European Journal of Pharmacology 748 (2015) 166–169
Contents lists available at ScienceDirect
European Journal of Pharmacology journal homepage: www.elsevier.com/locate/ejphar
Perspective
Apolipoprotein B, the villain in the drama? Qi Yu a,1, Yaping Zhang a,b,1, Cang-Bao Xu a,b,n a b
Institute of Basic and Translational Medicine, Xi'an Medical University, Shaanxi, Xi'an 710021, PR China Division of Experimental Vascular Research, Institute of Clinical Science in Lund, Lund University, BMC A13, SE-221 84 Lund, Sweden
art ic l e i nf o
a b s t r a c t
Article history: Accepted 20 August 2014 Available online 16 September 2014
Low-density lipoprotein (LDL) is the major atherogenic lipoprotein and the primary target of lipidlowering therapy for treating ischemic cardiovascular disease. Apolipoprotein B (apoB), an important structural component of LDL, plays a key role in cholesterol transport and removal in vascular wall. On the other hand, under pathological process, apoB interacts with the arterial wall to initiate the cascade of events that leads to atherosclerosis. However, interactions between apoB and vascular wall remain to be determined. Here, we address a pathological role of apoB per se and whole LDL particle in dysfunction of vascular endothelium and smooth muscle cells i.e. decreased endothelium-dependent vasodilation and increased receptor-mediated vasoconstriction. We intend to discuss: i) how apoB is responsible for the deleterious effects of LDL in the development of ischemic cardiovascular disease; ii) why vaccine based on peptides derived from apoB-100 is a promising therapy for treating ischemic cardiovascular disease, and iii) direct inhibition of apoB production should be a better therapeutic option than simple LDLcholesterol lowering therapy in the patients with severe hypercholesterolemia at high cardiovascular risk with statin intolerance. In conclusion, apoB, but not cholesterol, plays a major role in LDL-induced dysfunction of endothelium, suggesting that direct apoB-targeting agents might be a promising therapy for the treatment of ischemic cardiovascular disease. & 2014 Elsevier B.V. All rights reserved.
Keywords: Low-density lipoprotein Apolipoprotein B Cholesterol Vasoconstriction Vasodilation Ischemic cardiovascular disease
1. The “bad” cholesterol Atherosclerosis leads to ischemic cardiovascular disease, which is the major cause of mortality in the world. Low-density lipoprotein (LDL) is well-recognized to play a key role in the pathogenesis of atherosclerosis that is a chronic inflammatory disease of the arterial wall. The LDL particle consists of phospholipid, free cholesterol and apolipoprotein B (apoB) (Hevonoja et al., 2000). In contrast, highdensity lipoprotein (HDL) contains exchangeable apolipoproteins of the A, C and E families (Lund-Katz and Phillips, 2010). Under physiological conditions, LDL is a vehicle that transports cholesterol all over the body in order to maintain cell viability and to provide cholesterol for the synthesis of the steroid hormones (Grundy, 2002). However, under pathological conditions, fluid sheer stress affects the morphology of endothelial cells in regions of arterial branching or curvature, along with high plasma LDL, result in increased permeability of LDL to preferential sites (Chien, 2008). When LDL infiltrates into the artery wall, it can be modified to oxidized LDL (ox-LDL), and causes the lipid deposit in the intima with chronic inflammation in the
n
Corresponding author at: Institute of Basic and Translational Medicine, Xi'an Medical University, Shaanxi, Xi'an 710021, PR China. Tel./fax: þ 86 29 86177632. E-mail address: [email protected] (C.-B. Xu). 1 These authors equally contributed to this work. http://dx.doi.org/10.1016/j.ejphar.2014.08.037 0014-2999/& 2014 Elsevier B.V. All rights reserved.
arterial wall, which subsequently results in the formation of atherosclerotic plaques. In clinic, plasma LDL cholesterol levels in patients are found to be positively correlated with the risk of cardiovascular events in human populations and augment individual susceptibility to atherosclerosis and its complications (Boekholdt et al., 2012; Haimeur et al., 2013). Therefore, LDL cholesterol is often called the “bad” cholesterol, and has been as the primary target of lipid-lowering therapy for treating ischemic cardiovascular disease (Libby et al., 2011). However, LDL is more than just cholesterol, and the contribution of LDL to cardiovascular disease risk involves more than just the cholesterol effects contained in LDL. Almost all cells of the artery wall have the ability to pick up LDL particles through the LDL receptors. Accumulation of LDL in the vasculature can easily undergo oxidative modification catalyzed by any of the major cell types of artery wall (Steinberg, 1997). Although many studies pay more attention to interactions between LDL and its receptors on cells in the arterial wall, the molecular mechanism that how LDL leads to dysfunction of vascular endothelium and smooth muscle cells remain not fully understood. 2. Effects of LDL on receptor-mediated vasoconstriction and endothelium-dependent vasodilation Atherosclerosis is a chronic disease characterized by the accumulation of lipoprotein lipid within the arterial wall. LDL is one of
Q. Yu et al. / European Journal of Pharmacology 748 (2015) 166–169
