Behavior of Cardiac Cells When Exposed to Different Formulations of Decellularized Extracellular MatrixMSTP-SURF 2015Sarah Albrecht, University of Washington-Seatt leUCSD Department of Bioengineering, Christman Lab

Myocardial Infarction (MI) Cardiovascular disease is the leading cause of death in the United States killing an estimated 600,000 people each year.¹

Left Ventricle (LV) remodeling post-MI◦ Decrease in cardiomyocytes ◦ Degradation of the extracellular matrix of LV◦ Thinning of the LV wall◦ Eventually causes heart failure in the future

Only therapy for heart failure is a heart transplant

Infarcted heart and LV hours after MI

Infarcted heart and LV days after MI

1. Heron, M. Natl. Vital Stat. 2013

Therapies for Post-MI Ventricular Constraint

◦ Mechanical device or synthetic material that encompasses the heart to limit ventricular dilation

Injectable Biomaterial◦ Injecting a biomaterial gel in

the infarct area with specific cells or biomolecules

Cardiac Patch◦ Placement of a biomaterial

sheet seeded either with specific cells or biomolecules over the infarct area

Rane, A. a. & Christman, K. L. J. Am. Coll. Cardiol. 2011

Extracting Myocardial Decellularized ECM

Wang, R. M. & Christman, K. L. Adv. Drug Deliv. Rev. 2015

Reduced negative LV remodeling and improved heart function¹

Decellularized ECM has been shown to be biocompatible²

1. Sonya B. Seif-Naraghi et al. Science Translational Medicine. 20132. Jennifer M. Singelyn et al. Biomaterials. 2009

Extracting Myocardial Decellularized ECM

Can be separated into soluble and insoluble components

Project Overview Research Goal:

◦ Improve our understand of how the soluble component of the ECM hydrogel affects cell viability and migration.

Grow CPCs and HUVECs to a desired cell confluences

Isolate soluble ECM component

Plate cells with soluble ECM for

Viability and Migration assay

Analyze both cell lines appropriately to determine cell

survival and movement

1. Plate CPCs and HUVECs 2. Prepare various H2O2 concentrations (n=4 or 6)

3. Add Alamar Blue in wells and incubate for 4

hours

4. Record the fluorescence of each well with plate reader

Viability Assay

Migration Assay

1. Serum starve CPCs for 24 hours

2. Fluorescently label cells

3. Add media and cell solution into transwell of a 24-well plate

4. Count fluorescent cells to determine how many migrated

CPC Viability 4 hours at 50,000 cells

Results:◦ SS and soluble ECM have

higher viability than SF◦ Survival of CPCs at high

H2O2 concentrations

CPC Viability 8 hours at 25,000 cells

Parameters Changed:◦ Longer incubation time◦ Lower amount of cells per well

Results:◦ No viable cells at higher H2O2

◦ No significance between SS and SF

HUVEC Viability 4 hours at 50,000 cells

Results:◦ SF higher than SS◦ Viable cells at higher H2O2

concentrations

HUVEC Viability 8 hours at 25,000 cells

Parameters Changed:◦ Incubation time◦ Cell concentration◦ Media

Results:◦ No viable cells at higher

H2O2

◦ Significance between SS and SF no HE

Migration Assay with CPCSS SF

6 mg/mL 24 mg/mL

Parameters:◦ Cells counted twice to reduce error◦ Images captured at 6 hours after initial plating◦ n=3

Conclusion Viability Assay

◦ Increasing the incubation time, decreasing the cell concentration and using media without added molecules such as Heparin and E.C.G. affected cell survival

◦ Soluble ECM appears to not be detrimental to CPCs and HUVECs

Migration Assay◦ Soluble ECM encourages cell migration however it is not as affective as the SS media◦ At a higher concentration of the soluble ECM, more cell movement was observed compared to lower

concentrations of the ECM

The ECM can be a suitable injectable biomaterial therapy to treat the infarct region

Future Directions Soluble ECM

◦ Continue with cell studies for ECM hydrogel◦ Preform an angiogenesis assay to observe vessel formation

and cell-cell interactions

Insoluble ECM◦ Perform more cell studies on different

formulations of the insoluble ECM◦ Pellet form vs ECM hydrogel

Control Sol ECM

Images captured by JG Wang, R. M. & Christman, K. L. Adv. Drug Deliv. Rev. 2015

Acknowledgements Christman Lab

◦ Dr. Karen Christman◦ Dr. Mary Nguyen◦ Dr. Roberto Gaetani◦ Masaki Fujita◦ Rebecca Barden◦ Jessica Ungerleider◦ Melissa Hernandez◦ Raymond Wang◦ Heinz Strassle◦ Hillary Lam◦ Cori Espelien◦ Julian Garcia

MSTP-SURF Program◦ Dr. Robert Ross◦ Ryan Moore◦ Mary Alice Kiisel◦ Jennifer Dumdie◦ Aran Groves◦ Cindy Liu◦ Eulanca Liu

MSTP-SURF Participants◦ Pamela Capellan◦ Angel Diaz◦ Luis Gasca◦ Maryam Ige

◦ Kamren Livingston◦ Mauricio Mrquez

Palencia◦ Isaac Padilla◦ Assata Pyatt◦ Jasmine Rice◦ Ceciley Scarbrough