Investigation of inhibitory effect of sulfated chitosan oligomer on human heparanase enzyme: in silico and in vitro studies.

Deaths from cancer are widespread worldwide and the numbers continue to increase day by day. During the disease progression of cancer in cells, many of its metabolic activities change. Increased heparanase enzyme release is just one example. Following heparanase enzyme activity, many molecules interact with the remodeling of glycosaminoglycan structures, which triggers the release of different enzymes, cytokines, and growth factors, including fibroblast growth factors (FGF1 and FGF2), vascular endothelial growth factor (VEGF), hepatocyte growth factor, transforming growth factor ß and platelet-derived growth factor. These are the most important factors in metastasis due to the formation of new vascular structures caused by those elements. To reduce tumor growth and metastasis, various drugs have been designed by modifying chitosan and its derivatives. In this study, we used chitosan oligomer (A), sulfated chitosan oligomer (ShCsO) (B), heparin (C), phosphate monomer (D1) of PI-88 and sulfate monomer (D2) of PI-88 as heparanase inhibitors. We modified the chitosan oligomer with chlorosulfonic acid to synthesize ShCsO to investigate its inhibitory effects on human serum heparanase. Also examined were molecular docking; molecular dynamics (MD); adsorption, distribution, metabolism, elimination and toxicity (ADMET); and target prediction. ShCsO decreased enzyme activity at a concentration of 0.0001 mg/mL. The docking scores of A, B and C from in silico studies were -6.254, -6.936 and -6.980 kcal/mol, respectively, and the scores for the two different PI-88 monomers were -5.741 and -5.824 kcal/mol. These results show that ShCsO may be a potential drug candidate for treating cancer.Communicated by Ramaswamy H. Sarma.

Synthesis, Characterization, and Investigation of Doxorubicin Drug Release Properties of Poly(acrylamide-co-acrylic Acid/Maleic Acid)- Hydroxyapatite Composite Hydrogel.

BACKGROUND: Hydroxyapatite and its derivatives have been used for a lot of applications. One of them is drug release studies. Due to its low adhesion strength and lack of the strength and durability required for load-carrying applications, there is a need to improve the properties of hydroxyapatite. For this aim, the most important factors are increasing pH sensitivity and preventing coagulation. Mixing it with multifunctional polymers is the best solution. OBJECTIVES: The main objectives are: 1- preparing poly(acrylamide-co-acrylic acid/maleic acid)- hydroxyapatite (PAm-co-PAA/PMA-HApt), 2- assessment of (PAm-co-PAA/PMA-HApt) and dox-loaded poly(acrylamide-co-acrylic acid/maleic acid) (Dox-(PAm-co-PAA/PMA-HApt)) composite hydrogels, and 3- elucidating the difference in behavior of drug release studies between hydroxyapatite (HApt) and poly(acrylamide-co-acrylic acid/maleic acid) composite hydrogels. METHODS: A composite of PAm-co-PAA/PMA-HApt was prepared by direct polymerization of acrylamide-co-acrylic acid/maleic acid in a suspension of HApt. The drug loading and release features of PAm-co-PAA/PMA-HApt and HApt were then investigated for doxorubicin (dox) release. Using Fourier transform infrared spectroscopy (FTIR), X-ray diffraction (XRD), scanning electron microscopy (SEM), and thermogravimetric analysis (TG/DTA), this unique composite hydrogel has been physicochemically investigated. Also, a colorimetric assay was used to assess the in vitro biocompatible support and anticancer activity of HApt and the newly developed composite hydrogel XTT (2,3-Bis-(2-Methoxy-4-Nitro-5-Sulfophenyl)-2H-Tetrazolium-5-Carboxanilide) assay. RESULTS: According to the results of drug release studies of this new material, it is pH sensitive, and PAm-co-PAA/PMA-HApt demonstrated a faster release than HApt at 37 °C in the acidic solution of pH 4.5 than in the neutral solution of pH 7.4. The XTT assay outcomes also demonstrated the biocompatibility of PAm-co-PAA/PMA-HApt and HApt and the cytotoxic effect of dox-loaded PAm-co-PAA/PMA-HApt. CONCLUSION: It should be inferred that the drug release profile was improved at pH 4.5 by the newly produced pH-sensitive composite hydrogel.

