The nematode Pristionchus pacificus (Nematoda: Diplogastridae) is associated with the oriental beetle Exomala orientalis (Coleoptera: Scarabaeidae) in Japan

Pristionchus pacificus has been developed as a nematode satellite organism in evolutionary developmental biology. Detailed studies of vulva development revealed multiple differences in genetic and molecular control in P. pacificus compared to the model organism Caenorhabditis elegans. To place evolutionary developmental biology in a comprehensive evolutionary context, such studies have to be complemented with ecology. In recent field studies in western Europe and eastern North America we found 11 Pristionchus species that are closely associated with scarab beetles and the Colorado potato beetle. However, P. pacificus was not commonly found in association with scarab beetles in these studies. Here, we describe the results of a similar survey of scarab beetles in Japan. Pristionchus pacificus was the most common Pristionchus species on scarab beetles in Japan, with 40 out of 43 (93%) isolates. The other Pristionchus isolates represent three novel species, which we refer to as Pristionchus sp. 11, Pristionchus sp. 14, and Pristionchus sp. 15. Thirty-seven of the established P. pacificus strains were found on the oriental beetle Exomala orientalis. Laboratory studies with the sex pheromone (Z)-7-tetradecen-2-one of the oriental beetle revealed that P. pacificus shows strong olfactory attraction to the beetle's sex pheromone, which provides a potential mechanism for the recognition and interaction of P. pacificus and E. orientalis. Together, this study identifies P. pacificus as the most common Pristionchus nematode in field studies in Japan, identifies E. orientalis as an important host species, and provides the basis for the ecology of P. pacificus.     

ROK contribution to endothelin-mediated contraction in aorta and mesenteric arteries following intermittent hypoxia/hypercapnia in rats

We reported previously that intermittent hypoxia with CO(2) to maintain eucapnia (IH-C) elevates plasma endothelin-1 (ET-1) and arterial pressure. In small mesenteric arteries (sMA; inner diameter = 150 microm), IH-C augments ET-1 constrictor sensitivity but diminishes ET-1-induced increases in intracellular Ca(2+) concentration, suggesting IH-C exposure increases both ET-1 levels and ET-1-stimulated Ca(2+) sensitization. Because Rho-associated kinase (ROK) can mediate Ca(2+) sensitization, we hypothesized that augmented vasoconstrictor sensitivity to ET-1 in arteries from IH-C-exposed rats is dependent on ROK activation. In thoracic aortic rings, ET-1 contraction was not different between groups, but ROK inhibition (Y-27632, 3 and 10 microM) attenuated ET-1 contraction more in IH-C than in sham arteries (50 +/- 11 and 78 +/- 7% vs. 41 +/- 12 and 48 +/- 9% inhibition, respectively). Therefore, ROK appears to contribute more to ET-1 contraction in IH-C than in sham aorta. In sMA, ROK inhibitors did not affect ET-1-mediated constriction in sham arteries and only modestly inhibited it in IH-C arteries. In ionomycin-permeabilized sMA with intracellular Ca(2+) concentration held at basal levels, Y-27632 did not affect ET-1-mediated constriction in either IH-C or sham sMA and ET-1 did not stimulate ROK translocation. In contrast, inhibition of myosin light-chain kinase (ML-9, 100 microM) prevented ET-1-mediated constriction in sMA from both groups. Therefore, IH-C exposure increases ET-1 vasoconstrictor sensitivity in sMA but not in aorta. Furthermore, ET-1 constriction is myosin light-chain kinase dependent and mediated by Ca(2+) sensitization that is independent of ROK activation in sMA but not aorta. Thus ET-1-mediated signaling in aorta and sMA is altered by IH-C but is dependent on different second messenger systems in small vs. large arteries.     

TRAV gene usage in pig T-cell receptor alpha cDNA

Pig (Sus scrofa) TRA clones were isolated from cDNA libraries of total RNA from two different sources, the thymus of a 1-month-old LW strain pig and the peripheral blood lymphocytes of a 5-month-old Clawn strain pig. Among 103 complete TRA cDNA clones from both sources, 33 different TRAV genes were identified. By comparing their sequence identities against one another, these pig TRAV genes were grouped into 20 subgroups, including 13 subgroups, each containing only a single member. All of these pig subgroups gave corresponding human and mouse functional counterparts, suggesting their functional commonality. An exception was the Va01 gene segment, which lacked a functional human counterpart. The present report provides groundwork for studies on pig TRA expression.Electronic Supplementary Material Supplementary material is available for this article at .This report constitutes a part of the requirement for the doctorate thesis for R.Y. in Tohoku University.     

