Pioglitazone improves the phenotype and molecular defects of a targeted Pkd1 mutant

Mutations of either PKD1 or PKD2 are associated with autosomal dominant polycystic kidney disease (ADPKD). The molecular function of the gene product of PKD1, polycystin-1, in vitro has been elucidated recently, but the molecular pathological consequences of the loss of polycystin-1 in vivo have remained unclear. We have generated a mouse with a targeted deletion of exons 2-6 of Pkd1 to study the molecular defects in Pkd1 mutants. Homozygote embryos (Pkd1(-/-)) developed hydrops, cardiac conotruncal defects and renal cystogenesis. Total protein levels of beta-catenin in heart and kidney and c-MYC in heart were decreased in Pkd1(-/-) embryos. In the kidneys of Pkd1(-/-), the expression of E-cadherin and PECAM in basolateral membranes of renal tubules was attenuated, and tyrosine phosphorylation of epidermal growth factor receptor and Gab1 were constitutively enhanced when cystogenesis started on embryonic day (E) 15.5-16.5. Maternally administered pioglitazone, a thiazolidinedione compound, resolved these molecular defects of Pkd1(-/-). Treatment with pioglitazone improved survival of Pkd1(-/-) embryos and ameliorated the cardiac defects and the degree of renal cystogenesis. Long-term treatment with pioglitazone improved the endothelial function of adult Pkd1(+/-). These data indicated that molecular defects observed in Pkd1(-/-) embryos contributed to the pathogenesis of ADPKD and that thiazolidinediones had a compensatory effect on the pathway affected by the loss of polycystin-1. Pathways activated by thiazolidinediones may provide new therapeutic targets in ADPKD.     

Peptide analysis,stability studies,and structural modeling explain contradictory peptide motifs and unique properties of the NOD mouse MHC class II molecule H2-A(g7)

The MHC class II molecule H2-A(g7) is the chief genetic determinant in insulin-dependent diabetes mellitus of the non-obese diabetic (NOD) mice. Poor peptide binding ability, as well as presentation of a unique subset of peptides by this molecule was suggested to promote autoimmunity in this strain. However, several laboratories have presented results in favor of an H2-A(g7) molecule that can avidly bind many different peptides. The crystal structures of H2-A(g7) in complex with two different peptides did not completely resolve this issue. To analyze the peptide binding capacity and the motif requirements of H2-A(g7), we eluted natural ligands from purified H2-A(g7) molecules isolated from the H2-A(g7)-transfected M12-C3 cells. A low peptide yield dominated by a few peptide ligands was found. Pool sequencing and alignment of individual ligands on the basis of molecular modeling revealed a peptide-binding motif with basic/aliphatic/small hydrophilic amino acids at relative position 1 (p1), aliphatic amino acids at p4, Ala at p6, and acidic amino acids and Ser/Gly at p9, as well as acidic residues at p10/11. Though weak, the binding of individual ligands, as well as the importance of an acidic C-terminal residue was confirmed by peptide binding studies to isolated H2-A(g7) molecules. Furthermore, the H2-A(g7) molecule incompletely dissociated into its constituent chains in SDS-electrophoresis under nonreducing conditions. This provides additional evidence of its weak affinity for peptides, which probably arises from the combination of beta56His/beta57Ser/beta78Ala and other unique H2-A(g7) residues in contact with the antigenic peptide. These results allow a better understanding of the role of this molecule in the development of autoimmunity and the identification of epitopes relevant to diabetes.     

Synthesis and properties of thermotropic liquid-crystalline polymers containing transition metals

New thermotropic, liquid-crystalline polyesters ( 5a, b, 6a, b ) with mesogenic bipyridinediyl units were synthesized. They form complexes with Fe(II) and Cu(II) salts. Homopolyesters 5a, b , containing no metal or less than a certain amount of metal, are smectic. A nematic mesophase was observed in the case of copolyester 6a, b . The structural properties of a polyester complex 5a with Cu, as crystal, liquid-crystal and isotropic liquid, were examined by ESR.     

