Robust smooth segmentation approach for array CGH data analysis

MOTIVATION: Array comparative genomic hybridization (aCGH) provides a genome-wide technique to screen for copy number alteration. The existing segmentation approaches for analyzing aCGH data are based on modeling data as a series of discrete segments with unknown boundaries and unknown heights. Although the biological process of copy number alteration is discrete, in reality a variety of biological and experimental factors can cause the signal to deviate from a stepwise function. To take this into account, we propose a smooth segmentation (smoothseg) approach. METHODS: To achieve a robust segmentation, we use a doubly heavy-tailed random-effect model. The first heavy-tailed structure on the errors deals with outliers in the observations, and the second deals with possible jumps in the underlying pattern associated with different segments. We develop a fast and reliable computational procedure based on the iterative weighted least-squares algorithm with band-limited matrix inversion. RESULTS: Using simulated and real data sets, we demonstrate how smoothseg can aid in identification of regions with genomic alteration and in classification of samples. For the real data sets, smoothseg leads to smaller false discovery rate and classification error rate than the circular binary segmentation (CBS) algorithm. In a realistic simulation setting, smoothseg is better than wavelet smoothing and CBS in identification of regions with genomic alterations and better than CBS in classification of samples. For comparative analyses, we demonstrate that segmenting the t-statistics performs better than segmenting the data. AVAILABILITY: The R package smoothseg to perform smooth segmentation is available from http://www.meb.ki.se/~yudpaw.     

Feeding behaviour, swimming activity and boldness explain variation in feed intake and growth of sole (Solea solea) reared in captivity

The major economic constraint for culturing sole (Solea solea) is its slow and variable growth. The objective was to study the relationship between feed intake/efficiency, growth, and (non-) feeding behaviour of sole. Sixteen juveniles with an average (SD) growth of 2.7 (1.9) g/kg^0.8/d were selected on their growth during a 4-week period in which they were housed communally with 84 other fish. Selected fish were housed individually during a second 4-week period to measure individual feed intake, growth, and behaviour. Fish were hand-fed three times a day during the dark phase of the day until apparent satiation. During six different days, behaviour was recorded twice daily during 3 minutes by direct observations. Total swimming activity, frequency of burying and of escapes were recorded. At the beginning and end of the growth period, two sequential behavioural tests were performed: “Novel Environment” and “Light Avoidance”. Fish housed individually still exhibited pronounced variation in feed intake (CV = 23%), growth (CV = 25%) and behavior (CV = 100%). Differences in feed intake account for 79% of the observed individual differences in growth of sole. Fish with higher variation in feed intake between days and between meals within days had significantly a lower total feed intake (r = −0.65 and r = −0.77) and growth. Active fish showed significantly higher feed intake (r = 0.66) and growth (r = 0.58). Boldness during both challenge tests was related to fast growth: (1) fish which reacted with a lower latency time to swim in a novel environment had significantly higher feed intake (r = −0.55) and growth (r = −0.66); (2) fish escaping during the light avoidance test tended to show higher feed intake (P<0.1) and had higher growth (P<0.05). In conclusion, feeding consistency, swimming activity in the tank, and boldness during behavioral tests are related to feed intake and growth of sole in captivity.     

Visceral Leishmaniasis and HIV Coinfection in East Africa

Visceral Leishmaniasis (VL) is an important protozoan opportunistic disease in HIV patients in endemic areas. East Africa is second to the Indian subcontinent in the global VL caseload and first in VL-HIV coinfection rate. Because of the alteration in the disease course, the diagnostic challenges, and the poor treatment responses, VL with HIV coinfection has become a very serious challenge in East Africa today. Field experience with the use of liposomal amphotericin B in combination with miltefosine, followed by secondary prophylaxis and antiretroviral drugs, looks promising. However, this needs to be confirmed through clinical trials. Better diagnostic and follow-up methods for relapse and prediction of relapse should also be looked for. Basic research to understand the immunological interaction of the two infections may ultimately help to improve the management of the coinfection.     

Evaluation of injury thresholds for predicting severe head injuries in vulnerable road users resulting from ground impact via detailed accident reconstructions

Biomechanics and Modeling in Mechanobiology - The aim of this study was to evaluate the effectiveness of various head injury criteria and associated risk functions in prediction of vulnerable road...     

Chromophore distortions in the bacteriorhodopsin photocycle: evolution of the H-C14-C15-H dihedral angle measured by solid-state NMR.

