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The current mixed method study investigated the use of synchronous/asynchronous computer-mediated communication as a method of L2 pronunciation achievement among medical students during COVID-19 pandemic lockdown. In so doing, the study utilized an experimental comparative design with two groups of 17 participants. There were two types of tests in this study: word stress test, and a semi-structured interview. The results showed the positive effect of computer mediated communication in pronunciation development among students of medicine. Moreover, the findings revealed more positive effect of synchronous computer mediated communication-oriented instruction compared with asynchronous computer mediated communication-oriented instruction on medical students' English language pronunciation development. It can be concluded that computer mediated communication especially synchronous mode is effective for pronunciation development of medical students since technology provides the opportunities through which medical students can notice the gaps in their current English language and thus produce modified output.

Carbon dioxide (CO2) capture and storage (CCS) processes could benefit from thermostable carbonic anhydrases (CAs), which can significantly increase the rate of CO2 absorption into solvents. Since conventional CO2 absorption techniques are usually performed under high-temperature conditions in industrial plants, implementing highly thermostable CAs is very important to introduce a robust and economical process. The present study employed a combination of in silico tools to rationally engineer a thermostable CA (SspCA) originating from a thermostable bacterium Sulfurihydrogenibium yellowstonense. Based on the results, while the general folding of the enzyme and its catalytic efficiency was retained in the K100G mutant, the melting temperature increased by 3°C, and the CO2 hydration activity half-life at 85°C increased by two times. Molecular dynamics (MD) simulations were performed to determine the underlying causes of the thermostability improvement. The K100G mutation was shown to cause a reduction in protein local flexibility mainly by confining the flexible parts due to rearrangement of salt bridges and hydrogen interactions network. In sum, while strengthening the importance of rational design in thermostable enzymes, the present study showed the capability of merely one rationally-designed mutation which indirectly resulted in CA thermostability improvement by exerting local structural rigidification in its neighboring part.

Amygdalin induces apoptotic death in several carcinoma cells. Affibody is an engineered protein with a high affinity for human epidermal receptor 2 (HER2). We assessed the cytotoxic effects of the amygdalin-ZHER2 affibody conjugate on two breast carcinoma cell lines. The ZHER2 affibody gene was synthesized and transferred into E. coli BL21 as an expression host. After purification, the ZHER2 affibody was conjugated to amygdalin. The cytotoxic effects of amygdalin and its ZHER2 affibody conjugate on the SK-BR-3, with overexpression of HER2, and MCF-7 cells were evaluated by MTT assay. The effects of amygdalin and its conjugate on apoptotic death and expression of pro-apoptotic Bax and anti-apoptotic Bcl-2 proteins were measured. Amygdalin individually showed a potent cytotoxic effect against both MCF-7 (IC50 = 14.2 mg ml−1) and SK-BR-3 cells (IC50 = 13.7 mg ml−1). However, the amygdalin-ZHER2 affibody conjugate had a more cytotoxic effect on SK-BR-3 (IC50 = 8.27 mg ml−1) than MCF-7 cells (IC50 = 19.8 mg ml−1). Amygdalin had a significant apoptotic effect on both cell lines and the effect of its conjugate on SK-BR-3 cells was significantly more potent than MCF-7 cells. Amygdalin increased Bax and decreased Bcl-2 expression in both cell lines. However, the effect of its conjugate on the Bax and Bcl-2 expression in SK-BR-3 was more potent than MCF-7 cells. In conclusion, the amygdalin-ZHER2 affibody conjugate may be considered as a valuable candidate for specific treatment of breast cancer patients with overexpression of HER2. However, further in vivo studies are required to explain the antitumoral effects of constructed amygdalin-ZHER2 affibody conjugate.

In spite of several studies that have shown the cytotoxic effects of amygdalin on the different cancer cell lines, however, the chemopreventive potential of amygdalin on the breast cancer cell line is not completely understood. We investigated the effect of amygdalin on the cell death and the level of pro-apoptotic Bax protein and anti-apoptotic Bcl-2 protein in SK-BR-3 human breast cancer cell line. The cell viability of SK-BR-3 cells was evaluated by MTT assay in different concentration of amygdalin. The level of Bax and Bcl-2 in SK-BR-3 cells were measured by western blot analysis. For statistical analysis, One-way ANOVA was used for the comparison of Bax and Bcl-2 protein level and percent of cell viability between groups. The molecular docking studies of amygdalin within the Bcl-2 (PDB ID: 4LVT) and HER2 (PDB ID: 3RCD) active site, were performed using AutoDock 4.2.5. Amygdalin induced a significant reduction of cell viability in SK-BR-3 after 24-h treatment in a dose-dependent manner. Also, amygdalin causes an increase in pro-apoptotic Bax protein and a decrease in anti-apoptotic Bcl-2 protein expression in the SK-BR-3 cells. Molecular docking studies showed that amygdalin interacts with the active site amino acids of Bcl-2 and HER2 through hydrogen bonding and some hydrophobic interactions. Amygdalin can induce apoptotic death in SK-BR-3 cells by increasing pro-apoptotic Bax protein and decreasing anti-apoptotic Bcl-2 protein expression. The results suggest that amygdalin may be a valuable candidate for the treatment of breast cancer, especially in HER2 positive cells.

Overexpression of human epidermal growth factor receptor 2 (HER2) can be lead to cell transformation and consequently progression of various kinds of malignancies. Therefore, HER2 receptor is not just a specific target for immunotherapy using monoclonal antibodies such as trastuzumab (Herceptin®) but also can be considered as a targeting point for specific delivery of chemotherapeutic cargos to cancerous cells. Affibody ZHER2 with a great affinity for HER2 receptors can be used for specific targeting of HER2-overexpressing cells. In the current study, Purified ZHER2:342 affibody molecules were conjugated to gold nanoparticles (as X-ray radiosensitizer). Characteristics of the prepared GNPs and ZHER2 affibody-conjugated gold nanoparticles (GNP-ZHER2) were investigated by atomic force microscopy (AFM) and UV–Vis spectrophotometry. Efficacy of GNPs and GNP- ZHER2 conjugates in enhancement of X-ray radiotherapy of four malignant cell lines including MCF-7 (a cell line with a normal expression of HER2), SK-BR-3, HN-5, and SK-OV-3 (three different HER2-overexpressing cell lines) was investigated by measuring cell viability using MTT assay. Our in vitro results showed that although sensitivity of different examined cell lines to GNP, GNP-ZHER2 conjugate and X-ray radiation is different but in overall GNP-ZHER2 conjugate could enhance efficacy of X-ray radiation in ablation of HER2-overexpressed cancerous cell lines.

Cardiovascular diseases (CVDs) are the leading cause of death worldwide. The majority of cardiovascular complications are secondary to atherosclerosis. Extensive evidence has showed that environmental pollutants such as cigarette smoke and automobile exhaust increase the risk of developing atherosclerosis. Acrolein, a highly reactive unsaturated aldehyde, is found as a contaminant in air, food and water. Investigations during the last decades have shown that acrolein via various mechanisms such as oxidative stress, enhancement of inflammatory processes and the activation of matrix metalloproteases can initiate and accelerate atherosclerotic lesions formation. Furthermore, exposure to acrolein has been suggested to induce or exacerbate systemic dyslipidemia, an important risk factor for the development of atherosclerosis. Finally, there are reports which indicate acrolein can increase platelet activation and stimulation of the coagulation cascade which subsequently leads to thrombosis. Even a modest reduction of pollutants such as acrolein can have substantial effects on population health. Public health efforts to reduce acrolein exposures from known sources may lower the prevalence of vascular disease. This review focuses on the potential pathways and mechanisms behind the acrolein-induced atherothrombotic effects.

