Endothelin: a new inhibitor of renin release

Endothelin is a recently-discovered vasoconstrictor peptide which is produced by endothelium and acts on vascular smooth muscle cells. At present its actions on other organs or cells are unknown. We studied the effect of endothelin on renin release in a dynamic superfusion system of dispersed rat juxtaglomerular (JG) cells. Endothelin in concentrations of 10(-11) M or more inhibited renin release dose-dependently and this inhibitory action vanished in the absence of extracellular Ca. It is suggested that endothelin is an inhibitory regulator of renin secretion from JG cells and its action is Ca-dependent.     

Magnesium ion: a possible physiological regulator of aldosterone production

We examined the direct effect of magnesium ion on aldosterone production by adrenal cells using collagenase-dispersed zona-glomerulosa cells in rats. The effects of magnesium on aldosterone production stimulated by angiotensin II or ACTH were also investigated. Both magnesium sulphate (MgSO4) and magnesium chloride (MgCl2) (0 to 2 mM) decreased aldosterone production in a dose-dependent manner. In comparison with magnesium-free medium, 2 mM MgSO4 inhibited aldosterone production by 73% and MgCl2 by 65%. In addition, MgSO4 showed an inhibitory effect on aldosterone production stimulated by angiotensin II (10pM to 10nM), whereas it had no significant effect on aldosterone production due to ACTH stimulation (10pM to 10nM). These data suggest that magnesium has an inhibitory action on aldosterone production in vitro and may be a physiological regulator of aldosterone production.     

Distribution of antimicrobial‐resistant lactic acid bacteria in natural cheese in Japan

To determine and compare the extent of contamination caused by antimicrobial‐resistant lactic acid bacteria (LAB) in imported and domestic natural cheeses on the Japanese market, LAB were isolated using deMan, Rogosa and Sharpe (MRS) agar and MRS agar supplemented with six antimicrobials. From 38 imported and 24 Japanese cheeses, 409 LAB isolates were obtained and their antimicrobial resistance was tested. The percentage of LAB resistant to dihydrostreptomycin, erythromycin, and/or oxytetracycline isolated from imported cheeses (42.1%) was significantly higher than that of LAB resistant to dihydrostreptomycin or oxytetracycline from cheeses produced in Japan (16.7%; P = 0.04). Antimicrobial resistance genes were detected in Enterococcus faecalis (tetL, tetM, and ermB; tetL and ermB; tetM) E. faecium (tetM), Lactococcus lactis (tetS), Lactobacillus (Lb.), casei/paracasei (tetM or tetW), and Lb. rhamnosus (ermB) isolated from seven imported cheeses. Moreover, these E. faecalis isolates were able to transfer antimicrobial resistance gene(s). Although antimicrobial resistance genes were not detected in any LAB isolates from Japanese cheeses, Lb. casei/paracasei and Lb. coryniformis isolates from a Japanese farm‐made cheese were resistant to oxytetracycline (minimal inhibitory concentration [MIC], 32 µg/mL). Leuconostoc isolates from three Japanese farm‐made cheeses were also resistant to dihydrostreptomycin (MIC, 32 to > 512 µg/mL). In conclusion, the present study demonstrated contamination with antimicrobial‐resistant LAB in imported and Japanese farm‐made cheeses on the Japanese market, but not in Japanese commercial cheeses.     

Abnormal activation of glial cells in the brains of prion protein-deficient mice ectopically expressing prion protein-like protein, PrPLP/Dpl

BACKGROUND: Some lines of mice homozygous for a disrupted prion protein gene (Prnp), including Ngsk Prnp(0/0) mice, exhibit Purkinje cell degeneration as a consequence of the ectopic overexpression of the downstream gene for prion protein-like protein (PrPLP/Dpl) in the brain, but others, such as Zrch I Prnp(0/0) mice, show neither the neurodegeneration nor the expression of PrPLP/Dpl. In the present study, we found that Ngsk Prnp(0/0), but not Zrch I Prnp(0/0) mice, developed gliosis involving both astrocytes and microglia in the brain. MATERIALS AND METHODS: The brains from wild-type (Prnp(+/+)), Ngsk Prnp(0/0), Zrch I Prnp(0/0), and reconstituted Ngsk Prnp(0/0) mice carrying a mouse PrP transgene, designated Tg(P) Ngsk Prnp(0/0) mice, were subjected into Northern blotting and in situ hybridization using probes of glial fibrillary acidic protein (GFAP) and lysozyme M (LM) specific for astrocytes and microglia, respectively. Immunohistochemistry was also performed on the brain sections using anti-GFAP and anti-F4/80 antibodies. RESULTS: Northern blotting demonstrated upregulated expression of the genes for GFAP and LM in the brains of Ngsk Prnp(0/0), but not in Zrch I Prnp(0/0) mice. A transgene for normal mouse PrP(C) successfully rescued Ngsk Prnp(0/0) mice from the glial activation. In situ hybridization and immunohistochemistry revealed activated astrocytes and microglia mainly in the white matter of both the forebrains and cerebella. In contrast, there was no evidence of neuronal injury except for the Purkinje cell degeneration. Moreover, the glial cell activation was notable well before the onset of the Purkinje cell degeneration. CONCLUSIONS: These findings strongly suggest that ectopic PrPLP/Dpl in the absence of PrP(C) is actively involved in the glial-cell activation in the brain.     

