Label-free optical metabolic imaging of adipose tissues for prediabetes diagnosis.

Rationale: Prediabetes can be reversed through lifestyle intervention, but its main pathologic hallmark, insulin resistance (IR), cannot be detected as conveniently as blood glucose testing. In consequence, the diagnosis of prediabetes is often delayed until patients have hyperglycemia. Therefore, developing a less invasive diagnostic method for rapid IR evaluation will contribute to the prognosis of prediabetes. Adipose tissue is an endocrine organ that plays a crucial role in the development and progression of prediabetes. Label-free visualizing the prediabetic microenvironment of adipose tissues provides a less invasive alternative for the characterization of IR and inflammatory pathology. Methods: Here, we successfully identified the differentiable features of prediabetic adipose tissues by employing the metabolic imaging of three endogenous fluorophores NAD(P)H, FAD, and lipofuscin-like pigments. Results: We discovered that 1040-nm excited lipofuscin-like autofluorescence could mark the location of macrophages. This unique feature helps separate the metabolic fluorescence signals of macrophages from those of adipocytes. In prediabetes fat tissues with IR, we found only adipocytes exhibited a low redox ratio of metabolic fluorescence and high free NAD(P)H fraction a1. This differential signature disappears for mice who quit the high-fat diet or high-fat-high-sucrose diet and recover from IR. When mice have diabetic hyperglycemia and inflamed fat tissues, both adipocytes and macrophages possess this kind of metabolic change. As confirmed with RNA-seq analysis and histopathology evidence, the change in adipocyte's metabolic fluorescence could be an indicator or risk factor of prediabetic IR. Conclusion: Our study provides an innovative approach to diagnosing prediabetes, which sheds light on the strategy for diabetes prevention.

Thiolated Mesoporous Silica Nanoparticles as an Immunoadjuvant to Enhance Efficacy of Intravesical Chemotherapy for Bladder Cancer.

The characteristics of global prevalence and high recurrence of bladder cancer has led numerous efforts to develop new treatments. The spontaneous voiding and degradation of the chemodrug hamper the efficacy and effectiveness of intravesical chemotherapy following tumor resection. Herein, the externally thiolated hollow mesoporous silica nanoparticles (MSN-SH(E)) is fabricated to serve as a platform for improved bladder intravesical therapy. Enhanced mucoadhesive effect of the thiolated nanovector is confirmed with porcine bladder. The permeation-enhancing effect is also verified, and a fragmented distribution pattern of a tight junction protein, claudin-4, indicates the opening of tight junction. Moreover, MSN-SH(E)-associated reprogramming of M2 macrophages to M1-like phenotype is observed in vitro. The antitumor activity of the mitomycin C (MMC)-loaded nanovector (MMC@MSN-SH(E)) is more effective than that of MMC alone in both in vitro and in vivo. In addition, IHC staining is used to analyze IFN-γ, TGF-ß1, and TNF-α. These observations substantiated the significance of MMC@MSN-SH(E) in promoting anticancer activity, holding the great potential for being used in intravesical therapy for non-muscle invasive bladder cancer (NMIBC) due to its mucoadhesivity, enhanced permeation, immunomodulation, and prolonged and very efficient drug exposure.

Synergistic efficacy of telomerase-specific oncolytic adenoviral therapy and histone deacetylase inhibition in human hepatocellular carcinoma.