Due to the high affinity for binding to target molecules and also other unique attributes, affibodies have a great potential to be used in immunotherapeutic and diagnostic approaches. However, the possibility of undesirable immune response is still a great concern. In the current study, we investigated the possible antigenicity, allergenicity and cytotoxicity of the HER2-targeting affibody Z

Hyaluronidase (HAase) from bovine testes (BTH) has long been used in broad pharmaceutical areas, while it is associated with drawbacks in aspects of solubility, immunogenicity and pharmacokinetics. These issues can be addressed by gaining structural insights and designing rational modifications to the enzyme structure, as proposed in this study. A 3D structural model was built for HAase and underwent 40 ns of molecular dynamic simulation to examine its thermostability under normal, melting, and extreme conditions. The enzyme activity of BTH was measured against temperature and pH by kinetic assays. The interaction of bovine HAase with HA and chondroitin was defined by molecular docking. Furthermore, immunogenic properties of the enzyme were explored by immunoinformatics. Thermal effects on bovine HAase structural model and the HAase interactions with its substrates were described. We identified some B- and T-cell epitopes and showed that the protein could be recognized by human immune receptor molecules. Epitope masking by adding polyethylene glycol (PEG) to amine groups of residues presenting on the surface of the protein structure was adopted as a surface modification to enhance pharmacological properties of BTH. Assays showed that PEGylated BTH had higher thermostability and similar activity compared to the native enzyme.

Amygdalin induces apoptotic death in several carcinoma cells. Affibody is an engineered protein with a high affinity for human epidermal receptor 2 (HER2). We assessed the cytotoxic effects of the amygdalin-Z

Human epidermal growth factor receptor (HER) family is a four-member group of transmembrane receptors. Among them, HER2 over-expression is associated with various kinds of malignancies. Chemotherapy and immunotherapy are two main treatment approaches in the cancer therapy. In the current study our goal was to target over-expressing HER2 breast cancerous cell line (SK-BR-3) by idarubicin-loaded immunoliposome. Idarubicin-loaded vesicles were prepared and characterized using analytical methods. Levels of HER2 expression in breast cancerous cell lines (SK-BR-3 and MCF-7) was monitored by western blotting. Cell lines were treated by various concentrations of idarubicin, idarubicin-loaded liposome, and idarubicin-loaded trastuzumab-conjugated immunoliposome. Cell viability was evaluated by MTT assay. Idarubicin in the free form decreased proliferation of both SK-BR-3 and MCF-7 cells in a dose dependent manner although SK-BR-3 was more susceptible to idarubicin. Idarubicin-loaded liposome and immunoliposome in a similar way decreased viability of both cell lines. Loading of idarubicin in trastuzumab-conjugated immunoliposome improved its efficiency in decreasing viability of SK-BR-3 cells. In conclusion, it seems that immunoliposomal form of idarubicin is more potent in ablation of HER2-overexpressing cancerous cells and it has a great potential to be considered as a new candidate for treatment of HER2-positive malignancies.

Performances of five phenol-powered stacked microbial desalination cells (sMDC) with internally parallel and series continuous flow configurations were assessed. The effects of inter-membrane distance (5 and 10 mm) and flow configuration on water desalination were investigated in order to improve the flexibility and footprint of the system. Different sMDC configurations were examined with influent NaCl concentrations of 1–3% at flow rates of 3.5, 7, and 15 μL min−1. The results of the series flow configurations showed that at NaCl concentration of 1%, reducing the inter-membrane distance (from 10 to 5 mm) of the desalination chamber next to the anode compartment resulted in improved desalination efficiency and specific desalination rate (SDR) while at 3% NaCl concentration, the highest desalination efficiency was achieved with 10 mm inter-membrane distance of both desalination chambers. The results of the parallel flow configurations showed that decreasing the inter-membrane distance leads to distinct effects on desalination efficiency based on the initial NaCl concentration. At NaCl concentration of 1% in contrast to the higher concentrations of 2–3%, an increase in desalination efficiency (36%) was achieved by decreasing the inter-membrane distance. Overall, in parallel flow configurations, the SDR trend was incremental (16–277%). It can be concluded that the different behavior of the sMDC at high and low NaCl concentrations should be taken into consideration in order to optimize the design and operation of these desalination systems.

Chrysin (CH) is a natural flavone which possesses antioxidant, anti-cancer, and anti-inflammatory properties. The aim of the present study was to investigate the effects of CH on biochemical parameters, histopathological changes, and genotoxicity and hematological indices in diazinon (DZN)-induced toxicity in BALB/c mice. We induced sub-acute toxicity in mice using DZN (20 mg/kg/day) and treated them with CH at the 12.5, 25, and 50 mg/kg/day five times/week in 28 days. In our study, DZN increased lipid profile and liver function tests (LFTs) and creatinine (Cr) but decreased the red blood cell acetylcholinesterase (RBC-AChE) activity and glucose level. Also, CH when co-treated with DZN changed the LFTs, lipid profile, creatine phosphokinase (CPK), lactate dehydrogenase (LDH) and bilirubin total (Bili-T). Moreover, a significant decrease in RBCs, hemoglobin (Hgb), hematocrit (HCT) level, and platelet counts were seen in DZN group but WBCs, lymphocytes, and neutrophils count increased. CH 25 and 50 mg/kg significantly improved alterations of WBCs, RBCs, Hgb, HCT, lymphocytes, neutrophils, and reticulocytes count when co-treated with DZN. Moreover, the co-administration of CH plus DZN recovered histopathological alterations in liver and kidney, as well as, improved the absolute and relative weight of kidney and liver. DZN induced the formation of bone marrow micronuclei (MN) but CH 50 mg/kg decreased the MN formation when co-treated with DZN. These results suggest that CH not only restores renal and hepatic markers, and histopathological alterations but also improves hematological and genotoxicity indices induced by DZN in mice.

Inflammation and the pro-inflammatory cytokines are associated with numerous chronic diseases. Studies suggest that flavonoids, plant polyphenolic compound derivatives from natural origin, have a wide range of putative biological activities. Similar to other flavonoids, chrysin (CH) by its anti-oxidative and anti-inflammatory effects is a potential prophylactic agent in immunopathological and physicochemical injuries. This is an overview on putative immunomodulatory activities of flavonoids and beneficial health effects of these substances particularly, CH in the immune system. CH possesses potent immune-protective effects and suppresses inflammation in innate immune system which results to avoid damages induced by neutrophils and macrophages and suppresses immuno-inflammatory responses. Furthermore, beneficial effects of chrysin on inhibition of serum levels nuclear transcription factor κB (NF-κB) p65 unit, tumor necrosis factor alpha (TNF-α), interleukin-1β (IL-1β), IL-6, IL-12, IL-17A, interferon gamma (IFN-γ) was reported. Moreover, CH has been known as the antagonist of NF-kB and the agonist of peroxisome proliferator-activated receptor gamma (PPAR-γ) which in down regulation of the key pro-inflammatory enzymes such as myeloperoxidase (MPO), cyclooxygenase-2 (COX-2), inducible nitric oxide synthase (iNOS), phospholipase A2, and prostanoids. Therefore, CH can improve immune system and neurodegenerative diseases. In recent years, there has been an increasing interest in compound derivatives from natural origin. Taken together, evidences show that flavonoids may have health-promoting and disease-preventing dietary compounds with important benefits in modern life styles.