lipoproteins containing apoB-100, which makes LDL to be recognized and captured by LDL receptors on the cells in the arterial wall, but this does not induce cholesterol accumulation in monocyte/macrophages and transform into foam cells in the arterial intima. CD36 and scavenger receptor class A are mainly responsible for uptake of lipoprotein-derived cholesterol by macrophages, and the macrophages to be transformed into lipid-laden foam cells (Yu et al., 2013), which induces lipid deposition in and damage to the arterial wall. Since the discovery of endothelin-1 (ET-1) is a powerful endogenous vasoconstrictor, there have been intensive studies regarding the involvement of ET-1 and its receptors in the pathophysiology of ischemic cardiovascular diseases. In mammals, two different subtypes of endothelin receptors are known as the endothelin type A (ETA) and type B (ETB) receptors. The ETA receptors are located on the smooth muscle cells and mediate contractile effect, while the ETB receptors are limitedly expressed on the endothelial cells and respond with vasodilation (Xu et al., 2010). However, under pathological conditions like hypertension (Cao et al., 2013; Nilsson et al., 2008) and cerebral ischemia (Stenman et al., 2002), the ETB receptors can express on the
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smooth muscle cells and mediates strong vasoconstriction, suggesting the vasoconstrictor ETB receptor expression is inducible and pathogenic. Incubation of the arterial segments with native LDL for 24 h induces transcriptional upregulation of vasoconstrictor ETB receptor expression in the smooth muscle cells with significantly increased vasoconstriction. This vasoconstrictor receptor upregulation is demonstrated to be via activation of mitogen-activated protein kinase (MAPK) pathways (Xu et al., 2014). Minimally modified low-density lipoprotein (mmLDL) at 10 times lower concentration than native LDL can also induces upregulation of vasoconstrictor ETB receptor expression in rat coronary (Jie et al., 2012) arterial smooth muscle cells, suggesting that mmLDL has much stronger effects than native LDL. Thus, it is most likely there is a possibility that the upregulation of vasoconstrictor ETB receptor expression makes the subjects susceptible to vasospasm, and thus contributes to the pathogenesis of ischemic cardiovascular disease. On the other hand, endothelial cells maintain a relaxed vascular tone and low levels of oxidative stress by releasing mediators such as nitric oxide (NO), prostacyclin (PGI2) and endotheliumdependent hyperpolarizing factor (EDHF). Recently, we have reported that exposure of the arterial segments to native LDL for 24 h results in lipid peroxidation and impaired endotheliumdependent vasodilation with damaging the structure of endothelium (Zhang et al., 2014), which may, in addition to the LDL-increased vasoconstriction mediated by the receptors (Xu et al., 2014), contribute to the development of ischemic cardiovascular disease. More interestingly, similar effects are obtained from apoB per se at concentration equivalent to the amounts of apoB contained in LDL, suggesting that the detrimental effect of LDL is largely mediated by apoB (Zhang et al., 2014).
3. ApoB, the villain in the drama?
Vehicle Cholesterol 4.5mg/L
-10
167
-6
ACh (logM) Fig. 1. Acetylcholine (Ach)-induced vasodilatation in 5-HT (10 5.5 M) pre-contracted rat mesenteric artery segments after organ culture in presence of (A) cholesterol (4.5 mg/L), (B) ApoB (11 mg/L) (Zhang et al., 2014) or vehicle for 24 h. The used concentrations of cholesterol or ApoB were equivalent as in LDL 50 mg/L. Data are expressed as mean 7 S.E.M. (n¼ 8–12). Two-way ANOVA with Bonferroni post-test was used, ***Po 0.001 vs. vehicle.
ApoB is synthesized in the liver and occurs in two isoforms, apoB-48 and apoB-100. The apoB in LDL is apoB-100, which is essential for the binding of LDL particles to the LDL receptor, allowing cells to internalize LDL and thus to absorb cholesterol (Walldius and Jungner, 2006). Clinical studies demonstrate that hyperlipoproteinemia causes dysfunction of endothelium in the initial stage of atherogenesis with impairing endotheliumdependent vasodilation (Kraml et al., 2004), and in addition, apoB is proved to be an early marker of atherosclerosis (Lind, 2007). Not only cholesterol, but also other components contained in LDL, associate with the development of cardiovascular disease. Here, we have for the first time demonstrated that apoB (Zhang et al., 2014), but not cholesterol (Fig. 1), impairs endotheliumdependent vasodilation with attenuating the NO-mediated pathway. This agrees well with the findings from a large cohort study showing that the apoB/A1 levels are inversely related to endothelium-dependent vasodilation in a group of 1016 subjects (Lind, 2007). Considering that NO is a pivotal mediator of endothelium-dependent vasodilation, we have investigated the involvement of the NO pathway and shown that LDL induces decreased expression of both endothelial NO synthase (eNOS) and induced NO synthase (iNOS) as well as weaker staining of endothelial cell marker (CD31), indicating that structure and function of the endothelium are impaired (Zhang et al., 2014). To examine the effects of cholesterol, dimethyl sulfoxide (DMSO), an important polar aprotic solvent that is miscible both organic solvent and water, is used to dissolve cholesterol. At concentrations of apoB (11 mg/L) and cholesterol (4.5 mg/L), which are equivalent as in LDL 50 mg/L supplied as a lyophilized powder (5.0 mg protein/vial) and reconstituted with 1 mL of the solvent, apoB significantly attenuates vasodilatation with decreased
168
Q. Yu et al. / European Journal of Pharmacology 748 (2015) 166–169
maximal relaxation (Rmax) from 75% to 25% (Zhang et al., 2014), while cholesterol has no such effects (Fig. 1). This demonstrates that apoB is the malefactor of the LDL effects, which is further supported by the clinical findings that in type 2 diabetes, plasma apoB levels, but not LDL cholesterol, associate with coronary artery calcification (Martin et al., 2009). Therefore, apoB seems to be the villain in the drama. However, a meta-analysis of 62,154 patients treated with statins shows there is no significant difference between apoB and LDL cholesterol levels for increase in the risk of cardiovascular events (Boekholdt et al., 2012), suggesting that both of them are equally “bad”.