The telomerase-specific oncolytic adenovirus Telomelysin and the histone deacetylase inhibitor AR42 have demonstrated anticancer effects in preclinical models of human hepatocellular carcinoma (HCC). However, the clinical development of Telomelysin may be hindered by human antiviral immunity and tumor resistance. Combining oncolytic and epigenetic therapies is a viable approach for treating various cancers. This study investigated the potential synergism of Telomelysin and AR42 and the relevant underlying mechanisms. Telomelysin and AR42 exhibited synergistic antiproliferative effects in human HCC models in vitro and in vivo. Apoptosis induced by Telomelysin was significantly enhanced by AR42 in both PLC5 and Hep3B HCC cells. AR42 treatment unexpectedly attenuated the expression of the coxsackievirus and adenovirus receptor and the mRNA levels of human telomerase reverse transcriptase, which may be positively associated with the cytotoxicity of Telomelysin. Meanwhile, the cellular antiviral interferon response was not altered by AR42 treatment. Further, we found that Telomelysin enhanced Akt phosphorylation in HCC cells. AR42 reduced Telomelysin-induced phospho-Akt activation and enhanced Telomelysin-induced apoptosis. The correlation of Akt phosphorylation with drug-induced apoptosis was validated in HCC cells with upregulated or downregulated Akt signaling. Combination therapy with Telomelysin and AR42 demonstrated synergistic anti-HCC efficacy. Clinical trials investigating this new combination regimen are warranted.

Liposome-based artificial cells: From gene expression to reconstitution of cellular functions and phenotypes.

Bottom-up approaches in creating artificial cells that can mimic natural cells have significant implications for both basic research and translational application. Among various artificial cell models, liposome is one of the most sophisticated systems. By encapsulating proteins and associated biomolecules, they can functionally reconstitute foundational features of biological cells, such as the ability to divide, communicate, and undergo shape deformation. Yet constructing liposome artificial cells from the genetic level, which is central to generate self-sustained systems remains highly challenging. Indeed, many studies have successfully established the expression of gene-coded proteins inside liposomes. Further, recent endeavors to build a direct integration of gene-expressed proteins for reconstituting molecular functions and phenotypes in liposomes have also significantly increased. Thus, this review presents the development of liposome-based artificial cells to demonstrate the process of gene-expressed proteins and their reconstitution to perform desired molecular and cell-like functions. The molecular and cellular phenotypes discussed here include the self-production of membrane phospholipids, division, shape deformation, self-DNA/RNA replication, fusion, and intercellular communication. Together, this review gives a comprehensive overview of gene-expressing liposomes that can stimulate further research of this technology and achieve artificial cells with superior properties in the future.

Comprehensive Thione-Derived Perylene Diimides and Their Bio-Conjugation for Simultaneous Imaging, Tracking, and Targeted Photodynamic Therapy.

In this study, the chromophore 3,4,9,10-perylenetetracarboxylic diimide (PDI) is anchored with phenyl substituents at the imide N site, followed by thionation, yielding a series of thione products 1S-PDI-D, 2S-cis-PDI-D, 2S-trans-PDI-D, 3S-PDI-D, and 4S-PDI-D, respectively, with n = 1, 2, 3, and 4 thione. The photophysical properties are dependent on the number of anchored thiones, where the observed prominent lower-lying absorption is assigned to the S0 → S2(ππ*) transition and is red-shifted upon increasing the number of thiones; the lowest-lying excited state is ascribed to a transition-forbidden S1(nπ*) configuration. All nS-PDIs are non-emissive in solution but reveal an excellent two-photon absorption cross-section of >800 GM. Supported by the femtosecond transient absorption study, the S1(nπ*) → T1(ππ*) intersystem crossing (ISC) rate is > 1012 s-1, resulting in ∼100% triplet population. The lowest-lying T1(ππ*) energy is calculated to be in the order of 1S-PDI-D > 2S-cis-PDI-D ∼ 2S-trans-PDI-D > 3S-PDI-D > 4S-PDI-D, where the T1 energy of 1S-PDI-D (1.10 eV) is higher than that (0.97 eV) of the 1O2 1Δg state. 1S-PDI-D is further modified by either conjugation with peptide FC131 on the two terminal sides, forming 1S-FC131, or linkage with peptide FC131 and cyanine5 dye on each terminal, yielding Cy5-1S-FC131. In vitro experiments show power of 1S-FC131 and Cy5-1S-FC131 in recognizing A549 cells out of other three lung normal cells and effective photodynamic therapy. In vivo, both molecular composites demonstrate outstanding antitumor ability in A549 xenografted tumor mice, where Cy5-1S-FC131 shows superiority of simultaneous fluorescence tracking and targeted photodynamic therapy.

