High Density Lipoprotein Cholesterol & Cardioprotection: Role of Sphingosine-1-Phosphate
Dr Sarah Pedretti (Postdoctoral Fellow)
Supervisor: Assoc Prof. Sandrine Lecour
Year project started: 2011
Main purpose of study:
A recent World Health Organization report warns of the escalating global burden of cardiovascular diseases (CVD), projecting that it will become the major worldwide cause of death and disability by 2020. In the Western Cape region of South Africa, ischemic disease is already the leading cause of death, often killing by acute myocardial infarction. Surprisingly, the African black population from the heart of Soweto study presents a high prevalence of risk factors for atherosclerotic disease including diabetes but overall, ischemic heart disease apparently remains fairly uncommon compared with the other South African ethnic groups. This relative protective effect may be related to the rarity of hypercholesterolemia in the African black population as they may have a higher level of high density lipoprotein (HDL) relative to low density lipoprotein (LDL) than non-black South Africans.
Interestingly, a variety of studies exploring the mechanism, suggest a protective effect of HDL against ischemic heart disease. Well known for its ability to limit the formation of atherosclerotic plaque by favoring the cholesterol transport back to the liver, HDL has other beneficial cardiac effects including anti-inflammatory and anti-oxidant effects. However, the cardioprotective components of HDL and its exact mechanisms of cardiovascular protection remain unclear.
We have identified sphingosine-1-phosphate (S1P), a major component of HDL, as a powerful cardioprotective sphingolipid that limits ischemia-reperfusion injury.
Our preliminary work suggests that this protective effect is mediated via the activation of a prosurvival signaling pathway that we have newly discovered in the heart and that we have named the Survivor Activating Factor Enhancement (SAFE) pathway.
Since I started on that project I have demonstrated that, in a model of isolated adult mouse cardiomyocytes subjected to simulated ischemia, HDL/S1P cardioprotection is mediated via the SAFE pathway and subsequent inhibition of mitochondrial permeability transition pore.
The principal objective of our study is to evaluate the role of S1P in the cardioprotective capacity of HDL to counteract myocardial injury induced by ischemia/reperfusion in vitro (isolated perfused hearts or isolated cardiomyocytes). We will also address the importance of the SAFE pathway in S1P/HDL induced cardioprotection.
Experiments will be performed in isolated mouse cardiomyocytes and perfused hearts that will be subjected to an ischemia/reperfusion insult.
We have 2 main research aims:
1. To delineate the role of Sphingosine Kinases (SphK) in S1P-induced protection in hearts subjected to ischemia/reperfusion
SphK are the enzymes that catalyse the formation of endogenous S1P.
Using an isolated perfused heart model, hearts extracted from wildtype mice will be subjected to an ischemia/reperfusion insult with/without S1P and inhibitors of SphK. Measurements of SphK activities and S1P content in isolated hearts will be used as an endpoint.
2. To evaluate the mechanisms involved in HDL/S1P-induced cardioprotection
The prosurvival transcription factor signal transducer and activator of transcription 3 (STAT3) and tumor necrosis factor α (TNFα) are the 2 main components of the SAFE pathway.
To delineate whether the SAFE pathway is activated by S1P in HDL-induced cardioprotection, cardiomyocytes extracted from STAT3 and TNFα deficient mice will be subjected to a simulated ischemia/reperfusion insult with/without reconstituted HDL containing various concentrations of S1P. Western blot analysis will also be performed as an endpoint.
Papers published from study:
M. Brulhart-Meynet, V. Braunersreuther, J. Brinck, F. Montecucco, K. Galan, G. Pelli, S. Pedretti, F. Mach, S. Lecour, R.W James, M.A Frias “Post-ischemic treatment with native and reconstituted HDL limit reperfusion injury”, Basic Research in Cardiology, in revision
R.F Kelly-Laubscher, J.C King, D. Hacking, S. Somers, S. Pedretti, G. Maarman and S. Lecour «Preconditioning with sphingosine-1-phosphate requires activation of signal transducer and activator of transcription-3», Acta Physiologica, in revision
M.A Frias, S. Pedretti, D. Hacking, S. Somers, L. Lacerda, L.H Opie,R.W James and S. Lecour «HDL protects against ischemia reperfusion injury by preserving mitochondrial integrity», Atherosclerosis, May 2013; 228 (1):110-6
M.A Frias, S. Lecour, R.W James and S. Pedretti «High density lipoprotein/sphingosine-1 phosphate-induced cardioprotection: Role of STAT3 as part of the SAFE pathway», JAK/STAT, April 2012; 1 (2)
Techniques used in project:
- Isolation of adult mouse cardiomyocyte (in vitro)
- Measure of cell viability by trypan blue staining
- Measure of mPTP opening by TMRM fluorescence (FACS)
- Langendorff isolated perfused heart model (ex vivo)
- Measurement of infarct size
- Western blot: protein measurement
- Measurements of SphK activities
- Measurement of S1P content