Direct tandem mass spectrometry reveals limitations in protein profiling experiments for plasma biomarker discovery

The low molecular weight plasma proteome and its biological relevance are not well defined; therefore, experiments were conducted to directly sequence and identify peptides observed in plasma and serum protein profiles. Protein fractionation, matrix-assisted laser desorption ionization mass spectrometry (MALDI-MS) profiling, and liquid-chromatography coupled to MALDI tandem mass spectrometry (MS/MS) sequencing were used to analyze the low molecular weight proteome of heparinized plasma. Four fractionation techniques using functionally derivatized 96-well plates were used to extract peptides from plasma. Tandem TOF was successful for identifying peptides up to m/z 5500 with no prior knowledge of the sequence and was also used to verify the sequence assignments for larger ion signals. The peptides (n>250) sequenced in these profiles came from a surprisingly small number of proteins (n approximately 20), which were all common to plasma, including fibrinogen, complement components, antiproteases, and carrier proteins. The cleavage patterns were consistent with those of known plasma proteases, including initial cleavages by thrombin, plasmin and complement proteins, followed by aminopeptidase and carboxypeptidase activity. On the basis of these data, we discuss limitations in biomarker discovery in the low molecular weight plasma or serum proteome using crude fractionation coupled to MALDI-MS profiling.     

Thymosin beta10 inhibits cell migration and capillary-like tube formation of human coronary artery endothelial cells

Thymosin beta10 is a cytoplasm G-actin sequestering protein whose functions are largely unknown. To determine the direct effects of exogenous thymosin beta10 on angiogenic potentials as endothelial cell migration and capillary-like tube formation, human coronary artery endothelial cells (HCAECs) were incubated with increasing doses of thymosin beta10 (25-100 ng/ml). By using a modified Boyden chamber assay, thymosin beta10 inhibited cell migration in a dose- and time-dependent manner with the maximal effect being a 36% reduction at 100 ng/ml as compared to controls (P < 0.01). In addition, thymosin beta10 (100 ng/ml) significantly inhibited the capillary-like tube-formation of HCAECs on Matrigel, showing a 21% reduction of the total tube length as compared to negative controls (P < 0.01). Furthermore, by using real time PCR analysis, thymosin beta10 significantly decreased mRNA levels of vascular endothelial growth factor (VEGF), VEGF receptor-1 (VEGFR-1) and integrin alphaV after 24 h treatment in HCAECs. By contrast, thymosin beta4 significantly increased HCAEC migration. These results indicate that thymosin beta10, but not thymosin beta4, have direct inhibitive effects on endothelial migration and tube formation that might be mediated via downregulation of VEGF, VEGFR-1 and integrin alphaV in HCAECs. This study suggests a potential therapeutic application of thymosin beta10 to the diseases with excessive angiogenesis such as cancer.     

Screening of an α-amylase inhibitor peptide by photolinker-peptide array

Peptide arrays in which peptides were immobilized on cellulose membranes through photolinkers were synthesized. The peptides were subsequently detached from the arrays by ultraviolet (UV) photolysis for 3 h, and were used to search for functional peptides that inhibit the activity of α-amylase derived from human pancreatic juice. Amino acid replacement with high-molecular-size amino acids, Arg (R), Phe (F), Trp (W), or Tyr (Y), for the first and seventh residues of amylase inhibitor peptide, GHWYYRCW, as previous reported, led to enhancement of the inhibitory effect of the peptide on α-amylase. In particular, one of the resulting peptides, RHWYYRYW, showed a stronger inhibitory effect than acarbose (which is used as a hypoglycemic agent) or inhibitor peptide GHWYYRCW.     

Species diversity in native fish community in Japan: comparison between non-invaded and invaded ponds by exotic fish

The relationship between invasions by two exotic fishes (Micropterus salmoides and Lepomis macrochirus) and species diversity in native fish communities was studied in 14 Japanese farm ponds. We found that mean number of species in native fish communities was three times higher in the ponds without the exotic fish than in the ponds with them. Further, negative relationships were observed between abundance of the two exotic fish and the total abundance of native fish communities. Our results suggest that invasions by the two exotic fish caused serious depletion of native fish communities, although another process can also be considered , that is, that ponds with poor native fish communities were prone to colonization by these exotic fish.