Proteomics analysis revealed the presence of inflammatory and oxidative stress markers in the plasma of migraine patients during the pain period.

BACKGROUND: There is increasing evidence that some biomarkers are implicated in migraine pathogenesis. This study looks at plasma proteome in migraine patients for potential protein biomarkers. METHODS: This case-control study has two phases. In phase I, plasma samples were collected from three groups, including twenty-three episodic migraineurs, thirty-five chronic migraineurs, and twenty-nine healthy subjects. In phase II, plasma samples were prepared from two groups, including five episodic and five chronic migraine cases, during the pain and 24 h after the pain-free periods. Two-dimensional gel electrophoresis (2-DE) was performed on plasma proteins. The possible corresponding proteins for the differentially expressed spots between groups investigated by the Melanie software were predicted by 2-DE gels of the EXPASY database. LC-MS/MS additionally analyzed phase II data. RESULTS: Expression levels of haptoglobin, clusterin, fibrinogen alpha chain, fibrinogen beta chain, complement c3, transthyretin, α1-microglobulin, and retinol-binding protein 4 were shown considerable changes in migraine patients compared to controls or their pain-free period. CONCLUSION: Differences in expression levels for several proteins were observed across groups. Most of these are associated with inflammation, oxidative stress, and neuroprotection, which can be considered potential disease biomarkers. However, further research is necessary for this respect.

Crocetin ameliorates non-alcoholic fatty liver disease by modulating mitochondrial dysfunction in L02 cells and zebrafish model.

ETHNOPHARMACOLOGICAL RELEVANCE: Traditional Chinese medicine considers that the etiology and pathogenesis of non-alcoholic fatty liver disease (NAFLD) are related to liver depression and qi stagnation. Saffron and its active ingredient, crocetin (CCT), are used for the treatment of metabolic diseases owing to their "Liver deobstruent" and "Liver tonic" effects. However, the effect of CCT on NAFLD has not been fully elucidated. In the present study, the effect and potential molecular mechanism of CCT were explored in both in vivo and in vitro models of NAFLD. MATERIALS AND METHODS: CCT was isolated from saffron and purity and structure characterization were performed using HPLC, MS, 1H-NMR, and 13C-NMR. The effect of CCT on the viability of L02 cells and its maximum tolerable concentration (MTC) in zebrafish were investigated. Free fatty acids (FFA) and thioacetamide (TAA) were used to induce lipid accumulation in L02 cells and steatosis in zebrafish, respectively. The effects of CCT on indexes related to lipid metabolism, oxidative stress, and mitochondrial function in NAFLD models were explored using biochemical assay kits, Western blot analysis, Reverse Transcription-Polymerase Chain Reaction (RT-PCR), histopathology analysis, and determination of mitochondrial membrane potential (ΔΨm). Morphological analysis of mitochondria was performed using transmission electron microscopy (TEM). RESULTS: The levels of triglyceride (TG), total cholesterol (TC), malondialdehyde (MDA), and alanine/aspartate aminotransferases (ALT/AST) activities in FFA treated L02 cells were significantly reduced after CCT treatment. CCT treatment significantly increased ATP concentration, ΔΨm, and activities of superoxide dismutase (SOD), catalase (CAT), and cytochrome c oxidase (COX IV) in FFA treated L02 cells. TEM images showed restoration of mitochondrial morphology. CCT decreased ATP concentration and upregulated expression of B-cell lymphoma-2 (Bcl-2) and COX IV, whereas, CCT downregulated expression of BCL2-Associated X (Bax) and cleaved caspase-3 in TAA treated zebrafish. These findings indicated that mitochondrial dysfunction was alleviated after CCT treatment. Oil Red O staining of L02 cells and zebrafish showed that CCT treatment reversed the accumulation of lipid droplets. CONCLUSION: In summary, CCT treatment effectively alleviated the symptoms of NAFLD and restored mitochondrial function in L02 cells and zebrafish NAFLD model.

Preparation of a new nanobiosensor for the determination of some biogenic polyamines and investigation of their interaction with DNA.

