Effect of Bcl-2 antisense oligonucleotide on drug-sensitivity in association with apoptosis in undifferentiated thyroid carcinoma

Although attempts have been made to treat undifferentiated thyroid carcinoma using multidisciplinary therapeutic procedures including surgery, radiotherapy, and chemotherapy, the prognosis of undifferentiated thyroid carcinoma remains quite poor. New approaches to increase the sensitivity of patients to anticancer drugs and radiation will be needed to improve the survival rate for undifferentiated thyroid carcinoma. We examined the effect of Bcl-2 antisense oligonucleotide on drug-sensitivity in association with apoptosis in the 8305C undifferentiated thyroid carcinoma cell line. The drug sensitivity was evaluated by MTT assay for 48 h, while apoptosis was assessed according to the formation of internucleosomal DNA ladders. The Bcl-2 antisense was introduced into 8305C cells by using a 18-mer phosphorothioate oligonucleotide by lipopolyamine-mediated transfection twice for 12 h. The expression of apoptosis genes was assessed by Western blotting. The 8305C cells were sensitive to adriamycin (ADM), mitomycin (MMC), docetaxel (TXT), and paclitaxel (TXL), showing mean IC50 values of 0.72, 1.1, 1.3, and 4.1 microM, respectively. In contrast, the 8305C cells were resistant to cisplatin (CDDP) and 5-fluorouracil (5-FU), with mean IC50 values of 42.0 and 48.0 microM, respectively. Treatment with Bcl-2 antisense suppressed the protein level of Bcl-2 in 8305C cells in a dose-dependent manner up to 1.0 microM. Drug-sensitivity was increased by pretreatment with Bcl-2 antisense as assessed by the IC50 (x-fold): 0.48 (1.5-fold) in ADM; 0.42 (2.6-fold) in MMC, 0.56 (2.3-fold) in TXT, 1.5 (2.7-fold) in TXL, 8.6 (4.9-fold) in CDDP, and 25.0 (1.9-fold) in 5-FU, respectively. The increased drug-sensitivity was associated with the induction of apoptosis-related proteins, Fas, caspase 8, cytochrome c, caspase 3, and to subsequent apoptosis, as determined by the formation of internucleosomal DNA ladders and PARP in the treated cells. Susceptibility in apoptotic cell death following treatment with anticancer drugs was associated with induction of apoptosis-related genes in undifferentiated thyroid carcinoma cells, and induction of apoptosis was enhanced by pretreatment with Bcl-2 antisense oligonucleotide. These results imply a potential new strategy targeting an antiapoptotic protein, Bcl-2, by its antisense oligonucleotide for enhancement of chemotherapeutic efficacy in undifferentiated thyroid carcinomas.     

The fragile X in cattle

In search of an animal model for the human fragile X syndrome, the chromosomes of Holstein cows were examined. This breed was chosen because of previous studies on the baldy calf syndrome. An achromatic gap was observed at a specific site on the X chromosome closer to the centromere than that identified in humans. This unstained gap was found in 3%-4% of cells of the following four animals: an affected calf, her sister, their mother, and an unrelated Holstein cow. The bovine fragile X may not be analogous to the human fragile X but its location may be important as a genetic marker in linkage studies involving the loci for hypoxanthine phosphoribosyltransferase (HPRT) and glucose-6-phosphate dehydrogenase (G-6-PD).     

Correlation dimension analysis of the artificial circulation

Artificial circulation has been analyzed by decomposing it into parts. However, the sum of the decomposed parts is not equal to the whole system, especially in nonlinear dynamic systems such as biological systems. To evaluate prosthetic circulation as an entity, not as decomposed parts, nonlinear mathematical analytic techniques, including fractal dimension analyzing theory, were used. Two pneumatically actuated ventricular assist devices were implanted as biventricular bypasses (BVB) in chronic animal experiments using four healthy adult goats. For comparison between natural and prosthetic circulation in the same experimental animals, the BVB-type complete prosthetic circulation model with ventricular fibrillation was adopted. All hemodynamic parameters with natural and prosthetic circulation were recorded under awake conditions and calculated by a personal computer system. By the use of nonlinear mathematical techniques, time-series data of the hemodynamics were embedded into the phase space, and correlation dimension analysis was performed to evaluate the reconstructed attractor. Our results suggest that the correlation dimension of the arterial blood pressure does not linearly increase according to the increase of the embedding dimension, even during artificial circulation, suggesting those are the fractal time series data. Dimensional analysis of the hemodynamics revealed that lower dimensional fractal dynamics were observed during prosthetic circulation. Fractal time series data are suggested to have robustness and error resistance. Thus, our results suggest that the circulatory regulatory system with the artificial heart may have these desirable characteristics. Accepted: July 14, 1995     

Phase stability after aging and its influence on pin-on-disk wear properties of Ce-TZP/Al2O3 nanocomposite and conventional Y-TZP

