In the present study the effects of type 2 diabetes on coronary artery disease and cardiac function were investigated in 18-month-old mice after three months of Western diet. We found notable stenosis in proximal coronary arteries in both LDLR-/-ApoB100/100 and diabetic IGF-II/LDLR-/-ApoB100/100 mice and the stenoses were most severe in the left coronary artery, which was significantly narrowed in all animals of these groups despite compensatory vessel remodeling. Since type 2 diabetes increases significantly the risk of coronary events [2, 25], we wanted to study if the cardiac outcome would be worse in the diabetic animals. Regardless of increased intimal calcification in aortic lesions of the IGF-II/LDLR-/-ApoB100/100 mice, diabetes did not lead to a worsened CAD compared to hypercholesterolemic LDLR-/-ApoB100/100 mice. Also LV function was equally impaired in both hypercholesterolemic mouse strains compared to age- and diet-matched C57Bl/6J mice. Based on these findings the impaired cardiac function in IGF-II/LDLR-/-ApoB100/100 mice does not seem to be related to diabetes. This is likely subsequent to the high plasma cholesterol levels of the LDLR-/-ApoB100/100 background in both models, since it is widely recognized that cholesterol is the main driving force behind atherogenesis in mice. However, because the cholesterol levels in our study were equal in diabetic and non-diabetic LDLR-/-ApoB100/100 mice, detecting the effects of diabetes is possible, whereas the diabetes-induced elevation in plasma lipids seen in most models of diabetic atherosclerosis hampers dissecting the contribution of other metabolic factors on cardiovascular outcome .
Although genetically-modified mouse models of hypercholesterolemia are widely used in cardiovascular research [26, 27], the status of coronary arteries and potential manifestation of myocardial infarction have been studied to a lesser extent. In the commonly used LDLR-/- mice CAD seems to be absent even if atherogenic diet is used . Lesions in coronary arteries and myocardial fibrosis were described in Western diet fed ApoE-/- mice shortly after the model was generated , but since then controversial results have been reported: coronary artery occlusion and subsequent evidence of apical infarction are found by some , whereas in other studies the coronary arteries are found to be totally clean  or lesions are seen only in proximal parts of the arteries without signs of infracted myocardium [31–33]. The latter has been found also in the LDLR-/-ApoB100/100 mice by Saraste et al.  and verified in the present study. In the severely hypercholesterolemic ApoE-/-LDLR-/- mice both distal coronary artery lesions and myocardial infarction have been reported [35, 36]. The gravest phenotype is in double knockout mice lacking scavenger receptor class B type I (SR-BI) and ApoE - the mice die of CAD induced myocardial infarction by 8 weeks of age .
Positive or outward arterial remodeling is a compensatory increase in vessel size in order to maintain luminal area in atherosclerosis . Whereas clinical studies on vascular remodeling in different disease states are abundant, less is known about it in experimental models, especially mice. Positive remodeling has been observed in aortic sinus  as well as in carotid, thoracic and abdominal aortas of ApoE-/- mice [24, 40]. Consistent with this data, our results show enlargement of vessel areas in both aortic sinus and coronary arteries. In aortic sinus this compensatory remodeling resulted in maintenance of a lumen area equal to non-atherosclerotic C57Bl/6J controls, whereas in left coronary artery the lumen was significantly narrowed. Since stenosis was greater in coronary arteries (~80%) than in the aortic sinus (~60%), this might reflect a similar observation made in humans: once certain threshold in plaque deposition is exceeded, further enlargement might not be possible and lumen area is compromised by the growing lesion . To our knowledge, vascular remodeling has been previously studied neither in mouse coronary arteries nor in a model of diabetic atherosclerosis. Because of the increased risk for cardiac events in diabetes, this is a matter of interest. Association between positive remodeling and unstable lesions has been suggested [41, 42] and therefore it could be speculated that outward remodeling would be greater in diabetic patients with accelerated atherosclerosis and thus possibly also in the diabetic IGF-II/LDLR-/-ApoB100/100 mice with a more inflammatory vascular phenotype and advanced lesions . On the other hand, also negative remodeling could contribute to the increased incidence of cardiac events in diabetics. However, in the present study diabetes did not lead to either of these but the remodeling was equal compared to non-diabetic mice with similar atherosclerosis. Inconsistent findings have been done also in humans: both increased  and inadequate compensatory remodeling  have been detected with intravascular ultrasound, calling for more in-depth studies in experimental models.
