Downregulation of Stemness Genes and Induction of Necrosis in Rat LA7 Cancer Stem Cells Induced Tumors Treated with Starved Fibroblasts Culture Supernatant

Background:Stem cell differentiation therapy is a promising strategy in cancer treatment. we show that protein cocktail prepared from serum starved fibroblasts has therapeutic potential based on this strategy. Methods:The condition medium was prepared from foreskin isolated fibroblasts and analyzed by Liquid chromatography electrospray ionization mass spectrometry-mass spectrometry (LC-ESI-MS/MS). LA7 mammary gland cancer stem cells originated tumors were induced in Sprague Dawley rats. The rats treated subcutaneously with DMEM (group A), condition medium (group B), or normal saline (group C) once daily for 7 days. Then the tumors were removed and divided into the two parts, one part was used to quantify gene expression by stem-loop RT-qPCR assay and the other part was used for Hematoxylin & Eosin (H & E), Giemsa, and immunohistochemistry (IHC) staining.Results:All induced tumors appeared as sarcomatoid carcinoma (SC). Immunohistochemistry staining confirmed this conclusion by recognizing the tumor as Ki67⁺, cytokeratin⁺, vimentine⁺, and estrogen receptor negative SC. RT-qPCR analysis revealed that Oct4-, Sox-2, Nanog- gene expression was much reduced in the condition medium treated tumors versus proper controls (p< 0.05). Tissue necrosis was more prevalent in this group while tumors volume was diminished almost by 40%. The LC-ESI-MS/MS analysis unrevealed the stemness reducing and the cell death inducing proteins such as, pigment epithelium-derived factor (PEDF), insulin like growth factor binding protein-5 (IGFBP-5) and -7 (IGFBP-7) in the condition medium.Conclusion:This study showed that the substances released from starved human fibroblasts were able to down-regulate the stemness-related genes and induce necrosis in LA7 derived tumors.Key Words: Breast cancer, Cancer Stem cells, Cell differentiation, Fibroblasts, Gene expression     

Improvement of cytotoxicity of mitoxantrone and daunorubicin by candidone,tephrosin, and bavachinin

Molecular Biology Reports - Flavonoids have been demonstrated to have the ability of sensitizing cancer cells to chemotherapy and inverse multidrug resistance via various mechanisms, such as...     

MNPs-IHSPN nanoparticles in multi-application with absorption of bio drugs in vitro

The aim of this project is to investigate the method of using a common buffer to determine the degree of stabilization and secretion of two drug molecules that have been analyzed in vitro. First, magnetic nanoparticles were synthesized and their structure was identified by instruments such as XPS (X-ray photoelectron spectroscopy) and FT-IR (Fourier transform infrared spectroscopy). The main purpose of this study was to investigate the stabilization and release of methotrexate on the surface of magnetic nanoparticles. The two temperatures were 37 and 25°, respectively. After reaction with the biomolecules, the adsorption rate for both drug molecules was about 60–80. PBS buffer was also used for diffusion of biomolecules and the results were analyzed by spectrophotometer analysis. With these results, the adsorption of cysteine and MTX was more than 60% and its release rate in MNPS-IHSPN was up to 90%, which means that high-strength stabilization and release by magnetic nanoparticles under external magnetic field and in vitro confirmed. The result of this project for the exchange of drugs by the surface of magnetic nanoparticles to repair damaged cells in the body of living organisms can be generalized.     

Otomycosis in the South of Iran with a High Prevalence of Tympanic Membrane Perforation: A Hospital-Based Study

Mycopathologia - Otomycosis is a superficial infection of the external ear caused by fungal pathogens. The genera Aspergillus and Candida are considered the main fungal causative agents, with the...     

The effect of the glycolipoprotein extract (G‐90) from earthworm Eisenia foetida on the wound healing process in alloxan‐induced diabetic rats

Diabetes is now regarded as a major public health problem. The number of patients is estimated to increase to over 439 million cases by 2030. One of the major health clinical problems in patients with diabetes patients is impaired wound healing. Diabetic foot ulcer is a major complication of diabetes mellitus in 12 to 25% of patients, which increases the risk of damage in the limbs or amputation. The earthworm Eisenia foetida glycolipoprotein (as known G‐90) is a blend of macromolecules with some biological properties including mitogenicity, anticoagulation, fibrinolysis, bacteriostatic and antioxidatiaon. Given the biological properties of G‐90, this study was conducted to investigate the effect of extract obtained from the homogenate of Eisenia foetida (G‐90) on the wound healing process in alloxan‐induced diabetic rats. The results of the present study revealed that treatment by using G‐90 can speed up the wound healing process, which is exactly similar to the effect of D‐panthenol treatment in rats. These findings also demonstrated that G‐90 treatment decreases the risk of infection in the wound site compared with D‐panthenol treatment. In addition, histological analysis indicated that a better extracellular matrix formation with increased fibroblast proliferation, neovascularization, collagen synthesis and early epithelial layer formation was observed in G‐90 treated group. Therefore, the G‐90 could be considered as a new wound healing agent introducing promising therapeutic approaches in both human and veterinary medicine. Copyright © 2016 John Wiley & Sons, Ltd.     

