Associated factors of behavioural problems in children at preschool age: the Hokkaido study on environment and children's health.

BACKGROUND: Finding associated factors with childhood behavioural problems as early as preschool age is important. Studies have revealed several factors including socioeconomic factors, which may vary among different cultural background and population. However, investigation in general Japanese population of preschool age has not been well demonstrated. Thus, the objective of this study was to examine associated factors of childhood behavioural problems using Strengths and Difficulties Questionnaire (SDQ) in a prospective birth cohort study. METHODS: Total 3813 SDQ were distributed between October 2014 and December 2015 to the subpopulation of prospective birth cohort study, the Hokkaido Study on Environment and Children's Health. The subpopulation consisted of participants who had reached age 5 and were born between April 2008 and December 2010. Baseline questionnaire filled at recruitment and birth record were used to obtain participant information. Children with total difficulties score ≧ 13 were defined as likelihood of behavioural problems. A total of 2553 children with valid answers were included into the analysis. The response rate was 67.1%. RESULTS: Number of children with likelihood of behavioural problems was 521 (20.4%). Boys showed more problematic scores than girls. Multivariate analysis found that maternal pre-pregnancy BMI ≧ 30 kg/m2 , primipara, maternal education lower than high school, family income during pregnancy < 3 million yen/year and boy gender were the factors associated with increased odds ratio of likelihood of child behavioural problems. CONCLUSIONS: This study found that prenatal socioeconomic factors were associated with likelihood of child behavioural problems at preschool age in Japan.

Does tranexamic acid alter the risk of thromboembolism after total hip arthroplasty in the absence of routine chemical thromboprophylaxis?

Tranexamic acid (TXA) has been used to reduce blood loss during total hip arthroplasty (THA), but its use could increase the risk of venous thromboembolic disease (VTE). Several studies have reported that TXA does not increase the prevalence of deep vein thrombosis (DVT), but most of those used routine chemical thromboprophylaxis, thereby masking the potential increased risk of TXA on VTE. We wished to ascertain whether TXA increases the prevalence of VTE in patients undergoing THA without routine chemical thromboprophylaxis. We carried out a retrospective case-control study in 254 patients who underwent a primary THA, 127 of whom received TXA (1 g given pre-operatively) and a control group of 127 who did not. All patients had mechanical but no chemical thomboprophylaxis. Each patient was examined for DVT by bilateral ultrasonography pre-operatively and on post-operative days 1 and 7. TXA was found to statistically significantly increase the incidence of total DVT on post-operative day 7 compared with the control group (24 (18.9%) and 12 (9.4%), respectively; p < 0.05) but most cases of DVT were isolated distal DVT, with the exception of one patient with proximal DVT in each group. One patient in the control group developed a non-fatal symptomatic pulmonary embolism (PE). The use of TXA did not appear to affect the prevalence of either proximal DVT or PE.

Interlayer magnetoresistance due to chiral soliton lattice formation in hexagonal chiral magnet CrNb3S6.

We investigate the interlayer magnetoresistance (MR) along the chiral crystallographic axis in the hexagonal chiral magnet CrNb3S6. In a region below the incommensurate-commensurate phase transition between the chiral soliton lattice and the forced ferromagnetic state, a negative MR is obtained in a wide range of temperature, while a small positive MR is found very close to the Curie temperature. Normalized data of the negative MR almost falls into a single curve and is well fitted by a theoretical equation of the soliton density, meaning that the origin of the MR is ascribed to the magnetic scattering of conduction electrons by a nonlinear, periodic, and countable array of magnetic soliton kinks.

Anomalous magnetization behaviour in a single crystal of vanadium spinel FeV2O4.

Spinel oxide FeV2O4, having the orbital degrees of freedom at Fe(2+) and V(3+) ions, exhibits multi-step magnetic phase transitions and successive structural phase transitions at low temperatures. In order to clarify the magnetic properties of FeV2O4, we have measured the temperature dependence of magnetization, isothermal magnetization curves and specific heat using a single crystal of FeV2O4. Temperature-induced magnetization jumps below the 110 K were observed in the zero-field-cooled magnetization curves. Furthermore, we found that the behaviours of the isothermal magnetization curves were quite different between the zero-field-cooled and field-cooled conditions. We suggest that the change of the magnetic domain structure under the magnetic field associated with the orbital states of Fe(2+) ions is the possible origin of these intriguing and anomalous magnetic properties in a single crystal of FeV2O4.

