Role of heparin in the activation and migration of primary human umbilical vein endothelial cells (HUVECs) following signal transduction by CD44 and its ligand hyaluronic acid

Daphne Vincent Santhosh, Muddanna S. Rao

Research output: Contribution to journalArticle

Abstract

Background: CD44 is an extracellular matrix molecule that has hyaluronan as one of its principal ligands. The interaction between CD44 and its ligand initiates activation and migration of endothelial cells. Migration of endothelial cells holds importance in neovascularization during cancer metastasis where new blood vessels are formed in order to nurture metastasized cancerous tissue. Studies on bovine endothelial cells showed that greater degree of migrations occurred with addition of lower molecular weight hyaluronic acid fragments (HAF) than the whole molecule of hyaluronan (HA). Our study included addition of heparin to the seedings of HA and HAF at concentrations of 10μg/ml, 100μg/ml, 250μg/ml and 500μg/ml to endothelial cells and measuring the degrees of migrations obtained on migration assay. Methods: HUVECs were isolated from umbilical cords obtained from Kasturba Medical College, Manipal, India. The cells were standardized to 2000 cells/ml and cultured in co-star well plates and were seeded with HA and HAF at concentrations of 10μg/ml, 100μg/ml, 250μg/ml and 500μg/ml. Basal level of control were resting cells and PMA treated cells were positive control. Migration assay was done on which the endothelial cells were embedded. After treatments, the migrations were measured and compared. Expression of CD44 was done using SDS-PAGE and Western Blot analysis. Results: Heparin seems to play a role in the activation and migration of endothelial cells and their sustenance and viability in the medium. In Heparin M199 medium, the endothelial cells showed greater migrations when seeded with HAF than with HA. At 100μg/ml heparinised endothelial cells showed greater degree of migration with HAF than HA added at the same 100μg/ml. In heparin-free medium, the primary HUVECs migrated more at 10μg/ml for cells seeded with HA while they migrated more with HAF at a concentration of 250μg/ml. SDS-PAGE and western blot analysis showed CD44 molecules on the primary HUVECs. Conclusion: The assays were set up and studied in duplicate. Further analysis should be done in order to substantiate the role of heparin in the migration of primary human umbilical vein endothelial cells following signaling with CD44 and HA. This might hold promise for therapeutic research into cancer metastasis and neovascularisation.

Original languageEnglish
Pages (from-to)309-316
Number of pages8
JournalJournal of Research in Medical Sciences
Volume13
Issue number6
Publication statusPublished - 01-12-2008
Externally publishedYes

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Human Umbilical Vein Endothelial Cells
Hyaluronic Acid
Heparin
Signal Transduction
Ligands
Endothelial Cells
Polyacrylamide Gel Electrophoresis
Western Blotting
Neoplasm Metastasis
Therapeutic Human Experimentation
Umbilical Cord
Extracellular Matrix
Blood Vessels
India
Cultured Cells
Neoplasms

All Science Journal Classification (ASJC) codes

  • Medicine(all)

Cite this

@article{6cc8ea165f1a4a0198ccb2b61e5c67a8,
title = "Role of heparin in the activation and migration of primary human umbilical vein endothelial cells (HUVECs) following signal transduction by CD44 and its ligand hyaluronic acid",
abstract = "Background: CD44 is an extracellular matrix molecule that has hyaluronan as one of its principal ligands. The interaction between CD44 and its ligand initiates activation and migration of endothelial cells. Migration of endothelial cells holds importance in neovascularization during cancer metastasis where new blood vessels are formed in order to nurture metastasized cancerous tissue. Studies on bovine endothelial cells showed that greater degree of migrations occurred with addition of lower molecular weight hyaluronic acid fragments (HAF) than the whole molecule of hyaluronan (HA). Our study included addition of heparin to the seedings of HA and HAF at concentrations of 10μg/ml, 100μg/ml, 250μg/ml and 500μg/ml to endothelial cells and measuring the degrees of migrations obtained on migration assay. Methods: HUVECs were isolated from umbilical cords obtained from Kasturba Medical College, Manipal, India. The cells were standardized to 2000 cells/ml and cultured in co-star well plates and were seeded with HA and HAF at concentrations of 10μg/ml, 100μg/ml, 250μg/ml and 500μg/ml. Basal level of control were resting cells and PMA treated cells were positive control. Migration assay was done on which the endothelial cells were embedded. After treatments, the migrations were measured and compared. Expression of CD44 was done using SDS-PAGE and Western Blot analysis. Results: Heparin seems to play a role in the activation and migration of endothelial cells and their sustenance and viability in the medium. In Heparin M199 medium, the endothelial cells showed greater migrations when seeded with HAF than with HA. At 100μg/ml heparinised endothelial cells showed greater degree of migration with HAF than HA added at the same 100μg/ml. In heparin-free medium, the primary HUVECs migrated more at 10μg/ml for cells seeded with HA while they migrated more with HAF at a concentration of 250μg/ml. SDS-PAGE and western blot analysis showed CD44 molecules on the primary HUVECs. Conclusion: The assays were set up and studied in duplicate. Further analysis should be done in order to substantiate the role of heparin in the migration of primary human umbilical vein endothelial cells following signaling with CD44 and HA. This might hold promise for therapeutic research into cancer metastasis and neovascularisation.",
author = "Santhosh, {Daphne Vincent} and Rao, {Muddanna S.}",
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T1 - Role of heparin in the activation and migration of primary human umbilical vein endothelial cells (HUVECs) following signal transduction by CD44 and its ligand hyaluronic acid

AU - Santhosh, Daphne Vincent

AU - Rao, Muddanna S.