Recently zirconia/alumina composites have been examined by many researchers as the new generation of bearing materials in total joint replacements. In this study, the phase stability of a Ce-TZP/Al(2)O(3) nanocomposite and conventional Y-TZP after aging, and its influence on wear resistance, were investigated. Very slight phase transformation was observed in both types of ceramics 18 months after the implantation of Ce-TZP/Al(2)O(3) or Y-TZP samples into rabbit tibiae. However, Y-TZP showed marked phase transformation (approximately 80%) after aging in an autoclave (121 degrees C) for 190 h or in physiological saline at 62 degrees C for 18 months, whereas the new composite remained almost resistant to degradation. According to the results of self-pairing pin-on-disk wear tests using ceramic specimens with or without autoclave aging, the wear factor was almost the same between Ce-TZP/Al(2)O(3) samples with and without aging (6.74 +/- 0.36 x 10(-8) and 6.04 +/- 0.95 x 10(-8) mm(3)/Nm, respectively). In contrast, although non-aged Y-TZP had the lowest wear factor (4.88 +/- 0.51 x 10(-8) mm(3)/Nm) of all specimens tested, aged Y-TZP showed 10-fold greater wear than nonaged Y-TZP. The present study suggests that Ce-TZP/Al(2)O(3) nanocomposite has much greater phase stability than Y-TZP, and that its wear properties are not influenced by aging.     

Over-expression of TSC-22 (TGF-beta stimulated clone-22) markedly enhances 5-fluorouracil-induced apoptosis in a human salivary gland cancer cell line

We have recently isolated TSC-22 (transforming growth factor-beta-stimulated clone-22) cDNA as an anticancer, drug-inducible (with vesnarinone) gene in a human salivary gland cancer cell line, TYS. We have also reported that TSC-22 negatively regulates the growth of TYS cells and that down-regulation of TSC-22 in TYS cells plays a major role in salivary gland tumorigenesis (Nakashiro et al, 1998). In this study, we transfected TYS cells with an expression vector encoding the TSC-22-GFP (green fluorescent protein) fusion protein, and we established TSC-22-GFP-expressing TYS cell clones. Next, we examined (a) the subcellular localization of the fusion protein, (b) the sensitivity of the transfectants to several anticancer drugs (5-fluorouracil, cis-diaminedichloroplatinum, peplomycin), and (c) induction of apoptotic cell death in the transfectants by 5-fluorouracil treatment. The TSC-22-GFP fusion protein was clearly localized to the cytoplasm, but not to the nucleus. Over-expression of the TSC-22-GFP fusion protein did not affect cell growth, but significantly increased the sensitivity of the cells to the anticancer drugs (p < 0.01; one-way ANOVA). Furthermore, over-expression of the TSC-22-GFP fusion protein markedly enhanced 5-fluorouracil-induced apoptosis. These findings suggest that over-expression of TSC-22-GFP protein in TYS cells enhances the chemosensitivity of the cells via induction of apoptosis.     

Immunoperoxidase staining of hepatitis C virus in formalin-fixed,paraffin-embedded needle liver biopsies

The localization of hepatitis C virus (HCV) in the liver has not been well clarified. We report successful indirect immunoperoxidase staining of the HCV core antigen using polyclonal antibodies raised in rabbits and conventional formalin-fixed, paraffin-embedded needle biopsy sections of liver. The core antigen was distributed in a fine granular pattern diffusely, perisinusoidally, or focally within the hepatocellular cytoplasm of livers from patients with HCV infection. The staining tended to show a more heterogeneous pattern in terms of intensity and distribution in cases of more advanced disease. Hepatocellular carcinoma cells were also frequently stained. HCV immunostaining will provide important information on the pathogenesis and treatment of HCV-related liver diseases.     

Trials of some improved crystal vibrators for use in ultrasonic methods of gynecological diagnosis

Medical &; Biological Engineering &; Computing -     

Dynamics of gastrin-producing cells in the rat after truncal vagotomy. Evaluation by double immunostaining for bromodeoxyuridine and little gastrin.

The mechanism of hyperplasia of gastrin-producing cells (G-cells) in the rat antral mucosa after truncal vagotomy was studied using double immunostaining for bromodeoxyuridine (BrdU) and little gastrin (G17). With single labeling of BrdU, a few G-cells (less than 1%) showed positive immunostaining for BrdU in the nucleus throughout the experimental period in both vagotomized rats and those given a sham operation. The labeled cells in both groups demonstrated a linear increase of BrdU labeling in an identical number of cells for each experimental time-point. The labeling index of the G-cells increased rapidly from day 2 to day 6 and attained a maximum level of 44.0% on day 10 in the vagotomized group after cumulative labeling. Even in this group, however, many G-cells showed no BrdU immunoreactivity throughout the experimental period. These cells did not replicate during the experimental period, but showed an intense reaction product for G17 in their cytoplasm after vagotomy. The present study indicates that the most important factor involved in G-cell hyperplasia observed after truncal vagotomy is the activation of pre-existing G-cells to synthesize and release hormone, together with the rapid maturation of progenitor cells to mature G-cells.