, p53-dependent HIF-1α / autophagy-mediated glycolysis plays a role in facilitating Cr(VI)-induced carcinogenesis.

De Novo Generation and Identification of Novel Compounds with Drug Efficacy Based on Machine Learning.

One of the main challenges in small molecule drug discovery is finding novel chemical compounds with desirable activity. Traditional drug development typically begins with target selection, but the correlation between targets and disease remains to be further investigated, and drugs designed based on targets may not always have the desired drug efficacy. The emergence of machine learning provides a powerful tool to overcome the challenge. Herein, a machine learning-based strategy is developed for de novo generation of novel compounds with drug efficacy termed DTLS (Deep Transfer Learning-based Strategy) by using dataset of disease-direct-related activity as input. DTLS is applied in two kinds of disease: colorectal cancer (CRC) and Alzheimer's disease (AD). In each case, novel compound is discovered and identified in in vitro and in vivo disease models. Their mechanism of actionis further explored. The experimental results reveal that DTLS can not only realize the generation and identification of novel compounds with drug efficacy but also has the advantage of identifying compounds by focusing on protein targets to facilitate the mechanism study. This work highlights the significant impact of machine learning on the design of novel compounds with drug efficacy, which provides a powerful new approach to drug discovery.

Crebanine ameliorates ischemia-reperfusion brain damage by inhibiting oxidative stress and neuroinflammation mediated by NADPH oxidase 2 in microglia.

BACKGROUND: The urgent challenge for ischemic stroke treatment is the lack of effective neuroprotectants that target multiple pathological processes. Crebanine, an isoquinoline-like alkaloid with superior pharmacological activities, presents itself as a promising candidate for neuroprotection. However, its effects and mechanisms on ischemic stroke remain unknown. METHODS: The effects of crebanine on brain damage following ischemic stroke were evaluated using the middle cerebral artery occlusion and reperfusion (MCAO/R) model. Mechanism of action was investigated using both MCAO/R rats and lipopolysaccharide (LPS)-activated BV-2 cells. RESULTS: We initially demonstrated that crebanine effectively ameliorated the neurological deficits in MCAO/R rats, while also reducing brain edema and infarction. Treatment with crebanine resulted in the up-regulation of NeuN+ fluorescence density and down-regulation of FJB+ cell count, and mitigated synaptic damage. Crebanine attenuated the hyperactivation of nicotinamide adenine dinucleotide phosphate (NADPH) oxidase 2 (NOX2) by downregulating NADP+ and NADPH levels, suppressing gp91phox and p47phox expressions, and reducing p47phox membrane translocation in Iba-1+ cells. Additionally, crebanine reduced the quantity of Iba-1+ cells and protein expression. Correlation analysis has demonstrated that the inhibition of NOX2 activation in microglia is beneficial for mitigating I/R brain injuries. Moreover, crebanine exhibited significant antioxidant properties by down-regulating the expression of superoxide anion and intracellular reactive oxygen species in vivo and in vitro, and reducing lipid and DNA peroxidation. Crebanine exerted anti-inflammatory effect, as evidenced by the reduction in the expressions of nitric oxide, interleukin 1ß, tumor necrosis factor α, interleukin 6, and inducible nitric oxide synthase. The effect of crebanine was achieved through the suppression of nuclear factor-kappa B (NF-κB) and mitogen-activated protein kinases (MAPK) signaling pathway. This is supported by evidence showing reduced NF-κB p65 promoter activity and nucleus translocation, as well as suppressed IκBα phosphorylation and degradation. Additionally, it inhibited the phosphorylation of ERK, JNK, and p38 MAPKs. Importantly, the anti-oxidative stress and neuroinflammation effects of crebanine were further enhanced after silencing gp91phox and p47phox. CONCLUSION: Crebanine alleviated the brain damages of MCAO/R rats by inhibiting oxidative stress and neuroinflammation mediated by NOX2 in microglia, implying crebanine might be a potential natural drug for the treatment of cerebral ischemia.

1H NMR-guided isolation of hasubanan alkaloids from the alkaloidal extract of Stephania longa.