Background & Objective: Overexpression of human epidermal growth factor receptor 2 (HER2) causes cell transformation and development of various types of malignancies. Idarubicin is an effective anti-neoplastic drug but its specific delivery to the targeted cells is still a great challenge. Affibody as a cost-effective peptide molecule with low molecular weight has a high affinity for HER2 receptors. Breast and ovarian cancers as wide speared types of malignancies are associated with high expression of HER2. In the current study, we assessed the cytotoxic effects of idarubicin-ZHER2 affibody conjugate on the positive-HER2 cancer cell lines. Methods: The cytotoxic effects of constructed idarubicin-ZHER2 affibody conjugate on the SK-BR-3, SK-OV-3, and MCF-7 cells with various levels of HER2 expression were evaluated by MTT assay following 48 hours of incubation. Results: Idarubicin showed a potent and dose-dependent cytotoxic effect against all treated cell lines while the SK-OV-3 cells were significantly more sensitive. The dimeric form of the ZHER2 affibody molecule showed a mild effect on the cell viability of all treated cells at its optimum concentration. The constructed Idarubicin-ZHER2 affibody conjugate decreased the viability of SK-OV-3 cells at its optimal concentration, more efficiently and specifically than other treated cells. Conclusion: The ZHER2-affibody conjugate of idarubicin has a more specific cytotoxic effect compared with idarubicin alone against HER2-overexpressing ovarian cancerous cells. It appears the ZHER2-affibody conjugate of idarubicin has great potential to be implicated as an innovative anti-cancer agent in future clinical trials in patients with HER2-overexpressing ovarian cancer.

Carbon nanotubes (CNTs) have emerged as a new class of multifunctional nanoparticles in biomedicine, but their multiple in vivo effects remain unclear. Also, the impact of various functionalization types and duration of exposures are still unidentified. Herein, we report a complete toxicological study to evaluate the effects of single- and multiwalled carbon nanotubes (SWCNTs and MWCNTs) with either amine or carboxylic acid (COOH) surface functional groups. The results showed that significant oxidative stress and the subsequent cell apoptosis could be resulted in both acute and, mainly, in chronic intravenous administrations. Also, male reproductive parameters were altered during these exposures. The amino-functionalized CNTs had more toxic properties compared with the COOH functionalized group, and also, in some groups, the multiwalled nanotubes were more active in eliciting cytotoxicity than the single-walled nanotubes. Interestingly, the SWCNTs-COOH had the least alterations in most of the parameters. Evidently, it is concluded that the toxicity of CNTs in specific organs can be minimized through particular surface functionalizations.

Carbon nanotubes (CNTs) have great potential as novel diagnostic or therapeutic tools in biomedicine but, cellular toxicity must be well considered before widespread application of CNTs. Many chemical agents exert their toxicity through apoptotic pathways by induction of caspase biomolecules. In the current study, effects of carboxyl-functionalized single-walled (SW) and multi-walled (MW) CNTs at a single dose of 100 µg ml

This study aimed to evaluate the anti-inflammatory effect of Auraptene (AUR) and Umbelliprenin (UMB) in a rat model of rheumatoid arthritis (RA) induced by using complete Freund's adjuvant (CFA).Paw swelling of adjuvant arthritis rats measured at various times after CFA injection. Over 15 days of RA induction, mediator/cytokine-mediated processes involved in managing the regulation and resolving RA's inflammation were also quantified with ELISA. Histopathological changes were also assessed under a microscope 15 days after the CFA injection.AUR at all doses and UMB administration only at a 16 mM /kg administration dose significantly reduced CFA-induced paw edema level compared to the control group. UMB (64 and 32 mM) and AUR (64, 32, and 16 mM) could reduce the PGE2 (p .05). Unlike indomethacin and prednisolone, treatment of rats with AUR (16, 32, and 64 mM/kg) and UMB (16 and 32 mM/kg) reduced the level of IL-2 (p

Prediction of pharmacokinetics and drug targeting is a challenge in drug design. There are different types of software that can help to predict the pharmacokinetic profile of a drug. Quantitative structure-activity relationship (QSAR) modeling is used for drug design with less cost. Drug-excipient interactions are predicted by docking tools. Computerized drug target prediction and docking programs offer additional options to predict potential effects and adverse reactions of a given candidate as well as the best orientation of the compound on the receptor active site. Information on the absorption, distribution, metabolism and excretion of the drug in the body can enhance prediction of drug release and distribution in the blood and central nervous system (CNS). Computer- aided drug design and delivery can help to save the time and cost in the process of rational drug development.

A decrease in high-density lipoprotein (HDL) level and an increase in atherogenic low-density lipoprotein (LDL) concentration in plasma are involved in the progress of atherosclerosis and incidence of cardiovascular diseases. Cholesteryl ester transfer protein (CETP) passes cholesteryl ester and triglyceride molecules through a hydrophobic tunnel between lipoproteins and via that decreases the HDL/LDL ratio in plasma. Thus, inhibition of CETP activity by specific inhibitors could diminish atherosclerosis occurrence. Anacetrapib as a specific reversible inhibitor of CETP by stabilizing CETP–HDL complex significantly increases the HDL/LDL ratio in plasma. For better understanding the mechanism of CETP inhibition by Anacetrapib, after docking Anacetrapib against a CETP structure, molecular dynamics simulation of CETP and CETP–Anacetrapib in an explicit water cage was performed. Analyses of acquired trajectories show that Anacetrapib increases fluctuation of residues 421–442 of CETP through hydrophobic interactions and thus disrupts the integrity of an α-helix (helix B) structure in the C-terminal concave surface. Creation of a hinge by defeat of the helix B in the C-terminal of CETP, where CETP interacts with LDL, results in missing of an α-helix (residues 436–440) and conversion of banana-shaped CETP to a hemisphere conformation. According to the obtained results, bending of the CETP structure and conversion of the secondary structure in the C-terminal domain in the presence of Anacetrapib may destabilize CETP–VLDL complex and through that shift the dynamics of CETP to the formation of the CETP–HDL complex.

BACKGROUND Cellulose as a renewable biomaterial has forced attention on the use of cellulose-hydrolyzing enzymes for industrial bioconversion of lignocellulosic materials to glucose. This paper reports immobilization of cross-linked cellulase aggregates (CLEA) on the amine-functionalized Fe3O4@silica core-shell magnetic nanoparticles (MNPs). RESULTS The optimum pH of the cellulase cocktail upon immobilization (cellulase CLEA–MNP) shifted a little to the acidic side whereas the optimum temperature did not change significantly. The behavior of CMCase activity in the cellulase CLEA–MNP at pH and temperature values higher than the optimum was significantly different compared with free cellulase. Cellulase CLEA–MNP retained about 45% of its maximum activity at pH values higher than 4.8, while free cellulase lost its activity sharply. Immobilized cellulase in contrast to the free form retained about 65% of its maximum activity at 80°C. Cellulase CLEA–MNP had improved thermal stability at 65°C. Operational stability of the immobilized cellulase was also noticeable. After a sharp decrease during two cycles of CMC hydrolysis, cellulase CLEA–MNP retained 30% of its initial activity through six cycles of reuse. CONCLUSION Simple separation of MNPs from reaction medium and durability of CLEA–MNP composite during repeated use may overcome the major bottleneck against comprehensive applications of cellulase enzymes in industry. © 2014 Society of Chemical Industry

Iron overload in the amount of 10–20 mg/kg of body weight can induce toxicity. Iron causes its toxic effects in a form of oxidative stress and inhibition of key metabolic enzymes. It seems that montmorillonite clay can reduce absorption of elemental iron by iron fixation in the gastrointestinal tract. The potential effect of montmorillonite in decontaminating the gastrointestinal tract was investigated based on serum iron concentration. In the present experiment, ferrous sulfate (100 mg/kg) was administered to Wistar male rats followed by oral gavage of montmorillonite suspensions at three different doses (0.5, 1.0 and 1.5 g/kg). Blood for determination of serum iron concentrations was drawn at 1 h after montmorillonite administration. The results showed that montmorillonite clay with doses of 0.5 and 1.0 g/kg were the best for significantly decreasing the elevated serum iron concentration compared to 1.5 g/kg ( p