Biogenic polyamines are small organic polycations involving in a variety of biological processes. They form high affinity complexes with DNA. Here, we have followed two different novel approaches, either fabrication of an electrochemical nanobiosensor for determination of three of the most important biogenic polyamines; spermine (SPM), spermidine (SPD) and putrescine (PUT), or electrochemical investigation of their interaction with DNA. Strong binding of polyamines to DNA makes the DNA a suitable recognition element for construction of a sensitive biosensor. The fabricated biosensor responded to SPM, SPD and PUT over an extended dynamic range of 0.04-100 µM, 0.01-24 µM, and 0.08-100 µM respectively, with low detection limits of a few nM. We also studied the interaction of polyamines with three different DNA sequences with base composition of 100% AT, 80% AT and 100% GC in the presence of [Ru(NH3)6]3(+) as a redox probe. The highest kb values were obtained in the interaction of polyamines with 80% AT (mixed) DNA sequence. The kb values were 5.24 × 10(5), 4.17 × 10(5) and 1.46 × 10(5)M(-1) for SPM, SPD and PUT, respectively, which correlated well with their increasing number of amino groups. In addition, competition study showed the impotence of SPD to replace with histone H1 in histone H1-DNA complex, which indicates the more potent interaction of histone H1 with DNA. In this proof-of-principle study, we have proposed an approach for simple, cost-effective, miniaturizable, and direct-readout detection of polyamines, as well as the understanding of the modes of interaction between polyamines and DNA.

Electrochemical aptamer/antibody based sandwich immunosensor for the detection of EGFR, a cancer biomarker, using gold nanoparticles as a signaling probe.

Detection of epidermal growth factor receptor (EGFR) in biological fluids is of paramount importance, since it has significant application in cancer diagnosis, drug development, and therapy monitoring. EGFR is a cancer biomarker, and its overexpression is associated with the development of some types of cancer. Herein, we report on the development of a sensitive and selective electrochemical aptamer/antibody (Apt/Ab) sandwich immunosensor for detection of EGFR. In this study, a biotinylated anti-human EGFR Apt was immobilized on streptavidin-coated magnetic beads (MB) and served as a capture probe. A polyclonal anti-human EGFR Ab was conjugated to citrate-coated gold nanoparticles (AuNPs) and used as a signaling probe. In the presence of EGFR, an Apt-EGFR-Ab sandwich was formed on the MB surface. The extent of the complexation was evaluated by differential pulse voltammetry of AuNPs after their dissolution in HCl. Under optimal conditions, the dynamic concentration range of the immunosensor for EGFR spanned from 1 to 40 ng/mL, with a low detection limit of 50 pg/mL, and RSD percent of less than 4.2%. The proposed approach takes advantage of sandwich assay for high specificity, MBs for fast separation, and electrochemical method for cost-effective and sensitive detection. In this proof-of-principle study, we demonstrate the potential clinical efficacy of the immunosensor for monitoring of chemotherapy effectiveness in breast cancer samples.

Dual-frequency ultrasound activation of nanomicellar doxorubicin in targeted tumor chemotherapy.

INTRODUCTION: This study investigated the therapeutic effect of dual-frequency sonication (3 MHz and 28 kHz) at low intensity levels in combination with micellar doxorubicin in the treatment of a tumor model of spontaneous breast adenocarcinoma in Balb/c mice. METHODS: We used sonication frequencies 28 kHz and 3 MHz and their dual combinations in the progressive wave mode to enhance acoustic cavitation. Then, the antitumor effect of the simultaneous dual-frequency ultrasound (28 kHz and 3 MHz) at low intensity levels in combination with doxorubicin and micellar doxorubicin injection was investigated in a spontaneous model of breast adenocarcinoma in Balb/c mice. Sixty-three tumor-bearing mice were randomly divided into seven groups: control, sham, sonication with dual frequency, doxorubicin without sonication, doxorubicin with dual-frequency sonication, micellar doxorubicin without sonication, and micellar doxorubicin with dual-frequency sonication. The tumor volume change relative to the initial volume, tumor growth inhibition ratio, the required times for each tumor to reach two (T 2) and five (T 5) times its initial volume, and survival period were the tumor growth delay parameters which were calculated and recorded at various times after treatment. RESULTS: The results of the combination of frequencies 28 kHz (0.04 W/cm(2)) and 3 MHz (2.00 W/cm(2)) showed remarkable enhancement of the cavitation activity compared with single-frequency sonication (P < 0.05). The micellar doxorubicin injection with sonication group showed a significant difference in the relative volume percent parameter compared with the other groups (P < 0.05). Additionally, the T 2 and T 5 times in the micellar doxorubicin with sonication group were significantly higher than in the other groups (P < 0.05). Also, the survival period of the mice in the micellar doxorubicin with sonication group was significantly longer than in the other groups (P < 0.05). These findings were verified histopathologically. CONCLUSION: This study shows that simultaneous combined dual-frequency ultrasound sonication in continuous mode is effective in producing cavitation activity at low intensity. We conclude that dual-frequency sonication with micellar doxorubicin injection extends survival in a murine breast adenocarcinoma model.