医疗数据交换规格MML(Medical Markup Language)3.0汉化版的制作

MML(Medical Markup Language)是一套不同医疗设施间的数据交换规格。于1995年在日本被开发。MML从版本2.21开始使用XML(eXtensible Markup Language)作为标记语言。而最新版本3.0又遵循HL7Clincal Document Architecture(CDA),包含14模块和36个数据定义表格。目前在中国,还没有一个使用XML来结构整个病历内容的规格。鉴于MML的柔韧性,我们制作了一个基于3.0版本的汉化版。日本与中国虽然诊疗流程、病历记录的内容等都很相似,但是也有一些,比如民族的表现、中医诊断分类,医师资格分类等都是日本不存在的或者分类不同的信息。另外,因为国情不同,医疗保险制度也完全不同。为了使MML能在中国的医院适用,我们追加和更改了12个数据定义表格,并重新制作了医疗保险信息模块。MML汉化版不止是一个对原规格的翻译和说明,它还考虑了本地的需要。因此,使用MML汉化版在中国的医疗设施间进行医疗数据交换已经成为可能。     

A novel action of stargazin as an enhancer of AMPA receptor activity

Stargazin (γ-2) is disrupted in the ataxic and epileptic mutant mouse, stargazer (stg). The striking defect in the stg cerebellum is the lack of functional AMPA receptors on granule cells. Recently, it has been reported that γ-2 and its related molecules are crucial for the surface expression, synaptic targeting and recycling of AMPA receptors, being termed collectively as the transmembrane AMPA receptor regulatory proteins (TARPs). However, it is still unclear whether TARPs directly modulate AMPA receptor activity. Here we report that coexpression of GluR1 (GluR1) with γ-2 using HEK293 cells and Xenopus oocytes markedly enhanced glutamate-induced currents. This effect was far beyond the increase of AMPA receptor surface expression and accompanied by increased glutamate affinity and subunit cooperativity. Other member of TARPs (γ-3, γ-4, and γ-8) also enhanced the current response through the AMPA receptors. The enhancing effect by γ-2 coexpression was further observed for homomeric GluR2 (GluR2) channels, which, when expressed alone, are known to produce only a small or negligible current response. These results suggest that γ-2 not only promotes AMPA receptor surface expression but also directly modulates AMPA receptor activity.     

Ocular tracking of moving targets: effects of perturbing the background

Primates are able to track a moving target with their eyes, even when the target is seen against a stationary textured background. In this situation, the tracking eye movement induces motion of the background images on the retina (reafference) that competes with the motion of the target's retinal image, potentially disrupting the tracking of the target. Previous work on humans reported that brief perturbations of the background in the opposite direction to pursuit were much less disruptive than perturbations in the same direction as pursuit. Furthermore, if the background moved together with the pursuit target--so as to effectively eliminate the reafference--then the effects of a subsequent background perturbation showed less dependence on direction. This suggested that the direction selectivity to background perturbations during pursuit against a stationary background was due, at least in part, to the prior motion of the background secondary to the pursuit. We now report similar findings in monkeys, and in addition, have investigated the effect of moving the background while the animal was fixating a stationary target. In this situation, the ocular tracking responses to subsequent brief perturbations of the moving background were weaker when the perturbations were in the same direction as the prior background motion than when in the opposite direction. This suggests that the selective insensitivity to the reafferent visual input associated with pursuit across a stationary background is, at least in part, independent of pursuit per se and attributable to a progressive reduction in the sensitivity to sustained background motion.     