Chiral magnetic soliton lattice on a chiral helimagnet.

Using Lorenz microscopy and small-angle electron diffraction, we directly present that the chiral magnetic soliton lattice (CSL) continuously evolves from a chiral helimagnetic structure in small magnetic fields in Cr(1/3)NbS2. An incommensurate CSL undergoes a phase transition to a commensurate ferromagnetic state at the critical field strength. The period of a CSL, which exerts an effective potential for itinerant spins, is tuned by simply changing the field strength. Chiral magnetic orders observed do not exhibit any structural dislocation, indicating their high stability and robustness in Cr(1/3)NbS2.

Symmetry and broken-symmetry in molecular orbital descriptions of unstable molecules. 3. The nature of chemical bonds of spin frustrated systems.

Symmetry and broken symmetry in the molecular orbital description of spin frustration systems have been investigated in relation to the resonating valence bond (RVB) theory of the spin liquid state and non-BCS superconductivity. Broken symmetry (BS) and resonating BS (RBS) molecular orbital (MO) methods have been employed to obtain resonating valence bond (RVB)-type explanations of spin frustrated systems. RBS MO solutions are expanded using the localized molecular orbitals (LMO) to elucidate a universal MO-VB description. The BS and RBS MO descriptions of triangular spin frustrated systems corresponding to transition structures for exchange-forbidden radical insertions were investigated in comparison with the RVB-type explanations of such systems. The BS and RBS calculations by the use of three different axial (SDW) solutions or three noncollinear GSO (helical SDW) solutions of a triangular hydrogen cluster were performed to obtain potential curves with and without resonance (quantum) effects. The resonating GSO (noncollinear) state responsible for short-range correlation was found to be the most stable for the system. The reliability of the approximate spin projection (AP) procedure to eliminate the high-spin component was also elucidated, comparing with the AP BS and RBS potential curves. The BS GSO (GHF) computations of several triangular systems, N(CH(2))(3), (CH(2))(3), and Mn(II)(3)O(4), were performed to obtain total energies and total spin angular momentums and effective exchange integrals (J) between local spins, which are crucial for construction of effective spin Hamiltonian models. The exact diagonalization of the Heisenberg models was also performed to depict the energy levels and magnetic susceptibility curves for triangular and kagome lattices to elucidate spin frustration effects and related quantum spin behaviors. Implications of the computational results have been discussed in relation to magnetic properties of several triangular and kagome systems synthesized recently and the superconductivity of triangular systems discovered recently.

A resonating broken symmetry configuration interaction approach for double-exchange magnetic systems.

The correct description for ion-radical systems has recently attracted much attention from density functional theory (DFT) researchers. Although several hybridization schemes using exact (Hartree-Fock) exchange and DFT exchange-correlation functionals have been proposed, it has been reported that such treatments do not work for the description of ion-radical systems. In this study we show that combining the exact exchange term in the Kohn-Sham DFT (or the Hartree-Fock equation) with the following resonating configuration interaction method is effective for the description of double-exchange type molecular magnetic interactions. The results are analyzed in relation to the 'many-electron self-interaction' concept that was recently proposed by DFT researchers.

Characterization of non-linear relationship between total and unbound serum concentrations of valproic acid in epileptic children.

