Effects of carotenoids on mitochondrial dysfunction.

Oxidative stress, an imbalance between pro-oxidant and antioxidant status, favouring the pro-oxidant state is a result of increased production of reactive oxygen species (ROS) or inadequate antioxidant protection. ROS are produced through several mechanisms in cells including during mitochondrial oxidative phosphorylation. Increased mitochondrial-derived ROS are associated with mitochondrial dysfunction, an early event in age-related diseases such as Alzheimer's diseases (ADs) and in metabolic disorders including diabetes. AD post-mortem investigations of affected brain regions have shown the accumulation of oxidative damage to macromolecules, and oxidative stress has been considered an important contributor to disease pathology. An increase in oxidative stress, which leads to increased levels of superoxide, hydrogen peroxide and other ROS in a potentially vicious cycle is both causative and a consequence of mitochondrial dysfunction. Mitochondrial dysfunction may be ameliorated by molecules with antioxidant capacities that accumulate in mitochondria such as carotenoids. However, the role of carotenoids in mitigating mitochondrial dysfunction is not fully understood. A better understanding of the role of antioxidants in mitochondrial function is a promising lead towards the development of novel and effective treatment strategies for age-related diseases. This review evaluates and summarises some of the latest developments and insights into the effects of carotenoids on mitochondrial dysfunction with a focus on the antioxidant properties of carotenoids. The mitochondria-protective role of carotenoids may be key in therapeutic strategies and targeting the mitochondria ROS is emerging in drug development for age-related diseases.

Oxidative stress links periodontal inflammation and renal function.

AIMS: Patients with chronic kidney disease (CKD) are also susceptible to periodontitis. The causal link between periodontitis and CKD may be mediated via systemic inflammation/oxidative stress. Using structural equation modelling (SEM), this cross-sectional study aimed to explore the causal relationship between periodontal inflammation (PI) and renal function. MATERIALS AND METHODS: Baseline data on 770 patients with stage 3-5 (pre-dialysis) CKD from an ongoing cohort study were used. Detailed, bioclinical data on PI and renal function, as well as potential confounders and mediators of the relationship between the two, were collected. SEMs of increasing complexity were created to test the causal assumption that PI affects renal function and vice versa. RESULTS: Structural equation modelling confirmed the assumption that PI and renal function are causally linked, mediated by systemic oxidative stress. The magnitude of this effect was such that a 10% increase in PI resulted in a 3.0% decrease in renal function and a 10% decrease in renal function resulted in a 25% increase in PI. CONCLUSIONS: Periodontal inflammation represents an occult source of oxidative stress in patients with CKD. Further clinical studies are needed to confirm whether periodontal therapy, as a non-pharmacological approach to reducing systemic inflammatory/oxidative stress burden, can improve outcomes in CKD.

Paternal diet programs offspring health through sperm- and seminal plasma-specific pathways in mice.

The association between poor paternal diet, perturbed embryonic development, and adult offspring ill health represents a new focus for the Developmental Origins of Health and Disease hypothesis. However, our understanding of the underlying mechanisms remains ill-defined. We have developed a mouse paternal low-protein diet (LPD) model to determine its impact on semen quality, maternal uterine physiology, and adult offspring health. We observed that sperm from LPD-fed male mice displayed global hypomethylation associated with reduced testicular expression of DNA methylation and folate-cycle regulators compared with normal protein diet (NPD) fed males. Furthermore, females mated with LPD males display blunted preimplantation uterine immunological, cell signaling, and vascular remodeling responses compared to controls. These data indicate paternal diet impacts on offspring health through both sperm genomic (epigenetic) and seminal plasma (maternal uterine environment) mechanisms. Extending our model, we defined sperm- and seminal plasma-specific effects on offspring health by combining artificial insemination with vasectomized male mating of dietary-manipulated males. All offspring derived from LPD sperm and/or seminal plasma became heavier with increased adiposity, glucose intolerance, perturbed hepatic gene expression symptomatic of nonalcoholic fatty liver disease, and altered gut bacterial profiles. These data provide insight into programming mechanisms linking poor paternal diet with semen quality and offspring health.

