Time-multiplexed Laguerre-Gaussian holographic optical tweezers for biological applications

A ferroelectric liquid crystal spatial light modulator is used to generate up to 24 independently controllable traps in a holographic optical tweezers system using time-multiplexed Fresnel zone plates. For use in biological applications, helical zone plates are used to generate Laguerre-Gaussian laser modes. The high speed switching of the ferroelectric device together with recent advances in computer technology enable fast, smooth movement of traps that can be independently controlled in real time. This is demonstrated by the trapping and manipulation of yeast cells and fungal spores. (c) 2006 Optical Society of America.

Decision-making in a phase II clinical trial: a new approach combining Bayesian and frequentist concepts

The aim of a phase H clinical trial is to decide whether or not to develop an experimental therapy further through phase III clinical evaluation. In this paper, we present a Bayesian approach to the phase H trial, although we assume that subsequent phase III clinical trials will hat,e standard frequentist analyses. The decision whether to conduct the phase III trial is based on the posterior predictive probability of a significant result being obtained. This fusion of Bayesian and frequentist techniques accepts the current paradigm for expressing objective evidence of therapeutic value, while optimizing the form of the phase II investigation that leads to it. By using prior information, we can assess whether a phase II study is needed at all, and how much or what sort of evidence is required. The proposed approach is illustrated by the design of a phase II clinical trial of a multi-drug resistance modulator used in combination with standard chemotherapy in the treatment of metastatic breast cancer. Copyright (c) 2005 John Wiley & Sons, Ltd.

Bone morphogenetic protein (BMP) ligands and receptors in bovine ovarian follicle cells: actions of BMP-4,-6 and-7 on granulosa cells and differential modulation of Smad-1 phosphorylation by follistatin

Given the paucity of information on the potential roles of bone morphogenetic proteins (BMPs) in the ruminant ovary we conducted immunolocalization and functional studies on cells isolated from bovine antral follicles. Immunocytochemistry revealed expression of BMP-4 and -7 in isolated theca cells whereas granulosa cells and oocytes selectively expressed RMP-6. All three cell types expressed a range of BMP-responsive type-I (BMPRIB, ActRI) and type-II (BMPRII, ActRII, ActRIIB) receptors supporting autocrine/paracrine roles within the follicle. This was reinforced by functional experiments on granulosa cells which showed that BMP-4, -6 and -7 promoted cellular accumulation of phosphorylated Smad-1 but not Smad-2 and enhanced 'basal' and IGF-stimulated secretion of oestradiol (E2), inhibin-A, activin-A and follistatin (FS). Concomitantly, each BMP suppressed 'basal' and IGF-stimulated progesterone secretion, consistent with an action to prevent or delay atresia and/or luteinization. BMPs also increased viable cell number under 'basal' (BMP-4 and -7) and IGF-stimulated (BMP-4, -6 and -7) conditions. Since FS, a product of bovine granulosa cells, has been shown to bind several BMPs, we used the Biacore technique to compare its binding affinities for activin-A (prototype FS ligand) and BMP-4, -6 and -7. Compared with activin-A (K-d 0.28 +/- 0.02 nM; 100%), the relative affinities of FS for BMP-4, -6 and -7 were 10, 5 and 1% respectively. Moreover, studies on granulosa cells showed that preincubation of ligand with excess FS abolished activin-A-induced phosphorylation of Smad-2 and BMP-4-induced phosphorylation of Smad-1. However, FS only partially reversed BMP-6-induced Smad-1 phosphorylation and had no inhibitory effect on BMP-7-induced Smad-1 phosphorylation. These findings support functional roles for BMP-4, -6 and -7 as paracrine/autocrine modulators of granulosa cell steroidogenesis, peptide secretion and proliferation in bovine antral follicles. The finding that FS can differentially modulate BMP-induced receptor activation and that this correlates with the relative binding affinity of FS for each BMP type implicates FS as a potential modulator of BMP action in the ovary.

