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Methods and results The internal mammary artery (IMA) is the conduit of choice to bypass the left anterior descending (LAD) coronary artery in patients eligible for coronary artery bypass grafting (CABG). In case of graft occlusion, it is still not clear whether it is better to revascularize the graft or the recipient native vessel, on top of optimal medical therapy. A 55-year-old man, with prior CABG, underwent urgent coronary angiography for an acute coronary syndrome. LAD CTO, severe proximal circumflex artery stenosis, and mild atherosclerosis of the right coronary artery were found. Moreover, a proximal occlusion of the venous graft to the obtuse marginal branch and LIMA CTO within the body of the graft were shown as well. Surprisingly, many well-developed bridging collaterals, arising close to the proximal cap of the LIMA occlusion, supplied the bloodstream circulation distal to the occlusion within the LIMA. At that time, a left main–proximal circumflex artery stenting was performed (3.5×24-mm Xience Sierra, Abbott) because that was judged the culprit lesion. One month later the patient still complained of effort angina and dyspnoea despite optimized antianginal medical therapy. The patient was scheduled for elective antegrade LIMA CTO PCI, considered the most appropriate treatment strategy. Indeed, the presence of a prior stent implanted in the left main– circumflex artery would have made difficult a proximal cap negotiation and its penetration by guidewires in case of antegrade LAD CTO PCI. Thus, the LIMA CTO negotiation was initially attempted with a Fielder FC guidewire (Asahi Intecc) and a supporting Corsair Pro (Asahi Intecc) microcatheter. The polymeric soft wire prolapsed preferentially into the bridging collaterals without the chance to engage the CTO proximal cap. Finally, a moderate-weight hydrophilic GAIA second (Asahi Intecc) succeeded in cap penetration and final CTO crossing. After lesion predilatation with a 2.0×15-mm balloon (Emerge, Boston Scientific), 2 overlapped stents, respectively 2.75×44 mm and 3.0×18 mm (VIVO ISAR, Translumina) were deployed and postdilated by a 3.0×15-mm non-compliant balloon (Accuforce, Terumo). The final result was deemed very satisfactory with a TIMI 3 flow. Conclusions Many studies showed that IMA has active biological functions, providing a sort of protection against atherosclerosis and self-reparative properties. These biological properties might justify the IMA high long-term patency rates as CABG conduits. The development of bridging collaterals in this patient, represents a clear proof of the LIMA self-reparative properties. Whereas bridging collaterals are a common feature of coronary CTO, their development in a LIMA bypass graft represents a unique finding which requires further investigation.

The strongest genetic risk factor for sporadic Alzheimer’s disease (AD) is the presence of the ε4 allele of the apolipoprotein E (ApoE) gene, the major apolipoprotein involved in brain cholesterol homeostasis. Being astrocytes the main producers of cholesterol and ApoE in the brain, we investigated the impact of the ApoE genotype on astrocyte cholesterol homeostasis. Two mouse astrocytic cell lines expressing the human ApoE3 or ApoE4 isoform were employed. Gas chromatography–mass spectrometry (GC-MS) analysis pointed out that the levels of total cholesterol, cholesterol precursors, and various oxysterols are altered in ApoE4 astrocytes. Moreover, the gene expression analysis of more than 40 lipid-related genes by qRT-PCR showed that certain genes are up-regulated (e.g., CYP27A1) and others down-regulated (e.g., PPARγ, LXRα) in ApoE4, compared to ApoE3 astrocytes. Beyond confirming the significant reduction in the levels of PPARγ, a key transcription factor involved in the maintenance of lipid homeostasis, Western blotting showed that both intracellular and secreted ApoE levels are altered in ApoE4 astrocytes, as well as the levels of receptors and transporters involved in lipid uptake/efflux (ABCA1, LDLR, LRP1, and ApoER2). Data showed that the ApoE genotype clearly affects astrocytic cholesterol homeostasis; however, further investigation is needed to clarify the mechanisms underlying these differences and the consequences on neighboring cells. Indeed, drug development aimed at restoring cholesterol homeostasis could be a potential strategy to counteract AD.

Atherosclerosis is a degenerative disease characterized by lesions that develop in the wall of large- and medium-sized arteries due to the accumulation of low-density lipoproteins (LDLs) in the intima. A growing bulk of evidence suggests that cholesterol oxidation products, known as oxysterols, and the aldehyde 4-hydroxy-2-nonenal (HNE), the major pro-atherogenic components of oxidized LDLs, significantly contribute to atherosclerotic plaque progression and destabilization, with eventual plaque rupture. The involvement of certain members of the protein convertase subtilisin/kexin proteases (PCSKs) in atherosclerosis has been recently hypothesized. Among them, PCSK6 has been associated with plaque instability, mainly thanks to its ability to stimulate the activity of matrix metalloproteinases (MMPs) involved in extracellular matrix remodeling and to enhance inflammation. In U937 promonocytic cells and in human umbilical vein endothelial cells, an oxysterol mixture and HNE were able to up-regulate the level and activity of PCSK6, resulting in MMP-9 activation as demonstrated by PCSK6 silencing. Inflammation, enhanced by these lipid oxidation products, plays a key role in the up-regulation of PCSK6 activity as demonstrated by cell pretreatment with NS-398, with epigallocatechin gallate or with acetylsalicylic acid, all with anti-inflammatory effects. For the first time, we demonstrated that both oxysterols and HNE, which substantially accumulate in the atherosclerotic plaque, up-regulate the activity of PCSK6. Of note, we also suggest a potential association between PCSK6 activity and MMP-9 activation, pointing out that PCSK6 could contribute to atherosclerotic plaque development.

Chronic inflammation represents a main event in the onset and progression of atherosclerosis and is closely associated with oxidative stress in a sort of vicious circle that amplifies and sustains all stages of the disease. Key players of atherosclerosis are monocytes/macrophages. According to their pro- or anti-inflammatory phenotype and biological functions, lesional macrophages can release various mediators and enzymes, which in turn contribute to plaque progression and destabilization or, alternatively, lead to its resolution. Among the factors connected to atherosclerotic disease, lipid species carried by low density lipoproteins and pro-oxidant stimuli strongly promote inflammatory events in the vasculature, also by modulating the macrophage phenotyping. Therapies specifically aimed to balance macrophage inflammatory state are increasingly considered as powerful tools to counteract plaque formation and destabilization. In this connection, several molecules of natural origin have been recognized to be active mediators of diverse metabolic and signaling pathways regulating lipid homeostasis, redox state, and inflammation; they are, thus, considered as promising candidates to modulate macrophage responsiveness to pro-atherogenic stimuli. The current knowledge of the capability of nutraceuticals to target macrophage polarization and to counteract atherosclerotic lesion progression, based mainly on in vitro investigation, is summarized in the present review.

