Trajectory and Predictors of Mental Health Symptoms and Wellbeing in Newly Admitted Nursing Home Residents.

OBJECTIVES: This study determined changes in multiple aspects of mental health and wellbeing in newly admitted nursing home residents, and identified risk and protective factors. METHODS: Participants were 204 residents recently admitted to one of 42 nursing homes in Melbourne, Australia. A subgroup of 82 participants were followed up eight months post-admission. Depression, anxiety, stress, adjustment, and quality of life were assessed at baseline and follow-up. Predictive factors (demographics, health, transition factors, nursing home characteristics) were examined in multiple regression analyses. RESULTS: Rates of depression and anxiety were high at both baseline and follow-up. Low self-rated health and medical comorbidity predicted poor wellbeing at baseline. Higher perceived control in the relocation to the nursing home and engagement in meaningful activities were associated with better post-admission outcomes. Baseline psychotropic medication use predicted lower anxiety at follow-up but did not impact depressive symptoms. CONCLUSIONS: There were no significant changes in mental health and wellbeing from one to eight months post-admission. The negative effect of residing in a for-profit nursing home requires further investigation. CLINICAL IMPLICATIONS: Individual activity scheduling and an opportunity to participate in relocation decision-making and planning may support resident wellbeing post-admission.

Program to Enhance Adjustment to Residential Living (PEARL): Effect on Adjustment, Anxiety, Quality of Life, and Stress.

OBJECTIVES: The Program to Enhance Adjustment to Residential Living (PEARL) is a five session intervention primarily designed to address high rates of depression in newly admitted residents. This study reports the efficacy of PEARL on secondary outcomes of resident adjustment, symptoms of anxiety, quality of life, and stress. METHODS: A cluster randomized controlled trial was conducted with 219 newly admitted nursing home residents (M age = 85.5 years) from 42 nursing homes. Outcomes were assessed at baseline, post-intervention, and at two and six month post-intervention follow-up, compared to a standard care condition. RESULTS: There was a significant overall condition by time interaction for adjustment (p = .027) and quality of life (p = .015), but not for stress (p = .309). While the overall condition by time interaction was not significant for anxiety (p = .221), there was a significant interaction contrast six-month post-intervention, indicating a greater decrease in anxiety scores in the intervention group relative to control (p = .039). CONCLUSIONS: This study demonstrates the broad effects of PEARL on the wellbeing of newly admitted residents. CLINICAL IMPLICATIONS: PEARL is a brief intervention that may be feasible for routine use in nursing homes to facilitate adjustment and improve residents' quality of life.

Gypenosides mediate cholesterol efflux and suppress oxidized LDL induced inflammation in retinal pigment epithelium cells.

Age-related macular degeneration (AMD) is a predominant cause of visual deficit in aged population. Abnormal accumulation of cholesterol, including oxidized low-density lipoprotein (oxLDL), underneath the retinal pigment epithelium (RPE) cells contributes to the development of AMD. Gypenosides (Gyp) are glycosides extracted from Gynostemma pentaphyllum and have demonstrated protective effects against inflammation and oxidative stress. To determine the therapeutic potential of Gyp for AMD, we investigated its effect on cholesterol trafficking and metabolism and assessed the protective function of Gyp against oxLDL-induced damage in RPE cells. Cholesterol efflux to high-density lipoprotein (HDL) and human serum was significantly increased in RPE cells treated with Gyp when compared to untreated control cells. Expression of cholesterol metabolism (CYP27A1, CYP46A1) and trafficking (TSPO, ABCA1 and ABCG1) genes was also markedly increased in Gyp-treated RPE cells. OxLDL-treated RPE cells had significantly increased cholesterol accumulation and lipid droplet formation. There were marked increases in reactive oxygen species (ROS) generation and proinflammatory cytokines via NF-κB activation in RPE cells treated with oxLDL, while incubation with Gyp rectified these changes. These findings provide pharmacological evidence that Gyp has the potential to treat patients with early onset AMD by promoting cellular cholesterol removal from RPE cells and inhibiting inflammation and oxidative stress.

A feasibility and acceptability study of an adaptation of the Mindful Self-Compassion program for adult cancer patients.

