Season specificity in the cold-induced calcium signal and the volatile chemicals in the atmosphere

Cold-induced Ca²⁺ signals in plants are widely accepted to be involved in cold acclimation. Surprisingly, despite using Arabidopsis plants grown in a growth chamber, we observed a clear seasonal change in cold-induced Ca²⁺ signals only in roots. Ca²⁺ signals were captured using Arabidopsis expressing Yellow Cameleon 3.60. In winter, two Ca²⁺ signal peaks were observed during a cooling treatment from 20 to 0°C, but in summer only one small peak was observed under the same cooling condition. In the spring and autumn seasons, an intermediate type of Ca²⁺ signal, which had a delayed first peak and smaller second peaks compared with the those of the winter type, was observed. Volatile chemicals and/or particles in the air from the outside may affect plants in the growth chamber. This idea is supported by the fact that incubation of plants with activated carbon changed the intermediate-type Ca²⁺ signal to the summer-type. The seasonality was also observed in the freezing tolerance of plants cold-acclimated in a low-temperature chamber. The solar radiation intensity was weakly correlated, not only with the seasonal characteristics of the Ca²⁺ signal but also with freezing tolerance. It has been reported that the ethylene concentration in the atmosphere seasonally changes depending on the solar radiation intensity. Ethylene gas and 1-aminocyclopropane-1-carboxylic acid treatment affected the Ca²⁺ signals, the shape of which became a shape close to, but not the same as, the winter type from the other types, indicating that ethylene may be one of several factors influencing the cold-induced Ca²⁺ signal.     

Influence of substituent modifications on the binding of 2-amino-1,8-naphthyridines to cytosine opposite an AP site in DNA duplexes: thermodynamic characterization

Here, we report on a significant effect of substitutions on the binding affinity of a series of 2-amino-1,8-naphthyridines, i.e., 2-amino-1,8-naphthyridine (AND), 2-amino-7-methyl-1,8-naphthyridine (AMND), 2-amino-5,7-dimethyl-1,8-naphthyridine (ADMND) and 2-amino-5,6,7-trimethyl-1,8-naphthyridine (ATMND), all of which can bind to cytosine opposite an AP site in DNA duplexes. Fluorescence titration experiments show that the binding affinity for cytosine is effectively enhanced by the introduction of methyl groups to the naphthyridine ring, and the 1:1 binding constant (10⁶ M⁻¹) follows in the order of AND (0.30) < AMND (2.7) < ADMND (6.1) < ATMND (19) in solutions containing 110 mM Na⁺ (pH 7.0, at 20°C). The thermodynamic parameters obtained by isothermal titration calorimetry experiments indicate that the introduction of methyl groups effectively reduces the loss of binding entropy, which is indeed responsible for the increase in the binding affinity. The heat capacity change (ΔC_p), as determined from temperature dependence of the binding enthalpy, is found to be significantly different between AND (−161 cal/mol K) and ATMND (−217 cal/mol K). The hydrophobic contribution appears to be a key force to explain the observed effect of substitutions on the binding affinity when the observed binding free energy (ΔG_obs) is dissected into its component terms.     

A reproducible swine model of a surgically created saccular thoracic aortic aneurysm

A reproducible swine thoracic aortic aneurysm (TAA) model is useful for investigating new therapeutic interventions. We report a surgical method for creating a reproducible swine saccular TAA model. We used eight female swine weighing 20–25 kg (LWD; ternary species). All procedures were performed under general anesthesia and involved left thoracotomy. Following aortic cross-clamping, the thoracic aorta was surgically dissected and the media and intima were resected, and the dissection plane was extended by spreading the outer layer for aneurysmal space. Subsequently, only the adventitial layer of the aorta was sutured. At 2 weeks after these procedures, angiography and computed tomography were performed. After follow-up imaging, the model animals were euthanized. Macroscopic, histological, and immunohistological examinations were performed. All model animals survived, and a saccular TAA was confirmed by follow-up imaging in all cases. The mean length of the shorter and the longer aortic diameter after the procedure were 14.01 ± 1.0 mm and 18.35 ± 1.4 mm, respectively (P<0.001). The rate of increase in the aortic diameter was 131.7 ± 13.8%, and the mean length of aneurysmal change at thoracic aorta was 22.4 ± 1.9 mm. Histological examination revealed intimal tears and defects of elastic fibers in the media. Immunostaining revealed MMP-2 and MMP-9 expressions at the aneurysm site. We report our surgical method for creating a swine saccular TAA model. Our model animal may be useful to investigate new therapeutic interventions for aortic disease.     