Neurokinin B is a paracrine vasodilator in the human fetal placental circulation

Neurokinin (NK) B is a member of the tachykinin family of neurotransmitters, exerting hypotensive or hypertensive effects in the mammalian vasculature through synaptic release from peripheral neurons, according to either NK1 and NK2 or NK3 receptor subtype expression, respectively. There is recent evidence that NKB is expressed by the syncytiotrophoblast of the human placenta, an organ that is not innervated. We hypothesized that NKB is a paracrine modulator of tone in the fetal placental circulation. We tested this hypothesis using the in vitro perfused human placental cotyledon. Our data show that NKB is a dilator of the fetal vasculature, causing a maximal 25.1+/-4.5% (mean+/-SEM; n=5) decrease in fetal-side arterial hydrostatic pressure (5-muM NKB bolus; effective concentration in the circulation, 1.89 nM) after preconstriction with U-46619. RT-PCR demonstrated the presence of mRNA for NK1 and NK2 tachykinin receptors in the placenta. Using selective receptor antagonists, we found that NKB-induced vasodilation is through the NK1 receptor subtype. We found no evidence for the involvement of either nitric oxide or prostacyclin in this response. This study demonstrates a paracrine role for NKB in the regulation of fetal placental vascular tone.

The challenges for molecular nutrition research 1: linking genotype to healthy nutrition

Nutrition science finds itself at a major crossroad. On the one hand we can continue the current path, which has resulted in some substantial advances, but also many conflicting messages which impair the trust of the general population, especially those who are motivated to improve their health through diet. The other road is uncharted and is being built over the many exciting new developments in life sciences. This new era of nutrition recognizes the complex relation between the health of the individual, its genome, and the life-long dietary exposure, and has lead to the realisation that nutrition is essentially a gene - environment interaction science. This review on the relation between genotype, diet and health is the first of a series dealing with the major challenges in molecular nutrition, analyzing the foundations of nutrition research. With the unravelling of the human genome and the linking of its variability to a multitude of phenotypes from " healthy'' to an enormously complex range of predispositions, the dietary modulation of these propensities has become an area of active research. Classical genetic approaches applied so far in medical genetics have steered away from incorporating dietary effects in their models and paradoxically, most genetic studies analyzing diet-associated phenotypes and diseases simply ignore diet. Yet, a modest but increasing number of studies are accounting for diet as a modulator of genetic associations. These range from observational cohorts to intervention studies with prospectively selected genotypes. New statistical and bioinformatics approaches are becoming available to aid in design and evaluation of these studies. This review discusses the various approaches used and provides concrete recommendations for future research.

Impact of apoE genotype on oxidative stress, inflammation and disease risk

Although in developing countries an apolipoprotein E4 (apoE4) genotype may offer an evolutionary advantage, as it has been shown to offer protection against certain infectious disease, in Westernised societies it is associated with increased morbidity and mortality, and represents a significant risk factor for cardiovascular disease, late-onset Alzheimer's disease and other chronic disorders. ApoE is an important modulator of many stages of lipoprotein metabolism and traditionally the increased risk was attributed to higher lipid levels in E4 carriers. However, more recent evidence demonstrates the multifunctional nature of the apoE protein and the fact that the impact of genotype on disease risk may be in large part due to an impact on oxidative status or the immunomodulatory/anti-inflammatory properties of apoE. An increasing number of studies in cell lines, targeted replacement rodents and human volunteers indicate higher oxidative stress and a more pro-inflammatory state associated with the F,4 allele. The impact of genotype on the antioxidant and immunomodulatory/anti-inflammatory properties of apoE is the focus of the current review. Furthermore, current information on the impact of environment (diet, exercise, smoking status, alcohol) on apoE genotype-phenotype associations are discussed with a view to identifying particular lifestyle strategies that could be adapted to counteract the 'at-risk' E4 genotype.

System performance in IR atmospheric radiometry

System aspects of filter radiometer optics used to sense planetary atmospheres are described. Thus the lenses, dichroic beamsplitters and filters in longwave channels of the Mars Observer PMIRR Pressure Modulator Infrared radiometer instrument are assessed individually, and as systems at 20.7µm, 31.9µm, 47.2µm wavelength. A window filter and a longwave calibration filter of the SCARAB earth observer instrument are assessed similarly.