Aims The purpose of this study was to evaluate whether microvascular dysfunction is more present in patients with metabolic syndrome (MetS) compared to diabetics and hypertensive patients with two angiographic imaging methods, to evaluate the degree of microcirculation dysfunction, the TIMI frame count and Myocardial Blush grade. Both techniques of rapid use and relatively cheap, and allow us to have a good degree of evaluation referred to the function of the coronary microcirculation. Methods and results The study included 445 patients allocated into three groups, 157 in the MetS group, 128 in the diabetics group, and 160 patients in the hypertensive group. All patients accessed to the emergency room for anginal chest pain, all were hospitalized in the cardiac intensive care unit from 2015 to 2020. Inclusion criteria were the presences of chest pain and/or their positive stress test, and epicardial coronary arteries free from stenosis at coronary angiography. We compared the results obtained from the angiographic techniques (TIMI Frame Count and Myocardial Blush Grade) in the two subgroups: MetS vs. hypertensive, and MetS vs. diabetics. In the first subgroup, we analyzed the TFCs of the three vessels in patients with hypertension and comparing them with patients with MetS, we observed that the latter have a worse perfusion condition: the three epicardial coronary vessels have a higher TFC than the hypertensive population (TFC LAD 33.1 ± 5.6 vs. 28.4 ± 5.6, P = 0.018), (TFC RCA 27.2 ± 5.2 vs. 23.1 ± 5.2, P = 0.014) (TFC CX 27.9 ± 5.4 vs. 26.9 ± 5.4, P = 0.03). That indicates slow flow in patients with MetS coronary microcirculation. Analyzing the MBG, however, in the three coronary vessels of patients with hypertension compared to patients with metabolic syndrome, no difference was found in terms of worsening of the coronary microcirculation. Finally comparing the indices that summarize the values of the individual arteries both for the TFC and MBG, was seen as the TMBS is reduced in both groups (7.1 ± 0.49 vs. 7.1 ± 0.6, P-value = 0.04). The TTFC is instead higher in patients with MetS (83.9 ± 5.8 vs. 77.8 ± 6.7, P-value =0.024). Then we performed the same type of comparison between MetS and type 2 diabetic subgroup, in this comparison we observed how by analyzing the TFCs of the three coronary vessels, MetS patients have a slower coronary flow than patients with type 2 diabetes mellitus (TFC LAD 33.1 ± 5.6 vs. 30.6 ± 6.2, P = 0.04), (TFC RCA 27.2 ± 5.2 vs. 25 ± 5.3, P = 0.02), (TFC CX 27.9 ± 5.4 vs. 27.2 ± 5.6, P = 0.05). Comparing MBG of the three coronary vessels instead, the flow is lower in diabetic patients TTFC was higher in patients with metabolic syndrome (83.9 ± 5.8 vs. 82.7 ± 8.6, P-value = 0.02). While TMBS was lower in diabetic patients than in patients with metabolic syndrome (7.1 ± 0.49 vs. 6.7 ± 0.74, P-value = 0.01). Conclusions This study shows that patients with MetS had a major coronary microvascular dysfunction using TFC imaging technique, analysis compared to diabetics or hypertensive patients, these differences resulted to be statistically significant. A clinical evaluation of this parameters using TFC such in this study, might give further information about (CMD) in this patients in order to act to develop the best treatment to this patients and to improve their clinical condition.

Background and aims In the present study, we aimed to compare the clinical and coronary angiography features between South Asian and Caucasian patients with Acute Coronary Syndrome (ACS). In particular, we focused our analysis on the evaluation of recent cardiovascular risk markers, such as remnant cholesterol, corresponding to all plasma cholesterol minus HDL-C (high-density lipoprotein cholesterol) and LDL-C (low-density lipoprotein cholesterol), and the Monocyte-to-HDL-cholesterol ratio. We also compared values of several lipoprotein ratios and the Platelet-to-lymphocyte ratio, accurate predictors of coronary events and coronary artery disease. Methods and results We recruited 40 South Asian and 40 Caucasian patients admitted for ACS. Data were collected by consulting patients' medical records. We used Chi-square test and Student's t-test to analyse qualitative and quantitative variables, respectively. South Asian patients, compared to Caucasians, showed higher mean values of the parameters analysed: remnant cholesterol (32.6 ± 17 vs 26.5 ± 9.6), Monocyte-to-HDL-cholesterol ratio (26.4 ± 48.7 vs 16.5 ± 8.3), Platelet-to-lymphocyte ratio (124.7 ± 130.7 vs 120.5 ± 58.8). Moreover, higher mean values of several lipoprotein ratios were also found in South Asian patients compared to the control group. However, statistical significance was not reached for any of these differences observed. Conclusions The evaluation of the parameters analysed in this study might provide accurate information regarding the cardio-metabolic risk in South Asian patients. However, further studies with larger samples are needed to obtain more significant results.

High amounts of cholesterol have been definitely associated with the pathogenesis of several diseases, including metabolic and neurodegenerative disorders, cardiovascular diseases, and cancer. In all these pathologies the exacerbation of pro-oxidant and inflammatory responses is a consistent feature. In this scenario, species derived from enzymatic and non-enzymatic cholesterol oxidation, namely oxysterols, are strongly suspected to play a primary role. The consideration of these bioactive lipids is therefore helpful in investigating pathological mechanisms and may also acquire clinical value for the diagnosis and treatment of diseases. For this purpose and considering that a great number of oxysterols may be present together in the body, the employment of lipidomics technology certainly represents a powerful strategy for the simultaneous detection and characterization of these compounds in biological specimens. In this review, we will discuss the applicability of the lipidomics approach in the study of the association between oxysterols and diseases.

