Larval morphology of the Nemertean Carcinonemertes epialti (Nemertea: Hoplonemertea)

The morphology of the newly hatched larva of Carcinonemertes epialti Coe has been examined by light and electron microscopy. The newly hatched larva is covered with cilia and measures about 110 μm in length. Four types of epidermal cells are recognizable: (1) Multiciliated cells, (2) vacuolated cells, (3) mucous cells, and (4) “knob cells”. The knob cells protrude from the posterior end of the larva and contain granules and bundles of microfilaments. The gut is incomplete and is located ventral to the bipartite proboscis. A bilobed brain and two subepidermal ocelli are found in the anterior end of the larva. The anterior and posterior cirri are composed of long, tightly appressed cilia that arise from an invagination of the epidermis at each end of the larva. The anterior cirrus is surrounded by two types of glandular cells. It is proposed that the knob cells have a role in larval attachment, combining the functions of the adhesive cells and anchor cells described in the duo-gland system of turbellarians. The cirri are believed to be larval sensory structures that function in substrate selection. Histological and ultrastructural observations suggest that the larvae of Carcinonemertes are relatively long lived and develop into juveniles without a drastic metamorphosis.     

Evaluation of variant identification methods for whole genome sequencing data in dairy cattle

Background

Advances in human genomics have allowed unprecedented productivity in terms of algorithms, software, and literature available for translating raw next-generation sequence data into high-quality information. The challenges of variant identification in organisms with lower quality reference genomes are less well documented. We explored the consequences of commonly recommended preparatory steps and the effects of single and multi sample variant identification methods using four publicly available software applications (Platypus, HaplotypeCaller, Samtools and UnifiedGenotyper) on whole genome sequence data of 65 key ancestors of Swiss dairy cattle populations. Accuracy of calling next-generation sequence variants was assessed by comparison to the same loci from medium and high-density single nucleotide variant (SNV) arrays.

Results

The total number of SNVs identified varied by software and method, with single (multi) sample results ranging from 17.7 to 22.0 (16.9 to 22.0) million variants. Computing time varied considerably between software. Preparatory realignment of insertions and deletions and subsequent base quality score recalibration had only minor effects on the number and quality of SNVs identified by different software, but increased computing time considerably. Average concordance for single (multi) sample results with high-density chip data was 58.3% (87.0%) and average genotype concordance in correctly identified SNVs was 99.2% (99.2%) across software. The average quality of SNVs identified, measured as the ratio of transitions to transversions, was higher using single sample methods than multi sample methods. A consensus approach using results of different software generally provided the highest variant quality in terms of transition/transversion ratio.

Conclusions

Our findings serve as a reference for variant identification pipeline development in non-human organisms and help assess the implication of preparatory steps in next-generation sequencing pipelines for organisms with incomplete reference genomes (pipeline code is included). Benchmarking this information should prove particularly useful in processing next-generation sequencing data for use in genome-wide association studies and genomic selection.

Electronic supplementary material

The online version of this article (doi:10.1186/1471-2164-15-948) contains supplementary material, which is available to authorized users.     

Functional claudication distance: a reliable and valid measurement to assess functional limitation in patients with intermittent claudication

Background

Pharmacological inhibition of endothelial arginase-II has been shown to improve endothelial nitric oxide synthase (eNOS) function and reduce atherogenesis in animal models. We investigated whether the endothelial arginase II is involved in inflammatory responses in endothelial cells.

Methods

Human endothelial cells were isolated from umbilical veins and stimulated with TNFα (10 ng/ml) for 4 hours. Endothelial expression of the inflammatory molecules i.e. vascular cell adhesion molecule-1 (VCAM-1), intercellular adhesion molecule-1 (ICAM-1), and E-selectin were assessed by immunoblotting.

Results

The induction of the expression of endothelial VCAM-1, ICAM-1 and E-selectin by TNFα was concentration-dependently reduced by incubation of the endothelial cells with the arginase inhibitor L-norvaline. However, inhibition of arginase by another arginase inhibitor S-(2-boronoethyl)-L-cysteine (BEC) had no effects. To confirm the role of arginase-II (the prominent isoform expressed in HUVECs) in the inflammatory responses, adenoviral mediated siRNA silencing of arginase-II knocked down the arginase II protein level, but did not inhibit the up-regulation of the adhesion molecules. Moreover, the inhibitory effect of L-norvaline was not reversed by the NOS inhibitor L-NAME and L-norvaline did not interfere with TNFα-induced activation of NF-κB, JNK, p38mapk, while it inhibited p70s6k (S6K1) activity. Silencing S6K1 prevented up-regulation of E-selectin, but not that of VCAM-1 or ICAM-1 induced by TNFα.

Conclusion

The arginase inhibitor L-norvaline exhibits anti-inflammatory effects independently of inhibition of arginase in human endothelial cells. The anti-inflammatory properties of L-norvaline are partially attributable to its ability to inhibit S6K1.     