4. ApoB, a new and promising pharmacological target for treatment of ischemic cardiovascular disease Inhibitor of hydroxymethyl glutaryl coenzyme A reductase (statin therapy) lowers the plasma LDL levels and has been the core of guidelines for treating cardiovascular disease. However, apoB may be more useful clinically than LDL cholesterol in cardiovascular disease, because it captures greater information about atherogenic particles and is not influenced by heterogeneity of particle cholesterol content (Benn et al., 2007; Pischon et al., 2005). A considerable residual risk remains after intensive statin therapy and, a significant proportion of high-risk patients do not achieve the optimal LDL cholesterol goal recommended in the current guidelines (o1.8 mmol/L) (Sahebkar and Watts, 2013). Hence, new LDL cholesterol lowering agents are needed to address populations with these unique genetic conditions (Milani and Lavie, 2013). Mipomersen, a new drug, works as an antisense agent against mRNA expression of apoB and significantly reduces LDL cholesterol, apoB and lipoprotein(a) in patients with hypercholesterolemia with, or at risk for cardiovascular disease not controlled by existing therapies (Thomas et al., 2013). Recently, the US Food and Drug Administration has approved mipomersen for special use in the most dramatic elevations of LDL cholesterol that are usually seen in patients with genetic abnormalities in the LDL receptor gene on both chromosome pairs who are not fully responsive to current treatments (Brown et al., 2014). Since the concept of arterial inflammation in response to accumulation and oxidation of lipoproteins in the vascular wall became a new strategy for investigation of treating and/or preventing ischemic cardiovascular disease, plaque-antigen tolerogenic vaccines have been developed to dampen plaque inflammation and disease progression. A first generation of atherosclerosis vaccine based on peptides derived from apoB-100 has demonstrated promising results in animal studies (Herbin et al., 2012), and are now approaching clinical testing (Nilsson et al., 2013). Interestingly, high levels of autoantibodies against apoB100 peptides in clinic patients associates with less severe atherosclerosis in the coronary and lower risk of acute myocardial infarction (Sjogren et al., 2008). Not only apoB-100 modified by oxidation but also non-modified apoB-100 is targeted by autoimmune responses, suggesting that aggravation of plaque inflammation may occur as a result of a local loss of tolerance against LDL in the plaque, and thus restoration of lost tolerance represents a novel approach for treatment of cardiovascular disease (Nilsson et al., 2012). In conclusion, apoB, but not cholesterol, plays a major role in the LDL-induced dysfunction of vascular endothelium, suggesting that direct apoB-targeting agents might be a promising therapy for treating ischemic cardiovascular disease. Named “bad” cholesterol in LDL is insufficiently to describe the characters of LDL, and LDL has more detrimental effects than only transportation of cholesterol into artery walls. More importantly, there is still not fully
understood the pathological roles of LDL or apoB in ischemic cardiovascular disease. To explore these lipoproteins and apolipoproteins should be helpful for the establishment of novel pharmacological targets for translational medicine.
Acknowledgments This work was supported by grants from National Natural Science Foundation of China (no. 81470493), Natural Science Basic Research Plan in Shaanxi Province (Program no. 2013JM4022), the Shaanxi 100 Talents Program (Prof. Cang-Bao Xu) and Xi'an Medical University.
References Benn, M., Nordestgaard, B.G., Jensen, G.B., Tybjaerg-Hansen, A., 2007. Improving prediction of ischemic cardiovascular disease in the general population using apolipoprotein B: the Copenhagen City Heart Study. Arterioscler. Thromb. Vasc. Biol. 27, 661–670. Boekholdt, S.M., Arsenault, B.J., Mora, S., Pedersen, T.R., LaRosa, J.C., Nestel, P.J., Simes, R.J., Durrington, P., Hitman, G.A., Welch, K.M., DeMicco, D.A., Zwinderman, A.H., Clearfield, M.B., Downs, J.R., Tonkin, A.M., Colhoun, H.M., Gotto Jr., A. M., Ridker, P.M., Kastelein, J.J., 2012. Association of LDL cholesterol, non-HDL cholesterol, and apolipoprotein B levels with risk of cardiovascular events among patients treated with statins: a meta-analysis. JAMA 307, 1302–1309. Brown, W.V., Rader, D.J., Goldberg, A.C., 2014. JCL roundtable: drug treatment of severe forms of familial hypercholesterolemia. J. Clin. Lipidol. 