Using of the animal models to shed the light on the molecular mechanism of Atherosclerosis
Atherosclerosis is a disease of the arterial blood vessel due to accumulation of lipid in the wall of artery (Donners, 2004). It leads to the narrowing of the lumen of the artery and restriction of blood flow this called atherosclerotic plaque. There are many risk factors for atherosclerosis such as smoking, hypertension, diabetes mellitus, hypercholesterolemia, physical inactivity and stressful life. Also, high level of LDL is another risk factor for atherosclerosis. Atherosclerosis can affect any artery in the body, for example, arteries in the brain, heart, arm, legs and pelvis. It is caused by a response to damage to the endothelium from high cholesterol and high blood pressure and smoking. In addition, there are two mechanism to understand the mechanism of atherosclerosis, inflammatory mechanism and molecular mechanism. In molecular mechanism damage to endothelium lead to fatty streak formation by uptake of LDL to intima and then LDL is oxidized and it is taken by scavenger receptors on macrophage and finally leads to formation of foam cells that are died by apoptosis and accumulated. Moreover, vascular smooth muscle cells have a role in atherosclerosis mechanism in formation of fibrous cap. This essay will discuss the animal models that shed light on molecular mechanism of atherosclerosis.
Using rabbits as models to study atherosclerosis
Rabbits are used widely to study atherosclerosis because it has demonstrated that rabbits develop atherosclerosis within short period after eating high cholesterol diet (Yanni,2003). There is more than one type of rabbit but the most used one in the lab is New Zealand type (Yann, 2004). In this type of rabbit the normal cholesterol level is low but increase from 2 to 8 times after ingestion of high cholesterol diet (Bocan etal.1993). Moreover, after introduce the cholesterol to the rabbits the foam cells will produce and the early lesion are similar to fatty streak in human. However, this lesion will not develop to plaques which are a characteristic feature of human atherosclerosis. The work of Adams shows that Rabbit can develop atherosclerotic lesion similar to human lesion in late stage when they ingest low diet cholesterol for long period. Moreover, using of balloon injury in abdominal and thoracic aorta together with high cholesterol diet leads to formation of atherosclerotic lesion similar to human lesion (Aikawa et al,1998).
Transgenic rabbit models
Using of transgenic rabbit models is a new area to study molecular mechanism of atherosclerosis. Because high level of lipoprotein is consider as risk factor for atherosclerosis and it is associated with increase level of low density lipoprotein. So to study the relation between increase level of lipoprotein a and development of atherosclerosis the researchers generate watnabe heritable hyperlipidemia (WHHL) rabbits (Sun etal,2002) expressing human apolipoprotein(a).To demonstrate how lipoprotein(a) can induce vascular calcification , the researchers study the effect of human lipoprotein a on smooth muscle cells of cultured aortic rabbit and they established that there is increase in alkaline phosphatase activity in treated smooth muscle cells and that lead to accumulation of calcium. So it was estimated that LP (a) lead to formation of advanced atherosclerotic lesion. In this type of transgenic rabbits expresses human apolipoprotein a (Yanni.2004) lead to myocardial infarction as result of extensive atherosclerotic lesion in coronary artery in transgenic rabbit compared to control group. Another model is transgenic rabbits over expressing human apo-B-100and it was shown that plasma lipid such as cholesterol and triglyceride is increasing 2-3 times more than control group while HDL cholesterol is decreased (Fan etal.1995).
Using of mouse models to study molecular mechanism of atherosclerosis
-Diet induced model
This model is done by administrating diet containing 15% fat for C57BL/6J mice for a period of 10 weeks so that lead to development of atherosclerotic lesion. The drawback of using this model is that the lesion occupies the aortic root only so that make reduction in using this model to study the pathogenesis of atherosclerosis and use of genetically engineered mice widely (Ohashi etal.2004).
