Critical considerations in the development of an assay for sulfidopeptide leukotrienes in plasma

A sensitive and specific assay has been developed for measurement of total sulfidopeptide leukotrienes (LT) in plasma. LTC4 and LTD4 in plasma are converted to LTE4 which is then extracted by C18 Sep-Pak binding and elution. Total LTE4 is resolved by reverse phase high performance liquid chromatography (RP-HPLC) and quantitated by radioimmunoassay (RIA). A [3H]LTE4 internal standard is added to the starting plasma sample to allow overall recovery to be calculated and to define the fractions from RP-HPLC to be assayed for LTE4-like immunoreactivity. The correlation between the measured increase in LTE4 concentration after addition of incremental amounts of LTC4 and LTE4 to plasma was 0.989 and 0.978, respectively, with slopes of 1.05 and 1.11. Addition of 51 pg/ml LTE4 to 5 ml plasma was detectable; the measured increase was 48 +/- 12 pg/ml (mean +/- SE, n = 7). The intra-assay coefficient of variation for 341 pg/ml of added LTC4 was 3.2% (n = 6). Sulfidopeptide leukotrienes could not be detected in blood samples taken from 12 normal volunteers in whom the theoretical detection limit, calculated from the sensitivity of the RIA, the overall recovery of LTE4, and the volume of plasma extracted, was 83 +/- 4 pg LTE4/ml plasma (0.19 +/- 0.01 pmol sulfidopeptide leukotriene/ml plasma; mean +/- SE).     

Swim Stress Increases the Potency of Glycine at the N-Methyl-d-Aspartate Receptor Complex

Abstract: We have previously demonstrated that chronic administration of antidepressants results in a reduction in the potency of glycine to displace 5,7-[³H]dichlorokynurenic acid (5,7-[³H]-DCKA) from the strychnine-insensitive glycine recognition site of the NMDA receptor complex. We now report that exposure of rats to the forced swim test results in a 56% increase in the potency of glycine to displace 5,7-[³H]DCKA from frontal cortical homogenates. These data are consistent with the hypothesis that the forced swim test, a preclinical screen sensitive to acute administration of antidepressant drugs and NMDA receptor antagonists, also results in adaptation of the NMDA receptor complex. Moreover, these data lend further support to the hypothesis that glutamatergic pathways are involved in the neurobiological response to stress and, potentially, in the pathophysiology of depression.     

Effects of intravenous infusions of prostaglandin D2 in man

Prostaglandin D₂ (PGD₂) was infused intravenously into normal male volunteers. Seven subjects received infusions of 16, 32, 64 ng/kg/min and six of these a further dose of 128 ng/kg/min. Each individual's maximum dose was limited by discomfort caused by intense facial flushing and nasal congestion. At these doses there was no significant effect on systolic or diastolic blood pressure nor on spirometric measurements. There was a small but statistically significant tachycardia at 64 and 128 ng/kg/min. Collagen- and adenosine diphosphate (ADP)-induced platele aggregation was not affected at any of the infusion rates. Infused PGD₂ is unlikely to be a useful antithrombotic agent.     

THE PLACE OF GOD IN SYNTHETIC BIOLOGY: HOW WILL THE CATHOLIC CHURCH RESPOND?

Some religious believers may see synthetic biology as usurping God's creative role. The Catholic Church has yet to issue a formal teaching on the field (though it has issued some informal statements in response to Craig Venter's development of a 'synthetic' cell). In this paper I examine the likely reaction of the Catholic Magisterium to synthetic biology in its entirety. I begin by examining the Church's teaching role, from its own viewpoint, to set the necessary backround and context for the discussion that follows. I then describe the Church's attitude to science, and particularly to biotechnology. From this I derive a likely Catholic theology of synthetic biology. The Church's teachings on scientific and biotech research show that it is likely to have a generally positive disposition to synbio, if it and its products can be acceptably safe. Proper evaluation of, and protection against, risk will be a significant factor in determining the morality of the research. If the risks can be minimized through regulation or other means, then the Church is likely to be supportive. The Church will also critique the social and legal environment in which the research is done, evaluating issues such as the patenting of scientific discoveries and of life.     