Rhinocetin, a venom-derived integrin-specific antagonist inhibits collagen-induced platelet and endothelial cell functions

Snaclecs are small non-enzymatic proteins present in viper venoms reported to modulate haemostasis of victims through effects on platelets, vascular endothelial and smooth muscle cells. In this study, we have isolated and functionally characterised a snaclec which we named rhinocetin from the venom of West African gaboon viper, Bitis gabonica rhinoceros. Rhinocetin was shown to comprise α and β chains with the molecular masses of 13.5 and 13kDa respectively. Sequence and immunoblot analysis of rhinocetin confirmed this to be a novel snaclec. Rhinocetin inhibited collagen-stimulated activation of human platelets in dose dependent manner, but displayed no inhibitory effects on glycoprotein VI (collagen receptor) selective agonist, CRP-XL-, ADP- or thrombin-induced platelet activation. Rhinocetin antagonised the binding of monoclonal antibodies against the α2 subunit of integrin α2β1 to platelets and coimmunoprecipitation analysis confirmed integrin α2β1 as a target for this venom protein. Rhinocetin inhibited a range of collagen induced platelet functions such as fibrinogen binding, calcium mobilisation, granule secretion, aggregation and thrombus formation. It also inhibited integrin α2β1 dependent functions of human endothelial cells. Together, our data suggest rhinocetin to be a modulator of integrin α2β1 function and thus may provide valuable insights into the role of this integrin in physiological and pathophysiological scenarios including haemostasis, thrombosis and envenomation.

Dietary fat manipulation has a greater impact on postprandial lipid metabolism than the apolipoprotein E (epsilon) genotype: insights from the SATgenε study

Scope: Our aim was to determine the effects of chronic dietary fat manipulation on postprandial lipaemia according to apolipoprotein (APO)E genotype. Methods and results:Men (mean age 53 (SD 9) years), prospectively recruited for the APOE genotype (n = 12 E3/E3, n = 11 E3/E4), were assigned to a low fat (LF), high fat, high-saturated fat (HSF), and HSF diet with 3.45 g/day docosahexaenoic acid (HSF-DHA), each for an 8-week period in the same order. At the end of each dietary period, a postprandial assessment was performed using a test meal with a macronutrient profile representative of that dietary intervention. A variable postprandial plasma triacylglycerol (TAG) response according to APOE genotype was evident, with a greater sensitivity to the TAG-lowering effects of DHA in APOE4 carriers (p ≤ 0.005). There was a lack of an independent genotype effect on any of the lipid measures. In the groups combined, dietary fat manipulation had a significant impact on lipids in plasma and Svedberg flotation rate (Sf) 60–400 TAG-rich lipoprotein fraction, with lower responses following the HSF-DHA than HSF intervention (p < 0.05). Conclusion: Although a modest impact of APOE genotype was observed on the plasma TAG profile, dietary fat manipulation emerged as a greater modulator of the postprandial lipid response in normolipidaemic men.

Ovarian expression of insulin-like peptide 3 (INSL3) and its receptor (RXFP2) during development of bovine antral follicles and corpora lutea and measurement of circulating INSL3 levels during synchronized estrous cycles

Insulin-like peptide 3 (INSL3), a major product of testicular Leydig cells, is also expressed by the ovary but its functional role remains poorly understood. Here, we quantified expression of INSL3 and its receptor RXFP2 in theca interna (TIC) and granulosa (GC) compartments of developing bovine antral follicles and in corpora lutea (CL). INSL3 and RXFP2 mRNA levels were much higher in TIC than GC and increased progressively during follicle maturation with INSL3 peaking in large (11-18mm) estrogen-active follicles and RXFP2 peaking in 9-10mm follicles before declining in larger (11-18mm) follicles. Expression of both INSL3 and RXFP2 in CL was much lower than in TIC. In situ hybridization and immunohistochemistry confirmed abundant expression of INSL3 mRNA and protein in TIC. These observations indicate follicular TIC rather than CL as the primary site of both INSL3 production and action, implying a predominantly auto-/paracrine role in TIC. To corroborate the above findings, we showed that in vitro exposure of TIC to a luteinizing concentration of LH greatly attenuated expression of both INSL3 and its receptor while increasing progesterone secretion and expression of STAR and CYP11A1. Moreover, in vivo, a significant cyclic variation in plasma INSL3 was observed during synchronized estrous cycles. INSL3 and estradiol-17β followed a similar pattern, both increasing after luteolysis, before falling sharply after the LH surge. Thus, theca-derived INSL3, likely from the dominant pre-ovulatory follicle, is detectable in peripheral blood of cattle and expression is down-regulated during luteinisation induced by the pre-ovulatory LH surge. Collectively, these findings underscore the likely role of INSL3 as an important intrafollicular modulator of TIC function/steroidogenesis, whilst raising doubts about its potential contribution to CL function.