Monoclonal antibodies against synthesized short peptides corresponding to human AA amyloid protein

Monoclonal antibodies (McAbs) were raised against the synthesized short peptides corresponding to 37-47 residues in amino acid sequence of human AA protein. The McAbs reacted immunohistochemically to amyloid tissues from cow, mouse, swan, and human AA amyloidosis. We concluded that the McAbs were useful for identification of AA type amyloidosis of various species, and that the 37-47 residues were effective antigenic sites in AA protein.     

Cell lineage specificity of newly raised monoclonal antibodies against gastric mucins in normal, metaplastic, and neoplastic human tissues and their application to pathology diagnosis

The specificity of monoclonal antibodies against gastric mucins (designated as HIK1083, PGM 36, and PGM 37) was studied immunohistochemically in normal, metaplastic, and neoplastic human tissues. These antibodies labeled class III mucin-producing cells identified by paradoxical concanavalin A staining in normal stomach, duodenum (Brunner gland), biliary tract, and main pancreatic duct; in mucinous metaplasia of pancreas and gallbladder; and in adenocarcinomas of stomach (90%), bile duct (80%), gallbladder (100%), pancreas (80%), lung (100% of goblet cell type adenocarcinomas), ovary (67% of mucinous carcinomas), and uterine cervix (100% of adenoma malignum tumors). Normal and neoplastic cells of esophagus, colon, salivary gland, kidney, endometrium, breast, prostate, and liver, as well as normal small intestine, lung, and uterine cervix, were all negative. The antibodies used should be valuable for the detection of class III mucin and class III mucin-producing cells in normal, metaplastic, and neoplastic tissues.     

Mouse erythroblast formation is inhibited by anemia-inducing substance from the plasma of a patient with a malignant neoplasm

An anemia-inducing substance (AIS) was found as a protein, with a molecular weight of 50,000 on SDS electrophoresis gel, that decreased the osmotic resistance of erythrocytes. In this study, the plasma fraction containing AIS is shown to inhibit mouse erythroblast formation in vitro. The addition of the plasma fraction from a patient with an advanced malignant neoplasm to a liquid culture of mouse bone marrow cells with erythropoietin results in low numbers of erythroblast formation and low cellular yields of alpha- and beta-globin mRNAs. These inhibitions were not observed after immunoadsorption of the plasma fraction with an antiserum against AIS.     

Purification and characterization of a hemolysin produced by a clinical isolate of Kanagawa phenomenon-negative Vibrio parahaemolyticus and related to the thermostable direct hemolysin.

A clinical isolate (strain 4037) of Kanagawa phenomenon-negative Vibrio parahaemolyticus was studied. Although the strain was judged to be Kanagawa phenomenon-negative by various conventional tests, it produced a new hemolysin (named Vp-TRH, for thermostable direct hemolysin [Vp-TDH]-related hemolysin) that was related to the Vp-TDH produced by ordinary Kanagawa phenomenon-positive V. parahaemolyticus. Vp-TRH was purified by ammonium sulfate fractionation and successive column chromatographies on DEAE-cellulose, hydroxyapatite, and Mono Q. The molecular weight of Vp-TRH was estimated as 48,000 by Sephadex G-100 gel filtration, and the molecular weight of the subunit was estimated to be 23,000 by sodium dodecyl sulfate-slab gel electrophoresis. Thus, like Vp-TDH, Vp-TRH seems to be composed of two subunits. The isoelectric point of Vp-TRH was determined to be 4.6. Vp-TRH showed lytic activities different from those of Vp-TDH on erythrocytes from various animals, especially those from calves, chickens, and sheep. The hemolytic activity of Vp-TRH was labile on heat treatment at 60 degrees C for 10 min, unlike that of Vp-TDH. The immunological similarities, but not the identities of Vp-TRH and Vp-TDH, were demonstrated by Ouchterlony, neutralization, and latex agglutination tests. Thus, we conclude that this clinical isolate of Kanagawa phenomenon-negative V. parahaemolyticus produces a new type of hemolysin that is similar, but not identical, to Vp-TDH, which is usually produced by Kanagawa phenomenon-positive V. parahaemolyticus.     