Noninvasive assessment of liver function reserve with fluorescent dosimetry of indocyanine green.

Using in vivo multiphoton fluorescent dosimetry, we demonstrate that the clearance dynamics of Indocyanine Green (ICG) in the blood can quickly reveal liver function reserve. In normal rats, the ICG retention rate was below 10% at the 15-minute post-administration; While in the rat with severe hepatocellular carcinoma (HCC), the 15-minute retention rate is over 40% due to poor liver metabolism. With a 785 nm CW laser, the fluorescence dosimeter can evaluate the liver function reserve at a 1/10 clinical dosage of ICG without any blood sampling. In the future, this low-dosage ICG 15-minute retention dosimetry can be applied for the preoperative assessment of hepatectomy or timely perioperative examination.

Applications of triplex DNA nanostructures in sensor development.

Triplex DNA nanostructures are one of the most emerging and fascinating self-assembled nanostructures due to their unique nanoparticle-like organization and inherit characteristics. They have attracted numerous interests recently because of their versatile and powerful utility in diverse areas of science and technology, such as clinical or disease diagnosis and stimuli-based drug delivery. This review addresses particularly the utilization of DNA triplexes in the development of biosensors for detecting nucleic acid; strategies in sensing pH, protein activity, ions, or molecules. Finally, an outlook for potential applications of triplex DNA nanoswitches is provided.

Evaluation of the apolipoprotein E (apoE)-HDL-associated risk factors for coronary heart disease using duo-functional electrochemical aptasensor.

Apolipoprotein E containing high-density lipoprotein (apoE-HDL) and apoE-HDL cholesterol (apoE-HDL-C) are recently recognized as potential biomarkers for coronary heart disease (CHD). We herein developed a two-stage, enzyme-assisted, dual-signal aptasensor that enables a useful electrochemical sensing platform for simultaneous determination of apoE-HDL, apoE-HDL-C, and total HDL-C presented in the sample. The detection scheme consists of two subsystems. In subsystem (I), the level of apoE-HDL is evaluated upon the binding of apoE-specific aptamer and subsequently methylene blue (MB)-labeled DNA displacement occurs on the electrode surface, resulting in electrochemical reduction of methylene blue. In subsystem (II), two kinds of cholesterol levels (apoE-HDL-C and total HDL-C) can be measured. For apoE-HDL-C, the amount of cholesterol in apoE-HDL captured by the aptamer in the first step can be further determined with the aid of surfactant, cholesterol esterase, cholesterol oxidase, and p-aminophenol-mediated electrochemical signal amplification. As for total HDL-C, the amount of cholesterol is determined by the same approach as that used for apoE-HDL-C determination, but without washing (separation). The linear dynamic range for apoE-HDL determination is from 1 to 100 mg/dL (R2 = 1.00). For cholesterol standards, the linear dynamic range is determined to be 0-250 mg/dL (R2 = 0.98). Finally, serial dilutions of purified human HDL preparations were examined using the newly developed aptasensor; the percentage of apoE-HDL-C to HDL-C was found to be ~10%, which correlated well with previously reported values. In conclusion, we successfully developed an electrochemical aptasensor that allows concurrent quantification of apoE-HDL, apoE-HDL-C, and HDL-C on the same platform, offering an efficient, convenient, and purification-free sensing strategy for predictive CHD biomarkers.

Imaging of Cancer Cells and Dictated Cytotoxicity Using Aptamer-Guided Hybridization Chain Reaction (HCR)-Generated G-Quadruplex Chains.

