Poster J15 - Neutrophil Migration in Microfluidic Vascular Models

Friday, October 25, 2024

3:30 PM - 4:30 PM EST

Location: Exhibit Hall E, F & G

Primary Investigator(s)

SS

Sarah Shelton

Assistant Professor
North Carolina State University & University of North Carolina at Chapel Hill
Cary, North Carolina, United States

Presenting Author(s)

SD

Shania Dantes

Undergraduate Student
Biomedical Engineering/UNC & NC State, United States

Introduction: Neutrophils, a type of white blood cell, play a crucial role in the immune system as the first responders to infections in the body. However, their involvement in inflammatory diseases, such as asthma and cancer, can be both beneficial and detrimental. For example, neutrophils can help patients with asthma (a chronic inflammatory lung disease) breathe better by regulating the airway. On the other hand, an excess number of neutrophils can obstruct the airway and reduce the effectiveness of asthma treatments. Therefore, this research project aims to determine the conditions that shape neutrophil migration, using vascular microphysiological systems to model their path through the circulatory system into tissue. In these studies, we perfuse neutrophils through 3D, in vitro microvascular networks to observe their extravasation and migration to understand neutrophils' behavior in those networks.

Materials and

Methods: The microvascular networks were created by growing human umbilical vein endothelial cells (HUVECs) and normal human lung fibroblasts (NHLF) in fibrin within microfluidic devices. After the assembly of perfusable vascular networks, neutrophils were introduced by adding them to the media compartment of the devices and flowing them across the vascular compartment. The devices were then imaged on a confocal microscope, allowing us to observe neutrophil extravasation and migration.

Results, Conclusions, and Discussions: The results showed that neutrophils' perfusion and migration highly depended on the perfusability of the microvascular networks. Perfusability was influenced by the concentration of HUVECs and NHLF, as well as the fibrinogen density and thrombin concentration in the central hydrogel. Microfluidic devices with a concentration of 6.5x10^6to 1x10^6 (HUVECs to NHLF) and 3mg/mL fibrin had an average of 21.83 neutrophils in the microvascular networks and an average of 7.44 extravasated into the matrix. This was because the networks were relatively thin (average width of 21.76 +/- 6.28), decreasing the perfusability. When the concentration of HUVECs to NHLF was changed to 7x10^6to 1x10^6, the microvascular networks were thin (average width of 19.93 +/- 5.22) and less perfusable. They had an average of 18 neutrophils in the microvascular networks, and an average of 10.5 extravasated into the matrix. Whereas, when fibrin density was reduced to density of 2.75mg/mL with a concentration of 6.5x10^6to 1x10^6 (HUVECs to NHLF), the microvascular networks had an average diameter of 23.512 +/- 7.44, the larger vessels had improved perfusability. These findings highlight the factors influencing neutrophil perfusion and extravasation in microvascular networks. Lowering fibrin density promoted the formation of perfusable networks, allowing more neutrophils to circulate and extravasate. In ongoing work, we will compare neutrophil adhesion and extravasation in the presence of activating compounds (e.g. phorbol myristate acetate), inflammatory cytokines (e.g. TNF-a), and inhibitory compounds (e.g. corticosteroids or withaferin A). In addition to observing the behaviors of neutrophils in inflammatory conditions such as asthma, future research will also examine the role of neutrophils in cancer, enabling us to explore the dual roles of neutrophils in inflammatory diseases as well as their interaction with the tumor microenvironment.

Acknowledgements (Optional): Shania Dantes was supported by the Women and Minorities REU program at NC State University.