یدالله امیدی

Cytokine therapy is a fundamental step for the control of the most severe inflammatory diseases. Considering the importance of TNF-? in most of autoimmune and inflammatory diseases، it has been as an attractive target for many researchers and is an important candidate for cytokine therapy. Using monoclonal antibodies is a major approach in contrast to deleterious effects of TNF-. Antibody phage display technology is a major technique for the preparation of monoclonal antibodies. Our goal in current study was production and preparation of monoclonal antibody against TNF-. For Biopanning technique، we used ELISA plates for isolation of single chain variable fragments (scFv) from Tomlinson library. After coating of TNF-? protein in the wells، phage library was added. Specific phages were eluted using Trypsin after washing process and amplified in TG1 cells. Five rounds of panning were conducted for obtaining high specific phages. The evaluation of selected clones was done through monoclonal phage ELISA، PCR، RFLP and western blotting methods. Analysis of sequences using bioinformatic softwares showed that separated plasmids from TNF-? reactive clones contained insertions. Furthermore، most of variations are seen in three hot regions which can affect protein function. The results of this study showed that forth generation of monoclonal antibodies against human TNF-? were successfully produced.

Brevinin-2R، consisting of 25 amino acid residues، from the skin of the frog Rana ridibunda has potent antimicrobial and low hemolytic activity. From a structural point of view، this peptide has an N-terminal hydrophilic region، and a C-terminal loop region delineated by an intra-disulfide bridge، which is a common structural feature of antimicrobial peptides from Rana species. To investigate the structural features for antimicrobial، hemolytic activity and proteolytic stability، analogues diastereomer Sami-D brevinin-2R were synthesized and characterized. Structure-activity relationship studies revealed the importance of the amphipathic α-helical conformation of the reported peptides in inducing antimicrobial effects. Analogues D shows lower antimicrobial activity than brevinin-2R and did not show any hemolytic activity up 400 µg/ml whereas the brevinin-2R show low hemolytic activity. Circular dichroism spectra and the retention time on the C18 reverse phase column revealed that the brevinin-2R formed an amphipathic loop which increased hydrophobicity and helped to induce the α-helical structure in the membrane-mimetic environment. The hemolytic activity of brevinin-2R and its analogs also correlated well with the retention time and the α-helicity.

Nanotechnology will affect human being life impressively over the next decade in different fields, including medicine and pharmacy. Polymeric nanoparticles have been far and wide studied as particulate carriers in the pharmaceutical and medical fields since they show promise as drug delivery systems on account of their controlled and sustained- release properties, sub cellular size, biocompatibility with tissue and cells and enhancing the effectiveness of the loaded drugs. Numerous methods have been developed during the last two decades to formulate the pharmaceutical nanoparticles. These methods have been classified according to whether the particle formation implies a polymerization reaction or arises from a macromolecule or preformed polymer. In the current review the most important methods of preparation are explicated, more than ever those that make use of preformed synthetic polymers. Furthermore, the methods which can be commercialized as well as pharmaceutical aspects are discussed briefly. Pharmaceutical nanoparticles can be prepared using different methods depending on the physicochemical properties of the drug and polymers.

ECM33 gene encodes a protein which belongs to a large group of GPI anchored protein family. Although the precise molecular function of this protein is unknown, several studies have demonstrated an important role in fungal cell wall integrity and virulence. The aim of this study was to identify and clone ECM33 gene in Aspergillus niger. Also the expressional analysis of the gene was carried out. The complete genomic sequence of ECM33 gene along with 5' and 3' flanking regions has been identified in Aspergillus niger by means of bioinformatics and using ECM33 gene of A. fumigatus as template. The genomic fragment was PCR-amplified using specific primers and subsequently was cloned. The expression analysis was performed by RT-PCR. The Aspergillus niger ECM33 gene containing 1327 bp was cloned as a 5.2 kb fragment. Bioinformatic analysis predicted the presence of two introns which was confirmed experimentally. The gene product is a 397 amino acids protein with approximately 70% identity to the homologue proteins from other aspergilli. Expression analysis of ECM33 under different growth conditions including different temperatures, different carbon sources, and differentnitrogen sources confirmed a constitutive expression. This is the first report on characterization of ECM33 gene in an industrially important fungus, Aspergillus niger. The cloned fragment will be used in construction of gene disruption cassette which will be subsequently applied in gene knock out experiments.

Gene toxicology (particularly genocompatibility and/or toxicogenomics) has begun to be acknowledged as an important scientific discipline in drug delivery and targeting, in which the intrinsic genomic signature of chemicals and pharmaceuticals is going to be underlined in the heredity of living organisms for a better implementation of such pharmaceuticals. Viral and non-viral vectors are the two paradigms for transport of genome-based pharmaceuticals in vitro and in vivo. However, genomic impact(s) of these delivery systems in target cells/tissues are still in shed of their transfection potential. Despite substantial information on their cellular influence, little information is available about the genocompatibility and/or toxicogenomics of the non-viral vectors including cationic lipid (CL) delivery systems. In the current study, we investigated the genocompatibility of cationic lipid, Oligofectamine (OF) in human alvelolar epithelial A549 cell line. To screen the genomic impact(s) of these delivery systems, cytotoxicity assay using MTT and DNA microarray technology was utilized. RNA samples from treated (routinely used concentrations of CL) and untreated cells were converted to aminoallyl-cDNA, labeled with cyanine (Cy3/Cy5) and hybridized on target arrays housing 200 gene spots. Slid arrays were scanned, data was normalized and up- and down-regulated genes and their ontologies were detected. MTT assay showed cytotoxicity within A549 cells treated with the nanoliposomes. The gene expression profiles revealed marked changes (≥2-fold) in gene expression for 8-15% of genes from various genomic ontologies induced by OF. Among altered genes, some (e.g., il9r, ces111 and tnfsf6) were related to the apoptosis pathway; nevertheless the altered genes appeared to be within various gene ontologies. Our findings highlight pervasiveness of intrinsic genomic impacts by cationic nanoliposomes. Such impacts may interfere with the main goals of these delivery systems by masking/stimulating a cluster of non-specific genes that may affect the end point genotype and/or phenotype, thus we suggest genomic assessments for cationic liposome gene delivery systems.