Potent anti-R5 human immunodeficiency virus type 1 effects of a CCR5 antagonist, AK602/ONO4128/GW873140, in a novel human peripheral blood mononuclear cell nonobese diabetic-SCID, interleukin-2 receptor gamma-chain-knocked-out AIDS mouse model

We established human peripheral blood mononuclear cell (PBMC)-transplanted R5 human immunodeficiency virus type 1 isolate JR-FL (HIV-1(JR-FL))-infected, nonobese diabetic-SCID, interleukin 2 receptor gamma-chain-knocked-out (NOG) mice, in which massive and systemic HIV-1 infection occurred. The susceptibility of the implanted PBMC to the infectivity and cytopathic effect of R5 HIV-1 appeared to stem from hyperactivation of the PBMC, which rapidly proliferated and expressed high levels of CCR5. When a novel spirodiketopiperazine-containing CCR5 inhibitor, AK602/ONO4128/GW873140 (molecular weight, 614), was administered to the NOG mice 1 day after R5 HIV-1 inoculation, the replication and cytopathic effects of R5 HIV-1 were significantly suppressed. In saline-treated mice (n = 7), the mean human CD4(+)/CD8(+) cell ratio was 0.1 on day 16 after inoculation, while levels in mice (n = 8) administered AK602 had a mean value of 0.92, comparable to levels in uninfected mice (n = 7). The mean number of HIV-RNA copies in plasma in saline-treated mice were approximately 10(6)/ml on day 16, while levels in AK602-treated mice were 1.27 x 10(3)/ml (P = 0.001). AK602 also significantly suppressed the number of proviral DNA copies and serum p24 levels (P = 0.001). These data suggest that the present NOG mouse system should serve as a small-animal AIDS model and warrant that AK602 be further developed as a potential therapeutic for HIV-1 infection.     

Novel putative galactose operon involving lacto-N-biose phosphorylase in Bifidobacterium longum

A lacto-N-biose phosphorylase (LNBP) was purified from the cell extract of Bifidobacterium bifidum. Its N-terminal and internal amino acid sequences were homologous with those of the hypothetical protein of Bifidobacterium longum NCC2705 encoded by the BL1641 gene. The homologous gene of the type strain B. longum JCM1217, lnpA, was expressed in Escherichia coli to confirm that it encoded LNBP. No significant identity was found with any proteins with known function, indicating that LNBP should be classified in a new family. The lnpA gene is located in a novel putative operon for galactose metabolism that does not contain a galactokinase gene. The operon seems to be involved in intestinal colonization by bifidobacteria mediated by metabolism of mucin sugars. In addition, it may also resolve the question of the nature of the bifidus factor in human milk as the lacto-N-biose structure found in milk oligosaccharides.     

Dose-dependent effects of L-carnosine on the renal sympathetic nerve and blood pressure in urethane-anesthetized rats

The physiological function of L-carnosine (beta-alanyl-L-histidine) synthesized in mammalian muscles has been unclear. Previously, we observed that intravenous (i.v.) injection of L-carnosine suppressed renal sympathetic nerve activity (RSNA) in urethane-anesthetized rats, and L-carnosine administered via the diet inhibited the elevation of blood pressure (BP) in deoxycorticosterone acetate salt hypertensive rats. To identify the mechanism, we examined effects of i.v. or intralateral cerebral ventricular (l.c.v.) injection of various doses of L-carnosine on RSNA and BP in urethane-anesthetized rats. Lower doses (1 microg i.v.; 0.01 microg l.c.v.) of L-carnosine significantly suppressed RSNA and BP, whereas higher doses (100 microg i.v.; 10 microg l.c.v.) elevated RSNA and BP. Furthermore, we examined effects of antagonists of histaminergic (H1 and H3) receptors on L-carnosine-induced effects. When peripherally and centrally given, thioperamide, an H3 receptor antagonist, blocked RSNA and BP decreases induced by the lower doses of peripheral L-carnosine, whereas diphenhydramine, an H1 receptor antagonist, inhibited increases induced by the higher doses of peripheral L-carnosine. Moreover, bilateral lesions of the hypothalamic suprachiasmatic nucleus eliminated both effects on RSNA and BP induced by the lower (1 microg) and higher (100 microg) doses of peripheral L-carnosine. These findings suggest that low-dose L-carnosine suppresses and high-dose L-carnosine stimulates RSNA and BP, that the suprachiasmatic nucleus and histaminergic nerve are involved in the activities, and that L-carnosine acts in the brain and possibly other organs.     

The relationship between dietary intake and the number of teeth in elderly Japanese subjects

Objective: This study used a precise weighing method to assess whether tooth loss was related to nutrient intake in elderly Japanese subjects. Material and methods: Fifty‐seven subjects aged 74 years were randomly selected from a longitudinal interdisciplinary study of ageing. Complete 3‐day food intake data were obtained by a precise weighing method. The dietary intakes of energy and nutrients were calculated based on the Standard Tables of Food Composition in Japan (5th ed.). A clinical evaluation of the number of teeth present was carried out. Multiple regression standardised coefficients for each nutrient was estimated based on a continuous scale adjusted for gender, smoking habits, and educational level. After dividing the subjects into two groups according to the number of teeth present (0–19, 20+), the difference in the intake of nutrients and the amount of food consumed per day was evaluated. Results: The number of teeth present had a significant relationship with the intake of several nutrients. In particular, total protein, animal protein, sodium, vitamin D, vitamin B₁, vitamin B₆, niacin, and pantothenic acid were significantly associated with the number of teeth present and with the two groups (0–19, 20+). The intake of vegetables and fish, shellfish, and their products was significantly lower among subjects with fewer teeth. Conclusion: This study suggests that there was a significant relationship between nutrient intake, such as minerals and vitamins from food, and tooth loss.     

