Kinetic profiles of sequential gene expressions for chemokines in mice with contact hypersensitivity

Using cDNA microarray technology, the expression of chemokine genes in the elicitation site of 2,4,6-trinitrochlorobenzene-induced contact hypersensitivity (CHS) was examined in mice. Of the 33 genes analyzed, levels of 11 gene expressions changed, and these can be assigned to four groups based on their kinetic patterns; (1) LARC/CCL20 whose mRNA level increased rapidly at 3 h post-challenge and then gradually decreased, (2) JE/CCL2, MARC/CCL7, MIP-1gamma/CCL9, monocyte chemoattractant protein (MCP)-5/CCL12, ELC/CCL19 and BRAK/CXCL14 whose mRNA levels increased with time and reached the maximum at 6-9 h post-challenge, (3) LIX/CXCL5, Mig/CXCL9 and IP-10/CXCL10 whose mRNA levels increased gradually at least up to 12 h post challenge, and (4) SLC/CCL21 whose mRNA level decreased gradually with time after challenge. The findings suggest that sequential expression of chemokine genes is essential for orientating non-specific skin response to hapten-specific CHS response through the recruitment of inflammatory cells such as neutrophils, monocytes/macrophages and T-cells from the circulation into the tissue site.     

Analysis of 8-hydroxydeoxyguanosine 5'-monophosphate (8-OH-dGMP) as a reliable marker of cellular oxidative DNA damage after gamma-irradiation

In order to improve 8-hydroxyguanine (8-OH-Gua) detection in DNA, we digested isolated DNA with nuclease P1 and analyzed for 8-hydroxydeoxyguanosine 5'-monophosphate (8-OH-dGMP) using a high-performance liquid chromatography system equipped with an electrochemical detector (HPLC-ECD). The amount of 8-OH-Gua in the DNA was expressed as the ratio of 8-OH-dGMP to deoxycytidine monophosphate (dCMP). Using this analysis, the background level of 8-OH-Gua in DNA from human lung carcinoma cells (A549) was several-fold lower than that obtained by a previous method. A549 cells were exposed to 20-60 Gy of gamma-radiation and an increase in 8-OH-Gua concentration was observed with increasing gamma-ray dose (0.3 residues per 10(7) dCMP per Gy). Moreover, by an immunohistochemical procedure using a commercial FITC-kit, 8-OH-Gua was clearly detected in A549 cells and the fluorescence intensity of cells with oxidative DNA damage increased with the doses of gamma-irradiation. Using an endonuclease nicking assay, we also found that gamma-rays decreased 8-OH-Gua repair activity. The results indicate that 8-OH-dGMP is a useful and sensitive marker for estimating oxidative damage in DNA.     

Kissing reduces allergic skin wheal responses and plasma neurotrophin levels

The effect of kissing on allergen-induced skin wheal responses and plasma neurotrophin levels were studied in 30 normal subjects, 30 patients with allergic rhinitis (AR), and 30 patients with atopic dermatitis (AD). All of the patients with AR or AD are allergic to house dust mite (HDM) and Japanese cedar pollen (JCP). They are all Japanese and they do not kiss habitually. The subject kissed freely during 30 min with their lover or spouse alone in a room with closed doors while listening to soft music. Before and after kissing, skin prick tests were performed using commercial HDM allergen, JCP allergen, as well as histamine and control solution, and wheal responses were measured. Simultaneously, plasma levels of neurotrophin, including nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF), neurotrophin-3 (NT-3) and -4 (NT-4) were measured. Kissing significantly reduced wheal responses induced by HDM and JCP, but not by histamine, and decreased plasma levels of NGF, BDNF, NT-3, and NT-4 in patients with AR or AD, while it failed to do so in normal subjects. These finding indicate that kissing have some implication in the study of neuroimmunology in allergic patients.     

