Mathav Vignesh
Pronouns: He/him
Research Mentor(s): Andrea Thompson
Research Mentor School/College/Department: Department of Internal Medicine/Cardiovascular Division / Medicine
Program: UROP
Session: Session 1 (9:00am – 9:50am)
Authors: Mathav Vignesh, Jaime Yob, Andrea Thompson
Abstract
Hypertrophic cardiomyopathy (HCM) is a condition of the heart that causes the walls of the left ventricle to thicken and become stiff. This can lead to heart failure, certain arrhythmias, and even cardiac arrest. The most common cause of HCM are loss-of-function variants in the MYBPC3 gene that result in a reduction of MyBP-C protein in patients with HCM. Previous research has shown that cardiomyocytes without any MyBP-C (MYBPC3 -/-) have a hypercontractile phenotype. However, the relationship of MyBP-C protein levels and cell contractility has not been well defined in the range typically seen in human disease (40-60% reduction in MyBP-C compared to wild-type controls). This study attempts to map how the level of MyBP-C affects contractility of cardiomyocytes. In addition to testing MYBPC3 -/- cardiomyocytes, we will test the effect of varying levels of MYBP-C protein on iPSC-CM derived from patients with wild-type MYBP-C, reduced level of wild-type MyBP-C (+/c.2373dupG), and disease-causing missense variants (+/R502W). Contractility is measured using a micropatterned 2-D monolayer on PDMS and full time-series image capture. Thus, this platform is well suited to focus on the effect of MyBP-C protein level on contractile kinetics while controlling for geometry, size, and myofibrillar organization. In this platform MyBP-C protein levels can be controlled by introduction of transgene FKBP*-Flag-MYBPC3 which is unstable in the absence of the small molecule Shield and expresses FKBP*-Flag-MyBP-C protein which localizes to the sarcomere in dose-dependent manner after treatment with shield. Thus, within the same cardiomyocyte 2-D monolayer contractility can be measured at varying concentrations of MyBP-C. Finally, 2-D monolayers of cardiomyocytes can be fixed and stained with immunofluorescence to match contractility data to MyBP-C expression levels. Thus far we have successfully completed micropatterning and contractile imaging for WT iPSC-CMs. While we were able to obtain contractile data, we noticed clumping of cells on top of our 2-D monolayers, suggesting cell density during plating should be reduced to enable 2-D monolayers in the future.
