Atherosclerosis: Open Access
Open Access

Atherosclerosis, a systemic disease affecting arterial walls, is consistently present in the aorta [1]. Although aortic atherosclerosis can be extensive, it often remains asymptomatic throughout life. This is primarily due to the larger lumen size of the aorta, which prevents spontaneous ruptures of atherosclerotic plaques (SRAPs) from causing atherothrombotic occlusion. Consequently, examining the luminal wall of the aorta in vivo offers valuable insights into the complete and unabbreviated progression of atherosclerosis. Over the past five years, Komatsu et al. utilized non-obstructive general angioscopy to survey the aortic atherosclerotic landscape in patients with coronary disease, establishing it as the most sensitive technique for in vivo detection and characterization of aortic atherosclerotic plaques [2]. Initially, they identified and described various appearances of atherosclerotic plaques commonly observed during autopsies, including the consistent presence of plaques with features indicative of spontaneous rupture. Some SRAPs visibly affected the plaque cap through erosion, fissuring, rupture, and ulceration, while others impacted non-cap regions of the plaque, causing discoloration of the atherosclerotic surface, suggestive of intra-mural hemorrhage [3]. Subsequently, they safely retrieved debris from SRAPs, which contained large cholesterol crystals (CCs), necrotic gruel with calcium deposits, and cellular infiltrates consistent with the content of atherosclerotic plaque, as observed through confocal microscopy of fresh unprocessed debris from carotid plaques [4,5]. In this issue of Atherosclerosis, Komatsu et al. took an additional step by characterizing the cellular infiltrate in the debris associated with the so-called puff-chandelier lesions [6]. The analysis confirmed the presence of large CCs, contributing to the debris’s shimmering chandelier appearance. Additionally, the cellular infiltrate stained positive for CD68, NLRP3, IL-1β, and IL-6, markers typically expressed by activated macrophages. Neutrophils were also detected in a significant portion of the specimens. These findings provide strong circumstantial evidence of an innate inflammatory process triggered by CCs within these plaques [7].

The insights obtained from their in-vivo examination of the atherosclerotic landscape yield several important findings. Firstly, their observations confirm the prevalence of SRAPs in living subjects, highlighting the dynamic nature of the atherosclerotic process. These findings emphasize that as atherosclerosis advances; SRAPs commonly affect both the plaque cap and non-cap regions [8]. This observation is significant as it underscores the fact that the majority of plaques are situated deep within the vessel wall, with only a small portion of their surface exposed to the vessel lumen. While the occasional rupture of the plaque cap in smaller and medium-sized arteries can result in atherothrombosis leading to ischemic stroke and myocardial infarction, the frequent but clinically silent injury to the non-cap regions of atherosclerotic plaque is also consequential, as it contributes to disease progression [9,10]. The presence of large cholesterol crystals (CCs) within the debris expelled from SRAPs highlights their pivotal role in plaque rupture, offering further insights. While it is known that CCs can develop within the lipid-rich core of a plaque in living subjects, it is not widely recognized that CCs cannot continue to grow outside of such an environment. As free cholesterol molecules dissociate from lipoproteins and phospholipid structures, they spontaneously associate to form flexible meta-stable structures. Initially, these structures appear as filaments that gradually broaden into ribbons, twisting into helices and eventually forming tubular structures. In the presence of ample free cholesterol monohydrate, the meta-stable tubular structure expands, and numerous microscopic ridged flat plate CCs begin to detach from its end. This process is accompanied by the release of latent elastic energy, dispersing the CCs into their surroundings [11]. These microscopic flat plate CCs serve as platforms for additional free cholesterol molecules to attach, leading to the growth of macroscopic structures. The presence of calcium debris may accelerate the rate of CC growth [12]. When this process occurs within the core of a lipidrich plaque, it can cause damage to its walls as the sharp edges of flat plate CCs may puncture or perforate the plaque, and the aggregation of CC fragments can increase pressure and volume within the plaque, resulting in stretching, thinning, and even rupture of its walls. Plaque rupture exposes the plaque contents to either the systemic circulation or the interstitial space. However, since neither environment is rich in free cholesterol, CCs no longer enlarge once released from the plaque core. Therefore, it is plausible to speculate that the large flat plate CCs found in the debris expelled from ruptured atherosclerotic plaque must have formed and enlarged within the plaque core, rendering it vulnerable or even causing its rupture [13,14].

