The corrosion resistance of pure titanium in organic acids

The purpose of this study was to assess the corrosive properties of titanium at various pH values. Cast pure titanium specimens were immersed in 128 mmol/l of lactic and formic acids at pH 1.0-8.5 for 3 weeks at 37 degrees C. The solubility, color, weight and chemical binding state of specimens were observed. Titanium dissolved in all lactic acid. The amount of dissolved titanium tended to decrease with a higher pH. In formic acid, the amount of dissolved titanium at pH 1.0 was larger than that in lactic acid at the same pH, but less than the detectable limit at pH 4.0 or higher. Significant discoloration was macroscopically observed only in formic acid at pH 2.5 and 4.0. The weight of the titanium samples immersed in lactic acid all decreased, but it was not affected by pH. In formic acid, the weight decreased at pH 1.0 and increased at pH 2.5-5.5. Thickening of the TiO2 corresponding to that showing discoloration was observed in the superficial oxide film of the titanium samples. Our results show that the corrosive properties of titanium are markedly dependent on pH in formic acid, and relatively less dependent on pH in lactic acid in which titanium is dissolvable at pH 1.0-8.5.     

Affinity chromatography for purification of two urokinases from human urine

A new affinity chromatography (hydrophobic-mediated affinity chromatography), which was characterized by the matrix having both affinity site to urokinase and hydrophobic site, was established for the purification of urokinase from human urine. The hydrophobic affinity matrix (tentatively named PAS in the text) was prepared by immobilizing 6-aminocaproic acid on Sepharose CL-6B, followed by a coupling p-aminobenzamidine to a part of the hydrophobic site on the matrix. The PAS matrix was applied to the purification of urokinase from human urine, and high- and low-molecular weight pure urokinases were efficiently obtained in high yield by the present method.     

Transgenic rabbits with increased VEGF expression develop hemangiomas in the liver: a new model for Kasabach-Merritt syndrome

Clinical studies have provided ample evidence that high (either systemic or local) levels of vascular endothelial growth factor (VEGF) are associated with several pathophysiological disorders, including hemangiomas. To investigate whether elevated VEGF expression could directly affect these disorders, we created a transgenic (Tg) rabbit model with increased hepatic expression of the human VEGF(165) transgene under the control of the human alpha-antitrypsin promoter. Tg rabbits exhibited marked hepatomegaly, with livers 2.5-fold heavier than those of control rabbits. Histological analysis revealed that the livers of Tg rabbits showed prominent dilation of the sinusoids and formed various-sized blood vessel networks, a feature of diffuse hemangiomas. Immunohistochemical staining revealed that the hepatocytes produced VEGF(165), whereas plasma VEGF(165) was not detected. Furthermore, Tg rabbits suffered from hemolytic anemia, thrombocytopenia and splenomegaly, which was associated with marked extramedullary hematopoiesis. The manifestations of Tg rabbits mimic many of the features of hemangiomatous disorders in humans such as the Kasabach-Merritt syndrome, and therefore this model may be potentially useful for the study of the pathogenesis and complications of hemangiomas as well as the investigation of angiogenesis inhibitors.     

Prediction of sensitivity of advanced non-small cell lung cancers to gefitinib (Iressa, ZD1839)

Gefitinib (Iressa, ZD1839), an inhibitor of epidermal growth factor receptor-tyrosine kinase, has shown potent anti-tumor effects and improved symptoms and quality-of-life of a subset of patients with advanced non-small cell lung cancer (NSCLC). However, a large portion of the patients showed no effect to this agent. To establish a method to predict the response of NSCLC patients to gefitinib, we used a genome-wide cDNA microarray to analyze 33 biopsy samples of advanced NSCLC from patients who had been treated with an identical protocol of second to seventh line gefitinib monotherapy. We identified 51 genes whose expression differed significantly between seven responders and 10 non-responders to the drug. We selected the 12 genes that showed the most significant differences to establish a numerical scoring system (GRS, gefitinib response score), for predicting response to gefitinib treatment. The GRS system clearly separated the two groups without any overlap, and accurately predicted responses to the drug in 16 additional NSCLC cases. The system was further validated by the semi-quantitative RT-PCR, immunohistochemistry and ELISA for serological test. Moreover, we proved that the anti-apoptotic activity of amphiregulin, a protein that was significantly over-expressed in non-responders but undetectable in responders, leads to resistance of NSCLC cells to gefitinib in vitro. Our results suggested that sensitivity of a given NSCLC to gefitinib can be predicted according to expression levels of a defined set of genes that may biologically affect drug sensitivity and survival of lung cancer cells. Our scoring system might eventually lead to achievement of personalized therapy for NSCLC patients.     

