Functional Significance of the Upregulation of Adhesion Molecules on Irradiated Endothelial Cells
Year: 2006
Type: dissertation
Abstract: Ionizing radiation is a major therapy for cancer.Damage to the endothelium and upregulation of inflammatory responses are major side effects of radiotherapy, but the mechanisms behind these processes are not clear.A key component of the inflammatory response is leukocytes adhesion to the endothelium in a multi-step process involving leukocyte tethering to the endothelium, followed by rolling, firm adhesion/spreading and extravasation.Adhesion molecules mediate this process, particularly, Eselectin and ICAM-1, which are inducible adhesion molecules.E-selectin is involved in the initial tethering and rolling steps of the adhesion cascade while ICAM-1 appears to be involved in the latter steps.We have hypothesized that in response to irradiation, the endothelium becomes activated, increases its expression of adhesion molecules such as E-selectin and ICAM-1, and supports the increased interaction of leukocytes with the endothelium.To probe this hypothesis, we have investigated the expression of Eselectin and ICAM-1 on Human umbilical vein endothelial cells (HUVEC), human microvascular endothelial cells (HDMECs) and transformed microvascular endothelial cells (HMEC-1) at 5 hr, 24 hr, 48 hr and 72 hr post-irradiation.We have also developed an in vitro flow model to study the functional consequences of the radiation induced upregulation of adhesion molecules, primarily E-selectin and ICAM-1.Endothelial cells were grown in 35mm dishes and irradiated with a single dose of 10 Gy.HL60 (human promyelocytic leukemia) cells were perfused over the irradiated (IR) endothelial cells in a parallel plate flow chamber at shear stress of 0.5-2.0dynes/cm 2 .Flow cytometry analysis at 5 hr, 24hr, 48 hr and 72 hr post-irradiation showed upregulation of E-selectin at 5 hr post-IR only on HDMECs.ICAM-1 was upregulated at 24 hr (HDMECs), and 48 hr (HUVECs and HMEC-1).Flow assays revealed an increase in the rolling and adhesion of HL60 cells on 5 hr post-IR HDMEC.No rolling was observed on HUVECs and HMEC-1 monolayers 5 hr post-IR consistent with the absence of Eselectin from the flow cytometry data.Semi-static assays at 48 hr post-IR on HUVECs showed an increase in the number of adherent cells.There was a radiation dose dependent (5 Gy vs. 10 Gy) increase in the expression of adhesion molecules, which was significant at 72 hr post-IR.Our findings suggest that irradiation selectively impacts the functional aspects of the inflammatory response.Ph.D., for his constant guidance, encouragement, and patience during the tenure of this work.I would also like to thank my other committee members, Douglas Goetz, Ph.D., and Thomas Merchant, Ph.D., D.O., for their continued assistance throughout this study.In addition, I would like to thank Waleed Gaber, Ph.D., for performing the irradiation procedures and Felecia Waller for help with the flow cytometry data.Thanks are also due
Author: Balabhaskar Prabhakarpandian
Language: English
Subfield: Immunology and Allergy
Field: Medicine
Domain: Health Sciences
Open Access status: gold
Funder: National Institutes of Health
OpenAlex ID: W2182070087