Research Highlight: Harnessing the immune response for successful scaffold vascularization

Written by Marjan Rafat

Highlighting one of the latest articles in regenerative medicine

Evaluation of: Spiller KL, et al. The role of macrophage phenotype in vascularization of tissue engineering scaffolds. Biomaterials 35(15), 4477-4488 (2014)

One major challenge for tissue engineering scaffolds remains vascularization and subsequent anastamosis (1). Macrophages play a complex role in this process–“classically activated” M1 are generally thought to have a pro-inflammatory phenotype while “alternatively activated” M2 macrophages are associated with a pro-healing phenotype. This designation, however, remains controversial, and their involvement in angiogenesis and wound healing are poorly understood (2).

In this study, Spiller et al. describe the complexity of the macrophage response in the vascularization of scaffolds (3). Macrophages were derived from monocytes isolated from human blood samples and further polarized by incubation with chemokines known to induce diverse phenotypes. M1, M2a, and M2c macrophages were studied. Characterization of the polarized macrophages by RT-PCR and flow cytometry showed a unique marker signature for each phenotype. Gene expression and protein secretion of common angiogenic factors were also analyzed. M1 macrophages expressed genes involved in the initiation of angiogenesis while M2a macrophages expressed factors that support angiogenesis through pericyte recruitment and also regulating M1 signaling. Interestingly, M1 macrophages also expressed high levels of a pericyte-recruiting factor while all macrophages expressed extracellular matrix remodeling factors, indicating the overlapping roles of multiple phenotypes in vascularization. The ability of the macrophages to encourage angiogenesis was analyzed through in vitro sprouting assays. Surprisingly, M2 macrophages, thought to be “pro-angiogenic,” consistently produced fewer and shorter sprouts among the phenotypes. In a fibrin assay, it was found that changes in network formation and organization of endothelial cells (ECs) are initiated not by continuous exposure to factors secreted by M2a cells but when ECs experienced removal of those signals before exposure, which is consistent with findings in vivo where M1 macrophages are involved in early angiogenesis and wound healing while M2 macrophages appear in later stages. Finally, collagen scaffolds–either soaked in buffer as a control, soaked in lipopolysaccharide (LPS) to enhance the M1 response, or crosslinked with glutaraldehyde to preserve scaffold integrity and to stimulate a moderate inflammatory response–were implanted subcutaneously into immunocompetent mice. The unmodified collaged scaffold evoked the foreign body response and fibrous encapsulation, which was correlated to strong M2 marker staining. The unmodified or LPS-soaked scaffolds did not promote angiogenesis. The crosslinked scaffold, however, appeared well-vascularized. Although they did not distinguish between M2 types, it was clear that the crosslinked scaffolds induced infiltration of both M1 and M2-polarized macrophages, indicating the benefit and necessity of recruiting of both phenotypes in vascularization.

This study brings to light the importance of macrophage phenotype and kinetics in designing tissue engineering scaffolds. The authors propose a mechanism in which the M1, M2a, and M2c phenotypes interact to promote angiogenesis. Although some limitations of the paper include not fully characterizing or quantifying the response in vivo and not including other M2 phenotypes that may play a role in promoting angiogenesis, this work highlights the complexities of the immune response required for successful scaffold integration.


1. Novosel EC, Kleinhans C, Kluger PJ Vascularization is the key challenge in tissue engineering. Advanced drug delivery reviews 63(4-5), 300-311 (2011).

2. Kitajewski J Wnts heal by restraining angiogenesis. Blood 121(13), 2381-2382 (2013).

3. Spiller KL, et al. The role of macrophage phenotype in vascularization of tissue engineering scaffolds. Biomaterials 35(15), 4477-4488 (2014)

Scheduled to be published in Volume 10 Issue 01 of Regenerative Medicine.