Neuronal protein NP25 interacts with F-actin

Neuronal protein NP25 is a neuron-specific protein present in highly differentiated neural cells, but its functional properties have not been well characterized. NP25 shows high amino acid sequence homology with the smooth muscle cell cytoskeleton-associated proteins, SM22, mp20, and calponin. To gain an insight into the biological functions of NP25, we first examined its subcellular localization in the human neuroblastoma cell line, SK-N-SH. NP25 diffusely distributed in the cytoplasm and fiber-like staining was also observed. It showed that NP25 co-localized with F-actin on stress fibers. A co-sedimentation assay demonstrated that NP25 bound to filamentous actin. Further investigations using fluorescence resonance energy transfer (FRET) technique revealed intracellular binding of NP25 and actin. The significance of the interaction between NP25 and F-actin is discussed.     

Loss of heterozygosity on chromosome 9q22.3 in microdissected basal cell carcinomas around the Semipalatinsk Nuclear Testing Site, Kazakhstan

A high incidence of skin cancers has been noted around the Semipalatinsk Nuclear Testing Site (SNTS) in Kazakhstan. Recently, basal cell carcinoma (BCC) susceptibility genes, human homolog of the Drosophila pathed gene (PTCH), and the xeroderma pigmentosa group A-complementing gene (XPA), have been cloned and localized on chromosome 9q22.3. To clarify the effect of low-dose irradiation on the occurrence of BCC, we used microdissection and polymerase chain reaction to identify loss of heterozygosity (LOH) at 9q22.3 using BCC samples obtained from this region. Ten Japanese samples were analyzed as controls. LOH with at least 1 marker was identified in 5 of 14 cases from around SNTS, whereas only 1 case with 1 marker was identified among the 10 Nagasaki cases. The total number of LOH alleles from SNTS (8 of 45) was significantly higher than the number from Nagasaki (1 of 26) (P = 0.03). The higher incidence of LOH on 9q22.3 in BCC from around SNTS suggests involvement of chronic low-dose irradiation by fallout from the test site as a factor in the cancers.     

XIdax, an inhibitor of the canonical Wnt pathway, is required for anterior neural structure formation in Xenopus.

Wnt signaling pathways are involved during various stages in the development of many species. In Xenopus, the accumulation of beta-catenin on the dorsal side of embryo is required for induction of the organizer, while the head structure formation requires inhibition of Wnt signaling. Here, we report a role for xIdax, a negative regulator of Wnt signaling. XIdax is expressed in neural tissues at the neurula stage, and in the restricted region of the tadpole brain. Ectopic expression of xIdax inhibits the target gene expression, suggesting that xIdax can inhibit canonical Wnt signaling. To examine the function of xIdax, a morpholino oligo for xIdax (xIdaxMO) was designed. An injection into an animal pole cell caused a loss of forebrain. The anterior neural marker expression is decreased in xIdaxMO-injected embryo, suggesting that xIdax is required for anterior neural development. Moreover, a negative regulator that acts downstream of xIdax rescued this defect. We propose that Idax functions are dependent on the canonical Wnt pathway and are crucial for the anterior neural development.     

C and V proteins of Sendai virus target signaling pathways leading to IRF-3 activation for the negative regulation of interferon-beta production

We here report a molecular basis for downregulation of interferon (IFN)-beta production by V and C proteins of Sendai virus (SeV). The infection of HeLa cells with SeV poorly induced IFN-beta even if the expression of C/C' was disrupted. In contrast, when the expression of C/C'/Y1/Y2 or V/W was disrupted, SeV infection strongly induced IFN-beta production and significantly activated the interferon regulatory factor (IRF)-3 pathway. The independent expression of C or V inhibited the double-stranded (ds) RNA- or Newcastle disease virus (NDV)-induced activation of IRF-3 and NF-kappa B, as well as the IFN-beta promoter. This inhibitory effect was also observed when Y1, Y2, or a C-terminal half fragment (aa 85-204) of C was independently expressed. Phosphorylation and homodimer formation of IRF-3 were suppressed not only in cells infected with SeV capable of expressing both C/C'/Y1/Y2 (or Y1/Y2) and V/W, but also in HeLa cells constitutively expressing Y1. These results suggest that C, Y1, Y2, and V block signaling pathways leading to IRF-3 activation to downregulate IFN-beta production.