1H NMR-guided fractionation led to the isolation of 16 alkaloids from the alkaloidal extract of Stephania longa, including 11 new hasubanan alkaloids (1-11) and five known alkaloids (12-16). Interestingly, compounds 2 and 11 are typically considered protonated tertiary amine compounds, whereas compounds 1 and 10 are regarded as oxidized versions of the corresponding compounds. Their gross structures were determined through an extensive analysis of spectroscopic data (NMR (nuclear magnetic resonance) and HRESIMS (high resolution electrospray ionization mass spectroscopy)), and their absolute configurations were established by comparing their experimental and calculated electronic circular dichroism (ECD) spectra. The new (3) and a known (12) compounds in all isolates displayed stronger antineuroinflammatory effects (IC50 values of 1.8 and 11.1 µM, respectively) than minocycline (IC50 value of 15.5 µM) against NO production on LPS-activated BV2 cells.

Cbl induced ubiquitination of HER2 mediate immune escape from HER2-targeted CAR-T.

Breast cancer (BC) with high HER2 expression has higher recurrence rate and worse prognosis, and its immunotherapy is promising. Based on the high expression of HER2, develop Chimeric Antigen Receptor T-cell (CAR-T) and PDL-1 immunotherapy, and study the molecular pathways of related immune cells and recurrence. HER2-CAR-T cells were constructed using retroviruses, and their specific recognition and immune effects on HER2+ BC cells were verified by in vivo and in vitro experiments. PDL-1 was used as adjuvant immunotherapy, knocking down PDL-1 in tumor cells or dendritic cells, or depleted macrophages to study immune pathways. The negative regulation of HER2 by cbl was determined by IP, ubiquitination experiments, and segmented plasmids, elucidating the molecular mechanism of HER2+ BC recurrence after immunotherapy. HER2-CAR-T specifically recognizes HER2-positive tumor cells and inhibits tumor growth in vivo and in vitro, and anti-PDL1 treatment enhances the therapeutic effect of HER2-CAR-T on tumors. HER2-CART therapy eradicated solid tumors after PDL1 knockdown in dendritic cells. Immunotherapy of relapsed tumors lost HER2 expression by upregulating cbl. HER2-CAR-T shows specific recognition of HER2+ cells and can mediate immune response therapy with the cooperation of PDL-1.

Lignanamides from the stems of Piper hancei maxim. and their anti-inflammatory and cytotoxic activities.

Four new lignanamides, hancamides A - D (1-4), together with four known analogs (5-8), were isolated from the stems of Piper hancei Maxim. Their structures were determined based on 1D and 2D NMR, IR, UV, and HR-ESIMS spectroscopic analysis as well as by comparison with the reported data. All the isolates exhibited potential inhibitory effects on NO production in LPS-induced BV-2 microglial cells, with IC50 values of 4.26-40.68 µM. Moreover, compounds 2 and 8 displayed moderate cytotoxic activities against MGC-803, HepG2, SKOV-3, T24, and HeLa cells, with IC50 values ranging from 13.57 to 34.20 µM, respectively.

The Serum SIRT1 Protein is Associated with the Severity of Injury and Neurological Recovery in Mice with Traumatic Spinal Cord Injury.

The present study aimed to investigate the association between the serum SIRT1 protein and the severity of spinal cord injury (SCI) as well as the neurological recovery in mice. In this study, the wild-type (WT), Mx1-Cre+ SIRT1loxP/loxP (Mx1), and LCK-Cre+SIRT1loxP/loxP (LCK) mice were subjected to sham surgery, mild, moderate, or severe SCI, respectively. The serum was collected at intervals of 12 h, 1 day (d), 3 d, 5 d, 7 d, 10 d, 14 d, and 21 d after the injury. The locomotor function of all the animals was assessed using the Basso mouse scale (BMS) and the serum SIRT1 proteins were analyzed using enzyme-linked immunosorbent assay (ELISA). The results demonstrated that about 7-10 d after SCI, the levels of SIRT1 protein in the serum correlated significantly with the severity of the injury and at 28 d post-injury, there was a distant neurological recovery (BMS score). The serum SIRT1 concentration in both the Mx1 and LCK mice in the sham group was significantly reduced compared to that in the WT mice, and there was a delayed increase in the serum SIRT1 levels after injury. These findings indicate that the SIRT1 concentrations in the serum of the SCI mice closely correlated with the acute severity and neurological outcome.

