Macrophages are primarily known as phagocytic immune cells, but they also play a role in diverse processes, such as morphogenesis, homeostasis and regeneration. In this review, we discuss the influence of macrophages on angiogenesis, the process of new blood vessel formation from the pre‐existing vasculature. Macrophages play crucial roles at each step of the angiogenic cascade, starting from new blood vessel sprouting to the remodelling of the vascular plexus and vessel maturation. Macrophages form promising targets for both pro‐ and anti‐angiogenic treatments. However, to target macrophages, we will first need to understand the mechanisms that control the functional plasticity of macrophages during each of the steps of the angiogenic cascade. Here, we review recent insights in this topic. Special attention will be given to the TIE2‐expressing macrophage (TEM), which is a subtype of highly angiogenic macrophages that is able to influence angiogenesis via the angiopoietin–TIE pathway.

Tissue engineering aims to structurally and functionally regenerate damaged tissues, which requires the formation of new blood vessels that supply oxygen and nutrients by the process of angiogenesis. Stem cells are a promising tool in regenerative medicine due to their combined differentiation and paracrine angiogenic capacities. The study of their proangiogenic properties and associated potential for tissue regeneration requires complex in vivo models comprising all steps of the angiogenic process. The highly vascularized extraembryonic chorioallantoic membrane (CAM) of fertilized chicken eggs offers a simple, easy accessible, and cheap angiogenic screening tool compared to other animal models. Although the CAM assay was initially primarily performed for evaluation of tumor growth and metastasis, stem cell studies using this model are increasing. In this review, a detailed summary of angiogenic observations of different mesenchymal, cardiac, and endothelial stem cell types and derivatives in the CAM model is presented. Moreover, we focus on the variation in experimental setup, including the benefits and limitations of in ovo and ex ovo protocols, diverse biological and synthetic scaffolds, imaging techniques, and outcome measures of neovascularization. Finally, advantages and disadvantages of the CAM assay as a model for angiogenesis in tissue engineering in comparison with alternative in vivo animal models are described.

The myocardial sleeve of the porcine and equine pulmonary veins were histologically investigated and reconstructed three dimensionally. Moreover, the localization of neuron cell bodies at the veno-atrial junction and alongside the myocardial sleeve was light microscopically visualized to depict the organization of nerve, myocardial and fat tissue. Finally, the presence of telocytes inside the equine pulmonary veins was demonstrated by use of transmission electron microscopy. These structures are thought to play a role in the induction of atrial fibrillation, which is frequently seen in horses, while pigs are often used as a cardiovascular model in this context. This data fills in remaining gaps in the literature concerning the histological build-up of the pulmonary veins wall in pigs and horses. In-depth knowledge on the myocardial sleeve and its surrounding cell types are important to understand the possible outcome of an ablation therapy as an atrial fibrillation treatment. In pigs and horses, the layout of the pulmonary veins wall concerning these structures is comparable to humans. However, neuron cell bodies were recovered at the veno-atrial junction in both species but not alongside the myocardial sleeve in horses.

Recently, a new interstitial cell type called telocyte has been identified. Telocytes are found in many organs including the heart, where they are especially well described. However, their presence in the porcine heart has not yet been proven. The pig is a valuable animal model in research because of its resemblance with man, making it interesting to determine whether telocytes can be found in pigs as well. The focus of this study is the identification and ultrastructural description of telocytes in the heart tissue of pig. Using transmission electron microscopy, telocytes were found in both left and right atrium and ventricle, usually close to cardiomyocytes and/or blood vessels. Their most important characteristic is the long cytoplasmic processes called telopodes, which have a moniliform aspect, measure tens of μm and usually have a thickness below 0.2 μm. This unique morphological feature enables telocytes to be recognized from other interstitial cells such as fibroblasts. Additional observations include the ability to release extracellular vesicles and to make contacts with other structures such as endothelial cells, suggesting a role in intercellular communication.

Macrophages play an important but poorly understood role in angiogenesis. To investigate their role in vessel formation, relevant in vivo models are crucial. Although the chick chorioallantoic membrane (CAM) model has been frequently used as an angiogenesis assay, limited data are available on the involvement of chicken macrophages in this process. Here, we describe a method to deplete macrophages in the ex ovo chick CAM assay by injection of clodronate liposomes and show that this depletion directly affects vascularisation of collagen onplants. Chicken embryos were injected intravenously with either clodronate or phosphate-buffered saline (PBS) liposomes, followed by placement of collagen type I plugs on the CAM to quantify angiogenic ingrowth. Clodronate liposome injection led to a significant 3.4-fold reduction of macrophages compared with control embryos as measured by immunohistochemistry and flow cytometry. Furthermore, analysis of vessel ingrowth into the collagen plugs revealed a significantly lower angiogenic response in macrophage-depleted embryos compared with control embryos, indicating that chicken embryonic macrophages play an essential function in the development of blood vessels. These results demonstrate that the chick CAM assay provides a promising model to investigate the role of macrophages in angiogenesis.