An enzyme immunoassay for a metabolite of a new sleep inducer (450191-S), a ring-opened derivative of 1,4-benzodiazepine, in human serum

A competitive enzyme immunoassay was developed for metabolites of 450191-S, a ring-opened derivative of triazolobenzodiazepines, in human serum. Three kinds of labelled antigens, beta-D-galactosidase conjugates, and antisera from three kinds of immunogens, bovine serum albumin conjugates, were tested and the hapten heterologous assay was selected. B/F separation was performed using immobilized second antibody and the enzyme activity was measured using fluorescent substrate. One of the serum metabolites, the carboxyl form, was extracted from acidified serum and measured by the assay. The minimum detectable concentration was 1.5 ng/ml in serum, 15 pg/tube. The intra-assay and inter-assay variances were 5.5% and 7.7% at 6 ng/ml of serum, respectively. Standard curves of other related metabolites were assessed with various combinations of labelled antigens and antibodies.     

Duct-Islet Cell Tumor of the Pancreas. A Case Report with Immunohistochemical and Electron Microscopic Findings

A case of pancreatic tumor with features of both duct and islet cell components was found incidentally at autopsy in a 76 year old male who had died of intrahepatic cholan-giocarcinoma. The tumor, measuring about l.0cm in diameter, was located in the pancreatic tail. The tumor was composed of two distinct cell populations, islet cells and duct cells. Immunocytochemically, nearly all of the former cells were positive for insulin but negative for cytokeratin, carcinoembryonic antigen (CEA) and mucin, while the latter were positive for cytokeratin, CEA and mucin but negative for insulin. Additionally, a majority of the tumor cells that had formed islet-like structures were positive for neuron specific enolase (NSE), whereas NSE-positive cells were found only rarely in duct components. Electron microscopy confirmed the presence of two cell populations. Simultaneous occurrence of duct and islet cell components in a single pancreatic tumor indicates an intimate histogenetic relationship between pancreatic endocrine and duct cells. Acta Pathol Jpn 39: 328 335, 1989.     

Role of 5-HT1B receptors in entrainment disorder of Otsuka Long Evans Tokushima fatty (OLETF) rats

The role of 5-HT1A and 5-HT1B receptors in entrainment function was studied in Otsuka Long Evans Tokushima fatty (OLETF) rats and control Long Evans Tokushima Otsuka (LETO) rats. Light-induced (100 lux, 30 min) Fos expression in the suprachiasmatic nucleus was studied. Light-induced Fos expression was significantly decreased in OLETF rats compared to that in LETO rats. The decrease of light-induced Fos expression in OLETF rats was significantly reversed by pretreatment with the 5-HT1B receptor antagonist, isamoltan (3 mg/kg, i.p.). Simultaneous administration of CGS12066B (5 mg/kg, i.p.), a 5-HT1B agonist, blocked the reversal effect of isamoltan on Fos expression. Fos expression was not changed in LETO rats by pretreatment with isamoltan (3 mg/kg, i.p.). The Fos expression in LETO and OLETF rats was significantly decreased by pretreatment with the 5-HT1A antagonist, WAY-100,635. Phase shifts in locomotor activity paralleled the Fos expression. Light-induced phase shifts of locomotor activity in OLETF rats were significantly smaller than those in LETO rats. The phase shifts were significantly increased by isamoltan (3 mg/kg, i.p.) in OLETF rats. These results suggest that 5-HT1B receptors are involved in the reduced entrainment function of OLETF rats.     

Ultrastructural changes of bile duct epithelium in primary biliary cirrhosis in relation to progression of bile duct loss

