Effects Ofβ₃-adrenoceptor Activation On The Atherosclerosis

BackgroundAtherosclerotic （AS） disease is the leading cause of morbidity and mortality inthe western world and developing countries. It is associated with a metabolic andvascular cluster of disorders. The major risk factors of atherosclerosis includehyperlipidemia, glucose intolerance, hypertension, smoking and aging.Hyperlipidemia is summarized as raised trigIyceride （TG）, total cholesterol （TC） andlow density lipoprotein-cholesteroI （LDL-C） levels, reduced （high densitylipoprotein-cholesteroI） HDL-C level. Studies indicate that reducing LDL-C levelsand/or increasing HDL-C levels can decrease efficiently the incidence of AS disease.Recently, the epidemiology showed that the single standard of LDL-C level can onlyreduce the30%to45%of major coronary events. AS the raised LDL-C level, thereduced HDL-C levels also have high cardiovascular event risk. The increase ofHDL-C level with every1%would reduce the incidence of cardiovascular eventswith2%to3%. HDL involves in reverse cholesterol transport （RCT） which HDLinhibites the aggregation of cholesteryl ester in macrophages and prevents theformation of foam cells in the plaque. In addition, HDL also has anti-inflammatory,antioxidant, anticoagulant and improves the endothelial function. In recent years, thenew medcines are actively developed which could elevated HDL levels and enhanceits functionality.As the most important anti—atherosclerotic mechanism in vivo RCT，is aprocess which dynamically transport of excess cholesterol from the periphery into theliver, followed by excretion in the feces or the bile．Scavenger receptor class B type I （SR-BI） had already proved in molecular level as the sole HDL receptor whichlocates on celluar membrane．As the HDL specific receptor, SR-BI plays an importantrole in the HDL metabolism and the process of RCT. Apolipoprotein A-I （ApoA-I） isthe basic structure area of HDL combinated with SR-B1. It is essential that theappropriate position and content of ApoA-I in order to work effectively in the processof RCT. In addition, peroxisome proliferator-activated receptors （PPARs） recently areone of the main mechanisms of anti-atherosclerotic. PPARs belong to the members ofthe nuclear receptor superfamily including PPARα, PPARβand PPARγ. PPARαregulates fatty acid metabolism. PPARγ involve in lipid metabolism, glucosemetabolism and inflammation process by affecting fat cells differentiation andstorage.Thus PPARs interfere with the development of atherosclerosis.During the1980s, the classification of β-adrenoceptor （β-AR） was modifiedfollowing the discovery of a third β-AR primarily in rat adipose tissue. β₃-ARmediates the major effects of adrenaline and noradrenaline in adipose tissues, such aslipolysis in white adipose tissue and thermogenesis in brown adipose tissue. Itbelongs to the G-protein coupled receptor family which involves the activation ofadenylyl cyclase, cAMP/protein kinase A （PKA） pathway and recruits a PI3-kinasepathway. Previous studies have shown that chronic treatments of β₃-AR agonists wereeffective in improving glycaemic control, insulin sensitivity and reducing plasma TG,free fatty acid levels and body weight in obese diabetic animals. However the effectsof β₃-AR on atherosclerosis disease are largely unknown.ObjectiveTo study the effects of β₃-adrenoceptor （β₃-AR） activation on lipid, glucosemetabolism, the β₃-AR mRNAexpressions in withe adipose tissue, ApoA-I, ApoA-II,SR-BI expressions in liver tissue and atherosclerotic plaque development in agedapolipoprotein E-deficient (ApoE^-/-) mice. The study discussed β₃-AR possible mechanisms and provided the evidenceand which its function of HDL metabolism，the RCT process and atherosderosis．