Low-molecular RNA subcellular localization and its binding strength with cellular structures

Some fractions of low molecular weight (LMW) nuclear RNAs were shown to be present in the cytoplasm of rat liver cells. In addition to known 4S tRNA, 5S and 5,8S rRNAs U3 and 8S1 LMW nuclear RNAs, 8SII and 8SIII LMW RNAs have been detected in RNA preparations of free total and membrane-bound polysomes. The U3 and 8SI polysoma I RNAs seem to be associated with high molecular weight polysomal RNA. Using thermal phenol fractionation, that some LMW RNAs were shown to be slightly bound to the cellular structures whereas some others are bound more tightly. Considerable amounts of LMW RNAs are tightly bound to the chromosome-nucleolar apparatus. They can be extracted only at 85 degrees C. The data presented are discussed with regard to LMW nuclear and polysomal RNAs functions.     

Spectrophotometric determination of peptide transport with chromogenic peptide mimetics

A spectrophotometric assay to determine peptide transport has been developed. Using two chromogenic peptide mimetics, L-phenylalanyl-L-2-sulfanilylglycine (PSG) and L-phenylalanyl-L-3-thiaphenylalanine (PSP), the peptide transport patterns in individual cell species can be evaluated effectively. After the addition of PSG to a HeLa cell suspension, sulfanilic acid accumulated progressively inside, but not outside, the cells, demonstrating that PSG was transported wholly intact. The addition of PSP to the same cell suspension was followed immediately by extracellular thiophenol production. Measurement of the rate of thiophenol release thereby provided direct determination of PSP transport. The thiophenol release was consistent with Michaelis-Menten kinetics, with a K(m) of 0.016 mM and a V(max) of 5.07 nmol/min (1 x 10(6) cells/ml, pH 7.4, 37 degrees C). The resulting kinetic constants estimated were in agreement with values determined by single-substrate enzyme kinetics. Using PSP, transport kinetics of various dipeptides was examined by competitive spectrophotometry. As a result, dipeptides tested could be ranked in order of kinetic power for their transport.     

Increasing production in Korean shrimp farms with white-spot syndrome virus PCR-negative brood stock

White-spot syndrome virus (WSSV) is a devastating, infectious virus affecting shrimp. Although sensitive techniques involving PCR have been developed to assist farmers in screening shrimp (brood stock) for WSSV prior to stocking ponds, such practices have not yet been applied in Korea. Despite the rationality of implementing screening, there has been some doubt as to whether the stocking of WSSV-PCR-negative fry epidemiologically decreases white-spot disease outbreaks. Here, we report a retrospective analysis of data from shrimp farms in the western coast of Korea where WSSV-PCR-negative brood stocks were used to stock rearing ponds. A total of 366 shrimp from Heuksan Island were sampled for WSSV with PCR. Of the tested shrimp, 7.2% (28 brood stocks) were identified as WSSV positive; only WSSV-PCR-negative shrimp were used for brood stocks. Total unit production (final shrimp production/ the area of the ponds) was higher, at 1.96, in ponds where WSSV-PCR-negative shrimp were used, as compared with 1.02 in other ponds in Korea in 2004. This retrospective analysis of WSSV in Korea may be useful to the shrimp aquaculture industry, suggesting a testable hypothesis that may contribute to the eventual control of WSSV outbreaks.     

Polygoni cuspidati radix inhibits the activation of Syk kinase in mast cells for antiallergic activity

The antiallergic activity of Polygoni cuspidati radix (PR) and the mechanism of action by which it functions were investigated in this study. The extract of PR exhibited potent inhibitory activity in mast cells; its IC50 values were 62 +/- 2.1 microg/ml for RBL-2H3 mast cells and 46 +/- 3.2 microg/m for bone marrow-derived mast cells by antigen stimulation, and it also suppressed the expression of tumor necrosis factor-alpha and interleukin-4 in RBL-2H3 cells. According to the in vivo animal allergy model, it inhibited a local allergic reaction, passive cutaneous anaphylaxis, in a dose-dependent manner. With regard to its mechanism of action, PR inhibited the activating phosphorylation of Syk, a key signaling protein for the activation of mast cells. It also suppressed Akt and the mitogen-activated protein kinases ERK1/2, p38, and JNK, which are critical for the production of various inflammatory cytokines in mast cells. The results of the study indicate that the antiallergic activity of PR is mediated through the inhibition of histamine release and allergic cytokine production by the inhibition of Syk activating phosphorylation in mast cells.     