OBJECTIVES: Psychosocial interventions that mitigate psychosocial distress in cancer patients are important. The primary aim of this study was to examine the feasibility and acceptability of an adaptation of the Mindful Self-Compassion (MSC) program among adult cancer patients. A secondary aim was to examine pre-post-program changes in psychosocial wellbeing. METHOD: The research design was a feasibility and acceptability study, with an examination of pre- to post-intervention changes in psychosocial measures. A study information pack was posted to 173 adult cancer patients 6 months-5 years post-diagnosis, with an invitation to attend an eight-week group-based adaptation of the MSC program. RESULTS: Thirty-two (19%) consented to the program, with 30 commencing. Twenty-seven completed the program (mean age: 62.93 years, SD 14.04; 17 [63%] female), attending a mean 6.93 (SD 1.11) group sessions. There were no significant differences in medico-demographic factors between program-completers and those who did not consent. However, there was a trend toward shorter time since diagnosis in the program-completers group. Program-completers rated the program highly regarding content, relevance to the concerns of cancer patients, and the likelihood of recommending the program to other cancer patients. Sixty-three percent perceived that their mental wellbeing had improved from pre- to post-program; none perceived a deterioration in mental wellbeing. Small-to-medium effects were observed for depressive symptoms, fear of cancer recurrence, stress, loneliness, body image satisfaction, mindfulness, and self-compassion. SIGNIFICANCE OF RESULTS: The MSC program appears feasible and acceptable to adults diagnosed with non-advanced cancer. The preliminary estimates of effect sizes in this sample suggest that participation in the program was associated with improvements in psychosocial wellbeing. Collectively, these findings suggest that there may be value in conducting an adequately powered randomized controlled trial to determine the efficacy of the MSC program in enhancing the psychosocial wellbeing of cancer patients.

Retinal pigment epithelium cholesterol efflux mediated by the 18 kDa translocator protein, TSPO, a potential target for treating age-related macular degeneration.

Cholesterol accumulation beneath the retinal pigment epithelium (RPE) cells is supposed to contribute the pathogenesis of age-related macular degeneration (AMD). Cholesterol efflux genes (APOE and ABCA1) were identified as risk factors for AMD, although how cholesterol efflux influences accumulation of this lipid in sub-RPE deposits remains elusive. The 18 kDa translocator protein, TSPO, is a cholesterol-binding protein implicated in mitochondrial cholesterol transport. Here, we investigate the function of TSPO in cholesterol efflux from the RPE cells. We demonstrate in RPE cells that TSPO specific ligands promoted cholesterol efflux to acceptor (apo)lipoprotein and human serum, while loss of TSPO resulted in impaired cholesterol efflux. TSPO-/- RPE cells also had significantly increased production of reactive oxygen species (ROS) and upregulated expression of proinflammatory cytokines (IL-1ß and TNFα). Cholesterol (oxidized LDL) uptake and accumulation were markedly increased in TSPO-/- RPE cells. Finally, in aged RPE cells, TSPO expression was reduced and cholesterol efflux impaired. These findings provide a new pharmacological concept to treat early AMD patients by stimulating cellular cholesterol removal with TSPO specific ligands or by overexpression of TSPO in RPE cells.

Receipt and Perceived Helpfulness of Mental Illness Information: Findings from the Australian National Survey of Mental Health and Wellbeing.

The distribution of mental illness information is a crucial element of mental health promotion initiatives. We assessed the receipt and perceived helpfulness of such information in Australia. Data from the Australian National Survey of Mental Health and Wellbeing indicated that, during the year prior to the survey, 33.7% of Australians received mental illness information; of these, 51.2% found it helpful. Among people with a mental disorder, 46.1% received information; of these, 67.4% found it helpful. Non-English speakers and the socially disadvantaged were less likely to receive mental illness information. Older and less educated respondents were less likely to both receive mental illness information and find it helpful. Mental health service users were more likely to receive mental illness information perceived as helpful than those who had not accessed such services. Better targeted information interventions are required to ensure those most likely to benefit receive mental illness-related information.

The potential of human induced pluripotent stem cells for modelling diabetic wound healing in vitro.