Introduction: ABO and Rh are the most important blood group systems with various distributions reported for different populations. There are no reports referring to the distribution of the ABO/Rh blood groups in Iranian Kurdish population. Therefore, the aim of this study was to determine the frequency of these blood groups among this major Iranian ethnic group from Ilam city. Materials and Methods: 1,534 blood donors attending the blood transfusion service in Ilam, Iran, were randomly selected over a period of six months. The ABO/Rh blood groups were determined by the forward and reverse grouping. Results: The most frequent blood group in men and women was A and O; respectively, followed by B and AB. In the Rh system, 91.1% of the donors were positive and 8.9% were negative. Conclusion: This study provided new insight into distribution of the ABO/Rh blood groups in a major ethnic population of west Iran. Moreover, the presented data may also be useful in health care planning, population genetic studies, genetic counseling and medical diagnosis. Asian Journal of Medical Science, Volume-5(3) 2014: 26-29 http://dx.doi.org/10.3126/ajms.v5i3.9151

Trastuzumab (Herceptin(®) ) is a monoclonal antibody (mAb) for specific ablation of HER2-overexpressing malignant breast cancer cells. Intensification of antiproliferative activity of trastuzumab through construction of immunotoxins and nano-immunoconjugates is a promising approach for treatment of cancer. In this study, trastuzumab was directly conjugated to diphtheria toxin (DT). Also, conjugates of trastuzumab and multiwalled carbon nanotubes (MWCNT) were constructed by covalent immobilization of trastuzumab onto MWCNTs. Then, antiproliferative activity of the fusion constructs against HER2-overexpressing SK-BR-3 and also HER2-negative MCF-7 cancer cell lines were examined. Cells treated with trastuzumab-MWCNT conjugates were irradiated with near-infrared (NIR) light. Efficient absorption of NIR radiation and its conversion to heat by MWCNTs can be resulted to thermal ablation of cancerous cells. Our results strongly showed that both trastuzumab-MWCNT and trastuzumab-DT conjugates were significantly efficient in the specific killing of SK-BR-3 cells. Targeting of MWCNTs to cancerous cells using trastuzumab followed by exposure of cells to NIR radiation was more efficient in repression of cell proliferation than treatment for cancer cells with trastuzumab-DT. Our results also showed that conjugation linkers can significantly affect the cytotoxicity of MWCNT-immunoconjugates. In conclusion, our data demonstrated that trastuzumab-MWCNT is a promising nano-immunoconjugate for killing of HER2-overexpressing cancerous cells.

Most of the chemicals in the petrochemical sewages cause oxidative stress in marine organisms. Antioxidant enzymes (catalase (CAT) and superoxide dismutase (SOD)) as biomarkers of oxidative stress and liver histopathological alterations were investigated in the current study to evaluate the toxic effects of petrochemical pollutions in flatfish, Euryglossa orientalis. The enzymatic and histopathological changes were assessed in the liver of E. orientalis from Khowr-e Jafari (one of the creeks from Khowr-e Musa estuary) and Sajafi harbor as polluted and clean areas, respectively. A significant increase in the antioxidant enzyme activities was observed in response to aquatic pollutions of Khowr-e Jafari. Liver lesions were diagnosed and categorized using standard methods. The results of histopathological examinations showed more lesion scores in the fish from Khowr-e Jafari. Various histopathological changes including hepatocyte degeneration, inflammatory lesions, peliosis hepatis and pancreatic acinar cell adenoma, and increase in the number of pigmented macrophage aggregates were observed in the fish from polluted site. It is suggested that activities of CAT and SOD along with semi-quantitative histopathologic analysis of E. orientalis can be used for biomonitoring programs in Persian Gulf.

Human epidermal growth factor receptor 2 (HER2) contributes to the development of breast cancers and malignancies. On the other hand, engineered affibody Z(HER2:342) that binds to HER2 can be successfully used for both diagnostic purposes and specific ablation of malignant HER2-positive cell lines. In the current study, electrostatics-based prediction was applied for improving Z(HER2:342) binding affinity using computational design. The affibody Z(HER2:342) alone and in complex with HER2 was energetically minimized, solvated in explicit water, and neutralized. After heating and equilibration steps, the system was studied by isothermal-isobaric (NPT) MD simulation. According to trajectories, Z(HER2:342) specifically binds to HER2 through hydrogen bonds and salt bridges. Based on the electrostatic binding contributions, two affinity-matured variants namely V1 (Tyr35Arg) and V2 (Asn6Asp and Met9Glu) were rationally designed. More investigations through MD simulation show that V1 interacts with HER2 receptor more strongly, compared to Z(HER2:342) and V2.

Malfunction or overexpression of ErbB receptors (epidermal growth factor receptors) is associated with occurrence and severity of several types of cancer. Initiation of signal cascades by ErbB2 (also known as human epidermal growth factor receptor 2/neu) in breast cancer has been blocked by monoclonal antibodies such as Trastuzumab (Herceptin), which interact with the extracellular domain of human epidermal growth factor receptor 2. Due to some disadvantages of monoclonal antibodies, a new approach in blocking human epidermal growth factor receptor 2 dimerization and activation is designing peptidomimetic ligands based on human epidermal growth factor receptor 2-Trastuzumab interaction model. Growth of human epidermal growth factor receptor 2(+) SK-BR3 cells could be specifically inhibited by peptidomimetic herceptin-based peptidomimetic 5. In this study, herceptin-based peptidomimetic 5 was used as a benchmark peptidomimetic compound to perform a ligand-based virtual screening followed by structure-based screening by applying the molecular docking approach. The study aimed to explore more potent peptidomimetic molecules against extracellular part of human epidermal growth factor receptor 2. Our results showed that the newly designed peptidomimetic herceptin-based peptidomimetic n33B in comparison with herceptin-based peptidomimetic 5 binds more tightly to the extracellular domain of human epidermal growth factor receptor 2. Mechanistic aspects of herceptin-based peptidomimetic n33B interaction with human epidermal growth factor receptor 2 were more investigated through 20-ns molecular dynamic simulations. Additionally, a quantitative structure-activity relationships study was performed to develop a model for identification of structural determinants in herceptin-based peptidomimetic 5-based peptidomimetics.

Background:Expression of human epidermal growth factor receptor type 2 (HER2) in head and neck squamous cell carcinoma (HNSCC) cell line HN5 can be employed with great opportunities of success for specific targeting of anti-cancer chemotherapeutic agents.Objective:In the current study, HER2-specific affibody molecule, ZHER2:342 (an engineered protein with great affinity for HER2 receptors) was selected for conjugation to idarubicin (an anti-neoplastic antibiotic).Method:ZHER2:342 affibody gene with one added cysteine code at the its 5′ end was synthesized de novo and then inserted into pET302 plasmid and transferred to E. Coli BL21 hosting system. After induction of protein expression, the recombinant ZHER2 affibody molecules were purified using Ni- NTA resin and purity was analyzed through SDS-PAGE. Affinity-purified affibody molecules were conjugated to idarubicin through a heterobifunctional crosslinker, sulfosuccinimidyl 4-(Nmaleimidomethyl) cyclohexane-1-carboxylate (Sulfo-SMCC). Specific toxicity of idarubicin-ZHER2 affibody conjugate against two HER2-positive cells, HN5 and MCF-7 was assessed through MTT assay after an exposure time of 48 hours with different concentrations of conjugate.Results:Idarubicin in the non-conjugated form showed potent toxic effects against both cell lines, while HN5 cells were significantly more sensitive compared to MCF-7 cells. Dimeric ZHER2 affibody showed a mild decreasing effect on growth of both HN5 and MCF-7 cells at optimum concentration. Idarubicin-ZHER2 affibody conjugate at an optimum concentration reduced viability of HN5 cell line more efficiently compared to MCF-7 cell line.In conclusion, idarubicin-ZHER2 affibody conjugate in optimum concentrations can be used for specific targeting and killing of HN5 cells.

