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We hypothesized that patients with chronic kidney disease (CKD) display an altered plasma amino acid (AA) metabolomic profile that could contribute to abnormal vascular maintenance of peripheral circulation in uremia. The relationships between plasma AAs and endothelial and vascular smooth muscle function in the microcirculation of CKD patients are not well understood. The objective of this study is to investigate to what extent the levels of AAs and its metabolites are changed in CKD patients and to test their relationship with endothelial and vascular smooth muscle function. Patients with CKD stages 3 and 5 and non-CKD controls are included in this study. We report that there was a significant reduction of the biopterin (BH4/BH2) ratio, which was accompanied by increased plasma levels of BH2, asymmetric dimethylarginine (ADMA) and citrulline in patients with CKD-5 vs. CKD-3 vs. controls. In vivo augmentation index measurement showed a positive association with ADMA in all participants. The contribution of nitric oxide, assessed by ex vivo assay, showed a negative association with creatinine, ADMA and citrulline in all participants. In CKD-5, BH4 negatively correlated with ADMA and ornithine levels, and the ex vivo endothelium-mediated dilatation positively correlated with phenylalanine levels. In conclusion, uremia is associated with alterations in AA metabolism that may affect endothelium-dependent dilatation and vascular stiffness in microcirculation. Interventional strategies aiming to normalize the AA metabolism could be of interest as treatment options.

Increased saturated fatty acid levels in membrane phospholipids have been implicated in the development of metabolic disease. Here, we tested the hypothesis that increased saturated fatty acid (SFA) content in cell membranes negatively impacts adipocyte insulin signaling. Pre-adipocyte cell models with elevated SFA levels in phospholipids were generated by disrupting the ADIPOR2 locus, which resulted in a striking two-fold increase in SFA-containing phosphatidylcholines and phosphatidylethanolamines, which persisted in differentiated adipocytes. Similar changes in phospholipid composition were observed in white adipose tissues isolated from the ADIPOR2 knockout mice. The SFA levels in phospholipids could be further increased by treating ADIPOR2-deficient cells with palmitic acid and resulted in reduced membrane fluidity and endoplasmic reticulum stress in mouse and human pre-adipocytes. Strikingly, increased SFA levels in differentiated adipocyte phospholipids had no effect on adipocyte gene expression or insulin signaling in vitro. Similarly, increased adipocyte phospholipid saturation did not impair white adipose tissue function in vivo, even in mice fed a high saturated fat diet at thermoneutrality. We conclude that increasing SFA levels in adipocyte phospholipids is well tolerated and does not affect adipocyte insulin signaling in vitro and in vivo.

Sodium-glucose cotransporter 2 (SGLT2) inhibitors have been shown to increase ketone bodies in patients with type 2 diabetes; however, the underlying mechanisms have not been fully elucidated. Here we examined the effect of the SGLT2 inhibitor dapagliflozin (1 mg/kg/day, formulated in a water, PEG400, ethanol, propylene glycol solution, 4 weeks) on lipid metabolism in obese Zucker rats. Fasting FFA metabolism was assessed in the anesthetized state using a [9,10

A stabilized high drug load intravenous formulation could allow compounds with less optimal pharmacokinetic profiles to be developed. Polyethylene glycol (PEG)-ylation is a frequently used strategy for particle delivery systems to avoid the liver, thereby extending blood circulation time. The present work reports the mouse in vivo distribution after i.v. administration of a series of nanocrystals prepared with the bead milling technique and PEG-ylated with DSPE-PEG2000 and Pluronic F127, with and without polyvinylpyrrolidone K30 (PVP)/Aerosol OT (AOT) as primary stabilizers. While all formulations were cleared significantly faster than expected from nanocrystal dissolution alone, purely DSPE-PEG2000 PEG-ylated particles displayed prolonged circulation time (particles elimination half-life of 9 min) compared to DSPE-PEG2000/PVP/AOT formulation (half-life of 3 min). The two Pluronic F127 stabilized formulations displayed similar half-lives (9 min with and without PVP/AOT, respectively). Whole tissue kinetics shows that clearance of particles could be attributed to accumulation in the liver. A separate in vivo study addressed the liver cell distribution after administration. Dissolved compound accumulated in hepatocytes only, while particles were distributed between liver sinusoidal endothelial cells and Kupffer cells. More DSPE-PEG2000/PVP/AOT stabilized particles accumulated in the liver, preferably in Kupffer cells, compared to Pluronic F127/PVP/AOT stabilized particles. The present study extends the understanding of PEG-ylation and “stealth” behaviour to also include nanocrystals.

Heart failure with preserved ejection fraction (HFpEF) and HF with reduced ejection fraction (HFrEF) are associated with metabolic derangements, which may have different pathophysiological implications.In new-onset HFpEF (EF of ≥50%, n = 46) and HFrEF (EF of40%, n = 75) patients, 109 endogenous plasma metabolites including amino acids, phospholipids and acylcarnitines were assessed using targeted metabolomics. Differentially altered metabolites and associations with clinical characteristics were explored. Patients with HFpEF were older, more often female with hypertension, atrial fibrillation, and diabetes compared with patients with HFrEF. Patients with HFpEF displayed higher levels of hydroxyproline and symmetric dimethyl arginine, alanine, cystine, and kynurenine reflecting fibrosis, inflammation and oxidative stress. Serine, cGMP, cAMP, l-carnitine, lysophophatidylcholine (18:2), lactate, and arginine were lower compared with patients with HFrEF. In patients with HFpEF with diabetes, kynurenine was higher (P = .014) and arginine lower (P = .014) vs patients with no diabetes, but did not differ with diabetes status in HFrEF. Decreasing kynurenine was associated with higher eGFR only in HFpEF (PPatients with new-onset HFpEF compared with patients with new-onset HFrEF display a different metabolic profile associated with comorbidities, such as diabetes and kidney dysfunction. HFpEF is associated with indices of increased inflammation and oxidative stress, impaired lipid metabolism, increased collagen synthesis, and downregulated nitric oxide signaling. Together, these findings suggest a more predominant systemic microvascular endothelial dysfunction and inflammation linked to increased fibrosis in HFpEF compared with HFrEF.ClinicalTrials.gov NCT03671122 https://clinicaltrials.gov.