Preparation and characterization of lysozyme loaded cryogel for heavy metal removal.

In the present study, monolithic poly(N-isopropylacrylamide-acrylamide)-acrilic acid (poly(npam-aam)-aac) cryogels were made. Swelling tests, SEM, XRD, and ATR-FTIR analyses revealed distinct cryogel and lysozyme-loaded cryogel properties. The equilibrium swelling degree was 6.2 g H2O/g cryogel. The created poly(npam-aam)-aac with pores of 10-100 µm was obviously seen in SEM images. Lysozyme adsorption capacity on poly(npam-aam)-aac was found to be 260 mg/g at pH 7.4 and 40 °C. After that, we used lysozyme adsorbed cryogel for the removal of the model heavy metal ion (cadmium). A series of pH, duration, and ionic strengths were used to conduct Cd2+ adsorption experiments. The results showed that the new adsorbent had a considerable chemical affinity for Cd2+ ions in its ability to bind them under eye ocular conditions (pH 7.4, 32-36 °C, 0,15 M NaCl). The traditional Langmuir adsorption model was the most suitable, achieving maximum uptake of ∼185 mg/g. Chemical adsorption was found to be the rate-controlling step, and the process was also compatible with the pseudo-second-order model. For the treatment of ocular pathologies, the most effective enzyme, lysozyme, must show its function. That is why there is a need for using lysozyme, and lysozyme is selected as a lignad to adsorb heavy metal ions because of its high heavy metal binding affinity. This material could be used for the treatment of ocular pathologies in the future.

Magnetic chitosan oligomer-sulfonate-stearic acid triple combination as cisplatin carrier for site-specific targeted on MCF-7 cancer cells: Preparation, characterization and in vitro experiments.

In this study, a new amphiphilic target-specific adsorbent, chitosan oligomer-sulfonate-stearic acid triple combination (S-Cho-SA), and magnetic chitosan oligomer-sulfonate-stearic acid triple combination (M-S-Cho-SA) by oleic acid (OA)-modified Fe3 O4 via hydrophobic interaction are fabricated. By modifying the nanoparticle surfaces and having the ability to magnetically allow the target region, these particles attract attention as important particles used in targeting mechanisms in cancer therapy. With magnetic nanoparticles and an external magnetic field, it is possible to transport therapeutic agents to the target site and keep them in the desired effect zone for a longer period of time. These new adsorbents are characterized by scanning electron microscopy (SEM), attenuated total reflection Fourier transform infrared (ATR FT-IR) spectroscopy, nuclear magnetic resonance (NMR), X-ray diffraction (XRD), vibrating sample magnetometer (VSM), and thermogravimetric analysis (TG/DTA). After chemical characterization, it is complexed with cisplatin (CDDP). The magnetic adsorbents were loaded with high efficiency (>50%), and the release experiments exhibited that cisplatin is released more at pH 4.5 compared with pH 7.4 at 37°C. It showed better drug release results under a magnetic field for magnetic adsorbents (36% for pH 4.5 and 3.6% for pH 7.4). The biocompatibility of the prepared adsorbents was demonstrated via the XTT assay in MCF-7 cell lines. The results also exhibited that S-Cho-SA and M-S-Cho-SA were biocompatible, and free cisplatin and cisplatin-complexed adsorbents showed an antiproliferative effect. The results showed that these new cisplatin-loaded (M-S-Cho-SA) nanoparticles are good candidates for thermotherapy in cancer treatment in the future, as they can provide selectivity by site-specific targeting and hold onto an alternative magnetic field due to the magnetic nature of the nanoparticles.

Preparation of sulfatide mimicking oleic acid sulfated chitosan as a potential inhibitor for metastasis.