Myocardial infarction with non-obstructive coronary arteries (MINOCA) is defined by clinical evidence of myocardial infarction (MI) with normal or near-normal coronary arteries on angiography. This condition is present in about 5% to 25% of patients presenting with acute coronary syndromes. MINOCA is a working diagnosis. Current guidelines and consensus recommend identification of underlying causes of MINOCA in order to optimize treatment, improve prognosis, and promote prevention of recurrent myocardial infarction. An accurate evaluation of patient history, symptoms and use of invasive and non-invasive imaging should lead to identification of epicardial or microvascular causes of MINOCA and differentiation from non-ischemic myocardial injury due to both cardiac (e.g. myocarditis) and non-cardiac disease (e.g. pulmonary embolism). In this review, we highlight the role of coronary imaging in differential diagnosis of patients presenting with MINOCA. Intravascular ultrasound and optical coherence tomography are well known technologies used in different settings from acute to chronic coronary syndromes. In MINOCA patients, coronary imaging could help to identify pathological alterations of the epicardial vessels that are not visible by coronary angiography such as plaque disruption, coronary dissection, coronary thromboembolism, coronary spasm, and coronary artery disease in patients presenting with takotsubo syndrome. In future, the widespread use of these technologies, in the right clinical context, could lead to optimization and personalization of treatment, and to better prognosis of patients presenting with MINOCA.

Background: Among Alzheimer’s disease (AD) brain hallmarks, the presence of reactive astrocytes was demonstrated to correlate with neuronal loss and cognitive deficits. Evidence indeed supports the role of reactive astrocytes as mediators of changes in neurons, including synapses. However, the complexity and the outcomes of astrocyte reactivity are far from being completely elucidated. Another key role in AD pathogenesis is played by alterations in brain cholesterol metabolism. Oxysterols (cholesterol oxidation products) are crucial for brain cholesterol homeostasis, and we previously demonstrated that changes in the brain levels of various oxysterols correlate with AD progression. Moreover, oxysterols have been shown to contribute to various pathological mechanisms involved in AD pathogenesis.In order to deepen the role of oxysterols in AD, we investigated whether they could contribute to astrocyte reactivity, and consequently impact on neuronal health.Methods: Mouse primary astrocyte cultures were used to test the effect of two oxysterol mixtures, that represent the oxysterol composition respectively of mild or severe AD brains, on astrocyte morphology, markers of reactivity, and secretion profile. Co-culture experiments were performed to investigate the impact of oxysterol-treated astrocytes on neurons. Neuronal cultures were exposed to astrocyte conditioned media (ACM) deprived of lipocalin-2 (Lcn2) to investigate the contribution of this mediator to synaptotoxicity.Results: Results showed that oxysterols induce a clear morphological change in astrocytes, accompanied by the upregulation of some reactive astrocyte markers, including Lcn2. Moreover, ACM analysis revealed a significant increase in the release of Lcn2, cytokines, and chemokines in response to oxysterols. A significant reduction of postsynaptic density protein 95 (PSD95) and a concurrent increase in cleaved caspase-3 protein levels have been demonstrated in neurons co-cultured with oxysterol-treated astrocytes, pointing out that mediators released by astrocytes have an impact on neurons. Among these mediators, Lcn2 has been demonstrated to play a major role on synapses, affecting neurite morphology and decreasing dendritic spine density. Conclusions: These data demonstrated that oxysterols present in the AD brain promote astrocyte reactivity, determining the release of several mediators that affect neuronal health and synapses. Lcn2 has been shown to exert a key role in mediating the synaptotoxic effect of oxysterol-treated astrocytes.

A growing bulk of evidence suggests that cholesterol oxidation products, known as oxysterols, are potentially involved in the pathogenesis of major chronic diseases, including atherosclerosis, Alzheimer's disease, and inflammatory bowel disease. Oxysterols are involved in various key steps of these complex processes, mainly thanks to their ability to act through up-regulation of oxidative stress, inflammation, and cell toxicity. This review summarizes the current knowledge of the effects induced by these compounds on cells, after their accumulation in the arterial wall, brain, and intestine. This evidence might help to develop innovative strategies to counteract the progression of these chronic inflammatory human diseases.

In recent decades, the impairment of cholesterol metabolism in the pathogenesis of Alzheimer’s disease (AD) has been intensively investigated, and it has been recognized to affect amyloid β (Aβ) production and clearance, tau phosphorylation, neuroinflammation and degeneration. In particular, the key role of cholesterol oxidation products, named oxysterols, has emerged. Brain cholesterol metabolism is independent from that of peripheral tissues and it must be preserved in order to guarantee cerebral functions. Among the cells that help maintain brain cholesterol homeostasis, astrocytes play a starring role since they deliver de novo synthesized cholesterol to neurons. In addition, other physiological roles of astrocytes are to modulate synaptic transmission and plasticity and support neurons providing energy. In the AD brain, astrocytes undergo significant morphological and functional changes that contribute to AD onset and development. However, the extent of this contribution and the role played by oxysterols are still unclear. Here we review the current understanding of the physiological role exerted by astrocytes in the brain and their contribution to AD pathogenesis. In particular, we focus on the impact of cholesterol dysmetabolism on astrocyte functions suggesting new potential approaches to develop therapeutic strategies aimed at counteracting AD development.

In Alzheimer’s disease (AD), both cholesterol and glucose dysmetabolism precede the onset of memory deficit and contribute to the disease’s progression. It is indeed now believed that oxidized cholesterol in the form of oxysterols and altered glucose uptake are the main triggers in AD affecting production and clearance of Aβ, and tau phosphorylation. However, only a few studies highlight the relationship between them, suggesting the importance of further extensive studies on this topic. Recently, a molecular link was demonstrated between cholesterol oxidative metabolism and glucose uptake in the brain. In particular, 27-hydroxycholesterol, a key linker between hypercholesterolemia and the increased AD risk, is considered a biomarker for reduced glucose metabolism. In fact, its excess increases the activity of the renin-angiotensin system in the brain, thus reducing insulin-mediated glucose uptake, which has a major impact on brain functioning. Despite this important evidence regarding the role of 27-hydroxycholesterol in regulating glucose uptake by neurons, the involvement of other cholesterol oxidation products that have been clearly demonstrated to be key players in AD cannot be ruled out. This review highlights the current understanding of the potential role of cholesterol and glucose dysmetabolism in AD progression, and the bidirectional crosstalk between these two phenomena.