Low basal endogenous concentrations (20 pg/mL) of the 5-lipoxygenase (5-LO) pathway biomarker leukotriene E

Background Heart failure with preserved (HFpEF) and reduced (HFrEF) ejection fraction are both associated with metabolic derangements which may have different pathophysiological implications Purpose To identify metabolites and pathways differentially altered with the potential to differentiate HFpEF from HFrEF. Methods In the PREFERS Stockholm study (Preserved and Reduced Ejection Fraction Epidemiological Regional Study) 121 endogenous plasma metabolites were assessed by targeted mass spectrometry. Partial Least Squares Discriminant Analysis (PLS-DA) was used to identify metabolites differentially altered in new onset HF divided into HFpEF (EF ≥50%, n=46) versus HFrEF (ÈF Results Compared to HFrEF, HFpEF patients were older; 77 vs 65 years (p HFpEF patients had higher levels of hydroxyproline, cysteine, symmetric dimethyl arginine, alanine, kynurenine and acylcarinitines and lower levels of cAMP, lysoPC, L-carnitine, arginine, cGMP, serine and lactate (Figure). HFpEF was independently associated with reduced levels of cGMP (OR 0.98 [95% CI: 0.97–0.99; p=0.008]), serine (0.97 [0.95–1.00; 0.047]) and cAMP (0.97 [0.94–0.99; 0.009]). Figure 1 Conclusions In new onset HF patients, HFpEF was associated with decreased cGMP, cAMP and serine indicating increased oxidative stress, impaired innate immune function and myocardial hypertrophy. HFpEF patients displayed a distinct metabolic profile reflecting increased endothelial dysfunction, hypoxia, inflammation and myocardial fibrosis pointing towards more involvement of microvascular dysfunction compared to HFrEF. Acknowledgement/Funding Science for Life Laboratory–Astra Zeneca; Mölndal, Sweden collaborative grant No. 1377

Sodium-glucose cotransporter 2 inhibitors (SGLT2i) are effective treatments for improving glucose control and reducing cardiovascular events in patients with T2D. In the present study we examined the metabolic effects of chronic treatment of the SGLT2i dapagliflozin in obese insulin resistant Zucker rats. Three different studies were performed with the aims to assess: 1) liver CoA intermediates, 2) whole-body FA metabolism using a constant infusion of 3H-palmitate and 3) tissue-specific rates of FA utilization and storage using 3H-labelled bromo-palmitate and 14C-labelled palmitate, respectively. Four weeks treatment with dapagliflozin (1 mg/kg/d p.o.) markedly increased glucosuria, reduced HbA1c and fasting plasma glucose and insulin levels and increased pancreatic insulin content compared to vehicle treated rats. Liver acetyl-CoA levels were increased while the malonyl-CoA/acetyl-CoA ratio was decreased following dapagliflozin treatment, suggestive of an increased liver FA oxidation. Treatment with dapagliflozin also increased liver HMG-CoA levels, an intermediate in ketone body formation, indicating that the acetyl-CoA is diverted to ketone body formation. This was supported by increased fasting plasma levels of the ketone body β-hydroxybutyrate in the dapagliflozin-treated rats compared to vehicle. Dapagliflozin treatment resulted in increased plasma FA levels and rates of whole-body FA oxidation. The increased whole-body FA oxidation was subsequently found to be driven by an increase in liver FA utilization. In conclusion, dapagliflozin improves glucose homeostasis and enhances FA utilization in insulin resistant obese animals. These results suggest that dapagliflozin-induced glucose loss in urine signals a fasting state to the liver, which leads to an increased FA oxidation and ketone body production. Disclosure T. Kroon: Employee; Self; AstraZeneca. T. Hagstedt: None. A. Kjellstedt: None. A. Lindblom: Employee; Self; AstraZeneca. L. Löfgren: Employee; Self; AstraZeneca. J. Boucher: Employee; Self; AstraZeneca. M. Sörhede Winzell: Employee; Self; AstraZeneca. D. Linden: Employee; Self; AstraZeneca. K. Wallenius: Employee; Self; AstraZeneca. Stock/Shareholder; Self; AstraZeneca.

Low basal endogenous concentrations ( 80%) of plasma LTE4 level was observed, providing pharmacological evidence that endogenous 5-LO pathway activity could be assessed.