8, 10–17. Cao, L., Cao, Y.X., Xu, C.B., Edvinsson, L., 2013. Altered endothelin receptor expression and affinity in spontaneously hypertensive rat cerebral and coronary arteries. PLoS One 8, e73761. Chien, S., 2008. Effects of disturbed flow on endothelial cells. Ann. Biomed. Eng. 36, 554–562. Grundy, S.M., 2002. Low-density lipoprotein, non-high-density lipoprotein, and apolipoprotein B as targets of lipid-lowering therapy. Circulation 106, 2526–2529. Haimeur, A., Messaouri, H., Ulmann, L., Mimouni, V., Masrar, A., Chraibi, A., Tremblin, G., Meskini, N., 2013. Argan oil prevents prothrombotic complications by lowering lipid levels and platelet aggregation, enhancing oxidative status in dyslipidemic patients from the area of Rabat (Morocco). Lipids Health Dis. 12, 107. Herbin, O., Ait-Oufella, H., Yu, W., Fredrikson, G.N., Aubier, B., Perez, N., Barateau, V., Nilsson, J., Tedgui, A., Mallat, Z., 2012. Regulatory T-cell response to apolipoprotein B100-derived peptides reduces the development and progression of atherosclerosis in mice. Arterioscler. Thromb. Vasc. Biol. 32, 605–612. Hevonoja, T., Pentikainen, M.O., Hyvonen, M.T., Kovanen, P.T., Ala-Korpela, M., 2000. Structure of low density lipoprotein (LDL) particles: basis for understanding molecular changes in modified LDL. Biochim. Biophys. Acta 1488, 189–210. Jie, L., Yong-Xiao, C., Zu-Yi, Y., Cang-Bao, X., 2012. Minimally modified LDL upregulates endothelin type B receptors in rat coronary artery via ERK1/2 MAPK and NF-kappaB pathways. Biochim. Biophys. Acta 1821, 582–589. Kraml, P., Syrovatka, P., Stipek, S., Fialova, L., Koprivova, H., Potockova, J., Andel, M., 2004. Hyperlipoproteinemia impairs endothelium-dependent vasodilation. Physiol. Res. 53, 471–480. Libby, P., Ridker, P.M., Hansson, G.K., 2011. Progress and challenges in translating the biology of atherosclerosis. Nature 473, 317–325. Lind, L., 2007. Vasodilation in resistance arteries is related to the apolipoprotein B/ A1 ratio in the elderly: the Prospective Investigation of the Vasculature in Uppsala Seniors (PIVUS) study. Atherosclerosis 190, 378–384. Lund-Katz, S., Phillips, M.C., 2010. High density lipoprotein structure-function and role in reverse cholesterol transport. Subcell. Biochem. 51, 183–227. Martin, S.S., Qasim, A.N., Mehta, N.N., Wolfe, M., Terembula, K., Schwartz, S., Iqbal, N., Schutta, M., Bagheri, R., Reilly, M.P., 2009. Apolipoprotein B but not LDL cholesterol is associated with coronary artery calcification in type 2 diabetic whites. Diabetes 58, 1887–1892. Milani, R.V., Lavie, C.J., 2013. Lipid control in the modern era: an orphan's tale of rags to riches. J. Am. Coll. Cardiol. 62, 2185–2187. Nilsson, D., Wackenfors, A., Gustafsson, L., Edvinsson, L., Paulsson, P., Ingemansson, R., Malmsjo, M., 2008. Increased ET(A) and ET(B) receptor contraction in the left internal mammary artery from patients with hypertension. J. Hum. Hypertens. 22, 226–229. Nilsson, J., Bjorkbacka, H., Fredrikson, G.N., 2012. Apolipoprotein B100 autoimmunity and atherosclerosis - disease mechanisms and therapeutic potential. Curr. Opin. Lipidol. 23, 422–428. Nilsson, J., Wigren, M., Shah, P.K., 2013. Vaccines against atherosclerosis. Expert Rev. Vaccines 12, 311–321. Pischon, T., Girman, C.J., Sacks, F.M., Rifai, N., Stampfer, M.J., Rimm, E.B., 2005. Non-high-density lipoprotein cholesterol and apolipoprotein B in the prediction of coronary heart disease in men. Circulation 112, 3375–3383.
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Sahebkar, A., Watts, G.F., 2013. New LDL-cholesterol lowering therapies: pharmacology, clinical trials, and relevance to acute coronary syndromes. Clin. Ther. 35, 1082–1098. Sjogren, P., Fredrikson, G.N., Samnegard, A., Ericsson, C.G., Ohrvik, J., Fisher, R.M., Nilsson, J., Hamsten, A., 2008. High plasma concentrations of autoantibodies against native peptide 210 of apoB-100 are related to less coronary atherosclerosis and lower risk of myocardial infarction. Eur. Heart J. 29, 2218–2226. Steinberg, D., 1997. Low density lipoprotein oxidation and its pathobiological significance. J. Biol. Chem. 272, 20963–20966. Stenman, E., Malmsjo, M., Uddman, E., Gido, G., Wieloch, T., Edvinsson, L., 2002. Cerebral ischemia upregulates vascular endothelin ET(B) receptors in rat. Stroke 33, 2311–2316. Thomas, G.S., Cromwell, W.C., Ali, S., Chin, W., Flaim, J.D., Davidson, M., 2013. Mipomersen, an apolipoprotein B synthesis inhibitor, reduces atherogenic lipoproteins in patients with severe hypercholesterolemia at high
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cardiovascular risk: a randomized, double-blind, placebo-controlled trial. J. Am. Coll. Cardiol. 62, 2178–2184. Walldius, G., Jungner, I., 2006. The apoB/apoA-I ratio: a strong, new risk factor for cardiovascular disease and a target for lipid-lowering therapy–a review of the evidence. J. Intern Med. 259, 493–519. Xu, C.B., Sun, Y., Edvinsson, L., 2010. Cardiovascular risk factors regulate the expression of vascular endothelin receptors. Pharmacol. Ther. 127, 148–155. Xu, C.B., Zheng, J.P., Zhang, W., Liu, E., Edvinsson, L., Zhang, Y., 2014. Low density lipoprotein induces upregulation of vasoconstrictive endothelin type B receptor expression. Vascul. Pharmacol. 60, 42–48. Yu, X.H., Fu, Y.C., Zhang, D.W., Yin, K., Tang, C.K., 2013. Foam cells in atherosclerosis. Clin. Chim. Acta 424, 245–252. Zhang, Y., Zhang, W., Edvinsson, L., Xu, C.B., 2014. Apolipoprotein B of low-density lipoprotein impairs nitric oxide-mediated endothelium-dependent relaxation in rat mesenteric arteries. Eur. J. Pharmacol. 725, 10–17.
