A NEW METHOD FOR VITAL STAINING OF CENTRAL NERVOUS TISSUES WITH NEUTRAL RED

A new vital staining method with neutral red has been established where by cerebral ganglion cells can be stained in vivo .     

Glyceraldehyde 3-phosphate dehydrogenase is a novel autoantigen leading autoimmune responses to proliferating cell nuclear antigen multiprotein complexes in lupus patients

Using 2-dimensional electrophoresis and ion-pair chromatography, we have identified elements of proliferating cell nuclear antigen (PCNA) multiprotein complexes that are reactive to antibodies in sera from patients with systemic lupus erythematosus. Among the various elements of the complexes, a 37 kDa protein (PI 8.5) that specifically reacted with SLE sera, but not with sera from patients with other connective tissue diseases, was identified as glyceraldehyde 3-phosphate dehydrogenase (GAPDH). Immunoblot analysis showed that SLE sera reactive with the 37 kDa protein specifically reacted with GAPDH, as did anti-GAPDH mAbs. The purified autoantibodies to GAPDH from lupus serum showed both nuclear speckled and cytoplasmic staining patterns in immunofluorescence on Hep-2 cells. In addition, enzyme-linked immunosorbent assay (ELISA) revealed the presence of anti-GAPDH autoantibodies in 47% of lupus patients. Longitudinal analysis of the reactivity of lupus sera to PCNA complexes showed the autoimmune response to spread from GAPDH to other elements of PCNA complexes, and the presence of anti-GAPDH antibodies was significantly correlated with increased levels of serum PCNA. Taken together, these findings suggest that GAPDH interacting with PCNA in association with its cellular function is a novel autoantigen recognized by lupus sera, and that GAPDH thus plays an important role in the induction of autoimmune responses against the PCNA complex.     

Lysenin: A new tool for investigating membrane lipid organization

Sphingomyelin is a major sphingolipid species in animal cells and is a major lipid constituent of plasma membranes. Recent reports have established important roles for sphingomyelin and its metabolites as second messengers in signal transduction events during development and differentiation. Sphingomyelin is also a major component of sphingolipid, cholesterol-rich plasma membrane microdomains, known as 'lipid rafts'. However, little is known about the organization of sphingomyelin in biological membranes. Lysenin is a recently discovered sphingomyelin-specific toxin. In the present review, we summarize the current characterization of this protein and describe our recent attempt to elucidate the organization of sphingomyelin in cellular membranes using lysenin as a unique tool.     

Subcutaneous cryptococcosis due to Cryptococcus diffluens in a patient with sporotrichoid lesions case report, features of the case isolate and in vitro antifungal susceptibilities.

Environmental fungi, in particular primary pathogens and Cryptococcus spp. can be responsible for skin lesions mimicking sporotrichosis. In this paper, we report a case of subcutaneous cryptococcosis in an apparently healthy, young male patient due to a non-C. neoformans Cryptococcus species, C. diffluens. The isolate showed in vitro phenotypic switching that may affect virulence and host inflammatory and immune responses, and in vitro resistance to amphotericin B and 5-flucytosin. This species shares several phenotypic traits with C. neoformans, and, therefore, decisive diagnosis should be based on biopsy and culturing results followed by molecular identification.     

Expression of interferon-gamma-inducible protein-10 in human endometrial stromal cells

Human endometrial stromal cells (ESC) can produce a variety of chemokines, especially after inflammatory stimulation. Interferon-gamma-inducible protein-10 (IP-10) is a potent chemoattractant for lymphocytes, and belongs to the family of non-ELR CXC chemokines. The expression of IP-10 in ESC after stimulation with interferon-gamma (IFN-gamma), interleukin-1 beta (IL-1 beta), tumour necrosis factor-alpha (TNF-alpha), or lipopolysaccharide (LPS) was evaluated using an enzyme-linked immunosorbent assay and Northern blot analysis. A small amount of IP-10 protein was detected in the culture media of unstimulated ESC. The expression of IP-10 mRNA was detected in ESC. IFN-gamma, IL-1 beta, TNF-alpha and LPS significantly stimulated the expression of IP-10 mRNA and protein in ESC. These results suggest that the production of IP-10 by ESC is regulated by inflammatory mediators. The modulation of IP-10 concentrations in the local environment may contribute to the normal and pathological processes of human reproduction by regulating leukocyte trafficking in the endometrium.     