Impaired wound healing and ulceration caused by diabetes mellitus, is a significant healthcare burden, markedly impairs quality of life for patients, and is the major cause of amputation worldwide. Current experimental approaches used to investigate the complex wound healing process often involve cultures of fibroblasts and/or keratinocytes in vitro, which can be limited in terms of complexity and capacity, or utilisation of rodent models in which the mechanisms of wound repair differ substantively from that in humans. However, advances in tissue engineering, and the discovery of strategies to reprogramme adult somatic cells to pluripotency, has led to the possibility of developing models of human skin on a large scale. Generation of induced pluripotent stem cells (iPSCs) from tissues donated by diabetic patients allows the (epi)genetic background of this disease to be studied, and the ability to differentiate iPSCs to multiple cell types found within skin may facilitate the development of more complex skin models; these advances offer key opportunities for improving modelling of wound healing in diabetes, and the development of effective therapeutics for treatment of chronic wounds.

The Connexin Mimetic Peptide Gap27 and Cx43-Knockdown Reveal Differential Roles for Connexin43 in Wound Closure Events in Skin Model Systems.

In the epidermis, remodelling of Connexin43 is a key event in wound closure. However, controversy between the role of connexin channel and non-channel functions exist. We compared the impact of SiRNA targeted to Connexin43 and the connexin mimetic peptide Gap27 on scrape wound closure rates and hemichannel signalling in adult keratinocytes (AK) and fibroblasts sourced from juvenile foreskin (JFF), human neonatal fibroblasts (HNDF) and adult dermal tissue (ADF). The impact of these agents, following 24 h exposure, on GJA1 (encoding Connexin43), Ki67 and TGF-ß1 gene expression, and Connexin43 and pSmad3 protein expression levels, were examined by qPCR and Western Blot respectively. In all cell types Gap27 (100-100 µM) attenuated hemichannel activity. In AK and JFF cells, Gap27 (100 nM-100 µM) enhanced scrape wound closure rates by ~50% but did not influence movement in HNDF or ADF cells. In both JF and AK cells, exposure to Gap27 for 24 h reduced the level of Cx43 protein expression but did not affect the level in ADF and HNDF cells. Connexin43-SiRNA enhanced scrape wound closure in all the cell types under investigation. In HDNF and ADF, Connexin43-SiRNA enhanced cell proliferation rates, with enhanced proliferation also observed following exposure of HDNF to Gap27. By contrast, in JFF and AK cells no changes in proliferation occurred. In JFF cells, Connexin43-SiRNA enhanced TGF-ß1 levels and in JFF and ADF cells both Connexin43-SiRNA and Gap27 enhanced pSmad3 protein expression levels. We conclude that Connexin43 signalling plays an important role in cell migration in keratinocytes and foreskin derived fibroblasts, however, different pathways are evoked and in dermal derived adult and neonatal fibroblasts, inhibition of Connexin43 signalling plays a more significant role in regulating cell proliferation than cell migration.

Intracellular cholesterol transport proteins: roles in health and disease.

Effective cholesterol homoeostasis is essential in maintaining cellular function, and this is achieved by a network of lipid-responsive nuclear transcription factors, and enzymes, receptors and transporters subject to post-transcriptional and post-translational regulation, whereas loss of these elegant, tightly regulated homoeostatic responses is integral to disease pathologies. Recent data suggest that sterol-binding sensors, exchangers and transporters contribute to regulation of cellular cholesterol homoeostasis and that genetic overexpression or deletion, or mutations, in a number of these proteins are linked with diseases, including atherosclerosis, dyslipidaemia, diabetes, congenital lipoid adrenal hyperplasia, cancer, autosomal dominant hearing loss and male infertility. This review focuses on current evidence exploring the function of members of the 'START' (steroidogenic acute regulatory protein-related lipid transfer) and 'ORP' (oxysterol-binding protein-related proteins) families of sterol-binding proteins in sterol homoeostasis in eukaryotic cells, and the evidence that they represent valid therapeutic targets to alleviate human disease.

Social networks and links to isolation and loneliness among elderly HCBS clients.

OBJECTIVES: The purpose of this study was to explore the network types of HCBS clients based on the structural characteristics of their social networks. We also examined how the network types were associated with social isolation, relationship quality and loneliness. METHOD: Forty personal interviews were carried out with HCBS clients to assess the structure of their social networks as indicated by frequency of contact with children, friends, family and participation in religious and community organizations. Hierarchical cluster analysis was conducted to identify network types. RESULTS: Four network types were found including: family (n = 16), diverse (n = 8), restricted (n = 8) and religious (n = 7). Family members comprised almost half of participants' social networks, and friends comprised less than one-third. Clients embedded in family, diverse and religious networks had significantly more positive relationships than clients embedded in restricted networks. Clients embedded in restricted networks had significantly higher social isolation scores and were lonelier than clients in diverse and family networks. DISCUSSION: The findings suggest that HCBS clients' isolation and loneliness are linked to the types of social networks in which they are embedded. The findings also suggest that clients embedded in restricted networks are at high risk for negative outcomes.