- Apolipoprotein E knockout mice model
In this model the researchers used mice deficient in apo E gene and this model is produced for the first time in 1992 (Plump etal .1992).The most important function of apo E is to enhance the binding of lipoprotein to receptor so deficiency of apo E is associated with deposition of cholesterol ester containing particle. Moreover, there is increase in the level of cholesterol about 5 times more than normal and change in distribution of cholesterol containing plasma lipoprotein where the level of very low density lipoprotein is increase and level of high density lipoprotein is decrease. Mice that lack apo E when they are administrated diet containing 4.5% fat, they develop atherosclerotic lesion that are similar to human atherosclerosis. Atherosclerotic lesion in this model spreads throughout the arterial tree. The disadvantage of using this model is that lipoprotein profiles in apo E knockout mice are different from lipoprotein profiles of people who subjects to have atherosclerotic lesion (Ohashi etal.2004).
- Low density lipoprotein receptor knockout mice
It is a genetically engineered model in which the researchers produced mice lacking low density lipoprotein receptor gene. After administration of 10% fat diet this mice develops hypercholesterolemia and there is 2 times increase in total cholesterol level (Ishibashi etal.1993) because high fat diet and lacking of LDLR lead to increase in LDL and VLDL levels. LDLR knockout mice is different from the Apo E knockout mice in that the lipid profiles in LDLR knockout mice are the same for human hyperlipidemia while in Apo E -/- are different(Ohashi etal.2004). LDLR-/- mice develops more advanced and more extensive atherosclerotic lesion than diet induced atherosclerotic model (Donners etal.2004).Moreover, there are many types of modified atherosclerotic models have been developed to produce atherosclerotic lesion more similar to human atherosclerotic lesion . These modifications are hypertension, inflammation, glucose metabolism, proteinase and immune system (Ohashi etal.2004). In general using mouse as model has advantage of low cost and the disadvantage of using it is that the size of blood vessele is not the same of human blood vessel (Shi etal.2009).
Using of mice lack endothelial nitric oxide synthase (e NOS) gene
Mice that lack this gene will develop hypertension because endothelial nitric oxide synthase controls blood pressure by production of NO. It has estimated that when e NOS-/- mice crossbred with mice lack apo E gene, the offspring will develop hypertension and atherosclerosis. Moreover, e NOS or NO have proatherosclerotic features by producing superoxide anions that will oxidize LDL and produce atherosclerosis (Ohashi, 2004).
Moreover, ascorbic acid is important for collagen stabilization so the researchers generate mice lack ascorbic acid synthesis (Gulo-/-mice) and this mice crossbred with apo E-/- mice, the offspring has the same atherosclerotic lesion with the less collage formation (Nakata and Maeda 2002).In addition, macrophages have an important role in early stage of atherosclerosis because they infiltrate vessel wall due to endothelial dysfunction. To demonstrate the role of macrophages in atherosclerosis, the researchers generate mice with mutation in macrophage colony stimulating factor gene (osteopetrotic mice) and that lead to reduction in number of macrophages.The work of Smith reveals that when osteopetrotic mice crossbred with apo E-/- mice ,the offspring develop small atherosclerotic lesion compared to the control apoE-/- mice
Using of other animal models such as pig , swine
Atherosclerotic lesion that induced in pig is similar to human atherosclerotic lesion because pig has blood pressure and heart rate closely similar to human. Also, the size of pig blood vessel is similar to the size of blood vessel in human. However, the using of pig as model is restricted because it is more expensive than small animals and it is difficult to handle (Donners etal.2004). In study that was done by Shi, the researchers produced atherosclerotic lesion in swine model by partial surgical ligation of carotid artery and administrating high fat and high cholesterol diet to the swine. This diet contains 4% cholesterol, 20% saturated fat and 1.5 supplemental choline. All the characteristic features of advanced human atherosclerotic lesion such as necrosis, calcification, ulceration, interplaque hemorrhage are demonstrated in swine model (Shi etal,2009). In some studies the researchers demonstrate that chicken develop hypercholesterolemia , thicking in intima of the aorta and a hug number of foam cells were accumulated in the intima of aorta after administration of 1.0% cholesterol containing diet for 3 or 6 months. While the chicken that eat 0.1%cholesterol containing diet, there is a few amount of foam cells accumulated in aorta. In coronary artery of chicken that eats 1.0% cholesterol containing diet, there is marked increase in number of foam cells and marked stenosis of it. So the researchers estimated that chicken is a good experimental model to study atherosclerosis.