Contents lists available at ScienceDirect
European Journal of Pharmacology journal homepage: www.elsevier.com/locate/ejphar
Perspective
Apolipoprotein B, the villain in the drama? Qi Yu a,1, Yaping Zhang a,b,1, Cang-Bao Xu a,b,n a b
Institute of Basic and Translational Medicine, Xi'an Medical University, Shaanxi, Xi'an 710021, PR China Division of Experimental Vascular Research, Institute of Clinical Science in Lund, Lund University, BMC A13, SE-221 84 Lund, Sweden
art ic l e i nf o
a b s t r a c t
Article history: Accepted 20 August 2014 Available online 16 September 2014
Low-density lipoprotein (LDL) is the major atherogenic lipoprotein and the primary target of lipidlowering therapy for treating ischemic cardiovascular disease. Apolipoprotein B (apoB), an important structural component of LDL, plays a key role in cholesterol transport and removal in vascular wall. On the other hand, under pathological process, apoB interacts with the arterial wall to initiate the cascade of events that leads to atherosclerosis. However, interactions between apoB and vascular wall remain to be determined. Here, we address a pathological role of apoB per se and whole LDL particle in dysfunction of vascular endothelium and smooth muscle cells i.e. decreased endothelium-dependent vasodilation and increased receptor-mediated vasoconstriction. We intend to discuss: i) how apoB is responsible for the deleterious effects of LDL in the development of ischemic cardiovascular disease; ii) why vaccine based on peptides derived from apoB-100 is a promising therapy for treating ischemic cardiovascular disease, and iii) direct inhibition of apoB production should be a better therapeutic option than simple LDLcholesterol lowering therapy in the patients with severe hypercholesterolemia at high cardiovascular risk with statin intolerance. In conclusion, apoB, but not cholesterol, plays a major role in LDL-induced dysfunction of endothelium, suggesting that direct apoB-targeting agents might be a promising therapy for the treatment of ischemic cardiovascular disease. & 2014 Elsevier B.V. All rights reserved.
Keywords: Low-density lipoprotein Apolipoprotein B Cholesterol Vasoconstriction Vasodilation Ischemic cardiovascular disease
1. The “bad” cholesterol Atherosclerosis leads to ischemic cardiovascular disease, which is the major cause of mortality in the world. Low-density lipoprotein (LDL) is well-recognized to play a key role in the pathogenesis of atherosclerosis that is a chronic inflammatory disease of the arterial wall. The LDL particle consists of phospholipid, free cholesterol and apolipoprotein B (apoB) (Hevonoja et al., 2000). In contrast, highdensity lipoprotein (HDL) contains exchangeable apolipoproteins of the A, C and E families (Lund-Katz and Phillips, 2010). Under physiological conditions, LDL is a vehicle that transports cholesterol all over the body in order to maintain cell viability and to provide cholesterol for the synthesis of the steroid hormones (Grundy, 2002). However, under pathological conditions, fluid sheer stress affects the morphology of endothelial cells in regions of arterial branching or curvature, along with high plasma LDL, result in increased permeability of LDL to preferential sites (Chien, 2008). When LDL infiltrates into the artery wall, it can be modified to oxidized LDL (ox-LDL), and causes the lipid deposit in the intima with chronic inflammation in the
n
Corresponding author at: Institute of Basic and Translational Medicine, Xi'an Medical University, Shaanxi, Xi'an 710021, PR China. Tel./fax: þ 86 29 86177632. E-mail address: [email protected] (C.-B. Xu). 1 These authors equally contributed to this work. http://dx.doi.org/10.1016/j.ejphar.2014.08.037 0014-2999/& 2014 Elsevier B.V. All rights reserved.
arterial wall, which subsequently results in the formation of atherosclerotic plaques. In clinic, plasma LDL cholesterol levels in patients are found to be positively correlated with the risk of cardiovascular events in human populations and augment individual susceptibility to atherosclerosis and its complications (Boekholdt et al., 2012; Haimeur et al., 2013). Therefore, LDL cholesterol is often called the “bad” cholesterol, and has been as the primary target of lipid-lowering therapy for treating ischemic cardiovascular disease (Libby et al., 2011). However, LDL is more than just cholesterol, and the contribution of LDL to cardiovascular disease risk involves more than just the cholesterol effects contained in LDL. Almost all cells of the artery wall have the ability to pick up LDL particles through the LDL receptors. Accumulation of LDL in the vasculature can easily undergo oxidative modification catalyzed by any of the major cell types of artery wall (Steinberg, 1997). Although many studies pay more attention to interactions between LDL and its receptors on cells in the arterial wall, the molecular mechanism that how LDL leads to dysfunction of vascular endothelium and smooth muscle cells remain not fully understood. 2. Effects of LDL on receptor-mediated vasoconstriction and endothelium-dependent vasodilation Atherosclerosis is a chronic disease characterized by the accumulation of lipoprotein lipid within the arterial wall. LDL is one of
Q. Yu et al. / European Journal of Pharmacology 748 (2015) 166–169
