Dissertation Defense Announcement
To:  The George Mason University Community

Candidate: Rashmi Kumar
Program:    PhD Bioinformatics & Computational Biology

Date:   Thursday August 14, 2012
Time:   11:00 a.m.
Place:  George Mason University
           Occoquan Bldg., #204
           Prince William Campus
Dissertation Director/Committee Chair: Dr. M. Saleet Jafri
Committee members:
Dr. Dmitri Klimov, Dr. Patrick Gillevet, Dr. Siddhartha Sikdar

Title: "A Computational Analysis Of Mitochondrial Reactive Oxygen Species Dynamics In Cardiomyocytes"

The dissertation is on reserve in the Johnson Center Library, Fairfax campus.
The doctoral project will not be read at the meeting, but should be read in advance.

All members of the George Mason University community are invited to attend.


Mitochondria play an important role in the maintenance of ionic homeostasis and the control of ATP production, cellular redox potential and reactive oxygen species (ROS) production. Low levels of ROS are generated as a byproduct of energy metabolism by mitochondria. Experiments indicate that the two important sites contributing to ROS production are the Complex I (NADH: ubiquinone oxidoreductase) and Complex III (Ubiquinol: cytochrome C oxidoreductase) of mitochondrial respiratory chain.  These highly reactive molecules in excess can lead to oxidative stress and is linked to multiple pathological conditions like diabetes, neurodegenerative diseases and ischemia reperfusion. To ameliorate this risk, several antioxidant defenses play an important role in maintaining the redox balance. To better understand the complex system regulating ROS, we present a mechanistic model describing ROS production across the electron transport chain (ETC) as well as description of the pathways for the dissipation of ROS. The model incorporates detailed biochemical kinetics for electron fluxes across tricarboxylic acid cycle (TCA), mitochondrial calcium ([Ca2+]m) handling, membrane  ion transport processes. This is coupled to the mechanism of electron transfer across respiratory complex I, II and III, oxidative phosphorylation and  generation of ROS by complexes I and III. Additionally, the computational model presents a very detailed mechanism of various matrix and extramitochondrial antioxidant defenses (described by GSH/GSSG and NADPH/NADP+ redox pairs) and regulation of mitochondrial permeability transition pore (MPTP) dynamics by ROS. 

The model enables study of factors regulating ROS production and provides the first insights into the mechanism of succinate induced ROS production with increasing membrane potential (ΔѰm) which is quite distinct from the mechanism of reverse electron transport (RET) proposed by others. The association of substrate nature and presence of inhibitors in regulating ROS production is presented in this study.
The model suggests the mechanisms of mitochondrial acidification (pHm) in response to cytosolic acidification (pHe), and predicts an important role of mitochondrial respiration driven proton pumps in maintaining pHm in addition to the role of potassium-hydrogen (KHEm) exchanger suggested previously by others. Results from model also indicate that changes in extramitochondrial pH regulate ΔѰm, Ca2+ and ROS. The model identifies the factors modulating excess ROS formation during elevated [Na+]i, as observed during conditions of heart failure. The reduction in glutathione redox couple ratio (GSH/GSSG) is observed in parallel with progressive ΔѰm depolarization and increase in ROS levels in the model.
During experiments a transient increase in ROS levels has been observed concurrent with a decrease in membrane potential, however, the mechanism behind these remains controversial.  Others have proposed that there is positive feedback of ROS on ROS production (ROS-induced ROS release).  Our mechanistic model, however proposes an alternative as ROS production must decrease with decreasing membrane potential.  The model simulates ROS generation in the mitochondria due to laser induced photoactivation of TMRM. Under certain conditions a transient increase in ROS levels is observed that is termed a ROS burst.  It is hypothesized that this increase in mitochondrial ROS can lead to opening of MPTP which releases this burst of stored up ROS. The opening of MPTP leads to the decrease in mitochondrial membrane potential accompanying the ROS burst. The simulation results are qualitatively similar to those shown in experiments using isolated adult cardiac myocytes offering an alternative explanation for experimentally observed phenomenon of ROS burst in cardiac myocytes.