Synthesis of branched polysaccharides by polymerization of 6-O-t-butyldimethylsilyl-D-glucal through stereoregular haloglycosylation.

In this letter, we report synthesis of branched polysaccharide 2 by glycosylation of glucal-type monomer 1 with two free hydroxy groups at position 3 and 4. Monomer 1 polymerized with N-halosuccinimide promoter in acetonitrile solvent at room temperature--50 degrees C. The product was isolated as a petroleum ether insoluble fraction. The structure was determined by 1H and 13C NMR spectra as well as elemental analysis to be a polysaccharide consisting of 2-halo-2-deoxy-alpha-D-mannoside units, indicating that the polymerization proceeded via stereoregular glycosylation manner. The molecular weights determined by GPC with DMF were 3,300-4,000. The degree of branching was estimated by the NMR data of the product from the reaction of 2 with 3,5-dinitrobenzoyl chloride.

Skin antigens in the steady state are trafficked to regional lymph nodes by transforming growth factor-beta1-dependent cells.

Antigen capturing in the skin and antigen trafficking into regional lymph nodes (LN) initiate immune responses. In this study, employing melanin granule (MG) as an easily traceable antigen in two mouse strains that carried steel factor or hepatocyte growth factor transgenes and had melanocytosis in the epidermis or in the dermis respectively, we investigated the mechanism of antigen trafficking from the skin. MG captured in the epidermis or dermis accumulated in the regional LN, but not other tissues. Only in alymphoplastic mice did MG-laden cells pass through the lymphatics and reached many tissues. Since inflammatory regions were not observed in the skin of either type of transgenic mouse, our developmental system enables us to investigate constitutive capturing and trafficking of insoluble antigens in the steady state. Both dendritic cells and macrophages were laden with MG in the regional LN. To determine which cells traffic antigens to the LN, we prepared double mutants that carried the transgenes and lacked transforming growth factor (TGF)-beta1, since mice lacking TGF-beta1 are reported to be deficient of Langerhans cells. Few MG were observed in the regional LN of these double-mutant mice. We also showed that signaling via macrophage colony stimulating factor receptor or Flt3/Flk2 is not essential for development of the cells for this antigen trafficking. These results indicate that antigens in the epidermis and dermis in the steady state are trafficked into regional LN only by TGF-beta1-dependent cells, which may be a dendritic cell lineage.

Altered phase relation between sleep timing and core body temperature rhythm in delayed sleep phase syndrome and non-24-hour sleep-wake syndrome in humans.

Changes in the phase relation between sleep timing and the circadian pacemaker are suspected to have an etiological significance in circadian rhythm sleep disorders. Simultaneous recordings of rest-activity and rectal temperature in seven sighted delayed sleep phase syndrome (DSPS) patients, seven sighted non-24-h sleep-wake syndrome (non-24) patients, and 14 healthy controls were made for 10-14 days continuously in the subjects' homes. We found that sleep length and the interval from the body temperature (BT) trough to sleep offset were significantly longer in both non-24 and DSPS patients than in the controls, and that the interval between sleep onset and the BT trough was significantly less in the non-24 patients than in the DSPS patients and the controls. We postulate these alterations in phase relation to be associated with phase changes of the circadian pacemaker via different illumination timings.

New alpha-selective thermal glycosylation of acetyl-protected 2-acetamido-2-deoxy-beta-D-glucopyranosyl diphenylphosphinate.

This paper describes new alpha-selective thermal glycosylation using acetyl-protected 2-acetamido-2-deoxy-beta-D-glucopyranosyl diphenylphosphinate (4) as a glycosyl donor. When the glycosylation of 4 with 1-hexanol was carried out under various conditions, the conditions using trimethylsilyl trifluoromethanesulfonate as a promoter in nitromethane at reflux temperature were most suitable for the formation of the alpha anomer. The glycosylation of 4 with the other common alcohols gave corresponding alpha-glycosides in relatively high yields under the conditions. When cholesterol, a very steric hindered alcohol, was used as a glycosyl acceptor, alpha-glycoside was also produced predominantly.

Keratinocyte expression of transgenic hepatocyte growth factor affects melanocyte development, leading to dermal melanocytosis.

Using the epidermis-specific cytokeratin 14 promoter to deliver HGF exclusively from epidermal keratinocytes, we have examined the potential of hepatocyte growth factor (HGF) secreted from the normal environment to control morphogenesis. The transgenic mice displayed a significant increase of the number of melanocytes and their precursors in embryos starting not later than 16.5 dpc, and then after birth an explosive increase of dermal melanocytes started within 1 week, and these melanocytes were maintained throughout the entire life of the mice. Thus, HGF acts as a paracrine agent to promote survival, proliferation and differentiation of melanocyte precursors in vivo, and eventually causes melanocytosis. Loss of E-cadherin expression in dermal melanocyte precursors suggests that HGF caused dermal localization of melanocytes and their precursors by down-regulation of E-cadherin molecules.

