Nordexfenfluramine causes more severe pulmonary vasoconstriction than dexfenfluramine

The anorectic agent dexfenfluramine (dex) causes the development of primary pulmonary hypertension in susceptible patients by an unknown mechanism. We compared the effects of dex with those of its major metabolite, nordexfenfluamine (nordex), in the isolated perfused rat lung and in isolated rings of resistance pulmonary arteries. Nordex caused a dose-dependent and more intense vasoconstriction, which can be inhibited by the nonspecific 5-hydroxytryptamine type 2 (5-HT(2)) blocker ketanserin. Similarly a rise in cytosolic calcium concentration ([Ca(2+)](i)) in dispersed pulmonary artery smooth muscle cells (PASMCs) induced by nordex could be prevented by ketanserin. Unlike prior observations with dex, nordex did not inhibit K(+) current or cause depolarization in PASMCs. Removal of Ca(2+) from the tissue bath or addition of nifedipine (1 microM) reduced ring contraction to nordex by 60 +/- 9 and 63 +/- 4%, respectively. The addition of 2-aminoethoxydiphenyl borate (2-APB), a blocker of store-operated channels and the inositol 1,4,5-trisphosphate receptor, caused a dose-dependent decrease in the ring contraction elicited by nordex. The combination of 2-APB (10 microM) and nifedipine (1 microM) completely ablated the nordex contraction. Likewise the release of Ca(2+) from the sarcoplasmic reticulum by cyclopiazonic acid markedly reduced the nordex contraction while leaving the KCl contraction unchanged. We conclude that nordex may be responsible for much of the vasoconstriction stimulated by dex, through the activation of 5-HT(2) receptors and that the [Ca(2+)](i) increase in rat PASMCs caused by dex/nordex is due to both influx of extracellular Ca(2+) and release of Ca(2+) from the sarcoplasmic reticulum.     

Cardiac homeostasis is independent of calf venous compliance in subjects with paraplegia

The purpose of this study was to examine cardiac hemodynamics during acute head-up tilt (HUT) and calf venous function during acute head-down tilt (HDT) in subjects with paraplegia compared with sedentary nondisabled controls. Nineteen paraplegic males (below T6) and nine age-, height-, and weight-matched control subjects participated. Heart rate, stroke volume, and cardiac output were assessed using the noninvasive acetylene uptake method. Venous vascular function of the calf was assessed using venous occlusion plethysmography. After supine measurements were collected, the table was moved to 10 degrees HDT followed by the three levels of HUT (10, 35, and 75 degrees ) in random order. Cardiac hemodynamics were similar between the groups at all positions. Calf circumference was significantly reduced in the paraplegic group compared with the control group (P < 0.001). Venous capacitance and compliance were significantly reduced in the paraplegic compared with control group at supine and HDT. Neither venous capacitance (P = 0.37) nor compliance (P = 0.19) increased from supine with 10 degrees HDT in the paraplegic group. A significant linear relationship was established between supine venous compliance and supine cardiac output in the control group (r = 0.80, P < 0.02) but not in the paraplegic group. The findings of reduced calf circumference and similar venous capacitance at supine rest and 10 degrees HDT in the paraplegic group imply that structural changes may have limited venous dispensability in individuals with chronic paraplegia. Furthermore, the lack of a relationship between supine venous compliance and supine cardiac output suggests that cardiac homeostasis does not rely on venous compliance in subjects with paraplegia.     

Development and retroviral transduction of porcine neonatal pancreatic islet cells in monolayer culture

To learn more about the potential of neonatal porcine pancreatic duct and islet cells for xenotransplantation, the development of these cells when cultured as monolayers was evaluated. Immunostaining for islet hormones and cytokeratin-7 revealed that day eight monolayers consisted of approximately 70% duct cells and less than 10% beta cells. Using Ki-67 immunostaining as a proliferation marker, the fraction of beta cells in the cell cycle was shown to decrease from 20% at day three to 10% at day eight, and for duct cells from 36 to 19%. Insulin secretion increased 2.4-fold upon glucose stimulation, and 38-fold when 10 mm theophylline was added, showing the responsiveness of the neonatal beta cells. Reaggregated monolayers consisted mostly of duct cells, but 4 weeks after transplantation, grafts contained predominantly endocrine cells, with duct cells being almost absent, suggesting in vivo differentiation of duct cells to endocrine cells. Monolayer susceptibility to retroviral transduction was also investigated using a Moloney Murine Leukemia Virus-based vector. Approximately 60% of duct cells but less than 5% of beta cells expressed the transgene, indicating that precursor duct cells are better targets for transgene expression. These results show that porcine neonatal pancreatic cells can be cultured as monolayers in preparation for transplantation. Furthermore, in such a culture setting, precursor duct cells have a high rate of proliferation and are more efficiently transduced with a retrovirus-based reporter gene than are beta cells.     