Population dynamics of the swallowtail butterfly,Papilio xuthus L., in a deforested area

The long-term population dynamics of the swallowtail butterfly, Papilio xuthus, were studied by means of life table analysis in a deforested area, where the host tree, Zanthoxylum ailanthoides, was growing. The number of eggs laid on host trees in the deforested area decreases as secondary succession progresses. When the trees were classified into three groups, i. e. tall, medium and short, according to their height relative to the surrounding vegetation, less eggs were laid on tall than on short host trees. Life tables for the natural populations in each generation were developed on the basis of mean values for 6 years. Eggs and larvae in early stages were attacked chiefly by small- and middle-sized predators, such as ants, spiders, bugs and orthopterids, while later stage larvae were attacked chiefly by birds or Polistes wasps. An inchneumonid wasp was the most important mortality factor in the pupal stage. Key-factor analysis was tested on the life table data of each immature stage. The late larval stage was a key stage, and most mortality factors were considered to operate independently. The test for density dependency did not show any tendencies. The analysis of variance in two-way classifications was carried out for the difference of survival rate among the tree groups. It was suggested that tall host trees were unsuitable for P. xuthus. The construction of life tables for artificially inoculated populations on large host trees showed that high physiological death rate were characteristic of such trees in comparison with natural populations on smaller trees. The characteristics of the population dynamics of P. xuthus in the deforested area are compared among some kinds of the habitat, and the interrelationship between P. xuthus and its host plant are discussed.     

Rapid demonstration of diversity of sulfatide molecular species from biological materials by MALDI-TOF MS

By combining the partition method for enrichment of sulfatides without any chromatographic procedures and the preparation method of lysosulfatides, we succeeded in analyzing these sulfated glycosphingolipids from biological materials by matrix-assisted laser desorption and ionization time-of-flight mass spectrometry (MALDI-TOF MS) to reduce the complexity of mass fragmentation patterns within a day. We found that sulfated GalCer (HSO3-3Gal beta 1Cer) (SM4s [galactosylsulfatide]) was composed of different species. While composition of SM4s specifically depended on source materials, it always contained hydroxy fatty acids of various degrees. In addition to the common sphingoid 4-sphingenine (d18:1), uncommon/unusual sphingoids phytosphingosine (4-hydroxysphinganine) (t18:0), eicosasphinganine (d20:0), 4-eicosasphingenine (d20:1), and sphingadienine (d18:2) were easily detected. Finally, in addition to SM4s, sulfatide sulfated LacCer (HSO3-3Gal beta 4Glc beta 1Cer) (SM3 [sulfated lactosylceramide]) and sulfated Gg3Cer (GalNAc beta 4(HSO3-3)Gal beta 4Glc beta 1Cer) (SM2 [sulfated gangliotriaosylceramide]) were clearly detected in renal tubule cells. The major SM4s was composed of ceramides possessing d18:1 with C22 hydroxy fatty acids (C22:0 h), C23:0 h, and C24:0 h, whereas the major SM3/SM2 were composed of ceramides possessing t18:0 with C22 normal fatty acids (C22:0), C23:0, C24:0. Namely, in these two series of sulfatides, either fatty acids or sphingoids were hydroxylated, and chain lengths of these components were exactly the same, consequently resulting in a similar polarity of ceramide moieties in these sulfatide species. These results demonstrated diversities of sulfatide molecular species, not only with respect to sugar moieties but also to ceramide moieties, which are probably important for specific effective functions in particular microenvironments such as lipid membrane microdomains.     

NCYM,a Cis-Antisense Gene of MYCN,Encodes a De Novo Evolved Protein That Inhibits GSK3β Resulting in the Stabilization of MYCN in Human Neuroblastomas

The rearrangement of pre-existing genes has long been thought of as the major mode of new gene generation. Recently, de novo gene birth from non-genic DNA was found to be an alternative mechanism to generate novel protein-coding genes. However, its functional role in human disease remains largely unknown. Here we show that NCYM, a cis-antisense gene of the MYCN oncogene, initially thought to be a large non-coding RNA, encodes a de novo evolved protein regulating the pathogenesis of human cancers, particularly neuroblastoma. The NCYM gene is evolutionally conserved only in the taxonomic group containing humans and chimpanzees. In primary human neuroblastomas, NCYM is 100% co-amplified and co-expressed with MYCN, and NCYM mRNA expression is associated with poor clinical outcome. MYCN directly transactivates both NCYM and MYCN mRNA, whereas NCYM stabilizes MYCN protein by inhibiting the activity of GSK3β, a kinase that promotes MYCN degradation. In contrast to MYCN transgenic mice, neuroblastomas in MYCN/NCYM double transgenic mice were frequently accompanied by distant metastases, behavior reminiscent of human neuroblastomas with MYCN amplification. The NCYM protein also interacts with GSK3β, thereby stabilizing the MYCN protein in the tumors of the MYCN/NCYM double transgenic mice. Thus, these results suggest that GSK3β inhibition by NCYM stabilizes the MYCN protein both in vitro and in vivo. Furthermore, the survival of MYCN transgenic mice bearing neuroblastoma was improved by treatment with NVP-BEZ235, a dual PI3K/mTOR inhibitor shown to destabilize MYCN via GSK3β activation. In contrast, tumors caused in MYCN/NCYM double transgenic mice showed chemo-resistance to the drug. Collectively, our results show that NCYM is the first de novo evolved protein known to act as an oncopromoting factor in human cancer, and suggest that de novo evolved proteins may functionally characterize human disease.