1. Investigation of the Impact of Metformin on Metabolism and Function in Cardiac Ischaemia
and Reperfusion
Sophie Oakley, Department of Biological Science, University of Hull, Hull
INTRODUCTION
Ischaemia can come in many forms and varying degrees of severity but
ultimately leads to death of the myocardium and accounts for the morbidity
and mortality of coronary heart disease (1)
.
Damage caused by ischaemia is determined by the degree and length of
the disproportionate oxygen supply and demand. However the act of
reperfusing the heart comes with it's own consequences and exacerbates
the injury caused by ischaemia (2)
.
Cardioprotective agents such as Metformin aim to reduce the damage
caused my ischaemia-reperfusion injury. Thus leading to the hypothesis
that Metformin should show evidence of cardioprotection (3)
.
AIM
To investigate the potential cardioportive effects of Metformin on metabolism
and function during ischaemia reperfusion.
METHODS
Using male Sprague-Dawley rats, hearts were isolated and perfused in a
modifed isovolumic retrograde Langendorff mode using the apparatus in
Figure 1 (4)
.
Hearts were perfused with Krebs-Henseleit buffer containing (NaCl 118mM;
NaHCO3
25mM; KCl 4.5mM; KH2
PO4
1.2mM; MgSO4
◦ 7H2
O 25mM; CaCl2
◦
H2
O 1.25mM; Glucose 5mM; Palmitate 3mM; Lactate 1mM; Pyruvate 0.1mM;
Glutamine 0.55mM. 1MM of Metformin was added in the Metformin model
noted in Figure 2..(4)
PPARα expression was analysed by sodium dodecyl sulphate polyacrylamide
gel electrophoresis (SDS PAGE) and western blotting. Actin was used to
normalise the optical density values. (5)
Results are presented as the means ±SEM where appropriate with any
statistical significance identified using an unpaired student's T test for
statistical analysis.
Figure 1: The Langendorff perfusion apparatus. Figure 2:Control Perfusion Protocol in the absence and
presence of Metformin.
RESULTS
Figure 7 : Cessation of function (T1), Onset of Contracture (T2) and Time till max contracture
in the absence and presence of Metformin. N=2 in both groups.
Figure 3: Recorded trace of cardiac function. Diastolic Pressure (DP),
Systolic Pressure (SP), Left Ventricular Pressure (LVDP)
Figure 5: LVDP at different stages of the protocol in either the absence or
presence of Metformin. N=3 in each group.
CONCLUSIONS
During both normoxia and reperfusion no significant change was
seen between the control and normoxia models (Figure 4). This
was inconsistant with other studies(6)
suggesting that either the
sample size (n=3) was too small or the concentration of
Metformin (1mM) was not sufficient to elicite a change.
During ischaemia Metformin accelerated the cessation of function
indicating Metformin might affect contractile function within the
heart.
Metformin delayed the onset of contracture within ischaemia but
accelerated the time till max contracture and the extent of
contracture suggesting that Metformin reduced contractile injury.
Metformin increased expression of PPARα in reperfusion only
indicating that these changes may take longer than the 10mins in
normoxia to become apparent or an event during ischaemia -
reperfusion promoted this action.
Myocardial Contractile Function
Table 1.Myocardial Function in the presence and absence of Metformin
during normoxia.
Cardiac Function During Ischaemia
PPARα expression
Figure 6: Recorded trace of cardiac function during ischaemia.
Time till cessation of function (T1), onset of contracture (T2), time
till max contracture (T3) and extent of contracture (E1).
REFERENCES
Butany,J, Carlson,A, Chicoine,A, Courties,G,Dutta,P, Etzrodt,M Gorbatov,R,Heidt,T,
Iwamoto,Y, Katus,H, Lasitschka,F Leuschner,F, Libby,P, Lin,C, Majmudar, Moskowitz,M,
Morrow,D, Murphy,S,Nahrendorf,M, Niessen,M, Piek,J, Pittet,M, M, Robbins,C,
Rubin,B,Sabatine,M, Stone,J, Swirski,F, Thompson,B,Van der laan, A, Vinegoni,C,
Waring,M Waterman,P, Wei,Y and Weissleder,R. (2012) Myocardial infarction accelerates
athersclerosis. Nature, 111260.
Seymour, AM. (2013, October 16
th
). Cardioprotection 2.[PowerPoint slides]. Presented at
Clinical Chemistry lecture at Hull University.
Boengler,K, Heusch,G & Schulz,R. (2010) Inhibition of mitochondrial permability transition
pore opening: the holy grail or cardioprotection. Basic Research in Cardiology, 105,
(2),p151-154.
Cleland,J, Sample,J and Seymour,AM. (2006) Metabolic remodelling in the aging heart.
Journal of Molecular and Cellular Cardiology, 40, (1), p56-63.
Smith,K. (2006)The Impact of Erythropoietin on Uraemic Cardiomyopathy. PhD Thesis, Hull
University
.Barreto-Torres,G, Jaradov,S and Parodi-Rullan,R. (2012) The Role of PPAR α in
Metformin-induced attenuation of mitochondrial dysfunction in acute cardiac
ischemia/reperfuion in rats. International Journal of Molecular Sciences, 13, (6), p7694-
7709.
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2.
3.
4.
5.
6.
ACKNOWLEDGEMENTS
Many thanks and much gratitude goes to my supervisor Dr
Anne-Marie Seymour. Without her support and guidance this
project would not have been possible. Thank you Kath Bulmer,
your invaluable assistance and vast practical knowledge which
led to the success of both the perfusions and the analytical
processing.