Seasonal habitat use of three predatory fishes in a freshwater ecosystem

Hydrobiologia - To understand the spatiotemporal overlap in the habitat use of sympatric predators, we studied longitudinal activity and reservoir section and depth use of pike (Esox lucius),...     

Changes in Neuronal Dopamine Homeostasis following 1-Methyl-4-phenylpyridinium (MPP+) Exposure

1-Methyl-4-phenylpyridinium (MPP⁺), the active metabolite of the neurotoxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine, selectively kills dopaminergic neurons in vivo and in vitro via a variety of toxic mechanisms, including mitochondrial dysfunction, generation of peroxynitrite, induction of apoptosis, and oxidative stress due to disruption of vesicular dopamine (DA) storage. To investigate the effects of acute MPP⁺ exposure on neuronal DA homeostasis, we measured stimulation-dependent DA release and non-exocytotic DA efflux from mouse striatal slices and extracellular, intracellular, and cytosolic DA (DA_cyt) levels in cultured mouse ventral midbrain neurons. In acute striatal slices, MPP⁺ exposure gradually decreased stimulation-dependent DA release, followed by massive DA efflux that was dependent on MPP⁺ concentration, temperature, and DA uptake transporter activity. Similarly, in mouse midbrain neuronal cultures, MPP⁺ depleted vesicular DA storage accompanied by an elevation of cytosolic and extracellular DA levels. In neuronal cell bodies, increased DA_cyt was not due to transmitter leakage from synaptic vesicles but rather to competitive MPP⁺-dependent inhibition of monoamine oxidase activity. Accordingly, monoamine oxidase blockers pargyline and l-deprenyl had no effect on DA_cyt levels in MPP⁺-treated cells and produced only a moderate effect on the survival of dopaminergic neurons treated with the toxin. In contrast, depletion of intracellular DA by blocking neurotransmitter synthesis resulted in ∼30% reduction of MPP⁺-mediated toxicity, whereas overexpression of VMAT2 completely rescued dopaminergic neurons. These results demonstrate the utility of comprehensive analysis of DA metabolism using various electrochemical methods and reveal the complexity of the effects of MPP⁺ on neuronal DA homeostasis and neurotoxicity.     

Neutrophil activation by Candida glabrata but not Candida albicans promotes fungal uptake by monocytes

Candida albicans and Candida glabrata account for the majority of candidiasis cases worldwide. Although both species are in the same genus, they differ in key virulence attributes. Within this work, live cell imaging was used to examine the dynamics of neutrophil activation after confrontation with either C. albicans or C. glabrata. Analyses revealed higher phagocytosis rates of C. albicans than C. glabrata that resulted in stronger PMN (polymorphonuclear cells) activation by C. albicans. Furthermore, we observed differences in the secretion of chemokines, indicating chemotactic differences in PMN signalling towards recruitment of further immune cells upon confrontation with Candida spp. Supernatants from co‐incubations of neutrophils with C. glabrata primarily attracted monocytes and increased the phagocytosis of C. glabrata by monocytes. In contrast, PMN activation by C. albicans resulted in recruitment of more neutrophils. Two complex infection models confirmed distinct targeting of immune cell populations by the two Candida spp.: In a human whole blood infection model, C. glabrata was more effectively taken up by monocytes than C. albicans and histopathological analyses of murine model infections confirmed primarily monocytic infiltrates in C. glabrata kidney infection in contrast to PMN‐dominated infiltrates in C. albicans infection. Taken together, our data demonstrate that the human opportunistic fungi C. albicans and C. glabrata are differentially recognized by neutrophils and one outcome of this differential recognition is the preferential uptake of C. glabrata by monocytes.