Alzheimer's disease (AD) is a gradually debilitating disease that leads to dementia. The molecular mechanisms underlying AD are still not clear, and at present no reliable biomarkers are available for the early diagnosis. In the last several years, together with oxidative stress and neuroinflammation, altered cholesterol metabolism in the brain has become increasingly implicated in AD progression. A significant body of evidence indicates that oxidized cholesterol, in the form of oxysterols, is one of the main triggers of AD. The oxysterols potentially most closely involved in the pathogenesis of AD are 24-hydroxycholesterol and 27-hydroxycholesterol, respectively deriving from cholesterol oxidation by the enzymes CYP46A1 and CYP27A1. However, the possible involvement of oxysterols resulting from cholesterol autooxidation, including 7-ketocholesterol and 7β-hydroxycholesterol, is now emerging. In a systematic analysis of oxysterols in post-mortem human AD brains, classified by the Braak staging system of neurofibrillary pathology, alongside the two oxysterols of enzymatic origin, a variety of oxysterols deriving from cholesterol autoxidation were identified; these included 7-ketocholesterol, 7α-hydroxycholesterol, 4β-hydroxycholesterol, 5α,6α-epoxycholesterol, and 5β,6β-epoxycholesterol. Their levels were quantified and compared across the disease stages. Some inflammatory mediators, and the proteolytic enzyme matrix metalloprotease-9, were also found to be enhanced in the brains, depending on disease progression. This highlights the pathogenic association between the trends of inflammatory molecules and oxysterol levels during the evolution of AD. Conversely, sirtuin 1, an enzyme that regulates several pathways involved in the anti-inflammatory response, was reduced markedly with the progression of AD, supporting the hypothesis that the loss of sirtuin 1 might play a key role in AD. Taken together, these results strongly support the association between changes in oxysterol levels and AD progression., Highlights • Changes in brain oxysterol levels may influence AD progression. • Oxysterol accumulation in the brain may amplify neuroinflammation. • SIRT-1 loss during AD progression may favor neuroinflammation. • Oxysterols and SIRT-1 might be useful markers for early AD diagnosis.

Cholesterol oxidation products such as oxysterols are considered critical factors in the atherosclerotic plaque formation since they induce oxidative stress, inflammation and apoptotic cell death. 27-hydroxycholesterol (27-OH) is one of the most represented oxysterols in atherosclerotic lesions. We recently showed that relatively low concentrations of 27-OH generated a strong survival signaling through an early and transient increase of cellular ROS level, that enhanced MEK-ERK/PI3K-Akt phosphorylation, in turn responsible of a sustained quenching of ROS production. It remains to identify the link between ERK/Akt up-regulation and the consequent quenching effect on ROS intracellular level that efficiently and markedly delay the pro-apoptotic effect of the oxysterol. Here we report on the potent activation of Nrf2 redox-sensitive transcription factor by low micromolar amount of 27-OH added to U937 promonocytic cells. The 27-OH-exerted induction of Nrf2 and subsequently of the target genes, HO-1 and NQO-1, was proved to be: (i) dependent upon the activation of ERK and Akt pathways, (ii) directly responsible for the quenching of intracellular oxidative stress and by this way (iii) ultimately responsible for the observed oxysterol-induced pro-survival response. (C) 2015 Elsevier Inc. All rights reserved.

The ageing endothelium progressively loses its remarkable and crucial ability to maintain homeostasis of the vasculature, as it acquires a proinflammatory phenotype. Cellular and structural changes gradually accumulate in the blood vessels, and markedly in artery walls. Most changes in aged arteries are comparable to those occurring during the atherogenic process, the latter being more marked: pro-oxidant and proinflammatory molecules, mainly deriving from or triggered by oxidized low density lipoproteins (oxLDLs), are undoubtedly a major driving force of this process. Oxysterols, quantitatively relevant components of oxLDLs, are likely candidate molecules in the pathogenesis of vascular ageing, because of their marked pro-oxidant, proinflammatory and proapoptotic properties. An increasing bulk of experimental data point to the contribution of a variety of oxysterols of pathophysiological interest, also in the age-related genesis of endothelium dysfunction, intimal thickening due to lipid accumulation, and smooth muscle cell migration and arterial stiffness due to increasing collagen deposition and calcification. This review provides an updated analysis of the molecular mechanisms whereby oxysterols accumulating in the wall of ageing blood vessels may 'activate' endothelial and monocytic cells, through expression of an inflammatory phenotype, and 'convince' smooth muscle cells to proliferate, migrate and, above all, to act as fibroblast-like cells.

A novel innate immune strategy, involving specific cholesterol oxidation products as effectors, has begun to reveal connections between cholesterol metabolism and immune response against viral infections. Indeed, 25-hydroxycholesterol (25HC) and 27-hydroxycholesterol (27HC), physiologically produced by enzymatic oxidation of cholesterol, act as inhibitors of a wide spectrum of enveloped and non-enveloped human viruses. However, the mechanisms underlying their protective effects against non-enveloped viruses are almost completely unexplored. To get insight into this field, we investigated the antiviral activity of 25HC and 27HC against a non-enveloped virus causing acute gastroenteritis in children, the human rotavirus (HRV). We found that 25HC and 27HC block the infectivity of several HRV strains at 50% inhibitory concentrations in the low micromolar range in the absence of cell toxicity. Both molecules affect the final step of virus penetration into cells by preventing the association of two cellular proteins: the oxysterol binding protein (OSBP) and the vesicle-associated membrane protein-associated protein-A (VAP-A). By altering the activity of these cellular mediators, 25HC and 27HC disturb the recycling of cholesterol between the endoplasmic reticulum and the late endosomes which are exploited by HRV to penetrate into the cell. The substantial accumulation of cholesterol in the late endosomal compartment results in sequestering viral particles inside these vesicles thereby preventing cytoplasmic virus replication. These findings suggest that cholesterol oxidation products of enzymatic origin might be primary effectors of host restriction strategies to counteract HRV infection and point to redox active lipids involvement in viral infections as a research area of focus to better focus in order to identify novel antiviral agents targets.