Aims/hypothesis GPR44 (DP2, PTGDR2, CRTh2) is the receptor for the pro-inflammatory mediator prostaglandin D2 (PGD2) and it is enriched in human islets. In rodent islets, PGD2 is produced in response to glucose, suggesting that the PGD2-GPR44/DP2 axis may play a role in human islet function during hyperglycemia. Consequently, the aim of this work was to elucidate the insulinotropic role of GPR44 antagonism in vitro in human beta-cells and in type 2 diabetes (T2DM) patients. Methods We determined the drive on PGD2 secretion by glucose and IL-1beta, as well as, the impact on insulin secretion by pharmacological GPR44/DP2 antagonism (AZD1981) in human islets and beta-cells in vitro. To test if metabolic control would be improved by antagonizing a hyperglycemia-driven increased PGD2 tone, we performed a proof-of-mechanism study in 20 T2DM patients (average 54 years, HbA1c 9.4%, BMI 31.6 kg/m2). The randomized, double-blind, placebo-controlled cross-over study consisted of two three-day treatment periods (AZD1981 or placebo) separated by a three-day wash-out period. Mixed meal tolerance test (MMTT) and intravenous graded glucose infusion (GGI) was performed at start and end of each treatment period. Assessment of AZD1981 pharmacokinetics, glucose, insulin, C-peptide, glucagon, GLP-1, and PGD2 pathway biomarkers were performed. Results We found (1) that PGD2 is produced in human islet in response to high glucose or IL-1beta, but likely by stellate cells rather than endocrine cells; (2) that PGD2 suppresses both glucose and GLP-1 induced insulin secretion in vitro; and (3) that the GPR44/DP2 antagonist (AZD1981) in human beta-cells normalizes insulin secretion. However, AZD1981 had no impact on neither glucose nor incretin dependent insulin secretion in humans (GGI AUC C-peptide 1-2h and MMTT AUC Glucose 0-4h LS mean ratios vs placebo of 0.94 (80% CI of 0.90-0.98, p = 0.12) and 0.99 (90% CI of 0.94-1.05, p = 0.45), despite reaching the expected antagonist exposure. Conclusion/interpretation Pharmacological inhibition of the PGD2-GPR44/DP2 axis has no major impact on the modulation of acute insulin secretion in T2DM patients. Trial registration ClinicalTrials.gov NCT02367066.

Decrease of AMPK-related signal transduction and insufficient lipid oxidation contributes to the pathogenesis of obesity and type 2 diabetes. Previously, we identified that diacylglycerol kinase-δ (DGKδ), an enzyme involved in triglyceride biosynthesis, is reduced in skeletal muscle from type 2 diabetic patients. Here, we tested the hypothesis that DGKδ plays a role in maintaining appropriate AMPK action in skeletal muscle and energetic aspects of contraction. Voluntary running activity was reduced in DGKδ+/−mice, but glycogen content and mitochondrial markers were unaltered, suggesting that DGKδ deficiency affects skeletal muscle energetics but not mitochondrial protein abundance. We next determined the role of DGKδ in AMPK-related signal transduction and lipid metabolism in isolated skeletal muscle. AMPK activation and signaling were reduced in DGKδ+/−mice, concomitant with impaired lipid oxidation and elevated incorporation of free fatty acids into triglycerides. Strikingly, DGKδ deficiency impaired work performance, as evident by altered force production and relaxation dynamics in response to repeated contractions. In conclusion, DGKδ deficiency impairs AMPK signaling and lipid metabolism, thereby highlighting the deleterious role of excessive lipid metabolites in the development of peripheral insulin resistance and type 2 diabetes pathogenesis. DGKδ deficiency also influences skeletal muscle energetics, which may lead to low physical activity levels in type 2 diabetes.

Background: Several randomized trials have proven that preoperative chemotherapy is equivalent to adjuvant treatment, and allows for clinical and radiological assessment of efficacy in vivo. Recent results show that response-guided neoadjuvant therapy is a favorable policy for hormone receptor positive tumors, while pCR predicts prognosis for triple-negative (TNBC) and HER2 positive breast cancer (von Minckwitz 2013; Cortazar 2014). Since 2012 the FDA accepts pCR (ypT0 ypN0) as endpoint for accelerated drug approval. Methods: In total, 150 women 18 years or older with verified HER2 negative breast cancer suitable for primary medical treatment were included in the trial between September 2008 and December 2011. The patients received two courses of epirubicin and docetaxel (Taxotere®), both 75mg/m2 for the 1st two courses, followed by the same treatment with addition of bevacizumab (Avastin®) 15 mg/kg for 4 additional courses. Clinical and radiological evaluations with mammography and ultrasound were performed before start and after courses 2, 4 and 6. Core biopsies were taken before start, after 2 courses, and at time of surgery. Blood samples were drawn before and 24 hours after the first 4 courses. Results: Median age was 49 years, range: 27 to 70 years; 73% were reported as ductal, 15% as lobular, and 12% as rare histological types. Mean tumor size was 59 mm, median 55 mm, range: 20-180 mm; 3 tumors (2.0%) were reported as inflammatory, and 13 (8.7%) presented with skin involvement (T4b). Enlarged axillary nodes were found in 102 patients (68%) before start of treatment, 77 of these (64%) verified as metastatic. SNB in cases of normal axillary status was performed in 16 cases, in 9 cases (8%) with positive finding. Supra- or infraclavicular node involvement was verified in 20 cases (13%). 25% of all tumors were ER- and/or PR-negative, tumor grade based on a diagnostic biopsy was evaluable in only 83 cases. Of these, 4 (5%) were grade I, 46 (55%) grade II, and 33 (40%) grade III. Mean proliferation count (Ki67) was 37%, median 30%, range 1-90%. Breakdown into intrinsic subtypes based on immunohistochemistry defined 68 (46%) as luminal A-like, 36 (24%) as luminal B-like, and 44 (30%) as TNBC. pCR was achieved in 20 cases, 3 (2%) luminal A-like, 5 (3.4%) luminal B-like and 12 (8.2%) TNBC. After 2.2 years of follow-up, 35 patients (23.3%) have experienced recurrence and 18 of these (12%) have deceased due to breast cancer, among these 6 despite pCR, 2 classified as luminal B-like, and 4 as TNBC. The molecular subtype of the tumor predicted outcome, but pCR was not in our material, even after adjustment for tumor size at diagnosis, a predictor of favorable outcome. The number of events in relation to molecular subtypes is however limited. Updated outcome data will be presented. Conclusions: The present trial does not confirm previously reported observations that pCR is a marker of favorable prognosis. One possible explanation is that unfavorable biological characteristics, particularly heterogeneity, may increase with tumor burden. Genomic and proteomic analyses are currently ongoing. Citation Format: Thomas Hatschek, Judith Bjöhle, Elisabet Lidbrink, Tobias Lekberg, Niklas Loman, Anna von Wachenfeldt Väppling, Martin Söderberg, Zakaria Einbeigi, Lena Carlsson, Henrik Lindman, Irma Fredriksson, Jan Frisell, Lars Löfgren, Lisa Rydén, Mats Hellström, Mårten Fernö, Jonas Bergh. Is pathologic complete response (pCR) a valid marker of outcome even in large breast cancer? Clinical results from a neoadjuvant trial using a combination of epirubicin, docetaxel and bevacizumab (PROMIX) [abstract]. In: Proceedings of the Thirty-Seventh Annual CTRC-AACR San Antonio Breast Cancer Symposium: 2014 Dec 9-13; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2015;75(9 Suppl):Abstract nr P3-11-14.