DNA nanotechnology provides powerful tools for developing cancer theranostics. Here we introduce the autonomous surface-nucleolin-guided HCR that leads to the polymerization of G-quadruplex polymer chains, in which the ZnII -protoporphyrin IX is intercalated. We demonstrate that MDA-MB-231 (Triple Negative Breast Cancer cells, TNBC) with overexpressed surface nucleolin were able to induce HCR leading to the formation of the ZnII PPIX-loaded G-quadruplex polymer chains, while the M10 epithelial breast cells served as control. The ZnII PPIX-loaded nanowires allow the selective imaging of TNBC, and their permeation into the TNBC leads to selective cytotoxicity and guided photodynamic therapy toward the cancer cells due to structural perturbation of the membranes. The aptamer-guided HCR-generated G-quadruplex polymer chains may serve as a versatile tool to target TNBC featuring poor prognosis and high pathological risk of recurrence, thus offering a promising theranostic platform.

Amplification-free Detection of Cytomegalovirus miRNA Using a Modification-free Surface Plasmon Resonance Biosensor.

Cytomegalovirus (CMV) is the most frequent cause of congenital infection worldwide; congenital CMV may lead to significant mortality, morbidity, or long-term sequelae, such as sensorineural hearing loss. The current study presents a newly designed surface plasmon resonance (SPR) biosensor for CMV-specific microRNAs that does not involve extra care for receptor immobilization or treatment to prevent fouling on bare gold surfaces. The modification-free approach, which utilizes a poly-adenine [poly(A)]-Au interaction, exhibited a high affinity that was comparable to that of the gold-sulfur (Au-S) interaction. In addition, magnetic nanoparticles (MNPs) were used to separate the analyte from complex sample matrixes that significantly reduced nonspecific adsorption. Moreover, the MNPs also played an important role in SPR signal amplification due to the binding-induced change in the refractive index. Our SPR biosensing platform was used successfully for the multi-detection of the microRNAs, UL22A-5p, and UL112-3p, which were associated with CMV. Our SPR biosensor offered the detection limits of 108 fM and 24 fM for UL22A-5p and UL112-3p, respectively, with an R2 of 0.9661 and 0.9985, respectively. The precision of this biosensor has an acceptable CV (coefficient of variation) value of <10%. In addition, our sensor is capable of discriminating between serum samples collected from healthy and CMV-infected newborns. Taken together, we believe that our newly developed SPR biosensing platform is a promising alternative for the diagnosis of CMV-specific microRNA in clinical settings, and its application for the detection of other miRNAs may be extended further.

Targeting triple-negative breast cancer with an aptamer-functionalized nanoformulation: a synergistic treatment that combines photodynamic and bioreductive therapies.

BACKGROUND: Areas of hypoxia are often found in triple-negative breast cancer (TNBC), it is thus more difficult to treat than other types of breast cancer, and may require combination therapies. A new strategy that combined bioreductive therapy with photodynamic therapy (PDT) was developed herein to improve the efficacy of cancer treatment. Our design utilized the characteristics of protoporphyrin IX (PpIX) molecules that reacted and consumed O2 at the tumor site, which led to the production of cytotoxic reactive oxygen species (ROS). The low microenvironmental oxygen levels enabled activation of a bioreductive prodrug, tirapazamine (TPZ), to become a toxic radical. The TPZ radical not only eradicated hypoxic tumor cells, but it also promoted therapeutic efficacy of PDT. RESULTS: To achieve the co-delivery of PpIX and TPZ for advanced breast cancer therapy, thin-shell hollow mesoporous Ia3d silica nanoparticles, designated as MMT-2, was employed herein. This nanocarrier designed to target the human breast cancer cell MDA-MB-231 was functionalized with PpIX and DNA aptamer (LXL-1), and loaded with TPZ, resulting in the formation of TPZ@LXL-1-PpIX-MMT-2 nanoVector. A series of studies confirmed that our nanoVectors (TPZ@LXL-1-PpIX-MMT-2) facilitated in vitro and in vivo targeting, and significantly reduced tumor volume in a xenograft mouse model. Histological analysis also revealed that this nanoVector killed tumor cells in hypoxic regions efficiently. CONCLUSIONS: Taken together, the synergism and efficacy of this new therapeutic design was confirmed. Therefore, we concluded that this new therapeutic strategy, which exploited a complementary combination of PpIX and TPZ, functioned well in both normoxia and hypoxia, and is a promising medical procedure for effective treatment of TNBC.

