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Niemann Pick diseases types A (NPDA) and C (NPDC) are lysosomal storage disorders (LSDs) leading to cognitive impairment, neurodegeneration, and early death. NPDA and NPDC have different genetic origins, being caused by mutations in the acid sphingomyelinase (ASM) or the cholesterol transport protein NPC1, respectively. However, they share a common pathological hallmark in the accumulation of lipids in the endolysosomal compartment. Here, we tested the hypothesis that polyphenols reduce lipid overload in NPD cells by enhancing the secretion of extracellular vesicles (ECVs). We show that among the polyphenols tested, the ellagic acid metabolites, urolithin A and B, were the safest and most efficient in increasing ECV secretion. They reduced levels of accumulating lipids and lysosomal size and permeabilization in cultured bone marrow-derived macrophages and neurons from ASMko and NPC1 mutant mice, which mimic NPDA and NPDC, respectively. Moreover, oral treatment with ellagic acid reduced lipid levels, ameliorated lysosomal alterations, and diminished microglia activation in the brain of NPD mice. These results support the therapeutic value of ECV secretion and polyphenols for NPDs, which may also help treat other LSDs characterized by intracellular lipid overload.

Hypoxia is a crucial factor contributing to maintenance of atherosclerotic lesions. The ability of ABCA1 to stimulate the efflux of cholesterol from cells in the periphery, particularly foam cells in atherosclerotic plaques, is an important anti-atherosclerotic mechanism. The posttranscriptional regulation by miRNAs represents a key regulatory mechanism of a number of signaling pathways involved in atherosclerosis. Previously, miR-199a-5p has been shown to be implicated in the endocytic and retrograde intracellular transport. Although the regulation of miR-199a-5p and ABCA1 by hypoxia has been already reported independently, the role of miR-199a-5p in macrophages and its possible role in atherogenic processes such us regulation of lipid homeostasis through ABCA1 has not been yet investigated. Here, we demonstrate that both ABCA1 and miR-199a-5p show an inverse regulation by hypoxia and Ac-LDL in primary macrophages. Moreover, we demonstrated that miR-199a-5p regulates ABCA1 mRNA and protein levels by directly binding to its 3’UTR. As a result, manipulation of cellular miR-199a-5p levels alters ABCA1 expression and cholesterol efflux in primary mouse macrophages. Taken together, these results indicate that the correlation between ABCA1-miR-199a-5p could be exploited to control macrophage cholesterol efflux during the onset of atherosclerosis, where cholesterol alterations and hypoxia play a pathogenic role.

The evident implication of the insulin-degrading enzyme (IDE) in Alzheimer’s disease (AD) and type 2 diabetes mellitus (T2DM), among its capacity to degrade insulin and amyloid-β peptide (Aβ), suggests that IDE could be an essential link in the relation between hyperinsulinemia, insulin resistance and AD. However, little is known about the cellular and molecular regulation of IDE expression, and even less has been explored regarding the post-transcriptional regulation of IDE, although it represents a great molecular target of interest for therapeutic treatments. We recently described that miR-7, a novel candidate for linking AD and T2DM at the molecular level, regulates IDE and other key genes in both pathologies, including some key genes involved in the insulin signaling pathway. Here, we explored whether other miRNAs as well as other post-transcriptional regulators, such as RNA binding proteins (RBP), could potentially participate in the regulation of IDE expression in vitro. Our data showed that in addition to miR-7, miR-125, miR-490 and miR-199 regulate IDE expression at the post-transcriptional level. Moreover, we also found that IDE contains multiple potential binding sites for several RBPs, and a narrow-down prediction analysis led us to speculate on a novel regulation of IDE by RALY and HuD. Taken together, these results demonstrate the novel players controlling IDE expression that could represent potential therapeutical targets to treat several metabolic diseases with a high impact on human health, including AD and T2DM.

Exosomes/microvesicles originate from multivesicular bodies that allow the secretion of endolysosome components out of the cell. In the present work, we investigated the effects of rottlerin, a polyphenol, on exosome/microvesicle secretion in a model of intracellular lipid trafficking impairment, and elucidated the mechanism of action. In a model of lipid trafficking impairment in C6 glia cells, rottlerin increased ceramide levels, while decreasing hexosylceramide content. This was accompanied by increased exosome/microvesicle secretion, thereby reducing the concentration of lipids in the endolysosomal compartment. The reduction of hexosylceramide levels by rottlerin was attributed to the increase of β-glucosidase (glucosylceramidase) activity, and the effects of rottlerin were abrogated by β-glucosidase inhibitors such as isofagomine D-tartrate and AMP-deoxynojirimycin. Moreover, treatment with ML-266, a potent activator of the β-glucosidase enzyme, recapitulated the effects of rottlerin on the sphingolipid profile and exosome/microvesicle secretion. Finally, inhibition of AMPK (AMP-activated protein kinase) using compound C prevented both exosome/microvesicle secretion and the elimination of endolysosome lipids, which were promoted by rottlerin. The results showed that the decrease in intracellular lipid deposition induced by rottlerin was mediated by β-glucosidase activation and exosome/microvesicle release via the AMPK pathway. Rottlerin consumption could represent an additional health benefit in lysosomal deposition diseases.