Tuberculosis that is caused by Mycobacterium tuberculosis is one of the widespread infectious diseases with millions of death worldwide. Prevalence of Multi-drug-resistant (MDR) tuberculosis and emergence of extensively drug-resistant (XDR) species of M. tuberculosis have necessitated the discovery of new and potent antibiotics. Designing of specific inhibitors for 6-hydroxymethyl-7,8-dihydropteroate synthase (DHPS) has attracted a great interest because of its crucial role in de novo folate synthesis in prokaryotic but not in mammalian cells. In the current study, the ligand of DHPS, namely 6-hydroxymethylpterin monophosphate (PtP) was used as a template for structural-based inhibitor design. Virtual screening was performed between compounds with structural similarity to PtP. Between docked compounds against DHPS, 2-amino-6-(1,2,3-trihydroxybutyl)-1H-pteridin-4-one (PtP_1) showed the most negative free energy of binding. Binding of PtP and PtP_1 to DHPS was further investigated through molecular dynamic simulation. DHPS as alone or in complex with ligands was minimized, solvated in an explicit water cage, neutralized by addition of required ions and simulated for 50 ns. According to trajectories, PtP_1 shows high affinity for the active site of DHPS by H-bonds, hydrophobic forces, and cation-? interactions. Our results show that there could be a great chance for designing new inhibitors against DHPS from M. tuberculosis .

Lactate dehydrogenase C (LDHC) as the most abundant isoform of lactate dehydrogenase in testis and spermatozoa, has a critical role in energy metabolism of germ cells. Also, LDHC gene is expressed aberrantly with a high prevalence in various kinds of malignancies. So, LDHC is a promising target protein for development of contraceptive and anti-cancer drugs. In the current study, inhibitory effects of nicotine and cotinine on human LDHC were investigated. Partially purified LDHC was extracted from healthy human spermatozoa and treated with various concentrations of nicotine and cotinine at a range of 0.13-0.66 μM. The results showed that at a concentration of 0.6 μM, cotinine and nicotine could inhibit LDHC activity by 47% and 45%, respectively. Mechanisms of these inhibitory effects were further investigated by docking and molecular dynamic (MD) simulation studies. Nicotine and cotinine were docked against available crystal structure of mice LDHC (PDB ID: 2LDX). LDHC alone and in complex with docked ligands was minimized, solvated in explicit water cage, neutralized by addition of required ions, and finally simulated for 50 ns of MD simulation. Analysis of simulated structures revealed that nicotine and cotinine could affect LDHC structure near His192 which play a key role in the interaction of LDHC with pyruvate/lactate. Thus, it seems that nicotine and cotinine could induce their effects via an inhibitory competition mechanism. Enzymatic assays with various concentration of pyruvate in the presence of mean plasma concentration of nicotine and cotinine in active smokers confirmed MD simulation results.

Background: Over the past several years, the considerable attention has been progressively given to liposomal formulations of anthracyclines. SinaDoxosome® (Exir Nano Sina Company, Iran) is a pegylated liposomal doxorubicin (DOX) which approved by Food and Drug Administration of IRAN for treatment of some types of cancer. The aim of this study was to compare the biodistribution, efficacy, cardiotoxicity and hepatotoxicity of SinaDoxosome® with Caelyx® in mice bearing C-26 colon carcinoma. Methods and results: Mice tumor size evaluation during the experimental period (28 days) showed comparable therapeutic efficacy of nano-formulations. The biodistribution studies showed no significant difference in DOX tissue concentration between Caelyx® and SinaDoxosome®. DOX induced-ECG changes were not detected in nano-formulations. No significant alteration was found in biochemical indexes of myocardial injury in mice exposed to nano-formulations in comparison with control mice. The tissue oxidative parameters such as lipid peroxidation, glutathione, catalase and superoxide dismutase was significantly changed as the results of free DOX treatment. However, the oxidative status of Caelyx® and SinaDoxosome® treated animals did not showed any changes. The experiment also revealed that apoptotic pathway was not activated in the heart of mice exposed to nano-formulations. Conclusions: Although this investigation showed that Caelyx® and SinaDoxosome® are similar in terms of biodistribution, efficacy and toxicity, appropriate clinical evaluations in patients should be considered.

Microperoxidase-11 (also known as MP-11) was chemically modified by boron trifluoride–methanol reagent to obtain methylated MP-11 (mMP-11). Porphyrin transformation capability of mMP-11 in a ternary solvent system was investigated by spectroscopic and kinetic techniques and compared with those of native MP-11. Methyl-modified MP-11 showed higher porphyrin transformation efficiency for synthetic nickel- and vanadium-porphyrins than native MP-11. Upon the chemical modification, the initial rates of porphyrin transformation in a solvent mixture of methylene chloride, methanol and water (up to 5%, v/v) were increased by 60.0-, 4.1-, 7.0- and 1.13-fold for VTPP (V-tetraphenyl porphyrin), VOEP (V-octaethyl porphyrin), NiTPP (Ni-tetraphenyl porphyrin) and NiOEP (Ni-octaethyl porphyrin), respectively. Modified MP-11 also exhibited a good degradation potential in the presence of kerosene (a crude oil fraction). In an organic solvent mixture containing kerosene (35%, v/v), mMp-11 in compared to unmodified MP-11 showed 46.0-, 2.1-, 14.1- and 3.1-times higher activity for VTPP, VOEP, NiTPP and NiOEP, respectively. Experimental data showed that concentration and type of organic hydroperoxides, temperature, and pH of the added water play significant roles on the porphyrin degradation.

An electrochemical immunosensing method was developed based on a magnetic nanocomposite. The multiwalled carbon nanotubes (MWCNTs) were treated with nitric acid to produce carboxyl groups at the open ends. Then, Fe₃O₄ nanoparticles were deposited on COOH-MWCNTs by chemical coprecipitation of Fe²⁺ and Fe³⁺ salts in an alkaline solution. Goat anti-human IgG (anti-hIgG) was covalently attached to magnetic nanocomposite through amide bond formation between the carboxylic groups of MWCNTs and the amine groups of anti-hIgG. The prepared bio-nanocomposite was used for electrochemical sensing of human tetanus IgG (hIgG) as a model antigen. The anti-hIgG magnetic nanocomposite was fixed on the surface of a gold plate electrode using a permanent magnet. The hIgG was detected using horseradish peroxidase (HRP)-conjugated anti-hIgG in a sandwich model. Electrochemical detection of hIgG was carried out in the presence of H₂O₂ and KI as substrates of HRP. Using this method, hIgG was detected in a concentration range from 30 to 1000 ng ml⁻¹ with a correlation coefficient of 0.998 and a detection limit of 25 ng ml⁻¹ (signal/noise=3). The designed immunosensor was stable for 1 month.

Protozoan parasites of the genus Leishmania secrete a range of proteophosphoglycans (PPG) known to be important for successful colonization of Leishmania in the sandfly and for virulence in the mammalian host. PPGs are a large family of extensively glycosylated proteins with some unusual and unique features. In this study we purified PPG from culture supernatant of Leishmania major metacyclic promastigotes. In discontinuous SDS-PAGE, PPG could not enter the resolving gel but after mild acid hydrolysis several bands resolved. Agarose gel electrophoresis and immunoblot analysis using monoclonal antibody (WIC 79.3) indicated that the PPG preparation consisted of heterogeneous molecules. Compositional analysis showed that the PPG preparation contained 67% glycan, 28% protein and 5% phosphate. Additionally, the effect of PPG on reactive oxygen species (ROS) production and induction of IL-10, IL-12 and IFN-gamma secretion by human peripheral blood mononuclear cells (PBMCs) isolated from healthy individuals was investigated. The water-soluble secreted form of PPG at a concentration of 1 microg glycan/ml seems to be a potent inducer of ROS and IL-10 and to a lesser extent of IFN-gamma and IL-12. Cytokines and ROS production was decreased in a dose-dependent manner as the concentration of PPG was increased to 100 microg glycan/ml.