Hypercholesterolaemia-induced oxidative stress at the blood-brain barrier.

Blood cholesterol levels are not consistently elevated in subjects with age-related cognitive decline, although epidemiological studies suggest that Alzheimer's disease and cardiovascular diseases share common risk factors. These include the presence of an unusual genetic variant, the APOE4 (apolipoprotein E4) allele, which modulates LDL (low-density lipoproteins) metabolism, increases free radical formation and reduces plasma antioxidant concentrations. Together, these risk factors support a mechanism for increased LDL circulation time and free radical modification of LDL. Plasma oxycholesterols, hydroxylated metabolites of cholesterol, are carried by oxidized LDL, and elevated lipids in mid-life are associated with increased long-term risk of dementia. Although brain cholesterol metabolism is segregated from the systemic circulation, during oxidative stress, plasma oxycholesterols could have damaging effects on BBB (blood-brain barrier) function and consequently on neuronal cells. Cholesterol-lowering drugs such as statins may prevent the modifications to LDL in mid-life and might show beneficial effects in later life.

Oxysterol sulfates in fluids, cells and tissues: how much do we know about their clinical significance, biological relevance and biophysical implications?

Oxysterol sulfates are emerging as key players in lipid homeostasis, inflammation and immunity. Despite this, knowledge on their basal levels in fluids, cells and tissues and any changes associated with age, gender and diet in health and disease; as well as their spatio-temporal distribution in cell membranes and organelles have been greatly hampered by the lack of commercially available pure synthetic standards. Expansion of the panel of pure oxysterol sulfates standards is pivotal to improve our understanding on the impact of oxysterol sulfates at the membrane level and their role in cellular events. While the clinical significance, biophysical implications and biological relevance of oxysterol sulfates in fluids, cells and tissues remains largely unknown, knowledge already gathered on the precursors of oxysterol sulfates (e.g. oxysterols and cholesterol sulfate) can be used to guide researchers on the most relevant aspects to search for when screening for oxysterol sulfates bioavailability in (patho)physiological conditions which are crucial in the design of biophysical and of cell-based assays. Herein, we provide a review on the brief knowledge involving oxysterol sulfate and an overview on the biophysical implications and biological relevance of oxysterols and cholesterol sulfate useful to redirect further investigations on the role of oxysterol sulfates in health and disease.

Understanding exogenous factors and biological mechanisms for cognitive frailty: A multidisciplinary scoping review.

Cognitive frailty (CF) is the conjunction of cognitive impairment without dementia and physical frailty. While predictors of each element are well-researched, mechanisms of their co-occurrence have not been integrated, particularly in terms of relationships between social, psychological, and biological factors. This interdisciplinary scoping review set out to categorise a heterogenous multidisciplinary literature to identify potential pathways and mechanisms of CF, and research gaps. Studies were included if they used the definition of CF OR focused on conjunction of cognitive impairment and frailty (by any measure), AND excluded studies on specific disease populations, interventions, epidemiology or prediction of mortality. Searches used Web of Science, PubMed and Science Direct. Search terms included "cognitive frailty" OR (("cognitive decline" OR "cognitive impairment") AND (frail*)), with terms to elicit mechanisms, predictors, causes, pathways and risk factors. To ensure inclusion of animal and cell models, keywords such as "behavioural" or "cognitive decline" or "senescence", were added. 206 papers were included. Descriptive analysis provided high-level categorisation of determinants from social and environmental through psychological to biological. Patterns distinguishing CF from Alzheimer's disease were identified and social and psychological moderators and mediators of underlying biological and physiological changes and of trajectories of CF development were suggested as foci for further research.

Micronutrients, Frailty, and Cognitive Impairment: Design and Preliminary Results from the CogLife 2.0 Study.

