Continuous fluorometric assay of Ca2+ transport by liposomes with Quin 2 entrapped: effect of phospholipase A2 and unsaturated long-chain fatty acids

The amount of free calcium in the cytoplasm is important in stimulation coupled with a number of cellular functions. The putative ionophoretic action of membrane lipid metabolites on Ca2+ offers convenient explanation of the stimulation-coupled mobilization of cytoplasmic Ca2+. To analyze the ionophoretic action of the lipid metabolites, we devised a sensitive method to study Ca2+ transport that uses liposome-entrapped Quin 2. A calcium ionophore, A23187, increased the fluorescence intensity of the Ca2+-Quin 2 complex as a function of Ca2+ transport into liposomes. A similar Ca2+ flux into the liposomes was induced by phospholipase A2 (PLA2) and by various long-chain fatty acids in liposomes that consist of phospholipids containing unsaturated fatty acids. The potencies of the fatty acids for Ca2+ transport is inversely correlated with their melting points. The oxidized products of the unsaturated fatty acids increased the Ca2+ and nonspecific permeability of the biological membranes. These results suggest that stimulation-coupled PLA2 activation might mediates the mobilization of cytoplasmic Ca2+.     

Glucose-induced abnormal egg-laying rate in Caenorhabditis elegans

High glucose reduced the egg-laying rate of the nematode Caenorhabditis elegans and was dependent on serotonergic signaling. Antidiabetic drugs of the biguanide and thiazolidine classes ameliorated the detrimental effect of glucose on egg-laying rate, suggesting the possibility that this quick and easy assay system may be applicable to whole-animal screening for novel antidiabetic drugs, at least, of these classes.     

Purification of restriction endonuclease EcoO128I produced by an enteropathogenic Escherichia coli O128Ly3.

Restriction endonuclease EcoO128I, an isoschizomer of BstEII, was purified from a rough mutant of Escherichia coli O128Ly3. EcoO128I should be more convenient for recombinant DNA applications than BstEII, because of its improved cleavage activity at 37 degrees C.     

Pentanol and Benzyl Alcohol Attack Bacterial Surface Structures Differently

The genus Methylobacterium tolerates hygiene agents like benzalkonium chloride (BAC), and infection with this organism is an important public health issue. Here, we found that the combination of BAC with particular alcohols at nonlethal concentrations in terms of their solitary uses significantly reduced bacterial viability after only 5 min of exposure. Among the alcohols, Raman spectroscopic analyses showed that pentanol (pentyl alcohol [PeA]) and benzyl alcohol (BzA) accelerated the cellular accumulation of BAC. Fluorescence spectroscopic assays and morphological assays with giant vesicles indicated that PeA rarely attacked membrane structures, while BzA increased the membrane fluidity and destabilized the structures. Other fluorescent spectroscopic assays indicated that PeA and BzA inactivate bacterial membrane proteins, including an efflux pump for BAC transportation. These findings suggested that the inactivation of membrane proteins by PeA and BzA led to the cellular accumulation but that only BzA also enhanced BAC penetration by membrane fluidization at nonlethal concentrations.     

A new side effect of BCG immunotherapy — BCG-induced arthritis in man

Summary Ten of 159 patients showed arthritic symptoms during the course of BCG immunotherapy for advanced cancer. The arthritic symptoms occurring after BCG injections had the following characteristics: (1) The incidence of arthritis was closely correlated with the host immunologic responsiveness to BCG; (2) These symptoms usually occurred 1–5 months after the first BCG injection (7/10); (3) The arthritic symptoms usually started with morning stiffness (9/10), which was followed by acute inflammatory signs in the affected joints. They gradually subsided in response to treatment with nonsteroid antiinflammatory drugs, but were not completely cured while the effectiveness of BCG continued; (4) The symptoms were aggravated by additional BCG injections (8/10). (5) This form of arthritis could be differentiated from rheumatoid arthritis, tuberculous, or purulent arthritis by its clinical course and by roentgenograms of the affected joints. It is thought to be induced by the adjuvant effect of BCG, and is a new side effect of BCG immunotherapy.     

Improvement of the anoxia-induced mitochondrial dysfunction by membrane modulation

The mitochondrial dysfunction induced by anoxia in vitro was improved with chlorpromazine, cepharanthine, bromophenacyl bromide, and mepacrine without affecting phospholipid or adenine nucleotide metabolisms. The drugs inhibited lipid peroxidation by Fe²⁺, mitochondrial disruption by Ca²⁺, and membrane perturbation by lysolecithin, and retained the activity to control H⁺ permeability across mitochondrial membranes. The drugs appeared to preserve the functions by acting to suppress the development of membrane deterioration which may have resided in the deenergization of mitochondria in the absence of oxygen.     

Ca2+ translocation across liposomal membranes enhanced by unsaturated long-chain fatty acids

The putative ionophoretic action of phosphatidic acid or arachidonic acid metabolites for Ca2+ has offered an attractive explanation for stimulation-coupled mobilization of cytoplasmic Ca2+. We have examined the effects of Ca2+ ionophore and long-chain unsaturated fatty acids on the translocation of Ca2+ across the liposomal membrane by using Quin II-entrapped liposomes, a sensitive assay system for ionophoresis of Ca2+. A23187 increased Quin II fluorescence intensity corresponding to the translocation of Ca2+ into liposomes. Similar translocation was observed with unsaturated long-chain fatty acids but not with saturated fatty acids. Thus, when phospholipases of cell membrane are activated by certain stimuli, unsaturated long-chain fatty acids are liberated and might mediate the mobilization of cytoplasmic Ca2+.     

Diel patterns in singing activity of humpback whales in a winter breeding area in Okinawan (Ryukyuan) waters

Male humpback whales produce complex sounds known as songs during their breeding season. Previous studies have shown diel patterns of song in their breeding areas, but there had been no similar studies in the breeding area around Okinawa, Japan. To study diel patterns of song and the behavior of humpback whales in Okinawa, we conducted 24 hr recording with a fixed hydrophone in 2007, and vessel-based sighting surveys during 2014–2017. Song was monitored for 15 days, with peaks at sunrise and around 2200. Singing activity declined significantly between sunrise and sunset, then increased until 2200. Activity levels at night were higher and more stable than during the day. During 278 days of sighting surveys, 2,551 whales in 1,382 groups were observed. 79 individuals were confirmed as singers, all of which were lone whales. In six cases, singing individuals stopped singing before joining a group or began singing after leaving a group. Previous studies have shown that group size of humpback whales increases through the day. Considering the results from our study and the former studies, the decrease in singing activity as the day progresses may be a result of aggregation increasing, thus reducing the number of lone singers during the day.