Cellular self-assembly into 3D microtissues enhances the angiogenic activity and functional neovascularization capacity of human cardiopoietic stem cells
While cell therapy has been proposed as next-generation therapy to treat the diseased heart, current strategies display only limited clinical efficacy. Besides the ongoing quest for the ideal cell type, in particular the very low retention rate of single-cell (SC) suspensions after delivery remains a major problem. To improve cellular retention, cellular self-assembly into 3D microtissues (MTs) prior to transplantation has emerged as an encouraging alternative. Importantly, 3D-MTs have also been reported to enhance the angiogenic activity and neovascularization potential of stem cells. Therefore, here using the chorioallantoic membrane (CAM) assay we comprehensively evaluate the impact of cell format (SCs versus 3D-MTs) on the angiogenic potential of human cardiopoietic stem cells, a promising second-generation cell type for cardiac repair. Biodegradable collagen scaffolds were seeded with human cardiopoietic stem cells, either as SCs or as 3D-MTs generated by using a modified hanging drop method. Thereafter, seeded scaffolds were placed on the CAM of living chicken embryos and analyzed for their perfusion capacity in vivo using magnetic resonance imaging assessment which was then linked to a longitudinal histomorphometric ex vivo analysis comprising blood vessel density and characteristics such as shape and size. Cellular self-assembly into 3D-MTs led to a significant increase of vessel density mainly driven by a higher number of neo-capillary formation. In contrast, SC-seeded scaffolds displayed a higher frequency of larger neo-vessels resulting in an overall 1.76-fold higher total vessel area (TVA). Importantly, despite that larger TVA in SC-seeded group, the mean perfusion capacity (MPC) was comparable between groups, therefore suggesting functional superiority together with an enhanced perfusion efficacy of the neo-vessels in 3D-MT-seeded scaffolds. This was further underlined by a 1.64-fold higher perfusion ratio when relating MPC to TVA. Our study shows that cellular self-assembly of human cardiopoietic stem cells into 3D-MTs substantially enhances their overall angiogenic potential and their functional neovascularization capacity. Hence, the concept of 3D-MTs may be considered to increase the therapeutic efficacy of future cell therapy concepts.
KeywordsCardiopoietic stem cells Angiogenesis Neovascularization Perfusion capacity Chorioallantoic membrane (CAM) assay Microtissues Three dimensional
Acute myocardial infarction
Chronic heart failure
Haematoxylin and eosin
Human platelet lysate
Mean perfusion capacity
Magnetic resonance imaging
Mesenchymal stem cells
Regions of interest
Scanning electron microscope
Vascular endothelial growth factor
Total vessel area
We thank Carol De Simio for her excellent graphical support (University Hospital Zurich, Switzerland). We additionally thank André Fitsche, Christiane Mittmann, Ursula Süss, and Pia Fuchs for great support on histological processing (Institute of Pathology and Department of Surgical Research, University Hospital Zurich, Switzerland). Fatma Kivrak Pfiffner is acknowledged for technical assistance with the handling of the eggs (Clinic for Plastic Surgery and Hand Surgery, University Hospital Zurich, Switzerland). We kindly thank Peter De Weale, Aymeric Seron, Dorothee Daro, and Sebastien Mauen (Celyad, Belgium) for generation of the GMP-grade human cardiopoietic stem cells.
PW, JB, and MYE designed experiments; PW, AB, AW, and JB performed experiments; PW, AB, JB, and MYE analyzed all data; YT injected contrast agent for MRI measurements; OE performed SEM; MH evaluated all histological sections; PW, AB, JB, and MYE wrote manuscript; PW, AB, AW, MH, PG, MC, SPH, JB, and MYE edited and discussed manuscript.
This work was supported by the Hartmann Müller-Foundation and the Swiss Heart Foundation.
Compliance with ethical standards
Conflict of interest
The authors declare that they have no competing interests.
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