SummaryAMP-activated protein kinase (AMPK) plays a central role in regulating metabolism and energy homeostasis. It achieves its function by sensing fluctuations in the AMP:ATP ratio. AMP deaminase (AMPD) converts AMP into IMP, and the AMPD1 isoenzyme is expressed in skeletal muscles. Here, effects of pharmacological inhibition and genetic deletion of AMPD were examined in contracting skeletal muscles. Pharmacological AMPD inhibition potentiated rises in AMP, AMP:ATP ratio, AMPK Thr172, and acetyl-CoA carboxylase (ACC) Ser218 phosphorylation induced by electrical stimulation, without affecting glucose transport. In incubated extensor digitorum longus and soleus muscles from Ampd1 knockout mice, increases in AMP levels and AMP:ATP ratio by electrical stimulation were potentiated considerably compared with muscles from wild-type mice, whereas enhanced AMPK activation was moderate and only observed in soleus, suggesting control by factors other than changes in adenine nucleotides. AMPD inhibitors could be useful tools for enhancing AMPK activation in cells and tissues during ATP-depletion.

SummaryInhibition of AMP deaminase (AMPD) holds the potential to elevate intracellular adenosine and AMP levels and, therefore, to augment adenosine signaling and activation of AMP-activated protein kinase (AMPK). To test the latter hypothesis, novel AMPD pan inhibitors were synthesized and explored using a panel of in vitro, ex vivo, and in vivo models focusing on confirming AMPD inhibitory potency and the potential of AMPD inhibition to improve glucose control in vivo. Repeated dosing of selected inhibitors did not improve glucose control in insulin-resistant or diabetic rodent disease models. Mice with genetic deletion of the muscle-specific isoform Ampd1 did not show any favorable metabolic phenotype despite being challenged with high-fat diet feeding. Therefore, these results do not support the development of AMPD inhibitors for the treatment of type 2 diabetes.

Breast magnetic resonance imaging (MRI) has shown high sensitivity in determining tumor extent, multifocality, and occult contralateral breast cancer. Low specificity, unnecessary mastectomies, and costs are arguments against MRI. The purpose of this study was to determine whether preoperative breast MRI would affect primary surgical management, reduce reexcision/reoperation procedures, and influence the choice of neoadjuvant treatment in patients with newly diagnosed breast cancer. This prospective, randomized, multicenter study included 440 breast cancer patients younger than aged 56 years from three, Swedish, large-volume breast units. Patients were randomly allocated on a 1:1 basis to either preoperative staging with breast MRI (n = 220) or no breast MRI (n = 220) (control group). Treatment planning of all patients was discussed at multidisciplinary team conferences. In patients randomized to the MRI group, who had an observed higher percentage of planned breast-conserving surgery (BCS) compared with the control group, a change from suggested breast conservation to mastectomy occurred in 23 of 153 (15 %) patients. Breast MRI provided additional information in 83 of 220 (38 %) patients, which caused a change in treatment plan in 40 (18 %). The breast reoperation rate was significantly lower in the MRI group: 11 of 220 (5 %) versus 33 of 220 (15 %) in the control group (p

In this study we present a simple and rapid method for tissue lipid extraction. Snap-frozen tissue (15–150 mg) is collected in 2 ml homogenization tubes. 500 μl BUME mixture (butanol:methanol [3:1]) is added and automated homogenization of up to 24 frozen samples at a time in less than 60 seconds is performed, followed by a 5-minute single-phase extraction. After the addition of 500 μl heptane:ethyl acetate (3:1) and 500 μl 1% acetic acid a 5-minute two-phase extraction is performed. Lipids are recovered from the upper phase by automated liquid handling using a standard 96-tip robot. A second two-phase extraction is performed using 500 μl heptane:ethyl acetate (3:1). Validation of the method showed that the extraction recoveries for the investigated lipids, which included sterols, glycerolipids, glycerophospholipids and sphingolipids were similar or better than for the Folch method. We also applied the method for lipid extraction of liver and heart and compared the lipid species profiles with profiles generated after Folch and MTBE extraction. We conclude that the BUME method is superior to the Folch method in terms of simplicity, through-put, automation, solvent consumption, economy, health and environment yet delivering lipid recoveries fully comparable to or better than the Folch method.