Olive leaf extract prevents cartilage degeneration in osteoarthritis of STR/ort mice

The chondroprotective effect of olive leaf extract (OLE) on knee osteoarthritis (OA) was studied with STR/ort mice (n = 5). OLE was administrated with a dosage of 100 mg/kg for 8 weeks and the OA severity score of hind limb knee joints was then measured. The Mankin scores of the knee joints of the non-OA control group, OA control group and OLE-treated group were 3.50, 11.13 and 7.20, respectively. This suggests that oral OLE supplements help prevent cartilage degeneration in STR/ort mice. In vitro, the synthesis of high molecular weight hyaluronan in synovial cells (HIG-82) was increased by OLE stimulation. This suggests that OLE modulates hyaluronan metabolism in synovial cells and improves OA symptoms. Our findings indicate that OLE intake inhibits cartilage destruction by increasing high molecular weight hyaluronan and thus preventing OA progress.     

Prostaglandin stimulation of adenosine 3′,5′-monophosphate accumulation in cultured chondrocytes in the presence or absence of parathyroid hormone

The effect of prostaglandin analogues on the cycle AMP level in cultured chondrocytes were examined. Prostaglandin E₁ at 0.4 to 30 μM, increased the intracellular concentration of cyclic AMP in chondrocytes. Its effect was rapid, being evident within 1 min and reaching a maximum in 10 to 20 min. The maximum level was sustained until 30 min after its addition and then decreased gradually. Prostaglandin D₂ and E₂ also increased the cyclic AMP level in chondrocytes, but they had less effect than prostaglandin E₁. Prostaglandin A₁ had no effect on the nucleotide level in chondrocytes, although they markedly increased the level in fibroblasts. The time course of stimulation of cyclic AMP accumulation in chondrocytes by prostaglandin E₁, D₂ or E₂ was quite different from that by parathyroid hormone (PTH): the effect of prostaglandin was slower and more sustained than that of PTH. PTH potentiated the effect of prostaglandin E₁, E₂, or D₂ on the cyclic AMP level in chondrocytes and that the combined effects of prostaglandin, PTH or both produced a synergistic effect on the accumulation of cyclic AMP in the chondrocytes. These findings suggest that prostaglandin E₁, E₂, and D₂ increase the synthesis of cyclic AMP and that the combined effect of the prostaglandins and PTH on the cyclic AMP level in chondrocytes is partly attributed to the synergistic synthesis of cyclic AMP in the cells.     

Divergent synthesis of kinase inhibitor derivatives,leading to discovery of selective Gck inhibitors

We accomplished divergent synthesis of potent kinase inhibitor BAY 61-3606 (1) and 27 derivatives via conjugation of imidazo[1,2-c]pyrimidine and indole ring compounds with aromatic (including pyridine) derivatives by means of palladium-catalyzed cross-coupling reaction. Spleen tyrosine kinase (Syk) and germinal center kinase (Gck, MAP4K2) inhibition assays showed that some of the synthesized compounds were selective Gck inhibitors.     

Female-specific target sites for both oestrogen and androgen in the teleost brain

To dissect the molecular and cellular basis of sexual differentiation of the teleost brain, which maintains marked sexual plasticity throughout life, we examined sex differences in neural expression of all subtypes of nuclear oestrogen and androgen receptors (ER and AR) in medaka. All receptors were differentially expressed between the sexes in specific nuclei in the forebrain. The most pronounced sex differences were found in several nuclei in the ventral telencephalic and preoptic areas, where ER and AR expression were prominent in females but almost completely absent in males, indicating that these nuclei represent female-specific target sites for both oestrogen and androgen in the brain. Subsequent analyses revealed that the female-specific expression of ER and AR is not under the direct control of sex-linked genes but is instead regulated positively by oestrogen and negatively by androgen in a transient and reversible manner. Taken together, the present study demonstrates that sex-specific target sites for both oestrogen and androgen occur in the brain as a result of the activational effects of gonadal steroids. The consequent sex-specific but reversible steroid sensitivity of the adult brain probably contributes substantially to the process of sexual differentiation and the persistent sexual plasticity of the teleost brain.     

Detection of the Leghemoglobin Gene on Two Chromosomes of Phaseolus vulgaris by in situ PCR Linked-Fluorescent in situ Hybridization (FISH)

The leghemoglobin (Lb) gene on the metaphase chromosomes ofPhaseolus vulgaris was amplified by in situ PCR. The amplifiedLb gene could be detected on two chromosomes by fluorescentin situ hybridization (FISH) using the short Lb gene probe. (Received January 9, 1998; Accepted April 30, 1998)