Prevention of apoptosis of mammalian cells by the CED-3-cleaved form of CED-9

CED-9 prevents apoptosis in embryonic cells of Caenorhabditis elegans but not in mammalian cells. We show here that the prevention of apoptosis in mammalian cells requires a CED-3-cleaved form (68-280) of CED-9 which is localized in the inner mitochondrial membrane. The viability of PC12 and HeLa cells was significantly increased after death stimuli when truncated CED-9 was expressed in these cells but full-length CED-9 did not. The truncated CED-9 expressed in these cells was largely localized to the inner mitochondrial and the endoplasmic reticulum membranes, while full-length CED-9 was detected mainly in endoplasmic reticulum fractions. Moreover, truncated CED-9 in purified mitochondria was resistant to trypsin digestion, but full-length CED-9 was not. These results suggest that the CED-3-cleaved form of CED-9 prevents apoptosis in mammalian cells by localizing to the inner mitochondrial membrane.     

Histopathological study on the PTHrP-induced incisor lesions in rats

Parathyroid hormone related peptide (PTHrP) was discovered as a causative factor of humoral hypercalcemia of malignancy (HHM). The present study elucidates the histopathological characters of incisor lesions in the HHM rat model. Nude rats were implanted with PTHrP-expressing tumor (LC-6) cells, maintained for 12 weeks, after which the mandibular incisors were collected. Incisor fractures were observed grossly. Microscopically, hypercalcified dentin, dentin niche with osteodentin, and thinning of dentin were observed. Hypercalcified dentin was observed as a basophilic line of calcified dentin without associated odontoblastic changes, whereas dentin niche and thinning of dentin occurred with osteodentin and loss of cell height, respectively. In contrast with hypercalcified dentin, which was distributed throughout the dentin, dentin niche and thinning of dentin were localized to the labial area of the apical and middle region, and to the labial and lingual areas of the middle and incisal region, respectively. These results suggest that hypercalcemia affected the entire calcification process resulting in hypercalcified dentin, and that high PTHrP concentrations affected selective populations of odontoblasts resulting in formation of dentin niche and thinning of dentin. The localization of dentin niche and thinning of dentin also suggest that PTHrP may also be involved odontoblastic development in the rat.     

Subchronic exposure to arsenic through drinking water alters expression of cancer-related genes in rat liver

Although arsenic exposure causes liver disease and/or hepatoma, little is known about molecular mechanisms of arsenic-induced liver toxicity or carcinogenesis. We investigated the effects of arsenic on expression of cancer-related genes in a rat liver following subchronic exposure to sodium arsenate (1, 10, 100 ppm in drinking water), by using real-time quantitative RT-PCR and immunohistochemical analyses. Arsenic accumulated in the rat liver dose-dependently and caused hepatic histopathological changes, such as disruption of hepatic cords, sinusoidal dilation, and fatty infiltration. A 1-month exposure to arsenic significantly increased hepatic mRNA levels of cyclin D1 (10 ppm), ILK (1 ppm), and p27(Kip1) (10 ppm), whereas it reduced mRNA levels of PTEN (1 ppm) and beta-catenin (100 ppm). In contrast, a 4-month arsenic exposure showed increased mRNA expression of cyclin D1 (100 ppm), ILK (1 ppm), and p27(Kip1) (1 and 10 ppm), and decreased expression of both PTEN and beta-catenin at all 3 doses. An immunohistochemical study revealed that each protein expression accords closely with each gene expression of mRNA level. In conclusion, subchronic exposure to inorganic arsenate caused pathological changes and altered expression of cyclin D1, p27(Kip1), ILK, PTEN, and beta-catenin in the liver. This implies that arsenic liver toxicity involves disturbances of some cancer-related molecules.     

Metabolic polymorphisms and the micronucleus frequency in buccal epithelium of adolescents living in an urban environment

Micronuclei and other biomarkers were evaluated in oral cells from 11- to 16-year-old girls living in a foster home in the central area of México City. Variables analyzed for possible association with these biomarkers include smoking habits, body mass index, metabolic polymorphisms for NAT1 and GSTM1 and whether the cells were obtained from the cheek or pharynx. The results indicated that individuals having the NAT1*10 homozygous genotype showed a significant increase in chromatin buds and binucleated cells. When the damage in the cheek was compared with damage in the pharynx, a significant increase in micronuclei and binucleated cells was found for the latter tissue in all the individuals analyzed.     

