Inhibitory Effect of Glycerin on Vibrio parahaemolyticus and Salmonella

In a study of the effect of glycerin in transport media on Vibrio parahaemolyticus and Salmonella, it was found that a concentration of 30% glycerin was highly inhibitory for V. parahaemolyticus and to a lesser degree for Salmonella. The incorporation of peptone or human feces in media did not reduce the inhibitory effect of glycerin. In media with 15% glycerin, viable counts of V. parahaemolyticus and Salmonella increased after 24 hr of incubation both in the presence and absence of feces. Due to the concurrent increase in the total bacterial count in the media containing feces, no enrichment effect was noted.     

Direct Evidence for Changes in the Resistance of Legume Root Nodules to O2 Diffusion

Indirect evidence suggests that legumes can adjust rapidly theresistance of their root nodules to O₂ diffusion. Here we describeexperiments using O₂ specific micro-electrodes and dark fieldmicroscopy to study directly the operation of this diffusionbarrier. The O₂ concentration sensed by the electrode decreasedsharply in the region of the inner cortex and was less than1.0 mmol m^–3 throughout the infected tissue in nodulesof both pea (Pisum sativum) and french bean (Phaseolus vulgaris).In a number of experiments the ambient O₂ concentration wasincreased to 40% while the electrode tip was just inside theinner cortex. In 13 out of 21 cases the O₂ concentration atthis position either remained low and unchanged or increasedirreversibly to near ambient values. In the remaining casesthe O₂ concentration increased after 1 to 2.5 min and then decreasedto its former value. These results are ascribed to an increasein resistance of the barrier in response to increased O₂ fluxinto the nodule. It was shown microscopically that air spacesboth at the boundary between the infected zone and the innercortex, and within the infected zone started to disappear 3min after nodules were exposed to high ambient O₂ concentrationsand had disappeared completely after 8 min. These spaces werenot changed by exposure of the nodule for 10 min to either N₂or air. Key words: Oxygen, root nodules, air spaces     

Evolution of the 28S ribosomal RNA gene in anurans: regions of variability and their phylogenetic implications

Fifteen restriction sites were mapped to the 28S ribosomal RNA gene of individuals representing 54 species of frogs, two species of salamanders, a caecilian, and a lungfish. Eight of these sites were present in all species examined, and two were found in all but one species. Alignment of these conserved restriction sites revealed, among anuran 28S rRNA genes, five regions of major length variation that correspond to four of 12 previously identified divergent domains of this gene. One of the divergent domains (DD8) consists of two regions of length variation separated by a short segment that is conserved at least throughout tetrapods. Most of the insertions, deletions, and restriction-site variations identified in the 28S gene will require sequence-level analysis for a detailed reconstruction of their history. However, an insertion in DD9 that is coextensive with frogs in the suborder Neobatrachia, a BstEII site that is limited to representatives of two leptodactylid subfamilies, and a deletion in DD10 that is found only in three ranoid genera are probably synapomorphies.      

Black soldier fly (Hermetia illucens) larvae oil as an alternative fat ingredient to soybean oil in laying hen diets

ObjectiveThe objective of this study was to determine whether dietary black soldier fly (Hermetia illucens, HI) larvae oil (HILO) could serve as an alternative fat source to soybean oil (SBO) in laying hen diets.MethodsWe randomly assigned 25-week-old Hy-line Brown laying hens (n = 144) to receive (n = 6 hens/group; eight replicates) a control or an experimental diet in which SBO was replaced with 50% (50HILO) or 100% HILO (100HILO).ResultsDietary HILO did not negatively affect body weight or productive performance during the study. The eggs also had similar quality parameters, proximate composition, and cholesterol levels. However, the yolk color index was significantly higher (p<0.01) in the 100HILO than in the other groups. Dietary HILO significantly altered the composition of fatty acids (FAs) in abdominal fat and eggs. Total saturated fatty acids (SFAs) and total polyunsaturated FAs (PUFAs) were significantly increased and decreased in the 50HILO and 100HILO groups, respectively, compared with those in the control group (p<0.001 and p<0.0001, respectively). Specifically, the medium-chain FAs lauric and myristic acids were remarkably increased in the abdominal fat of laying hens fed HILO (p<0.0001), whereas only myristic acid increased in eggs (p<0.0001). Undesirable heavy metal (aluminum, fluorine, arsenic, lead, mercury, and cadmium) concentrations were below permissible limits in eggs.ConclusionWe considered that HILO could be an alternative dietary fat to SBO for laying hens with maintained productive performance and good egg quality.     

Cell cycle arrest induced by the vitamin D(3) analog EB1089 in NCI-H929 myeloma cells is associated with induction of the cyclin-dependent kinase inhibitor p27

EB1089, a 1,25-dihydroxyvitamin D(3) analog, has been known to have potent antiproliferative properties in a variety of malignant cells in vitro and in vivo. In the present study, we analyzed the effect of EB1089 on human myeloma cell lines. EB1089 inhibited the proliferation of NCI-H929 cells and RPMI8226 cells in a dose-dependent manner among three myeloma cell lines tested. The antiproliferative effect of EB1089 on myeloma cells was related to the expression level of vitamin D receptor. To investigate the mechanism of the antiproliferative effect of EB1089, cell cycle analysis was attempted in EB1089-sensitive NCI-H929 cells. EB1089 (1 x 10(-8) M) efficiently induced G(1) arrest of the cell cycle. Analysis of G(1) regulatory proteins demonstrated that protein levels of CDK2, CDK4, cyclin D1, and cyclin A were decreased in a time-dependent manner, but not those of CDK6 and cyclin E, by EB1089. In addition, EB1089 (1 x 10(-8) M, 72 h) increased the protein level of the CDKI p27 and markedly enhanced the binding of p27 with CDK2 compared to EB1089-untreated cells. Furthermore, the activity of CDK2-associated cyclin kinase was decreased, which was accompanied by the reduction of cyclin-D1-, cyclin-E-, and cyclin-A-associated kinase activities, resulting in the hypophosphorylation of Rb protein. These results suggest that EB1089 can inhibit the proliferation of human myeloma cells, especially NCI-H929 cells, via a G(1) block in association with the induction of p27 and the reduction of CDK2 activity.     