It is now established that cholesterol oxidation products (oxysterols) are involved in several events underlying Alzheimer's disease (AD) pathogenesis. Of note, certain oxysterols cause neuron dysfunction and degeneration but, recently, some of them have been shown also to have neuroprotective effects. The present study, which aimed to understand the potential effects of 24-hydroxycholesterol (24-OH) against the intraneuronal accumulation of hyperphosphorylated tau protein, stressed these latter effects. A beneficial effect of 24-OH was demonstrated in SK-N-BE neuroblastoma cells, and is due to its ability to modulate the deacetylase sirtuin 1 (SIRT1), which contributes to preventing the neurotoxic accumulation of the hyperphosphorylated tau protein. Unlike 24-OH, 7-ketocholesterol (7-K) did not modulate the SIRT1-dependent neuroprotective pathway. To confirm the neuroprotective role of 24-OH, in vivo experiments were run on mice that express human tau without spontaneously developing tau pathology (hTau mice), by means of the intracerebroventricular injection of 24-OH. 24-OH, unlike 7-K, was found to completely prevent the hyperphosphorylation of tau induced by amyloid β monomers. These data highlight the importance of preventing the loss of 24-OH in the brain, and of maintaining high levels of the enzyme SIRT1, in order to counteract neurodegeneration. Graphical abstract: A hypothetical scheme of the molecular mechanisms underlying the effects of 24-OH on hyperphosphorylated tau accumulation.fx1 Keywords: 24-hydroxycholesterol, Sirtuin 1, Tau, Oxysterols, Alzheimer's disease

Dietary oxysterols are cholesterol auto-oxidation products widely present in cholesterol-rich foods. They are thought to affect the intestinal barrier function, playing a role in gut inflammation. This study has characterized specific cell signals that are up-regulated in differentiated CaCo-2 colonic epithelial cells by a mixture of oxysterols representative of a hyper-cholesterolemic diet. p38 MAPK activation plays a major role, while other signal branches, i.e. the JNK and ERK pathways, make minor contributions to the intestinal inflammation induced by dietary oxysterols. p38 transduction might be the missing link connecting the known NADPH oxidase activation, and the induction of NF-κB-dependent inflammatory events related to oxysterols' action in the intestine. A NOX1/p38 MAPK/NF-κB signaling axis was demonstrated by the quenched inflammation observed on blocking individual branches of this signal with specific chemical inhibitors. Furthermore, all these signaling sites were prevented when CaCo-2 cells were pre-incubated with phenolic compounds extracted from selected wines made of typical Sardinian grape varieties: red Cannonau and white Vermentino. Notably, Cannonau was more effective than Vermentino. The effect of Sardinian wine extracts on intestinal inflammation induced by dietary oxysterols might mainly be due to their phenolic content, more abundant in Cannonau than in Vermentino. Furthermore, among different phenolic components of both wines, epicatechin and caffeic acid exerted the strongest effects. These findings show a major role of the NOX1/p38 MAPK/NF-κB signaling axis in the activation of oxysterol-dependent intestinal inflammation, and confirm the concept that phenolics act as modulators at different sites of pro-oxidant and pro-inflammatory cell signals.

Cholesterol oxidation products, termed oxysterols, have been shown to be more reactive than unoxidized cholesterol, possessing marked pro-inflammatory and cytotoxic effects in a number of cells and tissues. Oxysterols, absorbed with the diet as products of cholesterol auto-oxidation, have recently been suggested to potentially interfere with homeostasis of the mucosal intestinal epithelium, by promoting and sustaining irreversible damage. However, the treatment of colon cancer cells with a diet-compatible mixture of oxysterols does not elicit the same responses than individual components added to the cells at the same concentrations at which they are present in the mixture. Sixty μM oxysterol mixture showed a slight pro-apoptotic effect on human colon cancer CaCo-2 cell line, evaluated in terms of caspase-3 and caspase-7 activation; conversely, 7α-hydroxycholesterol, 7β-hydroxycholesterol and 5α,6α-epoxycholesterol were identified to be able to induce a significant pro-apoptotic effect if added to cell culture singly; 7β-hydroxycholesterol had stronger action than other compounds. The enhanced production of reactive oxygen species through up-regulation of the colonic NADPH-oxidase isoform NOX1 appeared to be the key event in oxysterol-induced apoptosis in these colon cancer cells. As regards pro-inflammatory effects of oxysterols, IL-8 and MCP-1 were evaluated for their chemotactic activity. Only MCP-1 production was significantly induced by 7β-hydroxycholesterol, as well as by cholesterol and oxysterol mixture. However, oxysterol-induced inflammation appeared to be NOX1-independent, suggesting a secondary role of this enzyme in inducing inflammation in colon cancer cells. A selective cell death induced by specific oxysterols against colon cancer cells, mainly exploiting their ability to activate NOX1 in generating oxidative reactions, might represent a promising field of investigation in colorectal cancer, and might bring new insights on strategies in anticancer therapy.

The hypercholesterolemia-atherosclerosis association is now established; hypercholesterolemia may induce vascular-cell activation, subsequently increasing expression of adhesion molecules, cytokines, chemokines, growth factors, and other key inflammatory molecules. Among inflammatory molecules expressed by vascular cells, integrins play a critical role in regulating macrophage activation and migration to the site of inflammation, by mediating cell-cell and cell-extracellular matrix interactions. The main lipid oxidation products present in oxidized LDL that may be responsible for inflammatory processes in atherogenesis, are cholesterol oxidation products, known as oxysterols. This study demonstrates the effect of an oxysterol mixture, compatible with that detectable in human hypercholesterolemic plasma, on the expression and synthesis of β1-integrin in cells of the macrophage lineage. The molecular signaling whereby oxysterols induce β1-integrin up-regulation is also comprehensively investigated. Over-expression of β1-integrin depends on activation of classic and novel members of protein kinase C and extracellular signal-regulated kinases 1 and 2, as well as of the up-stream G-protein (Gq and G13), c-Src, and phospholipase C. In addition, the localization of β1-integrin in advanced human carotid plaques is highlighted, marking its importance in atherosclerotic plaque progression.

Alzheimer's disease (AD), the most common form of dementia, is characterized by the progressive loss of neurons and synapses, and by extracellular deposits of amyloid-β (Aβ) as senile plaques, Aβ deposits in the cerebral blood vessels, and intracellular inclusions of hyperphosphorylated tau in the form of neurofibrillary tangles. Several mechanisms contribute to AD development and progression, and increasing epidemiological and molecular evidence suggests a key role of cholesterol in its initiation and progression. Altered cholesterol metabolism and hypercholesterolemia appear to play fundamental roles in amyloid plaque formation and tau hyperphosphorylation. Over the last decade, growing evidence supports the idea that cholesterol oxidation products, known as oxysterols, may be the missing link between altered brain cholesterol metabolism and AD pathogenesis, as their involvement in neurotoxicity, mainly by interacting with Aβ peptides, is reported.

Takotsubo cardiomyopathy, or broken heart syndrome, is characterized by transient left ventricular dysfunction associated to chest pain, elevation of cardiac enzymes, and electrocardiographic changes, mimicking an acute coronary syndrome, especially in older women after a physical or emotional stress. It is extremely infrequent in children as well as after infective stress. We described a celiac 4-year-old girl, following a gluten-free diet, who developed features of cardiac failure few days after episodes of acute diarrhea with fever. The patient was treated with oral anticongestive therapy and intravenous immunoglobulins, and she had a dramatic and rapid improvement; echocardiographic features normalized in 48 hours.