The presence of activated macrophages and, in some cases, neutrophils within the debris expelled from SRAPs sheds light on the events that occurred in the atherosclerotic region prior to plaque rupture, providing another important insight. When SRAPs affect non-cap regions of a plaque, CCs are released from the acellular core directly into the interstitial space. This release may lead to damage to the vasa vasorum, causing intra-plaque hemorrhage and triggering an immune response by complement and macrophages, recognizing the CCs as foreign. Fragments of CCs that are too large to be engulfed by phagocytes result in frustrated phagocytosis and promote chronic inflammatory injury [15]. The presence of CCs and inflammatory infiltrate in the debris associated with puff-chandelier ruptures suggests that these ruptures involve the plaque’s cap and shoulder regions, which are exposed to crystal-induced inflammation. These observations demonstrate that lipid-rich plaques are constantly at risk of injury, both from within due to the enlargement and aggregation of CCs, and from outside due to acute inflammatory flares triggered by intermittent release of CCs from the core. While most inflammatory flares resolve, leading to sequestration of CCs and progressive arterial sclerosis, there are instances where a flare may precipitate SRAP by further weakening portions of the plaque wall that have stretched and thinned, thus becoming more vulnerable to rupture in conjunction with the enlargement and aggregation of CCs in the core [16,17]. Furthermore, the frequent occurrence of SRAPs associated with the extrusion of atherosclerotic debris throughout the aortic landscape provides a plausible mechanism by which the chronic release of such debris into the periphery can contribute to gradual brain and renal injury that may eventually have clinical implications. While each of these observations in the atherosclerotic landscape supports the significance of CCs in the plaque core as primary drivers of disease progression and SRAPs. Compelling evidence now suggests that CCs also play a role in atherogenesis. When cells such as macrophages, smooth muscle cells, and endothelial cells become loaded with cholesterol, CCs have been observed to form within their membranes and lysosomes. Similar to what occurs in the plaque core, the growth of nascent CCs in cell membranes can lead to membrane dysfunction, while the growth of CCs in lysosomes can disrupt their membranes, resulting in the release of CCs and cathepsins into the cytosol. This, in turn, can activate the NLRP3 inflammasome and trigger apoptosis [18-20].

Conclusion

In conclusion, the work conducted by Komatsu et al. deserves recognition for their in-vivo investigations, which provide further support to the long-held belief derived from autopsy studies, laboratory experiments, and animal models. Their findings reinforce the notion that the transformation of cholesterol into its crystalline form within lipid-rich plaques initiates a cycle of repetitive traumatic and inflammatory injuries. This process ultimately shapes the arterial wall landscape, evolving it from a relatively flat surface to a complex terrain, resembling a volatile volcanic range that is constantly vulnerable to underlying factors.

1. Roberts JC, Moses C, Wilkins RH (1959) Autopsy studies in atherosclerosis I. Distribution and severity of atherosclerosis in patients dying without morphologic evidence of atherosclerotic catastrophe. Circulation 2:511-519.

Google Scholar

2. Komatsu S, Yutani C, Ohara T, Takahashi S, Takewa M, (2018) et al. (2018) Angioscopic evaluation of spontaneously ruptured aortic plaques. J Am Coll Cardiol 25:2893-2902.

Indexed at, Google Scholar, Crossref

3. Kojima K, Kimura S, Hayasaka K, Mizusawa M, Misawa T, et al. (2019) Aortic plaque distribution, and association between aortic plaque and atherosclerotic risk factors: an aortic angioscopy study. J Atherosclerosis Thromb 26:997-1006.

Indexed at, Google Scholar, Crossref

4. Komatsu S, Yutani C, Ohara T, Takahashi S, Takewa M, et al. (2018) Angioscopic evaluation of spontaneously ruptured aortic plaques. J Am Coll Cardiol 25:2893-2902.

Indexed at, Google Scholar, Crossref

5. Nasiri M, Janoudi A, Vanderberg A, Frame M, Flegler C, et al. (2015) Role of cholesterol crystals in atherosclerosis is unmasked by altering tissue preparation methods. Microsc Res Tech 78:969-974.