OBJECTIVE: To establish a regression equation to properly estimate the unbound serum concentration of valproic acid (VPA) from its total serum concentration; the relationship between total and unbound serum VPA concentrations was retrospectively characterized. METHODS: Data were obtained from the clinical examination records that were routinely archived during therapeutic drug monitoring. The screening encompassed 342 records of 108 paediatric patients whose total and unbound VPA concentrations had been determined. The relationship between total and unbound VPA concentrations was characterized according to the Langmuir equation by taking account of inter-individual variability with the nonmem program. RESULTS: The total VPA concentration (C(t)) in the screened patients ranged from 5.5 to 179.8 microg/mL, and the unbound VPA concentration (C(f)) increased in a non-linear manner as the total VPA concentration increased. Taking account of the effects of antiepileptics concurrently administered, the VPA dissociation constant (K(d)) and maximum binding site concentration (B(m)) were 7.8 +/- 0.7 and 130 +/- 4.5 microg/mL respectively, for the regression equation, C(t) = C(f) + B(m) x C(f)/(K(d) + C(f)). An alteration in the unbound concentration was seen in patients who were treated with the combination of VPA and ethosuximide and in those who received two additional antiepileptics. CONCLUSIONS: A regression equation for estimation of the unbound VPA concentration, based on total VPA concentration collected during routine therapeutic drug monitoring was established. Use of two additional antiepileptics and ethosuximide treatment was considered as potential factors affecting unbound VPA concentration.

Quasi-one-dimensional molecular magnets based on derivatives of (fluorobenzyl)pyridinium with the [M(mnt)2] monoanion (M = Ni, Pd or Pt; mnt2- = maleonitriledithiolate): syntheses, crystal structures and magnetic properties.

The syntheses, structural characterizations and magnetic behaviors of three new complexes, 1-(3',4',5'-trifluorobenzyl)pyridinium [M(mnt)2]- [M = Ni (1), Pd (2) or Pt (3)], are reported. These complexes are isomorphous and their prominent structural character is that the [M(mnt)2]- anions form columnar stacks, in which the dimerization was observed. Complexes 2 and 3 are diamagnetic, while 1 possesses an energy gap of 2474 K. For crystal 4, 1-(4'-fluorobenzyl)pyridinium [Ni(mnt)2] (its structure and magnetic susceptibility were briefly reported earlier), the magnetic behavior can be divided into two regimes, namely, weakly ferromagnetic coupling above 93 K and strongly antiferromagnetic coupling below 93 K. A transition occurs at 93 K which switches the magnetic exchange nature from ferromagnetic to antiferromagnetic. A sharp thermal abnormality with lambda-shape, associated with the transition, appears from its heat capacity measurement to indicate that the transition is first order. The temperature dependences of the superlattice diffractions revealed the existence of the pretransitional phenomena up to at least 140 K. The unusual magnetic behavior of 4, such as the origin of the ferromagnetic interaction in the high temperature phase and what causes the spin transition, are discussed further.

Structural and magnetic investigations for the doping effect of nonmagnetic impurity on the spin-Peierls-like transition in a quasi-one-dimensional magnet: 1-(4'-nitrobenzyl)pyridinium bis(maleonitriledithiolato)nickelate.

A nonmagnetic compound, [NO(2)BzPy][Au(mnt)(2)] (NO(2)BzPy(+) = 1-(4'-nitrobenzyl)pyridinium; mnt(2-) = maleonitriledithiolate), was synthesized and characterized structurally, which is isostructural with [NO(2)BzPy][Ni(mnt)(2)] that is a quasi-one-dimensional magnet and possesses a spin-Peierls-like transition with J = 192 K in the gapless state and spin energy gap = 738 K in the dimerization state, respectively. Further, ten nonmagnetic impurity doped compounds with a formula [NO(2)BzPy][Au(x)Ni(1-x)(mnt)(2)] (x = 0.01-0.73) were prepared and investigated by crystal structural determinations and magnetic susceptibility measurements. The nonmagnetic doping causes the suppression of the spin transition with an average rate of 221(12) K/percentage of dopant concentration. From the plots of chi(m)-T, the transition collapse (the characteristic of the transition is the sudden drop of chi(m) upon cooling, and the disappearance of this characteristic is considered as the criterion for the transition collapse) is estimated at around x > 0.27. In heavier doped system x = 0.49, the spin gap vanishes and a gapless phase is achieved again.

Design of a magnetic bistability molecular system constructed by H-bonding and pi...pi-stacking interactions.