Regulation of Superoxide by BAP31 through Its Effect on p22phox and Keap1/Nrf2/HO-1 Signaling Pathway in Microglia.

Reactive oxygen species (ROS) production by activation of microglia is considered to be a major cause of neuronal dysfunction, which can lead to damage and death through direct oxidative damage to neuronal macromolecules or derangement of neuronal redox signaling circuits. BAP31, an integral ER membrane protein, has been defined as a regulatory molecule in the CNS. Our latest studies have found that BAP31 deficiency leads to activation of microglia. In this study, we discovered that BAP31 deficiency upregulated LPS-induced superoxide anion production in BV2 cells and mice by upregulating the expression level of p22phox and by inhibiting the activation of Nrf2-HO-1 signaling. Knockdown of p22phox/keap1 or use of an NADPH oxidase inhibitor (apocynin) reversed the production of superoxide anion and inflammatory cytokines, which then reduced neuronal damage and death in vitro and in vivo. These results suggest that BAP31 deficiency contributes to microglia-related superoxide anion production and neuroinflammation through p22phox and keap1. Furthermore, the excess superoxide anion cooperated with inflammatory cytokines to induce the damage and death of neurons. Thus, we determined that BAP31 is an important regulator in superoxide anion production and neuroinflammation, and the downstream regulators or agonists of BAP31 could therefore be considered as potential therapeutic targets in microglial-related superoxide anion production and neuroinflammation.

Complexation of Cyclic Glutarimidedioxime with Cerium: Surrogating for Redox Behavior of Plutonium.

The complexation of cerium with glutarimidedioxime (H2L) was studied by potentiometry, ESI-mass spectrometry, and cyclic voltammetry. Crystallization of [CeIV(HL)3]+ from Ce3+ starting reactant indicated spontaneous complexation-driven oxidation. In aqueous solution, Ce3+ ions form three successive complexes, Ce(HL)2+, Ce(HL)2+, and Ce(HL)3 (where HL- stands for the singly deprotonated ligand). The interactions of glutarimidedioxime with metal ions are dominantly electrostatic in nature, and the stability constants of the complexes are correlated to the charge density of metal ions. Extrapolations of predicted stability constant (log ß) values were made from plotting effective charge and the ionic radius of the metal ion for Pu3+ and Pu4+. The stability constants of PuIV(HL)3+ and PuIII(HL)2+ are estimated to be 27.74 and 19.75, respectively. The differences of stability constants mean that glutarimidedioxime selectively binds Pu4+ over Pu3+ by a factor of about 8 orders of magnitude, suggesting Pu4+ would be stabilized by chelation with glutarimidedioxime. The mechanism of reduction of Pu4+ to Pu3+ in acidic solution is explained by decomposition of glutarimidedioxime through acid hydrolysis rather than a chelation-driven mechanism.

Inflammatory mechanism of cerebral ischemia-reperfusion injury with treatment of stepharine in rats.

BACKGROUND: Increasing evidence has shown that microglia-induced neuroinflammation is involved in the pathogenesis of ischemic stroke. Stepharine, one of the alkaloids extracted from Stephania japonica (Thunb.) Miers, exhibited strong inhibitory effect on microglial overactivation. However, it is not known whether it has the potential to prevent ischemic stroke. METHODS: The neuroprotective and anti-neuroinflammatory effects of stepharine were investigated in vivo and in vitro, using a rat model of middle cerebral artery occlusion (MCAO) and lipopolysaccharide (LPS)-stimulated BV-2 cells, respectively. RESULTS: In vivo, stepharine (500 µg/kg) suppressed neurological deficits scores, brain water content and cerebral infarct volume induced by MCAO. Moreover, stepharine (500 µg/kg) inhibited NeuN+ cells loss and Iba-1+ cells increase in the MCAO ischemic cortex. In vitro, stepharine (10, 30 µM) substantially inhibited nitric oxide release as well as the mRNA and protein expression of pro-inflammatory mediators [inducible nitric oxide synthase, interleukin (IL)-6, tumor necrosis factor (TNF)-α, IL-1ß] in LPS-activated BV-2 cells. LPS-induced increase of TLR4 expression, IκBα phosphorylation, and NF-κB p65 nuclear translocation was inhibited by stepharine (10, 30 µM). Molecular docking analysis showed that stepharine directly interacted with TLR4. SPR assay further confirmed that stepharine could bind to the TLR4/MD2 complex. Meanwhile, stepharine exhibited neuroprotective effects on SH-SY5Y cells cultured with LPS-treated conditioned medium. CONCLUSION: Our study demonstrated for the first time that stepharine improved the outcomes in MCAO rats, reduced neuronal loss, and suppressed microglial overactivation via the inhibition of TLR4/NF-κB pathway. These results suggest that stepharine might be a potential therapeutic agent for the treatment of ischemic stroke.