Simple and Cost-effective Enzymatic Detection of Cholesterol Using Flow Injection Analysis.

A flow-injection analytical (FIA) system was developed for the determination of cholesterol concentrations based on enzymatic reactions that occurred in a cholesterol oxidase (CHOx)-immobilized, fused-silica capillary followed by electrochemical detection. The production of hydrogen peroxide from cholesterol in an enzymatic reaction catalyzed by CHOx was subsequently oxidized electrochemically at an electrode. Our FlA system demonstrated its cost-effectiveness and utility at an applied potential of 0.6 V (vs. Ag/AgCl), a flow rate of 100 µL/min and, under optimal conditions, the resulting signal demonstrated a linear dynamic range from 50 µM to 1.0 mM with a limit of detection (LOD) of 12.4 µM, limit of quantification (LOQ) of 44.9 µM, and the coefficient of variation of 5.17%. In addition, validation of our proposed system using a reference HDL-cholesterol kit used for clinical diagnosis suggested our FIA system was comparable to commercial kits for the determination of the cholesterol incorporation amount in various aqueous liposomal suspensions. These good analytical features achieved by FIA could make the implementation of this methodology possible for on-line monitoring of cholesterol in various types of samples.

Simple aminophenol-based electrochemical probes for non-enzymatic, dual amperometric detection of NADH and hydrogen peroxide.

Non enzymatic detection of NADH and H2O2 is of practical significance for both environmental and biological prospective. However, there is no simple, straight forward electrochemical sensor available for sensing of them in real samples. Addressing this challenge, we report a simple stimuli responsive aminophenol, pre-anodized screen printed carbon electrode (SPCE*/AP) based electrochemical probes for dual detection of NADH and H2O2. Aminophenol prepared and adsorbed on the electrode from aminophenylboronic acid via boronic acid deprotection with H2O2. The SPCE*/AP fabricated with this process was characterized by cyclic voltammetry (CV), scanning electron microscope (SEM), Raman spectroscopy, UV-visible spectroscopy, and X-ray photoelectron spectroscopy (XPS). Amperometric detection results showed that SPCE*/AP electrodes exhibited linearity from 50 µM to 500 µM and from 200 µM to 2 mM with a detection limit (S/N = 3) of 4.2 µM and 28.9 µM for NADH and H2O2, respectively. Excellent reproducibility and selectivity for NADH and H2O2 were observed for this electrochemical platform. In addition, the matrix effect was investigated further using the same technique to analyze NADH and H2O2 in human urine samples, human serum samples, cell culture medium (containing 10% fetal bovine serum, FBS), and environmental water samples (tap water and rain water). Also, the present sensor demonstrated promising outcomes with living cells (normal cells and cancer cells).

Diagnosing the RGS11 Lung Cancer Biomarker: The Integration of Competitive Immunoassay and Isothermal Nucleic Acid Exponential Amplification Reaction.