Gold nanoparticles are potentially very attractive components for therapeutic delivery since they can be synthesized with any diameter from 1 to 200 nm to carry a payload of therapeutic molecules into a cell without triggering an immune response. Gold nanoparticles must undergo surface transformations before coupling to therapeutic molecules to become eligible for this purpose. It is now more understood that amine groups can bind to gold nanoparticles strongly, which has enabled surface modification of gold nanoparticles with amino acid lysine through its amine group. These lysine capped gold nanoparticles can further be coupled to therapeutic molecules for delivery purposes. In this study gold nanoparticles were first synthesized and capped with lysine molecules. TEM and FTIR measurements demonstrated the synthesis of lysine-capped gold nanoparticles with an average diameter of 10 nanometers. Interferon alpha molecules-one of the most important therapeutic protein were then chemically bound to lysine-cap...

Aims: The metabolism of phenanthrene and anthracene by a moderate thermophilic Nocardia otitidiscaviarum strain TSH1 was examined. Methods and Results: When strain TSH1 was grown in the presence of anthracene, four metabolites were identified as 1,2-dihydroxy-1,2-dihydroanthracene, 3-(2-carboxyvinyl)naphthalene-2-carboxylic acid, 2,3-dihydroxynaphthalene and benzoic acid using gas chromatography-mass spectrometry (GC-MS), reverse phase-high performance liquid chromatography (RP-HPLC) and thin-layer chromatography (TLC). Degradation studies with phenanthrene revealed 2,2′-diphenic acid, phthalic acid, 4-hydroxyphenylacetic acid, o-hydroxyphenylacetic acid, benzoic acid, a phenanthrene dihydrodiol, 4-[1-hydroxy(2-naphthyl)]-2-oxobut-3-enoic acid and 1-hydroxy-2-naphthoic acid (1H2NA), as detectable metabolites. Conclusions: Strain TSH1 initiates phenanthrene degradation via dioxygenation at the C-3 and C-4 or at C-9 and C-10 ring positions. Ortho-cleavage of the 9,10-diol leads to formation of 2,2′-diphenic acid. The 3,4-diol ring is cleaved to form 1H2NA which can subsequently be degraded through o-phthalic acid pathway. Benzoate does not fit in the previously published pathways from mesophiles. Anthracene metabolism seems to start with a dioxygenation at the 1 and 2 positions and ortho-cleavage of the resulting diol. The pathway proceeds probably through 2,3-dicarboxynaphthalene and 2,3-dihydroxynaphthalene. Degradation of 2,3-dihydroxynaphthalene to benzoate and transformation of the later to catechol is a possible route for the further degradation of anthracene. Significance and Impact of the Study: For the first time, metabolism of phenanthrene and anthracene in a thermophilic Nocardia strain was investigated.

The thermophilic bacterium Nocardia otitidiscaviarum strain TSH1, originally isolated in our laboratory from a petroindustrial wastewater contaminated soil in Iran, grows at 50 degrees C on a broad range of hydrocarbons. Transformation of naphthalene by strain TSH1 which is able to use this two ring-polycyclic aromatic hydrocarbon (PAH) as a sole source of carbon and energy was investigated. The metabolic pathway was elucidated by identifying metabolites, biotransformation studies and monitoring enzyme activities in cell-free extracts. The identification of metabolites suggests that strain TSH1 initiates its attack on naphthalene by dioxygenation at its C-1 and C-2 positions to give 1,2-dihydro-1,2-dihydroxynaphthalene. The intermediate 2-hydroxycinnamic acid, characteristic of the meta-cleavage of the resulting diol was identified in the acidic extract. Apart from typical metabolites of naphthalene degradation known from mesophiles, benzoic acid was identified as an intermediate for the naphthalene pathway of this Nocardia strain. Neither phthalic acid nor salicylic acid metabolites were detected in culture extracts. Enzymatic experiments with cell extract showed the catechol 1,2-dioxygenase activity while transformation of phthalic acid and protocatechuic acid was not observed. The results of enzyme activity assays and identification of benzoic acid in culture extract provide strong indications that further degradation goes through benzoate and beta-ketoadipate pathway. Our results indicate that naphthalene degradation by thermophilic N. otitidiscaviarum strain TSH1 differs from the known pathways found for the thermophilic Bacillus thermoleovorans Hamburg 2 and mesophilic bacteria.

Indigenous thermophilic hydrocarbon degraders are of special significance for the bioremediation of oil-contaminated desert soils with ambient temperature of 45-50 degrees C. The first objective of this study was to demonstrate the hydrocarbon-degrading capability of Nocardia otitidiscaviarum TSH1 (DSM 45,036) which grows optimally at 50 degrees C. Analysis of the metabolic profile of the strain TSH1 showed that it could metabolize phenol, intermediate-chain-length n -alkanes and some polycyclic aromatic hydrocarbons (PAHs) ranging in size from two to four fused rings efficiently, but not toluene and xylene. N. otitidiscaviarum TSH1 was able to survive and grow at phenol concentrations up to 875 mg l(-1). For the first time, the physiological response of a thermophilic Nocardia strain to poorly available hydrophobic compounds was also investigated. When grown on a mineral salt medium with hexadecane, N. otitidiscaviarum TSH1 showed very high affinity for the organic phase. Additionally, PAH-grown cells were considerably hydrophobic. The capacity of PAH-utilizing N. otitidiscaviarum TSH1 isolate to produce biosurfactants was also investigated. Fatty acids (C(14)-C(18)) were detected by GC-MS analysis during bacterial growth in PAH supplemented mineral media. High cell surface hydrophobicity and capability of N. otitidiscaviarum TSH1 to degrade different hydrocarbons at 50 degrees C may make it an ideal candidate to treat oil-contaminated desert soils.

The low stability of recombinant human interferon-γ (rhIFN-γ) therapeutic protein imposes some restrictions in its medical applications. In the current study, the effect of oxygen tension on the stability of purified rhIFN-γ was investigated. The rhIFN-γ was purified (>99%) by a two-step chromatographic process. Storage vials were filled by purified formulated product under normal atmospheric oxygen and low oxygen tension conditions. At different time intervals, the amounts of rhIFN-γ covalent dimers and deamidated forms were analyzed using analytical high-performance liquid chromatography (HPLC; size exclusion and cation exchange) and sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) methods. To determine the biological activity of purified rhIFN-γ, an antiviral activity assay against vesicular stomatitis virus (VSV) was performed. Upon rhIFN-γ long-term storage in a low oxygen tension condition, the amounts of rhIFN-γ covalent dimers and deamidated forms and also the biological activity of rhIFN-γ changed a little. In contrast, by 9 months of storage of rhIFN-γ preparations under normal atmospheric condition, the amount of covalent dimers and deamidated forms increased with time and reached to approximately 3.5% and 11.5% of the initial amount, respectively. The antiviral specific activity of 9-month-old rhIFN-γ preparations decreased to 41% of the initial amount at normal storage condition, while no significant reduction was seen at the low oxygen tension condition. In conclusion, oxygen tension during storage could have a significant impact on rhIFN-γ stability and finally on the quality of pharmaceutical rhIFN-γ product.

