Short half-lives of ataxia-associated aprataxin proteins in neuronal cells

Early-onset ataxia with ocular motor apraxia and hypoalbuminemia (EAOH)/ataxia with oculomotor apraxia type 1 (AOA1) is caused by mutations in the gene encoding aprataxin (APTX). Although several in vitro findings proposed that impaired enzymatic activities of APTX are responsible for EAOH/AOA1, potential instability of mutant proteins has also been suggested as the pathogenesis based on in vivo finding that mutant proteins are almost undetectable in EAOH/AOA1 tissues or cells. The present study aimed to experimentally prove instability of mutant proteins in neuronal cells, the cell type preferentially affected by this disease. Results of pulse-chase experiments demonstrated that all of the disease-associated mutants had extremely shorter half-lives than the WT. We further found that mutants were targeted for rapid proteasome-mediated degradation. These results help establish pathogenic and physiological protein characteristics of APTX in neuronal cells.     

Characterization of factor XII Tenri, a rare CRM-negative factor XII deficiency

Factor XII Tenri (Y34C), a rare cross-reacting material (CRM)-negative factor XII deficiency, was identified in a 71-yr-old Japanese woman with angina pectoris. In the patient's plasma, factor XII activity and antigen levels were only 1.6% and 5.0%, respectively, of those seen in a normal subject. Immunoblot analysis showed that the secreted factor XII Tenri existed not only as a monomer (76 kDa), but also in complexes with apparent molecular weights of approximately 115, 140, 190, 215, and 225 kDa. After reduction with 2-mercaptoethanol, the factor XII Tenri contained in the complexes was completely converted to monomeric form on immunoblot patterns. It appeared that some of the secreted factor XII Tenri formed several types of disulfide-linked complexes, including a factor XII-alpha1-microglobulin complex, through a newly generated Cys residue. The monomeric form of factor XII Tenri, like normal factor XII, was degraded into 2 major fragments with molecular weights of approximately 45 kDa and 30 kDa following mixing with activated partial-thromboplastin-time measuring reagent (cephalin and ellagic acid), whereas the factor XII Tenri that formed the complexes was not. This indicates that the factor XII Tenri present in disulfide-linked complexes with other proteins (and itself) is not converted to active forms, suggesting that attached proteins obstruct or delay the activation of factor XII via an inhibition of its binding to a negatively charged surface in vitro.     

A neural network-based similarity index for clustering DNA microarray data

A common approach to the analysis of gene expression data is to define clusters of genes that have similar expression. A critical step in cluster analysis is the determination of similarity between the expression levels of two genes. We introduce a neural network-based similarity index as a non-linear similarity index and compare the results with other proximity measures for Saccharomyces cerevisiae gene expression data. We show that the clusters obtained using Euclidean distance, correlation coefficients, and mutual information were not significantly different. The clusters formed with the neural network-based index were more in agreement with those defined by functional categories and common regulatory motifs.     

Renal glomerulogenesis in medaka fish, Oryzias latipes

We provide an overview of glomerulogenesis in medaka from the embryo to the adult by means of in situ hybridization with the wt1 gene as a marker as well as histology and three-dimensional images. The pronephric glomus starts to develop in the intermediate mesoderm during early somitogenesis, is completed before hatching, and persists throughout the lifetime of the fish. Within 5 days after hatching, mesonephric glomerulus formation begins in the caudomedial end of the pronephric sinus and duct area. The number of glomeruli reaches approximately 200-300 in each kidney within 2 months after hatching. wt1 expression during nephron maturation served as a marker for the formation of the mesenchymal condensate and the nephrogenic body. Existence of mesenchymal condensates and persistence of wt1 expression in the adult kidney suggest that the mesonephros retains precursor cells that may be capable of contributing to neoglomerulogenesis during adulthood. Developmental Dynamics 237:2342-2352, 2008. (c) 2008 Wiley-Liss, Inc.     

Contribution of BCAP to maintenance of mature B cells through c-Rel

Mice deficient in the B cell adaptor for phosphoinositide 3-kinase (BCAP) have reduced numbers of mature B lymphocytes, which show defects in cell survival and proliferation. We found here that the NF-kappa B (Rel) pathway was impaired in BCAP-deficient mature B cells and that NF-kappa B target genes, indispensable for cell survival and division, were not induced in response to B cell receptor (BCR) stimulation. Among the NF-kappa B (Rel) family, expression of c-Rel was specifically reduced in BCAP-deficient B cells. Retrovirus-mediated reintroduction of c-Rel restored the pool size of immunoglobulin (Ig)M(lo)IgD(hi) mature B cells in the spleen as well as proliferative responses to BCR stimulation. These results indicate BCAP is essential in the maintenance of mature B cells through functional coupling with c-Rel.     

Gain-of-function polymorphism in mouse and human Ltk: implications for the pathogenesis of systemic lupus erythematosus

Systemic lupus erythematosus (SLE), a complex multigenic disease, is a typical antibody-mediated autoimmune disease characterized by production of autoantibodies against a variety of autoantigens and immune complex-type tissue inflammation, most prominently in the kidney. Evidence suggests that genetic factors predisposing to aberrant proliferation/maturation of self-reactive B cells initiate and propagate the disease. In SLE-prone New Zealand Black (NZB) mice and their F1 cross with New Zealand White (NZW) mice, B cell abnormalities can be ascribed mainly to self-reactive CD5+ B1 cells. Our genome-wide scans to search for susceptibility genes for aberrant activation of B1 cells in these mice showed evidence that the gene, Ltk, encoding leukocyte tyrosine kinase (LTK), is a possible candidate. LTK is a receptor-type protein tyrosine kinase, belonging to the insulin receptor superfamily, and is mainly expressed in B lymphocyte precursors and neuronal tissues. Sequence and functional analyses of the gene revealed that NZB has a gain-of-function polymorphism in the LTK kinase domain near YXXM, a binding motif of the p85 subunit of phosphatidylinositol 3-kinase (PI3K). SLE patients also had this type of Ltk polymorphism with a significantly higher frequency compared with the healthy controls. Our findings suggest that these polymorphic LTKs cause up-regulation of the PI3K pathway and possibly form one genetic component of susceptibility to abnormal proliferation of self-reactive B cells in SLE.     