Sulfatide is associated with numerous health problems, affecting different parts of the human body, including the metastasis; however, the underlying mechanisms are yet to be fully elucidated. Sulfatide has been used to potential inhibitor for tumor cell metastasis. In the present study we synthesized oleic acid sulfated chitosan (OlcShCs). It shows structural similarity to sulfatide because of its functional groups (sulfate and fatty acyl chains). Chitosan has smart properties such as biocompatibility, biodegradability and non-toxicity. We have prepared oleic acid sulfated chitosan (OlcShCs) by chitosan modification to mimic sulfatide. Its structure was characterized by FT-IR, H-NMR, and thermogravimetric analysis. After characterization studies its antimicrobial, antifungal and cytotoxic properties were investigated. Oleic acid sulfated chitosan (OlcShCs) was tested for its anti-cancer potential against human cancer cell lines (HeLa (ATCC® CCL-2™)) for 24 h, 48 h and 72 h using the MTT assays. This new material which is soluble at physiological conditions, is a potential candidate for further metastasis inhibition investigations.

Wi-Fi decreases melatonin protective effect and increases hippocampal neuronal damage in pentylenetetrazole induced model seizures in rats.

AIM: Epilepsy is a common brain disorder in which the seizures could cause a neuronal loss in the hippocampus. Oxidative stress has an important role in the pathology of epilepsy. Some studies indicate that Wi-Fi increases oxidative stress and suppresses antioxidant systems. The aim of this study is to investigate the effect of Wi-Fi on melatonin anticonvulsive effect and oxidative damage in pentylenetetrazole-induced epileptic seizures in rats. METHODS: In our study, we used 30 male Wistar Albino rats, 230-250 grams of the body weight. The animals were divided into five groups as control, saline (1 ml/kg/day olive oil for 30 days), Wi-Fi (12 h/day for 30 days), melatonin (10 mg/kg/day for 30 days) and melatonin + Wi-Fi (10 mg/kg/day +12 h/day for 30 days). In the thirtieth day, thirty minutes after the last drugs administration at the indicated doses, PTZ in 45 mg/kg was administered to induce epileptic seizure. The animals were observed for 30 min during the seizure stages (according to the Racine Scale) and first myoclonic jerk times (FMJ). Twenty-four hours after PTZ injection, brain tissues were removed for biochemical and histopathological evaluation. The hippocampal Cornu Ammonis (CA) 1, CA3 and DG (dentate gyrus) regions were histopathologically evaluated in terms of a neuronal damage in addition that oxidative stress markers (total antioxidant status (TAS), total oxidant status (TOS) and oxidative stress index (OSI)) were measured in brain tissues. RESULTS: Wi-Fi was not found to affect behavioral changes associated with epilepsy (p > 0.05). However, Wi-Fi reduced anticonvulsive and antioxidant effect of melatonin (p < 0.05). Moreover, Wi-Fi increased neuronal damage in hippocampus (p < 0.05). CONCLUSION: Wi-Fi did not directly affect epileptic seizures. Nevertheless, it inhibits the positive effects of melatonin on epilepsy and it also has negative effects on hippocampal neuronal damage. These effects of Wi-Fi may occur via oxidative pathways.

Mercury leads to abnormal red blood cell adhesion to laminin mediated by membrane sulfatides.

Exposure to mercury is associated with numerous health problems, affecting different parts of the human body, including the nervous and cardiovascular systems in adults and children; however, the underlying mechanisms are yet to be fully elucidated. We investigated the role of membrane sulfatide on mercuric ion (Hg2+) mediated red blood cell (RBC) adhesion to a sub-endothelial matrix protein, laminin, using a microfluidic system that mimics microphysiological flow conditions. We exposed whole blood to mercury (HgCl2), at a range of concentrations to mimic acute (high dose) and chronic (low dose) exposure, and examined RBC adhesion to immobilized laminin in microchannels at physiological flow conditions. Exposure of RBCs to both acute and chronic levels of Hg2+ resulted in elevated adhesive interactions between RBCs and laminin depending on the concentration of HgCl2 and exposure duration. BCAM-Lu chimer significantly inhibited the adhesion of RBCs that had been treated with 50 µM of HgCl2 solution for 1 h at 37 °C, while it did not prevent the adhesion of 3 h and 24 h Hg2+-treated RBCs. Sulfatide significantly inhibited the adhesion of RBC that had been treated with 50 µM of HgCl2 solution for 1 h at 37 °C and 0.5 µM of HgCl2 solution for 24 h at room temperature (RT). We demonstrated that RBC BCAM-Lu and RBC sulfatides bind to immobilized laminin, following exposure of RBCs to mercuric ions. The results of this study are significant considering the potential associations between sulfatides, red blood cells, mercury exposure, and cardiovascular diseases.