Background: Among preventive strategies against dementia, nutrition is considered a powerful one and the recently established "nutritional cognitive neuroscience of aging" is a highly active research field. Objective: The present study was designed to deeply characterize older adults across the continuum from cognitive integrity to mild cognitive impairment (MCI) and better elucidate the prognostic role of lipophilic micronutrients within their lipidomic signature. Methods: 123 participants older than 65 years across the continuum from cognitive integrity to MCI were included [49 with subjective cognitive impairment, 29 women, 72.5±5.4 years, 26 MCI, 9 women, 74.5±5.8 years and 50 without cognitive impairment, 21 women, 70.8±4.3 years]. All participants underwent neuropsychological and nutritional examination as well as comprehensive geriatric assessment with calculation of the Multidimensional Prognostic Index (MPI) as a proxy of frailty and biological age and blood withdrawal for the analyses of lipophilic micronutrients, metabolomics and oxylipidomics. One year after the evaluation, same tests are ongoing. Results: After adjustment for age, sex, daily fruit and vegetable intake and cholesterol, we found a significant positive correlation between lutein and the number of correct words in category fluency (p = 0.016). Conclusions: This result supports the importance of carotenoids as robust biomarkers of cognitive performance independent of the nutritional status and frailty of the participants, as the entire present study collective was robust (MPI 0-0.33). The complete analyses of the metabolome and the oxylipidome will hopefully shed light on the metabolic and prognostic signature of cognitive decline in the rapidly growing population at risk of frailty.

Cholesterol homeostasis in hair follicle keratinocytes is disrupted by impaired ABCA5 activity.

The importance of cholesterol in hair follicle biology is underscored by its links to the pathogenesis of alopecias and hair growth disorders. Reports have associated defects in ABCA5, a membrane transporter, with altered keratinocyte cholesterol distribution in individuals with a form of congenital hypertrichosis, yet the biological basis for this defect in hair growth remains unknown. This study aimed to determine the impact of altered ABCA5 activity on hair follicle keratinocyte behaviour. Primary keratinocytes isolated from the outer root sheath of plucked human hair follicles were utilised as a relevant cell model. Following exogenous cholesterol loading, an increase in ABCA5 co-localisation to intracellular organelles was seen. Knockdown of ABCA5 revealed a dysregulation in cholesterol homeostasis, with LXR agonism leading to partial restoration of the homeostatic response. Filipin staining and live BODIPY cholesterol immunofluorescence microscopy revealed a reduction in endo-lysosomal cholesterol following ABCA5 knockdown. Analysis of oxysterols showed a significant increase in the fold change of 25-hydroxycholesterol and 7-ß-hydroxycholesterol following cholesterol loading in ORS keratinocytes, after ABCA5 knockdown. These data suggest a role for ABCA5 in the intracellular compartmentalisation of free cholesterol in primary hair follicle keratinocytes. The loss of normal homeostatic response, following the delivery of excess cholesterol after ABCA5 knockdown, suggests an impact on LXR-mediated transcriptional activity. The loss of ABCA5 in the hair follicle could lead to impaired endo-lysosomal cholesterol transport, impacting pathways known to influence hair growth. This avenue warrants further investigation.

Type 2 Diabetes mellitus alters the cargo of (poly)phenol metabolome and the oxidative status in circulating lipoproteins.

The incidence of diabetes on the worldwide population has tripled in the past 5 decades. While drug-based therapies are valuable strategies to treat and ease the socio-economic burden of diabetes, nutritional strategies offer valuable alternatives to prevent and manage diabetes onset and contribute to the sustainability of health budgets. Whilst, intervention studies have shown that (poly)phenol-rich diets improve fasting glucose levels and other blood parameters, very little is known about the distribution of ingested polyphenols in circulation and the impact of diabetes on its cargo. In this study we investigate the impact of type 2 diabetes on the cargo of plasma (poly)phenols. Our results show that phenolic compounds are heterogeneously distributed in circulation though mainly transported by lipoprotein populations. We also found that diabetes has a marked effect on the phenolic content transported by VLDL resulting in the decrease in the content of flavonoids and consequently a decrease in the antioxidant capacity. In addition to the reduced bioavailability of (poly)phenol metabolites and increase of oxidative status in LDL and HDL populations in diabetes, cell-based assays show that sub-micromolar amounts of microbial (poly)phenol metabolites are able to counteract the pro-inflammatory status in glucose-challenged endothelial cells. Our findings highlight the relevance of triglyceride-rich lipoproteins in the transport and delivery of bioactive plant-based compounds to the endothelium in T2DM supporting the adoption of nutritional guidelines as an alternative strategy to drug-based therapeutic approaches.

