Novel Plaque Enriched Long Noncoding RNA in Atherosclerotic Macrophage Regulation (PELATON)

Abstract

Objective: Long noncoding RNAs (lncRNAs) are an emergent class of molecules with diverse functional roles, widely expressed in human physiology and disease. Although some lncRNAs have been identified in cardiovascular disease, their potential as novel targets in the prevention of atherosclerosis is unknown. We set out to discover important lncRNAs in unstable plaque and gain insight into their functional relevance. Approach and Results: Analysis of RNA sequencing previously performed on stable and unstable atherosclerotic plaque identified a panel of 47 differentially regulated lncRNAs. We focused on LINC01272, a lncRNA upregulated in unstable plaque previously detected in inflammatory bowel disease, which we termed PELATON (plaque enriched lncRNA in atherosclerotic and inflammatory bowel macrophage regulation). Here, we demonstrate that PELATON is highly monocyte- and macrophage-specific across vascular cell types, and almost entirely nuclear by cellular fractionation (90%–98%). In situ hybridization confirmed enrichment of PELATON in areas of plaque inflammation, colocalizing with macrophages around the shoulders and necrotic core of human plaque sections. Consistent with its nuclear localization, and despite containing a predicted open reading frame, PELATON did not demonstrate any protein-coding potential in vitro. Functionally, knockdown of PELATON significantly reduced phagocytosis, lipid uptake and reactive oxygen species production in high-content analysis, with a significant reduction in phagocytosis independently validated. Furthermore, CD36, a key mediator of phagocytic oxLDL (oxidized low-density lipoprotein) uptake was significantly reduced with PELATON knockdown. Conclusions: PELATON is a nuclear expressed, monocyte- and macrophage-specific lncRNA, upregulated in unstable atherosclerotic plaque. Knockdown of PELATON affects cellular functions associated with plaque progression.

Role of Macrophages and RhoA Pathway in Atherosclerosis

“…Many in vitro and rodent model studies showed that the macrophage-derived foam cells have inflammatory functions. The internalized oxLDLs induce, through the binding to the scavenger receptor/Toll-like receptors (CD36/TLR4/TLR6) complex, transcription and translation of the pro-inflammatory cytokines [38,[49][50][51]. Moreover, the cholesterol and the lipids isolated from human atheromas activated NOD-, LRR-, and pyrin domain-containing protein 3 (NLRP3) inflammasome, induced endoplasmic reticulum (ER) stress, and the production of pro-inflammatory IL-1β in the in vitro cultured macrophages [33,[44][45][46].…”

“…There are also other, recently discovered factors, which regulate lipid uptake, functions of macrophages, and the progression and stability of the atherosclerotic plaque. The RNA sequencing of stable and unstable atherosclerotic plaques identified 47 differentially expressed long non-coding RNAs (lncRNAs) [ 50 ]. It is known that only a small fraction of the mammalian genome is transcribed into the protein-coding mRNAs, while about 80% of the genome is transcribed into non-coding RNAs.…”

“…Studies showed that the expression of one of these RNAs, the LINC01272 (also called PELATON), is highly upregulated in the nuclei of monocytes and macrophages upregulated in the unstable plaques, especially in the regions of plaque inflammation. The knockdown studies showed that although the PELATON RNA is not translated into protein it regulates phagocytosis, lipid uptake, and production of the reactive oxygen species [ 50 ]. These findings open new avenues for the studies of factors regulating macrophage functions in atherosclerosis.…”

The Long Non-coding Road to Atherosclerosis

“…However, how exactly lncRNA HOXA-AS2 regulates inflammatory marker and especially the feedback loop with NF-κB is not yet known. LncRNA PELATON (Plaque enriched lncRNA in atherosclerotic and inflammatory bowel macrophage regulation) was found to be implicated in plaque instability and is enriched in unstable compared with stable atherosclerotic plaques [82]. With its high nuclear expression in monocytes and macrophages, it was shown to regulate phagocytosis, oxLDL uptake, and ROS production in differentiated primary human monocytes, in part due to changes in CD36 expression.…”

Macrophage Long Non-Coding RNAs in Pathogenesis of Cardiovascular Disease

“…As anticipated, in vitro experiments demonstrated that PELATON knockdown effectually impaired macrophage phagocytosis, lipid uptake, and reactive oxygen species production, as well as CD36 expression. Detailed mechanistic studies providing greater insights into the PELATON-mediated regulation of CD36 were not conducted [72]. Regardless of the many advancements in RNA biology to date, no known studies are investigating the impact of lncRNAs on those other receptors (e.g., SR-A1 and LOX-1) in influencing foam cell formation.…”