Production, purification, and characterization of metalloprotease from Candida kefyr 41 PSB.

A thermostable metalloprotease, produced from an environmental strain of Candida kefyr 41 PSB, was purified 16 fold with a 60% yield by cold ethanol precipitation and affinity chromatography (bentonite-acrylamide-cysteine microcomposite). The purified enzyme appeared as a single protein band at 43kDa. Its optimum pH and temperature points were found to be 7.0 and 105°C, respectively. Km and Vmax values of the enzyme were determined to be 3.5mg/mL and 4.4µmolmL-1min-1, 1.65mg/mL and 6.1µmolmL-1min-1, using casein and gelatine as the substrates, respectively. The activity was inhibited by using ethylenediamine tetraacetic acid (EDTA), indicating that the enzyme was a metalloprotease. Stability of the enzyme was investigated by using thermodynamic and kinetic parameters. The thermal inactivation profile of the enzyme conformed to the first order kinetics. The half life of the enzyme at 95, 105, 115, 125 and 135°C was 1310, 610, 220, 150, and 86min, respectively.

Characterization of thermostable ß-amylase isozymes from Lactobacillus fermentum.

A strain of Lactobacillus fermentum producing two isozymes of a 20kDa ß-amylase was isolated from the faecal sample of a newborn. The starin was identified by sequencing its 16S rRNA gene. The two ß-amylase isozymes were resolved and visualized by two dimensional protein gel electrophoresis (2-D gel electrophoresis). Some of the physical and biochemical properties of the enzymes were characterized. The ß-amylase displayed two optimum pH s, 5.0 and 10.0 and two optimum temperatures, 45°C and 37°C, respectively. The isozymes hydrolyzed different substrates: glycogen at pH 5.0, and corn starch at pH 10.0. The activity did not require Ca2+, though the activity at pH 10.0 was enhanced in the presence of 5.0mM and 10.0mM CaCl2, 110% and 130%, respectively.

Production and immobilization of a novel thermoalkalophilic extracellular amylase from bacilli isolate.

A Thermoalkalophilic amylase was produced from an environmental bacterial isolate. The enzyme was then immobilized through its amino groups onto the epoxy rings of magnetic poly glycidyl methacrylate [m-poly (GMA)] beads. The free enzyme was active within a large pH range, between 7 and 12 and displayed the optimum activity at 95°C and pH 10. The immobilization appeared to increase the stability of the enzyme as its bound form showed optimum activity at 105°C and pH 11.0. Kinetic studies demonstrated that immobilized enzyme had higher K(m) and lower V(max) values. The activity of the free and bound enzyme was determined, at 37°C and pH 10.0 and pH 11.0, respectively, in the presence of various organic solvents and detergents (5%, v/v). Results obtained indicated that detergents, sodium dodecyl sulfate (SDS) and TritonX-100, caused six fold increase and that various organic solvents also increased the activity of the amylase.

IgG purification by bentonite-acrylamide-histidine microcomposite.

In this work, a new microcomposite composed of bentonite, acrylamide and histidine, as a pseudospecific ligand, was synthesized by bulk polymerization. The aim of this study was to improve IgG adsorption capacity of bentonite by incorporating histidine. The surface areas of the bentonite and bentonite-acrylamide-histidine microcomposites were 33.4 and 1.42 m(2)/g, respectively. The amount of histidine was found to be 50 µmol/g bentonite via elemental analysis. Adsorption capacity was at the value of 100mg/g from aqueous solution while adsorption capacity was 108 mg/g from human plasma with a purity of 90%. IgG biomolecules were able to be adsorbed and desorbed five times by using the same microcomposites without significant loss in their adsorption capacity.