MITOCHONDRIAL COMPLEX 1 ACTIVITY IN MYOCARDIAL-SPECIFIC CHF1 DEFICIENT MICE
Jeffrey Gordon11/23/2010
CHF1 – A Brief Overview
• CHF1 is a basic-HLH transcription repressor which plays an important role in cardiovascular development
• CHF1 deficient mice are more susceptible to heart failure
• CHF1 myocardial transgenic mice are less susceptible to heart failure
My Project
• Hypothesis: differences of mitochondrial complex 1 activity may explain the phenotypic difference in susceptibility to heart failure in the myocardial specific CHF1 deficient mouse model
• Why investigate complex 1?
• Who cares / to what end?
In heart failure, decreased mitochondrial content and overall mitochondrial dysfunction exist
There are increased RNA transcripts of complex 1 in CHF1 transgenic mice
Preliminary data suggests decreased mitochondrial respiration in CHF1 deficient mice
Complex I is a manageable target
Why Test Complex 1 ?
Mitochondria
Mitochondria generate most of a cell’s ATP.
Mitochondria have a more broad function Made of 4 complexes and ATP synthase
http://en.wikipedia.org/wiki/File:ETC_electron_transport_chain.svg
Complex I
Largest complex – 45 different polypeptides
Redox reaction –
NADH + H(+) + ubiquinone (Q)
NAD(+) + dihydroubiquinone (QH2)
Who Cares / to what end?
Further characterize the mechanism responsible for increased susceptibility to heart failure of CHF1 deficient mice
Further understand mitochondria, which may offer therapeutic targets in the future
Overview of My Project
1. Breed wild-type controls and myocardial-specific CHF1 deficient mice
2. Isolate left ventricular mitochondrial tissue
3. Measure complex 1 activity via spectrophotometry
4. Normalize this to total mitochondrial activity
5. Compare the two groups
Spectrophotometry for Complex 1 Beer - Lambert Law Measure light absorbance of a material
to calculate concentration. For complex 1:
NADH + H(+) + ubiquinone (Q)
NAD(+) + dihydroubiquinone (QH2)
http://www.adinstruments.com/solutions/experiments/labchart_experiments/full.php?exp_id=187§ion_id=13&name_id=245&template=teaching
Results
Genotype C1 activity C1/CSF/F aMHC-Cre 639.34 1.602F/F aMHC-Cre 567.73 1.836F/F aMHC-Cre 255.73 2.178F/F aMHC-Cre 506.35 1.61F/F aMHC-Cre 673.43 1.757F/F 427.93 1.932F/F 603.53 2.168F/F 499.54 2.055F/F 368.26 1.921F/F 334.16 1.802F/F 339.27 1.857F/F 521.7 2.3F/F 663.21 1.985F/F 579.24 1.428
-C1 = complex 1. CS = citrate synthase. -C1 Activity measured in nm/ (min *mg)
Results
0
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Complex 1 Activity
CHF1 DeficientWT
P Value: 0.314564499
Conclusions
Complex 1 activity is not significantly different between wild-type and myocardial specific CHF-1 deficient mice
Thus, differences in complex 1 are unlikely to explain the increased susceptibility to heart failure between these groups
Future Avenues
CHF1 deficient mice have abnormal with calcium signaling and apoptosis. Further studies into mitochondrial dysfunction may lead to better understandings of these mechanisms.