Asymmetries of prefrontal cortex in human episodic memory: effects of transcranial magnetic stimulation on learning abstract patterns

Functional neuroimaging suggests asymmetries of memory encoding and retrieval in the prefrontal lobes, but different hypotheses have been presented concerning the nature of prefrontal hemispheric specialization. We studied an associative memory task involving pairs of Kanji (Chinese) pictographs and unfamiliar abstract patterns. Subjects were ten Japanese adults fluent in Kanji, so only the abstract patterns represented novel material. During encoding, transcranial magnetic stimulation (TMS) was applied over the left and right dorsolateral prefrontal cortex (DLPFC). A significant (P<0.05) reduction in subsequent recall of new associations was seen only with TMS over the right DLPFC. This result suggests that the right DLPFC contributes to encoding of visual-object associations, and is consistent with a material-specific rather than a process-specific model of mnemonic function in DLPFC.     

Further evidence for excitability changes in human primary motor cortex during ipsilateral voluntary contractions

The present study aimed to further investigate whether the intracortical neural circuits within the primary motor cortex (M1) are modulated during ipsilateral voluntary finger movements. Single- and paired-pulse (interstimulus intervals, ISIs; 3 ms and 12 ms) transcranial magnetic stimulations of the left M1 were applied to elicit motor evoked potential (MEP) in the right first dorsal interosseous (Rt-FDI) muscle during voluntary contractions (10% and 30% maximum voluntary contraction) of the left FDI (Lt-FDI) muscle. F-waves of Rt-FDI muscle were recorded under these left index-finger conditions for ensuring that the excitability changes occur at the supraspinal level. MEPs were also recorded during motor imagery of the left index-finger abduction instead of overt movement. The results showed that, in single-pulse transcranial magnetic stimulation (TMS) paradigm, MEPs in Rt-FDI muscle were markedly enhanced during voluntary contractions of Lt-FDI muscle compared with the complete resting state. In paired-pulse TMS paradigm, the short intracortical inhibition was significantly reduced in proportion to increments of the ipsilateral muscle contraction, whereas the intracortical facilitation had no change. F-wave of Rt-FDI muscle was unchanged under these conditions, while MEP in Rt-FDI muscle was also enhanced during motor imagery of the left index-finger abduction. Based on the present results, it is suggested that the intracortical inhibitory neural circuits may be modulated in the transition from rest to activity of the ipsilateral homonymous muscle. The excitability changes in M1 might be induced by overflows of voluntary drive given to the ipsilateral limb, probably via the transcallosal pathway.     

Further characterization of memory T cells existing in a case of CD8 deficiency

CD8 deficiency is a rare primary immunodeficiency caused by a defect of ZAP-70, which plays a pivotal role in T cell activation. We previously reported the existence of memory phenotype-CD4+ T cells in a case of CD8 deficiency, which demonstrates that activation signals through ZAP-70 are not essential to the phenotypic conversion of T cells from "naive" to "memory." In this study, we further characterized CD45RO+ T cells in a CD8 deficient patient. We showed that the patient's CD45RO+ T cell population had a wide variety of T cell receptor Vbeta-chain gene usage, and contained few clonally expanded T cells, while many clonally expanded T cells were present in the memory T cell population of age-matched healthy children. These results suggest that various kinds of antigens were involved in the differentiation of the patient's T cells, and that the differentiation into memory T cells was not accompanied by profound T cell proliferation. Moreover, our findings confirmed that the patient's CD45RO+CD4+ T cells had acquired effector-cytokine producing ability, indicating that there exists an alternative activation pathway which is independent of ZAP-70 for the acquisition of effector-cytokine producing ability.     

Observation of cavitation in a mechanical heart valve in a total artificial heart

Recently, cavitation on the surface of mechanical heart valves has been studied as a cause of fractures occurring in implanted mechanical heart valves. The cause of cavitation in mechanical heart valves was investigated using the 25 mm Medtronic Hall valve and the 23 mm Omnicarbon valve. Closing of these valves in the mitral position was simulated in an electrohydraulic totally artificial heart. Tests were conducted under physiologic pressures at heart rates from 60 to 100 beats per minute with cardiac outputs from 4.8 to 7.7 L/min. The disk closing motion was measured by a laser displacement sensor. A high-speed video camera was used to observe the cavitation bubbles in the mechanical heart valves. The maximum closing velocity of the Omnicarbon valve was faster than that of the Medtronic Hall valve. In both valves, the closing velocity of the leaflet, used as the cavitation threshold, was approximately 1.3-1.5 m/s. In the case of the Medtronic Hall valve, cavitation bubbles were generated by the squeeze flow and by the effects of the venturi and the water hammer. With the Omnicarbon valve, the cavitation bubbles were generated by the squeeze flow and the water hammer. The mechanism leading to the development of cavitation bubbles depended on the valve closing velocity and the valve stop geometry. Most of the cavitation bubbles were observed around the valve stop and were generated by the squeeze flow.