Inhibition of thirst when dehydrated rats drink water or saline

The present experiments sought to identify the physiological signals that inhibit thirst when dehydrated rats drink water or NaCl solution. Rats were deprived of drinking fluid but not food overnight. When allowed to drink again, the dehydrated animals consumed water or saline (0.05 M, 0.10 M, 0.15 M, or 0.20 M NaCl solution) almost continuously for 5-8 min before stopping. The volumes consumed were similar regardless of which fluid they ingested, but blood analyses indicated that increased plasma osmolality and decreased plasma volume, or both, still remained when drinking terminated. These results suggest that the composition of the ingested fluid is less significant than its volume in providing an early signal that inhibits thirst and fluid consumption by dehydrated rats. Analyses of the gastrointestinal tracts revealed that the cumulative volume in the stomach and small intestine correlated highly with the amount consumed regardless of which fluid was ingested. These and other results suggest that the volume of fluid ingested by dehydrated rats is sensed by stretch receptors detecting distension of the stomach and small intestine, which provide an early inhibitory stimulus of thirst.     

Structural reorganizations of the endoplasmic reticulum during egg maturation and fertilization

The endoplasmic reticulum (ER) of eggs is a major internal store of calcium ions that must be properly mobilized at fertilization for development to proceed. In most species, the ER develops distinct clusters in the cortical ooplasm as the oocyte matures into a fertilizable egg. Following fertilization, the structure of the ER rapidly reorganizes in eggs that produce a single fertilization-induced calcium wave, whereas ER clusters persist for relatively long periods in eggs that generate multiple calcium oscillations. This review considers such pre- and post-fertilization reorganizations of the ER and what effects these changes might have on calcium signaling patterns.     

Effects of myostatin deletion in aging mice

Inhibitors of myostatin, a negative regulator of skeletal muscle mass, are being developed to mitigate aging-related muscle loss. Knock-out (KO) mouse studies suggest myostatin also affects adiposity, glucose handling and cardiac growth. However, the cardiac consequences of inhibiting myostatin remain unclear. Myostatin inhibition can potentiate cardiac growth in specific settings ( Morissette et al., 2006) , a concern because of cardiac hypertrophy is associated with adverse clinical outcomes. Therefore, we examined the systemic and cardiac effects of myostatin deletion in aged mice (27–30 months old). Heart mass increased comparably in both wild-type (WT) and KO mice. Aged KO mice maintained twice as much quadriceps mass as aged WT; however, both groups lost the same percentage (36%) of adult muscle mass. Dual-energy X-ray absorptiometry revealed increased bone density, mineral content, and area in aged KO vs. aged WT mice. Serum insulin and glucose levels were lower in KO mice. Echocardiography showed preserved cardiac function with better fractional shortening (58.1% vs. 49.4%, P  = 0.002) and smaller left ventricular diastolic diameters (3.41 vs. 2.71, P  = 0.012) in KO vs. WT mice. Phospholamban phosphorylation was increased 3.3-fold in KO hearts ( P  < 0.05), without changes in total phospholamban, sarco(endo)plasmic reticulum calcium ATPase 2a or calsequestrin. Aged KO hearts showed less fibrosis by Masson's Trichrome staining. Thus, myostatin deletion does not affect aging-related increases in cardiac mass and appears beneficial for bone density, insulin sensitivity and heart function in senescent mice. These results suggest that clinical interventions designed to inhibit skeletal muscle mass loss with aging could have beneficial effects on other organ systems as well.     

Cytokine profile of human adipose-derived stem cells: expression of angiogenic, hematopoietic, and pro-inflammatory factors

Adipose tissue serves as a source of adipokines and cytokines with both local and systemic actions in health and disease. In this study, we examine the hypothesis that multipotent human adipose-derived stem cells (ASCs), capable of differentiating along the adipocyte, chondrocyte, and osteoblast pathways, contribute to adipose tissue-derived cytokine secretion. Following exposure to basic fibroblast growth factor (bFGF) or epidermal growth factor (EGF), the ASCs significantly increase their secretion of hepatocyte growth factor (HGF), a cytokine implicated in hematopoiesis, vasculogenesis, and mammary epithelial duct formation. Ascorbic acid synergizes with these inductive factors, further increasing HGF levels. Following exposure to lipopolysaccharide, ASCs increase their secretion of both hematopoietic (granulocyte/monocyte, granulocyte, and macrophage colony stimulating factors, interleukin 7) and proinflammatory (interleukins 6, 8, and 11, tumor necrosis factor alpha) cytokines based on ELISA and RT-PCR. In co-cultures established with umbilical cord blood-derived CD34(+) cells, the ASCs support long-term hematopoiesis in vitro. Furthermore, in short-term 12-day co-cultures, the ASC maintain and expand the numbers of both myeloid and lymphoid progenitors. These observations are consistent with the functionality of the secreted cytokines and confirm recent reports by other laboratories concerning the hematopoietic supportive capability of ASCs. We conclude that the ASCs display cytokine secretory properties similar to those reported for bone marrow-derived mesenchymal stem cells (MSCs).     

Confocal microscopy of fertilization-induced calcium dynamics in sea urchin eggs.

Although confocal microscopy has typically been utilized in studies of fixed specimens, its potential for exploring dynamic processes in living cells is rapidly being realized. In this report, confocal laser scanning microscopy is used to analyze the calcium wave that occurs following fertilization in living sea urchin eggs microinjected with the calcium-sensitive fluorescent probes fluo-3 or calcium green. Time-lapse recordings of optical sections depicting calcium dynamics within the eggs are also subjected to volumetric reconstructions. Such analyses indicate that (1) cytoplasmic free calcium levels become elevated throughout the fertilized egg, (2) fertilization also causes the egg nucleus to undergo a transient increase in free calcium, and (3) normal cleavage can be obtained following time-lapse imaging of the calcium waves.