Abstract Atherosclerosis, the major cause of cardiovascular disease, is a chronic inflammatory disease characterized by the accumulation of lipids and inflammatory cells in the artery wall. Aberrant expression of microRNAs has been implicated in the pathophysiological processes underlying the progression of atherosclerosis. Here, we define the contribution of miR‐21 in hematopoietic cells during atherogenesis. Interestingly, we found that miR‐21 is the most abundant miRNA in macrophages and its absence results in accelerated atherosclerosis, plaque necrosis, and vascular inflammation. miR‐21 expression influences foam cell formation, sensitivity to ER‐stress‐induced apoptosis, and phagocytic clearance capacity. Mechanistically, we discovered that the absence of miR‐21 in macrophages increases the expression of the miR‐21 target gene, MKK3, promoting the induction of p38‐CHOP and JNK signaling. Both pathways enhance macrophage apoptosis and promote the post‐translational degradation of ABCG1, a transporter that regulates cholesterol efflux in macrophages. Altogether, these findings reveal a major role for hematopoietic miR‐21 in atherogenesis.

Haloperidol, a typical antipsychotic, has been shown to inhibit cholesterol biosynthesis by affecting Δ7-reductase, Δ8,7-isomerase, and Δ14-reductase activities, which results in the accumulation of different sterol intermediates. In the present work, we investigated the effects of atypical or second-generation antipsychotics (SGA), such as clozapine, risperidone, and ziprasidone, on intracellular lipid metabolism in different cell lines. All the SGAs tested inhibited cholesterol biosynthesis. Ziprasidone and risperidone had the same targets as haloperidol at inhibiting cholesterol biosynthesis, although with different relative activities (ziprasidone > haloperidol > risperidone). In contrast, clozapine mainly affected Δ24-reductase and Δ8,7-isomerase activities. These amphiphilic drugs also interfered with the LDL-derived cholesterol egress from the endosome/lysosome compartment, thus further reducing the cholesterol content in the endoplasmic reticulum. This triggered a homeostatic response with the stimulation of sterol regulatory element-binding protein (SREBP)-regulated gene expression. Treatment with SGAs also increased the synthesis of complex lipids (phospholipids and triacylglycerides). Once the antipsychotics were removed from the medium, a rebound in the cholesterol biosynthesis rate was detected, and the complex-lipid synthesis further increased. In this condition, apolipoprotein B secretion was also stimulated as demonstrated in HepG2 cells. These effects of SGAs on lipid homeostasis may be relevant in the metabolic side effects of antipsychotics, especially hypertriglyceridemia.

There is a tight relationship between fertility and changes in cholesterol metabolism during spermatogenesis. In the testis, class B scavenger receptors (SR-B) SR-BI, SR-BII, and LIMP II mediate the selective uptake of cholesterol esters from HDL, which are hydrolyzed to unesterified cholesterol by hormone-sensitive lipase (HSL). HSL is critical because HSL knockout (KO) male mice are sterile. The aim of the present work was to determine the effects of the lack of HSL in testis on the expression of SR-B, lipid raft composition, and related cell signaling pathways. HSL-KO mouse testis presented altered spermatogenesis associated with decreased sperm counts, sperm motility, and infertility. In wild-type (WT) testis, HSL is expressed in elongated spermatids; SR-BI, in Leydig cells and spermatids; SR-BII, in spermatocytes and spermatids but not in Leydig cells; and LIMP II, in Sertoli and Leydig cells. HSL knockout male mice have increased expression of class B scavenger receptors, disrupted caveolin-1 localization in lipid raft plasma membrane microdomains, and activated phospho-ERK, phospho-AKT, and phospho-SRC in the testis, suggesting that class B scavenger receptors are involved in cholesterol ester uptake for steroidogenesis and spermatogenesis in the testis.

First- and second-generation antipsychotics (FGAs and SGAs, respectively), both inhibit cholesterol biosynthesis and impair the intracellular cholesterol trafficking, leading to lipid accumulation in the late endosome/lysosome compartment. In this study we examined if curcumin, a plant polyphenol that stimulates exosome release, can alleviate antipsychotic-induced intracellular lipid accumulation.HepG2 hepatocarcinoma cells were treated with antipsychotics or placebo and DiI-labelled LDL for 18 h and then exposed to curcumin for the last 2 h. Cells and media were collected separately and used for biochemical analyses, electron microscopy and immunocytochemistry. Exosomes were isolated from the incubation medium by ultracentrifugation.Curcumin treatment reduced the number of heterolysosomes and shifted their subcellular localization to the periphery, as revealed by electron microscopy, and stimulated the release of lysosomal β-hexosaminidase and exosome markers flotillin-2 and CD63 into the media. The presence of DiI in exosomes released by cells preloaded with DiI-LDL demonstrated the endolysosomal origin of the microvesicles. Furthermore, curcumin increased the secretion of cholesterol as well as LDL-derived DiI and [3H]-cholesterol, in association with a decrease of intracellular lipids. Thus, the disruption of lipid trafficking induced by FGAs or SGAs can be relieved by curcumin treatment. This polyphenol, however, did not mitigate the reduction of cholesterol esterification induced by antipsychotics.Curcumin stimulates exosome release to remove cholesterol (and presumably other lipids) accumulated within the endolysosomal compartment, thereby normalizing intracellular lipid homeostasis. This action may help minimize the adverse metabolic effects of antipsychotic treatment, which should now be evaluated in clinical trials.