Fertilization and early embryology: Effects of embryo density and co-culture of unfertilized oocytes on embryonic development of in-vitro fertilized mouse embryos

We have evaluated the effects of embryo density and the co-cultureof unfertilized (degenerating) oocytes on the development ofin-vitro fertilized (IVF) mouse embryos. In experiment 1, groupsof one, five, 10 or 20 zygotes were cultured in 20 µldrops of modified human tubal fluid (HTF) medium for 168 h at38.7°C in 5% CO₂ and 95% air. As the embryo density increased,significantly (P < 0.05) higher rates of embryos reachedhatched blastocyst stage. In addition, the time required forhatching after IVF was significantly (P < 0.05) shortenedby the increase in embryo density. In experiment 2, 10 IVF zygoteswere cultured with or without 10 unfertilized (degenerating)oocytes in 20 µl drops of HTF medium. The rates of IVFembryos that developed to morula, blastocyst, expanded blastocystand hatched blastocyst stages were decreased significantly (P< 0.01) by culturing embryos with unfertilized oocytes comparedwith culturing embryos alone. In experiment 3, groups of oneor 10 IVF zygotes or 10 IVF zygotes plus 10 unfertilized oocyteswere cultured in 20 µl drops of HTF medium and the numberof cells per blastocyst was examined at 120 h after IVF. Increasingembryo density resulted in a significant (P < 0.05) increasein the number of cells per blastocyst. In contrast, the cellnumber of IVF embryos that developed to blastocyst decreasedsignificantly (P < 0.05) when they were cultured with unfertilizedoocytes. The results suggest that in-vitro development of IVFmouse embryos is enhanced by increasing embryo density and isimpaired by co-culture with unfertilized (degenerating) oocytes.     

Regulation of the type I IFN induction: a current view

The type I IFN-alpha/beta gene family was identified about a quarter of a century ago as a prototype of many cytokine gene families, which led to the subsequent burst of studies on molecular mechanisms underlying cytokine gene expression and signaling. Although originally discovered for their activity to confer an antiviral state on cells, more evidence has recently been emerging regarding IFN-alpha/beta actions on cell growth, differentiation and many immunoregulatory activities, which are of even greater fundamental biological significance. Indeed, much attention has recently been focused on the induction and function of the IFN-alpha/beta system regulated by Toll-like receptors (TLRs), which are critical for linking the innate and adaptive immunities. The understanding of the regulatory mechanisms of IFN-alpha/beta gene induction by TLRs and viruses is an emerging theme, for which much new insight has been gained over the past few years.     

Effects of hypocapnia on respiratory timing and inspiratory activities of the superior laryngeal, hypoglossal, and phrenic nerves in the vagotomized rat

Effects of acute hypocapnia on respiratory timing (inspiratory and expiratory times (TI, TE) ) and on inspiratory activities of the efferent superior laryngeal (Xs1), hypoglossal (XII), and phrenic (Phr) nerves were studied in artificially ventilated vagotomized, and anesthetized rats. Hyperventilation induced a decrease in respiratory frequency exclusively due to prolongation of TE and resulted in expiratory apnea. Inspiratory activities of three nerves decreased with reduction in CO2 concentration of end-tidal gas (FETCO2), and disappeared simultaneously at a threshold FETCO2 for apnea. The decrease in the peak inspiratory activity by hypocapnia was larger in the XII than in the Phr or Xs1 nerve (XII greater than Phr greater than Xs1). The results suggest that the CO2 stimulus (mainly via a central chemosensor) plays an important role in the process of terminating expiration or of expiratory-inspiratory phase switching and that the responses of the XII or Xs1 motoneurons to variation in CO2 stimulus differ from that of the Phr motoneurons (or of the Phr driving medullary neurons). A possible functional significance of these observations is discussed.     