Antioxidant Micronutrients and Oxidative Stress Biomarkers.

Chronic inflammatory diseases are the major causes of mortality in humans and recent research has improved our understanding of the major impact of lifestyle factors upon inflammatory diseases and conditions. One of the most influential of these is nutrition, which may drive both pro-inflammatory as well as anti-inflammatory cascades at molecular and cellular levels. There are a variety of model systems that may be employed to investigate the impact of micronutrients and macronutrients upon inflammatory pathways, many of which operate through oxidative stress, either at the level of controlling the redox state of the cell and downstream redox-regulated gene transcription factors, and other acting as free radical generating or scavenging agents. This chapter focuses upon biological sample preparation prior to assay and details methods for analyzing certain antioxidant micronutrients and biomarkers of oxidative stress.

TNF-α-Mediated Endothelial Cell Apoptosis Is Rescued by Hydrogen Sulfide.

Endothelial dysfunction is implicated in the development and aggravation of cardiovascular complications. Among the endothelium-released vasoactive factors, hydrogen sulfide (H2S) has been investigated for its beneficial effects on the vasculature through anti-inflammatory and redox-modulating regulatory mechanisms. Reduced H2S bioavailability is reported in chronic diseases such as cardiovascular disease, diabetes, atherosclerosis and preeclampsia, suggesting the value of investigating mechanisms, by which H2S acts as a vasoprotective gasotransmitter. We explored whether the protective effects of H2S were linked to the mitochondrial health of endothelial cells and the mechanisms by which H2S rescues apoptosis. Here, we demonstrate that endothelial dysfunction induced by TNF-α increased endothelial oxidative stress and induced apoptosis via mitochondrial cytochrome c release and caspase activation over 24 h. TNF-α also affected mitochondrial morphology and altered the mitochondrial network. Post-treatment with the slow-releasing H2S donor, GYY4137, alleviated oxidising redox state, decreased pro-caspase 3 activity, and prevented endothelial apoptosis caused by TNF-α alone. In addition, exogenous GYY4137 enhanced S-sulfhydration of pro-caspase 3 and improved mitochondrial health in TNF-α exposed cells. These data provide new insights into molecular mechanisms for cytoprotective effects of H2S via the mitochondrial-driven pathway.

Identification and quantification of ionising radiation-induced oxysterol formation in membranes of lens fibre cells.

Ionising radiation (IR) is a cause of lipid peroxidation, and epidemiological data have revealed a correlation between exposure to IR and the development of eye lens cataracts. Cataracts remain the leading cause of blindness around the world. The plasma membranes of lens fibre cells are one of the most cholesterolrich membranes in the human body, forming lipid rafts and contributing to the biophysical properties of lens fibre plasma membrane. Liquid chromatography followed by mass spectrometry was used to analyse bovine eye lens lipid membrane fractions after exposure to 5 and 50 Gy and eye lenses taken from wholebody 2 Gy-irradiated mice. Although cholesterol levels do not change significantly, IR dose-dependant formation of the oxysterols 7ß-hydroxycholesterol, 7-ketocholesterol and 5, 6-epoxycholesterol in bovine lens nucleus membrane extracts was observed. Whole-body X-ray exposure (2 Gy) of 12-week old mice resulted in an increase in 7ß-hydroxycholesterol and 7-ketocholesterol in their eye lenses. Their increase regressed over 24 h in the living lens cortex after IR exposure. This study also demonstrated that the IR-induced fold increase in oxysterols was greater in the mouse lens cortex than the nucleus. Further work is required to elucidate the mechanistic link(s) between oxysterols and IR-induced cataract, but these data evidence for the first time that IR exposure of mice results in oxysterol formation in their eye lenses.

