مجله پژوهش های سلولی مولکولی (زیست شناسی ایران)
سال سی و پنجم شماره 4 (زمستان 1401)

Pseudomonas aeruginosa is a prevalent bacteria in nosocomial infections that are resistant to various types of antibiotics by several mechanisms. Nowadays, there utilizes drug delivery methods such as liposomes for boosting the efficiency of treatment. In this study, pipractam, were encapsulated in a variety of liposomal formulations and evaluated for their efficacy by MIC assay. Different liposomes of which incl. neutral, cationic and anionic types were prepared by solvent phase evaporation and their biophysical properties including shape, size and charge were evaluated using electron microscope, DLS and zeta-sizer respectively. The antimicrobial effect of liposomal formulations and drug solution was investigated on two clinical isolates and a standard strain of pseudomonas aeruginosa by serial dilution broth and the rate of encapsulation was measured by HPLC technique.The spherical liposome is shown by electron microscope. .The HPLC technique evaluated the encapsulation of neutral, cationic and anionic liposomes by 5.26%, 8.31% and 6.37%, respectively. Results of the lowest inhibitory concentration on the standard strain and two isolates 62 and 60 was evaluated for free drug 40, 8, 8; neutral liposomes 4, 32, 16; cationic liposomes 4, 8, 32 and anionic liposomes 1, 2 and 1 μg / ml. Solvent phase evaporation technique made liposomal particles that had small dimensions and a suitable encapsulation percentage. By Comparison of minimum concentration inhibition of different formulations is revealed pipractam anionic liposome was the best formulation that could reduce four folds MIC of pipracatam than one’s free form on pseudomonas aeruginosa in this study

Site-directed mutagenesis is one of the most effective methods for enhancing the activity and stability of industrial enzymes. In this study, the influence of Alanine 56 substitution to isoleucine on the thermostability of a metalloprotease produced by a moderately halophilic bacterium named Salinivibrio Proteolyticus (SVP) Strain AF-2004 was investigated. The given mutation was constructed using the mutagenic primers and DpnI restriction enzyme treatment. The resulted mutant gene in the pQE-80L vector was transformed into the various expression strains of Escherichia coli (XL1-Blue، BL21 (DE3) pLysS, and Top10). Then, the enzyme expression was optimized using different cultural conditions. Subsequently, the wild and mutant enzymes were extracted from zymography gel. Finally, the thermostability and activity of wild and mutant enzymes were assayed. The maximum expression of the enzyme was achieved using E.coli BL21 (DE3) pLysS, in Terrific broth medium, in the presence of 1 mM IPTG, and after 24 h induction at 37 ºC. The extracted wild and mutant enzymes lost 42 and 24 percent of their activity after incubation at 70 ºC for 20 min, respectively. However, the activity of the mutant enzyme was 11 percent more than the wild enzyme. Therefore, the activity and thermostability of the mutant SVP metalloprotease (A56I) were improved compared to the wild enzyme.

Proteomics is a new technique in plant-insect interaction studies. Identification of effective proteins in the insect- plant systems is main challenge in the evolutionary ecology. In this study, accumulated plant proteins in the gut of Aelia acuminate were visualized using two dimensional electrophoresis. Targets spots were selected to cut, digest and blast in the mascot software using the National Center of Biotechnology Information database, respectively. Results showed that accumulated proteins were serpin, alpha- amylase inhibitors as plant inhibitors; peroxidase, dehydroascorbate reductase and deoxymuogenic acid synthase as antioxidant proteins; beta-amylase and agglutinin lecithin 3 as stored proteins; calmodulin as regulation protein and hypothetical protein as unknown protein. Accumulation of plant inhibitor proteins in the gut of adults proved that gut proteases of insect can be overcome to these toxic proteins. Results suggest that proteomics is effective technique to identify changed proteins in the insect and plants tissues and it will be helpful to discuss their physiological roles in the biochemical process.

Identification of genetic loci controlling quantitative trait is an important subject in genetics and breeding programs, particularly under stress conditions. Broomrape is a holophrastic weed in tobacco (Nicotiana tabacum L.) fields with devastating effects on its dry leaf yield production. In the present study, the reaction of 89 tobacco genotypes was evaluated against broomrape (Orobanche aegyptiaca) in randomized complete block design with three replications during two successive years in pot conditions. In each year, the genotypes were planted separately in non-inoculated and inoculated conditions. In broomrape conditions the soil was mixed with 0.06 g of broomrape seed. In the molecular experiment, 26 SSR primers pairs was used for preparing the molecular profile of genotypes. Studying population structure as a prerequisite for association analysis with bayesian method revealed that there are two possible subgroups (K=2) in the studied population. Based on D′ statistics, 4.31% of possible SSR locus pairs showed significant level of linkage disequilibrium (P