CO2 dissociation curves of oxygenated whole blood obtained at rest and in exercise

Summary In vitro CO₂ dissociation curves for oxygenated whole blood were determined in 19 healthy male subjects at rest and during submaximal and maximal bicycle work. Hemoglobin concentration and blood lactate increased with increasing work load and accordingly buffer value of the whole blood increased while bicarbonate and Base Excess (BE) decreased, resulting in a downward shift of the CO₂ dissociation curve during exercise. Despite the marked increase in buffer values of the blood, the slopes of the CO₂ dissociation curves during exercise were found to be about the same as those obtained at rest. It was inferred that the increasing effect of increased buffer value, on the dissociation slope, was essentially compensated by the decreasing effect of diminished bicarbonate content. The advantages of this relatively constant CO₂ dissociation slope for the indirect measurement of cardiac output by the Fick principle are discussed.     

Increased release of glucuronoxylomannan antigen and induced phenotypic changes in Trichosporon asahii by repeated passage in mice

Clinically important fungi such as Candida albicans and Cryptococcus neoformans are known to undergo phenotypic changes after repeated subculture or passages in vivo. However, there are no reports describing this phenomenon in Trichosporon species. This study investigated whether in-vivo passages of environmental isolates of Trichosporon asahii in mice changes their phenotype; three environmental isolates and 14 clinical isolates (from deep-seated infections) were used. The shape of the colony and cell type were observed, and the titre of glucuronoxylomannan (GXM) antigen and concentration of (1-->3)-beta-D-glucan were measured for each isolate. Changes in these features were also examined after three passages of the environmental isolates in mice. The shape of colonies and cell types were clearly different in environmental and clinical isolates. Furthermore, the clinical isolates released significantly higher levels of GXM antigen than environmental isolates (titre: log2 9.4 SD 0.7 versus log2 5.4 SD 1.4). The phenotype of passaged isolates was significantly different from the original environmental isolates with respect to the morphology of colonies and cell type and GXM release (titre: log2 10.0 SD 0.7 versus log2 5.4 SD 1.4). These results suggest that the phenotypic changes in T. asahii occur as a result of in-vivo passages. This process may allow a proportion of the fungal population to escape eradication by the host immune system, as GXM antigen is considered to protect the fungi against phagocytosis by polymorphonuclear leucocytes and monocytes in vivo.     

Role of nectin in organization of tight junctions in epithelial cells

BACKGROUND: In polarized epithelial cells, cell-cell adhesion forms specialized membrane structures comprised of claudin-based tight junctions (TJs) and of E-cadherin-based adherens junctions (AJs). These structures are aligned from the apical to the basal side of the lateral membrane, but the mechanism of this organization remains unknown. Nectin is a Ca2+ independent immunoglobulin-like cell-cell adhesion molecule which localizes at AJs. Nectin is associated with E-cadherin through their respective cytoplasmic tail-binding proteins, afadin and catenins, and involved in the formation of AJs in cooperation with E-cadherin. We show here that nectin is also involved in the formation of TJs. RESULTS: During the formation of the junctional complex consisting of AJs and TJs in Madin-Darby canine kidney (MDCK) cells, claudin and occludin accumulated at the apical sites of the nectin-based cell-cell adhesion sites. This accumulation of claudin and occludin was inhibited by inhibitors acting on the trans interaction of nectin. The barrier function of TJs was also impaired by the nectin inhibitors. It has been shown that a phorbol ester promotes the formation of a TJ-like structure in an E-cadherin-independent manner. This phorbol ester-induced formation of the TJ-like structure was also inhibited by the nectin inhibitors. CONCLUSIONS: These results suggest a role of the nectin-afadin system in the organization of TJs as well as AJs in epithelial cells.