The BPV-1 E5 oncoprotein expressed in Schizosaccharomyces pombe exhibits normal biochemical properties and binds to the endogenous 16-kDa component of the vacuolar proton-ATPase

The 44-amino-acid E5 oncoprotein of bovine papillomavirus type 1 transforms immortalized murine fibroblast cell lines. This highly hydrophobic protein forms homodimers, localizes to intracellular membrane compartments (including the Golgi apparatus), and forms a complex with the 16-kDa membrane-embedded constituent (16k) of the vacuolar proton-ATPase. To develop a system for the genetic and biochemical analysis of the E5/16k interaction, the E5 gene was cloned into a new vector which was designed for expression in the fission yeast Schizosaccharomyces pombe. The E5 protein synthesized in this system dimerized normally and bound to endogenous and overexpressed S. pombe 16k protein. Comparison of the S. pombe and mammalian 16k proteins showed strong conservation in carboxyl-terminal amino acids but greater variation in the amino-terminal sequences, suggesting that E5 was interacting with the 16k carboxyl domains. Finally, a new protein epitope tag is described which permitted for the first time the coprecipitation of E5 with antibodies directed against the 16k protein.     

Local secretory immunoglobulin A and postimmunization gastritis correlate with protection against Helicobacter pylori infection after oral vaccination of mice

C57BL/6 mice were orally immunized with five weekly doses of 2 mg, 200 microgram, or 2 microgram of Helicobacter pylori (Sydney strain) whole-cell sonicate combined with cholera toxin. One week after the last vaccination, mice were challenged with 5 x 10(7) CFU of live H. pylori three times at 2-day intervals. At 6 or 18 weeks after the challenge, mice were sacrificed and bacterial cultures and histological studies of the stomach were performed. Vaccination with 2 mg/session or 200 microgram/session inhibited H. pylori colonization by 90 and 100%, respectively. These mice were considered protected. Lower levels of H. pylori-specific immunoglobulin A (IgA) were detected in fecal and saliva samples before challenge. However, a significant increase in IgA secretion in mucosal tissue and a higher labeling index for IgA-positive lumina of pyloric glands were noted in these mice in response to challenge and in a vaccine dose-dependent manner. In protected mice, however, severe gastritis characterized by marked infiltration of inflammation mononuclear cells was noted at 6 weeks after challenge, compared with the gastritis seen in unprotected mice or nonvaccinated, ordinarily infected mice. Marked expression of gamma interferon mRNA was detected in the stomach of all protected mice, and 50% of these mice expressed interleukin 4 (IL-4) or IL-5 mRNA. Our findings suggest that local secretory IgA antibody and severe postimmunization gastritis correlate well with protection of mice against H. pylori infection.     

Ras homolog gene family H (RhoH) deficiency induces psoriasis-like chronic dermatitis by promoting TH17 cell polarization

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Electrical polarization of plasma-spray-hydroxyapatite coatings for improvement of osteoconduction of implants

We have established the electrical polarization method of hydroxyapatite (HA) coatings (HAC) for clinical use, such as dental and orthopedic implants. The HAC examined in the current study was prepared in titanium substrates by plasma spraying of beta-tricalcium phosphate (TCP) powders followed by hydrothermal treatment. The prepared HAC consisted of a single phase of HA, because the starting TCP phase was completely transformed to the HA phase during the posthydrothermal treatment. Polarization was carried out at the elevated temperature of 400 degrees C under a d.c. field of 1 kV . cm(-1). The electrical measurements showed that the stored charges of the polarized HAC were greater than the reported value of the sintered ceramic HA. The enhanced bioactivity of the polarized HAC was demonstrated using 1.5 simulated body fluid (SBF). The crystal growth from the SBF solution was accelerated on the negatively charged surface in comparison with the untreated HAC. Similar to the polarized ceramic HA, the current results confirmed that the bioactivity of HAC would be effective for improving the initial fixation by polarization.     

Effects of Alkyl Alcohols and Related Chemicals on Rat Liver Structure and Function: III. Physicochemical Properties of Ethanol-, Propanol- and Butanol-treated Rat Liver Mitochondrial Membranes

The physicochemical properties of mitochondria in liver tissue obtained from rats given 32% ethanol, 32% propanol or 6.9% butanol in drinking water for up to 3 months were investigated using differential scanning calo-rimetry and fluorescence polarization measurements. The results obtained were as follows: 1) Phospholipids extracted from mitochondria showed increases in the relative amounts of phosphatidylcholine, phosphatidylinositol and phosphatidylserine, and a decrease in the relative amount of phosphatidylethanolamine. An increase in the unsatu-rated/saturated fatty acid ratio of phospholipids was also observed. 2) Elevation of the thermotropic lipid phase transition temperature with a decrease in the enthalpy value (δ H ) was revealed by differential scanning calo-rimetry. 3) The elevation of the lipid phase transition temperature was detected also by fluorescence polarization measurements using 1,6 diphenyl 1, 3, 5 hexatriene (DPH) as a probe. Elevation of mitochondrial membrane fluidity was found in some of the experimental animals, but most showed no changes in comparison with the control. A possible role of membrane fusion in the mechanism of formation of ethanol-, propanol- and butanol induced hepatic megamitochondria is discussed on the basis of these results. Acta Pathol Jpn 42: 549–557, 1992.