Intracellular cholesterol transporters and modulation of hepatic lipid metabolism: Implications for diabetic dyslipidaemia and steatosis.

AIMS/HYPOTHESES: To examine hepatic expression of cholesterol-trafficking proteins, mitochondrial StarD1 and endosomal StarD3, and their relationship with dyslipidaemia and steatosis in Zucker (fa/fa) genetically obese rats, and to explore their functional role in lipid metabolism in rat McArdle RH-7777 hepatoma cells. METHODS: Expression of StarD1 and StarD3 in rat liver and hepatoma samples were determined by Q-PCR and/or immunoblotting; lipid mass by colorimetric assays; radiolabelled precursors were utilised to measure lipid synthesis and secretion, and lipidation of exogenous apolipoprotein A-I. RESULTS: Hepatic expression of StarD3 protein was repressed by genetic obesity in (fa/fa) Zucker rats, compared with lean (Fa/?) controls, suggesting a link with storage or export of lipids from the liver. Overexpression of StarD1 and StarD3, and knockdown of StarD3, in rat hepatoma cells, revealed differential effects on lipid metabolism. Overexpression of StarD1 increased utilisation of exogenous (preformed) fatty acids for triacylglycerol synthesis and secretion, but impacted minimally on cholesterol homeostasis. By contrast, overexpression of StarD3 increased lipidation of exogenous apoA-I, and facilitated de novo biosynthetic pathways for neutral lipids, potentiating triacylglycerol accumulation but possibly offering protection against lipotoxicity. Finally, StarD3 overexpression altered expression of genes which impact variously on hepatic insulin resistance, inducing Ppargcla, Cyp2e1, Nr1h4, G6pc and Irs1, and repressing expression of Scl2a1, Igfbp1, Casp3 and Serpine 1. CONCLUSIONS/INTERPRETATION: Targeting StarD3 may increase circulating levels of HDL and protect the liver against lipotoxicity; loss of hepatic expression of this protein, induced by genetic obesity, may contribute to the pathogenesis of dyslipidaemia and steatosis.

How People with Depression Receive and Perceive Mental Illness Information: Findings from the Australian National Survey of Mental Health and Wellbeing.

Despite the recognised importance of accurate mental illness information in help-seeking and improving recovery, little is known about the dissemination of such information to people with depression. With a view to informing effective communication to those most in need, we explored the extent to which mental illness information is received by people with depression, its perceived helpfulness and we characterise those who do not receive such information. Using data from the Australian National Survey of Mental Health and Wellbeing we observed that mental illness information was received by 54.7 % of those with depression. Most (76.7 %) found it helpful. Pamphlets were the most frequently cited source of information. People who did not receive information were less educated, unlikely to have accessed mental health services and unlikely to believe they had mental health needs. Targeted information campaigns which shape perceptions of need in relation to depression have the potential to reduce the resultant disease burden.

Targeting mitochondrial 18 kDa translocator protein (TSPO) regulates macrophage cholesterol efflux and lipid phenotype.