Lung cancer is the primary cause of cancer-associated mortality worldwide, which makes the identification of reliable lung cancer biomarkers a pressing need for early diagnosis and prognosis. RGS11, which is a regulator of G-protein signaling and also a lung cancer biomarker, plays an important role in cancer-related metastasis. However, trace levels of RGS11 (in the range of pg/mL) in serum samples make it difficult to quantify using currently available enzyme-linked immunosorbent assay (ELISA) kits and, therefore, this hinders progress in the discovery of new approaches for treating lung cancer. The aim of this study is to develop a rapid, sensitive, and reliable platform for the detection of RGS11 lung cancer biomarker based on a suspension immunoassay coupled with an isothermal exponential amplification strategy. Our study was initiated by the functionalization of magnetic beads with anti-RGS11 antibodies (Ab-MB) by EDC (1-ethyl-3-(3-(dimethylamino)propyl)-carbodiimide)/NHS ( N-hydroxysulfosuccinimide) activation. Ab-MB served as a sensing probe for the competitive immunorecognitions between known concentrations of His-tag RGS11 and unknown concentrations of target RGS11 in serum. The reporter anti-His antibodies, which were modified with primers that induced an isothermal exponential amplification reaction, were subsequently introduced to the reaction mixture that resulted in the formation of immunosandwich complexes. The exponentially amplified DNA duplex that was intercalated with SYBR Green was designated as a signal reporter for the assessment of RGS11 in an inversely proportional relationship. The sensing platform was excellent for the determination of RGS11 with an exceptional detection limit of 148 fg/mL and a linear dynamic range of 0.1-10 pg/mL using a minimal sample volume (20 µL) and with a reaction time of 1.5 h. In addition, we challenged the sensing platform with RGS11-spiked samples (in 2× diluted serum), and an acceptable recovery rate (>90%) was observed. Finally, 24 clinical samples acquired from patients with advanced lung cancer (C), inflammation (I), and heart failure (H) were analyzed by this newly developed sensing platform and a commercial ELISA kit for validation. This sensing platform has potential in biomedical applications for clinically diagnosing liquid biopsy samples for patients with lung cancer. Moreover, the universal design of our proposed system is easily adapted to detect any other protein if a His-tag recombinant protein is available.

Chemopreventive Potential of Ethanolic Extracts of Luobuma Leaves (Apocynum venetum L.) in Androgen Insensitive Prostate Cancer.

Luobuma (Apocynum venetum L. (AVL)) is a popular beverage in Asia and has been reportedly to be associated with the bioactivities such as cardiotonic, diuretic, antioxidative, and antihypertensive. However, its biofunction as chemoprevention activity is seldom addressed. Herein, we aimed to characterize the anti-androgen-insensitive-prostate-cancer (anti-AIPC) bioactive compounds of Luobuma, and to investigate the associated molecular mechanisms. Activity-guided-fractionation (antioxidative activity and cell survivability) of Luobuma ethanolic extracts was performed to isolate and characterize the major bioactive compounds using Ultra Performance Liquid Chromatography (UPLC), Liquid Chromatography-Mass Spectrometry (LC-MS), and Nuclear Magnetic Resonance (NMR). Plant sterols (lupeol, stigamasterol and ß-sitosterol) and polyphenolics (isorhamnetin, kaempferol, and quercetin) were identified. Lupeol, a triterpene found in the fraction (F8) eluted by 10% ethyl acetate/90% hexane and accounted for 19.3% (w/w) of F8, inhibited the proliferation of PC3 cells. Both lupeol and F8 induced G2/M arrest, inhibition of ß-catenin signaling, regulation of apoptotic signal molecules (cytochrome c, Bcl-2, P53, and caspase 3 and 8), and suppression DNA repair enzyme expression (Uracil-DNA glycosylase (UNG)). To our knowledge, our study is the first report that lupeol inhibited the expression of UNG to elicit the cytotoxicity against androgen-insensitive-prostate-cancer cells. Collectively, Luobuma, which contains several antitumor bioactive compounds, holds the potential to be a dietary chemopreventive agent for prostate cancer.

Tyramine detection using PEDOT:PSS/AuNPs/1-methyl-4-mercaptopyridine modified screen-printed carbon electrode with molecularly imprinted polymer solid phase extraction.

