A Unique In Vitro Migration Model for Leukocyte Trans-Lymphatic Endothelial Migration.
1University of Maryland, Baltimore
2University of Minnesota, Minneapolis.
Meeting: 2016 American Transplant Congress
Abstract number: A25
Keywords: Adhesion molecules, Endothelial cells, Gene expression, Leukocytes
Session Information
Session Type: Poster Session
Date: Saturday, June 11, 2016
Session Time: 5:30pm-7:30pm
Presentation Time: 5:30pm-7:30pm
Location: Halls C&D
Trans-endothelial migration (TEM) is an essential process for leukocyte circulation. While much is known about lymphocyte blood to tissue migration, far less is known about tissue to lymphatic migration. There are in vivo models for assessing lymphatic TEM such as ear pinna and footpad assays but hard to quantitate, and in vitro models that require highly complex equipment. Our aim was to create a robust in vitro system to evaluate lymphatic TEM for various cell types, migration to different chemo-attractants, and have the capacity to integrate stromal variability and fluid flow.
Methods: MS-1 blood endothelial (BEC) and SVEC4-10 lymphatic endothelial cell (LEC) lines and human and murine primary LEC were used in transwell assays. Human and murine CD4 T cells placed in the upper chamber were migrated across endothelium to chemokines, cytokines, or S1P; while stromal fibers and fluid flow variables were assessed.
Results: Human and murine CD4 T cells migrated through LEC but not BEC, in a vectorial fashion, migrating from the abluminal to the luminal surface but not in the reverse direction. The migration was chemotactic and dose-dependent to chemoattractants. Under interstitial-type flow conditions, CD4 T cells migrated more toward S1P and other chemokines across LEC than without flow. Mouse LEC lines and primary cells were characterized by high expression of Lyve-1, Prox1, Vegfr3 and gp38, but not PNAd, confirming their LEC origin and phenotype. Stromal support of the LEC with collagens (gelatin), laminin a4, or laminin a5 were equally effective in enhancing LEC growth and layer integrity and assuring leukocyte vectorial migration. Expression of VCAM-1 and ICAM-1 by LEC was increased by the inflammatory cytokines TNFa or IFNg; and CD4 T cell migration toward S1P and CCL19 were increased across TNFa pretreated LEC. CD8 T cells also migrated across LEC to S1P and chemokines, but exhibited some differences from CD4 cells with regard to vectorial migration.
Conclusion: We devised a simple system that modeled lymphatic TEM through LEC monolayers, validated vectorial migration, established the ability to assess migration of different cell types to chemo-attractants, showed that human and murine cells behaved identically, and that lymphatic flow and stromal structural element can be easily incorporated into the model. This system paves the way for high throughput screening of drugs for regulating lymphatic migration in immunity or tolerance.
CITATION INFORMATION: Xiong Y, Brinkman C, Hippen K, Blazar B, Bromberg J. A Unique In Vitro Migration Model for Leukocyte Trans-Lymphatic Endothelial Migration. Am J Transplant. 2016;16 (suppl 3).
To cite this abstract in AMA style:
Xiong Y, Brinkman C, Hippen K, Blazar B, Bromberg J. A Unique In Vitro Migration Model for Leukocyte Trans-Lymphatic Endothelial Migration. [abstract]. Am J Transplant. 2016; 16 (suppl 3). https://atcmeetingabstracts.com/abstract/a-unique-in-vitro-migration-model-for-leukocyte-trans-lymphatic-endothelial-migration/. Accessed December 2, 2024.« Back to 2016 American Transplant Congress