The age-related increased impedance in large arteries overloads the senescent heart, and the myocardial phenotype is hypertrophic. Together with qualitative changes observed in the senile heart, this can be responsible for impaired diastolic function. A restricted diet providing adequate nutrient intake, e.g. alternate-day fasting (ADF), has been shown to extend life-span and decrease incidence and progression of age-associated diseases in laboratory rodents, and to ameliorate some metabolic markers of aging in rhesus monkeys and humans. This study reports an age-related increase of some biological and morphological hypertrophy markers in the rat heart, together with increased plasma BNP, a well known marker of heart failure. The tissue modifications might likely be related to hyper-activation of two of the signaling pathways associated with myocardial pathological hypertrophy: ERK1/2 and PI3Kγ. Increased ERK1/2 activation might be in part related to the disturbance of STAT3, with a consequent decrease of SOCS3. In this context, the down-modulation of ERK1/2 and PI3Kγ signaling, together with the restoration of STAT3 activity and SOCS3 content, both observed with ADF, might help to reduce pathological hypertrophy stimuli and to rescue an important cardioprotective pathway, possibly opening new preventive and therapeutic perspectives in age-related heart failure.

All three cholesterol oxidation products implicated thus far in the pathogenesis of Alzheimer's disease, 7β-hydroxycholesterol, 24-hydroxycholesterol, and 27-hydroxycholesterol, markedly enhance the binding of amyloid-beta (Aβ) to human differentiated neuronal cell lines (SK-N-BE and NT-2) by up-regulating net expression and synthesis of CD36 and β1-integrin receptors. However, only 24-hydroxycholesterol markedly potentiates the pro-apoptotic and pro-necrogenic effects of Aβ(1-42) peptide on these cells: 7β-hydroxycholesterol and 27-hydroxycholesterol, like unoxidized cholesterol, show no potentiating effect. This peculiar behavior of 24-hydroxycholesterol at physiologic concentrations (1 μm) depends on its strong enhancement of the intracellular generation of NADPH oxidase-dependent reactive oxygen species (ROS), mainly H(2) O(2) , and the consequent impairment of neuronal cell redox equilibrium, measured in terms of the GSSG/GSH ratio. Cell incubation with antioxidants quercetin or genistein prevents 24-hydroxycholesterol's pro-oxidant effect and potentiation of Aβ-induced necrosis and apoptosis. Thus, the presence of 24-hydroxycholesterol in the close vicinity of amyloid plaques appears to enhance the adhesion of large amounts of Aβ to the plasma membrane of neurons and then to amplify the neurotoxic action of Aβ by locally increasing ROS steady-state levels. This report further supports a primary involvement of altered brain cholesterol metabolism in the complex pathogenesis of Alzheimer's disease.

4-hydroxynonenal (HNE), a lipid peroxidation product, is a promising anti-neoplastic drug due to its remarkable anti-cancer activities. However, this possibility has not been explored, because the delivery of HNE is very challenging as a result of its low solubility and its poor stability. This study intentionally designed a new type of lipid nanocapsules specifically for HNE delivery. They consist of a medium chain triglyceride liquid oil core surrounded by a polymer shell. A β-cyclodextrin-poly(4-acryloylmorpholine) conjugate was selected as the shell component. HNE-loaded nanocapsules were about 350 nm in size with a negative surface charge. They were stable for two years when stored in suspensions at 4 degrees C. In vitro experiments showed that HNE was released from the nanocapsules at a considerable rate. Nanocapsule uptake into cells was evaluated using a fluorescent formulation that revealed rapid internalisation. Cytotoxicity studies demonstrated the safety of the formulation. Enhanced anti-tumoral activity against various cell lines, depending on increased HNE stability, was obtained by using HNE-loaded nanocapsules. In particular, we have demonstrated an increase in anti-proliferative, pro-apoptotic and differentiative activity in several tumour cell lines from different tissues. Moreover, we evaluated the effects of these new nanocapsules on a three-dimensional human reconstructed model of skin melanoma. Interestingly, the encouraging results obtained with topical administration on the epidermal surface could open new perspectives in melanoma treatments.

Objective: to evaluate the incidence of coronary anomalies (CAA) and risk factors for coronary involvement in children with KD from 2000 through 2013 in Emilia-Romagna. Design: an 11-centers retrospective study was conducted using data from 132 patients diagnosed with KD. Results: during the acute phase, CAA developed in 13/132 pts (9.8%): 2 pts had isolated ectasia of right coronary artery (RCA), 10 pts had ectasia of left anterior descending CA (LAD), 1 pt had aneurysm of LAD, 1 pt had ectasia of circonflex CA (CX). During the subacute phase 14/117 pts (11.9%) presented CAA: 3 had aneurysm of LAD, 1 had aneurysm of LAD and RCA, 1 had isolated ectasia of RCA, 9 had ectasia of LAD. During the convalescent phase CAA were detected in 18/83 pts (21%): 3 had aneurysm of LAD and RCA, 2 isolated aneurysm of LAD, 1 had aneurysm of RCA, 12 ectasia of LAD. 3 out of 6 aneurysms detected in theconvalescent phase developed in the subacute stage. Univariate analyses identified WBC and RBC in the subacute phase, and PLT in chronic phase as predictors of chronic CAA (respectively p 0.04, 0.013 and 0.002). A multivariate logistic regression analysis revealed that only PLT of the chronic stage was predictor for chronic CAA. ROC curve for PLT identified a value superior to 652000/mm3 to be the cut off value for CAA, with an 72% sensitvity and 87% specificity. Incomplete or atipical clinical presentation was correlated with CAA (p 0.02). Younger age (< 1 yr) at diagnosis, seasonal distribution, IVIG responsivness, race, elevation of ALT, percentage of neutrophil, hemoglobin, CRP and ESR, albumin and sodium were not correlated with risk for CAL. Conclusions: atipical/incomplete clinical presentation, higher WBC and RBC in the subacute phase and PLT of the convalescent phase were predictors of CAA in our region, while blood exams of the acute stage, IVIG responsivness, sex and younger age were not.

