Effect of lung-protective ventilation on severe Pseudomonas aeruginosa pneumonia and sepsis in rats

Pneumonia caused by Pseudomonas aeruginosa carries a high rate of morbidity and mortality. A lung-protective strategy using low tidal volume (V(T)) ventilation for acute lung injury improves patient outcomes. The goal of this study was to determine whether low V(T) ventilation has similar utility in severe P. aeruginosa infection. A cytotoxic P. aeruginosa strain, PA103, was instilled into the left lung of rats anesthetized with pentobarbital. The lung-protective effect of low V(T) (6 ml/kg) with or without high positive end-expiratory pressure (PEEP, 10 or 3 cmH(2)O) was then compared with high V(T) with low PEEP ventilation (V(T) 12 ml/kg, PEEP 3 cmH(2)O). Severe lung injury and septic shock was induced. Although ventilatory mode had little effect on the involved lung or septic physiology, injury to noninvolved regions was attenuated by low V(T) ventilation as indicated by the wet-to-dry weight ratio (W/D; 6.13 +/- 0.78 vs. 3.78 +/- 0.26, respectively) and confirmed by histopathological examinations. High PEEP did not yield a significant protective effect (W/D, 4.03 +/- 0.32) but, rather, caused overdistension of noninvolved lungs. Bronchoalveolar lavage revealed higher concentrations of TNF-alpha in the fluid of noninvolved lung undergoing high V(T) ventilation compared with those animals receiving low V(T). We conclude that low V(T) ventilation is protective in noninvolved regions and that the application of high PEEP attenuated the beneficial effects of low V(T) ventilation, at least short term. Furthermore, low V(T) ventilation cannot protect the involved lung, and high PEEP did not significantly alter lung injury over a short time course.     

Configuration effects on texture transparency

This study examined the factors producing the perception of transparency between overlaid regions composed of Gabor micro-patterns as functions of their spatial frequency, separation of overlaid regions, and types of orientation modulation. The results showed that the likelihood of perceiving transparency was high both when (1) the difference in Gabor spatial frequency between regions was large, and (2) the region boundary, which was formed by short-range orientation differences in the Gabor micro-patterns, clearly emerged. We conclude that texture transparency appears to result from an interaction between a boundary-detection mechanism defining the shape of each region and a surface-detection mechanism assigning the boundary.     

Multiple points of interaction between retinoic acid and FGF signaling during embryonic axis formation

Anteroposterior (AP) patterning of the developing CNS is crucial for both regional specification and the timing of neurogenesis. Several important factors are involved in AP patterning, including members of the WNT and FGF growth factor families, retinoic acid receptors, and HOX genes. We have examined the interactions between FGF and retinoic signaling pathways. Blockade of FGF signaling downregulates the expression of members of the RAR signaling pathway, RARalpha, RALDH2 and CYP26. Overexpression of a constitutively active RARalpha2 rescues the effects of FGF blockade on the expression of XCAD3 and HOXB9. This suggests that RARalpha2 is required as a downstream target of FGF signaling for the posterior expression of XCAD3 and HOXB9. Surprisingly, we found that posterior expression of FGFR1 and FGFR4 was dependent on the expression of RARalpha2. Anterior expression was also altered with FGFR1 expression being lost, whereas FGFR4 expression was expanded beyond its normal expression domain. RARalpha2 is required for the expression of XCAD3 and HOXB9, and for the ability of XCAD3 to induce HOXB9 expression. We conclude that RARalpha2 is required at multiple points in the posteriorization pathway, suggesting that correct AP neural patterning depends on a series of mutually interactive feedback loops among FGFs, RARs and HOX genes.     