Tspan6 stimulates the chemoattractive potential of breast cancer cells for B cells in an EV- and LXR-dependent manner.

The immune microenvironment in breast cancer (BCa) is controlled by a complex network of communication between various cell types. Here, we find that recruitment of B lymphocytes to BCa tissues is controlled via mechanisms associated with cancer cell-derived extracellular vesicles (CCD-EVs). Gene expression profiling identifies the Liver X receptor (LXR)-dependent transcriptional network as a key pathway that controls both CCD-EVs-induced migration of B cells and accumulation of B cells in BCa tissues. The increased accumulation oxysterol ligands for LXR (i.e., 25-hydroxycholesterol and 27-hydroxycholesterol) in CCD-EVs is regulated by the tetraspanin 6 (Tspan6). Tspan6 stimulates the chemoattractive potential of BCa cells for B cells in an EV- and LXR-dependent manner. These results demonstrate that tetraspanins control intercellular trafficking of oxysterols via CCD-EVs. Furthermore, tetraspanin-dependent changes in the oxysterol composition of CCD-EVs and the LXR signaling axis play a key role in specific changes in the tumor immune microenvironment.

Oxysterols and Oxysterol Sulfates in Alzheimer's Disease Brain and Cerebrospinal Fluid.

BACKGROUND: Brain cholesterol levels are tightly regulated but increasing evidence indicates that cholesterol metabolism may drive Alzheimer's disease (AD)-associated pathological changes. Recent advances in understanding of mitochondrial dysfunction in AD brain have presented a vital role played by mitochondria in oxysterol biosynthesis and their involvement in pathophysiology. Oxysterol accumulation in brain is controlled by various enzymatic pathways including sulfation. While research into oxysterol is under the areas of active investigation, there is less evidence for oxysterol sulfate levels in human brain. OBJECTIVE: This study investigates the hypothesis that AD brain oxysterol detoxification via sulfation is impaired in later stages of disease resulting in oxysterol accumulation. METHODS: Lipids were extracted from postmortem frozen brain tissue and cerebrospinal (CSF) from late- (Braak stage III-IV) and early- (Braak stage I-II) stage AD patients. Samples were spiked with internal standards prior to lipid extraction. Oxysterols were enriched with a two-step solid phase extraction using a polymeric SPE column and further separation was achieved by LC-MS/MS. RESULTS: Oxysterols, 26-hydroxycholesterol (26-OHC), 25-hydroxycholesterol (25-OHC), and 7-oxycholesterol levels were higher in brain tissue and mitochondria extracted from late-stage AD brain tissue except for 24S-hydroxycholesterol, which was decreased in late AD. However, oxysterol sulfates are significantly lower in the AD frontal cortex. Oxysterols, 25-OHC, and 7-oxocholesterol was higher is CSF but 26-OHC and oxysterol sulfate levels were not changed. CONCLUSION: Our results show oxysterol metabolism is altered in AD brain mitochondria, favoring synthesis of 26-OHC, 25-OHC, and 7-oxocholesterol, and this may influence brain mitochondrial function and acceleration of the disease.

The Use of Retinal Microvascular Function and Telomere Length in Age and Blood Pressure Prediction in Individuals with Low Cardiovascular Risk.