The most common method for peptide synthesis is to synthetize it on a solid resin. Among the variety of resins, Wang resin is one of the highly use ones. In this study, we introduced improved protocols for anchoring an amino acid on the Wang resin, a famous resin for peptide synthesis, and N-terminally labeling peptide with (5/6)-carboxyfluorescein, a fluorescent dye. The peptide tetra arginine, RRRR, was synthetized on Wang resin with solid phase peptide synthesis (SPPS) method and the dye was attached to its N-terminus. The attachment yield of the first arginine residue (Fmoc-Arg(pbf)-OH) were quantified by liberating the Fmoc groups from the amino acid/resin complex following measure its UV absorption at 278 nm. The labeling was monitored by Ninhydrin test that reveals presence of primary, unbound N-terminally amines. In anchoring the first arginine residue on the resin, DIC (N,N’-Diisopropyl carbodiimide) works more efficiently than HBTU (N,N,N′,N′-Tetramethyl-O-(1H-benzotriazol-1-yl)uronium hexafluorophosphate), only if the incubation time increases to 17 hours; while, HBTU undeniably enhances the efficiency of the fluorescence labeling compared to DIC. However, in most available protocols, DIC was recommended for labeling the peptides with (5/6)-carboxyfluorescein and the recommended incubation time for attaching the first amino acid on the Wang resin is usually far shorter than 17 hours.

Iranian drug CinnoVex (Interferon beta) is an important drug for preventing the progress of M.S. disease and increase of its stability is very important. In this research, we designed five mutations in the structure of interferon and performed molecular dynamics simulation and docking and the best stabilizer mutation was determined. At first, the wild type structure of beta interferon was subjected to Pop music server for prediction of stabilizer mutations. Then the first four proper score mutations were selected and mutated structures and a mix mutation (mut 2+ mut 3) (mut 5) were made. Then, molecular dynamics simulation performed via Gromacs 5 package on wild and mutated structures in five temperatures (300, 350, 400, 450, 500 K) separately for 20 ns (in total 600 ns). The simulation parameters were calculated in each temperature for all structures during the last 10 ns of simulations. The plots of temperature against averages of parameters were drawn and the regression slope of parameters was calculated to find the more stable structure. The results revealed that mutation 3 (G127E) is the best mutation compare to wild interferon. Also, the binding power of mutation 3 with its receptor is more than wild type and other mutations. It seems that the increase of hydrophilicity of protein around position 127 in mutation 3 on interferon surface leads to increase the stability and solubility of it ex vivo.

The majority of neurogenesis is terminated soon after birth and in adults new neurons generated only in the sub ventricular zone and the hippocampus accordingly neurodegenerative disorders and nerve damage are vital problem. Although long non-coding RNAs (lncRNAs) are involved in several cellular process, their function in the regulation of neurogenesis should be further clarified. In this regards, this study aim to analyze expression levels of LncRNA RMST during neuronal differentiation of human pluripotent embryonal carcinoma NT2 cells. In this study, human pluripotent embryonal carcinoma NT2 cells were neurally differentiated though retinoic acids and a cocktail of neural. The expression levels of neural markers were analyzed by RT-PCR. During neural differentiation of NT2 cells, the expression levels of neural specific markers NSE and MAP2 were observed (***P

Nanoscale science and technology the production of matter in nanometer dimensions and exploitation of its properties in modern systems. Today, one of the most efficient methods of nanoparticle synthesis is the method of green synthesis or biosynthesis of nanoparticles by plants. The purpose of this study was to synthesize and develop a green method for the preparation of iron nanoparticles. In this study, iron nanoparticles were synthesized in a process by the extract of Russian knapweed (Acroptilon repens L.) which containing phenolic compounds. The extract of the plant was obtained by maceration method. The co-precipitation method was used to prepare iron nanoparticles with the extract of Acroptilon repens L. The samples were analyzed by Thermo Gravimetric Analysis (TGA), Fourier transform infrared (FT-IR), Vibrating Sample Manetometer (VSM), Scanning Electron Microscope (SEM) and X-Ray Diffraction (XRD). In Spectrophotometric analysis, the presence of a peak in the range of 490 to 600 nm indicates the biological synthesis of these nanoparticles with extract of Acroptilon repens. The size and morphology of biologically synthesized nanoparticles were determined by scanning electron microscopy, and it was found that the shape of the polygonal and round particles and their average size are about 39 to 50 nanometers. The results of X-ray diffraction analysis also showed the nanocrystals synthesized extract of Acroptilon repens. According to the results, it can be said that the polyphenolic compounds in the Acroptilon repens. extract were acted as iron-metal reducing agents, and also as a complexing agent synthesize iron nanoparticles and make them sustainable