Recombinant Tandem Repeated Expression of S3 and SΔ3 Antimicrobial Peptides

Background:Antimicrobial peptides (AMPs) are promising candidates for new generations of antibiotics to overcome the threats of multidrug-resistant infections as well as other industrial applications. Recombinant expression of small peptides is challenging due to low expression rates and high sensitivity to proteases. However, recombinant multimeric or fusion expression of AMPs facilitates cost-effective large-scale production of AMPs. In This project, S3 and SΔ3 AMPs were expressed as fusion partners. S3 peptide is a 34 amino acid linear antimicrobial peptide derived from lipopolysaccharide (LPS) binding site of factor C of horseshoe crab hemolymph and SΔ3 is a modified variant of S3 possessing more positive charges.Methods:Two copy tandem repeat of the fusion protein (named as SΔ3S3-2mer-GS using glycine- serine linker was expressed in E. coli. BL21 (DE3). After cell disruption and solubilization of inclusion bodies, the protein was purified by Ni -NTA affinity chromatography. Antimicrobial activity and cytotoxic properties of purified SΔ3S3-2mer-GS were compared with a previously produced tetramer of S3 with the same glycine- serine linker (S3-4mer-GS) and each of monomeric blocks of S3 and SΔ3. Results:SΔ3S3-2mer-GS was successfully expressed with an expression rate of 26%. The geometric average of minimum inhibitory concentration (MIC GM) of SΔ3S3-2mer-GS was 28%, 34%, and 57% lower than SΔ3, S3-4mer-GS, and S3, respectively. SΔ3S3-2mer-GS had no toxic effect on eukaryotes human embryonic kidney cells at its MIC concentration.Conclusion:tandem repeated fusion expression strategy could be employed as an effective technique for recombinant production of AMPs.Key Words: Antimicrobial Peptide, S3, SΔ3 Fusion Expression, Tandem Repeat Expression     

Multi-spectroscopic and molecular docking studies on the interaction of neotame with calf thymus DNA

Abstract

In this paper, we have studied the in vitro binding of neotame (NTM), an artificial sweetener, with native calf thymus DNA using different methods including spectrophotometric, spectrofluorometric, competition experiment, circular dichroism (CD), and viscosimetric techniques. From the spectrophotometric studies, the binding constant (K_b) of NTM-DNA was calculated to be 2?×?10³ M^?1. The quenching of the intrinsic fluorescence of NTM in the presence of DNA at different temperatures was also used to calculate binding constants (K_b) as well as corresponding number of binding sites (n). Moreover, the obtained results indicated that the quenching mechanism involves static quenching. By comparing the competitive fluorimetric studies with Hoechst 33258, as a known groove probe, and methylene blue, as a known intercalation probe, and iodide quenching experiments it was revealed that NTM strongly binds in the grooves of the DNA helix, which was further confirmed by CD and viscosimetric studies. In addition, a molecular docking method was employed to further investigate the binding interactions between NTM and DNA, and confirm the obtained results.     

Spermine as a porcine pancreatic elastase activator: spectroscopic and molecular simulation studies

Abstract

The aim of this study was to investigate the spermine effect on the thermal denaturation, conformation and activity of elastase at three temperatures of 303, 313 and 323?K in the Tris buffer, at pH 8.5, using UV–vis spectrophotometry, spectrofluorometry and circular dichroism as well as molecular docking and molecular simulation. The increased absorption of elastase in the presence of spermine suggested a change in the environment of tryptophan. It was found that under the influence of spermine, the emission intensity of elastase extremely was reduced, and the use of the Stern-Volmer equation showed that some static quenching had occurred. The thermodynamic parameters values (enthalpy and entropy) and the molecular docking technique also revealed that van der Waals forces or hydrogen bonding interactions played an important role in the binding process. The spermine–elastase complex formation led to increasing the value of the catalytic constant (k_cat). So it could be considered as an activator. Slight changes were observed in the second structure of elastase (1.06% increase for the α-helix and 0.048% decrease the β-sheet) and the thermal stability effect. Molecular docking results also demonstrated that spermine could bind to porcine pancreatic elastase, and van der Waals forces or hydrogen bonding interactions played the major role in the binding process. Overall, our results showed that spermine could induce structural alterations in elastase, acting as a partial stabilizer and an activator for the enzyme.

Communicated by Ramaswamy H. Sarma