A novel 3D dual hollow fibre bioreactor for the production of human red blood cells

Abstract

Blood shortage is one of the biggest concerns of the World Health Organization. The reason: while around 30% of the industrialized world population will require a life-saving transfusion sometime during their lives, only 6% of the total population actually donates blood that can only be stored for up to 42 days. Despite the huge efforts that have been made to awaken the population to proactively donate blood (according to the World Health Organization 2009a, apporximately 85 million units of blood are collected annually worldwide), there is still a large shortage around the world. Hence, other alternatives have been suggested to address this. Since the first attempts, in the late 1940s, to produce an alternative to blood donation in the laboratory, several breakthroughs were made, most of them focusing on the synthetic route: the search for a chemical molecule that could replace the main vital function of red blood cells (RBCs) - oxygenation of the body cells (Kimball 1994). Nevertheless, several issues, mainly regarding the stability and controlled oxygen release by these molecules, still pose serious barriers for the success of this option (Kimball 1994). Nevertheless, several issues, mainly regarding the stability and controlled oxygen release by these molecules, still pose serious barriers for the success of this option (Kimball 1994). Hence, the focus on blood substitutes is now being directed to blood itself. My PhD project in the Department of Chemical Engineering of Imperial College London aimed at mimicking nature's bioreactor to produce human blood: the Bone Marrow (BM), a three-dimensional (3D) structure comprised of a vascularized support matrix, cellular constituents, and humoral factors. The BM 3D spatial configuration generates micro-concentration gradients that modulate cellular self-renewal, differentiation and apoptosis - mimicked by a 3D scaffold that can be rendered bioactive by coating with extracellular proteins. On the other hand, the BM vascular system ensures the supply of nutrients and removal of harmful metabolites, as well as the collection of maturing cells formed in the marrow cavity - mimicked by an intricate selectively-permeable membrane system that both renews the microenvironment and harvests mature RBCs. I have combined these two bio-inspired characteristics of the marrow into the first ex vivo 3D dual hollow fibre bioreactor (DHFB) that allows addressing mass transfer challenges faced by state-of-the-art technology and the continuous production and release of RBCs. This system was shown to be biocompatible, and allowed the differentiation of cord blood stem cells into mature enucleated RBCs under a cocktail composed of 100ng.mL-1 stem cell factor and 2,000U.ml-1 erythropoietin over 31 days. Mature enucleated RBCs could be prpduced exclusively ex vivo in a 3-dimension feeder-free culture. This technology has the potential to allow cost-effective production of clinically-relevant numbers of red blood cells with selective cell harvesting in a closed hematopoietic system.