Northeastern Society of Plastic Surgeons

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Intraoperative Bioprinting and Microsurgery: Defining Cell Types within a Vascularized Scaffold
Jessica C. El-Mallah1, Miji Yeo2, Summer N. Horchler3, Olivia Waldron3, Mary Landmesser3, MIngjie Sun3, Mohammad Hossein Asgardoon3 , Ibrahim Ozbolat2, Dino Ravnic1
1Department of Surgery, Penn State Hershey Medical Center, Hershey, PA, United States. 2Department of Biomedical Engineering, Penn State University, Hershey, PA, United States. 3Penn State College of Medicine, Hershey, PA, United States.

Background: Intraoperative bioprinting (IOB) of scaffolds offers a new frontier in reconstructive surgery. Scaffold success relies on rapid vascularization, which can be enhanced through endothelial cell seeding. We have described micropuncture ("MP"), a surgical technique in which the recipient blood vessel wall is precisely disrupted to expedite scaffold vascularization. We have demonstrated that MP increases endothelial cell migration as well as macrophage recruitment to promote angiogenesis. We have found that the combined effects of MP and cell-seeding ("cells") in a collagen-based bioink promote vascularization at short and long timepoints. In this study we seek to identify the cellular contributors to this enhanced vascularization, hypothesizing that there will greater endothelial cell migration and macrophage recruitment in our Cells+MP group and this cellular recruitment will continue to increase with time.
Methods: In a rat model MP was performed on femoral vessels of one hindlimb while the other served as a non-MP control. Bioink scaffolds with or without rat aortic endothelial cells were deposited over the vessels using an extrusion-based bioprinter engineered at our institution. On POD10 or 40, the bioink scaffolds were explanted and prepared for thin section histology. Immunohistochemistry was performed with nuclear marker DAPI, endothelial marker PECAM-1 and macrophage marker F4/80 (n=3 per group). Images were acquired with microscopy and ImageJ was used to process the images for quantification.
Results: MP significantly accelerated cellular infiltration by Day 40, while cell-involved groups exhibited higher levels of PECAM-1 and F4/80 expression compared to the Bioink group, this trend continued to heighten at a later time point (Figure 1).
Conclusion: This study introduces a new approach that combines MP and IOB to create vascularized tissue. Both MP and cell seeding show potential as effective pro-angiogenic strategies. Future research will focus on utilizing these techniques to develop in-situ flaps for tissue reconstruction.


Figure 1. Cellular Dynamics Within an IOB scaffold utilizing MP and Cell-Seeding technology. IHC staining was performed with nuclear marker DAPI, endothelial marker PECAM-1 and macrophage marker F4/80 (n=3). Images were examined for stained area and quantified. One-on-one comparison between the Bioink (control) and experimental groups was conducted via independent-samples T test (p# < 0.05, p## < 0.01, p### < 0.001). One-on-one comparison between two timepoints within the same group was conducted via paired-samples T test (p"  < 0.05, p" "  < 0.01, p" " "  < 0.001).
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