We updated our regional database for KD patients from 2011 to 2014 to investigate differences in epidemiological, clinical and outcome data between 2011-2014 pts and those diagnosed between 2000-2010 in Emilia Romagna. We had 77 KD patients during last 4 years ( vs 81 pts during 2000-2010), mean age at diagnosis 40 months+- 36,4 (vs 31,2 mths ), 10% were younger than 6 months at diagnosis (vs 14%), male were 60 % (vs 59%). Seasonality was as follows: spring 27 (% vs 33%), winter 25% (vs 28%), autumn 25% (vs 26%), summer 23% (vs 14%). Clinical presentation was complete in 69 % (vs vs 74%), incomplete in 30% (vs 26%), atipical 1 % (vs 0%). IVIG responder were 77% (vs 67%), non responders 16 % (vs 12%), late treatment 2 % (vs 12% P 0.002), not treated with IVIG 5 % (vs 9%). 18/77 pts ( vs 7 of 2000-2010,P 0.004 ) had alteration of ALT ( v> 45 U/) and AST ( v > 55 U/l) and among those with transminases alteration 8 ( vs 4 pts) had cholestasis. Coronary arteries anomalies (CAA) were present in 15% ( 9 % ectasia and 6% (vs 2 %) aneurysms ( mean age 12,6 months, 2 pts non responders to standard therapy, 1 pts with late treatment, 1 pts not treated, and 1 pts well treated. The 2 patients who developed giant aneurysms were both non responder to standard therapy, 1 pts had associated cholestasis. Conclusion: during the last four years we observed an higher number of diagnosis of KD than 2000-2010; incomplete and atipical cases, and non-responder pts were more frequent but we noted a decreased of pts not treated or delayed treated; more cases of cholestasis associated to KD and an increased number of patients with aneurysms.

It is now thought that atherosclerosis, although due to increased plasma lipids, is mainly the consequence of a complicated inflammatory process, with immune responses at the different stages of plaque development. Increasing evidence points to a significant role of Toll-like receptor 4 (TLR4), a key player in innate immunity, in the pathogenesis of atherosclerosis. This study aimed to determine the effects on TLR4 activation of two reactive oxidized lipids carried by oxidized low-density lipoproteins, the oxysterol 27-hydroxycholesterol (27-OH) and the aldehyde 4-hydroxynonenal (HNE), both of which accumulate in atherosclerotic plaques and play a key role in the pathogenesis of atherosclerosis. Secondarily, it examined their potential involvement in mediating inflammation and extracellular matrix degradation, the hallmarks of high-risk atherosclerotic unstable plaques. In human promonocytic U937 cells, both 27-OH and HNE were found to enhance cell release of IL-8, IL-1β, and TNF-α and to upregulate matrix metalloproteinase-9 (MMP-9) via TLR4/NF-κB-dependent pathway; these actions may sustain the inflammatory response and matrix degradation that lead to atherosclerotic plaque instability and to their rupture. Using specific antibodies, it was also demonstrated that these inflammatory cytokines increase MMP-9 upregulation, thus enhancing the release of this matrix-degrading enzyme by macrophage cells and contributing to plaque instability. These innovative results suggest that, by accumulating in atherosclerotic plaques, the two oxidized lipids may contribute to plaque instability and rupture. They appear to do so by sustaining the release of inflammatory molecules and MMP-9 by inflammatory and immune cells, for example, macrophages, through activation of TLR4 and its NF-κB downstream signaling.

A still growing bulk of evidence underlies the multifaceted biochemical properties of oxysterols, several of them of clear relevance to human pathophysiology. Taken up by cells through both vesicular and non vesicular ways or often generated intracellularly, oxysterols contribute to modulate the inflammatory response of a given tissue but also cell viability, metabolism and function. The signaling pathways and the transcription factors whose activation they can influence, often through redox-mediated reactions, are quite a number. Moreover, the outcome of the complex network of intracellular reactions promoted by oxysterols appears to be largely dependent upon specific features and dynamic conditions of the cellular and tissue environment but also on their actual concentration in the site of action. Here focus was given to their potential implication in the advanced progression of atherosclerosis, whosehypercholesterolemia is a recognized primary risk factor. A significant contribution to the possible rupture of atherosclerotic plaque could be provided by defined oxysterols, in particular 27-hydroxycholesterol. Indeed, the latter oxysterol was shown in a promonocytic cell line to amplify the inflammatory reaction and activate several pathways leading to a marked up-regulation of metalloproteases, the key enzymes involved in cap matrix degradation and weakening.

Altered cholesterol metabolism in the brain is implicated both in the initiation and in the progression of Alzheimer’s disease (AD), and hypercholesterolemia is a potential risk factor. In particular, oxidized cholesterol, in the form of oxysterols, is believed to be one of the main triggers of AD. Oxysterols can accumulate in the brain, where they can behave as friends or foes: they cause neuron dysfunction and degeneration and contribute to neuroinflammation and amyloidogenesis, but they can also be neuroprotective. One of the main oxysterols most closely involved in AD pathogenesis is 24-hydroxycholesterol (24OH), also called cerebrosterol. The levels of 24OH are markedly reduced in the brain during AD progression, and it is likely that its reduction may accelerate the disease development, considering that it’s known from literature that 24OH may have neuroprotective effects. Moreover, the loss of 24OH proceeds in parallel with the reduction of sirtuin 1 (SIRT1), a deacetylase promoting neuroprotection. We demonstrated that 24OH activates the neuroprotective axis ROS/SIRT1/PGC1a in neuroblastoma cells, thus favoring Tau deacetylation and preventing Tau hyperphosphorylation. Thus, it would be clinically important to prevent the loss of 24OH in the brain, in order to stimulate long upon the expression of SIRT1 with the consequent delay of Tau pathology.

The oxysterol 27-hydroxycholesterol (27-OH) is increasingly considered to be involved in a variety of pathophysiological processes, having been shown to modulate cell proliferation and metabolism, and also to exert proinflammatory and proapoptotic effects. This study aimed to elucidate the molecular pathways whereby 27-OH may generate survival signals in cells of the macrophage lineage, and to clarify whether its known prooxidant effect is involved in that process. A net up-regulation of survival signaling, involving the extracellular signal-regulated kinase (ERIC) and phosphoinositide 3-kinase (PI3K)/Akt phosphorylation pathways, was observed in U937 promonocytic cells cultivated over time in the presence of a low micromolar concentration of the oxysterol. Interestingly, the up-regulation of both kinases was shown to be closely dependent on an early 27-OH-induced intracellular increase of reactive oxygen species (ROS). In turn, stimulation of ERR and PI3K/Akt both significantly quenched ROS steady state and markedly phosphorylated Bad, thereby determining a marked delay of the oxysterol's proapoptotic action. The 27-OH-induced survival pathways thus appear to be redox modulated and, if they occur within or nearby inflammatory cells during progression of chronic diseases such as cancer and atherosclerosis, they could significantly impact the growth and evolution of such diseases. (C) 2014 Elsevier Inc. All rights reserved.