lipoproteins containing apoB-100, which makes LDL to be recognized and captured by LDL receptors on the cells in the arterial wall, but this does not induce cholesterol accumulation in monocyte/macrophages and transform into foam cells in the arterial intima. CD36 and scavenger receptor class A are mainly responsible for uptake of lipoprotein-derived cholesterol by macrophages, and the macrophages to be transformed into lipid-laden foam cells (Yu et al., 2013), which induces lipid deposition in and damage to the arterial wall. Since the discovery of endothelin-1 (ET-1) is a powerful endogenous vasoconstrictor, there have been intensive studies regarding the involvement of ET-1 and its receptors in the pathophysiology of ischemic cardiovascular diseases. In mammals, two different subtypes of endothelin receptors are known as the endothelin type A (ETA) and type B (ETB) receptors. The ETA receptors are located on the smooth muscle cells and mediate contractile effect, while the ETB receptors are limitedly expressed on the endothelial cells and respond with vasodilation (Xu et al., 2010). However, under pathological conditions like hypertension (Cao et al., 2013; Nilsson et al., 2008) and cerebral ischemia (Stenman et al., 2002), the ETB receptors can express on the
Relaxation (% of 5-HT)
0
25
50
75
100 -9
-8
-7
-6
-5
-4
***
***
-5
-4
ACh (logM) 0
Relaxation (% of 5-HT)
***
***
25
50
75
Vehicle ApoB 11mg/L
100 -10
-9
-8
-7
smooth muscle cells and mediates strong vasoconstriction, suggesting the vasoconstrictor ETB receptor expression is inducible and pathogenic. Incubation of the arterial segments with native LDL for 24 h induces transcriptional upregulation of vasoconstrictor ETB receptor expression in the smooth muscle cells with significantly increased vasoconstriction. This vasoconstrictor receptor upregulation is demonstrated to be via activation of mitogen-activated protein kinase (MAPK) pathways (Xu et al., 2014). Minimally modified low-density lipoprotein (mmLDL) at 10 times lower concentration than native LDL can also induces upregulation of vasoconstrictor ETB receptor expression in rat coronary (Jie et al., 2012) arterial smooth muscle cells, suggesting that mmLDL has much stronger effects than native LDL. Thus, it is most likely there is a possibility that the upregulation of vasoconstrictor ETB receptor expression makes the subjects susceptible to vasospasm, and thus contributes to the pathogenesis of ischemic cardiovascular disease. On the other hand, endothelial cells maintain a relaxed vascular tone and low levels of oxidative stress by releasing mediators such as nitric oxide (NO), prostacyclin (PGI2) and endotheliumdependent hyperpolarizing factor (EDHF). Recently, we have reported that exposure of the arterial segments to native LDL for 24 h results in lipid peroxidation and impaired endotheliumdependent vasodilation with damaging the structure of endothelium (Zhang et al., 2014), which may, in addition to the LDL-increased vasoconstriction mediated by the receptors (Xu et al., 2014), contribute to the development of ischemic cardiovascular disease. More interestingly, similar effects are obtained from apoB per se at concentration equivalent to the amounts of apoB contained in LDL, suggesting that the detrimental effect of LDL is largely mediated by apoB (Zhang et al., 2014).
3. ApoB, the villain in the drama?
Vehicle Cholesterol 4.5mg/L
-10
167
-6
ACh (logM) Fig. 1. Acetylcholine (Ach)-induced vasodilatation in 5-HT (10 5.5 M) pre-contracted rat mesenteric artery segments after organ culture in presence of (A) cholesterol (4.5 mg/L), (B) ApoB (11 mg/L) (Zhang et al., 2014) or vehicle for 24 h. The used concentrations of cholesterol or ApoB were equivalent as in LDL 50 mg/L. Data are expressed as mean 7 S.E.M. (n¼ 8–12). Two-way ANOVA with Bonferroni post-test was used, ***Po 0.001 vs. vehicle.
ApoB is synthesized in the liver and occurs in two isoforms, apoB-48 and apoB-100. The apoB in LDL is apoB-100, which is essential for the binding of LDL particles to the LDL receptor, allowing cells to internalize LDL and thus to absorb cholesterol (Walldius and Jungner, 2006). Clinical studies demonstrate that hyperlipoproteinemia causes dysfunction of endothelium in the initial stage of atherogenesis with impairing endotheliumdependent vasodilation (Kraml et al., 2004), and in addition, apoB is proved to be an early marker of atherosclerosis (Lind, 2007). Not only cholesterol, but also other components contained in LDL, associate with the development of cardiovascular disease. Here, we have for the first time demonstrated that apoB (Zhang et al., 2014), but not cholesterol (Fig. 1), impairs endotheliumdependent vasodilation with attenuating the NO-mediated pathway. This agrees well with the findings from a large cohort study showing that the apoB/A1 levels are inversely related to endothelium-dependent vasodilation in a group of 1016 subjects (Lind, 2007). Considering that NO is a pivotal mediator of endothelium-dependent vasodilation, we have investigated the involvement of the NO pathway and shown that LDL induces decreased expression of both endothelial NO synthase (eNOS) and induced NO synthase (iNOS) as well as weaker staining of endothelial cell marker (CD31), indicating that structure and function of the endothelium are impaired (Zhang et al., 2014). To examine the effects of cholesterol, dimethyl sulfoxide (DMSO), an important polar aprotic solvent that is miscible both organic solvent and water, is used to dissolve cholesterol. At concentrations of apoB (11 mg/L) and cholesterol (4.5 mg/L), which are equivalent as in LDL 50 mg/L supplied as a lyophilized powder (5.0 mg protein/vial) and reconstituted with 1 mL of the solvent, apoB significantly attenuates vasodilatation with decreased
168
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maximal relaxation (Rmax) from 75% to 25% (Zhang et al., 2014), while cholesterol has no such effects (Fig. 1). This demonstrates that apoB is the malefactor of the LDL effects, which is further supported by the clinical findings that in type 2 diabetes, plasma apoB levels, but not LDL cholesterol, associate with coronary artery calcification (Martin et al., 2009). Therefore, apoB seems to be the villain in the drama. However, a meta-analysis of 62,154 patients treated with statins shows there is no significant difference between apoB and LDL cholesterol levels for increase in the risk of cardiovascular events (Boekholdt et al., 2012), suggesting that both of them are equally “bad”.
