Accumulation of hydroxyapatite in the kidney of streptozotocin-induced diabetic rat fed a low-zinc diet

Calcification occurred in the kidney of streptozotocin (STZ)-induced diabetic rats fed a low-zinc diet. The deposits were stained by the von Kossa method and were detected intracellularly in the tubular cells, mainly in the cortico-medullar region. The deposits were found to be a heterogenous substance on electron microscopy. There were various sizes of deposits, and the surfactant was very much distorted. Many deposits grew up to bind small particles, and the core-like substance was observed in the center of the deposit. The component of the deposit was analyzed by an X-ray microanalyzer, and was found to be calcium and phosphorus. The ratio of calcium to phosphorus was 2.159, which coincided with the ratio of standard hydroxyapatite. From these observations, the deposit is believed to be hydroxyapatite. It is thought that the core is formed at first, many particles are then bound to the core, and deposits grow up.     

GENETIC CORRELATIONS BETWEEN LIFE-HISTORY AND BEHAVIORAL TRAITS CAN CAUSE REPRODUCTIVE ISOLATION

Reproductive isolation may often evolve as an indirect (pleiotropic) consequence of populations adapting to different environments or habitats. For example, niches that are temporally or seasonally offset can select for organisms with different developmental characteristics. These developmental differences can inadvertently cause reproductive isolation by a variety of means including shifts in mating activity patterns. Here, we show a genetic correlation between a life-history trait (developmental period) and a behavioral trait (time of mating) that causes significant premating isolation in the melon fly, Bactrocera cucurbitae (Diptera: Tephritidae). Fly lines selected for short and long developmental periods differ in their preferred times of mating during the evening. This difference translates into significant prezygotic isolation, as measured by mate choice tests. If the time of mating between two populations differed more than one hour, the isolation index was significantly higher than zero. These indicate that premating isolation can be established if the developmental period is divergently selected for. If such genetic correlations are ubiquitous in many organisms, multifarious divergent selection for life-history traits would often accelerate the evolution of reproductive isolation. We speculate that reproductive isolation may have been evolved via genetic correlations among time-related traits, for example, developmental period and time of mating, as in other organisms.     

Regulation of Transforming Growth Factor-��-dependent Cyclooxygenase-2 Expression in Fibroblasts

Abnormal transforming growth factor-β (TGF-β) signaling is a critical contributor to the pathogenesis of various human diseases ranging from tissue fibrosis to tumor formation. Excessive TGF-β signaling stimulates fibrotic responses. Recent research has focused in the main on the antiproliferative effects of TGF-β in fibroblasts, and it is presently understood that TGF-β-stimulated cyclooxygenase-2 (COX-2) induction in fibroblasts is essential for antifibroproliferative effects of TGF-β. Both TGF-β and COX-2 have been implicated in tumor growth, invasion, and metastasis, and therefore tumor-associated fibroblasts are a recent topic of interest. Here we report the identification of positive and negative regulatory factors of COX-2 expression induced by TGF-β as determined using proteomic approaches. We show that TGF-β coordinately up-regulates three factors, heterogeneous nuclear ribonucleoprotein A/B (HNRPAB), nucleotide diphosphate kinase A (NDPK A), and nucleotide diphosphate kinase A (NDPK B). Functional pathway analysis showed that HNRPAB augments mRNA and protein levels of COX-2 and subsequent prostaglandin E₂ (PGE₂) production by suppressing degradation of COX-2 mRNA. In contrast, NDPK A and NDPK B attenuated mRNA and protein levels of COX-2 by affecting TGF-β-Smad2/3/4 signaling at the receptor level. Collectively, we report on a new regulatory pathway of TGF-β in controlling expression of COX-2 in fibroblasts, which advances our understanding of pathophysiological mechanisms of TGF-β.     