In recent years, structural information about bacteriorhodopsin has grown substantially with the publication of several crystal structures. However, precise measurements of the chromophore conformation in the various photocycle states are still lacking. This information is critical because twists about the chromophore backbone chain can influence the Schiff base nitrogen position, orientation, and proton affinity. Here, we focus on the C14-C15 bond, using solid-state nuclear magnetic resonance spectroscopy to measure the H-C14-C15-H dihedral angle. In the resting state (bR(568)), we obtain an angle of 164 +/- 4 degrees, indicating a 16 degrees distortion from a planar all-trans chromophore. The dihedral angle is found to decrease to 147 +/- 10 degrees in the early M intermediate (M(o)) and to 150 +/- 4 degrees in the late M intermediate (M(n)). These results demonstrate changes in the chromophore conformation undetected by recent X-ray diffraction studies.     

A Rab8-specific GDP/GTP exchange factor is involved in actin remodeling and polarized membrane transport

The mechanisms mediating polarized delivery of vesicles to cell surface domains are poorly understood in animal cells. We have previously shown that expression of Rab8 promotes the formation of new cell surface domains through reorganization of actin and microtubules. To unravel the function of Rab8, we used the yeast two-hybrid system to search for potential Rab8-specific activators. We identified a coil-coiled protein (Rabin8), homologous to the rat Rabin3 that stimulated nucleotide exchange on Rab8 but not on Rab3A and Rab5. Furthermore, we show that rat Rabin3 has exchange activity on Rab8 but not on Rab3A, supporting the view that rat Rabin3 is the rat equivalent of human Rabin8. Rabin8 localized to the cortical actin and expression of Rabin8 resulted in remodeling of actin and the formation of polarized cell surface domains. Activation of PKC by phorbol esters enhanced translocation of both Rabin8 and Rab8-specific vesicles to the outer edge of lamellipodial structures. Moreover, coexpression of Rabin8 with dominant negative Rab8 (T22N) redistributes Rabin8 from cortical actin to Rab8-specific vesicles and promotes their polarized transport to cell protrusions. The C-terminal region of Rabin8 plays an essential role in this transport. We propose that Rabin8 is a Rab8-specific activator that is connected to processes that mediate polarized membrane traffic to dynamic cell surface structures.     

2‐photon excitation fluorescence microscopy enables deeper high‐resolution imaging of voltage and Ca2+ in intact mice,rat, and rabbit hearts

We describe a novel two‐photon (2P) laser scanning microscopy (2PLSM) protocol that provides ratiometric transmural measurements of membrane voltage (V_m) via Di‐4‐ANEPPS in intact mouse, rat and rabbit hearts with subcellular resolution. The same cells were then imaged with Fura‐2/AM for intracellular Ca²⁺ recordings. Action potentials (APs) were accurately characterized by 2PLSM vs. microelectrodes, albeit fast events (<1 ms) were sub‐optimally acquired by 2PLSM due to limited sampling frequencies (2.6 kHz). The slower Ca²⁺ transient (CaT) time course (>1ms) could be accurately described by 2PLSM. In conclusion, V_m ‐ and Ca²⁺‐sensitive dyes can be 2P excited within the cardiac muscle wall to provide AP and Ca²⁺ signals to ～400 µm. (© 2013 WILEY‐VCH Verlag GmbH &Co. KGaA, Weinheim)     

Population-based cervical screening with a 5-year interval in The Netherlands. Stabilization of the incidence of squamous cell carcinoma and its precursor lesions in the screened population

OBJECTIVE: To evaluate the outcome of population-based cervical screening at 5-year intervals. STUDY DESIGN: Results from the west region of the Netherlands (population 2 million) were used. The 1995-2000 round was compared with the first 2 years of the second (2001-2002). All results were prospectively collected in a central database. Positive cytologies and histoscores per 1,000 screened for preinvasive squamous cervical intraepithelial neoplasia (CIN) lesions and invasive squamous cell carcinoma were calculated. RESULTS: In the first round, 378,081 women were screened; in the second round, 100,561 women were screened. In both rounds the youngest screenees had the highest cytoscores. Cytoscores in the first round did not differ significantly from those in the second. The histoscore for CIN 1 and 2 was 1.42 per 1,000 in the first round and 1.18 per 1,000 (NS, P < .01) in the second. The histoscore for CIN 3 was 2.07 per 1,000 in the first round and 2.13 per 1,000 (NS, P < .01) in the second. Histoscores for invasive squamous cell carcinoma remained virtually the same (0.16 per 1,000 in the first, 0.14 per 1,000 in the second round). CONCLUSION: Population-based screening at 5-year intervals in the Netherlands may result in stabilization of positive cytology and of the incidence of CIN and (histologic) invasive squamous cell carcinoma. The program seems more cost effective than that of 2 decades ago, with a screening interval of 3 years.     

