Dicyanovinylnaphthalenes for neuroimaging of amyloids and relationships of electronic structures and geometries to binding affinities

The positron-emission tomography (PET) probe 2-(1-[6-[(2-fluoroethyl)(methyl)amino]-2-naphthyl]ethylidene) (FDDNP) is used for the noninvasive brain imaging of amyloid-β (Aβ) and other amyloid aggregates present in Alzheimer’s disease and other neurodegenerative diseases. A series of FDDNP analogs has been synthesized and characterized using spectroscopic and computational methods. The binding affinities of these molecules have been measured experimentally and explained through the use of a computational model. The analogs were created by systematically modifying the donor and the acceptor sides of FDDNP to learn the structural requirements for optimal binding to Aβ aggregates. FDDNP and its analogs are neutral, environmentally sensitive, fluorescent molecules with high dipole moments, as evidenced by their spectroscopic properties and dipole moment calculations. The preferred solution-state conformation of these compounds is directly related to the binding affinities. The extreme cases were a nonplanar analog t-butyl-FDDNP, which shows low binding affinity for Aβ aggregates (520 nM K_i) in vitro and a nearly planar tricyclic analog cDDNP, which displayed the highest binding affinity (10 pM K_i). Using a previously published X-ray crystallographic model of 1,1-dicyano-2-[6-(dimethylamino)naphthalen-2-yl]propene (DDNP) bound to an amyloidogenic Aβ peptide model, we show that the binding affinity is inversely related to the distortion energy necessary to avoid steric clashes along the internal surface of the binding channel.     

Increased lipolysis and altered lipid homeostasis protect γ-synuclein–null mutant mice from diet-induced obesity

Synucleins are a family of homologous proteins principally known for their involvement in neurodegeneration. γ-Synuclein is highly expressed in human white adipose tissue and increased in obesity. Here we show that γ-synuclein is nutritionally regulated in white adipose tissue whereas its loss partially protects mice from high-fat diet (HFD)–induced obesity and ameliorates some of the associated metabolic complications. Compared with HFD-fed WT mice, HFD-fed γ-synuclein–null mutant mice display increased lipolysis, lipid oxidation, and energy expenditure, and reduced adipocyte hypertrophy. Knockdown of γ-synuclein in adipocytes causes redistribution of the key lipolytic enzyme ATGL to lipid droplets and increases lipolysis. γ-Synuclein–deficient adipocytes also contain fewer SNARE complexes of a type involved in lipid droplet fusion. We hypothesize that γ-synuclein may deliver SNAP-23 to the SNARE complexes under lipogenic conditions. Via these independent but complementary roles, γ-synuclein may coordinately modulate lipid storage by influencing lipolysis and lipid droplet formation. Our data reveal γ-synuclein as a regulator of lipid handling in adipocytes, the function of which is particularly important in conditions of nutrient excess.Understanding the link between increased adiposity and the development of metabolic disease may reveal novel therapeutic targets to counter the rising pandemic of obesity. Inhibiting adipose tissue expansion alone is likely to worsen metabolic outcome, as evidenced by human syndromes of lipodystrophy, whereby inappropriately decreased adipose mass causes severe metabolic disorders (1). Indeed, adipose tissue dysfunction and/or exceeded adipose storage capacity may underlie ectopic lipid accumulation and lipotoxicity in obesity (2). Therefore, a major challenge is to identify pathways via which adiposity can be reduced without concomitant increases in circulating lipids and attendant metabolic disease. Achieving this goal requires a better understanding of the molecular mechanisms that regulate lipid metabolism and storage in adipocytes, particularly in times of energy surplus.γ-Synuclein belongs to the synuclein family of proteins, whose founder member α-synuclein is best known for its links with neurodegenerative diseases, most notably Parkinson disease (3). To date, no clear cellular role is attributed to γ-synuclein, and ablation of γ-synuclein causes only minor changes in the nervous system (4–7). Recently, we and others have reported high levels of γ-synuclein expression in adipose tissue of humans and other mammals (8, 9). Moreover, expression of γ-synuclein is increased in the adipose tissue of obese humans and decreased during caloric restriction (8).Here we demonstrate that γ-synuclein–null mice display significantly reduced adiposity and fewer metabolic derangements compared with WT mice following high-fat feeding. This appears to result from increased adipocyte lipolysis coupled to enhanced whole-body lipid oxidation and energy expenditure. At a molecular level, we identify dual roles for γ-synuclein independently regulating lipid droplet fusion and adipocyte lipolysis to coordinately regulate triglyceride (TG) storage in adipocytes. Together, our observations reveal that γ-synuclein is a regulator of lipid metabolism and, hence, a potential therapeutic target for treatment of obesity and associated metabolic diseases.     