Lipid extraction from biological samples is a critical and often tedious preanalytical step in lipid research. Primarily on the basis of automation criteria, we have developed the BUME method, a novel chloroform-free total lipid extraction method for blood plasma compatible with standard 96-well robots. In only 60 min, 96 samples can be automatically extracted with lipid profiles of commonly analyzed lipid classes almost identically and with absolute recoveries similar or better to what is obtained using the chloroform-based reference method. Lipid recoveries were linear from 10–100 µl plasma for all investigated lipids using the developed extraction protocol. The BUME protocol includes an initial one-phase extraction of plasma into 300 µl butanol:methanol (BUME) mixture (3:1) followed by two-phase extraction into 300 µl heptane:ethyl acetate (3:1) using 300 µl 1% acetic acid as buffer. The lipids investigated included the most abundant plasma lipid classes (e.g., cholesterol ester, free cholesterol, triacylglycerol, phosphatidylcholine, and sphingomyelin) as well as less abundant but biologically important lipid classes, including ceramide, diacylglycerol, and lyso-phospholipids. This novel method has been successfully implemented in our laboratory and is now used daily. We conclude that the fully automated, high-throughput BUME method can replace chloroform-based methods, saving both human and environmental resources.

Background: A side effect of diagnostic needle biopsies is the possibility to disseminate tumour cells into the needle track, which may cause concern in certain malignant tumour types. Methods: In order to prevent tumour cell dissemination we developed a technology that uses radiofrequency (RF) pulses to sterilise the needle track and denaturate tumour cells. To determine feasibility, we applied this technology to fine needle aspiration biopsy (FNAB) and used breast cancer as a model tumour. Routine FNAB was performed in 88 patients with adenocarcinoma and blood droplets passing the skin orifice were cytomorphologically analysed for the presence of tumour cells. Results: The analysis showed the presence of tumour cells in 65/88 cases (74%). When using an experimental anti-seeding device in a subset of patients viable tumour cells were found in 0/31 cases (P

The purpose of this study was to determine the safety and efficacy of percutaneous ultrasound (US) guided preferential radiofrequency ablation (PRFA) of unifocal human invasive breast carcinoma with largest radiological diameters of up to 16 mm. Thirty-three patients were enrolled in a study to be treated prior to scheduled partial mastectomy. A needle-shaped treatment electrode, successively developed in two different sizes, was placed into the center of the lesions using ultrasound guidance. A temperature of 85 degrees C was maintained for 10 min. The analysis of the resected specimen was performed using conventional histopathological methods with the aim to determine the size of the lesion as well as the potential viability of tumor cells. Of the 33 patients enrolled 31 were treated. In 26 (84%) patients a complete ablation of the tumor was achieved. Ultrasound guided preferential radiofrequency ablation of small breast carcinoma is feasible and patient friendly. The success rate depends on accurate preoperative diagnostic imaging as well as an exact position of the needle electrode.

Estrogen hormones have a large impact on both normal development and tumorigenesis of the breast.Breast tissue samples from 49 women undergoing surgery were included. The estrogen receptors (ERα and ERβ), ERα36 and G-coupled estrogen receptor-1 (GPER) were determined in benign and malignant breast tissue.The ERα36 and ERα mRNA levels were highest in malignant tumors. Stromal ERβ immunostaining in benign tumors was higher than in the paired normal tissue. GPER expression was lowest in benign tumors. In the malignant tumors, the Nottingham Prognostic Index (NPI) correlated positively with stromal GPER and the serum testosterone level. The serum insulin-like growth factor-1 (IGF-1) level correlated negatively with GPER mRNA and glandular ERα.The expression of ERα36 is stronger in malignant breast tissue. The strong positive correlation between NPI and GPER in malignant breast stroma indicates an important role for GPER in breast cancer prognosis.

We present a new core needle biopsy and treatment electrode precision placement technique which, regardless of needle size, target lesion hardness and elasticity, makes it possible to precisely place an image guided device inside the abnormal tissue. Once inside the abnormal lesion, multiple tissue samples can be collected using a dedicated trocar and collecting system. Our unique "Fourier" driver substitutes the commonly used spring-loaded device or complements the jerky insertion technique used by experienced interventional physicians. It enables the physician to precisely and with extreme tactility maneuver even large diameter core needles or treatment-electrodes into the lesion using only a diminutive external force. This is achieved by applying supporting servo-controlled mechanical high-acceleration micro-pulses, proportional to the average vector directed by the physician. The Fourier-needle or Fourier-electrode stands completely non-moving when the system automatically goes into full idling. This means that the angle of attack successively and arbitrary can be aligned to hit the target, becoming successively symmetrically inserted into even small tumors to be treated as well as exactly hit any point outlined by real time ultrasound guiding. This kind of biopsy needle or treatment electrode placement results in a uniquely accurate and less traumatic procedure. Due to the risk of disseminating viable tumor cells the precision placement device can be combined with a computer controlled anti-seeding system, denaturizing tumor cells detached during penetration of the biopsy needle or treatment electrode.

The objective of this study was to assess efficacy and safety of percutaneous ultrasound (US) guided preferential radiofrequency ablation (PRFA) in early breast carcinoma under local anesthesia and to evaluate a new assessment protocol. Eighteen breast cancer patients were enrolled in order to receive PRFA treatment three weeks prior to resection. Pain assessment was performed using the visual analoge scale. Analysis of treatment success was performed using magnetic resonance imaging (MRI) as well as histological assays for hematoxylin & eosin (H&E) and cytokeratine 8 (CK8). In a subset of patients contrast enhanced ultrasound (CEUS) was performed before and after treatment. MRI showed no residual tumor growth in 100% (18/18) of cases. Complete tumor devitalization was indicated in 83% (15/18) of patients as judged by H&E staining and in 89% (16/18) as judged by immunostaining for CK8. In 100% (18/18) at least one histologic method showed devitalization in the entire tumor. Treatment was well tolerated. Pain experienced during the procedure was mild. US-guided PRFA of small breast carcinoma is feasible under local anesthesia. MRI and CK8 have proven valuable additions to the RF breast tumor ablation protocol. CEUS shows potential as a modality for radiological follow-up.

