TY - JOUR

T1 - A pre-conditioning protocol of peripheral blood derived endothelial colony forming cells for endothelialization of tissue engineered constructs

AU - Kraus, Xenia

AU - Pflaum, Michael

AU - Thoms, Stefanie

AU - Jonczyk, Rebecca

AU - Witt, Martin

AU - Scheper, Thomas

AU - Blume, Cornelia

N1 - Funding Information: This study was funded by the Deutsche Forschungsgemeinschaft (DFG – Grant Number: 388094931 ). This work has been carried out within the framework of the SMART BIOTECS alliance between the Technische Universität Braunschweig and the Leibniz Universität Hannover. This initiative is supported by the Ministry of Economy and Culture (MWK) of Lower Saxony, Germany.

PY - 2021/3

Y1 - 2021/3

N2 - In regenerative medicine, autologous endothelial colony forming cells (ECFCs) bear the greatest potential to be used for surface endothelialization of tissue engineered constructs, as they are easily attainable and possess a high proliferation rate. The aim of this study was to develop a standardized pre-conditioning protocol under dynamic conditions simulating the physiology of human circulation to improve the formation of a flow resistant monolayer of ECFCs and to enhance the antithrombogenicity of the endothelial cells. The main focus of the study was to consequently compare the cellular behavior under a steady laminar flow against a pulsatile flow. Mononuclear cells were isolated out of peripheral blood (PB) buffy coats and plated on uncoated tissue culture flasks in anticipation of guidelines for Advanced Therapy Medicinal Products. ECFCs were identified by typical surface markers such as CD31, CD146 and VE-Cadherin. To explore the effects of dynamic cultivation, ECFCs and human umbilical vein endothelial cells were comparatively cultured under either laminar or pulsatile (1 Hz) flow conditions with different grades of shear stress (5 dyn/cm2 versus 20 dyn/cm2). High shear stress of 20 dyn/cm2 led to a significant upregulation of the antithrombotic gene marker thrombomodulin in both cell types, but only ECFCs orientated and elongated significantly after shear stress application forming a confluent endothelial cell layer. The work therefore documents a suitable protocol to pre-condition PB-derived ECFCs for sustainable endothelialization of blood contacting surfaces and provides essential knowledge for future cultivations in bioreactor systems.

AB - In regenerative medicine, autologous endothelial colony forming cells (ECFCs) bear the greatest potential to be used for surface endothelialization of tissue engineered constructs, as they are easily attainable and possess a high proliferation rate. The aim of this study was to develop a standardized pre-conditioning protocol under dynamic conditions simulating the physiology of human circulation to improve the formation of a flow resistant monolayer of ECFCs and to enhance the antithrombogenicity of the endothelial cells. The main focus of the study was to consequently compare the cellular behavior under a steady laminar flow against a pulsatile flow. Mononuclear cells were isolated out of peripheral blood (PB) buffy coats and plated on uncoated tissue culture flasks in anticipation of guidelines for Advanced Therapy Medicinal Products. ECFCs were identified by typical surface markers such as CD31, CD146 and VE-Cadherin. To explore the effects of dynamic cultivation, ECFCs and human umbilical vein endothelial cells were comparatively cultured under either laminar or pulsatile (1 Hz) flow conditions with different grades of shear stress (5 dyn/cm2 versus 20 dyn/cm2). High shear stress of 20 dyn/cm2 led to a significant upregulation of the antithrombotic gene marker thrombomodulin in both cell types, but only ECFCs orientated and elongated significantly after shear stress application forming a confluent endothelial cell layer. The work therefore documents a suitable protocol to pre-condition PB-derived ECFCs for sustainable endothelialization of blood contacting surfaces and provides essential knowledge for future cultivations in bioreactor systems.

KW - Actin cytoskeleton

KW - Bioreactor culture

KW - Endothelial colony-forming cells (ECFC)

KW - Endothelial progenitor cells (EPC)

KW - Endothelialization

KW - Pulsatile flow

KW - Thrombomodulin

UR - http://www.scopus.com/inward/record.url?scp=85097386738&partnerID=8YFLogxK

U2 - 10.1016/j.mvr.2020.104107

DO - 10.1016/j.mvr.2020.104107

M3 - Article

C2 - 33212112

AN - SCOPUS:85097386738

VL - 134

JO - Microvascular research

JF - Microvascular research

SN - 0026-2862

M1 - 104107

ER -