Crystal structures and magnetic properties were determined for two novel polymorphs of the complex [H2DABCO][Ni(mnt)2] [(H2DABCO)2+ = diprotonated 1,4-diazabicyclo[2.2.2]octane; mnt2- = maleonitriledithiolate]. For each polymorph, anions form a layered structure in which two kinds of dimers were observed. The adjacent anionic sheets are held together by cations via H-bonding interactions between protons of cations and CN groups of anions. Two polymorphs possess spin bistability; namely, upon cooling, a magnetic transition happens at around 120 K with about 1 K hysteresis on heating for the alpha phase and at 112 K with about 10 K hysteresis for the beta phase. Above the transition, the magnetic behaviors of two polymorphs can be approximately interpreted by a singlet-triplet model of an antiferromagnetically coupled S = 1/2 dimer, which is supported by the crystal structures and spin dimer analyses based on extended Hückel molecular orbital calculations.

Structural phase transition driven by spin-lattice interaction in a quasi-one-dimensional spin system of [1-(4'-iodobenzyl)pyridinium][Ni(mnt)2].

Crystal structures and magnetic properties were determined for two novel compounds, [1-(4'-iodobenzyl)pyridinium][M(mnt)2] (mnt2- = maleonitriledithiolate; M = Ni (1) or Cu (2)). At room temperature, single crystals of 1 and 2 were isostructural, featuring the formation of segregated columnar structures with regular stacks of cations and anions. For crystal 1, a magnetic transition was observed at approximately 120 K; furthermore, its magnetic behavior was consistent with that of a regular Heisenberg antiferromagnetic (AFM) chain of S = 1/2 in the high-temperature phase (HT phase) and that of a spin-gap system in the low-temperature phase (LT phase). Such a phenomenon is similar to the spin-Peierls transition. However, the crystal structure of 1 in the LT phase at 100 K revealed that its structural transition is associated with the magnetic transition. Because crystal 2 (S = 0) did not exhibit a structural transition, the structural transition of 1 is driven by spin-lattice interaction.

Gene expression relevant to osteoclastogenesis in the synovium and bone marrow of mature rats with collagen-induced arthritis.

OBJECTIVES: To investigate gene expression relevant to osteoclastogenesis in the synovium and bone marrow during the development of collagen-induced arthritis (CIA) in mature rats. METHODS: Total messenger RNAs (mRNAs) were obtained from CIA synovium and bone marrow after immunization. First, reverse transcriptase-polymerase chain reactions (RT-PCR) were carried out to detect the mRNA encoding receptor activator of NF-kappaB (RANK), RANK ligand (RANKL), osteoprotegerin (OPG), tumour necrosis factor-alpha (TNF-alpha), interleukin (IL)-1beta, IL-6 and the osteoclast markers tartrate-resistant acid phosphatase (TRAP) and cathepsin K. Secondly, the genes detected clearly by RT-PCR were quantified using real-time PCR. RESULTS: In the synovium, expression of all genes was confirmed by specific single bands in RT-PCR. In real-time PCR, the expression levels of TNF-alpha, IL-1beta, IL-6, RANKL, TRAP and cathepsin K mRNA increased, whereas the expression levels of RANK and OPG were unchanged and decreased respectively. RANKL expression was highly correlated with the two osteoclast markers. In the bone marrow, RT-PCR did not clearly detect the expression of IL-6, RANKL or OPG mRNA. Quantitative real-time PCR showed that TNF-alpha, RANK and TRAP mRNA expression did not change significantly with time, and that IL-1beta and cathepsin K changed slightly compared with those in the synovium. CONCLUSIONS: In the early stages of arthritis, synovial RANKL is closely involved in osteoclastogenesis, and various changes in synovial cytokines, including down-regulation of OPG, probably accelerate osteoclast formation. In contrast, cytokine mRNA in the bone marrow showed little fluctuation. We suggest that synovial cytokines affect osteoclastogenesis not only in the synovium but in the bone marrow.

Expression and activity of Apaf1 and caspase-9 in granulosa cells during follicular atresia in pig ovaries.

