Advances in Endothelial Cell Isolation and Culture

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Advances in Endothelial Cell Isolation and Culture

The endothelium plays crucial roles in physiological processes as diverse as angiogenesis, vasoconstriction, haemostasis, leucocyte trafficking and antigen presentation. Therefore, manipulation of endothelial cell (EC) gene expression and function is very attractive for therapeutic intervention. This meeting will highlight exciting new areas in EC research, including isolation and culture of EC from different vascular beds, culture of EC at haemodynamic shear stress and novel techniques for EC targeted gene transfection.

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Agenda
Morning Session

09:30 10:00 Registration Tea, coffee and biscuits

10:00 10:30 Introduction by the Chair: Dr Charlotte Lawson Royal Veterinary College

10:30 11:00 Isolation and characterisation of Equine digital vein endothelial cells (EDVEC)
Dr. Yoel Berhane
EDVEC were isolated from hind limbs obtained from mixed breed adult horses killed at an abattoir. Briefly, after ligation of the digital arteries, EDVEC were isolated following collagenase digestion (collagenase type II, 1mg/ml) under sterile conditions and were cultured with DMEM in humidified incubator containing 5% CO2 at 37oC. EDVEC reached confluency in 5 10 days and were identified by forming a monolayer in culture and having the cobblestone morphology, characteristic of endothelial cells. The phenotype of EDVEC was further confirmed by positive immunofluorescent staining for the endothelial cell marker, von Willebrand factor.

11:00 11:20 Morning Tea/Coffee with the companies

11:20 11:50 Culture of endothelial cells in hollow fiber modules under conditions of defined chronic shear stress
Mr. John Cadwell
Endothelial cells behave very differently when cultured under conditions of shear stress as opposed to standard flask culture. They lay down flat, form a monolayer, create tight junctions and express approximately a dozen genes that are not expressed in static culture. Endothelial cells can be seeded onto the inside of hollow fibers where the defined flow of medium over them produces chronic shear stress. Additionally the fibers are manufactured of a material that allows for the binding of specific matrix proteins onto the surface of the fiber. A description of the technique and examples of the types of data obtained from it will be presented.

11:50 12:05 Evaluation of circulating EPC levels in various disease groups
Dr Anna Dulic-Sills

EPCs are present in the peripheral blood homing to sites of ischaemia and contributing to postnatal vasculogenesis.

It is not clear at present whether and how the number of circulating EPCs vary in different diseases and how this number correlates with the prognosis, progression and clinical outcome. We aim to provide a database of the effect of a number of diseases on t he levels of circulating EPCs and to correlate the numbers and function with the disease activity and clinical outcomeл

12:05 13:00 Lunch and meet the companies

Afternoon Session

13:00 13:20 Primary Porcine Blood-Brain Barrier Endothelial Cells: A Model System for Therapeutic Screening, Transport and Uptake Studies
Mr Mathew Smith

The blood-brain barrier (BBB) remains a significant obstacle to the delivery of biologics to the central nervous system (CNS) and the need for a robust, simple and readily available BBB in-vitro model is undeniable. We have developed an in-vitro primary porcine model that shows promise for use in therapeutic screening programs and for understanding transport and uptake systems within the BBB. Here we present supporting data.

13:20 13:50 Immunolipsomes for efficient gene transfer to primary human endothelial cells
Prof. Andrew George

Gene therapy directed at endothelial cells has great potential in the context of inflammatory diseases. At present viral vectors, when used to transduce endothelial cells, result in activation of the cells and upregulation of adhesion molecules. We have therefore developed novel methods for endothelial transfection using antibodies linked to liposomes. This allows both efficient gene transfer and also targeted delivery for example to endothelial cells expressing E-selectin. This approach has been developed using primary human saphenous vein endothelial cells, which have advantages as a model when compared to the more commonly used umbilical vein endothelial cells.

13:50 14:20 Afternoon Tea/Coffee and cakes with the companies

14:20 14:50 Simulation of blood vessels in cell culture biochips
Dr. Ulf Rdler
Vessel diseases as for instance Arteriosclerosis belong in the affluent society to the most common causes of death.

Here we describe a biomimetic system for the simulation of blood vessels, which is based on disposable flow chambers. It allows cultivation and high end microscopy investigation of endothelial cells on adjustable flow conditions. In contrast to the round cross section of natural blood vessels, a rectangular cross section is favourable for microscopic applications, which is realised in the Slides. Surface modifications have been tested to evaluate optimal shear rate analysis for HUCECs under flow conditions.

14:50 15:20 BD BioCoat TM Solutions for Angiogenesis Cell Culture Assays
Dr Mathias Beckmann
Angiogenesis, the formation of new blood vessels, is essential for normal growth and homeostasis. In addition, since the balance in angiogenesis can be key in certain disease status, angiogenesis is a rapidly expanding field for the drug discovery and basic research scientist.
We have developed several in vitro assays to understand the mechanisms of angiogenesis and to identify potential therapeutic molecules. These BioCoat cell-based assay platforms for routine use addressing endothelial invasion, migration and tube formation will be described in the talk.

15:20 Close

Standard fee - 160
Student fee - 80 (subject to proof of studies)

This meeting has been granted CPD creditation