Summary Using wedge liver biopsies from patients with primary biliary cirrhosis (PBC), ultrastructural features of the intrahepatic bile ducts in livers with slight or no bile duct loss were compared with those in livers with advanced bile duct loss and in extrahepatic cholestasis (EHC).Most changes in the biliary epithelium in PBC were similar to those in EHC. Microvillous loss and bleb formation, mitochondrial damage and increase in endoplasmic reticulum and ribosomes were found in PBC irrespective of the degree of bile duct loss, and also in EHC. These changes were present almost equally at any level of the biliary tree, and are presumed to represent a variety of non-specific lesions of biliary epithelial cells. As the loss of bile ducts in PBC progressed, cytoskeletal filaments and cytophagosomes increased in number and basement membranes were more thickened and reduplicated. These changes were more or less conspicuous in smaller branches of the biliary tree, and were also prominent in EHC. They might be causally related to the bile flow disturbance in the liver. Lateral intercellular spaces were irregularly dilated and contained osmiophilic membranous and/or granular material, similar to that found in duct lumena, within and without the basement membrane, and in the cytoplasm of periductal macrophages. Furthermore, pinocytotic vesicles were increased in the biliary cytoplasm facing periphery. These findings suggest possible alteration of the permeability of biliary epithelial cells, probably in the direction from the lumena to the periductal tissue. Such changes were found in PBC livers with virtual absence of bile duct loss, and the significance of this phenomenon is discussed.     

Oligomeric immunoglobulin A antibody response to rubella virus infection.

Pooled sera from rubella patients in the early convalescent stage, containing a high titer of hemagglutination-inhibiting (HI) antibody, were treated with protein A-conjugated gel to reduce immunoglobulin G (IgG) antibody and then centrifuged in sucrose gradients. This treatment resulted in the detection of an HI activity peak sedimenting at a rate intermediate between 7S and 19S. In contrast to the 19S antibody, the HI activity of this peak was not abolished by 2-mercaptoethanol, but sedimented at 7S after this treatment. The activity was considered to consist of IgA oligomers, since it was removed by anti-IgA immunosorbent. The appearance of the oligomeric IgA antibody after the infection was then studied using serum samples collected sequentially from five rubella patients. Shortly after the onset of the disease, the HI activity appeared at high titer and thereafter gradually decreased in titer until it could no longer be detected in the sera. The time of its disappearance varied with each patient.     

Gingival mononuclear cells from chronic inflammatory periodontal tissues produce interleukin (IL)-5 and IL-6 but not IL-2 and IL-4.

A unique characteristic of the localized inflammatory tissue in the periodontium (e.g., adult periodontitis [AP]) is the accumulation of IgG (IgG1 > IgG2 > IgG3 > or = IgG4) followed by IgA plasma cells (IgA1 > IgA2). However, the exact molecular mechanisms contributing to these elevated B-cell responses at the local disease site are still unknown. Thus, this study has examined the production of cytokines of importance in B-cell responses, e.g., interleukin (IL)-2, IL-4, IL-5, and IL-6 by gingival mononuclear cells (GMC) isolated from patients in severe stages of AP. These cytokines were assessed at the protein and messenger (m)RNA levels to understand their importance for the observed increased B-cell responses present in these tissues. Among the four cytokines tested by respective cytokine-specific, polymerase chain reaction and dot-blot hybridization, high levels of IL-5- and IL-6-specific mRNA were noted in GMC freshly isolated from AP patients. On the other hand, specific message for IL-2 and IL-4 were not present. Further, the analysis of culture supernatants of GMC also revealed that cells from AP patients spontaneously produced IL-5 and IL-6 but not IL-2 and IL-4. In contrast, when peripheral blood mononuclear cells isolated from the same patients were examined for these cytokines, no detectable levels of mRNA or secreted cytokines were noted. These results showed that GMC from localized inflammatory tissues in severe stages of AP possess a distinct cytokine profile represented by high levels of IL-5 and IL-6 mRNA expression and protein synthesis, whereas IL-2 and IL-4 were not detected. Further, this study supports the concept that AP is a localized inflammatory disease, because GMC from the inflamed tissue actively produce IL-5 and IL-6, whereas peripheral blood mononuclear cells from the same patients do not.     

Fibronectin binds to C1q: possible mechanisms for their co-precipitation in cryoglobulins from patients with systemic lupus erythematosus

Fibronectin and C1q frequently co-precipitated in cryoglobulins from patients with SLE. As a portion of the C1q molecule is similar to collagen to which fibronectin has a high affinity, we studied whether fibronectin specifically bound to C1q. Fibronectin was found to bind to both native and heat-inactivated C1q. The binding was enhanced by Ca++ and low ionic strength. We have also demonstrated that fibronectin is capable of binding to C1q fixed to immune complexes. The interaction between fibronectin and C1q may play a role in cryoglobulin formation and in clearance of immune complexes by reticuloendothelial system.