Site‐specific cytomorphology of disseminated PC‐3 prostate cancer cells visualized in vivo with fluorescent proteins

Circulating tumor cells (CTC) may reach multiple organ sites. However, CTC seeding and growth in distant organs is not random. Each metastatic site may contain a specific subpopulation of the original metastatic tumor capable of growing at that site. The fluorescent orthotopic prostate cancer model (PC‐3‐GFP) model was used for immunomagnetic capture of CTC. The captured CTC were efficiently cultivated in vitro. PC‐3‐GFP cells were also isolated from various metastatic sites, grown in vitro and examined under fluorescence microscopy. The differential morphology was compared of primary tumor cells, CTC and disseminated (DTC) from multiple metastatic sites, from nude mice with orthotopic PC‐3‐GFP. The cultured captured CTC and DTC from various organs have distinctive morphologies. Distinct cancer cell morphologies were observed at different metastatic sites as well as among CTC. The distinct morphologies were maintained during in vitro culture. The results demonstrate extensive tumor heterogeneity that could account for the widely different behavior of cancer cells in a single tumor. Further hetereogeneity testing would be a big promise for personalizing the cancer treatment in the future. Diagn. Cytopathol. 2013. © 2012 Wiley Periodicals, Inc.     

Treatment for brain arteriovenous malformation in the 1998–2011 period and review of the literature

Purpose

The results of the treatment of pial AVM provided at our neurosurgical centre are presented. Based on these results and on an overview of literary data on the efficacy and complications of each therapeutic modality, the algorithm of indications, as used at our institution, is presented.

Cohort of patients

The series comprises 195 patients, aged 9 to 87 years and treated in the years 1998–2011. The surgical group consists of 76 patients; of these, 49 patients solely received endovascular treatment, 25 were consulted and referred directly to the radiosurgical unit, and the remaining 45 were recommended to abide by the strategy of “watch and wait”.

Results

In the surgical group, serious complications were 3.9 %, at a 96.1 % therapeutic efficacy. As for AVM treated with purely endovascular methods, serious procedural complications were seen in 4.1 % of patients, with efficacy totalling 32.7 %. One observed patient suffered bleeding, resulting in death. For comparison with literary data for each modality, a survival analysis without haemorrhage following monotherapy for AVM with each particular modality was carried out.

Conclusions

Based on our analysis, we have devised the following algorithm of treatment:

1. We regard surgical treatment as the treatment of choice for AVM of Spetzler-Martin (S-M) grades I and II, and only for those grade III cases that are surgically accessible.
2. Endovascular intervention should mainly be used for preoperative embolisation, as a curative procedure for lower-grade AVM in patients with comorbidities, and as palliation only for higher-grade cases.
3. Stereotactic irradiation with Leksell Gamma Knife (LGK) is advisable, mainly for poorly accessible, deep-seated grade-III AV malformations. In the case of lower grades, the final decision is left to the properly informed patient.
4. Observation should be used as the method of choice in AVM of grades IV and V, where active therapy carries greater risk than the natural course of the disease.

     

Skeletal Muscle Triacylglycerol Hydrolysis Does Not Influence Metabolic Complications of Obesity