Inhibitory effect of heterologous ribosome recycling factor on growth of Escherichia coli

Ribosome recycling factor (RRF) of Thermotoga maritima was expressed in Escherichia coli from the cloned T. maritima RRF gene and purified. Expression of T. maritima RRF inhibited growth of the E. coli host in a dose-dependent manner, an effect counteracted by the overexpression of E. coli RRF. T. maritima RRF also inhibited the E. coli RRF reaction in vitro. Genes encoding RRFs from Streptococcus pneumoniae and Helicobacter pylori have been cloned, and they also impair growth of E. coli, although the inhibitory effect of these RRFs was less pronounced than that of T. maritima RRF. The amino acid sequence at positions 57 to 62, 74 to 78, 118 to 122, 154 to 160, and 172 to 176 in T. maritima RRF differed totally from that of E. coli RRF. This suggests that these regions are important for the inhibitory effect of heterologous RRF. We further suggest that bending and stretching of the RRF molecule at the hinge between two domains may be critical for RRF activity and therefore responsible for T. maritima RRF inhibition of the E. coli RRF reaction.     

Rapid and Quantitative Assay of Amyloid-Seeding Activity in Human Brains Affected with Prion Diseases

The infectious agents of the transmissible spongiform encephalopathies are composed of amyloidogenic prion protein, PrP^Sc. Real-time quaking-induced conversion can amplify very small amounts of PrP^Sc seeds in tissues/body fluids of patients or animals. Using this in vitro PrP-amyloid amplification assay, we quantitated the seeding activity of affected human brains. End-point assay using serially diluted brain homogenates of sporadic Creutzfeldt–Jakob disease patients demonstrated that 50% seeding dose (SD₅₀) is reached approximately 10¹⁰/g brain (values varies 10^8.79–10.63/g). A genetic case (GSS-P102L) yielded a similar level of seeding activity in an autopsy brain sample. The range of PrP^Sc concentrations in the samples, determined by dot-blot assay, was 0.6–5.4 μg/g brain; therefore, we estimated that 1 SD₅₀ unit was equivalent to 0.06–0.27 fg of PrP^Sc. The SD₅₀ values of the affected brains dropped more than three orders of magnitude after autoclaving at 121°C. This new method for quantitation of human prion activity provides a new way to reduce the risk of iatrogenic prion transmission.     

Population ecology of Thrips palmi (Thysanoptera:Thripidae) in orchid farms in Thailand

The population dynamics and in-farm distribution of Thrips palmi Karny were evaluated in two orchid farms, one in the Bangkok metropolitan area and the other in Ratchaburi Province, Thailand. Yellow sticky traps were hung above the orchid canopy (three traps/220 m²), and the trapped adults were counted weekly from November 2009 to October 2010 at the first location and from January to December of 2012 at the second location. T. palmi nymphs and adults were both counted weekly on 300 orchid panicles in an area of 2,200 m² at both locations. We found that the T. palmi population dynamics at the first location was represented by a W-shaped curve, with the distribution clumped throughout the year, whereas the curve was J-shaped at the second location, with the distribution mostly clumped. Qualitative surveys of thrips species were conducted on ten different orchid genera in ten locations and on 20 different Dendrobium hybrids in 20 locations. Five random sampling plots (1 × 4 m²) were examined on each farm. The results indicated that only T. palmi was found on every farm. Moreover, quantitative comparison among five orchid types revealed that Dendrobium flowers with darker color are more attractive to T. palmi. However, this phenomenon does not coincide with Oncidium sp. and Mokara sp. because of their unfavorable morphology for thrips behavior.     

Ultrasensitive human prion detection in cerebrospinal fluid by real-time quaking-induced conversion

The development of technologies for the in vitro amplification of abnormal conformations of prion protein (PrP(Sc)) has generated the potential for sensitive detection of prions. Here we developed a new PrP(Sc) amplification assay, called real-time quaking-induced conversion (RT-QUIC), which allows the detection of ≥1 fg of PrP(Sc) in diluted Creutzfeldt-Jakob disease (CJD) brain homogenate. Moreover, we assessed the technique first in a series of Japanese subjects and then in a blind study of 30 cerebrospinal fluid specimens from Australia, which achieved greater than 80% sensitivity and 100% specificity. These findings indicate the promising enhanced diagnostic capacity of RT-QUIC in the antemortem evaluation of suspected CJD.     

TLR-dependent induction of IFN-beta mediates host defense against Trypanosoma cruzi

Host resistance to the intracellular protozoan parasite Trypanosoma cruzi depends on IFN-gamma production by T cells and NK cells. However, the involvement of innate immunity in host resistance to T. cruzi remains unclear. In the present study, we investigated host defense against T. cruzi by focusing on innate immunity. Macrophages and dendritic cells (DCs) from MyD88(-/-)TRIF(-/-) mice, in which TLR-dependent activation of innate immunity was abolished, were defective in the clearance of T. cruzi and showed impaired induction of IFN-beta during T. cruzi infection. Neutralization of IFN-beta in MyD88(-/-) macrophages led to enhanced T. cruzi growth. Cells from MyD88(-/-)IFNAR1(-/-) mice also showed impaired T. cruzi clearance. Furthermore, both MyD88(-/-)TRIF(-/-) and MyD88(-/-)IFNAR1(-/-) mice were highly susceptible to in vivo T. cruzi infection, highlighting the involvement of innate immune responses in T. cruzi infection. We further analyzed the molecular mechanisms for the IFN-beta-mediated antitrypanosomal innate immune responses. MyD88(-/-)TRIF(-/-) and MyD88(-/-)IFNAR1(-/-) macrophages and DCs exhibited defective induction of the GTPase IFN-inducible p47 (IRG47) after T. cruzi infection. RNA interference-mediated reduction of IRG47 expression in MyD88(-/-) macrophages resulted in increased intracellular growth of T. cruzi. These findings suggest that TLR-dependent expression of IFN-beta is involved in resistance to T. cruzi infection through the induction of IRG47.