A Novel Quinolyl-Substituted Analogue of Resveratrol Inhibits LPS-Induced Inflammatory Responses in Microglial Cells by Blocking the NF-κB/MAPK Signaling Pathways.

SCOPE: The anti-neuroinflammatory effect of a novel quinolyl-substituted analogue of resveratrol (RV01) on lipopolysaccharide (LPS)-induced microglial activation is investigated, as well as the possible underlying mechanisms. METHODS AND RESULTS: Cell viability is measured using an MTT assay. Nitric oxide (NO) release is determined by nitrite assay. The interaction between RV01 and inducible nitric oxide synthase (iNOS) is studied using molecular docking. Free radical scavenging activity and reactive oxygen species (ROS) production are determined by DPPH reduction assay and DCFH-DA assay. Pretreatment with RV01 (1-30 µm) prior to LPS (1 µg mL-1 ) stimulation decreased NO release and iNOS expression without observable cytotoxicity. RV01 reduced the mRNA levels and secretion of tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6). RV01 also inhibited LPS-induced ROS production and nicotinamide adenine dinucleotide phosphate (NADPH) oxidase activation. Furthermore, RV01 decreases the protein expression of toll-like receptor 4 (TLR4) and inhibits the LPS-induced activation of the mitogen-activated protein kinase (MAPK) and nuclear transcription factor-κB (NF-κB) signaling pathways. Additionally, conditioned medium from microglia co-treated with LPS and RV01 alleviates the death of SH-SY5Y cells induced by conditioned medium from activated N9 microglial cells. Lastly, a mouse neuroinflammation model is further used to confirm the effect of RV01 in vivo. CONCLUSION: These results show that RV01 suppresses microglia-mediated neuroinflammation and protects neurons from inflammatory damage, which indicates that RV01 has great potential as a nutritional preventive strategy for neuroinflammation-related diseases.

Tamarix hohenackeri Bunge exerts anti-inflammatory effects on lipopolysaccharide-activated microglia in vitro.

BACKGROUND: Tamarix species are well known as the main host plants of Herba Cistanches, a valuable Traditional Chinese Medicine. They are also traditional medicinal plants themselves and are used to treat spleen problems, leucoderma and ocular conditions. PURPOSE: The aim of the present study was to investigate the anti-inflammatory effect of Tamarix hohenackeri Bunge. METHODS: In the present study, BV-2 microglial cells were used and stimulated with lipopolysaccharide (LPS). Cell viability was tested using the MTT assay. The release of nitric oxide (NO) was determined using the Griess assay. The mRNA level of inducible nitric oxide synthase (iNOS), tumor necrosis factor α (TNF-α), interleukin (IL)-1ß and IL-6 were investigated by quantitative real-time PCR (qRT-PCR). The protein levels of phosphorylated of IκBα, ERK and MEK, as well as the cytoplasmic and nuclear NF-κB p65 were tested by Western blot analysis. The translocation of the NF-κB p65 subunit from the cytosol to the nucleus was investigated by immunofluorescence staining. RESULTS: Ethyl acetate (EtOAc) extract of Tamarix hohenackeri Bunge significantly inhibited the release of NO. Phytochemical research was performed to produce 13 main constituents. Among them, compounds 6, 7, 10 and 13 were identified to be the effective components with anti-inflammatory activity. These compounds significantly inhibited the production of NO by LPS-activated BV-2 microglial cells. qRT-PCR showed that compounds 6 and 7 significantly suppressed the LPS-induced transcription of genes encoding pro-inflammatory mediators, including iNOS, TNF-α, IL-1ß and IL-6. Western blot analysis showed that compound 7 inhibited the LPS-induced phosphorylation of IκBα and antagonized the LPS-induced reduction of cytoplasmic NF-κB p65 and the increase of nuclear NF-κB p65. Immunofluorescence staining showed that nuclear translocation of NF-κB p65 was suppressed by compound 7. Western blot analysis showed that compound 7 inhibited the LPS-induced phosphorylation of ERK and MEK. CONCLUSION: The present study revealed, for the first time, the effective anti-inflammatory agents from T. Hohenackeri. Compound 7 exerted potent anti-inflammatory effects and its underlying mechanism may be associated with its capacity to inhibit NF-κB signaling pathway and the MEK/ERK activation in activated microglia. The compound may be potential candidate therapeutic agent for neurodegenerative diseases.

