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Recently, a subpopulation of monocytes expressing Tie2 receptors has been identified, playing an important role in tumor angiogenesis. Selective depletion of Tie2-expressing monocytes in tumor-bearing mice can inhibit tumor angiogenesis. Some of these macrophages have been shown to be located near the tumor blood vessels, forming vessels in these areas paracrinely during the angiogenesis process.

This study aims to investigate the role of putative phosphorylation sites on Tie2 receptor in tumor- associated macrophages connected to human endothelial cells.

In this study, we used a series of Tie2 mutants. After transfection of tumor-associated macrophages  with these mutants, they were evaluated for physical connection using the surface plasmon resonance technique and by non-contact co-culture of these macrophages with endothelial cells.

Mutation in the tyrosine residues 1106 and 1111 had an inhibitory effect on macrophage binding to endothelial cells, resulting in deterioration of the angiogenic activity of these cells.

Tie2 receptor and its downstream molecular pathways such as AKT/PI3 have a role in the interaction of tumor-associated macrophages with human endothelial cells, directly (via physical binding) and indirectly (through secretion of factors affecting angiogenesis). This emphasize the importance of the molecular mechanisms of Tie2 receptor activation in the interactions of endothelial cells with tumor-associated macrophages and as an anti-angiogenic therapy for cancer.

Cartilage damages, including degenerative joint diseases, are a current medical‏ ‏problem. Once damaged, adult ‎human articular cartilage has a limited capacity‏ ‏for intrinsic repair. Tissue engineering is becoming promising for ‎cartilage‏ ‏repair. Nano fibers‏ ‏have considered as suitable candidates for improving the tissue engineering‏ ‏scaffold, ‎since they can mimic microarchitecture of tissue extracellular matrix, ‎perfectly. In this research, a biodegradable ‎nanofiber scaffold of 10% silk fibroin and 5% ‎chitosan solution with a ratio of 70-30 or 50-50 respectively, made by ‎electrospinning technique ‎for articular cartilage tissue engineering. By Scanning electron microscope imaging, ‎nanofibers morphology and average ‎diameter were determined. Fourier Transform Infrared spectroscopy was ‎performed to ‎investigate the functional groups of the scaffold surface. Mechanical properties of these ‎scaffolds were ‎determined by measuring of Young´s Module, Elongation at break and Breaking ‎tenacity. PBS buffer was used for ‎in-vitro degradation assay. Scanning electron microscope images analysis showed that ‎the fibers were bead-less with ‎uniform morphology with an average diameter of 148.8 ± 52.32‎‏ ‏and 138.5± 24.51 nm respectively for 70-30 and 50-‎‎50 scaffolds. After 12 weeks‎‏ ‏assay, the percentage of degradation for 70-30 and 50-50 scaffold respectively‎‏ ‏were ‎‎6.16 ± 0.9 and 6.82±0.7% of total scaffold weight. As a result, silk fibroin-chitosan nanofiber scaffolds due to ‎containing cell adhesion domains caused by‏ ‏their protein nature and because of their slow degradation, suitable ‎physical‏ ‏structure and mechanical properties are suitable candidates for articular‏ ‏cartilage repair.‎

Sperm-mediated gene transfer (SMGT) based on the intrinsic ability of sperms to bind and take-up of exogenous DNA was introduced in 1989. From that time on, it has been a challenging topic. One of the serious challenges was the motility of transfected sperms and hence their ability to fertilize the oocyte. The present study aimed to determine the effects of media and incubation conditions and also sperm parameters on the transfection of ovine spermatozoa by foreign DNA. In this study, the effects of various incubation temperature (5, 20 and 37 ºC), three different media (PBS, TCM and DMEM), presence of FBS in medium, viability and motility of sperms and sperm capacitation in DNA absorption rate and intensity were assessed by using rhodamine-labeled EGFP plasmid. Results showed that incubation of spermatozoa with plasmid in 37ºC leads to more transfection rate but various incubation media and presence or absence of FBS had no significant effect on DNA uptake rate and intensity. Motility rate and capacitation of sperms had no significant effects too. However, in sperms with a damaged membrane, the DNA uptake rate increased significantly. All of the spermatozoa with true DNA absorption (post acrosome absorption) were immotile and none of the examined treatments in this study could produce motile transfected spermatozoa. Regarding the results of this study, it seems that membrane-damaged spermatozoa incubated with foreign DNA can be used for SMGT-ICSI to produce transgenic animals.