Normal phase high-performance liquid chromatography of pneumocandins: in situ modification of silica with L-proline to separate structural analogues

Lipopeptides such as pneumocandin B(0) are often produced by fermentation processes. Many compounds with similar structures (structural analogues), and hence similar physiochemical properties, are coproduced in the fermentation. We employed high performance liquid chromatography using silica gel as the stationary phase and a ternary ethyl acetate/MeOH/water mobile phase to separate pneumocandin B(0) from these structural analogues. Despite extensive efforts to optimize this system, two key structural analogues, pneumocandin E(0) and pneumocandin B(5), continued to be poorly resolved from the main product peak (pneumocandin B(0)). As a result, feed load was restricted and productivity was limited. In situ modification of the silica gel stationary phase with l-proline or other amino acids significantly enhances the resolution of the two key structural analogues from the compound of interest, enabling a two-fold increase in productivity. Results of a systematic study showed that the amine group in l-proline and other amino acids plays a key role in the modification of the surface of the silica gel to mediate the selectivity enhancement.     

Lipogenesis Is Decreased by Grape Seed Proanthocyanidins According to Liver Proteomics of Rats Fed a High Fat Diet

Bioactive proanthocyanidins have been reported to have several beneficial effects on health in relation to metabolic syndrome, type 2 diabetes, and cardiovascular disease. We studied the effect of grape seed proanthocyanidin extract (GSPE) in rats fed a high fat diet (HFD). This is the first study of the effects of flavonoids on the liver proteome of rats suffering from metabolic syndrome. Three groups of rats were fed over a period of 13 weeks either a chow diet (control), an HFD, or a high fat diet supplemented for the last 10 days with GSPE (HFD + GSPE). The liver proteome was fractionated, using a Triton X-114-based two-phase separation, into soluble and membrane protein fractions so that total proteome coverage was considerably improved. The data from isobaric tag for relative and absolute quantitation (iTRAQ)-based nano-LC-MS/MS analysis revealed 90 proteins with a significant (p < 0.05) minimal expression difference of 20% due to metabolic syndrome (HFD versus control) and 75 proteins due to GSPE treatment (HFD + GSPE versus HFD). The same animals have previously been studied (Quesada, H., del Bas, J. M., Pajuelo, D., Díaz, S., Fernandez-Larrea, J., Pinent, M., Arola, L., Salvadó, M. J., and Bladé, C. (2009) Grape seed proanthocyanidins correct dyslipidemia associated with a high-fat diet in rats and repress genes controlling lipogenesis and VLDL assembling in liver. Int. J. Obes. 33, 1007–1012), and GSPE was shown to correct dyslipidemia observed in HFD-fed rats probably through the repression of hepatic lipogenesis. Our data corroborate those findings with an extensive list of proteins describing the induction of hepatic glycogenesis, glycolysis, and fatty acid and triglyceride synthesis in HFD, whereas the opposite pattern was observed to a large extent in GSPE-treated animals. GSPE was shown to have a wider effect than previously thought, and putative targets of GSPE involved in the reversal of the symptoms of metabolic syndrome were revealed. Some of these novel candidate proteins such as GFPT1, CD36, PLAA (phospholipase A₂-activating protein), METTL7B, SLC30A1, several G signaling proteins, and the sulfide-metabolizing ETHE1 and SQRDL (sulfide-quinone reductase-like) might be considered as drug targets for the treatment of metabolic syndrome.An increase in high calorie diets and a sedentary lifestyle are considered the key factors in explaining the epidemic rise in obesity in developed countries (1). Obese patients, especially those with abdominal obesity due to visceral adipose tissue accumulation, run a higher risk of impaired glucose tolerance, which frequently evolves into insulin resistance (2). Obesity and insulin resistance are frequently associated with hypertension, proatherogenic dyslipidemia, chronic inflammation, a prothrombotic state, and recently also fatty liver (3), conditions that together make up what is known as metabolic syndrome and lead to an increased risk of developing cardiovascular disease (CVD)¹ and type 2 diabetes (4). Conversely, some dietary patterns and specific food components have been associated with a lower prevalence of obesity, type 2 diabetes, and CVD. In this sense, the traditional Mediterranean