Apoptosis in granulosa cells plays a crucial role in ovarian follicular atresia, but the intracellular regulating mechanism, especially the mitochondrion-dependent apoptosis signalling pathway, is still largely unknown. This study examined whether the mitochondrial pathway is associated with granulosa cell apoptosis during atresia in pig ovaries. Both mRNAs of caspase-9 and apoptotic protease-activating factor 1 (Apaf1), which are major signal transducing components in the mitochondrial pathway, were detected in granulosa cells in healthy, early atretic and progressed atretic follicles by RT-PCR. No changes in the expression of Apaf1 mRNA were seen during follicular atresia, but the expression of caspase-9 mRNA increased during atresia. Apaf1 protein was steadily detected in granulosa cells prepared from healthy, early atretic and progressed atretic follicles by western blot analysis, but high expression of the precursor of caspase-9 (procaspase-9) was detected only in granulosa cells of healthy follicles. Decreased procaspase-9 protein was demonstrated during follicular atresia. Proteolytic activity of caspase-9 increased during atresia, in agreement with the diminution of procaspase-9 protein. Intensive expression of caspase-9 mRNA was demonstrated in the granulosa cells of early atretic and progressed atretic follicles but not in those of healthy follicles. These results indicate that the mitochondrial signalling pathway, which is mediated by Apaf1 and caspase-9, plays a crucial role in determining the fate of granulosa cells during atresia in pig ovaries.

Immunohistochemical study of p53, c-erbB-2, and PCNA in barrett's esophagus with dysplasia and adenocarcinoma arising from experimental acid or alkaline reflux model.

PURPOSE: An immunohistochemical study of p53, c-erbB-2, and proliferating cell nuclear antigen (PCNA) in Barrett's esophagus with dysplasia and adenocarcinoma, arising from experimental acid or alkaline reflux, was performed in dogs. METHODS: Cardiectomy was performed in group A (n = 26) as an acid reflux model, and total gastrectomy was performed in group B (n = 24) as an alkaline reflux model. After surgery, the esophageal mucosa was observed and biopsied endoscopically every 3 months over a period of 6 years. Immunohistochemical staining of p53. c-erbB-2, and PCNA was performed, using biopsied specimens. RESULTS: In group A, Barrett's esophagus developed in 14 of the 26 dogs. Low-grade dysplasia occurred in 5 of the 26 dogs, and in 1 of these 5 dogs, it developed into high-grade dysplasia. In this animal, adenocarcinoma arose 63 months after the operation. In group B, Barrett's esophagus developed in 10 of the 24 dogs. Low-grade dysplasia was observed in 4 of the 24 dogs. In 1 of these 4 dogs, the dysplasia became high-grade and adenocarcinoma occurred 66 months after the operation. In group A, PCNA was positive in adenocarcinoma; the PCNA labeling index (LI) was 58. c-erbB-2 and p53 were negative in all animals in group A. In group B, PCNA was positive in Barrett's esophagus with high-grade dysplasia and adenocarcinoma; the PCNA LI was 77. p53 was positive in adenocarcinoma. c-erbB-2 was negative in adenocarcinoma. CONCLUSIONS; The results of this study provided evidence of the dysplasia-carcinoma sequence arising from alkaline reflux, as well as from acid reflux. To the best of our knowledge, this is the first report of the use of an alkaline reflux model and a 6-year study using dogs to observe the course of Barrett's esophagus.

Polymorphisms of two fucosyltransferase genes (Lewis and Secretor genes) involving type I Lewis antigens are associated with the presence of anti-Helicobacter pylori IgG antibody.