Effect of crocin on the insulin resistance and lipid profile of streptozotocin-induced diabetic rats.

Crocin is the only water soluble carotenoid in nature, and it has a known powerful antioxidant activity. The aim of this work was to investigate the hypoglycemic and hypolipidemic effects of crocin in streptozotocin (STZ)-induced type 2 diabetic rats. Neonatal male Wistar rats (2-5 days old) were randomly divided into five groups. Three groups were intraperitoneally injected with STZ (90 mg/kg body weight). Among them, two groups were treated with intraperitoneal injection of crocin (50 or 100 mg/kg), and the third group was treated with vehicle only. Two control groups were also considered, and one of them was treated with crocin. After 5 months, their blood and urine samples were collected, and the animals were sacrified. The results indicate a significant lower body weight (P < 0.001) and abnormal parameters in the diabetic rats compared with the normal group. An administration of both doses of crocin significantly decreased the levels of serum glucose, advanced glycation end products, triglyceride, total cholesterol, and low-density lipoprotein and increased the high-density lipoprotein in the diabetic rats. The treatments were also effective in decreasing HbA1c and microalbuminuria, as well as homeostatic model assessment for insulin resistance as a measure of insulin resistance in the diabetic rats.

Electrochemical studies of DNA immobilization onto the azide-terminated monolayers and its interaction with taxol.

A surface modification procedure for the creation of self-assembled monolayers (SAMs) that can be used as a scaffold for double-stranded DNA (dsDNA) incorporation onto the gold surfaces is described. The SAMs of an azidohexane thiol derivative were prepared on the Au electrode and then used for the immobilization of dsDNA. The electrochemical characteristics of dsDNA onto the SAM-modified gold electrode were investigated by cyclic voltammetry and electrochemical impedance spectroscopy, and the surface concentration of dsDNA onto the SAMs surface was estimated. The interaction of dsDNA with the anticancer drug, taxol (paclitaxel), was also studied on the surface of DNA/SAM/Au electrode. The observed decrease in the guanine oxidation peak current was used to monitor the interaction of taxol with DNA. The resulting Langmuir isotherm for taxol binding to DNA at the modified electrode was used to evaluate the binding constant of taxol-DNA. The results obtained supported the groove binding interaction of taxol with DNA. The modified electrode was used as a sensitive sensor for quantification of taxol in human serum sample.

DNA as an enzyme. The effect of imidazole derivatives as cofactors and metal ions as activators or inhibitors.

OBJECTIVE: A highly sensitive spectrofluorometric method using dichlorofluorescin (LDCF) was employed to study the rate of electron transfer reaction in presence of DNA and some imidazole derivatives. RESULTS: In our experiments, DNA possessed a unique enzyme like catalytic function in oxidative conversion of nonfluorescent LDCF to fluorescent dichlorofluorescein (DCF). The rate enhancement was associated with the turn over constant: k(p) = 10 s(-1) for DNA and cinnamoyl imidazole as a cofactor. A biphasic saturation curve was observed when the reaction velocities were measured at fixed concentrations of DNA and variable amounts of carnosine. Each of the biphasic trends gave the Scatchard values of V(m1)/K(m1) = 3.1 x 10(-5) and V(m2)/K(m2) = 5.1 x 10(-6) with K(m1) = 2.7 x 10(-5) M and K(m2) = 4.2 x 10(-4) M for carnosine. Although Ni (II) and Pb (II) induced inhibition in the rate of electron transfer reaction in presence of DNA and cinnamoyl imidazole or carnosine, metal ions such as Mg (II), Cd (II), Zn (II) and Fe (II) caused activation of DNA. The rates of the reactions showed strong dependency on electronegativity and conductivity of metal ions, namely the increase in activity of DNA in presence of each metal ion correlated inversely with the electronegativities of the metal and was also related directly to the conductivities of individual metal. These effects were observed both in activation and also inhibition of DNA reaction. Imidazole compounds, e.g., Histidine, N-trans cinnamoyl imidazole and imidazole along with Cd (II) produced further rate enhancement. The increase was several times greater with N-trans cinnamoyl imidazole. CONCLUSIONS: This effect could provide additional evidence for the importance of an intermediary cofactor that could facilitate the transfer of the electron from the reaction site to the DNA conductive chord. This was most guaranteed by the conjugated system provided by a compound such as N-trans cinnamoyl imidazole.