4. ApoB, a new and promising pharmacological target for treatment of ischemic cardiovascular disease Inhibitor of hydroxymethyl glutaryl coenzyme A reductase (statin therapy) lowers the plasma LDL levels and has been the core of guidelines for treating cardiovascular disease. However, apoB may be more useful clinically than LDL cholesterol in cardiovascular disease, because it captures greater information about atherogenic particles and is not influenced by heterogeneity of particle cholesterol content (Benn et al., 2007; Pischon et al., 2005). A considerable residual risk remains after intensive statin therapy and, a significant proportion of high-risk patients do not achieve the optimal LDL cholesterol goal recommended in the current guidelines (o1.8 mmol/L) (Sahebkar and Watts, 2013). Hence, new LDL cholesterol lowering agents are needed to address populations with these unique genetic conditions (Milani and Lavie, 2013). Mipomersen, a new drug, works as an antisense agent against mRNA expression of apoB and significantly reduces LDL cholesterol, apoB and lipoprotein(a) in patients with hypercholesterolemia with, or at risk for cardiovascular disease not controlled by existing therapies (Thomas et al., 2013). Recently, the US Food and Drug Administration has approved mipomersen for special use in the most dramatic elevations of LDL cholesterol that are usually seen in patients with genetic abnormalities in the LDL receptor gene on both chromosome pairs who are not fully responsive to current treatments (Brown et al., 2014). Since the concept of arterial inflammation in response to accumulation and oxidation of lipoproteins in the vascular wall became a new strategy for investigation of treating and/or preventing ischemic cardiovascular disease, plaque-antigen tolerogenic vaccines have been developed to dampen plaque inflammation and disease progression. A first generation of atherosclerosis vaccine based on peptides derived from apoB-100 has demonstrated promising results in animal studies (Herbin et al., 2012), and are now approaching clinical testing (Nilsson et al., 2013). Interestingly, high levels of autoantibodies against apoB100 peptides in clinic patients associates with less severe atherosclerosis in the coronary and lower risk of acute myocardial infarction (Sjogren et al., 2008). Not only apoB-100 modified by oxidation but also non-modified apoB-100 is targeted by autoimmune responses, suggesting that aggravation of plaque inflammation may occur as a result of a local loss of tolerance against LDL in the plaque, and thus restoration of lost tolerance represents a novel approach for treatment of cardiovascular disease (Nilsson et al., 2012). In conclusion, apoB, but not cholesterol, plays a major role in the LDL-induced dysfunction of vascular endothelium, suggesting that direct apoB-targeting agents might be a promising therapy for treating ischemic cardiovascular disease. Named “bad” cholesterol in LDL is insufficiently to describe the characters of LDL, and LDL has more detrimental effects than only transportation of cholesterol into artery walls. More importantly, there is still not fully
understood the pathological roles of LDL or apoB in ischemic cardiovascular disease. To explore these lipoproteins and apolipoproteins should be helpful for the establishment of novel pharmacological targets for translational medicine.
Acknowledgments This work was supported by grants from National Natural Science Foundation of China (no. 81470493), Natural Science Basic Research Plan in Shaanxi Province (Program no. 2013JM4022), the Shaanxi 100 Talents Program (Prof. Cang-Bao Xu) and Xi'an Medical University.
References Benn, M., Nordestgaard, B.G., Jensen, G.B., Tybjaerg-Hansen, A., 2007. Improving prediction of ischemic cardiovascular disease in the general population using apolipoprotein B: the Copenhagen City Heart Study. Arterioscler. Thromb. Vasc. Biol. 27, 661–670. Boekholdt, S.M., Arsenault, B.J., Mora, S., Pedersen, T.R., LaRosa, J.C., Nestel, P.J., Simes, R.J., Durrington, P., Hitman, G.A., Welch, K.M., DeMicco, D.A., Zwinderman, A.H., Clearfield, M.B., Downs, J.R., Tonkin, A.M., Colhoun, H.M., Gotto Jr., A. M., Ridker, P.M., Kastelein, J.J., 2012. Association of LDL cholesterol, non-HDL cholesterol, and apolipoprotein B levels with risk of cardiovascular events among patients treated with statins: a meta-analysis. JAMA 307, 1302–1309. Brown, W.V., Rader, D.J., Goldberg, A.C., 2014. JCL roundtable: drug treatment of severe forms of familial hypercholesterolemia. J. Clin. Lipidol. 