Intramedullary and extramedullary B lymphopoiesis in osteopetrotic mice.

Adult bone marrow is a major site for hematopoiesis, and reduction of the bone marrow cavity induces hematopoiesis in extramarrow tissues. To investigate the rudimentary intramarrow and the compensatory extramarrow hematopoiesis, particularly B lymphopoiesis, we used 3 osteopetrotic mouse strains [op/op, mi/mi, and Fos (-/-)], which are severely deficient in functional osteoclasts and therefore form inadequate bone marrow cavities. We found that bone marrow in these osteopetrotic mice supports myelopoiesis but not B lymphopoiesis, although cells that have the potential to differentiate into B lineage cells are present in the bone marrow. Although B lymphopoiesis normally occurs both in the spleen and liver of newborn mice, compensatory B lymphopoiesis in adult op/op and mi/mi mice is observed only in the liver, while myelopoiesis is enhanced in both organs. Interestingly, mice lacking the Fos proto-oncogene exhibit B lymphopoiesis in the spleen as well as liver. The amounts of expression of steel factor, Flt3/Flk-2 ligand, and interleukin-7 in the bone marrow, spleen, or liver were not significantly affected in these osteopetrotic mutants. These findings suggest that the volume of the bone marrow cavity regulates B lymphopoiesis without affecting the production of certain hematopoietic growth factors. The splenic microenvironments that support both myelopoiesis and B lymphopoiesis in the neonatal stage are lost in adults and are not reactivated even in the osteopetrotic adults unless the Fos gene is disrupted.

Temporal EEG dynamics of non-REM sleep episodes in humans.

The process of the human non-rapid eye movement (non-REM) sleep period has not been clarified. Time-based analysis on sleep EEG may provide an explanation. We focused on chronological aspects of initiation and termination of non-REM episodes, using spectral analysis of sleep EEG. The subjects were healthy male volunteers (n14 Hz) and longer in lower frequency ranges (<14 Hz). There were significant differences in the rise and decay latencies between low and high sigma ranges, indicating that the whole frequency ranges were clearly separated at the middle of the sigma range (14 Hz). The rise and decay latencies were significantly different in lower frequency ranges. The clock time of the night significantly affected only the rise latencies of the delta (0.78-3.9 Hz), alpha (8.2-11.7 Hz) and low sigma (12.1-13.7 Hz) ranges. In conclusion, initiation and termination of non-REM sleep was represented by higher frequency ranges, whereas further evolution and devolution of non-REM sleep was represented by lower frequency ranges, and only the evolution process was affected by the clock time of the night.

Diurnal preference, sleep habits, circadian sleep propensity and melatonin rhythm in healthy human subjects.

After 24-h sleep deprivation, 33 healthy young subjects entered the 10/20 min ultra-short sleep-wake schedule for 26 h. Melatonin rhythm was hourly assessed simultaneously. Results indicated that morning preference was significantly correlated with habitual sleep onset (r=-0.41, P=0.04), habitual sleep offset (r=-0.52, P=0.002), melatonin peak time (r=-0.36, P=0.04), and sleep propensity onset time (r=-0.36, P=0.04). The intervals between habitual sleep mid-point and melatonin peak time and between habitual sleep mid-point and sleep propensity onset time were significantly longer in morning-preference subjects than in evening-preference subjects (P<0.05). These findings suggest that the variance of diurnal preference may be related to differences in phase relations between habitual sleep timing and the circadian pacemaker.

Tooth-specific expression conferred by the regulatory sequences of rat dentin sialoprotein gene in transgenic mice.

We have isolated a 3.8-kb DNA fragment containing the 5' flanking region, 1st exon, and 1st intron of the rat dentin sialoprotein (rDsp) gene and produced transgenic mice carrying a LacZ reporter gene under the control of this fragment. Expression of the transgene transcript and beta-galactosidase activity were restricted to dentin and odontoblasts with spatial and temporal patterns comparable to those of the endogenous mouse Dsp transcript, although beta-galactosidase activity could not be detected visually during embryonal stages. Other tissues tested, such as alveolar bones, ameloblasts and dental pulps, did not express the transgene. This indicates that the regulatory elements necessary for tooth-specific expression are present in the fragment, which contains a TATA box and several consensus sequences for binding sites of transcription factors related to tooth development, such as TCF-1/LEF-1, MSX-1 and Dlx-1. The regulatory sequences and the transgenic mice described here provide useful information for the study of tooth development.