Control of the ciliary beat by cyclic nucleotides in intact cortical sheets from Paramecium

The locomotor behavior of Paramecium depends on the ciliary beat direction and beat frequency. Changes in the ciliary beat are controlled by a signal transduction mechanism that follows changes in the membrane potential. These events take place in cilia covered with a ciliary membrane. To determine the effects of second messengers in the cilia, cortical sheets were used with intact ciliary membrane as a half-closed system in which each cilium is covered with a ciliary membrane with an opening to the cell body. Cyclic nucleotides and their derivatives applied from an opening to the cell body affected the ciliary beat. cAMP and 8-Br-cAMP increased the beat frequency and the efficiency of propulsion and acted antagonistically to the action of Ca(2+). cGMP and 8-Br-cGMP increased the efficiency of propulsion accompanying clear metachronal waves but decreased the beat frequency. These results indicate that the cyclic nucleotides affect target proteins in the ciliary axonemes surrounded by the ciliary membrane without a membrane potential and increase the efficiency of propulsion of the ciliary beat. In vitro phosphorylation of isolated ciliary axonemes in the presence of cyclic nucleotides and their derivatives revealed that the action of cAMP was correlated with the phosphorylation of 29-kDa and 65-kDa proteins and that the action of cGMP was correlated with the phosphorylation of a 42-kDa protein.     

Inhibitory NK receptor Ly49Q is expressed on subsets of dendritic cells in a cellular maturation- and cytokine stimulation-dependent manner

Ly49Q is a member of the Ly49 family that is expressed on Gr-1+ cells but not on NK and NKT cells. Ly49Q appears to be involved in regulating cytoskeletal architectures through ITIM-mediated signaling. We provide evidence that dendritic cells (DCs) of certain maturational states expressed Ly49Q, and that IFN-alpha plays an important role in its regulation. Freshly prepared murine plasmacytoid pre-DCs as well as Flt3L-induced plasmacytoid pre-DCs expressed Ly49Q, whereas freshly prepared myeloid DCs did not. However, GM-CSF-induced myeloid DCs showed low levels of Ly49Q expression, and this was significantly enhanced by IFN-alpha. In contrast, other cytokines and ligands for TLRs such as TNF-alpha, IL-6, LPS, and CpG-ODN had little or no effect on Ly49Q expression. Plasmacytoid pre-DCs in all mouse strains examined expressed Ly49Q. Constitutive expression of Ly49Q on myeloid DCs was observed in three restricted mouse strains including 129, NZB, and NZW. As can be seen in other Ly49 family members, Ly49Q expression was affected by MHC class I expression. At the same time, Ly49Q possessed polymorphisms, including at least three alleles. The polymorphic residues lay within the stalk and carbohydrate recognition domain, and two of them, in loop 3 and loop 6 of the carbohydrate recognition domain, are located in the region implicated in the interaction of Ly49A with H-2D(d). Therefore, depending on IFN-alpha, our results imply that Ly49Q serves a role for the biological functions of certain DC subsets through recognition of MHC class I or related molecules.     

Vitellogenin-immunohistochemistry in the liver and the testis of the medaka, Oryzias latipes, exposed to 17beta-estradiol and p-nonylphenol

Vitellogenin (VTG) produced in male fish has been used for a biomarker to study endocrine disrupters. However, the characteristics of VTG produced in male fish have not been studied well. In this study, we investigated the localization of VTG in the liver and the testis of male medaka (Oryzias latipes) treated with 17beta-estradiol (E2) and p-nonylphenol (NP). The male fish were exposed to 1 microg/L E2 and 500 microg/L NP for 1-12 days. Control groups were kept in water including only vehicle. The frozen sections of the liver and the testis were stained with immunohistochemical methods using an antiserum against medaka VTG as the first antibody. In the E2 and NP treated liver, the hepatocytes showed immunoreactivity. In particular, the cytoplasm close to the cell membrane surrounding the sinusoids was strongly immunopositive. In the testis of both treatments, the interstitial tissues and the cells (spermatocytes) in the seminiferous tubules were immunopositive. The concentration of VTG became gradually higher in both tissues with longer treatments. These results suggest that germ cells in the testis treated with E2 and NP are able to incorporate and accumulate VTG.     