Ticagrelor, a P2Y12 antagonist, is an antiplatelet agent approved for the treatment of acute coronary syndromes; it also inhibits adenosine uptake by erythrocytes and other cells.To test whether ticagrelor inhibits platelet aggregation (PA) in whole blood (WB) by increasing the extracellular levels of adenosine, which inhibits PA via the A2A receptor.Collagen-induced PA was measured in WB or platelet-rich plasma (PRP) from 50 healthy subjects and two patients with inherited P2Y12 deficiency, in presence/absence of adenosine concentrations that by themselves marginally affected PA in WB, and ZM241385 (A2A antagonist). The effects of ticagrelor, the active metabolite of prasugrel (PAM) (P2Y12 antagonist), and dipyridamole (adenosine uptake inhibitor) on PA and on adenosine clearance in WB were compared.For PA in WB, adenosine contributed to drug-induced inhibition of PA; the adenosine contribution was similar for dipyridamole and ticagrelor but was significantly greater for ticagrelor than for PAM (P 0.01). For PA in PRP (no adenosine uptake by erythrocytes), adenosine contributed to inhibition of PA in the presence/absence of all tested drugs. ZM241385 reversed the inhibition by adenosine in WB and PRP. Similar results were obtained with WB and PRP from P2Y12 -deficient patients. Adenosine (7.1 μmol L(-1) ) added to WB, was detectable for 0.5 min in the presence of vehicle or PAM, for 3-6 min in the presence of ticagrelor, and for 60 min in the presence of dipyridamole.This study provides the first evidence of an additional antiplatelet mechanism by ticagrelor, mediated by the induced increase of adenosine levels.

Metabolic flexibility was assessed in male Zucker rats: lean controls, obese controls, and obese rats treated with the dual peroxisome proliferator activated receptor (PPAR) agonist, tesaglitazar, 3 μmol/kg/day for 3 weeks. Whole body glucose disposal rate () and hepatic glucose output (HGO) were assessed under basal fasting and hyperinsulinemic isoglycemic clamp conditions using [3,3H]glucose. Indices of tissue specific glucose utilization () were measured at basal, physiological, and supraphysiological levels of insulinemia using 2-deoxy-D-[2,6-3H]glucose. Finally, whole body and tissue specific FFA and glucose utilization and metabolic fate were evaluated under basal and hyperinsulinemic conditions using a combination of [U-13C]glucose, 2-deoxy-D-[U-14C]glucose, [U-14C]palmitate, and [9,10-3H]-(R)-bromopalmitate. Tesaglitazar improved whole body insulin action by greater suppression of HGO and stimulation of compared to obese controls. This involved increased insulin stimulation of in fat and skeletal muscle as well as increased glycogen synthesis. Tesaglitazar dramatically improved insulin mediated suppression of plasma FFA level, whole body turnover (), and muscle, liver, and fat utilization. At basal insulin levels, tesaglitazar failed to lower HGO or compared to obese controls. In conclusion, the results demonstrate that tesaglitazar has a remarkable ability to improve insulin mediated control of glucose and FFA fluxes in obese Zucker rats.

Introduction. Ticagrelor, a P2Y 12 antagonist, inhibits adenosine uptake by erythrocytes and increases adenosine-induced coronary blood flow. We hypothesiszed that ticagrelor inhibits platelet function not only by antagonizing P2Y 12 , but also by increasing the levels of adenosine, which inhibits platelets by interacting with the platelet A 2A receptor. Methods. Collagen-induced platelet aggregation (PA) was measured in citrate-anticoagulated whole blood (WB) (Multiplate) or platelet-rich plasma (PRP) (light-transmission aggregometry) from 12 healthy controls and 2 patients with congenital deficiency of P2Y 12 , in the presence or absence of adenosine (7.1 μM), which marginally affected PA in WB, and the A 2A inhibitor ZM241385. The effects on PA of ticagrelor, the active metabolite of prasugrel (PAM) (P2Y 12 antagonist) and dipyridamole (DYP) (ENT-1 inhibitor) were compared. The effects of these drugs on the clearance of adenosine after its in vitro addition to WB were also studied. Results. PA in WB: Adenosine contributed to drug-induced inhibition of PA in healthy controls. The magnitude of the contribution (absolute difference in % aggregation) was greater in the presence of ticagrelor (20%, p 12 -deficient patients was in agreement, but due to low numbers no statistical analysis was performed. Adenosine, added at 7.1μM to WB in vitro, was detectable up to 0.5 min in the presence of vehicle or PAM, up to 3 min in the presence of ticagrelor, and up to 60 min in the presence of DYP. These results are compatible with the hypothesis that ticagrelor and DYP, but not PAM, increase the adenosine levels by inhibiting its uptake by erythrocytes. Conclusion. This study provides the first evidence of an additional anti-platelet mechanism by ticagrelor, mediated by the induced increase of adenosine levels in WB. It remains to be established if and to what extent this additional mechanism of action contributes to the strong clinical benefit observed with ticagrelor in patients with acute coronary syndromes.

