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Biomedical Engineering Education

Biomedical Engineering Education - Poster Session C

Poster E8 - Developing Bulk and Microporous Annealed Based Decellularized Uterine Tissue ECM Scaffolds

Friday, October 25, 2024

10:00 AM - 11:00 AM EST

Location: Exhibit Hall E, F & G

Presenting Author(s)

Lily Ro

Undergraduate
North Carolina State University
Raleigh, North Carolina, United States

Co-Author(s)

AS

Aryssa Simpson

Graduate
North Carolina State University, United States
SW

Shelby White

Post-Doc
Joint Department of Biomedical Engineering, North Carolina State and University of North Carolina-Chapel Hill, Raleigh, NC, United States of America, United States
Donald Freytes

Associate Professor
Joint Department of Biomedical Engineering, North Carolina State and University of North Carolina-Chapel Hill, Raleigh, NC, United States of America, United States

Primary Investigator(s)

AB

Ashley Brown

Associate Professor
NC State University, United States

Introduction: Cesarean section (or delivery) is the surgical delivery of babies through an abdominal and uterine incision. In rare cases, abnormal healing of the uterine muscle occurs, resulting in the rupture of the uterine musculature, but the uterine serosa is still intact. This condition is called uterine scar dehiscence, with current treatment options including resuturing of the uterine incision line or a hysterectomy. This work aims to develop a decellularized porcine uterine tissue surgical additive to encourage wound healing and reduce the incidence of uterine dehiscence. Methods include dissecting porcine tissue and performing a manual decellularization using previously established methods. Following successful decellularization, the decellularized ECM (dECM) tissue will be further digested and formed into a bulk decellularized uterine tissue hydrogel. As a surgical additive alternative, the bulk uterine dECM will be chopped and sonicated to develop microgel-based dECM. This microgel-based dECM will be utilized to formulate a microporous annealed-based decellularized uterine tissue ECM (dut-ECM) scaffold. The scaffold aims to provide a bioactive environment that promotes tissue regeneration, reduces inflammation, and enhances the mechanical integrity of the uterine wall. This ultimately decreases the risk of uterine dehiscence. By mimicking the natural extracellular matrix, the dut-ECM scaffold facilitates cellular adhesion, proliferation, and differentiation. Its microporous structure ensures efficient nutrient and oxygen diffusion that support tissue healing. This approach offers a biologically compatible solution to improve clinical outcomes for patients undergoing cesarean sections.

Materials and

Methods: In this study, we dissected an 18-month-old porcine model and focused on the distal, middle, and proximal uterine horns. The tissue samples underwent decellularization using paraffin and OCT embedding methods to remove cellular components while preserving the extracellular matrix (dECM). The decellularized tissue was then powdered, and luteolin was introduced as a cross-linking agent to form a bulk uterine dECM hydrogel.

Initially, we filtered samples using 70 µm and a combination of 70 & 40 µm filters to refine particle size. This filtration aimed to prepare the samples for further analysis, but it did not achieve the desired microgel consistency when evaluated by Atomic Force Microscopy (AFM). To address this, we prepared new samples and employed sonication to further enhance particle uniformity. Sonication was performed at two settings: 10% amplitude for 1 minute and 20% amplitude for 2 minutes. Following sonication, the samples were filtered through 40 µm cell strainers to achieve a more refined particle size distribution.

Then, Atomic Force Microscopy (AFM) was used to characterize the mechanical and surface properties of the resulting dECM scaffolds. For AFM imaging, samples were prepared by depositing a small volume of the dECM suspension onto a clean glass slide and allowing it to air dry. The AFM was operated to obtain high-resolution topographical images of the scaffold surfaces. This technique allowed for the measurement of particle size, surface roughness, and structural uniformity.

Results, Conclusions, and Discussions: Filtration reduced particle size by removing larger aggregates. Higher concentration (1mgmL) also
produced smaller particles than lower concentrations (0.1mgmL). This suggests that higher
concentrations could result in more effective breakdown into finer particles.

Similarly, sonication decreased particle size, but higher intensity and longer duration (20% for 2 minutes)
resulted in larger particles compared to lower intensity and shorter duration (10% for 1 minute).
Increase in energy could have caused particles to clump together or aggregate.

Filtration alone reduced particle size, but combining it with sonication ensured more consistent
particle sizes, highlighting the advantage of using both methods for optimal microporous scaffolds.

In future studies, we will evaluate polymerization rates via rheology and conduct tensile testing for
dmECM gels, depending on positive LIVE/DEAD results. These studies will assess how particle size
and uniformity from sonication and filtration impact scaffold polymerization behavior and mechanical
strength to optimize scaffold performance.

Acknowledgements (Optional): Advanced Wound Healing Lab

REdesign Lab

Sarah Courtney Cavatoni

Audrey Clark

Chloe Hincher

Dr. Olgha Qaqish