Intramyocellular triacylglycerol (IMTG) accumulation is highly associated with insulin resistance and metabolic complications of obesity (lipotoxicity), whereas comparable IMTG accumulation in endurance-trained athletes is associated with insulin sensitivity (the athlete’s paradox). Despite these findings, it remains unclear whether changes in IMTG accumulation and metabolism per se influence muscle-specific and systemic metabolic homeostasis and insulin responsiveness. By mediating the rate-limiting step in triacylglycerol hydrolysis, adipose triglyceride lipase (ATGL) has been proposed to influence the storage/production of deleterious as well as essential lipid metabolites. However, the physiological relevance of ATGL-mediated triacylglycerol hydrolysis in skeletal muscle remains unknown. To determine the contribution of IMTG hydrolysis to tissue-specific and systemic metabolic phenotypes in the context of obesity, we generated mice with targeted deletion or transgenic overexpression of ATGL exclusively in skeletal muscle. Despite dramatic changes in IMTG content on both chow and high-fat diets, modulation of ATGL-mediated IMTG hydrolysis did not significantly influence systemic energy, lipid, or glucose homeostasis, nor did it influence insulin responsiveness or mitochondrial function. These data argue against a role for altered IMTG accumulation and lipolysis in muscle insulin resistance and metabolic complications of obesity.Obesity is a global public health problem and a major risk factor for insulin resistance and type 2 diabetes. These disorders are characterized by excess lipid accumulation in multiple tissues, primarily as triacylglycerols (TAGs). The lipotoxicity hypothesis suggests that this lipid excess promotes cellular dysfunction and cell death, which ultimately contribute to insulin resistance and metabolic disease (1). However, intracellular TAG accumulation is not always associated with adverse metabolic outcomes, suggesting that TAGs themselves are not pathogenic (2). In contrast, other non-TAG lipid metabolites such as fatty acids (FAs), diacylglycerols (DAGs), and ceramides have been shown to influence glucose homeostasis and insulin action by interfering with insulin signaling and glucose transport, promoting endoplasmic reticulum stress and mitochondrial dysfunction, and activating inflammatory and apoptotic pathways (reviewed in ref. 3). Nevertheless, the precise identities and sources of these bioactive lipid intermediates remain elusive (4,5). Furthermore, whether intracellular TAGs serve as a protective sink or a toxic source of deleterious lipid metabolites that contribute to insulin resistance remains unclear (6).Since skeletal muscle is the major contributor to insulin-mediated glucose disposal, lipid excess in this tissue could have serious implications for systemic glucose homeostasis and insulin responsiveness (7). Indeed, numerous studies have demonstrated a strong association between intramyocellular triacylglycerol (IMTG) accumulation and insulin resistance (reviewed in ref. 8). In contrast, endurance exercise training is characterized by IMTG accumulation and insulin sensitivity (the athlete’s paradox) (2). This variable association between IMTG accumulation and insulin responsiveness has largely been attributed to differences in the balance between lipid delivery and muscle oxidative capacity (8–10). Not surprisingly then, most studies have focused on the impact of muscle FA uptake and/or oxidation on glucose homeostasis and insulin action (11). However, experimental manipulations of these parameters cannot distinguish among the effects of IMTGs, IMTG metabolism, and other lipid intermediates. Furthermore, accumulating evidence suggests that muscle oxidative capacity cannot entirely explain differences in IMTGs or insulin responsiveness (12). These findings have led to speculation that dynamic IMTG metabolism (i.e., TAG synthesis or hydrolysis) may be critically involved in lipid-induced insulin resistance (6). However, few studies have specifically addressed the contribution of IMTG metabolism per se to this process.The regulated storage and release of IMTGs remain poorly understood, but require the coordinated action of synthetic enzymes (i.e., diacylglycerol acyltransferases [DGATs]), hydrolytic enzymes (i.e., adipose triglyceride lipase [ATGL] and hormone sensitive lipase [HSL]), and other lipid droplet proteins (6). Specifically, modulating IMTG synthesis in murine skeletal muscle alters IMTG content and systemic glucose homeostasis, supporting a role for IMTG metabolism in metabolic disease (13–15). However, the metabolic impact of modulating IMTG hydrolysis in vivo remains unclear. Global deletion of either ATGL (16–19) or HSL (20) has produced variable results. The former, but not the latter, results in massive IMTG accumulation with improvement in systemic glucose homeostasis, suggesting that inhibition of ATGL-mediated TAG hydrolysis protects against insulin resistance. In contrast, recent studies in cardiac muscle (21) and other tissues (22,23) indicate that ATGL-mediated TAG hydrolysis is required for mitochondrial function such that enhancing, rather than inhibiting, ATGL action may improve metabolic outcomes. Nevertheless, the autonomous role of skeletal muscle TAG catabolism in influencing muscle-specific and systemic metabolic phenotypes remains unknown.The goal of the current study was to understand the contribution of IMTG hydrolysis to tissue-specific and systemic metabolic phenotypes, particularly glucose homeostasis and insulin action, in the context of obesity. We therefore generated animal models with decreased (skeletal muscle-specific ATGL knockout [SMAKO] mice) and increased (muscle creatine kinase [Ckm]-ATGL transgenic [Tg] mice) ATGL action exclusively in skeletal muscle, and assessed the metabolic consequences at baseline and in response to chronic high-fat feeding. Interestingly, modulation of IMTG hydrolysis via ATGL action did not significantly influence glucose homeostasis, insulin action, or other metabolic phenotypes in the context of obesity despite dramatic changes in IMTG content.