Chemokine-independent preference for T-helper-1 cells in transendothelial migration

We analyzed differences in the transendothelial migration (TEM) ability of T-helper (Th)-1 and Th2 cells across a murine endothelial cell line (F-2) under static conditions. The TEM abilities of Th1 cells from mice bearing autoimmune diseases and antigen-specific Th1 cell lines were severalfold higher than those of Th2 cells and lines of the same origin. These preferences were observed without exogenous chemoattractant and were insensitive to pertussis toxin, which completely blocks TEM induced by exogenous chemoattractants. Antibodies against LFA-1 and ICAM-1 as well as CD44 markedly blocked the TEM of Th1 cells. TEM ability was also blocked by pharmacological inhibitors of Src family protein-tyrosine kinases (PP2 and herbimycin A), phosphatidylinositol 3-kinase (wortmannin), and phosphatidylinositol-specific phospholipase C (). Cross-linking of CD44 strongly induced highly elongated morphology in Th1 lines, but weakly in Th2 lines. The pharmacological inhibitors that blocked TEM also inhibited this morphological change, whereas pertussis toxin did not. These data indicate that there are signaling pathways for TEM independent of chemokine attraction, but through adhesion molecules including CD44, and that the preferential TEM ability of Th1 over Th2 cells is formed, at least in part, by intrinsic differences in these pathways.     

Regeneration processes of a boreal forest in Kamchatka with special reference to the contribution of sprouting to population maintenance

We studied regeneration patterns of three tree species Picea ajanensis, Betula platyphylla and Populus tremula from 1998 to 2000 in the Central Depression of the Kamchatka Peninsula. We paid special attention to the contribution of sprouting to their regeneration. P. ajanensis was the only species that regenerated by seedling. In a 40 × 40 m study plot, the density of P. ajanensis saplings < 2.0 cm in diameter at basal area (DBH) was 1132, and this was the highest among the three species studied. The number of saplings 2 cm in DBH declined sharply with size class. The spatial distribution of P. ajanensis saplings (< 2 cm in DBH) showed a significant positive correlation with that of adult trees and a negative correlation with that of gaps. These trends were not changed after re-measurement in 2000, although nearly half of the juveniles had died or been injured during the two years. These results suggest that small Picea saplings prefer habitats under the canopy of adult trees rather than in gaps for establishment. Most small individuals of B. platyphylla were produced from sprouts. The number of saplings in the smallest size class (< 2 cm in DBH) was much less than that of P. ajanensis, although the number of larger individuals did not decrease remarkably. The spatial distribution of B. platyphylla saplings showed a positive correlation with that of adult trunks and a negative correlation with that of canopy trees of P. ajanensis. These results suggest an effective contribution of sprouts to the regeneration of B. platyphylla. P. tremula was the only species that could invade big gaps and produce many root suckers efficiently. There were 181 suckers of P. tremula in the smallest size class (< 2 cm in DBH) in the study plot, although the number of saplings 2 cm in DBH declined abruptly. The spatial distribution of saplings of this species showed a slight positive correlation with that of gaps, and negative correlation with that of adult trees of B. platyphylla, P. ajanensis, and P. tremula. The root suckering strategy of P. tremula might be adaptive under severe conditions in high-latitude regions. Our data suggest, however, that it does not necessarily contribute to regeneration in mature forests. The three component species in this forest did not seem to utilize canopy gaps for regeneration; we suggest that gap dynamics do not work in this forest. The sparse canopy, which is a typical character of forests in high-latitude regions, might be a consequence of high mortalities of seedlings and root suckers inside gaps.     

Characterization of citrate transport through the plasma membrane in a carrot mutant cell line with enhanced citrate excretion

The superior ability of citrate excretion in a carrot (Daucus carota L.) mutant cell line, namely IPG (insoluble phosphate grower) [Takita et al. (1999a) Plant Cell Physiol. 40: 489] cells has been characterized in terms of citrate transport at the plasma membrane. IPG cells released about a 20-fold increase in citrate in comparison with malate, while the concentration of malate was only 35% lower than that of citrate in the cell sap. Citrate excretion was sensitive to anion channel blockers, such as niflumic acid and anthracene-9-carboxylic acid. These results indicate that IPG cells release citrate through the plasma membrane using citrate specific anion channels. The rate of citrate release from IPG cells was not affected by the concentration of aluminum (0 and 50 micro M), soluble P(i) (0 or 2 mM) and the pH (4.5-5.6) of the medium, suggesting that anion channels would not be regulated by such external conditions. Citrate excretion correlated with the H(+) efflux, possibly from the action of H(+)-ATPase on the plasma membrane. The activity of plasma membrane H(+)-ATPase was about three times higher in IPG cells than in wild-type cells, and might be involved in the high citrate excretion ability.     

A new ecdysteroid, 2-deoxy-5beta,20-dihydroxyecdysone from the fruits of Diploclisia glaucescens

Chemical investigation of ethyl acetate extract of the fruits of Diploclisia glaucescens of the family Menispermaceae furnished a new ecdysteroid 2-deoxy-5beta,20-dihydroxyecdysone, together with 20-hydroxyecdysone, 3-deoxy-1beta,20-dihydroxyecdysone, 2-deoxy-20-hydroxyecdysone, 24-ethyl-20-hydroxyecdysone (makisterone C). Latter two ecdysteroids are reported first time from the family Menispermaceae.     