Evidence supporting an association between the DRB1 gene and schizophrenia in Japanese.

The authors attempted a replication of earlier studies that detected an association of HLA-DR4 and DR1 with schizophrenia. Japanese patients with schizophrenia (n = 266, DSM-III-R criteria) and Japanese controls (n = 283) were genotyped for DR1 and DR4 alleles using a combination of group-specific polymerase chain reaction (PCR) amplification and PCR-restriction fragment length polymorphism. Significant positive association with HLA-DR1 [odds ratio (OR) = 1.87, corrected p = 0.04] and a negative association with HLA-DR4 (OR = 0.63, corrected p = 0.02) was noted. DR1 and DR4 were independently associated with schizophrenia. The association of the DR1-positive/DR4-negative genotype with schizophrenia was modest (OR = 2.60, 95% confidence intervals = 1.38-4.89, corrected p = 0.008). Thus, these findings support an association of the HLA DRB1 gene locus with schizophrenia in the Japanese population. Since both DR4 and DR1 are positively associated with rheumatoid arthritis, our findings are not simply consistent with the known negative association between schizophrenia and rheumatoid arthritis.

Commitment and differentiation of stem cells to the osteoclast lineage.

Osteoclasts are hematopoietic cells which play important roles in bone remodeling and resorption. They have phenotypic characteristics of the monocyte/macrophage lineages. In this review we first describe the phylogeny of osteoclasts. Osteoclast generation is closely linked to the presence of bone tissues. The formation of bone cavities in aquatic animals is underdeveloped, even though they have cells which have the potential to differentiate into osteoclasts. Next we describe recent advances in our understanding of osteoclastogenesis that have resulted from the identification of critical molecules and mutated genes of osteopetrotic mice. Reports that transcriptional factors PU.1 and c-Fos are essential for commitment and (or) differentiation into the osteoclast lineage and novel culture systems, which have clarified some characteristics of osteoclast precursors, are also described. We are now able to induce mature osteoclasts from hematopoietic stem cells and even from totipotent embryonic stem cells. Cell lines that differentiate into osteoclasts are also available. Using these culture systems and cell lines, the interactions of osteoclasts with osteoblastic stromal cells, which produce critical molecules for osteoclastogenesis, have been studied. Very recently, one of these critical molecules, osteoclast differentiation factor/osteoprotegerin-ligand, was cloned. The presence of this factor and macrophage-colony-stimulating factor is sufficient to induce osteoclast development in cultures inoculated only with an osteoclast precursor cell line. We review the present status and the remaining questions in osteoclast biology.

Progesterone-induced changes in sleep in male subjects.

Progesterone administration induces a reduction of the vigilance state in humans during wakefulness. It has been been suggested that this effect is mediated via neuroactive metabolites that interact with the gamma-aminobutyric, acidA (GABAA) receptor complex. To investigate the effects of progesterone administration on the sleep electroencephalogram (EEG) in humans we made polysomnographic recordings, including sleep stage-specific spectral analysis, and concomitantly measured plasma concentrations of progesterone and its GABA-active metabolites 3 alpha-hydroxy-5 alpha-dihydroprogesterone (allopregnanolone) and 3 alpha-hydroxy-5 beta-dihydroprogesterone (pregnanolone) in nine healthy male subjects in a double-blind placebo-controlled crossover study. Progesterone administration at 9:30 PM induced a significant increase in the amount of non-rapid eye movement (REM) sleep. The EEG spectral power during non-REM sleep showed a significant decrease in the slow wave frequency range (0.4-4.3 Hz), whereas the spectral power in the higher frequency range (> 15 Hz) tended to be elevated. Some of the observed changes in sleep architecture and sleep-EEG power spectra are similar to those induced by agonistic modulators of the GABAA receptor complex and appear to be mediated in part via the conversion of progesterone into its GABA-active metabolites.

Association of dopamine D2 receptor molecular variant with schizophrenia.

We have examined a variant of the dopamine D2 receptor gene (Ser311-->Cys) in 156 Japanese schizophrenic patients and 300 controls. The allele frequency of Cys311 was significantly higher in the whole patient group (0.054), among patients with onset before age 25 (0.090), and among those with a family history (0.135) than in the controls (0.018). 3 patients were homozygous for Cys311. The patients with Cys311 showed significantly less severe thought disorder and negative symptoms of schizophrenia than those without Cys311. The Cys311 variant of the D2 receptor may be a genetic risk factor for some types of schizophrenia.