Tyramine (4-hydroxyphenethylamine), which is a monoamine metabolized by monoamine oxidase (MAO), exists widely in plants, animals, fermented foods, and salted foods. The incidence of hypertension, or "cheese effect", which is associated with a large dietary intake of tyramine while taking MAO inhibitors has been reported; therefore, the measurement of tyramine is an urgent concern. Herein, an efficient approach that integrates a molecular imprinting polymer for solid phase extraction (MISPE) technique with a sensitive electrochemical sensing platform (SPCE/PEDOT: PSS/AuNP/1-m-4-MP) for the quantification of tyramine is presented. Enhanced electrode conductivity was achieved sequentially by constructing a conductive polymer (PEDOT: PSS) on a screen-printed carbon electrode (SPCE), followed by electrodeposition with gold nanoparticles (AuNPs) and, finally, by modification with positively charged 1-methyl-4-mercaptopyridine (1-m-4-MP) using an Au-S bond. Tyramine was isolated selectively and pre-concentrated by the MISPE technique; electroanalysis that used differential pulse voltammetry (DPV) in NaOH (0.1M, pH 13) was conducted successively. Experimental parameters (such as modes of electrode modification, ratio of PEDOT: PSS, pH of electrolyte, time required for AuNP deposition, and 1-m-4-MP concentrations) that were associated with optimal detection conditions were evaluated also. We obtained a linear concentration range (5-100nM, R2=0.9939) with LOD and sensitivity at 2.31nM, and 3.11µAnM-1cm-2, respectively. The applicability of our technique was demonstrated by analyzing tyramine in spiked serum and milk. The feature of our newly developed analytical methods that coupled sample pre-treatment (sample clean-up and pre-concentration) with sensitive detection makes it a promising tool for quantifying of tyramine.

Green Synthesis of Novel Polyaniline Nanofibers: Application in pH Sensing.

An optically active polyaniline nanomaterial (PANI-Nap), doped with (S)-naproxen, was developed and evaluated as a potent pH sensor. We synthesized the material in one pot by the addition of the dopant, (S)-naproxen, prior to polymerization, followed by the addition of the oxidizing agent (ammonium persulfate) that causes polymerization of the aniline. This green chemistry approach allowed us to take only 1 h to produce a water-soluble and stable nanomaterial. UV-visible spectroscopy, fluorescence spectroscopy, FT-IR spectroscopy, Raman spectroscopy, scanning electron microscopy (SEM), transmission electron microscopy (TEM), and X-ray photoelectron spectroscopy (XPS) were used to characterize the designed nanomaterial. This nanomaterial exhibited excellent pH sensing properties and showed long term stability (up to one month) without loss of sensor performance.

Gold Nanocluster-Assisted Fluorescent Detection for Hydrogen Peroxide and Cholesterol Based on the Inner Filter Effect of Gold Nanoparticles.

We developed a simple, sensitive inner filter effect (IFE)-based fluorescent assay for sensing H2O2 and cholesterol. In the process, poly(vinylpyrrolidone)-protected gold nanoparticles (PVP-AuNPs) and fluorescent BSA-protected gold nanoclusters (BSA-AuNCs) were used as an IFE absorber/fluorophore pair. PVP-AuNPs can be a powerful absorber to influence the emission of the fluorophore, BSA-AuNCs, in the IFE-based fluorescent assays. That is due to the high extinction coefficient of AuNPs and the complementary overlap between the surface plasmon resonance (SPR) absorption of PVP-AuNPs and the excitation of BSA-AuNCs. The PVP-Au seeds, produced by directly mixing PVP with HAuCl4, were able to catalyze H2O2 to enlarge AuNPs. The SPR absorption of PVP-AuNPs was enhanced with an increased concentration of H2O2 and, subsequently, induced significant fluorescence quenching of BSA-AuNCs. The IFE-based fluorescent assay enabled the detection of H2O2 and generation of H2O2 in the presence of O2/cholesterol and cholesterol oxidase (ChOx) by the fluorescence response of BSA-AuNCs. The present IFE-based approach can detect H2O2 ranging from 1 to 100 µM with a detection limit of 0.8 µM and cholesterol ranging from 1 to 100 µM with a detection limit of 1.4 µM.

