Showing 3 results for Endothelial Cells
Hesari M, Mansouri K, Mostafaie A, Bidmeshkipour A,
Volume 68, Issue 3 (6-2010)
Abstract
Background: Proteolytic enzymes, especially collagenases, are used for digestion of
extracellular matrix, cell isolation and primary culture. Because of the problems in purification and low amount of collagenases in bacterial or animal sources, it is important to find new sources of the enzymes. So, in the present study actinidin, a plentiful protein in kiwifruit was purified and a mixture of actinidin and trypsin was applied to isolate rat aortic endothelial cells.
Methods: Aortic endothelial cells were isolated using digestion solution containing different concentrations of actinidin (from 2 to 16 mg/ml) and trypsin (0.3, 0.6, 1.2 and 2.4 mg/ml) in different times (from 15 to 90 minute). Isolated cells were cultured in
DMEM culture medium. Isolated cells were identified by morphological characteristics and immunocytochemical staining viability of separated cells was estimated by trypan blue exclusion test.
Results: Actinidin in concentration of 10 mg/ml with trypsin in concentration of 1.2 mg/ml for one hour could isolate rat aortic endothelial cells. In this condition the viability of cells was estimated 90%. Morphological and immunocytochemical charac- teristics confirmed the isolated cells as endothelial cells.
Conclusion: The results showed that the mentioned mixture of actinidin and trypsin has not considerable toxic effects on separated cells and is a novel and suitable option for isolation of rat aortic endothelial cells
Gholami K, Nejati V, Delirezh N, Ganji Bakhsh M, Asadi M,
Volume 69, Issue 3 (6-2011)
Abstract
Background: The innate and adaptive immune responses are dependent on the
migration of leukocytes across endothelial cells. Dendritic cells (DCs) play an important role in the initiation of cellular immune responses during their migration from tissues into the lymph nodes where they interact with endothelial cells of lymphatic vessels. We investigated the effects of surface-adherent and non-activated endothelial cells on phenotypic and functional characteristics of dendritic cells. Methods: Immature dendritic cells were generated from the isolation of peripheral blood mononuclear cells and their subsequent culture in DC-RPMI 1640 medium containing 10% FCS, interleukin-4 and granulocyte-macrophage colony-stimulating factor (GM-CSF) for five days. On day five, a maturation factor (composed of monocyte-conditioned medium, tumor necrosis factor-α (TNF-α) and poly I:C) was added to the RPMI medium where immature DCs were co-cultured with endothelial cell monolayer for 24 h. The maturation of harvested DCs on day seven was evaluated via flow cytometry, a beta-counter and an ELISA kit.
Results: This study showed that the endothelial cells interact with dendritic cells generated from peripheral blood monocytes via cell-to-cell interaction. This interaction inhibits the maturation of DCs via decrease in the expression of CD83, CD86, CD80, HLA-DR and up-regulation of CD14. The interaction also inhibits the stimulation of T-lymphocytes resulting in a decrease in their proliferation.
Conclusion: According to the findings of this study, it could be concluded that the endothelial cells can act as a potent regulator for DCs differentiation and function at the encounter made between them during the migration of DCs from tissues to lymph
nodes.
Adeleh Poursaleh, Mohammad Najafi , Farahnaz Sadegh Beigee ,
Volume 77, Issue 3 (6-2019)
Abstract
Background: The immune-mediated responses in vascular cells may include the increased expression of endothelial adhesion molecules, leukocyte rolling and infiltration, cellular lipid dysregulation and vascular smooth muscle cells (VSMCs) differentiation. Investigating the cellular and molecular events involved in the rolling process is useful for treatment or prevention of the vessel stenosis especially in coronary arteries. MiRNAs are small and single-stranded noncoding RNAs with about 19-23 nucleotides. In this study, the role of microRNA-125 was predictably selected and experimentally investigated on the changes of expression level of adhesion molecule in endothelial cells isolated from human aorta and on the monocyte cells isolated from whole blood human with endothelial cells adhesion. The aim of this study was to determine the effect of miRNA-125 repression on cell adhesion in leukocyte rolling process to reduce or suppress artery stenosis in susceptible individuals.
Methods: This experimental study was performed in Cellular-Molecular Research Center of Iran University of Medical Sciences, Tehran, Iran from July to December 2017. Normal aortic samples were prepared from subjects with brain death in Masih Daneshvari Hospital and under strictly sterile conditions, it was transferred as soon as possible. The endothelial cells were isolated from aorta of subjects with brain death using collagenase. The monocytes were isolated from whole blood. The microRNA-125 was transfected into ECs with use of polyethyleneimine (PEI). The expression level of adhesion molecule and monocyte recruitment were identified by quantitative reverse transcription polymerase chain reaction (qRT-PCR) technique and CytoSelect™ leukocyte-endothelium adhesion assay kit (Cell Biolabs, San Diego, CA, USA), respectively.
Results: The results showed the microRNA-125 suppresses significantly integrin beta 2 (ITGB2) expression level (P=0.008). In addition, the monocyte-EC adhesion was shown in the aortic miRNA-treated endothelial cells. The adhesive rate between cells reduced significantly with microRNA-125 as compared with miR-synthetic (P=0.02). Thereby, there were the associations between the ITGB2 and miR-125a. Downregulation of ITGB2 may be reduced the adhesion of endothelial cells and moderating the process rolling.
Conclusion: This study suggested that the suppression of leukocyte rolling process might be more due to the function of ITGB2. However, the functional effects of this miRNA should be directly investigated on the studied gene.