Development of Outer Hair Cells in Ames Waltzer Mice: Mutation in Protocadherin 15 Affects Development of Cuticular Plate and Associated Structures

The Ames waltzer (av) mouse mutant harbors a mutation in the protocadherin 15 gene (Pcdh15) and is a model for deafness in Usher syndrome 1F and nonsyndromic deafness DFNB23. Mutation in Pcdh15 affects stereocilia morphogenesis and polarity. Disruptions of apical cellular components in outer hair cells have also been described in av mutants. Organization of stereocilia and cell polarization may be dependent on proper orientation of structural components residing in the apical portion of the cell during development. We used electron and immunofluorescent microscopy to examine structural maturation of outer hair cells in av3J mice with emphasis on the fonticulus, basal body/centriole complex, actin mesh, and the microtubule network during initiation of bundle organization, between embryonic day (E) 16.5 and postnatal day 5 (P5). We found major ultrastructural rearrangements near the hair cell surface in av3J mice. Earliest changes were in kinocilia, basal body, and stereocilia positioning and microtubule arrangement once the kinocilia had lateralized to the side of the cell (between E16.5 and postnatal day [P] 0, before cuticular plate formation and stereocilia elongation). By P0, the developing fonticulus in av mice appeared enlarged, with a normal vesicle density. Stereocilia bundle disorganization increased after P0, with disruptions of the actin mesh within the cuticular plate. These observations support the hypothesis that mutations in Pcdh15 in av3J mice adversely affect coordinated maturation of apical cell components, resulting in disturbed stereocilia bundle polarity in av mice. Anat Rec, 2007. © 2008 Wiley‐Liss, Inc.     

Up-regulation of transferrin receptor 1 in chronic hepatitis C: Implication in excess hepatic iron accumulation.

BACKGROUND/AIMS:: To clarify the mechanism of excess hepatic iron accumulation in chronic hepatitis C, we investigated the expressions of transferrin receptor 1 and divalent metal transporter 1 in hepatocytes, both of which are involved in cellular iron uptake, in relation to the degree of hepatic iron accumulation and hepatic fibrosis by immunohistochemistrical study. METHODS:: Forty-six hepatic tissues with chronic hepatitis C and five normal hepatic tissues were examined. Chemical detection of hepatic iron accumulation was performed by Perl's Prussian blue stain. The immunohistochemistrical study was performed by avidin-biotin complex method with alkaline phosphatase. RESULTS:: In chronic hepatitis C: (1) Hepatic iron accumulation was significantly increased in relation to the advance of the fibrosis. (2) Divalent metal transporter 1 decreased significantly in relation to the advance of hepatic fibrosis. (3) Transferrin receptor 1 expression was always detected, although not in normal hepatic tissues; there was no relation between expression levels and the degree of hepatic fibrosis. CONCLUSIONS:: These data demonstrated that the transferrin receptor 1 expression was up-regulated irrespective of the degree of hepatic iron accumulation, suggesting that the up-regulation of transferrin receptor 1 might act as one of the key mechanisms implicated in the accumulation of hepatic iron in chronic hepatitis C.     

Serum levels of soluble form of receptor for advanced glycation end products (sRAGE) are positively associated with circulating AGEs and soluble form of VCAM-1 in patients with type 2 diabetes

We have recently found that soluble form of receptor for advanced glycation end products (sRAGE) levels are positively associated with inflammatory biomarkers and the presence of coronary artery disease (CAD) in type 2 diabetic patients. Since advanced glycation end products (AGEs) up-regulate RAGE expression and endogenous sRAGE could be generated from the cleavage of cell surface RAGE, it is conceivable that sRAGE is positively associated with circulating AGEs levels in diabetes. In this study, we examined whether sRAGE were correlated to circulating levels of AGEs and soluble forms of vascular cell adhesion molecule-1 (sVCAM-1) and intercellular adhesion molecule-1 (sICAM-1) in patients with type 2 diabetes. Eighty-two Japanese type 2 diabetic patients underwent a complete history and physical examination, determination of blood chemistries, sRAGE, AGEs, sVCAM-1 and sICAM-1. Multiple regression analysis revealed that serum levels of AGEs and sVCAM-1 were independently correlated with sRAGE. This study demonstrated that serum levels of sRAGE were positively associated with circulating AGEs and sVCAM-1 levels in type 2 diabetic patients. Our present observations suggest sRAGE level may be elevated in response to circulating AGEs, thus being a novel marker of vascular injury in patients with type 2 diabetes.