Resuscitation Using Liposomal Vasopressin in an Animal Model of Uncontrolled Hemorrhagic Shock.

BACKGROUND: Current research suggests that administration of vasopressin to patients with uncontrolled hemorrhagic shock (UHS) can avoid the detrimental effects associated with aggressive fluid resuscitation. However, vasopressin has a short half-life of 10~35 minutes in in vivo use and precludes its use in the pre-hospital setting. To increase the half-life of vasopressin, we proposed to synthesize liposome-encapsulated vasopressin and test it in a rat model of UHS. METHODS: The film hydration method was used to prepare liposomal vasopressin consisting of: Dipalmitoylphosphatidylcholine, cholesterol, and dipalmitoyl phosphatidylethanolamine (20:20:1 mole ratio). 42 rats were subjected to UHS and randomly received 5 different treatments (vasopressin, liposomal vasopressin, lactate ringer (LR), liposome only and sham). Outcome of UHS were measured using 4 common prognostic tests: mean arterial pressure (MAP), serum lactate level, inflammatory profile and pulmonary edema. RESULTS: The dynamic light scattering results confirmed that we had prepared a successful liposomal vasopressin complex. Comparing the serum vasopressin concentration of liposomal vasopressin and vasopressin treated animals by ELISA, we found that the concentration of vasopressin for the liposomal vasopressin treated group is higher at 60 minutes. However, there was no significant difference between the MAP profile of rats treated with vasopressin and liposomal vasopressin in UHS. We also observed that animals treated with liposomal vasopressin performed indifferently to vasopressin treated rats in serum lactate level, inflammatory profile and edema profile. For most of our assays, the liposome only control behaves similarly to LR resuscitation in UHS rats. CONCLUSION: We have synthesized a liposomal vasopressin complex that can prolong the serum concentration of vasopressin in a rat model of UHS. Although UHS rats treated with either liposomal vasopressin or vasopressin showed no statistical differences, it would be worthwhile to repeat the experiments with different liposomal compositions.

Gallic acid induces necroptosis via TNF-α signaling pathway in activated hepatic stellate cells.

Gallic acid (3, 4, 5-trihydroxybenzoic acid, GA), a natural phenolic acid widely found in gallnuts, tea leaves and various fruits, possesses several bioactivities against inflammation, oxidation, and carcinogenicity. The beneficial effect of GA on the reduction of animal hepatofibrosis has been indicated due to its antioxidative property. However, the cytotoxicity of GA autoxidation causing cell death has also been reported. Herein, we postulated that GA might target activated hepatic stellate cells (aHSCs), the cell type responsible for hepatofibrosis, to mitigate the process of fibrosis. The molecular cytotoxic mechanisms that GA exerted on aHSCs were then analyzed. The results indicated that GA elicited aHSC programmed cell death through TNF-α-mediated necroptosis. GA induced significant oxidative stress through the suppression of catalase activity and the depletion of glutathione (GSH). Elevated oxidative stress triggered the production of TNF-α facilitating the undergoing of necroptosis through the up-regulation of key necroptotic regulatory proteins TRADD and receptor-interacting protein 3 (RIP3), and the inactivation of caspase-8. Calmodulin and calpain-1 activation were engaged, which promoted subsequent lysosomal membrane permeabilization (LMP). The TNF-α antagonist (SPD-304) and the RIP1 inhibitor (necrostatin-1, Nec-1) confirmed GA-induced TNFR1-mediated necroptosis. The inhibition of RIP1 by Nec-1 diverted the cell death from necroptosis to apoptosis, as the activation of caspase 3 and the increase of cytochrome c. Collectively, this is the first report indicating that GA induces TNF signaling-triggered necroptosis in aHSCs, which may offer an alternative strategy for the amelioration of liver fibrosis.