The aim of the present study was to establish mitochondrial cholesterol trafficking 18 kDa translocator protein (TSPO) as a potential therapeutic target, capable of increasing macrophage cholesterol efflux to (apo)lipoprotein acceptors. Expression and activity of TSPO in human (THP-1) macrophages were manipulated genetically and by the use of selective TSPO ligands. Cellular responses were analysed by quantitative PCR (Q-PCR), immunoblotting and radiolabelling, including [3H]cholesterol efflux to (apo)lipoprotein A-I (apoA-I), high-density lipoprotein (HDL) and human serum. Induction of macrophage cholesterol deposition by acetylated low-density lipoprotein (AcLDL) increased expression of TSPO mRNA and protein, reflecting findings in human carotid atherosclerosis. Transient overexpression of TSPO enhanced efflux (E%) of [3H]cholesterol to apoA-I, HDL and human serum compared with empty vector (EV) controls, whereas gene knockdown of TSPO achieved the converse. Ligation of TSPO (using PK11195, FGIN-1-27 and flunitrazepam) triggered increases in [3H]cholesterol efflux, an effect that was amplified in TSPO-overexpressing macrophages. Overexpression of TSPO induced the expression of genes [PPARA (peroxisome-proliferator-activated receptor α), NR1H3 (nuclear receptor 1H3/liver X receptor α), ABCA1 (ATP-binding cassette A1), ABCG4 (ATP-binding cassette G4) and APOE (apolipoprotein E)] and proteins (ABCA1 and PPARα) involved in cholesterol efflux, reduced macrophage neutral lipid mass and lipogenesis and limited cholesterol esterification following exposure to AcLDL. Thus, targeting TSPO reduces macrophage lipid content and prevents macrophage foam cell formation, via enhanced cholesterol efflux to (apo)lipoprotein acceptors.

Mindfulness-based cognitive therapy for recurrent depression: A translational research study with 2-year follow-up.

OBJECTIVE: While mindfulness-based cognitive therapy (MBCT) has demonstrated efficacy in reducing depressive relapse/recurrence over 12-18 months, questions remain around effectiveness, longer-term outcomes, and suitability in combination with medication. The aim of this study was to investigate within a pragmatic study design the effectiveness of MBCT on depressive relapse/recurrence over 2 years of follow-up. METHOD: This was a prospective, multi-site, single-blind trial based in Melbourne and the regional city of Geelong, Australia. Non-depressed adults with a history of three or more episodes of depression were randomised to MBCT + depression relapse active monitoring (DRAM) (n=101) or control (DRAM alone) (n=102). Randomisation was stratified by medication (prescribed antidepressants and/or mood stabilisers: yes/no), site of usual care (primary or specialist), diagnosis (bipolar disorder: yes/no) and sex. Relapse/recurrence of major depression was assessed over 2 years using the Composite International Diagnostic Interview 2.1. RESULTS: The average number of days with major depression was 65 for MBCT participants and 112 for controls, significant with repeated-measures ANOVA (F(1, 164)=4.56, p=0.03). Proportionally fewer MBCT participants relapsed in both year 1 and year 2 compared to controls (odds ratio 0.45, p<0.05). Kaplan-Meier survival analysis for time to first depressive episode was non-significant, although trends favouring the MBCT group were suggested. Subgroup analyses supported the effectiveness of MBCT for people receiving usual care in a specialist setting and for people taking antidepressant/mood stabiliser medication. CONCLUSIONS: This work in a pragmatic design with an active control condition supports the effectiveness of MBCT in something closer to implementation in routine practice than has been studied hitherto. As expected in this translational research design, observed effects were less strong than in some previous efficacy studies but appreciable and significant differences in outcome were detected. MBCT is most clearly demonstrated as effective for people receiving specialist care and seems to work well combined with antidepressants.

Mitochondrial (dys)function and regulation of macrophage cholesterol efflux.

Cholesterol trafficking from the outer to the cholesterol-poor inner mitochondrial membrane requires energized, polarized and actively respiring mitochondria, mediated by a highly regulated multimeric (140-200 kDa) protein complex comprising StAR (steroidogenic acute regulatory protein), mitochondrial TSPO (translocator protein), VDAC (voltage-dependent anion channel), ANT (adenine nucleotide transporter) and associated regulatory proteins. Mitochondrial cholesterol transport is rate-limiting in the CYP27A1 (sterol 27-hydroxylase)-dependent generation of oxysterol ligands for LXR (liver X receptor) transcription factors that regulate the expression of genes encoding proteins in the cholesterol efflux pathway, such as ABC transporters (ATP-binding cassette transporters) ABCA1 and ABCG1. These transporters transfer cholesterol and/or phospholipids across the plasma membrane to (apo)lipoprotein acceptors, generating nascent HDLs (high-density lipoproteins), which can safely transport excess cholesterol through the bloodstream to the liver for excretion in bile. Utilizing information from steroidogenic tissues, we propose that perturbations in mitochondrial function may reduce the efficiency of the cholesterol efflux pathway, favouring accumulation of cholesteryl ester 'foam cells' and allowing the toxic accumulation of free cholesterol at the interface between the endoplasmic reticulum and the mitochondrial membrane. In turn, this will trigger opening of the permeability transition pore, allowing unregulated production of oxysterols via CYP27A1, allowing the accumulation of esterified forms of this oxysterol within human atherosclerotic lesions. Defective cholesterol efflux also induces endoplasmic reticulum stress, proteasomal degradation of ABCA1 and Fas-dependent apoptosis, replicating findings in macrophages in advanced atherosclerotic lesions. Small molecules targeted to mitochondria, capable of sustaining mitochondrial function or improving cholesterol trafficking may aid cholesterol efflux from macrophage 'foam' cells, regressing and stabilizing the atherosclerotic plaque.