Atherosclerosis as a progressive disease and aging is associated with general cardiovascular changes . Compared to our former characterization of 15-month-old animals , older age of mice in the present study reflected as an analogous augmentation of atherosclerotic calcification in both diabetic and non-diabetic LDLR-/-ApoB100/100 mice. Also plasma triglyceride levels of the IGF-II/LDLR-/-ApoB100/100 mice were now higher than in LDLR-/-ApoB100/100 mice and could in theory contribute to increased calcification. However, this is unlikely, since the level was still in the normal range and numerically only marginally higher than in the LDLR-/-ApoB100/100 mice. In addition, in our previous study the situation was vice versa and yet there was more calcification in the IGF-II/LDLR-/-ApoB100/100 mice. Thus the difference in plasma triglycerides most probably describes natural fluctuation. Moreover, no significant correlation between lipid levels and level of calcification were noted. Therefore the physiological significance of minor changes in triglycerides is unlikely, especially in the presence of severe hypercholesterolemia.
Despite prominent stenosis in proximal coronary arteries and observed regional akinesia in LVAW of LDLR-/-ApoB100/100 and IGF-II/LDLR-/-ApoB100/100 mice, signs of myocardial infarction were not found. Thus, the observed severe LV dysfunction most likely results from adaptation to chronic hypoperfusion rather than acute ischemic events. In support of this the resting perfusion in LVAW was only slightly reduced, which is in line with the study observing reduced coronary flow reserve in old, Western diet fed LDLR-/-ApoB100/100 mice . Cardiac functional reserve after dobutamine challenge was preserved, although depressed compared to C57Bl/6 controls, indicating that the dysfunctional myocardium of LDLR-/-ApoB100/100 and IGF-II/LDLR-/-ApoB100/100 mice was not scarred but viable. Dobutamine stress echocardiography is a feasible non-invasive method to gain information about cardiac function and capacity, and to predict reversibility of wall motion after revascularization . In ApoE-/- mice the cardiac functional reserve has been found to be reduced . Also in diabetic db/db  and ob/ob mice  the response to β-adrenergic stimulation has been shown to be decreased. So far the only study done in a mouse model of diabetic atherosclerosis showed impaired cardiac reserve in the ob/ob/LDLR-/- mice .
In patients with ischemic cardiomyopathy, hibernating myocardium is an important clinical entity with significant prognostic value when considering revascularization procedures. Myocardial hibernation is referred as a state of reduced contractile function in the setting of CAD that is reversible with revascularization. Interestingly, various results in the present study suggest myocardial hibernation: LV dysfunction distal to coronary stenosis, preserved inotropic reserve, absence of necrosis and minor reduction in myocardial perfusion. Also the ultrastructural findings in LDLR-/-ApoB100/100 and IGF-II/LDLR-/-ApoB100/100 mice support the possibility of hibernation, since the morphological hallmarks of chronic myocardial hibernation include signs of atrophy (especially contractile myofibrils and loss of cardiomyocytes) and degenerative changes (disorganization of cytoskeleton, cellular debris in interstitium, increased collagen and glycogen) without necrosis . As a limitation to the hypothesis of myocardial hibernation, some of the changes seen in LDLR-/-ApoB100/100 and IGF-II/LDLR-/-ApoB100/100 mice could be induced also by aging. However, there is a general lack of suitable animal models that has hampered the studies of myocardial hibernation. Because achieving reproducible and chronic partial coronary artery stenosis without infarction is technically very challenging, most experimental studies of myocardial hibernation have been done in larger animals, such as dogs and pigs [49, 50]. The few mouse models reported are mainly based a short-term hypoperfusion  or genetic manipulation of e.g. myocardial angiogenesis . Since chronic CAD and the resulting gradual intravascular occlusion are the major causes of ischemic cardiomyopathy in humans, it would be aspired to have them manifested also in the experimental animal model. Even if the model would not perfectly reproduce the human condition and the actual revascularization procedures as an affirmation of the hibernation are not technically feasible in mice, animal models can provide unique and valuable information.