Introduction A simple and efficient method for producing multi-transgenic animals is required for medical and veterinary applications. The principal technique for the production of transgenic animals is pronuclear microinjection, which has a low efficiency for the generation of transgenic farm animals expressing a single transgene. Recently, nuclear transfer has been used to clone large animals, and could allow multiple genetic manipulations to be undertaken in vitro, prior to a single nuclear transfer, rather than complex and time consuming breeding programs. However, at present the frequency of success in cloning large animals is very low and is very expensive. The production of transgenic livestock by sperm mediated gene transfer (SMGT) has a number of advantages compared to other transgenic techniques such as nuclear transfer and microinjection. Both nuclear transfer and microinjection techniques are technically demanding and labor intensive. In contrast, SMGT requires no sophisticated equipment or technical expertise. Furthermore, bovine genetics are distributed through sperm in the dairy industry consequently making it easy to distribute genetically modified sperm. SMGT is based on the ability of sperm to bind to exogenous DNA molecules and transfer it to the oocyte during fertilization. The major benefits of the SMGT technique were found to be its high efficiency, low cost and ease of use compared to other methods. SMGT was first described in a small animal model, with high efficiency reported in the mouse. Recently the technique has been successfully adapted and optimized for use in large animals. Studies have shown that spermatozoa from numerous species, including bovine, can bind and take up foreign DNA and transfer it to the embryo. In bovine studies, the efficiency of SMGT can vary widely depending on both the transgene and the gene transfer method. Liposomes have been shown to be particularly effective in transferring DNA into bovine sperm. However, not all embryos derived from transfected sperm contain the transgene, suggesting that mechanisms exist, which impede SMGT. The aim of this study was to investigate the possibility of gene transfer to bull spermatozoa by using lipofection.
Materials and Methods Ram testes collected from Meysam abattoir slaughterhouse immediately after slaughter and were brought to the laboratory in an ice chest. In the laboratory, the testes were rinsed twice with normal saline and were then trimmed to remove the extra testicular tissue and washed properly with saline containing 0.1% streptomycin sulphate. Connective tissue covering the cauda epididymis was removed by careful dissection, with care to avoid rupturing blood vessels or the epididymal duct. For detection of transfected spermatozoa, they stained with Rodamine. In order to transfection of sperm, 2 μg of Rodamine labeld DNA and 0.5 μl of TurboFect were diluted in 25 μl of transfection medium separately, and incubated for 5 min at room temperature. Then, the diluted DNA was added to diluted TurboFect (total volume=50 μl) and incubated for 20 min at room temperature. 1×106 sperm were added to 50 μl of DNA- TurboFect complexes and mixed gently by rocking the plate back and forth. To evaluation of transfected spermatozoa motility, acridine orange staining was used. Each experiment was replicated at least three times, and for each replicate, at least 50 ES cell colonies were used. Data were analysed with a statistical software program (SPSS 16). Comparisons between two treatments and multiple numeric datasets were performed using t-test and one-way ANOVA followed by Duncan multiple-range test, respectively. Results are expressed as mean±SEM and statistical significance was accepted at P0.05). The comparison of transfected and normal spermatozoa reveal that, motility of transfected spermatozoa at 60 minutes after transfection was significantly lower than normal ones (p

Spermatogonial Stem Cells (SSCs) are the only stem cells in adults that can transfer genetic information to future generations. Considering that a single SSC gives rise to a vast number of spermatozoa, genetic manipulation of these cells is a potential novel technology with practical application to various animal species.
The aim of this study was to evaluate Enhanced Green Fluorescent Protein (EGFP) gene transfection into bovine spermatogonial colonies via liposome carrier and assess the best incubation day in uptake exogenous gene by spermatogonial colonies.
Transfection efficiency EGFP gene through lipofection was determined different in three days (day 4, 6 and 8) after the beginning of the culture by fluorescent microscope. Immunofluorescent staining against OCT4 and vimentin led to the confirmation of the nature of both SSCs and sertoli cells.
Results showed that the transfected colonies through lipofection increased significantly (p It was concluded that lipofectamine can be used safely for direct loading of exogenous DNA to spermatogonila colony, particularly during the fourth day of culture.

As we all know, sperm has the capacity to take up foreign DNA, therefore, sperm mediated gen transfer can be an inexpensive and simple method in animal transgenesis in various species. However, there is not sufficient evidence of DNA uptake by ovine spermatozoa. The purpose of the present study was to examine the uptake of human lysozyme gene contained plasmid (pEGFP-IRES-hLys) by ovin spermatozoa. In the first experiment, semen was prepared from three ram (each ram two times) by electrical method. After removal of seminal plasma, 1x106 spermatozoa were incubated with rhodamin-labled pEGFP-IRES-hLys in TCM199 for 15, 30, 60 and 120 minutes and then observed for motility, uptake percent and uptake intensity by florescent microscopy. Also after 60 minutes incubation sperms were treated by DNaseI to assay adsorption or uptake of pEGFP-IRES-hLys. In the second experiment, washed and unwashed sperms were incubated with rhodamin-labled pEGFP-IRES-hLys in TCM199 for 30 and 60 minutes to evaluate the effect of presence seminal plasma on sperm uptake and motility. The findings showed that increasing incubation time increased number/percentage of spermatozoa carrying exogenous DNA and its intensity. But this different was significant only up to 30 minutes. We found that 60.16% of the cells were bound to DNA after 120 minutes incubation. Incubation with exogenous DNA induced a slightly decrease in sperm total and progressive motility. But no post acrosom uptaked sperm was motile. After treatment with DNaseI, strong florescent emission from post acrosom indicated absorption of pEGFP-IRES-hLys by spermatozoa. Presence of seminal plasma induced a slightly decrease in percent of DNA absorbed spermatozoa and absorption intensity, but did not inhibit completely. Ram spermatozoa showed a high capacity to bind DNA quickly and reach a maximum after 30 min. However, no sperm with real uptake (post acrosomal) was motile. Incubation with lower DNA concentration and/or shorter time may be helpful.