Modulation of the Cellular microRNA Landscape: Contribution to the Protective Effects of High-Density Lipoproteins (HDL).

High-density lipoproteins (HDL) play an established role in protecting against cellular dysfunction in a variety of different disease contexts; however, harnessing this therapeutic potential has proved challenging due to the heterogeneous and relative instability of this lipoprotein and its variable cargo molecules. The purpose of this study is to examine the contribution of microRNA (miRNA; miR) sequences, either delivered directly or modulated endogenously, to these protective functions. This narrative review introduces the complex cargo carried by HDL, the protective functions associated with this lipoprotein, and the factors governing biogenesis, export and the uptake of microRNA. The possible mechanisms by which HDL can modulate the cellular miRNA landscape are considered, and the impact of key sequences modified by HDL is explored in diseases such as inflammation and immunity, wound healing, angiogenesis, dyslipidaemia, atherosclerosis and coronary heart disease, potentially offering new routes for therapeutic intervention.

Increased central and peripheral inflammation and inflammatory hyperalgesia in Zucker rat model of leptin receptor deficiency and genetic obesity.

This study investigated whether sensitivity to nociceptive stimuli is altered in obese rats using established models of inflammatory pain, and using real-time PCR, profiled alterations in expression of key adipokine and inflammatory mediator mRNA (adiponectin, tumor necrosis factor-α, interleukin-1ß, cyclooxygenase-2, inducible nitric oxide synthase (iNOS)) in spinal cord with obesity. Responses to thermal and mechanical stimulation of the hindpaw and paw oedema were assessed in adult male Zucker fatty rats (fa/fa) and their lean littermates (fa/-; n = 6-9 per group) in the absence of inflammation (acute nociception), then in response to intradermal hindpaw injection of carrageenan (3%; 50 µl) or capsaicin (10 µg; 50 µl) or hindpaw incision. The analgesic potency of morphine (1, 2.5 or 5 mg kg(-1) or vehicle; s.c.) was also assessed. Acute nociception was unaltered in obese animals, but following carrageenan-induced inflammation the obese rats were significantly more sensitive to mechanical and thermal stimulation of the inflamed paw, and displayed greater paw oedema. No difference in the capsaicin- or paw-incision-induced pain sensitivity or in the analgesic potency of morphine was observed between groups. Levels of adiponectin and inducible nitric oxide synthase mRNA were downregulated in spinal cord from obese rats, whereas tumour necrosis factor-α mRNA was upregulated; interleukin-1ß and cyclo-oxygenase were unchanged. The increased pain sensitivity and inflammatory response together with changes in spinal adipokine expression in obese rats fit well with the hypothesis that obesity is a chronic low-grade inflammatory disorder, producing a state where responses to subsequent inflammatory challenge are potentiated.

Mitochondrial function is involved in regulation of cholesterol efflux to apolipoprotein (apo)A-I from murine RAW 264.7 macrophages.