MethodsTen10-week-old wild-type C57BL/6J mice （A group） were fed normal chow.Fifty10-week-old ApoE^-/-mice were fed high-fat diet. At36-week-old, ApoE^-/-micewere randomly given normal saline （B group）, atorvastatin （10mg kg-1d-1, C group）,β₃-AR agonist BRL37344（1.65μg kg-1, twice a week, D group）, BRL37344（3.30μg kg-1, twice a week, E group） and β₃-AR antagonist SR52390A （50μg kg-1,twice a week, F group） for12weeks. The serum lipid, glucose and insulin levels weremeasured. The protein expressions of ApoA-I, SR-BI in the liver tissue by westernblot．Real-time quantitative PCR was used to determine the expressions of ApoA-ImRNA, ApoA-II mRNA, SR-BI mRNAand PPARα mRNA in the liver tissue and theexpression of β₃-AR mRNA and PPARγ mRNA in withe adipose tissue of mice. Theatherosclerotic plaque area in the thoracic aortic was determined withHematoxy-Eosin stain, and level of fibrosis in the plaque was assessed with Massonstain.Results1. Compared with wild-type C57BL/6J mice, ApoE^-/-mice showed a significanthyperlipidemia, insulin résistance, the downregulation expression of β₃-AR mRNAand SR-BI （P <0.01）, the upregulation expression of ApoA-I, ApoA-II and PPARαmRNA （P <0.01）, PPARγ mRNA levels in adipose tissue was no significant changes（P＞0.05）, atherosclerotic plaque formation with obvious fibrosis in the thoracicaortic （P <0.01）.2. Compared withApoE^-/-control mice with saline, atorvastatin obviouslydecreased the serum levels ofTC, VLDL/LDL-C, HDL-C and Ins （P <0.01）, but hadno effect on TG and Glu （P＞0.05）. In additionally, atorvastatin significiantly upregulated the expression ofApoA-I, ApoA–II, SR-BI and PPARα in liver tissueand the expression of PPARγ mRNA in adipose tissue （P <0.01）. β₃-AR mRNAlevelsof adipose tissue did not change significantly （P>0.05）.Atorvastatin could decreasethe atherosclerotic plaque size （P <0.01）.3. Compared with ApoE^-/-control mice with saline, low or high dose β₃-ARagonist significantly decreased the serum levels of TG, Glu and Ins, and obviouslyelevated HDL-C level and HDL-C/TC ratio. Simultaneously, β₃-AR agonist coulddistinctly up-regulate β₃-AR mRNA, ApoA-I and SR-BI levels, down-regulateApoA-II and PPARα mRNA levels and decrease the atherosclerotic plaque size（P<0.01）. PPARγ mRNA levels of adipose tissue did not change significantly （P>0.05）. Effect of high dose β₃-AR agonist was significantly superior to low dose（P<0.01）.4. Compared with atorvastatin group, the effects of high dose β₃-AR agonistwere similar in decreasing the atherosclerotic plaque size（P>0.05）, but high doseβ₃-AR agonist was better than atorvastatin in increasing levels of HDL-C, HDL-C/TCand in reducing levels of insulin（P<0.01）. β₃-AR agonist obviously up-regulated theβ₃-AR expression and down-regulated the expression ofApoA-I, ApoA-II, SR-BI,PPARα and PPARγ （P<0.01）.5. Compared with ApoE^-/-control mice，β₃-AR antagonist had no effect on lipids、Glu、Ins and the atherosclerotic plaque size （P>0.05）. PPARγ mRNA levels wasincreased significiantly in β₃-AR antagonist group（P<0.01）. There was no obviouschange about ApoA-I, ApoA-II, SR-BI, β₃-AR and PPARα expression （P>0.05）. Conclusion1. β₃-AR activation can improve the aged ApoE^-/-mice lipid metabolism,reducing levels of TC, VLDL/LDL-C and TG，increasing level of HDL；2. β₃-AR activation can improve the aged ApoE^-/-mice glucose metabolism,reducing levels of glucose and insulin；3. β₃-AR activation can up-regulate levels of β₃-AR mRNAexpression in whiteadipose tissue, ApoA-I and SR-BI expression in the liver tissue；4. β₃-AR activation can down-regulate levels of ApoA-II and PPARα mRNAexpression in the liver tissue；5. β₃-AR activation had no obvious effect on PPARγ mRNA expression of theaged ApoE^-/-mice；6. β₃-AR activation can significantly decrease the atherosclerotic plaque sizeand play a role of anti-atherosclerosis.