Ageing represents a major risk factor for many pathologies that limit human lifespan, including cardiovascular diseases. Biological ageing is a good biomarker to assess early individual risk for CVD. However, finding good measurements of biological ageing is an ongoing quest. This study aims to assess the use retinal microvascular function, separate or in combination with telomere length, as a predictor for age and systemic blood pressure in individuals with low cardiovascular risk. In all, 123 healthy participants with low cardiovascular risk were recruited and divided into three groups: group 1 (less than 30 years old), group 2 (31-50 years old) and group 3 (over 50 years old). Relative telomere length (RTL), parameters of retinal microvascular function, CVD circulatory markers and blood pressure (BP) were measured in all individuals. Symbolic regression- analysis was used to infer chronological age and systemic BP measurements using either RTL or a combination of RTL and parameters for retinal microvascular function. RTL decreased significantly with age (p = 0.010). There were also age-related differences between the study groups in retinal arterial time to maximum dilation (p = 0.005), maximum constriction (p = 0.007) and maximum constriction percentage (p = 0.010). In the youngest participants, the error between predicted versus actual values for the chronological age were smallest in the case of using both retinal vascular functions only (p = 0.039) or the combination of this parameter with RTL (p = 0.0045). Systolic BP was better predicted by RTL also only in younger individuals (p = 0.043). The assessment of retinal arterial vascular function is a better predictor than RTL for non-modifiable variables such as age, and only in younger individuals. In the same age group, RTL is better than microvascular function when inferring modifiable risk factors for CVDs. In older individuals, the accumulation of physiological and structural biological changes makes such predictions unreliable.

Paternal low protein diet perturbs inter-generational metabolic homeostasis in a tissue-specific manner in mice.

The underlying mechanisms driving paternally-programmed metabolic disease in offspring remain poorly defined. We fed male C57BL/6 mice either a control normal protein diet (NPD; 18% protein) or an isocaloric low protein diet (LPD; 9% protein) for a minimum of 8 weeks. Using artificial insemination, in combination with vasectomised male mating, we generated offspring using either NPD or LPD sperm but in the presence of NPD or LPD seminal plasma. Offspring from either LPD sperm or seminal fluid display elevated body weight and tissue dyslipidaemia from just 3 weeks of age. These changes become more pronounced in adulthood, occurring in conjunction with altered hepatic metabolic and inflammatory pathway gene expression. Second generation offspring also display differential tissue lipid abundance, with profiles similar to those of first generation adults. These findings demonstrate that offspring metabolic homeostasis is perturbed in response to a suboptimal paternal diet with the effects still evident within a second generation.

Activation of the neutrophil respiratory burst by plasma from periodontitis patients is mediated by pro-inflammatory cytokines.

AIM: To determine the effect of periodontitis patients' plasma on the neutrophil oxidative burst and the role of albumin, immunoglobulins (Igs) and cytokines. MATERIALS AND METHODS: Plasma was collected from chronic periodontitis patients (n=11) and periodontally healthy controls (n=11) and used with/without depletion of albumin and Ig or antibody neutralization of IL-8, GM-CSF or IFN-α to prime/stimulate peripheral blood neutrophils, isolated from healthy volunteers. The respiratory burst was measured by lucigenin-dependent chemiluminescence. Plasma cytokine levels were determined by ELISA. RESULTS: Plasmas from patients were significantly more effective in both directly stimulating neutrophil superoxide production and priming for subsequent formyl-met-leu-phe (fMLP)-stimulated superoxide production than plasmas from healthy controls (p<0.05). This difference was maintained after depletion of albumin and Ig. Plasma from patients contained higher mean levels of IL-8, GM-CSF and IFN-α. Individual neutralizing antibodies against IL-8, GM-CSF or IFN-α inhibited the direct stimulatory effect of patients' plasma, whereas the ability to prime for fMLP-stimulated superoxide production was only inhibited by neutralization of IFN-α. The stimulating and priming effects of control plasma were unaffected by antibody neutralization. CONCLUSIONS: This study demonstrates that plasma cytokines may have a role in inducing the hyperactive (IL-8, GM-CSF, IFN-α) and hyper-reactive (IFN-α) neutrophil phenotype seen in periodontitis patients.

Current and Future Directions for Targeting Lipoxin A4 in Alzheimer's Disease.