Increased visceral fat, as opposed to subcutaneous/gluteal, most strongly relates to key metabolic dysfunctions including insulin resistance, hepatic steatosis, and inflammation. Mesenteric fat hypertrophy in patients with Crohn's disease and in experimental rodent models of gut inflammation suggest that impaired gut barrier function with increased leakage of gut-derived antigens may drive visceral lipid deposition. The aim of this study was to determine whether increased intestinal permeability is associated with visceral adiposity in healthy humans. Normal to overweight female subjects were recruited from a population-based cohort. Intestinal permeability was assessed using the ratio of urinary excretion of orally ingested sucralose to mannitol (S/M). In study 1 (n = 67), we found a positive correlation between waist circumference and S/M excretion within a time frame of urine collection consistent with permeability of the lower gastrointestinal tract (6-9 hours post-ingestion; P = 0.022). These results were followed up in study 2 (n = 55) in which we used computed tomography and dual energy X-ray absorptiometry to measure visceral and subcutaneous fat areas of the abdomen, liver fat content, and total body fat of the same women. The S/M ratio from the 6-12 h urine sample correlated with visceral fat area (P = 0.0003) and liver fat content (P = 0.004), but not with subcutaneous or total body fat. This novel finding of an association between intestinal permeability and visceral adiposity and liver fat content in healthy humans suggests that impaired gut barrier function should be further explored as a possible mediator of excess visceral fat accumulation and metabolic dysfunction.

Inhibition of acetyl-CoA carboxylases has the potential for modulating long chain fatty acid biosynthesis and mitochondrial fatty acid oxidation. Hybridization of weak inhibitors of ACC2 provided a novel, moderately potent but lipophilic series. Optimization led to compounds 33 and 37, which exhibit potent inhibition of human ACC2, 10-fold selectivity over inhibition of human ACC1, good physical and in vitro ADME properties and good bioavailability. X-ray crystallography has shown this series binding in the CT-domain of ACC2 and revealed two key hydrogen bonding interactions. Both 33 and 37 lower levels of hepatic malonyl-CoA in vivo in obese Zucker rats.

Samples of urban air hydrocarbons were taken on specifically made adsorbent cartridges and analysed by gas chromatography after thermal desorption. The four isomeric butenes and 1,3-butadiene were favourably resolved and separated from the abundant alkanes on an aluminium oxide PLOT column. The concentrations of butadiene, reflecting outdoor urban exposure, were in the range of 0.5-5 micrograms/m3. An approximate 1:4 ratio was observed between butadiene and propene which both originate predominantly from vehicle exhaust. The four butenes made up approximately 50% of the propene concentration in exhaust-polluted air, with methylpropene greater than 1-butene greater than trans-2-butene greater than cis-2-butene. Petrol vapour contributed less than exhaust but about five times more to the 2-butenes than to methylpropene and 1-butene. The highest exposure levels of butadiene and butenes were consistently observed in the vicinity of exhaust pipes and petrol-fuelled vehicles.

A recently introduced technique based on differential optical absorption spectroscopy (DOAS) was compared with conventional adsorption sampling and gas chromatography for the determination of aromatic hydrocarbons in air. Measurements were made in Switzerland and Sweden, at sites with benzene and toluene concentrations up to 20 and 40 μg m−3, respectively. The hydrocarbons determined originated mainly from traffic exhaust. The results demonstrate that the version of DOAS studied is a versatile tool for continuous monitoring of benzene and toluene in air at concentrations higher than 10 μg m−3.

Hydrocarbons of particular toxicological interest were determined in Scandinavian urban air as their fraction of total hydrocarbons except methane. Samples were taken on triple-layer adsorbent cartridges and the C2-C8 hydrocarbons were analyzed by gas chromatography on an Al2O3/KCl column. The approximate ratios 10:4:1 were observed for ethene, propene and butadiene. The proportion of these combustion-derived alkenes was low for cold starts, and highest for fast traffic.

A system is described for the specific determination in air of ethene, propene, the four isomeric butenes, cyclopentene and the six isomeric pentenes. The butenes, pentenes and hexenes are well separated from the commonly occurring matrix of alkanes in much higher concentrations. Field samples were taken using easily carried equipment consisting of an air pump connected to an adsorbent cartridge. An advantageous combination of triple-layer adsorbents was found to be Tenax + Carbotrap + Carbosieve S-III. In the laboratory, the hydrocarbons were thermally desorbed into a cold trap. The trap was directly linked to the aluminium oxide PLOT column which effected the alkene separations.

The purpose of the study reported here was to develop a method for the determination of lipid classes in intestinal fluids, including bile acids (BAs). A solid-phase extraction (SPE) method using C18 and silica columns for the separation of BAs, phospholipids (PLs), and neutral lipids (NLs), including free fatty acids, has been developed and validated. Fed-state small intestinal fluid collected from humans was treated with orlistat to inhibit lipolysis and mixed with acetic acid and methanol before SPE to maximize lipid recoveries. BAs, PLs, and NLs were isolated using lipophilic and polar solvents to promote elution from the SPE columns. The different lipid classes were subsequently analyzed using three separately optimized HPLC methods with evaporative light-scattering detectors. High recoveries (>90%) of all lipids evaluated were observed, with low coefficients of variation (

To investigate the gastrointestinal secretory and enzymatic responses to a liquid meal during in vivo perfusion of the proximal human jejunum. Human intestinal fluid was collected from the proximal jejunum by single-pass in vivo perfusion (Loc-I-Gut). The fluid was quantitatively collected at 10-min intervals during 90 min while perfusing a nutritional drink at 2 mL/min. Quantification of lipids in the fluid leaving the segment was performed by using novel chromatographic methods. The overall bile acid concentration varied between 0.5 and 8.6 mM with a peak level 40 min after the start of the liquid meal perfusion. The total concentration of phospholipids was between 0.1 and 3.9 mM and there was a rapid degradation of phosphatidylcholine to lysophosphatidylcholine. The tri-, di-, monoglycerides and free fatty acid levels increased sharply in the beginning and reached steady-state levels between 7 and 9.5 mM. There is a rapid secretion of bile in response to food. Most of the dietary lipids are found in the form of their degradation products in vivo in human jejunum. This novel in vivo characterization, based on direct and high-recovery sampling of intestinal fluids, forms a basis for further development of improved in vitro drug dissolution test media.