Helicobacter pylori attach to the gastric mucosa with adhesin, which binds to Lewis b (Le(b)) or H type I carbohydrate structures. The Secretor (Se) gene and Lewis (Le) gene are involved in type I Le antigen synthesis. The present study was performed to investigate the possibility that Se and Le gene polymorphisms alter the risk of H. pylori infection. Two hundred thirty-nine participants were genotyped for Se and Le and tested for the presence of anti-H. pylori IgG antibodies. Using the normal gastric mucosa from 60 gastric cancer patients, we assessed immunohistochemically whether type I Le antigen expression depended on the Se and Le genotypes. The H. pylori infection rate was positively associated with the number of Se alleles (se/se group, 45.1%; Se/se group, 64.6%; and Se/Se group, 73.3%) and negatively associated with the number of Le alleles (le/le group, 76.4%; Le/le group, 68.3%; and Le/Le group, 55.6%). When the subjects were classified into three groups [low risk, (se/se, Le/Le) genotype; high risk, (Se/Se, le/le), (Se/Se, Le/le), and (Se/se, le/le) genotypes; moderate risk, other than low- or high-risk group], the odds ratio relative to the low-risk group was 3.30 (95% confidence interval, 1.40-7.78) for the moderate-risk group and 10.33 (95% confidence interval, 3.16-33.8) for the high-risk group. Immunohistochemical analysis supported the finding that Se and Le genotypes affected the expression of H. pylori adhesin ligands. We conclude that Se and Le genotypes affect susceptibility to H. pylori infection.

A remodeling system of the 3'-sulfo-Lewis a and 3'-sulfo-Lewis x epitopes.

It has been reported that the chemically synthesized 3'-sulfo-Le(a) and 3'-sulfo-Le(x) epitopes have a high potential as a ligand for selectins. To elucidate the physiological functions of 3'-sulfated Lewis epitopes, a remodeling system was developed using a combination of a betaGal-3-O-sulfotransferase GP3ST, hitherto known alpha1,3/1,4-fucosyltransferases (FucT-III, IV, V, VI, VII, and IX) and arylsulfatase A. The pyridylaminated (PA) lacto-N-tetraose (Galbeta1-3GlcNAcbeta1-3Galbeta1-4Glc) was first converted to 3'-sulfolacto-N-fucopentaose II (sulfo-3Galbeta1-3(Fucalpha1-4)GlcNAcbeta1-3Galbeta1-4Glc)-PA by sequential reactions with GP3ST and FucT-III. The 3'-sulfolacto-N-fucopentaose III (sulfo-3Galbeta1-4(Fucalpha1-3)GlcNAcbeta1-3Galbeta1-4Glc)-PA was then synthesized from lacto-N-neotetraose (Galbeta1-4GlcNAcbeta1-3Galbeta1-4Glc)-PA by GP3ST and FucT-III, -IV, -V, -VI, -VII, or -IX in a similar manner. The substrate specificity for the 3'-sulfated acceptor of the alpha1,3-fucosyltransferases was considerably different from that for the non-substituted and 3'-sialylated varieties. When the GP3ST gene was introduced into A549 and Chinese hamster ovary cells expressing FucT-III, they began to express 3'-sulfo-Le(a) and 3'-sulfo-Le(x) epitopes, respectively, suggesting that GP3ST is responsible for their biosynthesis in vivo. The expression of the 3'-sialyl-Le(x) epitope on Chinese hamster ovary cells was attenuated by the introduction of GP3ST gene, indicating that GP3ST and alpha2,3-sialyltransferase compete for the common Galbeta1-4GlcNAc-R oligosaccharides. Last, arylsulfatase A, which is a lysosomal hydrolase that catalyzes the desulfation of 3-O-sulfogalactosyl residues in glycolipids, was found to hydrolyze the sulfate ester bond on the 3'-sulfo-Le(x) (type 2 chain) but not that on the 3'-sulfo-Le(a) (type 1 chain). The present remodeling system might be of potential use as a tool for the study of the physiological roles of 3'-sulfated Lewis epitopes, including interaction with selectins.

Epigenetic silencing of PEG3 gene expression in human glioma cell lines.

Genomic imprinting, the phenomenon in which alleles of genes are expressed differentially depending on their parental origins, has important consequences for mammalian development, and disturbance of normal imprinting leads to abnormal embryogenesis and some inherited diseases and is also associated with various cancers. In the context of screening for novel imprinted genes on human chromosome 19q13.4 with mouse A9 hybrids, we identified a maternal allele-specific methylated CpG island in exon 1 of paternally expressed imprinted gene 3 (PEG3), a gene that exhibits paternal allele-specific expression. Because PEG3 expression is downregulated in some gliomas and glioma cell lines, despite high-level expression in normal brain tissues, we investigated whether the loss of PEG3 expression is related to epigenetic modifications involving DNA methylation. We found monoallelic expression of PEG3 in all normal brain tissues examined and five of nine glioma cell lines that had both unmethylated and methylated alleles; the remaining four glioma cell lines exhibited gain of imprinting with hypermethylated alleles. In addition, treatment of glioma cell lines with the DNA demethylating agent 5-aza-2'-deoxycytidine reversed the silencing of PEG3 biallelically. In this article, we report that the epigenetic silencing of PEG3 expression in glioma cell lines depends on aberrant DNA methylation of an exonic CpG island, suggesting that PEG3 contributes to glioma carcinogenesis in certain cases.