Empirical Bayes estimation of gene-specific effects in micro-array research

Micro-array technology allows investigators the opportunity to measure expression levels of thousands of genes simultaneously. However, investigators are also faced with the challenge of simultaneous estimation of gene expression differences for thousands of genes with very small sample sizes. Traditional estimators of differences between treatment means (ordinary least squares estimators or OLS) are not the best estimators if interest is in estimation of gene expression differences for an ensemble of genes. In the case that gene expression differences are regarded as exchangeable samples from a common population, estimators are available that result in much smaller average mean-square error across the population of gene expression difference estimates. We have simulated the application of such an estimator, namely an empirical Bayes (EB) estimator of random effects in a hierarchical linear model (normal-normal). Simulation results revealed mean-square error as low as 0.05 times the mean-square error of OLS estimators (i.e., the difference between treatment means). We applied the analysis to an example dataset as a demonstration of the shrinkage of EB estimators and of the reduction in mean-square error, i.e., increase in precision, associated with EB estimators in this analysis. The method described here is available in software that is available at .     

Benzimidazole derivatives as novel nonpeptide luteinizing hormone-releasing hormone (LHRH) antagonists. Part 2: Benzimidazole-5-sulfonamides

The 2-cyclopropyl substituted benzimidazole 2 has been used as a starting point for further optimization of an LHRH antagonist series. SAR studies revealed that a tert-butyl urea fragment connected through a simple carbon chain would improve activity. Further modification of the benzylsulfonamide moiety led to the discovery of 23 (IC(50): 4.2 nM).     

Potential involvement of oxidative stress in cartilage senescence and development of osteoarthritis: oxidative stress induces chondrocyte telomere instability and downregulation of chondrocyte function

Oxidative stress leads to increased risk for osteoarthritis (OA) but the precise mechanism remains unclear. We undertook this study to clarify the impact of oxidative stress on the progression of OA from the viewpoint of oxygen free radical induced genomic instability, including telomere instability and resulting replicative senescence and dysfunction in human chondrocytes. Human chondrocytes and articular cartilage explants were isolated from knee joints of patients undergoing arthroplastic knee surgery for OA. Oxidative damage and antioxidative capacity in OA cartilage were investigated in donor-matched pairs of intact and degenerated regions of tissue isolated from the same cartilage explants. The results were histologically confirmed by immunohistochemistry for nitrotyrosine, which is considered to be a maker of oxidative damage. Under treatment with reactive oxygen species (ROS; 0.1 μmol/l H₂O₂) or an antioxidative agent (ascorbic acid: 100.0 μmol/l), cellular replicative potential, telomere instability and production of glycosaminoglycan (GAG) were assessed in cultured chondrocytes. In tissue cultures of articular cartilage explants, the presence of oxidative damage, chondrocyte telomere length and loss of GAG to the medium were analyzed in the presence or absence of ROS or ascorbic acid. Lower antioxidative capacity and stronger staining of nitrotyrosine were observed in the degenerating regions of OA cartilages as compared with the intact regions from same explants. Immunostaining for nitrotyrosine correlated with the severity of histological changes to OA cartilage, suggesting a correlation between oxidative damage and articular cartilage degeneration. During continuous culture of chondrocytes, telomere length, replicative capacity and GAG production were decreased by treatment with ROS. In contrast, treatment with an antioxidative agent resulted in a tendency to elongate telomere length and replicative lifespan in cultured chondrocytes. In tissue cultures of cartilage explants, nitrotyrosine staining, chondrocyte telomere length and GAG remaining in the cartilage tissue were lower in ROS-treated cartilages than in control groups, whereas the antioxidative agent treated group exhibited a tendency to maintain the chondrocyte telomere length and proteoglycan remaining in the cartilage explants, suggesting that oxidative stress induces chondrocyte telomere instability and catabolic changes in cartilage matrix structure and composition. Our findings clearly show that the presence of oxidative stress induces telomere genomic instability, replicative senescence and dysfunction of chondrocytes in OA cartilage, suggesting that oxidative stress, leading to chondrocyte senescence and cartilage ageing, might be responsible for the development of OA. New efforts to prevent the development and progression of OA may include strategies and interventions aimed at reducing oxidative damage in articular cartilage.