Preparation, mechanical property and cytocompatibility of freeze-cast porous calcium phosphate ceramics reinforced by phosphate-based glass.

In this study, phosphate-based glass (PG) was used as a sintering aid for freeze-cast porous biphasic calcium phosphate (BCP) ceramic, which was sintered under a lower temperature (1000°C). The phase composition, pore structure, compressive strength, and cytocompatibility of calcium phosphate composite ceramics (PG-BCP) were evaluated. The results indicated that PG additive reacted with calcium phosphate during the sintering process, forming ß-Ca2P2O7; the ions of sodium and magnesium from PG partially substituted the calcium sites of ß-calcium phosphate in BCP. The PG-BCP showed good cytocompatibility. The pore width of the porous PG-BCP ceramics was around 50µm, regardless of the amount of PG sintering aid. As the content of PG increased from 0wt.% to 15wt.%, the compressive strength of PG-BCP increased from 0.02 MP to 0.28MPa. When the PG additive was 17.5wt.%, the compressive strength of PG-BCP dramatically increased to 5.66MPa. Addition of 15wt.% PG was the critical point for the properties of PG-BCP. PG is considered as an effective sintering aid for freeze-cast porous bioceramics.

Positive correlation between neovascularization degree of carotid atherosclerosis determined by contrast-enhanced ultrasound and level of serum C-reactive protein.

BACKGROUND: Neovascularization is one of the most important risk factors for unstable plaque. This study was designed to correlate plaque thickness, artery stenosis and levels of serum C-reactive protein with the degree of intraplaque enhancement determined by contrast-enhanced ultrasound. PATIENTS AND METHODS: Contrast-enhanced ultrasound was performed on 72 carotid atherosclerotic plaques in 48 patients. Contrast enhancement within the plaque was categorized as grade 1, 2 or 3. Maximum plaque thickness was measured in short-axis view. Carotid artery stenosis was categorized as mild, moderate or severe. RESULTS: Plaque contrast enhancement was not associated with the degree of artery stenosis or with plaque thickness. Serum C-reactive protein levels were positively correlated with the number of new vessels in the plaque. C-reactive protein levels increased in the three groups(Grade 1: 3.72±1.79mg/L; Grade 2: 7.88±4.24 mg/L; Grade 3: 11.02±3.52 mg/L), with significant differences among them (F=10.14, P<0.01), and significant differences between each two groups (P<0.05). Spearman’s rank correlation analysis showed that serum C-reactive protein levels were positively correlated with the degree of carotid plaque enhancement (Rs =0.69, P<0.01). CONCLUSIONS: The combination of C-reactive protein levels and intraplaque neovascularization detected by contrast-enhanced ultrasound may allow more accurate evaluation of plaque stability.

Fluorescent BINOL-based sensor for thorium recognition and a density functional theory investigation.

A novel 1,1'-bi-2-naphthol (BINOL) derivative fluorescence sensor L-1 for the recognition of thorium ion with a fluorescence quench response. This ligand showed high selectivity and sensitivity for thorium ion recognition. Coordination effects were investigated by DFT calculations, and the coordination modes and sites were confirmed. Moreover, the coordination abilities of the L-1 ligand with Th(4+) and UO2(2+) were evaluated.