Oxysterols are a family of 27-carbon molecules originated from cholesterol oxidation and the atherogenic potential of oxysterols is linked to their ability to induce oxidative stress, inflammation and apoptotic cell death. Apparently, these compounds are able to modulate not only pro-apoptotic but also anti-apoptotic signals in targeted cells; however, their anti-apoptotic effect has not been investigated in depth. Hence, we aimed to elucidate the molecular mechanisms underlying the survival signaling elicited by 27-hydroxycholesterol (27-OH) which is the most represented oxysterol in human blood. Using human promonocytic cells (U937) challenged with a relatively low concentration of 27-OH, a strong survival signaling through an early and transient increase of cellular reactive oxygen species (ROS) level, that enhanced MEK-ERK/PI3K-Akt phosphorylation, in turn responsible of a sustained quenching of ROS production was observed. Notably, involvement of antioxidant Nrf2 and its target genes, HO-1 and NQO-1 in this early survival response were shown. It thus appears that Nrf2 is responsible for the quenching of the oxidative imbalance exerted in 27-OH challenged cells. On the other hand, relatively high micromolar amount of 27- OH did not stimulate at all the investigated survival signaling with a constant prooxidant effect. The results of ongoing experiments on the actual role of autophagy in modulating the survival response will be also presented and discussed.The data obtained highlight oxysterols’ ability to promote cell survival that might contribute to the pathogenesis of inflammation-driven chronic diseases such as atherosclerosis.

The hypercholesterolemia-atherosclerosis association is now established; hypercholesterolemia may induce vascular-cell activation, subsequently increasing expression of adhesion molecules, cytokines, chemokines, growth factors, and other key inflammatory molecules. Among inflammatory molecules expressed by vascular cells, integrins play a critical role in regulating macrophage activation and migration to the site of inflammation, by mediating cell-cell and cell-extracellular matrix interactions. The main lipid oxidation products present in oxidized LDL that may be responsible for inflammatory processes in atherogenesis, are cholesterol oxidation products, known as oxysterols. This study demonstrates the effect of an oxysterol mixture, compatible with that detectable in human hypercholesterolemic plasma, on the expression and synthesis of β(1)-integrin in cells of the macrophage lineage. The molecular signaling whereby oxysterols induce β(1)-integrin up-regulation is also comprehensively investigated. Over-expression of β(1)-integrin depends on activation of classic and novel members of protein kinase C and extracellular signal-regulated kinases 1 and 2, as well as of the up-stream G-protein (Gq and G13), c-Src, and phospholipase C. In addition, the localization of β(1)-integrin in advanced human carotid plaques is highlighted, marking its importance in atherosclerotic plaque progression.

Percutaneous aortic valvuloplasty is an effective means of treatment for congenital aortic valve stenosis. The aim of this study was to evaluate the immediate results of valvuloplasty, to analyze the medium to long-term outcome and to assess its efficacy in preventing or postponing a new percutaneous valvuloplasty or aortic valve surgery.We retrospectively analyzed the reports of 37 patients aged1 month (mean age 6.3 years) who underwent aortic valvuloplasty for severe aortic stenosis. Associated congenital cardiac defects were present in 16% of the patients. The average time of follow-up was 5.07 years. Particular attention was focused on occurrence and progression of aortic regurgitation.Hemodynamic gradient after aortic valvuloplasty decreased from 58.5 to 22.5 mmHg, with an average decrease of 61.5%. On echography, the maximum gradient decreased from 93.0 to 40.5 mmHg, with an average decrease of 56.5%; mean gradient decreased from 52.0 to 20.5 mmHg with an average decrease of 60.6%. At last follow-up the average maximum and mean gradient on echo were 50.0 and 27.0 mmHg. A reintervention was needed in 21.6% of cases: a second valvuloplasty in 8.1% and aortic surgery in 13.5%. The mortality rate was 2.7%. Survival after 14 years was 97.2%; freedom from aortic valve surgery was 85.5%, from a second valvuloplasty was 89.5%, and from any type of procedure was 76.1%.Percutaneous aortic valvuloplasty is a safe and effective treatment for congenital aortic stenosis in patients aged1 month. Aortic regurgitation is the main concern in the follow-up. Nonetheless, 14 years after valvuloplasty, over 75% of patients are free from any type of aortic valve reintervention.

Chronic inflammatory events appear to play a fundamental role in Alzheimer's disease (AD)-related neuropathological changes, and to result in neuronal dysfunction and death. The inflammatory responses observed in the AD brain include activation and proliferation of glial cells, together with up-regulation of inflammatory mediators and of free radicals. Along with glial cells, neurons themselves can also react and contribute to neuroinflammatory changes in the AD brain, by serving as sources of inflammatory mediators. Because excess cholesterol cannot be degraded in the brain, it must be excreted from that organ as cholesterol oxidation products (oxysterols), in order to prevent its accumulation. Among risk factors for this neurodegenerative disease, a mechanistic link between altered cholesterol metabolism and AD has been suggested; oxysterols appear to be the missing linkers between the two, because of their neurotoxic effects. This study shows that 24-hydroxycholesterol, 27-hydroxycholesterol, and 7β-hydroxycholesterol, the three oxysterols potentially implicated in AD pathogenesis, induce some pro-inflammatory mediator expression in human neuroblastoma SH-SY5Y cells, via Toll-like receptor-4/cyclooxygenase-2/membrane bound prostaglandin E synthase (TLR4/COX-2/mPGES-1); this clearly indicates that oxysterols may promote neuroinflammatory changes in AD. To confirm this evidence, cells were incubated with the anti-inflammatory flavonoid quercetin; remarkably, its anti-inflammatory effects in SH-SY5Y cells were enhanced when it was loaded into β-cyclodextrin-dodecylcarbonate nanoparticles, versus cells pretreated with free quercetin. The goal of loading quercetin into nanoparticles was to improve its permeation across the blood-brain barrier into the brain, and its bioavailability to reach target cells. The findings show that this drug delivery system might be a new therapeutic strategy for preventing or reducing AD progression.