Reproductive toxicity of carboxyl-functionalised carbon nanotubes (CNT-COOH), as the most commonly used form of water-soluble CNTs, is not clearly studied. The aim of this study was to investigate in vitro toxicity of carboxylated single-walled and multi-walled CNTs (SWCNT-COOH and MWCNT-COOH) against human spermatozoa. Sperm cells from healthy donors were incubated with 0.1-100 μg/ml of SWCNT-COOH or MWCNT-COOH at 37°C for up to 5 hr. Viability of sperm cells was assessed using MTT test, and sperm motility was evaluated following World Health Organization guideline. Production of reactive oxygen species (ROS) and nitric oxide (NO) in sperm was also assessed. We showed that both MWCNT-COOH and SWCNT-COOH following incubation in vitro with human spermatozoa did not exert negative effect on viability while motility was significantly (p

Malfunction or overexpression of ErbB receptors (epidermal growth factor receptors) is associated with occurrence and severity of several types of cancer. Initiation of signal cascades by ErbB2 (also known as human epidermal growth factor receptor 2/neu) in breast cancer has been blocked by monoclonal antibodies such as Trastuzumab (Herceptin), which interact with the extracellular domain of human epidermal growth factor receptor 2. Due to some disadvantages of monoclonal antibodies, a new approach in blocking human epidermal growth factor receptor 2 dimerization and activation is designing peptidomimetic ligands based on human epidermal growth factor receptor 2-Trastuzumab interaction model. Growth of human epidermal growth factor receptor 2(+) SK-BR3 cells could be specifically inhibited by peptidomimetic herceptin-based peptidomimetic 5. In this study, herceptin-based peptidomimetic 5 was used as a benchmark peptidomimetic compound to perform a ligand-based virtual screening followed by structure-based screening by applying the molecular docking approach. The study aimed to explore more potent peptidomimetic molecules against extracellular part of human epidermal growth factor receptor 2. Our results showed that the newly designed peptidomimetic herceptin-based peptidomimetic n33B in comparison with herceptin-based peptidomimetic 5 binds more tightly to the extracellular domain of human epidermal growth factor receptor 2. Mechanistic aspects of herceptin-based peptidomimetic n33B interaction with human epidermal growth factor receptor 2 were more investigated through 20-ns molecular dynamic simulations. Additionally, a quantitative structure-activity relationships study was performed to develop a model for identification of structural determinants in herceptin-based peptidomimetic 5-based peptidomimetics.

A bacterial strain, FBHYA2, capable of degrading naphthalene, was isolated from the American Petroleum Institute (API) separator of the Tehran Oil Refinery Complex (TORC). Strain FBHYA2 was identified as Achromobacter sp. based on physiological and biochemical characteristics and also phylogenetic similarity of 16S rRNA gene sequence. The optimal growth conditions for strain FBHYA2 were pH 6.0, 30 °C and 1.0% NaCl. Strain FBHYA2 can utilize naphthalene as the sole source of carbon and energy and was able to degrade naphthalene aerobically very fast, 48 h for 96% removal at 500 mg/L concentration. The physiological response of Achromobacter sp., FBHYA2 to several hydrophobic chemicals (aliphatic and aromatic hydrocarbons) was also investigated. No biosurfactant was detected during bacterial growth on any aliphatic/aromatic hydrocarbons. The results of hydrophobicity measurements showed no significant difference between naphthalene- and LB-grown cells. The capability of the strain FBHYA2 to degrade naphthalene completely and rapidly without the need to secrete biosurfactant may make it an ideal candidate to remediate polycyclic aromatic hydrocarbon (PAH)-contaminated sites.

An electrochemical immunosensing method was developed based on a magnetic nanocomposite. The multiwalled carbon nanotubes (MWCNTs) were treated with nitric acid to produce carboxyl groups at the open ends. Then, Fe₃O₄ nanoparticles were deposited on COOH-MWCNTs by chemical coprecipitation of Fe²⁺ and Fe³⁺ salts in an alkaline solution. Goat anti-human IgG (anti-hIgG) was covalently attached to magnetic nanocomposite through amide bond formation between the carboxylic groups of MWCNTs and the amine groups of anti-hIgG. The prepared bio-nanocomposite was used for electrochemical sensing of human tetanus IgG (hIgG) as a model antigen. The anti-hIgG magnetic nanocomposite was fixed on the surface of a gold plate electrode using a permanent magnet. The hIgG was detected using horseradish peroxidase (HRP)-conjugated anti-hIgG in a sandwich model. Electrochemical detection of hIgG was carried out in the presence of H₂O₂ and KI as substrates of HRP. Using this method, hIgG was detected in a concentration range from 30 to 1000 ng ml⁻¹ with a correlation coefficient of 0.998 and a detection limit of 25 ng ml⁻¹ (signal/noise=3). The designed immunosensor was stable for 1 month.

Bacterial lipopolysaccharide (LPS) has T-helper 1 (Th1) immunostimulatory activities but because of toxicity and pyrogenicity cannot be used as an adjuvant. Brucella abortus LPS has less toxicity and no pyrogenic properties in comparison to other bacterial LPS. In the current study, the immunostimulatory properties of B. abortus LPS were evaluated for its adjuvant activity. Tuberculin purified protein derivative (PPD) from Mycobacterium tuberculosis was extracted and after anion-exchange chromatography on Q-sepharose column, two fractions (17 and 23), which dominantly contained 30- and 70-kDa antigens, were collected for immunological studies. BALB/c mice were immunized with four different antigen preparations (BCG, PPD, 17th and 23rd PPD fractions) along with complete Freund’s adjuvant or B. abortus LPS. The T-cell immune response of mice was assessed by measurement of Th1-type cytokine (IFN-γ) and Th2-type cytokines (IL-5 and IL-10) levels. Also, the humoral immunity was evaluated by measuring the specific IgG levels. Our results showed that immunization of mice with 17th PPD fraction along with B. abortus LPS can induce a Th1-type cytokine response characterized with a high IFN-γ/IL-5 ratio, while immunization with PPD or 23rd PPD fraction along with the same adjuvant resulted to a mixed Th1/Th2-type cytokine response.

Tuberculosis (TB) represents one of the leading killers among all infectious disease. Protection against TB depends on the activation of T-helper type I (Th1) immune response. Carbon nanotubes (CNTs) have attracted considerable attention because of their potential applications as new nanovehicle. In the current study, tuberculin purified protein derivative (PPD) was conjugated to carboxylated single-walled carbon nanotubes (SWCNTs). Cytotoxicity of the carboxylated SWCNT and SWCNT-PPD conjugate was analyzed with MTT assay and by reactive oxygen species (ROS) and nitric oxide (NO) generation. Male BALB/c mice were immunized with BCG, PPD, SWCNT-PPD conjugate and PPD in complete Freund's adjuvant (CFA). Induction of cellular immune response was analyzed by measuring the levels of Th1 cytokines (IFN-gamma and IL-12) and Th2 cytokines (IL-10 and IL-5). Immunization with non-conjugated PPD or PPD in Freund's adjuvant induced a Th2 cytokine response while immunization with BCG resulted to a mixed Th1/Th2 cytokine response. In contrast, PPD in conjugation with SWCNT generated preferentially a Th1-type cytokine response in the absence of potential cytotoxic effects.

Indigenous thermophilic hydrocarbon degraders are of special significance for the bioremediation of oil-contaminated desert soils with ambient temperature of 45-50 degrees C. The first objective of this study was to demonstrate the hydrocarbon-degrading capability of Nocardia otitidiscaviarum TSH1 (DSM 45,036) which grows optimally at 50 degrees C. Analysis of the metabolic profile of the strain TSH1 showed that it could metabolize phenol, intermediate-chain-length n -alkanes and some polycyclic aromatic hydrocarbons (PAHs) ranging in size from two to four fused rings efficiently, but not toluene and xylene. N. otitidiscaviarum TSH1 was able to survive and grow at phenol concentrations up to 875 mg l(-1). For the first time, the physiological response of a thermophilic Nocardia strain to poorly available hydrophobic compounds was also investigated. When grown on a mineral salt medium with hexadecane, N. otitidiscaviarum TSH1 showed very high affinity for the organic phase. Additionally, PAH-grown cells were considerably hydrophobic. The capacity of PAH-utilizing N. otitidiscaviarum TSH1 isolate to produce biosurfactants was also investigated. Fatty acids (C(14)-C(18)) were detected by GC-MS analysis during bacterial growth in PAH supplemented mineral media. High cell surface hydrophobicity and capability of N. otitidiscaviarum TSH1 to degrade different hydrocarbons at 50 degrees C may make it an ideal candidate to treat oil-contaminated desert soils.