Indexed at, Google Scholar, Crossref

6. Komatsu S, Yutani C, Takahashi S, Takewa M, Ohara T, et.al. Debris collected in-situ from spontaneously ruptured atherosclerotic plaque invariably contains large cholesterol crystals and evidence of activation of innate inflammation: Insights from non-obstructive general angioscopy. Atherosclerosis 352:96-102.

Indexed at, Google Scholar, Crossref

7. Galozzi P, Bindoli S, Luisetto R, Sfriso P, Ramonda R, et al. (2021) Regulation of crystal induced inflammation: current understandings and clinical implications. Jul Expet Rev Clin Immunol 17:773-787.

Indexed at, Google Scholar, Crossref

8. Konikoff FM, Chung DS, Donovan JM, Small DM, Carey MC (1992) Filamentous, helical, and tubular microstructures during cholesterol crystallization from bile. Evidence that cholesterol does not nucleate classic monohydrate plates. J Clin Invest 90:1155-1160.

Indexed at, Google Scholar, Crossref

9. Varsano N, Beghi F, Elad N, Pereiro E, Dadosh T, et al. (2018) Two polymorphic cholesterol monohydrate crystal structures form in macrophage culture models of atherosclerosis. Proc Natl Acad Sci Unit States Am 115:7662-7669.

Indexed at, Google Scholar, Crossref

10. Abela GS (2010) Cholesterol crystals piercing the arterial plaque and intima trigger local and systemic inflammation. J Clin Lipidol 4:156-164.

Indexed at, Google Scholar, Crossref

11. Park S, Sut TN, Ma GJ, Parikh AN, Cho NJ (2020) Crystallization of cholesterol in phospholipid membranes follows ostwald's rule of stages. J Am Chem Soc 142:1872-1882.

Indexed at, Google Scholar, Crossref

12. Frincu MC, Fleming SD, Rohl AL, Swift JA (2004) The epitaxial growth of cholesterol crystals from bile solutions on calcite substrates. J Am Chem Soc 126:7915-7924.

Indexed at, Google Scholar, Crossref

13. Abela GS, Aziz K (2006) Cholesterol crystals rupture biological membranes and human plaques during acute cardiovascular events-a novel insight into plaque rupture by scanning electron microscopy. Scanning 28:1-10.

Indexed at, Google Scholar, Crossref

14. Abela GS, Kalavakunta JK, Janoudi A, Leffler D, Dhar G, et al. (2017) Frequency of cholesterol crystals in culprit coronary artery aspirate during acute myocardial infarction and their relation to inflammation and myocardial injury. Am J Cardiol 120:1699-1707.

Indexed at, Google Scholar, Crossref

15. Abela GS, Aziz K, Vedre A, Pathak DR, Talbott, JD, et al. (2009) Effect of cholesterol crystals on plaques and intima in arteries of patients with acute coronary and cerebrovascular syndromes. Am J Cardiol 103:959-968.

Indexed at, Google Scholar, Crossref

16. Andrade-Oliveira V, Foresto-Neto O, Watanabe IKM, Zatz R, Câmara NOS (2019) Inflammation in renal diseases: new and old players. Front Pharmacol 10:1192.

Google Scholar, Crossref

17. Brown WR, Thore CR (2011) Review: cerebral microvascular pathology in ageing and neurodegeneration. Feb Neuropathol Appl Neurobiol 37:56-74.

Indexed at, Google Scholar, Crossref

18. Janoudi A, Shamoun FE, Kalavakunta JK, Abela GS (2016) Cholesterol crystal induced arterial inflammation and destabilization of atherosclerotic plaque. Eur Heart J 37:1959-1967.

Indexed at, Google Scholar, Crossref

19. Duewell P, Kono H, Rayner KJ, Sirois CM, Vladimer G, et al. (2010) NLRP3 inflammasomes are required for atherogenesis and activated by cholesterol crystals. Nature 464:1357-1361.

Indexed at, Google Scholar, Crossref

20. Kellner-Weibel G, Yancey PG, Jerome WG, Walser T, Mason RP, et al. (1999) Crystallization of free cholesterol in model macrophage foam cells. Arterioscler Thromb Vasc Biol 19:1891-1898.

Indexed at, Google Scholar, Crossref