Mechanism and role of high-potassium-induced reduction of intracellular Ca2+ concentration in rat osteoclasts

Osteoclasts are multinucleated, bone-resorbing cells that show structural and functional differences between the resorbing and nonresorbing (motile) states during the bone resorption cycle. In the present study, we measured intracellular Ca²⁺ concentration ([Ca²⁺]_i) in nonresorbing vs. resorbing rat osteoclasts. Basal [Ca²⁺]_i in osteoclasts possessing pseudopodia (nonresorbing/motile state) was around 110 nM and significantly higher than that in actin ring-forming osteoclasts (resorbing state, around 50 nM). In nonresorbing/motile osteoclasts, exposure to high K⁺ reduced [Ca²⁺]_i, whereas high K⁺ increased [Ca²⁺]_i in resorbing state osteoclasts. In nonresorbing/motile cells, membrane depolarization and hyperpolarization applied by the patch-clamp technique decreased and increased [Ca²⁺]_i, respectively. Removal of extracellular Ca²⁺ or application of 300 µM La³⁺ reduced [Ca²⁺]_i to 50 nM in nonresorbing/motile osteoclasts, and high-K⁺-induced reduction of [Ca²⁺]_i could not be observed under these conditions. Neither inhibition of intracellular Ca²⁺ stores or plasma membrane Ca²⁺ pumps nor blocking of L- and N-type Ca²⁺ channels significantly reduced [Ca²⁺]_i. Exposure to high K⁺ inhibited the motility of nonresorbing osteoclasts and reduced the number of actin rings and pit formation in resorbing osteoclasts. These results indicate that in nonresorbing/motile osteoclasts, a La³⁺-sensitive Ca²⁺ entry pathway is continuously active under resting conditions, keeping [Ca²⁺]_i high. Changes in membrane potential regulate osteoclastic motility by controlling the net amount of Ca²⁺ entry in a "reversed" voltage-dependent manner, i.e., depolarization decreases and hyperpolarization increases [Ca²⁺]_i. membrane depolarization; resorbing and motile activities; bone resorbing cycle     

Macrophage ubiquitin-specific protease 2 modifies insulin sensitivity in obese mice

We previously reported that ubiquitin-specific protease (USP) 2 in macrophages down-regulates genes associated with metabolic diseases, suggesting a putative anti-diabetic role for USP2 in macrophages. In this study, we evaluate this role at both cellular and individual levels. Isolated macrophages forcibly expressing Usp2a, a longer splicing variant of USP2, failed to modulate the insulin sensitivity of 3T3-L1 adipocytes. Similarly, macrophage-selective overexpression of Usp2a in mice (Usp2a transgenic mice) had a negligible effect on insulin sensitivity relative to wild type littermates following a three-month high-fat diet. However, Usp2a transgenic mice exhibited fewer M1 macrophages in their mesenteric adipose tissue. Following a six-month high-fat diet, Usp2a transgenic mice exhibited a retarded progression of insulin resistance in their skeletal muscle and liver, and an improvement in insulin sensitivity at an individual level. Although conditioned media from Usp2a-overexpressing macrophages did not directly affect the insulin sensitivity of C2C12 myotubes compared to media from control macrophages, they did increase the insulin sensitivity of C2C12 cells after subsequent conditioning with 3T3-L1 cells. These results indicate that macrophage USP2A hampers obesity-elicited insulin resistance via an adipocyte-dependent mechanism.     

A case with glucagonoma syndrome--heterogeneity of glucagon and insulin

The heterogeneity of glucagon and insulin in plasma and tissue extracts from a 57-year-old female with glucagonoma syndrome with surgically and autopsy verified islet-cell tumors was studied by Bio-Gel P-10 filtration. The preoperative plasma immunoreactive glucagon (IRG) level was 20.2 ng/ml, and plasma glucagon-like immunoreactivity(GLI) 25.8 ng/ml. The column chromatography of the preoperative plasma revealed three or four IRG components and four GLI components. Among these, peak II, the large glucagon immunoreactivity (LGI) peak, considered a candidate for proglucagon, was prominent, along with peak III. The resected metastatic liver tumor contained an enormous amount of IRG and an appreciable amount of immunoreactive insulin (IRI), indicating that the elevated plasma IRG was mainly of tumor origin. The IRG pattern of the tumor tissue extract revealed a small quantity of IRG in peaks I and II, and a large amount in peak III; control pancreatic tissue extract manifested a similar elution pattern. The IRI elution pattern of the tumor tissue extract revealed two major IRI peaks which migrated close to the elution volume of cytochrome C and insulin, respectively. This is a quite different pattern from the control pancreatic tissue extract in which the RI peak was localized in the elution volume of the insulin. We conclude that the present metastatic liver tumor produced not only enormous amounts of glucagon but heterogeneous peptides which contained immunological insulin determinants within their.     