A functional polymorphism in the promoter/enhancer region of the<Emphasis Type="Italic"> FOXP3/Scurfin</Emphasis> gene associated with type 1 diabetes

FOXP3/Scurfin, a member of forkhead/winged-helix proteins, is involved in the regulation of T-cell activation, and essential for normal immune homeostasis. The FOXP3/Scurfin gene is located on chromosome Xp11.23, which includes one of the type 1 diabetes susceptible loci. Therefore, we investigated whether the human FOXP3/Scurfin gene might be a new candidate gene for type 1 diabetes. We first screened the human FOXP3/Scurfin gene for microsatellite and single nucleotide polymorphisms. Next, we performed an association study between the FOXP3/Scurfin gene and type 1 diabetes. Then, the evaluation of promoter/enhancer activity of the intron with (GT)(n) polymorphism was performed by dual luciferase reporter assay. We demonstrated two regions contained microsatellite polymorphisms; one was (GT)(n), located on intron zero and the other (TC)(n) on intron 5, which were under linkage-disequilibrium. The (GT)(15) allele showed a significantly higher frequency in patients with type 1 diabetes than in controls (43.1% vs 32.6%, P=0.0027). The genotype frequencies of (GT)(15)/(GT)(15) in female patients and of (GT)(15) in male patients tended to be higher than those in female ( P=0.064) and male ( P=0.061) controls, respectively. A significant difference in the enhancer activity between (GT)(15) and (GT)(16) dinucleotide repeats was detected. In conclusion, the FOXP3/Scurfin gene appears to confer a significant susceptibility to type 1 diabetes in the Japanese population.     

Molecular mechanism of insulin resistance and obesity

Obesity and insulin resistance have been recognized as leading causes of major health issues. We have endeavored to depict the molecular mechanism of insulin resistance, focusing on the function of adipocyte. We have investigated a role of PPARgamma on the pathogenesis of Type II diabetes. Heterozygous PPARgamma-deficient mice were protected from the development of insulin resistance due to adipocyte hypertrophy under a high-fat diet. Moreover, a Pro12Ala polymorphism in the human PPARgamma2 gene was associated with decreased risk of Type II diabetes in Japanese. Taken together with these results, PPARgamma is proved to be a thrifty gene mediating Type II diabetes. Pharmacological inhibitors of PPARgamma/RXR ameliorate high-fat diet-induced insulin resistance in animal models of Type II diabetes. We have performed a genome-wide scan of Japanese Type 2 diabetic families using affected sib pair analysis. Our genome scan reveals at least 9 chromosomal regions potentially harbor susceptibility genes of Type II diabetes in Japanese. Among these regions, 3q26-q28 appeared to be very attractive one, because of the gene encoding adiponectin, the expression of which we had found enhanced in insulin-sensitive PPARgamma-deficient mice. Indeed, the subjects with the G/G genotype of SNP276 in the adiponectin gene were at increased risk for Type II diabetes compared with those having the T/T genotype. The plasma adiponectin levels were lower in the subjects with the G allele, suggesting that genetically inherited decrease in adiponectin levels predispose subjects to insulin resistance and Type II diabetes. Our work also confirmed that replenishment of adiponectin represents a novel treatment strategy for insulin resistance and Type II diabetes using animal models. Further investigation will be needed to clarify how adiponectin exerts its effect and to discover the molecular target of therapies.     

SOCS-3 regulates onset and maintenance of T(H)2-mediated allergic responses

Members of the suppressor of cytokine signaling (SOCS) family are involved in the pathogenesis of many inflammatory diseases. SOCS-3 is predominantly expressed in T-helper type 2 (T(H)2) cells, but its role in T(H)2-related allergic diseases remains to be investigated. In this study we provide a strong correlation between SOCS-3 expression and the pathology of asthma and atopic dermatitis, as well as serum IgE levels in allergic human patients. SOCS-3 transgenic mice showed increased T(H)2 responses and multiple pathological features characteristic of asthma in an airway hypersensitivity model system. In contrast, dominant-negative mutant SOCS-3 transgenic mice, as well as mice with a heterozygous deletion of Socs3, had decreased T(H)2 development. These data indicate that SOCS-3 has an important role in regulating the onset and maintenance of T(H)2-mediated allergic immune disease, and suggest that SOCS-3 may be a new therapeutic target for the development of antiallergic drugs.     

Comparative study of the defensive behavior and morphology of the gland opening area among harvestmen (Arachnida, Opiliones, Gonyleptidae) under a phylogenetic perspective

The arachnids of the order Opiliones (harvestmen) have a pair of scent glands opening at the sides of the body, the substances of which are used in defense. Several types of behavioral, morphological and chemical defensive mechanisms have been assigned to the order as a whole, although some of these tactics were restricted to particular groups. Only around 25 species have been studied from this perspective so far. In the present paper, we analyzed 33 species (mostly from the largest harvestmen family, the Gonyleptidae) aiming at recognizing the usefulness of the defensive characters in taxonomy and evolutionary biology. We observed the morphology of the gland opening (ozopore) area and the defensive behavior, and their relationship, and mapped these traits on an available phylogenetic hypothesis of relationship within Gonyleptidae. As outgroups, we analyzed Cosmetidae and Stygnidae. Combining the observed behavioral characters of the emission of defensive secretion (near the ozopore, with liquid displacement through an integumentary groove, or in form of a jet) with the morphological types of the gland opening (direction of the integumentary dome that surrounds the gland opening, presence of two openings and the relationship between their sizes, and presence of a V-shaped cut at the anterior opening), we recognized eight patterns. In addition, we could examine the evolution of such traits within Gonyleptidae.