8, 10–17. Cao, L., Cao, Y.X., Xu, C.B., Edvinsson, L., 2013. Altered endothelin receptor expression and affinity in spontaneously hypertensive rat cerebral and coronary arteries. PLoS One 8, e73761. Chien, S., 2008. Effects of disturbed flow on endothelial cells. Ann. Biomed. Eng. 36, 554–562. Grundy, S.M., 2002. Low-density lipoprotein, non-high-density lipoprotein, and apolipoprotein B as targets of lipid-lowering therapy. Circulation 106, 2526–2529. Haimeur, A., Messaouri, H., Ulmann, L., Mimouni, V., Masrar, A., Chraibi, A., Tremblin, G., Meskini, N., 2013. Argan oil prevents prothrombotic complications by lowering lipid levels and platelet aggregation, enhancing oxidative status in dyslipidemic patients from the area of Rabat (Morocco). Lipids Health Dis. 12, 107. Herbin, O., Ait-Oufella, H., Yu, W., Fredrikson, G.N., Aubier, B., Perez, N., Barateau, V., Nilsson, J., Tedgui, A., Mallat, Z., 2012. Regulatory T-cell response to apolipoprotein B100-derived peptides reduces the development and progression of atherosclerosis in mice. Arterioscler. Thromb. Vasc. Biol. 32, 605–612. Hevonoja, T., Pentikainen, M.O., Hyvonen, M.T., Kovanen, P.T., Ala-Korpela, M., 2000. Structure of low density lipoprotein (LDL) particles: basis for understanding molecular changes in modified LDL. Biochim. Biophys. Acta 1488, 189–210. Jie, L., Yong-Xiao, C., Zu-Yi, Y., Cang-Bao, X., 2012. Minimally modified LDL upregulates endothelin type B receptors in rat coronary artery via ERK1/2 MAPK and NF-kappaB pathways. Biochim. Biophys. Acta 1821, 582–589. Kraml, P., Syrovatka, P., Stipek, S., Fialova, L., Koprivova, H., Potockova, J., Andel, M., 2004. Hyperlipoproteinemia impairs endothelium-dependent vasodilation. Physiol. Res. 53, 471–480. Libby, P., Ridker, P.M., Hansson, G.K., 2011. Progress and challenges in translating the biology of atherosclerosis. Nature 473, 317–325. Lind, L., 2007. Vasodilation in resistance arteries is related to the apolipoprotein B/ A1 ratio in the elderly: the Prospective Investigation of the Vasculature in Uppsala Seniors (PIVUS) study. Atherosclerosis 190, 378–384. Lund-Katz, S., Phillips, M.C., 2010. High density lipoprotein structure-function and role in reverse cholesterol transport. Subcell. Biochem. 51, 183–227. Martin, S.S., Qasim, A.N., Mehta, N.N., Wolfe, M., Terembula, K., Schwartz, S., Iqbal, N., Schutta, M., Bagheri, R., Reilly, M.P., 2009. Apolipoprotein B but not LDL cholesterol is associated with coronary artery calcification in type 2 diabetic whites. Diabetes 58, 1887–1892. Milani, R.V., Lavie, C.J., 2013. Lipid control in the modern era: an orphan's tale of rags to riches. J. Am. Coll. Cardiol. 62, 2185–2187. Nilsson, D., Wackenfors, A., Gustafsson, L., Edvinsson, L., Paulsson, P., Ingemansson, R., Malmsjo, M., 2008. Increased ET(A) and ET(B) receptor contraction in the left internal mammary artery from patients with hypertension. J. Hum. Hypertens. 22, 226–229. Nilsson, J., Bjorkbacka, H., Fredrikson, G.N., 2012. Apolipoprotein B100 autoimmunity and atherosclerosis - disease mechanisms and therapeutic potential. Curr. Opin. Lipidol. 23, 422–428. Nilsson, J., Wigren, M., Shah, P.K., 2013. Vaccines against atherosclerosis. Expert Rev. Vaccines 12, 311–321. Pischon, T., Girman, C.J., Sacks, F.M., Rifai, N., Stampfer, M.J., Rimm, E.B., 2005. Non-high-density lipoprotein cholesterol and apolipoprotein B in the prediction of coronary heart disease in men. Circulation 112, 3375–3383.
Q. Yu et al. / European Journal of Pharmacology 748 (2015) 166–169
Sahebkar, A., Watts, G.F., 2013. New LDL-cholesterol lowering therapies: pharmacology, clinical trials, and relevance to acute coronary syndromes. Clin. Ther. 35, 1082–1098. Sjogren, P., Fredrikson, G.N., Samnegard, A., Ericsson, C.G., Ohrvik, J., Fisher, R.M., Nilsson, J., Hamsten, A., 2008. High plasma concentrations of autoantibodies against native peptide 210 of apoB-100 are related to less coronary atherosclerosis and lower risk of myocardial infarction. Eur. Heart J. 29, 2218–2226. Steinberg, D., 1997. Low density lipoprotein oxidation and its pathobiological significance. J. Biol. Chem. 272, 20963–20966. Stenman, E., Malmsjo, M., Uddman, E., Gido, G., Wieloch, T., Edvinsson, L., 2002. Cerebral ischemia upregulates vascular endothelin ET(B) receptors in rat. Stroke 33, 2311–2316. Thomas, G.S., Cromwell, W.C., Ali, S., Chin, W., Flaim, J.D., Davidson, M., 2013. Mipomersen, an apolipoprotein B synthesis inhibitor, reduces atherogenic lipoproteins in patients with severe hypercholesterolemia at high
169
cardiovascular risk: a randomized, double-blind, placebo-controlled trial. J. Am. Coll. Cardiol. 62, 2178–2184. Walldius, G., Jungner, I., 2006. The apoB/apoA-I ratio: a strong, new risk factor for cardiovascular disease and a target for lipid-lowering therapy–a review of the evidence. J. Intern Med. 259, 493–519. Xu, C.B., Sun, Y., Edvinsson, L., 2010. Cardiovascular risk factors regulate the expression of vascular endothelin receptors. Pharmacol. Ther. 127, 148–155. Xu, C.B., Zheng, J.P., Zhang, W., Liu, E., Edvinsson, L., Zhang, Y., 2014. Low density lipoprotein induces upregulation of vasoconstrictive endothelin type B receptor expression. Vascul. Pharmacol. 60, 42–48. Yu, X.H., Fu, Y.C., Zhang, D.W., Yin, K., Tang, C.K., 2013. Foam cells in atherosclerosis. Clin. Chim. Acta 424, 245–252. Zhang, Y., Zhang, W., Edvinsson, L., Xu, C.B., 2014. Apolipoprotein B of low-density lipoprotein impairs nitric oxide-mediated endothelium-dependent relaxation in rat mesenteric arteries. Eur. J. Pharmacol. 725, 10–17.