Bioconversion of 2,6-dimethylpyridine to 6-methylpicolinic acid by Exophiala dermatitidis (Kano) de Hoog DA5501 cells grown on n-dodecane

Alkane-assimilating microorganisms were isolated from enrichment cultures using n-octane, n-dodecane, n-hexadecane, or pristane (2,6,10,14-tetramethylpentadecane) as a sole carbon source to find microbial catalysts oxidizing methyl groups of 2,6-dimethylpyridine. The cells of Exophiala dermatitidis (Kano) de Hoog DA5501, an n-dodecane-assimilating fungus, oxidized a single methyl group of 2,6-dimethylpyridine to produce 6-methylpicolinic acid (6-methylpyridine-2-carboxylic acid) without the formation of dipicolinic acid (pyridine-2,6-dicarboxylic acid); 67 mM 6-methylpicolinic acid (9.2 g/l) accumulated with a molar conversion yield of 89% by 54-h incubation. The fungus cells also oxidized the methyl group of 2,6-dimethylpyrazine and 2,4,6-trimethylpyridine regioselectively.     

Paecilotoxin production in clinical or terrestrial isolates of Paecilomyces lilacinus strains

The production of paecilotoxin from various isolates of Paecilomyces lilacinus was studied using three different media and high performance liquid chromatography (HPLC). Alkaline medium was found to be suitable for the production of the toxins. Among 20 strains tested, 19 including four clinical isolates were found to produce the toxins. Production patterns of paecilotoxins were very similar in each strain and the main toxins were A and B.     

Direct isolation of Yersinia pseudotuberculosis from fresh water in Japan.

Yersinia pseudotuberculosis was recovered from KOH-treated precipitates of 20.6% of 500 freshwater samples. KOH treatment of precipitates is a simple and expedient means of recovering Y. pseudotuberculosis from such samples.     

Controlled-release system of single-stranded DNA triggered by the photothermal effect of gold nanorods and its in vivo application

Gold nanorods have strong absorption bands in the near-infrared region, in which light penetrates deeply into tissues. The absorbed light energy is converted into heat by gold nanorods, the so-called 'photothermal effect'. Hence, gold nanorods are expected to act not only as on-demand thermal converters for photothermal therapy but also as controllers of a drug-release system responding to irradiation by near-infrared light. To achieve a controlled-release system that can be triggered by light irradiation, double-stranded DNA (dsDNA) was modified on gold nanorods. When the dsDNA-modified gold nanorods were irradiated by near-infrared light, the single-stranded DNA (ssDNA) was released from gold nanorods due to the photothermal effect. The amount of released ssDNA was dependent upon the power and exposure time of light irradiation. Release of ssDNA was also observed in tumors grown on mice after light irradiation. Such a controlled-release system of oligonucleotide triggered by the photothermal effect could expand the applications of gold nanorods that have unique optical characteristics in medicinal fields.     

Comparative study on kynurenic acid production in the rat striatum by tryptophan enantiomers: an in vivo microdialysis study

Kynurenic acid (KYNA), an endogenous antagonist of N-methyl-D-aspartate and α(7) nicotinic acetylcholine receptors, is one of the L-tryptophan (Trp) metabolites. To compare the level of KYNA produced in the striatum of rats after independent administration of L-Trp and D-Trp, rats were intraperitoneally administered L-Trp and/or D-Trp (100 mg/kg), and a microdialysis (MD) probe was implanted in the striatum. The KYNA level in the MD samples was determined using the column-switching high-performance liquid chromatography system. KYNA levels in the MD samples increased by approximately twofold in rats that were administered D-Trp or L-Trp; this result suggests that just as L-Trp, D-Trp was also metabolized to KYNA in the striatum. Additionally, 30 min before the administration of D-Trp, rats were administered 3-methyl pyrazole-5-carboxylic acid (MPC) (50 mg/kg), which is a specific inhibitor of D-amino acid oxidase (DAAO). Pretreatment with MPC suppressed striatal KYNA production; this result suggests that DAAO, encoded by one of the susceptible genes for schizophrenia, may contribute to the production of KYNA from D-Trp in the striatum of rats.