Lung cancer is one of the most common types of cancer in the world and Iran. Herbs are a source of a variety of antioxidant compounds that can be used to produce anti-cancer drugs. In the present study, the effect of cytotoxicity of essential oil and hydroalcoholic extract of Juniperus excelsa on A549 lung cancer cells was investigated. The branches of male rootstocks of Aras plant were collected from Chehel Cheshmeh in Firoozkooh and Sirachal in Karaj and the essential oil and extract were prepared by Clevenger and soaking methods, respectively. The degree of cytotoxicity, different concentrations of essential oil and extract of the two regions, on A549 cells in three times of 24, 48, and 72 hours Was evaluated using MTT method. Flow cytometry was used to evaluate cell death and cell cycle in cells treated for 72 hours with 10 µg/ml of essential oil and extract of two regions. The results of cytotoxicity showed that the lethal power of extracts was significantly higher than essential oils, but there was no significant difference between different regions (P <0.01). Also, a further decrease in the number of viable cells and an increase in cell entry into the Sub G1 phase, resulting in an increase in apoptosis treated with male root extract, was proven. Thus, the extract of the branches of this plant has anti-cancer properties by stopping the cell cycle in the G2 / M stage.

Peptides Aβ(1-40) and Aβ(1-42) are effective peptides in Alzheimer's disease. With the aggregation of these peptides outside the nerve cell, disease occurs. Drugs that reduce this accumulation prevent the progression of Alzheimer's. In this regard, herbal medicines are receiving more attention due to their less side effects. Due to its antioxidant properties, boswellia is effective in reducing the progression of neurological diseases. In this work, effects of Acetyl-11-keto-β-boswellic acid derivatives on the monomer and dimer structure of Aβ (1-40) and Aβ (1-42) peptides are monitored by using molecular dynamics simulation. The results showed that in the presence of ligand, the flexibility of the C-terminal residues of Aβ (1-40) is higher. The ligand also reduces the amount of peptide sheets. By calculating the conformation factor, the afinity of the ligand to interact with each of the peptide residues was calculated. The results of this calculation showed that the residues that tend to be binded have less flexibility. By calculating the free binding energy, the binding site of the ligands was determined.

Nanoparticles (NPs) have broad applications in various industries, that it can be expected them find a way to enter into the environment and affected the living organism with unexpected consequences. According to the broad applications of the Fe2O3 nanoparticles (nFe2O3) in different industries, in this study, its interaction with human serum albumen (HSA), as an abundant protein in blood, was investigated using intrinsic fluorescence at different temperature ranges, extrinsic fluorescence and circular dichroism (CD) methods. Results showed, with increasing NP concentration in media, intrinsic fluorescence intensity of the protein decreased in all temperatures. Based on the quenching rate constant (Kq), it revealed that interaction between HAS and nFe2O3 take place through static mechanism. Thermodynamic parameters indicate the sign of enthalpy and entropy are negative, which implies the role of hydrogen bands and Van der Waals forces in this interaction. The negative sign of free energy (∆G°), revealed this reaction is exergonic and spontaneously. On the other hand, ANS fluorescence intensity increased more for HSA in interaction with NP in contrast to the ANS. Furthermore, Far-UV CD spectra shows an alteration in the secondary structure of this protein in interaction with the NP. The results all together revealed changes in the structure of HSA upon exposed to this nanoparticle which can affected its performance.

The root endophytic fungus Serendipita indica has useful and unique features to enhance growth, yield and resistance of plants against biotic and abiotic stresses. The recent studies have shown that plant growth, physiology and production of phytochemicals are significantly affected by nanoparticles. The purpose of this study was to investigate the effects of S. indica and zinc oxide nanoparticles on the symbiosis amount, growth parameters and the production of some phytochemicals in pharmaceutical plant Physalis alkekengi under in vitro conditions. The treatments were two fungus level (presence of fungus and absence of fungus), and five concentrations of zinc oxide nanoparticles (0, 5, 10, 15 and 20 mg/L). By increasing zinc nanoparticles in media, symbiosis percentage and the growth parameters were significantly increased. Application of nanoparticle in some levels, increased total flavonoid, total phenol, anthocyanins and antioxidant capacity of leaf and stem, in compare to control. Presence of S. indica significantly increased growth parameters, stem flavonoids, leaf and stem anthocyanins, leaf total phenol and antioxidant capacity of the stem and leaf. The results from this study showed that zinc nanoparticles, especially at 20 mg/L, as abiotic elicitor and S. indica as biotic elicitor can increase the growth and production of some phytochemicals in pharmaceutical plant Physalis alkekengi.