Scientific knowledge develops in an increasingly fragmentary way. A multitude of scientific disciplines branch out. Curiosity for this development leads into quests for a unifying understanding. To a certain extent, foundational studies provide such unification. There is a tendency, however, also of a fragmentary growth of foundational studies, like in a multitude of disciplinary foundations. We suggest to look at the foundational problem, not primarily as a search for foundations for one discipline in another, as in some reductionist approach, but as a steady revelation of presuppositions for individual scientific theories -- which are bound to meet, sooner or later, in a common language. A decisive point here is our holistic conception of language, as a whole of description-interpretation processes which are entangled (complementary) in the language itself. For every language there is a linguistic complementarity. We suggest this unique form of entanglement as a unifying presupposition, ultimately foundational for all communicable knowledge. Involved is a linguistic realism, in terms of which we critically examine "language-world" problems, as exposed by Wittgenstein, and Russell, about a foundational interdependence of language and reality (world). Throughout, we attach to the development of foundational studies of mathematics, logics, and the natural sciences. In particular, we study the interpretation problem for axioms of infinity in some detail. We emphasize that the holistic concept of language contradicts Carnap’s semiotic fragmentation thesis (thus, no clean cut between syntax, semantics, pragmatics). This is the final, accepted and revised manuscript of this article. Use alternative location to go to the published version. Requires subscription.

We explain autonomy with full-fledged reference to the complementaristic conception of language and its introspective capabilities. This field that we refer to is understood to belong to a wider epistemic category than that of (the received view of) semiotics as well as of systems science. It follows from the linguistic complementarity that autonomies (self-references; independencies) can never be absolutely complete. Rather, we speak of autonomies as partial. By the tension view of the linguistic complementarity, there are possibilities of realizing high interpretability at the cost of low describability and conversely. Correspondingly, more complete autonomies can be achieved at the price of a lowered describability in the language where the phenomenon of autonomy occurs. The tension aspect is illuminated for various languages.

Publisher Summary This chapter presents the understanding of time in complementaristic language. Time cannot be completely described without some reference to time itself. Both time and language need complementaristic comprehension. The insight that science needs a complementary way of knowing, to be able to cope with time (without distorting it by fragmentation) was of course a very bold thought at the time when analytic thinking was dominant, which it has also continued to be. On the basis of several earlier investigations of language phenomena, from genetic language, through programming language, formal language, observation language, inner cerebral language, to external communication language, it is concluded that there is a common concept of language, of which all these phenomena are species. Holistic language consists of description and interpretation processes as complementaristic parts. Semiotics is fragmented into various parts—namely, syntax, semantics, and pragmatics.

Much like the way we distinguish between formalism and experimentalism, we distinguish between ascertainment by proof and ascertainment by measurement. We argue that quantum mechanics, which characteristically encompasses both kinds of ascertainment, is too complex to be fully captured by formalism alone, and needs relativization to language in its complementaristic conception. In particular, we argue that there is a partial tie between the two ascertainments. Although, at higher levels, inferences or proofs may well be accepted as less constructive than direct measurements, they are tied at a basic level of constructivity. An inference is here of the same constructive nature as that of a direct measurement. The levelled approach is helpful, e.g., for understanding Bohr´s wave-particle complementarity and its recent challenge by the double-prism experiment (as well as, e.g., for understanding a thesis of a programmable experimentability within "quantum computation").

In a large variety of disciplines, fundamental studies often straddle in self-referential situations, in need of relativization to language and complementaristic resolution. In such attempts, the languages at play are hardly visible themselves. Only shadows of language, somewhat characteristic of the various disciplines, become visible and tractable. We select two domains, quantum mechanics and cybernetics, for a comparative study of their complementarity concepts with consequence for understandings of observability, describability, and objectivity. In particular, we compare Bohr-Pauli’s "nondetachability of the observer" and von Foerster’s aphorisms for objectivity. In quantum mechanics, with its emphasis on experimentability and measurability, Bohr's primary view of complementarity takes the form of a tension between definability and measurability. In cybernetics, we have a central interest in inferribility above the more constructive measurability, and the linguistic complementarity takes the form of a tension between describability and interpretability. In quests for a complete quantum mechanical measurement language, we face an interesting situation, that of simulating semantic measurability by syntactic inferribility. It calls for a cybernetic tie, whereby the two processes of assertibility, by measurability and by provability, become united under complementarity. Although semantic and syntactic processes are complementary within the measurement language, they may be unfolded in levels of constructivity, allowing identification of the lowest levels. Namely, identification of the constructivity level of a basic measurement sentence, i.e., a sentence which can be affirmed by a direct measurement (without involving further inferences), with the lowest constructivity level of syntactic provability. We exemplify with an explanation of a recent challenge against Bohr’s wave-particle compementarity. Namely, by the so called double-prism experiment of Ghose, Home, and Agarwal. We find the experiment quite interesting, not by the alleged challenge, but by realizing that its interpretation requires a levelled approach to quantum theory.