Type-specific evolution of amyloid plaque and angiopathy in APPsw mice

To clarify how Aβ deposits start in the brain, we examined the early to late stages of senile plaques and amyloid angiopathy in APPsw mice. All types of human senile plaques were observed in the mouse brains. The premature forms of cored plaques appeared first in the cerebral cortex of mice at 7–8 months old. Then, amyloid angiopathy emerged, followed by diffuse plaques consisting of Aβ1–42. Modifications of the N-terminus of Aβ were late phase phenomena. The premature forms of cored plaques were composed of central Aβ1–40 amyloid cores, surrounding amorphous Aβ1–42 deposits, and accumulation of Aβ1–42 in some peripheral cells. These cells were incorporated in amyloid cores, and these plaques developed to large cored plaques composed of Aβ1–40 and Aβ1–42. The size and number of cored plaques were increased with age. These findings indicate different evolution paths for cored plaques and diffuse plaques, and suggest the presence of a pathway that initiates with the intracellular accumulation of Aβ1–42 and leads to the development of classic plaques in human brain tissues.     

Signal transduction through interferon-gamma receptor on human eosinophils

BACKGROUND: We reported on the constitutive interferon-gamma receptor (IFN-gammaR) expression on eosinophils. But signal transduction through IFN-gammaR on eosinophils remains to be elucidated. In this study, we examined the involvement of the Jak/Stat pathway in the signaling of eosinophils after IFN-gammaR conjugation by the ligand binding. METHODS: Purified peripheral eosinophils were stimulated with IFN-gamma at 37 degrees C for 1-60 min. Tyrosine phosphorylation of IFN-gammaR, Jak1, Jak2, and Stat1alpha was examined by immunoblotting. Gel-shift assay was also examined to show the formation of Stat1alpha-DNA complexes. RESULTS: We show that binding of IFN-gamma to human eosinophils initiated a series of events that resulted in the rapid tyrosine phosphorylation of not only the IFN-gammaRalpha chain but also Jak1, Jak2, and Stat1alpha. In addition, IFN-gamma enhanced the DNA-binding activity of Stat1alpha. CONCLUSION: These data indicate that IFN-gamma affects eosinophils through its specific receptor and utilizes the Jak/Stat pathway as its mode of signaling.     

Differences in distribution of myofiber types between the supraspinatus and infraspinatus muscles of sheep

Background: The m. supraspinatus stabilizes the shoulder joint to bear the body weight, and the m. infraspinatus assists in extension and flexion of the joint in sheep. Postural muscles have many SO myofibers, whereas locomotory muscles have numerous fast-twitch myofibers. In sheep the distribution of myofiber types within the two muscles, necessary for a better understanding of postural function, remains to be clarified. Methods: Muscle samples were removed from the whole transverse sections of the dorsal, middle, and ventral compartments of the m. supraspinatus and m. infraspinatus of sheep. Myofibers were classified into FG, FOG, SO-1, and SO-2 myofibers by histochemical methods. Results: The distribution of SO myofibers changed more greatly in the m. supraspinatus (15.0–99.1%) than in the m. infraspinatus (24.5–62.3%). SO myofibers were concentrated markedly in the caudal and deep regions near the spine and fossa of the scapula in the m. supraspinatus and distributed more in the medial part than in the lateral part in the m. infraspinatus. Such changes were caused by increases in percentage of SO-2 myofibers and not SO-1 myofibers. The craniolateral regions of the m. supraspinatus and the caudolateral regions of the m. infraspinatus had many fast-twitch (FOG plus FG) myofibers suited for rapid extension and flexion of the shoulder joint. Conclusions: The m. supraspinatus has the compartmentalized, deep, and caudal regions occupied by SO myofibers, which seem to be specialized for maintenance of the joint extension. The medial region of the m. infraspinatus may assist in the joint stabilization. © 1995 Wiley-Liss, Inc.     