Neuroinflammation has been implicated in Alzheimer's disease onset and progression. Chronic neuroinflammation is initiated by amyloid-ß-activated microglial cells that secrete immuno-modulatory molecules within the brain and into the vasculature. Inflammation is normally self-limiting and actively resolves by "switching off" the generation of pro-inflammatory mediators and by non-phlogistic clearance of spent cells and their debris to restore tissue homeostasis. Deficits in these anti-inflammatory/pro-resolution pathways may predispose to the development of chronic inflammation. The synthesis of endogenous lipid mediators from arachidonic acid, lipoxins via cyclooxygenase 2 and lipoxygenases, and conversion of exogenous polyunsaturated fatty acids, namely docosahexaenoic acid and eicosapentaenoic acid, to resolvins contributes to effective, timely resolution of acute inflammation. Work by Xiuzhe et al., 2020 in the Journal of Alzheimer's Disease reported that plasma level of LXA4 is related to cognitive status in ischemic stroke patients suggesting that decreased LXA4 may be a potential risk factor for post post-stroke cognitive impairment. As evident by recent clinical trials and development of drug analogues, there is recent drive to search for lipoxin analogues as therapeutics for inflammatory diseases. Understanding how bioactive lipid signaling is involved in resolution will increase our understanding of controlling inflammation and may facilitate the discovery of new classes of therapeutic pro-resolution agents for evaluation in AD prevention studies.

Oxysterols and Retinal Microvascular Dysfunction as Early Risk Markers for Cardiovascular Disease in Normal, Ageing Individuals.

The aim of the present paper is to assess the relationship between oxysterol levels and retinal microvascular function in individuals of various age groups, free of clinically evident diseases. Forty-two apparently healthy individuals were included in the present study (group 1: 19-30 years, group 2: 31-50 years, and group 3: 51-70 years). Retinal microvascular function was assessed using the dynamic retinal vessel analyzer (DVA, IMEDOS GmbH, Jena, Germany). Fasting plasma was obtained from all subjects and quantification of monohydroxy and dihydroxy oxysterols assessment was performed using LC-MS/MS following reverse phase chromatography. A Griess assay was used to evaluate the Nitric Oxide (NO) concentration in all individuals. The glutathione redox ratio was also analyzed by means of whole blood glutathione recycling assay. In all participants, the levels of 7-Ketocholesterol, 25-hydroxycholesterol and 7ß-hydroxycholesterol correlated significantly and positively with the time to maximum arteriolar dilation. In addition, 25-hydroxycholesterol and 7ß-hydroxycholesterol negatively correlated to the percentage of maximum arteriolar dilation. A negative correlation was observed for 27-hydroxycholesterol and 7ß-hydroxycholesterol with microvascular arteriolar constriction. These results suggest that, with age, abnormal oxysterol levels correlate with early changes in microvascular bed function. This relationship could signal early risk for cardiovascular diseases (CVDs) in an ageing population.

Cholesterol and oxysterol sulfates: Pathophysiological roles and analytical challenges.

Cholesterol and oxysterol sulfates are important regulators of lipid metabolism, inflammation, cell apoptosis, and cell survival. Among the sulfate-based lipids, cholesterol sulfate (CS) is the most studied lipid both quantitatively and functionally. Despite the importance, very few studies have analysed and linked the actions of oxysterol sulfates to their physiological and pathophysiological roles. Overexpression of sulfotransferases confirmed the formation of a range of oxysterol sulfates and their antagonistic effects on liver X receptors (LXRs) prompting further investigations how are the changes to oxysterol/oxysterol sulfate homeostasis can contribute to LXR activity in the physiological milieu. Here, we aim to bring together for novel roles of oxysterol sulfates, the available techniques and the challenges associated with their analysis. Understanding the oxysterol/oxysterol sulfate levels and their pathophysiological mechanisms could lead to new therapeutic targets for metabolic diseases. LINKED ARTICLES: This article is part of a themed issue on Oxysterols, Lifelong Health and Therapeutics. To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v178.16/issuetoc.