Stimulation of In Vivo Angiogenesis Through Dual Growth Factor Loaded Hyaluronic Acid Hydrogels Celeste Riley Mentor: Dr. Robert Peattie Department of Chemical Engineering

Importance Engineered tissues Organ transplants Ischemic Pathologies (Diabetes)

Components of the Project Preparation of Hydrogels Surgery on living mouse models Count formed blood vessels and analyze data

Hydrogel Preparation What is a hydrogel? Natural or Synthetic Biocompatible Crosslinked to form a gel-like substance Dehydration

Chemical structure of hyaluronic acid Hydrogel Preparation What is Hyaluronic Acid? Glycosaminoglycan Complexes H20 molecules Chief component of extracellular matrix Contributes to cell proliferation and migration Chemical structure of hyaluronic acid http://www.hyalogic.com/aboutha.htm

Addition of Growth Factors What are growth factors? Diagram of Growth Factor Binding Signaling proteins Bind receptors of target cells Promote differentiation and maturation http://www.grahamstudios.net/medcon.html

Addition of Growth Factors Which Growth Factors? Model of VEGF Vascular Endothelial Growth Factor (VEGF) Angiopoietin 1 (Ang-1) http://kerrins.net/ud/proteins.html

Goal of Project GFs + Make hydrogel from hyaluronic acid Impregnate hydrogel with growth factors Ang-1 and VEGF Dehydrate and implant hydrogel into mouse ear Determine if implant, loaded with growth factors, stimulates angiogenesis GFs +

Project Setup Conditions to consider Spontaneous Growth Original Vessels Individual combinations: control cases (chart)

Treatment Cases 6 animals per treatment group Treatment 1 2 3 4 5 6 7 8 HA gel - + VEGF Ang-1

Determining Blood Vessel Growth Thin-section both surgical and contralateral ear Count blood vessels under 400X magnification Contralateral Ear

Data Analysis Define a statistical index to account for What do we want to measure? How do we measure it? Define a statistical index to account for effect of sham surgery number of vessels present before surgery Additional vessels due only to the particular surgical treatment

Neovascularization Index NI = (treatment-CL) – (sham-CL) mean CL (treatment-CL) = average # of additional blood vessels in the implanted ear of treatment group (sham-CL) = average # of additional vessels due to the surgical procedure itself mean CL = average vessel count in all contralateral ears: effectiveness measure

Results Treatment 7 day NI 14 day NI - 0.12 1.80 0.95 1.30 2.97 6.74 Sham - HA gel only 0.12 1.80 VEGF only 0.95 1.30 Ang-1 only Ang-1 + VEGF HA gel + Ang-1 HA gel + VEGF 2.97 6.74 HA gel + Ang-1 + VEGF

Conclusion Currently, data collection is still ongoing When calculated, compare NI of experimental case to NIs of control cases At this point, no definite conclusion can be drawn about the effect of Ang-1 and VEGF delivered through HA hydrogel on angiogenesis

Hughes Medical Institute Acknowledgments Dr. Robert Peattie Dr. Kevin Ahern Howard Hughes Medical Institute Co-researcher Peter Fuegy