Two case reports of childhood liver cell adenomas harboring beta-catenin abnormalities

The benign epithelial neoplasm liver cell adenoma is rare, especially in childhood. We report 2 such cases, 1 of which was associated with Prader-Willi syndrome. Differential diagnosis of the liver cell adenomas on the basis of histopathologic findings proved difficult and was based on the absence of cellular and nuclear atypia, mitotic activity, and invasive growth. In both cases, immunohistochemical staining demonstrated the nuclear accumulation of beta-catenin, and in 1 case, the tumor cells carried a mutation of the beta-catenin gene. Recently, disregulation of the Wnt/beta-catenin pathway, attributable to abnormalities of the beta-catenin gene, has been reported to be a major event in the development of hepatocellular carcinomas and hepatoblastomas. Our report may be the first to describe the beta-catenin abnormalities in childhood liver cell adenoma. These findings imply that abnormalities of beta-catenin can be an early initiating event in human liver tumorigenesis.     

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.     

Diffusion coefficient of water through dental composite resin

Water sorption of polymer filling materials affects dimensional stability, mechanical properties and bonding strength with tooth structures. To clarify the effect of the degradation on service life and micro-leakage, the diffusion coefficient of water through the resin should be identified. Distributions of time-dependent water concentrations in the resin were computed. Water sorption of composite resin discs with different thicknesses was measured and compared with the solution of Fick's second law. The diffusion coefficient of water through the resin discs was computed to be D=3.9-5.0 x 10(-13)m(2)/s from the measurements of specimens with different thicknesses. Results of water sorption measurements for the discs with different thicknesses were in good agreement with the theoretical results. The relationship among the thickness of the disc, the diffusion coefficient and the water sorption ratio was shown clearly. The testing method for water sorption by International Standard ISO 4049 for resin-based filling materials was discussed.     

The effect of peroxisome proliferator-activated receptor-gamma ligand on urological cancer cells

Peroxisome proliferator activator-receptor (PPAR)-gamma ligand induces growth arrest of cancer cells through apoptosis. In this study, we examined the effects of PPAR-gamma inhibitors on cell proliferation in renal cell carcinoma (RCC), bladder tumor (BT), and prostatic carcinoma (PC) cell lines. We investigated the inhibitory effect of PPAR-gamma ligands, troglitazone and 15-deoxy-Delta12,14-prostaglandin J2 (15dPGJ2) on RCC, BT and PC-derived cell lines using MTT assay and Hoechst staining. PPAR-gamma ligands (troglitazone and 15dPGJ2) induced the reduction of cell viability with the half-maximal concentration of growth inhibition of RCC, BT, and PC cell lines. Furthermore, counting cells at days 1, 2 and 3, clearly showed marked inhibition of cell proliferation using troglitazone and 15dPGJ2. All PPAR-gamma inhibitors stopped the growth of all RCC, BT and PC cells. Cells treated with PPAR-gamma inhibitors showed chromatin condensation, cellular shrinkage, small membrane-bound bodies (apoptotic bodies), and cytoplasmic condensation. These cellular changes were typically redundant characteristics of apoptosis. PPAR-gamma ligands may mediate potent antiproliferative effects against RCC, BT and PC cells through differentiation. Thus, PPAR-gamma may become a new target in treatment of urological tumors.     

Deposition of complement protein C3b on mixed self-assembled monolayers carrying surface hydroxyl and methyl groups studied by surface plasmon resonance

Since complement activation is recognized as a common response of the host defense system when an artificial medical device is applied to a patient, great effort has been devoted to studies on the interaction of the complement system with artificial materials. However, some uncertainties remain, partially because of the lack of well characterized surfaces and suitable analytic methods for study of the surface phenomena that occur on artificial materials under physiologic conditions. In this study, we employed self-assembled monolayers (SAMs) and the surface plasmon resonance (SPR) technique to study interactions of the serum complement with well characterized surfaces. Self-assembled monolayers carrying various concentrations of hydroxyl groups were prepared using 11-mercapto-1-undecanol (C11-OH) and one of n-nonanethiol, n-dodecanethiol, and n-hexadecanethiol. The amount of NHS deposition on the SAMs increased with increasing C11-OH content of the SAMs, and the amount of anti-C3b antibody immobilization formed on the NHS deposition layers increased with increasing C11-OH content of the SAMs. These results clearly demonstrate that a large amount of C3b, produced through the activation of the complement system, binds covalently to and is adsorbed by hydroxyl-group-rich surfaces. The combination of SAMs and the SPR technique is suitable for studying the interaction of the complement system with solid surfaces, and the results should give basic information needed for a rational design of biocompatible surfaces on synthetic materials.