Aims: Our goal was the characterization of a new moderate thermophilic polycyclic aromatic hydrocarbon (PAH)-utilizing Nocardia strain. Methods and Results: A thermophilic bacterium, strain TSH1, was isolated from a contaminated soil. The macroscopic and microscopic features fit well with the description of Nocardia species. The results of 16S rRNA gene analysis showed 100% match to the type strain of N. otitidiscaviarum DSM 43242T. Strain TSH1 showed the same mycolic acid pattern as the type strain of N. otitidiscaviarum but its fatty acid profile did not permit identification to the species level. The carbon utilization profile of strain TSH1 was different from N. otitidiscaviarum. The results of hydrophobicity measurements showed that PAHs-grown cells were significantly more hydrophobic than LB-grown cells. Furthermore, biosurfactant production was detected during bacterial growth on different culture media. Conclusions: Strain TSH1 is capable of growing on a range of PAHs. When grown in PAHs-supplemented media, strain TSH1 showed a high affinity for the organic phase, suggesting that it can develop a hydrophobic surface. Significance and Impact of the Study: High cell surface hydrophobicity and capability of strain TSH1 to degrade different PAHs at 50°C may make it an ideal candidate to treat PAH-contaminated desert soils.

Experimental murine models with high, intermediate and low levels of genetically based susceptibility to Leishmania major infection reproduce almost entire spectrum of clinical manifestations of the human disease. There are increasing non-comparative studies on immune responses against isolated antigens of L. major in different murine strains. The aim of the present study was to find out whether there is an antigen that can induce protective immune response in resistant and susceptible murine strains. To do that, crude antigenic extract of procyclic and metacyclic promastigotes of L. major was prepared and subjected to SDS-PAGE electrophoresis. Western-blotting was used to search for antigen(s) capable of raising high antibody level of IgG2a versus IgG1 in the sera of both infected resistant and susceptible strains. Two novel antigens from metacyclic promastigotes of L. major (140 and 152 kDa) were potentially able to induce specific dominant IgG2a responses in BALB/c and C57BL/6 mice. The 2 antigens also reacted with IgG antibody of cutaneous leishmaniasis patients. We confirm that 140 and 152 kDa proteins of L. major promastigotes are inducing IgG production in mice and humans.

Carbon nanotubes (CNTs) could serve as nanovehicle for drug delivery. It is proposed that this class of emerging materials is capable to penetrate into cells without causing cell death. However, relatively little is known about the potential biological risks from CNTs. Nanoparticles present enormous surface areas and are found to enhance the rate of protein fibrillation by decreasing the lag time for nucleation. Protein fibrillation is associated with protein misfolding diseases including Type II Diabetes, Alzheimer’s, Parkinson's, Creutzfeld-Jacob disease and Dialysis-related amyloidosis. In the present study, via Congo red binding assay and circular dichroism (CD) spectroscopy we investigated the amyloidogenic effects of crude and carboxylated-CNT on the in-vitro fibrillogenesis of hen-egg wight lysozyme at acidic condition. Our results showed that the formation of lysozyme amyloid fibrils at pH 2.0 and 55°C was remarkably increased by the addition of crude unmodified CNTs, whereas carboxylated CNTs did not enhance the probability of appearance of a critical nucleus for nucleation of protein fibrils. Introduction: Recently, considerable effort has been generated to characterize intermediate structures accumulated during protein folding and unfolding (1). Interestingly growth of toxic amyloid structures, is supposed to be associated with various late-onset neurodegenerative disorders, initiated from intermediate structures interaction (2, 3). The potential of nanoparticles to promote protein fibrillation is demonstrated (4). Previous investigations by different research groups showed that exposure of mammalian cells to CNTs caused inhibition of cell proliferation (5), activation of nuclear transcription factor-κB (6) and also increase in oxidative stress and accumulation of peroxidative products (7). Our previous results showed no significant decrease in viability of cells exposed to different concentrations of carboxylated-CNTs (8). Additionally; no significant increase in ROS was seen at CNT concentrations ranging from 1 to 15μgml−1 (8). Also, our finding confirmed that carboxylated-CNTs probably have no interaction with the expression of the iNOS protein (8). In the present study, acidic fibrillation of hen-egg wight lysozyme (HEWL) was analyzed in the presence of prepared crude and carboxylfunctionalized nanotubes. Materials & Methods: Preparation of carbon nanotubes: As-received multi-walled CNTs (10 mg) were suspended in concentrated H2SO4:HNO3 solution (3:1 v/v) and sonicated in a sonication bath for 4 h at 40°C. The CNT suspension was diluted with distilled water and a CNT mat was obtained after filtration through a 0.45 μm polycarbonate membrane. The CNT mat was washed until no acid was detected and then was suspended in 25 ml deionized water (di-H2O) by sonication. IR-spectroscopy was used to verify the CNT functionalization. Amyloid fibril formation condition: A sample solution of 2 mg/ml HEWL in di-H2O (pH 2.0) with salts (136 mM NaCl, 2.7 mM KCl) was prepared. Nanotubes at two final concentrations (2.5 and 10 μg/ml) were added to HEWL solution. HEWL sample solutions were incubated at 55°C for different times. Congo red binding assay: Increase in the absorbance intensity combined with a red shift in the absorbance spectrum of Congo red, is a method of choice to characterize amyloid fibrillation (9). The absorbance spectrum was acquired on Shimadzu UV-visible Spectrophotometer (Kyoto, Japan) in the 400to 600-nm region. Far UV-Circular Dichroism (CD): Far-UV CD spectra were acquired on Aviv 215 spectropolarimeter (Aviv Associates, Lakewood, NJ). A quartz cell with 0.1 mm light path and a protein concentration of 0.6 mg/ml was used to record CD spectrum from 190 to 260 nm. Results and Discussion: The IR spectrum of the acid-treated multi-walled CNTs showed peaks at 3424, 1719 and 1626cm attributed to the hydroxyl, carboxyl and carbonyl functional groups, respectively. Nanoparticles present enormous surface areas and are found to enhance the rate of protein fibrillation by decreasing the lag time for nucleation (4). In this study, effect of nano tubes with polar and non-polar surfaces on the process of amyloid fibrillation was investigated. Results from Congo red absorption spectroscopy demonstrated the formation of fibrillar amyloid structures at 72 hours after the initiation of HEWL incubation in amyloidogenic condition. In addition, CD spectra of HEWL exhibited a characteristic pattern of β-sheet conformation (Fig. 1). In exploring if CNT exerted a stimulatory action on the formation of HEWL fibrils, we monitored the changes of Congo red binding and CD spectra in amyloidogenic conditions. Our observations indicated that crude CNT could outstandingly enhance amyloid fibril formation while CNT in carboxylated form had no significant effect on fibrillation process. It is concluded, introduction of polar groups on the hydrophobic surfaces may play an important inhibitory role on surface-assisted nucleation. References: [1] Dobson C.M., Nature, 426 (2003) 884-890. [2] Dobson C.M., Trends Biochem. Sci, 24 (1999) 329-332 [3] Chiti F., Dobson C.M., Nature Chemical Biology 5 (2009) 15 – 22. [4] Linse S., Cabaleiro-Lago C., Xue W.F., Lynch I., Lindman S., Thulin E., Radford S. E., Dawson K. A., PNAS 104 (2007) 8691-9696. [5] Shvedova A.A., Castranova V., Kisin E.R., Schwegler-Berry D., Murray A.R., Gandelsman V., J Toxicol Environ Health A 66 (2003) 1909–26. [6] Kam N.W.,,Jessop T.C., Wender P.A., Dai H., J Am Chem Soc 126 (2004) 6850-1. [7] Manna S.K., Sarkar S., Barr J., Wise K., Barrera E.V., Jejelowo O., Nano Lett 5 (2005) 1676–84. [8] Zeinalia M., Jammalana M., Ardestania S.K., Mosaveri N., Immunology Letters 126 (2009) 48-53. [9] Naiki H., Higuchi K., Hosokawa M., Takeda T., Anal Biochem 177 (1989) 244–249.