Oleate and linoleate enhance the growth-promoting effects of insulin-like growth factor-I through a phospholipase D-dependent pathway in arterial smooth muscle cells

Diabetes causes accelerated atherosclerosis and subsequent cardiovascular disease through mechanisms that are poorly understood. We have previously shown, using a porcine model of diabetes-accelerated atherosclerosis, that diabetes leads to an increased accumulation and proliferation of arterial smooth muscle cells in atherosclerotic lesions and that this is associated with elevated levels of plasma triglycerides. We therefore used the same model to investigate the mechanism whereby diabetes may stimulate smooth muscle cell proliferation. We show that lesions from diabetic pigs fed a cholesterol-rich diet contain abundant insulin-like growth factor-I (IGF-I), in contrast to lesions from non-diabetic pigs. Furthermore, two fatty acids common in triglycerides, oleate and linoleate, enhance the growth-promoting effects of IGF-I in smooth muscle cells isolated from these animals. These fatty acids accumulate predominantly in the membrane phospholipid pool; oleate accumulates preferentially in phosphatidylcholine and phosphatidylethanolamine, whereas linoleate is found mainly in phosphatidylethanolamine. The growth-promoting effects of oleate and linoleate depend on phospholipid hydrolysis by phospholipase D and subsequent generation of diacylglycerol. Thus, concurrent increases in levels of IGF-I and triglyceride-derived oleate and linoleate in lesions may contribute to accumulation and proliferation of smooth muscle cells and lesion progression in diabetes-accelerated atherosclerosis.     

Deletion of UCP2 in iNOS deficient mice reduces the severity of the disease during experimental autoimmune encephalomyelitis

Uncoupling protein 2 is a member of the mitochondrial anion carrier family that is widely expressed in neurons and the immune cells of humans. Deletion of Ucp2 gene in mice pre-activates the immune system leading to higher resistance toward infection and to an increased susceptibility to develop chronic inflammatory diseases as previously exemplified with the Experimental Autoimmune Encephalomyelitis (EAE), a mouse model for multiple sclerosis. Given that oxidative stress is enhanced in Ucp2-/- mice and that nitric oxide (NO) also plays a critical function in redox balance and in chronic inflammation, we generated mice deficient for both Ucp2 and iNos genes and submitted them to EAE. Mice lacking iNos gene exhibited the highest clinical score (3.4+/-0.5 p<0.05). Surprisingly, mice deficient for both genes developed milder disease with reduced immune cell infiltration, cytokines and ROS production as compared to iNos-/- mice.     

Assessment of the humoral immune response to cancer

One of the deadly hallmarks of cancer is its ability to prosper within the constraints of the host immune system. Recent advances in immunoproteomics and high-throughput technologies have lead to profiling of the antibody repertoire in cancer patients. This in turn has lead to the identification of tumour associated antigens/autoantibodies. Autoantibodies are extremely attractive and promising biomarker entities, however there has been relatively little discussion on how to interpret the humoral immune response. It may be that autoantibody profiles hold the key to ultimately uncovering neoplastic associated pathways and through the process of immunosculpting the tumour may have yielded an immune response in the early stages of malignant tumour development. The aim of this review is to discuss the utility of the autoantibody response that is elicited as a result of malignancy and discuss the advantages and limitations of autoantibody profiling. This article is part of a Special Issue entitled: Translational Proteomics.     

Epidemiology of squamous cell carcinomas in rudd Scardinius erythrophthalmus from SE Ireland

An epidemiological study was carried out to investigate the possible aetiology of squamous cell carcinomas which occur in a population of rudd Scardinius erythrophthalmus (L.) from Lough Aderry in south-east Ireland. A total of 1343 rudd were sampled from Lough Aderry and 2 nearby small lakes in spring, summer, autumn, and winter over 2 yr, 1986 to 1988. Fish were weighed, measured, sexed, aged, and examined for lesions. Water quality parameters and natural radioactivity were assessed as possible influencing factors in the disease. The prevalence of squamous cell carcinoma was 6.1% overall, with no significant difference between the lakes, seasons or years. Both male and female fish were affected, of ages from 1+ to 5+ yr. No fish of 6+ or 7+ yr was found with tumours, indicating that the neoplasm caused premature death. The female:male sex ratio of rudd with tumours was higher than that of healthy rudd overall, suggesting that males are more susceptible to the neoplasm. Of the parasites observed, Posthodiplostomum cuticola was common, but rarely found in rudd with tumours. Sphaerospora sp.was also common, but not in sufficient densities for statistical inference, and Argulus sp. was present on 7 fish. No viral particles were found. Natural radiation levels in the vicinity of the lakes were low. The lakes sampled are in an agricultural catchment, and the waters were eutrophic. While rudd are tolerant, it is likely that environmental conditions were stressful at least some of the time, with possible consequences for the immunocompetence of the rudd. The aetiology of the carcinoma is still unknown, but the possibilities of carcinogenic compounds resulting from the high nutrient levels, or of carcinogenic algal toxins produced during algal blooms, should be examined.