Umbilical Cord Blood-Derived Monocytes as A Reliable Source of Functional Macrophages for Biomedical Research

Abstract

Objective: Macrophages are multifunctional immune cells widely used in immunological research. While autologousmacrophages have been widely used in several biomedical applications, allogeneic macrophages have alsodemonstrated similar or even superior therapeutic potential. The umbilical cord blood (UCB) is a well-describedsource of abundant allogenic monocytes and macrophages that is easy to collect and can be processed withoutinvasive methods. Current monocyte isolation procedures frequently result in heterogenous cell products, with limitedyields, activated cells, and high cost. This study outlines a simple isolation method that results in high yields and puremonocytes with the potential to differentiate into functional macrophages.Materials and Methods: In the experimental study, we describe a simple and efficient protocol to isolate highpuritymonocytes. After collection of human UCB samples, we used a gradient-based procedure composed of threeconsecutive gradient steps: i. Hydroxyethyl starch-based erythrocytes sedimentation, followed by ii. Mononuclearcells (MNCs) isolation by Ficoll-Hypaque gradient, and iii. Separation of monocytes from lymphocytes by a slighthyperosmolar Percoll gradient (0.573 g/ml). Then the differentiation potential of isolated monocytes to pro- and antiinflammatorymacrophages were evaluated in the presence of granulocyte colony-stimulating factor (GM-CSF) andmacrophage CSF (M-CSF), respectively. The macrophages were functionally characterized as well.Results: A high yield of monocytes after isolation (25 to 50 million) with a high purity (>95%) could be obtained fromevery 100-150 ml UCB. Isolated monocytes were defined based on their phenotype and surface markers expressionpattern. Moreover, they possess the ability to differentiate into pro- or anti-inflammatory macrophages with specificphenotypes, gene/surface protein markers, cytokine secretion patterns, T-cell interactions, and phagocytosis activity.Conclusion: Here we describe a simple and reproducible procedure for isolation of pure monocytes from UCB, whichcould be utilized to provide functional macrophages as a reliable and feasible source of allogenic macrophages forbiomedical research.

Keywords

• macrophage polarization
• monocytes
• umbilical cord blood