Influenza virus sequence feature variant type analysis: evidence of a role for NS1 in influenza virus host range restriction

Genetic drift of influenza virus genomic sequences occurs through the combined effects of sequence alterations introduced by a low-fidelity polymerase and the varying selective pressures experienced as the virus migrates through different host environments. While traditional phylogenetic analysis is useful in tracking the evolutionary heritage of these viruses, the specific genetic determinants that dictate important phenotypic characteristics are often difficult to discern within the complex genetic background arising through evolution. Here we describe a novel influenza virus sequence feature variant type (Flu-SFVT) approach, made available through the public Influenza Research Database resource (www.fludb.org), in which variant types (VTs) identified in defined influenza virus protein sequence features (SFs) are used for genotype-phenotype association studies. Since SFs have been defined for all influenza virus proteins based on known structural, functional, and immune epitope recognition properties, the Flu-SFVT approach allows the rapid identification of the molecular genetic determinants of important influenza virus characteristics and their connection to underlying biological functions. We demonstrate the use of the SFVT approach to obtain statistical evidence for effects of NS1 protein sequence variations in dictating influenza virus host range restriction.     

Alpha-hydroxyketones as inhibitors of urease

A variety of alpha-hydroxyketones (1-13) and alpha-diketones (14-20) were evaluated for their effect on the jack bean urease. Of 13 alpha-hydroxyketones (1-13) tested, 2,2'-thenoin (10) (IC(50)=0.18 mM), furoin (9) (IC(50)=0.36 mM), 2-hydroxy-1-phenylethanone (5) (IC(50)=0.47 mM) and acetol (1) (IC(50)=2.9 mM) showed potent inhibitory activity against the enzyme, comparable with hydroxyurea (IC(50)=0.1 mM). The inhibitory effects were completely blocked by 2-mercaptoethanol or dithiothreitol. A nickel ion influenced the inhibitory effects of 5 and 9 in a dose-dependent manner, but not of 1 and 10. On the other hand, the corresponding alpha-diketones such as 2,2'-thenil (20), furil (19) and PhCOCHO (14) exhibited little or no ability to inhibit the urease. We have demonstrated for the first time that some alpha-hydroxyketone derivatives show urease inhibitory activity, possibly by binding to cysteinyl residues in the active site.     

Highly functionalized 7-azaindoles as selective PPAR gamma modulators

A series of highly functionalized 3-aroyl and 3-phenoxy-2-methyl-7-azaindoles have been identified, which are potent selective PPARγ modulators (SPPARγMs). Addition of substituents at the 6-position of the 7-azaindoles improves in vitro potency and pharmacokinetics. 7-Azaindoles have significantly improved off-target profiles compared to the parent indole series.     

Neutral endopeptidase inhibitor suppresses the early phase of atrial electrical remodeling in a canine rapid atrial pacing model

Introduction

We examined the acute effects of neutral endopeptidase inhibitor on the hemodynamics and electrical properties of dogs subjected to rapid atrial pacing.

Methods

Ten beagle dogs were used and divided into two groups with and without candoxatril, a neutral endopeptidase inhibitor preadministration. Before and after the 6 hours rapid atrial pacing from the right atrial appendage, the hemodynamics, atrial effective refractory period, and monophasic action potential duration of the right atrial appendage were measured and blood samples were collected. Atrial tissue was also excised after the experiment.

Results

Candoxatril significantly increased plasma ANP levels (Control: 88.4 ± 50.25 vs. Candoxatril: 197.1 ± 32.09 pg/ml, p = 0.004) and prevented reductions in atrial effective refractory period and monophasic action potential duration. We further demonstrated that the treated animals exhibited significantly higher levels of atrial tissue cyclic GMP (Control: 28.1 ± 1.60 fmol/mg vs. Candoxatril: 44.5 ± 12.28 fmol/mg, p = 0.034) as well as that of plasma cyclic GMP (Control: 32 ± 5.5 vs. Candoxatril: 42 ± 7.1 pg/ml, p = 0.028).

Conclusion

Candoxatril suppressed the shortening of atrial effective refractory period and monophasic action potential duration in the rapid atrial pacing model. As plasma ANP and the atrial tissue levels of cyclic GMP were higher in the Candoxatril group than the control, this effect was considered to appear through the reduction of calcium overload caused by ANP and cyclic GMP.     