Encoding-related brain activity during deep processing of verbal materials: a PET study

The recent advent of neuroimaging techniques provides an opportunity to examine brain regions related to a specific memory process such as episodic memory encoding. There is, however, a possibility that areas active during an assumed episodic memory encoding task, compared with a control task, involve not only areas directly relevant to episodic memory encoding processes but also areas associated with other cognitive processes for on-line information. We used positron emission tomography (PET) to differentiate these two kinds of regions. Normal volunteers were engaged in deep (semantic) or shallow (phonological) processing of new or repeated words during PET. Results showed that deep processing, compared with shallow processing, resulted in significantly better recognition performance and that this effect was associated with activation of various brain areas. Further analyses revealed that there were regions directly relevant to episodic memory encoding in the anterior part of the parahippocampal gyrus, inferior frontal gyrus, supramarginal gyrus, anterior cingulate gyrus, and medial frontal lobe in the left hemisphere. Our results demonstrated that several regions, including the medial temporal lobe, play a role in episodic memory encoding.     

CD27/CD70 interaction directly drives B cell IgG and IgM synthesis

CD27 is a T cell activation antigen expressed on a majority of peripheral blood T cells. CD27 is also expressed on a subpopulation of human B cells, and it is reported that CD27⁺ B cells secrete both IgG and IgM. CD70, a ligand for CD27, is expressed on activated T and B cells, suggesting an interaction between T and B cells via CD27/CD70 ligation. Here, we analyze B cell immunoglobulin synthesis using a CD70 transfectant and present functional data showing that B cells secrete large amounts of IgG and IgM as a result of the CD27/CD70 interaction. A flow cytometric analysis showed that CD27 expression was increased and CD70 was expressed on tonsillar and peripheral blood B cells after activation with Staphylococcus aureus Cowan strain (SAC) plus interleukin (IL-2). In addition, the proliferation of B cells was enhanced mildly by the addition of CD70 transfectant, and its proliferation was blocked by anti-CD70 mAb. More importantly, the CD70 transfectant enhanced IgG and IgM production by purified B cells greatly in the presence of SAC plus IL-2. The enhancement was completely blocked by the addition of either anti-CD70 mAb or anti-CD27 mAb. Strongly suggesting that the interaction of CD27 with its ligand, CD70, on B cells plays an important role in B cell growth and differentiation to produce IgG and IgM.     

Equine herpesvirus-1-induced encephalomyelitis in mice: a comparative study of neuroadapted virus and its parental strain

Little is known about the neuropathogenicity of equine herpesvirus-1 (EHV-1) in mice. No neurological signs were observed in 6-day-old mice inoculated intracerebrally with the HH1 strain (HH1) of EHV-1. However,6-day-old mice inoculated intracerebrally with a variant derived by serial passage of HH1 in mouse brain showed severe neurological symptoms and eventually died. Histological analyses were performed on 6-day-old mice inoculated with the neuroadapted HH1 (NHH1) and the parental HH1 strain by the intracerebral, intranasal or intraperitoneal route. All routes of inoculation with NHH1 caused encephalitis, but myelitis was observed only in mice inoculated intraperitoneally. Prominent histological findings were perivascular cuffing sometimes associated with small fibrin thrombi, neuronal and glial degeneration and necrosis, and intranuclear inclusion bodies in neurons, glial cells and ependymal cells. Intracerebral and intranasal inoculation, but not intraperitoneal inoculation, with HH1 induced central nervous system (CNS) lesions that were milder than those in mice inoculated with NHH1. The distribution of viral antigen was more widespread in mice inoculated with NHH1 than with HH1. No viral antigen was detected in the CNS of mice inoculated intraperitoneally with HH1. These results indicate that increased viral multiplication and spreading in the CNS were responsible for the enhanced neurovirulence of NHH1. Although EHV-1 has been considered to be primarily endotheliotropic in horses, both NHH1 and HH1 showed tropism for the parenchymal cells of the CNS in mice, namely neurons, glial cells and ependymal cells.     

Analysis of E-cadherin in diffuse-type gastric cancer using a mutation-specific monoclonal antibody