PY - 2008/12/1

Y1 - 2008/12/1

N2 - Background: CD44 is an extracellular matrix molecule that has hyaluronan as one of its principal ligands. The interaction between CD44 and its ligand initiates activation and migration of endothelial cells. Migration of endothelial cells holds importance in neovascularization during cancer metastasis where new blood vessels are formed in order to nurture metastasized cancerous tissue. Studies on bovine endothelial cells showed that greater degree of migrations occurred with addition of lower molecular weight hyaluronic acid fragments (HAF) than the whole molecule of hyaluronan (HA). Our study included addition of heparin to the seedings of HA and HAF at concentrations of 10μg/ml, 100μg/ml, 250μg/ml and 500μg/ml to endothelial cells and measuring the degrees of migrations obtained on migration assay. Methods: HUVECs were isolated from umbilical cords obtained from Kasturba Medical College, Manipal, India. The cells were standardized to 2000 cells/ml and cultured in co-star well plates and were seeded with HA and HAF at concentrations of 10μg/ml, 100μg/ml, 250μg/ml and 500μg/ml. Basal level of control were resting cells and PMA treated cells were positive control. Migration assay was done on which the endothelial cells were embedded. After treatments, the migrations were measured and compared. Expression of CD44 was done using SDS-PAGE and Western Blot analysis. Results: Heparin seems to play a role in the activation and migration of endothelial cells and their sustenance and viability in the medium. In Heparin M199 medium, the endothelial cells showed greater migrations when seeded with HAF than with HA. At 100μg/ml heparinised endothelial cells showed greater degree of migration with HAF than HA added at the same 100μg/ml. In heparin-free medium, the primary HUVECs migrated more at 10μg/ml for cells seeded with HA while they migrated more with HAF at a concentration of 250μg/ml. SDS-PAGE and western blot analysis showed CD44 molecules on the primary HUVECs. Conclusion: The assays were set up and studied in duplicate. Further analysis should be done in order to substantiate the role of heparin in the migration of primary human umbilical vein endothelial cells following signaling with CD44 and HA. This might hold promise for therapeutic research into cancer metastasis and neovascularisation.

AB - Background: CD44 is an extracellular matrix molecule that has hyaluronan as one of its principal ligands. The interaction between CD44 and its ligand initiates activation and migration of endothelial cells. Migration of endothelial cells holds importance in neovascularization during cancer metastasis where new blood vessels are formed in order to nurture metastasized cancerous tissue. Studies on bovine endothelial cells showed that greater degree of migrations occurred with addition of lower molecular weight hyaluronic acid fragments (HAF) than the whole molecule of hyaluronan (HA). Our study included addition of heparin to the seedings of HA and HAF at concentrations of 10μg/ml, 100μg/ml, 250μg/ml and 500μg/ml to endothelial cells and measuring the degrees of migrations obtained on migration assay. Methods: HUVECs were isolated from umbilical cords obtained from Kasturba Medical College, Manipal, India. The cells were standardized to 2000 cells/ml and cultured in co-star well plates and were seeded with HA and HAF at concentrations of 10μg/ml, 100μg/ml, 250μg/ml and 500μg/ml. Basal level of control were resting cells and PMA treated cells were positive control. Migration assay was done on which the endothelial cells were embedded. After treatments, the migrations were measured and compared. Expression of CD44 was done using SDS-PAGE and Western Blot analysis. Results: Heparin seems to play a role in the activation and migration of endothelial cells and their sustenance and viability in the medium. In Heparin M199 medium, the endothelial cells showed greater migrations when seeded with HAF than with HA. At 100μg/ml heparinised endothelial cells showed greater degree of migration with HAF than HA added at the same 100μg/ml. In heparin-free medium, the primary HUVECs migrated more at 10μg/ml for cells seeded with HA while they migrated more with HAF at a concentration of 250μg/ml. SDS-PAGE and western blot analysis showed CD44 molecules on the primary HUVECs. Conclusion: The assays were set up and studied in duplicate. Further analysis should be done in order to substantiate the role of heparin in the migration of primary human umbilical vein endothelial cells following signaling with CD44 and HA. This might hold promise for therapeutic research into cancer metastasis and neovascularisation.

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