Analytical validation of quantitative immunohistochemical assays of tumor infiltrating lymphocyte biomarkers

Tumor infiltrating lymphocytes (TIL), especially T-cells, have both prognostic and therapeutic applications. The presence of CD8+ effector T-cells and the ratio of CD8+ cells to FOXP3+ regulatory T-cells have been used as biomarkers of disease prognosis to predict response to various immunotherapies. Blocking the interaction between inhibitory receptors on T-cells and their ligands with therapeutic antibodies including atezolizumab, nivolumab, pembrolizumab and tremelimumab increases the immune response against cancer cells and has shown significant improvement in clinical benefits and survival in several different tumor types. The improved clinical outcome is presumed to be associated with a higher tumor infiltration; therefore, it is thought that more accurate methods for measuring the amount of TIL could assist prognosis and predict treatment response. We have developed and validated quantitative immunohistochemistry (IHC) assays for CD3, CD8 and FOXP3 for immunophenotyping T-lymphocytes in tumor tissue. Various types of formalin fixed, paraffin embedded (FFPE) tumor tissues were immunolabeled with anti-CD3, anti-CD8 and anti-FOXP3 antibodies using an IHC autostainer. The tumor area of stained tissues, including the invasive margin of the tumor, was scored by a pathologist (visual scoring) and by computer-based quantitative image analysis. Two image analysis scores were obtained for the staining of each biomarker: the percent positive cells in the tumor area and positive cells/mm² tumor area. Comparison of visual vs. image analysis scoring methods using regression analysis showed high correlation and indicated that quantitative image analysis can be used to score the number of positive cells in IHC stained slides. To demonstrate that the IHC assays produce consistent results in normal daily testing, we evaluated the specificity, sensitivity and reproducibility of the IHC assays using both visual and image analysis scoring methods. We found that CD3, CD8 and FOXP3 IHC assays met the fit-for-purpose analytical acceptance validation criteria and that they can be used to support clinical studies.     

Peripheral Afferent Mechanisms Underlying Acupuncture Inhibition of Cocaine Behavioral Effects in Rats

Administration of cocaine increases locomotor activity by enhancing dopamine transmission. To explore the peripheral mechanisms underlying acupuncture treatment for drug addiction, we developed a novel mechanical acupuncture instrument (MAI) for objective mechanical stimulation. The aim of this study was to evaluate whether acupuncture inhibition of cocaine-induced locomotor activity is mediated through specific peripheral nerves, the afferents from superficial or deep tissues, or specific groups of nerve fibers. Mechanical stimulation of acupuncture point HT7 with MAI suppressed cocaine-induced locomotor activity in a stimulus time-dependent manner, which was blocked by severing the ulnar nerve or by local anesthesia. Suppression of cocaine-induced locomotor activity was elicited after HT7 stimulation at frequencies of either 50 (for Meissner corpuscles) or 200 (for Pacinian corpuscles) Hz and was not affected by block of C/Aδ-fibers in the ulnar nerve with resiniferatoxin, nor generated by direct stimulation of C/Aδ-fiber afferents with capsaicin. These findings suggest that HT7 inhibition of cocaine-induced locomotor activity is mediated by A-fiber activation of ulnar nerve that originates in superficial and deep tissue.     

Anti-inflammatory effects of anethole in lipopolysaccharide-induced acute lung injury in mice

Aims

Anethole, the major component of the essential oil of star anise, has been reported to have antioxidant, antibacterial, antifungal, anti-inflammatory, and anesthetic properties. In this study, we investigated the anti-inflammatory effects of anethole in a mouse model of acute lung injury induced by lipopolysaccharide (LPS).

Main methods

BALB/C mice were intraperitoneally administered anethole (62.5, 125, 250, or 500 mg/kg) 1 h before intratracheal treatment with LPS (1.5 mg/kg) and sacrificed after 4 h. The anti-inflammatory effects of anethole were assessed by measuring total protein and cell levels and inflammatory mediator production and by histological evaluation and Western blot analysis.

Key findings

LPS significantly increased total protein levels; numbers of total cells, including macrophages and neutrophils; and the production of inflammatory mediators such as matrix metalloproteinase 9 (MMP-9), tumor necrosis factor-alpha (TNF-α), interleukin-6 (IL-6), and nitric oxide (NO) in bronchoalveolar lavage fluid. Anethole (250 mg/kg) decreased total protein concentrations; numbers of inflammatory cells, including neutrophils and macrophages; and the inflammatory mediators MMP-9, TNF-α and NO. In addition, pretreatment with anethole decreased LPS-induced histopathological changes. The anti-inflammatory mechanism of anethole in LPS-induced acute lung injury was assessed by investigating the effects of anethole on NF-κB activation. Anethole suppressed the activation of NF-κB by blocking IκB-α degradation.

Significance

These results, showing that anethole prevents LPS-induced acute lung inflammation in mice, suggest that anethole may be therapeutically effective in inflammatory conditions in humans.