Atherosclerosis is a serious disease and the incidence of the disease is increasing sharply. So the researches try to understand the pathology and molecular mechanism of the disease by developing different experimental models. These experimental models include diet induced mice model, LDLR-/- knockout mice, Apo E-/-knockout mice and using of large animal such as rabbit, swine, dog and chicken. The aim of develop these models is to produce a gold therapy to prevent mortality of the disease and to decrease the morbidity of the disease. Although these models have some drawbacks such as the cost of using pig and atherosclerotic lesion that produced in mice is different from human atherosclerotic lesion, the cardiovascular researches has a great benefit from using these models.
Aikawa, M., Rabkin,e., Okada, Y., Voglic, S.J., et al.(1998). Lipid lowering by diet reduces matrix metalloproteinase activity and increase collagen content of rabbit atheroma ; a potential mechanism of lesion stabilization. Circulation,97,2433-2444.
Adams, C.W., Miller, N.,E., Morgan R., S.,& Rao, S.,N.(1982). Lipoprotein level and tissue lipids in fatty-fibrous atherosclerosis induced in rabbits by two year's cholesterol feeding at a low level. Atherosclerosis, 44, 1-8.
Bocan, T. M., Mueller, S.B., Mazur, M. J.,Uhlendorf, P.D., BBrown, E. Q., Kieft, K.A.(1993). The relationship between the degree of dietary- induced hypercholesterolemia in the rabbit and atherosclerotic lesion formation. Atherosclerosis,102,9-22.
Donners, M.M., Heeneman, S.,& Daemen,M.J.(2004). Models of atherosclerosis and transplant arteriosclerosis :the quest for the best. Elsevier 1(3),257-263.
Fan,J.,McCormick, S.P., Krauss R. M., Taylor, S., et al. (1995). Overexpression of human apolipoprotein B-100 in transgenic rabbits results in increased levels of LDL and decreased levels of HDL. Atherosclerosis Thrombosis and vascular Biology,15,1889-1899.
Ishibashi, S., Brown, M.S., Goldstein, JL., etal. (1993). Hypercholesterolemia in low density lipoprotein receptor knockout mice and its reversal by adenovirus-mediated gene delivery. J Clin Invest ,92:883-893.
Kolodgie, F. D., Katocs, A.S., Largis, E.E.,Wrenn S.M.,Cornhill, J.F., Herderick ,E.E.,LEE,S.J., &Virmani ,R.(1996). Hypercholesterolemia in the rabbit induced by feeding graded amounts of low level cholesterol. Atherosclerosis Thrombosis and Vascular Biology,16,1454-64.
Nakata, Y., Maeda, N. (2002). Vulnerable atherosclerotic plaque morphology in apolipoprotein E-deficient mice unable to make ascorbic acid. Circulation, 105, 1485-1490.
Ohashi, R., Mu, H., Yao, Q.,Chen ,C.(2004). Cellular and molecular mechanisms of atherosclerosis with mouse models. Trends Cardiovasc Med , 14,187-190.
Plump, A.S., Smith, J.D., Hayek, T., et al .(1998). Severe hypercholesterolemia and atherosclerosis in apolipoprotein E- deficient mice created by homologous recombination in ES cells. Cell, 71: 343-353.
Sun, H., Unoki, H., Wang, X., et al.(2002). Lipoprotein(a) enhances advanced atherosclerosis and vascular calcification in WHHL transgenic rabbits expressing human apolipoprotein (a). Journal of Biological Chemistry , 277(49),47486-47492.
Shi, Z.S., Feng, L., X., Ishii, A., Goldstine, J. et al. (2009). Vulnerable plaque in a swine model of carotid atherosclerosis . Ajnr Am J Neuroradiol, 30,469-472.
Smith, J.D.,Trogan,E., Ginsberg,M., etal.(1995). Decreased atherosclerosis in mice deficient in both macrophage colony-stimulating factor (op) and apolipoprotein E. Proc Natl Acad Sci USA 92,8264-8268.
Yanni, A.E., Yatzidis, H., Kavantazas, N., Agapitos, E., Perrea, D.,& Karayannacos, P.E. (2003). Dietary L-aspartate and L-aglutamate inhibit fatty streak initiation in cholesterol-fed rabbit. Nutrition Metabolism and Cardiovascular Diseases,13,80-6.
Yanni, A. E.(2004). The laboratory rabbit :an animal model of atherosclerosis research. Laboratory Animals ,38,246-256.