In-frame deletions from the E-cadherin mRNA, coding for a homophilic cell adhesion molecule, are characteristic for diffuse-type gastric carcinomas. Using immunohistochemical analysis the mutant form cannot be distinguished from normal E-cadherin, making results difficult to interpret. In this study, a rat monoclonal antibody, designated E-cad delta 9-1, was generated against a peptide spanning the fusion junction region between exons 8 and 10. This new epitope is present in an E-cadherin variant that lacks exon 9 from the mRNA due to different splice-site gene mutations. Using Western blotting and immunohistochemistry of E-cadherin-transfected cells, we demonstrate that E-cad delta 9-1 specifically reacts with E-cadherin lacking exon 9 but not with the wild-type protein. No immunoreactivity was observed in 31 nontumorous and embryonal tissues analyzed. In gastric carcinoma specimens known to express mutant E-cadherin mRNA lacking exon 9, E-cad delta 9-1 targets exclusively tumor cells in routine formalin-fixed and paraffin-embedded material from biopsies, primary tumors, and lymph node metastases. In a retrospective series of 172 diffuse-type gastric carcinomas expressing E-cadherin, E-cad delta 9-1 reacted with 22 tumors (13%). This new tumor marker-monoclonal antibody system could open novel avenues for selective diagnosis and specific therapy of a subgroup of diffuse-type gastric cancer patients.     

Identification of neural genes using Xenopus DNA microarrays.

To isolate novel genes regulating neural induction, we used a DNA microarray approach. As neural induction is thought to occur by means of the inhibition of bone morphogenetic protein (BMP) signaling, BMP signaling was inhibited in ectodermal cells by overexpression of a dominant-negative receptor. RNAs were isolated from control animal cap explants and from dominant-negative BMP receptor expressing animal caps and subjected to a microarray experiment using newly generated high-density Xenopus DNA microarray chips representing over 17,000 unigenes. We have identified 77 genes that are induced in animal caps after inhibition of BMP signaling, and all of these genes were subjected to whole-mount in situ hybridization analysis. Thirty-two genes showed specific expression in neural tissues. Of the 32, 14 genes have never been linked to neural induction. Two genes that are highly induced by BMP inhibition are inhibitors of Wnt signaling, suggesting that a key step in neural induction is to produce Wnt antagonists to promote anterior neural plate development. Our current analysis also proves that a microarray approach is useful in identifying novel candidate factors involved in neural induction and patterning.     

The effect of calcium ion concentration on osteoblast viability, proliferation and differentiation in monolayer and 3D culture

Our research group aims to develop an osteochondral composite using type II collagen gel with hydroxyapatite (HAp) deposited on one side. Soaking gels in Ca2+ and phosphate solution is indispensable to HAp deposition, so relationships between cell behavior and Ca2+ concentration were examined in two- and three-dimensional cultures. The present results indicate that 2-4 mM Ca2+ is suitable for proliferation and survival of osteoblasts, whereas slightly higher concentrations (6-8 mM) favor osteoblast differentiation and matrix mineralization in both 2- and 3-dimensional cultures. Higher concentrations (>10 mM) are cytotoxic. Purely from the perspective of calcium deposition, higher concentrations lead to increased accumulation of Ca2+. Culturing cells in phosphate-containing gel in media with Ca2+ also leads to time-dependent formation of HAp in the gel. Considering the viability of embedded cells, culturing scaffolds in media with Ca2+ concentrations around 5mM is useful for both HAp deposition and osteoblast behavior.     

Analysis of proliferating biliary epithelial cells in human liver disease using a monoclonal antibody against DNA polymeraseα

The proliferative activity and ultrastructural characteristics of proliferating biliary epithelial cells were analysed immunohistocytochemically in 39 biopsied liver specimens from patients with acute viral hepatitis, chronic hepatitis and liver cirrhosis using a monoclonal antibody against DNA polymerase (DNA-PA). In acute viral hepatitis with perivenular confluent necrosis, proliferation of typical bile ducts was found frequently in portal areas. In chronic aggressive hepatitis and cirrhosis, ductular proliferation of both typical and atypical forms was found in enlarged portal and periportal areas and in confluent necrotic areas. The number of proliferating biliary epithelial cells that stained positive for DNA-PA was small. There were very few positively stained cells in atypical bile ducts in confluent necrotic areas of cirrhosis. Atypical bile ducts seen in chronic aggressive hepatitis, cirrhosis and acute hepatitis with confluent necrosis were positively stained for both cytokeratins 8 and 19. In cirrhosis, the number of stained biliary epithelial cells in typical bile ducts was larger than the number of such cells in atypical bile ducts (P< 0.01). By electron microscopy, the cells positively stained for DNA-PA were mostly so-called clear cells with irregular nuclei containing coarse nucleoplasm, and a few small cells with scanty cytoplasm and few organelles.