Regulation of platelet dense granule secretion by the Ral GTPase-exocyst pathway

Non-hydrolyzable GTP analogues, such as guanosine 5'-(beta, gamma-imido)triphosphate (GppNHp), induce granule secretion from permeabilized platelets in the absence of increased intracellular Ca(2+). Here, we show that the GppNHp-induced dense granule secretion from permeabilized platelets occurred concomitantly with the activation of small GTPase Ral. This secretion was inhibited by the addition of GTP-Ral-binding domain (RBD) of Sec5, which is a component of the exocyst complex known to function as a tethering factor at the plasma membrane for vesicles. We generated an antibody against Sec5-RBD, which abolished the interaction between GTP-Ral and the exocyst complex in vitro. The addition of this antibody inhibited the GppNHp-induced secretion. These data indicate that Ral mediates the GppNHp-induced dense granule secretion from permeabilized platelets through interaction with its effector, the exocyst complex. Furthermore, GppNHp enhanced the Ca(2+) sensitivity of dense granule secretion from permeabilized platelets, and this enhancement was inhibited by Sec5-RBD. In intact platelets, the association between Ral and the exocyst complex was induced by thrombin stimulation with a time course similar to that of dense granule secretion and Ral activation. Taken together, our results suggest that the Ral-exocyst pathway participates in the regulation of platelet dense granule secretion by enhancing the Ca(2+) sensitivity of the secretion.     

Uptake of dipeptide and beta-lactam antibiotics by the basolateral membrane vesicles prepared from rat kidney

The transport of dipeptides and beta-lactam antibiotics across the rat renal basolateral membrane was examined. The initial uptake of glycylsarcosine and cefadroxil by rat renal basolateral membrane vesicles was inhibited by the presence of all the di- and tripeptides and beta-lactam antibiotics that were tested in this study. However, the uptake of both substrates was not inhibited by glycine, an amino acid. The initial uptake of zwitterionic beta-lactam antibiotics, cefadroxil, cephradine, and cephalexin, was stimulated by preloaded glycylsarcosine (countertransport effect). On the other hand, the uptake of dianionic beta-lactam antibiotics, ceftibuten and cefixime, was not affected. A concentration-dependent initial uptake of glycylsarcosine and cefadroxil suggested the existence of a carrier-mediated mechanism, whereas the transport of ceftibuten did not show any saturated uptake. The transporter that participates in the permeation of dipeptides and beta-lactam antibiotics across basolateral membranes showed lower affinity than did PEPT1 and PEPT2. This is the first study that showed an evidence for a peptide transporter, expressed in the rat renal basolateral membrane, that recognizes zwitterionic beta-lactam antibiotics using basolateral membrane vesicles isolated from normal rat kidney.     

Bidirectional ephrinB2-EphB4 signaling controls bone homeostasis

Bone homeostasis requires a delicate balance between the activities of bone-resorbing osteoclasts and bone-forming osteoblasts. Various molecules coordinate osteoclast function with that of osteoblasts; however, molecules that mediate osteoclast-osteoblast interactions by simultaneous signal transduction in both cell types have not yet been identified. Here we show that osteoclasts express the NFATc1 target gene Efnb2 (encoding ephrinB2), while osteoblasts express the receptor EphB4, along with other ephrin-Eph family members. Using gain- and loss-of-function experiments, we demonstrate that reverse signaling through ephrinB2 into osteoclast precursors suppresses osteoclast differentiation by inhibiting the osteoclastogenic c-Fos-NFATc1 cascade. In addition, forward signaling through EphB4 into osteoblasts enhances osteogenic differentiation, and overexpression of EphB4 in osteoblasts increases bone mass in transgenic mice. These data demonstrate that ephrin-Eph bidirectional signaling links two major molecular mechanisms for cell differentiation--one in osteoclasts and the other in osteoblasts--thereby maintaining bone homeostasis.