BACKGROUND: Mitochondrial DNA damage, increased production of reactive oxygen species and progressive respiratory chain dysfunction, together with increased deposition of cholesterol and cholesteryl esters, are hallmarks of atherosclerosis. This study investigated the role of mitochondrial function in regulation of macrophage cholesterol efflux to apolipoprotein A-I, by the addition of established pharmacological modulators of mitochondrial function. METHODS: Murine RAW 264.7 macrophages were treated with a range of concentrations of resveratrol, antimycin, dinitrophenol, nigericin and oligomycin, and changes in viability, cytotoxicity, membrane potential and ATP, compared with efflux of [3H]cholesterol to apolipoprotein (apo) A-I. The effect of oligomycin treatment on expression of genes implicated in macrophage cholesterol homeostasis were determined by quantitative polymerase chain reaction, and immunoblotting, relative to the housekeeping enzyme, Gapdh, and combined with studies of this molecule on cholesterol esterification, de novo lipid biosynthesis, and induction of apoptosis. Significant differences were determined using analysis of variance, and Dunnett's or Bonferroni post t-tests, as appropriate. RESULTS: The positive control, resveratrol (24 h), significantly enhanced cholesterol efflux to apoA-I at concentrations ≥30 µM. By contrast, cholesterol efflux to apoA-I was significantly inhibited by nigericin (45%; p<0.01) and oligomycin (55%; p<0.01), under conditions (10 µM, 3 h) which did not induce cellular toxicity or deplete total cellular ATP content. Levels of ATP binding cassette transporter A1 (ABCA1) protein were repressed by oligomycin under optimal efflux conditions, despite paradoxical increases in Abca1 mRNA. Oligomycin treatment did not affect cholesterol biosynthesis, but significantly inhibited cholesterol esterification following exposure to acetylated LDL, and induced apoptosis at ≥30 µM. Finally, oligomycin induced the expression of genes implicated in both cholesterol efflux (Abca1, Abcg4, Stard1) and cholesterol biosynthesis (Hmgr, Mvk, Scap, Srebf2), indicating profound dysregulation of cholesterol homeostasis. CONCLUSIONS: Acute loss of mitochondrial function, and in particular Δψm, reduces cholesterol efflux to apoA-I and dysregulates macrophage cholesterol homeostasis mechanisms. Bioavailable antioxidants, targeted to mitochondria and capable of sustaining effective mitochondrial function, may therefore prove effective in maintenance of arterial health.

Homo Sapiens (Hsa)-microRNA (miR)-6727-5p Contributes to the Impact of High-Density Lipoproteins on Fibroblast Wound Healing In Vitro.

Chronic, non-healing wounds are a significant cause of global morbidity and mortality, and strategies to improve delayed wound closure represent an unmet clinical need. High-density lipoproteins (HDL) can enhance wound healing, but exploitation of this finding is challenging due to the complexity and instability of these heterogeneous lipoproteins. The responsiveness of primary human neonatal keratinocytes, and neonatal and human dermal fibroblasts (HDF) to HDL was confirmed by cholesterol efflux, but promotion of 'scrape' wound healing occurred only in primary human neonatal (HDFn) and adult fibroblasts (HDFa). Treatment of human fibroblasts with HDL induced multiple changes in the expression of small non-coding microRNA sequences, determined by microchip array, including hsa-miR-6727-5p. Intriguingly, levels of hsa-miR-6727-5p increased in HDFn, but decreased in HDFa, after exposure to HDL. Delivery of a hsa-miR-6727-5p mimic elicited repression of different target genes in HDFn (ZNF584) and HDFa (EDEM3, KRAS), and promoted wound closure in HDFn. By contrast, a hsa-miR-6727-5p inhibitor promoted wound closure in HDFa. We conclude that HDL treatment exerts distinct effects on the expression of hsa-miR-6727-5p in neonatal and adult fibroblasts, and that this is a sequence which plays differential roles in wound healing in these cell types, but cannot replicate the myriad effects of HDL.

Transmembrane stem cell factor protein therapeutics enhance revascularization in ischemia without mast cell activation.

Stem cell factor (SCF) is a cytokine that regulates hematopoiesis and other biological processes. While clinical treatments using SCF would be highly beneficial, these have been limited by toxicity related to mast cell activation. Transmembrane SCF (tmSCF) has differential activity from soluble SCF and has not been explored as a therapeutic agent. We created novel therapeutics using tmSCF embedded in proteoliposomes or lipid nanodiscs. Mouse models of anaphylaxis and ischemia revealed the tmSCF-based therapies did not activate mast cells and improved the revascularization in the ischemic hind limb. Proteoliposomal tmSCF preferentially acted on endothelial cells to induce angiogenesis while tmSCF nanodiscs had greater activity in inducing stem cell mobilization and recruitment to the site of injury. The type of lipid nanocarrier used altered the relative cellular uptake pathways and signaling in a cell type dependent manner. Overall, we found that tmSCF-based therapies can provide therapeutic benefits without off target effects.