Expression and intracellular localization of FKHRL1 in mammary gland neoplasms

FKHRL1 (FOXO3a), a member of the Forkhead family of genes, has been considered to be involved in the development of breast tumors; however, the in vivo expression and activation status of FKHRL1 in breast tumors still remains unclear. We immunohistochemically demonstrated the expression and intracellular localization of FKHRL1 in human breast tumors by the novel anti-FKHRL1 antibody which is available for formalin-fixed paraffin-embedded specimens. In a total of 51 cases of benign tumors, FKHRL1 was diffusely expressed in all cases, and its intracellular localization was revealed to be cytoplasmic (inactive form) in 94% of cases of intraductal papillomas (16/17) and 91% cases of fibroadenomas (31/34), with a similar pattern to normal glandular epithelium. In invasive ductal carcinomas, 83% of the cases (93/112) diffusely expressed FKHRL1; however, unlike benign tumors, 71% of the cases (66/93) showed the nuclear-targeted, active form of FKHRL1. Moreover, activated FKHRL1 was predominantly observed in scirrhous (29/36, 81% of the cases) and papillotubular (30/38, 79% of the cases) subtypes, compared to the solid-tubular subtype (7/19, 37% of the cases). Furthermore, the cases with nuclear-targeted FKHRL1 showed a tendency to have lymph nodal metastasis with statistical significance (P < 0.0001). Thus, the activation of FKHRL1 seems to be recognized as one of the specific features of invasive ductal carcinoma of the breast.     

Purification, cDNA cloning, and secretory properties of FLRG protein from PC12 cells and the distribution of FLRG mRNA and protein in rat tissues

A 35 kD protein was isolated and purified from conditioned media of Bcl-2 cDNA-transfected PC12 cells and its cDNA cloned. A database analysis showed that the 35 kD protein is a rat homologue of the human FLRG protein. The biochemical as well as morphological properties of the rat FLRG protein in PC12 cells were examined and its distribution in rat tissues determined. The levels of FLRG mRNA expressed were low during the fetal period, compared with those of follistatin mRNA. The distribution of FLRG and follistatin mRNAs differed from each other after birth; the expression levels of FLRG mRNA were abundant in the adrenal gland and testis, whereas those of follistatin mRNA and activin A were markedly high in the ovary. The presence of FLRG mRNA and/or protein was confirmed in spermatocytes at various differentiating stages andin endocrine cells of both the adrenal cortex and medulla. When overexpressed in PC12 cells, the FLRG protein was found to be stored in secretory granules of the cells and largely secreted by a regulated pathway, while activin A enhancedthe constitutive secretion of the FLRG protein from wild-typpe PC12 cells, indicating that the FLRG protein possesses dualproperties in secretory pathways. The different distribution between FLRG and follistatin mRNA suggests that, like follistatin in the ovary, the FLRG protein may be involved in the maintenance of spermatogenesis in the testis and the growth and function of adrenal tissue cells, probably by regulating the functions of its binding partners such as the TGF-beta ( superfamily members.     

In vivo study of the effects of cryopreservation on heart valve xenotransplantation.

BACKGROUND: Recent reports have suggested that cryopreservation reduces the immunogenicity of donor tissue. The immunomodulation by cryopreservation might influence on the tissue durability after xenotransplantation. We investigated the in vivo morphologic changes in cryopreserved xenograft (CXG) heart valves. MATERIAL AND METHOD: We transplanted a fresh (fresh xenograft; FXG) and a cryopreserved (CXG) porcine aortic root and a cryopreserved canine (cryopreserved allograft; CAG) aortic root into the abdominal aorta of a dog without any immunosuppressive agents. Explanted grafts on the 21st to 49th days after implantation were analyzed morphologically with light microscopy using some special stains, immunohistochemical analysis, and scanning electron microscopy (SEM). RESULT: Light microscopy showed the absence of smooth muscle cells in the media of the aorta in any group after transplantation. FXG valves did not maintain any cellularity after transplantation. CXG valves contained cellular infiltration in themselves. CAG valves contained numerous fibroblasts, which showed the maintenance of tissue integrity without allowing cellular infiltration. The structure of elastic fibers was well maintained, even in the part of CXG valve with cellular infiltration. Immunohistochemical studies documented the infiltration of T lymphocytes in CXG valves that were labeled by anti-CD3 antibodies. SEM demonstrated that no endothelia were seen on the surface of the valves in any group after transplantation. CONCLUSION: We concluded that the cryopreservation method might provide an immunomodulation of xenogeneic heart valves for transplantation.     

Heat-shock induced nuclear retention and recycling inhibition of importin alpha

Heat-shock induces a strong stress response and modifies all aspects of cellular physiology, which involves dynamic changes in the nucleocytoplasmic distributions of a variety of proteins. Many distinct nucleocytoplasmic transport pathways exist in eukaryotic cells, but how a particular transport pathway is regulated under different cellular conditions remains elusive. The finding of this study indicate that conventional nuclear import, which is mediated by importin alpha/beta, is down-regulated, while the nuclear import of 70 kD heat-shock cognate protein is up-regulated in heat-shock cells. Among the factors involved in the mediation of the conventional nuclear import, significant levels of importin alpha accumulate in the nucleus in response to heat-shock. An analysis of the behaviour of importin alpha with fluorescence recovery after photobleaching and fluorescence loss in photobleaching studies show that nuclear importin alpha becomes less mobile and its nucleocytoplasmic recycling is impaired in heat-shock cells. These data coincided well with biochemical and cytological studies. Our present data show that heat-shock induces the nuclear accumulation, nuclear retention, and recycling inhibition of importin alpha, resulting in the suppression of conventional nuclear import. This suggests a new regulatory mechanism for the adaptation of cells to environmental changes, such as heat-shock.