Construction of 700 human/mouse A9 monochromosomal hybrids and analysis of imprinted genes on human chromosome 6.

As an in vitro assay system for the identification of human imprinted genes, a library of human/mouse A9 monochromosomal hybrids containing a single, intact bsr-tagged human chromosome of known parental origin, derived from normal human fibroblasts, has been previously generated by microcell-mediated chromosome transfer (MMCT). To supplement this assay system, we constructed additional 700 A9 monochromosomal hybrids, using a pSTneo or pPGKneo selection marker. To validate the A9 hybrids, we screened them with chromosome-specific polymorphic markers, and identified the hybrids containing either human chromosome 6, 7, 14, 18, or 21 of known parental origin. Matching paternal and maternal chromosome pairs of A9 hybrids were identified for chromosomes 6, 7, 14, and 18. The paternal-specific expression of ZAC (zinc finger protein, which regulates apoptosis and cell cycle arrest) and HYMAI (hydatidiform mole-associated and imprinted transcript), and the maternal-specific methylation of a CpG island within an imprinted domain on human chromosome 6q24, were maintained in A9 hybrids. For an example, we profiled the expression of expressed sequence tags (ESTs) and the methylation of CpG islands in the 300-kb imprinted domain around 6q24, which may be associated with cancers and transient neonatal diabetes mellitus (TNDM). Thus, the 700 A9 hybrids should be useful for various aspects of imprinting studies.

Molecular cloning and characterization of UDP-GlcNAc:lactosylceramide beta 1,3-N-acetylglucosaminyltransferase (beta 3Gn-T5), an essential enzyme for the expression of HNK-1 and Lewis X epitopes on glycolipids.

A new member of the UDP-N-acetylglucosamine:beta-galactose beta1,3-N-acetylglucosaminyltransferase (beta3Gn-T) family having the beta3Gn-T motifs was cloned from rat and human cDNA libraries and named beta3Gn-T5 based on its position in a phylogenetic tree. We concluded that beta3Gn-T5 is the most feasible candidate for lactotriaosylceramide (Lc(3)Cer) synthase, an important enzyme which plays a key role in the synthesis of lacto- or neolacto-series carbohydrate chains on glycolipids. beta3Gn-T5 exhibited strong activity to transfer GlcNAc to glycolipid substrates, such as lactosylceramide (LacCer) and neolactotetraosylceramide (nLc(4)Cer; paragloboside), resulting in the synthesis of Lc(3)Cer and neolactopentaosylceramide (nLc(5)Cer), respectively. A marked decrease in LacCer and increase in nLc(4)Cer was detected in Namalwa cells stably expressing beta3Gn-T5. This indicated that beta3Gn-T5 exerted activity to synthesize Lc(3)Cer and decrease LacCer, followed by conversion to nLc(4)Cer via endogenous galactosylation. The following four findings further supported that beta3Gn-T5 is Lc(3)Cer synthase. 1) The beta3Gn-T5 transcript levels in various cells were consistent with the activity levels of Lc(3)Cer synthase in those cells. 2) The beta3Gn-T5 transcript was presented in various tissues and cultured cells. 3) The beta3Gn-T5 expression was up-regulated by stimulation with retinoic acid and down-regulated with 12-O-tetradecanoylphorbol-13-acetate in HL-60 cells. 4) The changes in beta3Gn-T5 transcript levels during the rat brain development were determined. Points 2, 3, and 4 were consistent with the Lc(3)Cer synthase activity reported previously.