Association studies under general disease models

There is great expectation that the levels of association found between genetic markers and disease status will play a role in the location of disease genes. This expectation follows from regarding association as being proportional to linkage disequilibrium and therefore inversely related to recombination value. For disease genes with more than two alleles, the association measure is instead a weighted average of linkage disequilibria, with the weights depending on allele frequencies and genotype susceptibilities at the disease loci. There is no longer a simple relationship, even in expectation, with recombination. We adopt a general framework to examine association mapping methods which helps to clarify the nature of case-control and transmission/disequilibrium-type tests and reveals the relationship between measures of association and coefficients of linkage disequilibrium. In particular, we can show the consequences of additive and nonadditive effects at the trait locus on the behavior of these tests. These concepts have a natural extension to marker haplotypes. The association of two-locus marker haplotypes with disease phenotype depends on a weighted average of three-locus disequilibria (two markers with each disease locus). It is likely that these two-marker analyses will provide additional information in association mapping studies.     

Response of Acala cotton to nitrogen rates in the San Joaquin Valley of California

The responses of Acala cotton (Gossypium hirsutum L.) in California to a range of applied nitrogen (N) treatments were investigated in a 5-year, multisite experiment. The experiment's goals were to identify crop growth and yield responses to applied N and provide information to better assess the utility of soil residual N estimates in improving fertilizer management. Baseline fertilizer application rates for the lowest applied N treatments were based on residual soil nitrate-N (NO3-N) levels determined on soil samples from the upper 0.6 m of the soil collected prior to spring N fertilization and within 1 week postplanting each year. Results have shown positive cotton lint yield responses to increases in applied N across the 56 to 224 kg N/ha range in only 41% (16 out of 39) of test sites. Soil NO3-N monitoring to a depth of 2.4 m in the spring (after planting) and fall (postharvest) indicate most changes in soil NO3- occur within the upper 1.2 m of soil. However, some sites (those most prone to leaching losses of soluble nutrients) also exhibited net increases in soil NO3-N in the 1.2- to 2.4-m depth zone when comparing planting time vs. postharvest data. The lack of yield responses and soil NO3-N accumulations at some sites indicate that more efforts should be put into identifying the amount of plant N requirements that can be met from residual soil N, rather than solely from fertilizer N applications.     

Retinoid signalling and gene expression in neuroblastoma cells: RXR agonist and antagonist effects on CRABP-II and RARbeta expression

9-cis Retinoic acid (RA) induces gene expression in neuroblastoma cells more effectively and with different kinetics than other RA isomers, and could be acting in part through Retinoid X Receptors (RXRs). The aim of this study was to characterise the effects of an RXR agonist and RXR homodimer antagonist on the induction of cellular RA binding protein II (CRABP-II) and RA receptor-beta (RARbeta) in neuroblastoma cells in response to different retinoids. The RXR agonist, LDG1069, was as effective as all-trans RA in inducing gene expression, but less effective than 9-cis RA. The RXR-homodimer antagonist, LG100754, inhibited the induction of CRABP-II mRNA in SH-SY5Y neuroblastoma cells by 9-cis RA or the RXR-specific agonist LGD1069, but had no effect when used with all-trans RA. Conversely, LG100754 did not inhibit induction of RARbeta mRNA by 9-cis or all-trans RA, or by LGD1069. RAR- and RXR-specific ligands used together induced CRABP-II and RARbeta as effectively as 9-cis RA. These results demonstrate the value of combining RXR- and RAR-specific ligands to regulate RA-inducible gene expression. The possibility that RXR-homodimers mediate, in part, the induction of CRABP-II by 9-cis RA and RXR-specific ligands is discussed.     

The taxonomic status of Corethromyces bicolor from New Zealand, as inferred from morphological, developmental, and molecular studies

Due to the absence of antheridial characters in collected material the precise placement of Corethromyces bicolor has remained troublesome up until now. Recent re-examination of receptacular and appendage characters present in the holotype led to its transfer to the genus Mimeomyces. Fresh collections of this fungus have provided the opportunity to re-assess its taxonomic position. Based on a combination of morphological and molecular characters, this species is re-instated within the genus Corethromyces.     

Influence of temperature on Cryptosporidium parvum oocyst infectivity in river water samples as detected by tissue culture assay

Cryptosporidium parvum oocysts were stored in 1-ml aliquots of filtered river water at -20, 4, 10, and 21-23 C in the dark. Oocysts were also added to filter-sterilized river water samples and stored at 21-23 C. The infectivity of oocysts stored under different conditions was assayed at weekly intervals through infection of human adenocarcinoma ileocecal (HCT-8) cell monolayers. Wells containing between 10 and 100 foci of infection were enumerated by immunofluorescent microscopy, and the number of infective oocysts was calculated. No infectious oocysts were detected after 1 wk at -20 C. The number of infective oocysts stored at 4 C decreased 5-fold, and the number of those stored at 10 C decreased 2.5-fold after 14 wk. The infectivity of oocysts stored in potassium dichromate (positive control) at 4 C decreased 2-fold over 14 wk. The number of infective oocysts in filter-sterilized and non-filter-sterilized river water stored at 21-23 C decreased by 3.3 and 2.6 log units, respectively, over 12 wk, and no foci of infection were detected at 14 wk. The results show that as temperature increased from 4 to 23 C, the duration of oocyst infectivity decreased.