diet (characterized by a high fiber content, low glycemic index carbohydrates, unsaturated fats, vitamins, and antioxidant polyphenols) has been linked to a lower incidence of CVD, obesity, and type 2 diabetes (5–8). Moreover, the French population presents a very low prevalence of death due to CVD despite consuming a diet rich in saturated fats and cholesterol. This phenomenon, known as “the French paradox” (9), has been ascribed to the moderate consumption of red wine and specifically to its content of polyphenols (10–12).Polyphenols include flavonoids of which flavan-3-ols and their oligomeric forms (proanthocyanidins) have been reported to exhibit several beneficial health effects by acting as antioxidant, anticarcinogen, cardioprotective, antimicrobial, antiviral, and neuroprotective agents (for a review, see Ref. 13). Specifically, grape and wine proanthocyanidins have a cardioprotective effect through increasing plasma high density lipoprotein cholesterol, decreasing low density lipoprotein-derived atherosclerotic foam cell lesions, attenuating oxidant formation by quenching harmful radicals, increasing endothelium-dependent vasorelaxation, etc. (13). In this context, our group has been working for years on the effect of a grape seed proanthocyanidin extract (GSPE) (containing monomers and oligomers of flavan-3-ols) in relation to metabolic syndrome. In previous works, we have found that GSPE prevents oxidative injury (14), has an insulinomimetic effect on adipocytes and adipose tissue (15), modulates glucose homeostasis (16), decreases plasma levels of triglycerides (TGs) and apolipoprotein B in normolipidemic rats (17), and acts as an in vitro (18, 19) and in vivo (20) anti-inflammatory. We have also shown that GSPE decreases postprandial plasma TG and apolipoprotein B in mice through a hepatic induction of a farnesoid X receptor (FXR) and the small heterodimer partner (SHP) that in turn down-regulates SREBP1c and other lipogenic genes in the liver (21, 22). Furthermore, we have demonstrated that the molecules responsible for the reduced TG synthesis in HepG2 cells treated with GSPE are the sum of a proanthocyanidins trimer and a dimer gallate because they reproduce the GSPE effect (23).The effect of GSPE on metabolic syndrome has been studied in our laboratory by feeding rats a “cafeteria diet.” This diet is an experimental model of a western high sugar and high fat diet extensively used to produce obesity in rats because its palatability induces the animals to increase their energy intake (24). In a recent study conducted by our group (25) as well as this study, the rats were fed a high fat diet (HFD) (cafeteria diet) for 13 weeks, and one group of the animals was treated with a daily dose of GSPE (25 mg/kg of body weight) for the last 10 days (HFD + GSPE). In that study, HFD was shown to cause the animals to be overweight and to suffer from fatty liver, dyslipidemia, and hepatic overexpression of key genes involved in lipogenesis and VLDL assembly, whereas GSPE treatment corrected dyslipidemia and down-regulated some of the genes up-regulated by HFD (25).To better investigate the mechanism behind the changes observed in HFD- and HFD + GSPE-fed rats, we analyzed protein expression in the liver. Because GSPE treatment and obesity have multiple effects, a proteome-wide approach is needed to map proteins from different pathways. Proteomics studies related to obesity, metabolic syndrome, fatty liver, or insulin resistance have previously been performed on the liver (26–32). Two such studies looked into the effects of flavonoids in mouse livers (33, 34), but to our knowledge, this is the first hepatic proteome analysis of the effect of flavonoids in rats suffering from metabolic syndrome. To improve the proteome coverage of the complex liver samples, we performed a proteome fractionation according to protein solubility using a two-phase detergent protocol (35). This strategy was advantageous because it captured membrane proteins that otherwise would have been difficult to detect. The resulting soluble and membrane protein fractions were digested, iTRAQ-labeled, fractionated according to isoelectric point, and analyzed by nano-LC-MS/MS. The proteomics study presented here reports a differential expression due to HFD or HFD + GSPE for approximately 140 proteins, indicating that both conditions were potent modifiers of the liver proteome. We have focused on the sugar and lipid metabolism data, which confirmed the repression of hepatic lipogenesis in HFD + GSPE rats. Additionally, new proteins have been revealed as putative GSPE targets.