Human acetylcholinesterase and butyrylcholinesterase are encoded by two distinct genes

1. Various hybridization approaches were employed to investigate structural and chromosomal interrelationships between the human cholinesterase genes CHE and ACHE encoding the polymorphic, closely related, and coordinately regulated enzymes having butyrylcholinesterase (BuChE) and acetylcholinesterase (AChE) activities. 2. Homologous cosmid recombination with a 190-base pair 5' fragment from BuChEcDNA resulted in the isolation of four overlapping cosmid clones, apparently derived from a single gene with several introns. The Cosmid CHEDNA included a 700-base pair fragment known to be expressed at the 3' end of BuChEcDNA from nervous system tumors and which has been mapped by in situ hybridization to the unique 3q26-ter position. In contrast, cosmid CHEDNA did not hybridize with full-length AChEcDNA, proving that the complete CHE gene does not include AChE-encoding sequences either in exons or in its introns. 3. The chromosomal origin of BuChE-encoding sequences was further examined by two unrelated gene mapping approaches. Filter hybridization with DNA from human/hamster hybrid cell lines revealed BuChEcDNA-hybridizing sequences only in cell lines including human chromosome 3. However, three BuChEcDNA-homologous sequences were observed at chromosomal positions 3q21, 3q26-ter, and 16q21 by a highly stringent in situ hybridization protocol, including washes at high temperature and low salt. 4. These findings stress the selectivity of cosmid recombination and chromosome blots, raise the possibility of individual differences in BuChEcDNA-hybridizing sequences, and present an example for a family highly similar proteins encoded by distinct, nonhomologous genes.     

Cytokines IL-1α, IL-6, and GM-CSF constitutively secreted by oral squamous carcinoma induce down-regulation of CD80 costimulatory molecule expression: restoration by interferon γ

We previously characterized the expression of CD80 in different murine head and neck squamous cell carcinoma (HNSCC) clones derived following tumor progression in the absence of T cell–mediated immunity in severe combined immunodeficient (SCID) mice. We found that HNSCCs that did not express CD80 grew as progressors, while those that expressed CD80 were regressors when grown in immune-competent animals. In the present study, we characterized expression of a repertoire of immunoregulatory cytokines in these HNSCC lines, and found that HNSCCs that express cytokines IL-1, IL-6, and GM-CSF do not express CD80, suggesting the hypothesis that these cytokines may down-modulate expression of CD80. Cytokine-conditioned medium from progressor HNSCC and recombinant IL-1, IL-6, and GM-CSF caused a reduction of CD80 expression in regressor HNSCCs without affecting proliferation. Conversely, the decrease in CD80 expression in progressor HNSCCs could be restored by IFN-, a known inducer of CD80 expression. These data strongly suggest that high levels of cytokines IL-1, IL-6, and GM-CSF expressed by tumor cells can down-regulate CD80 expression in HNSCC, and that IFN- can independently stimulate expression. These data provide evidence for a novel mechanism of cytokine-mediated down-modulation of CD80 during malignant progression of HNSCC that can be restored by IFN-.     

Inhibition of cell proliferation in human breast tumor cells by antisense oligonucleotides against facilitative glucose transporter 5

In recent years, successful examples of antisense oligonucleotide (AS) therapy for genetic diseases have stimulated scientists to investigate its application on cancer diseases. AS can be used to down-regulate the mRNA and protein expression by annealing to specific region of the target mRNA which is responsible for the malignancy. Glucose transporter 5 (Glut5) is a tissue specific transporter that can be found on breast cancer tissues but not on normal breast tissues. Therefore, it is of clinical interest to investigate whether AS against Glut5 mRNA can tackle breast cancer. In this study, two cell lines, MCF-7 which is estrogen-receptor positive and MDA-MB-231 which is estrogen-receptor negative, were used to mimic breast cancer tissues at early and late stages, respectively. A 15-base sequence around the start codon of Glut5 was used. It was found that AS against Glut5 exerted anti-proliferative effect on both of these two breast tumor cell lines and seemed to exert its effect via the suppression of expression of Glut5 proteins in the cells. AS against Glut5 exhibited no effect on human hepatoma HepG2 cells which do not possess any Glut5. The results imply an alternative way in treating breast tumor as the AS against Glut5, unlike tamoxifen, takes effect on breast tumor cells via suppressing the expression of Glut5 that they specifically possess, and regardless whether the breast tumors are estrogen dependent or not.     

Thyrotropin and serum regulate thyroid cell proliferation through differential effects on p27 expression and localization

Thyroid cell proliferation is regulated by the concerted action of TSH/cAMP and serum growth factors. The specific contributions of cAMP-dependent vs. -independent signals to cell cycle progression are not well understood. We examined the molecular basis for the synergistic effects of TSH and serum on G1/S phase cell cycle progression in rat thyroid cells. Although strictly required for thyroid cell proliferation, TSH failed to stimulate G1 phase cell cycle progression. Together with serum, TSH increased the number of cycling cells. TSH enhanced the effects of serum on retinoblastoma protein hyperphosphorylation, cyclin-dependent kinase 2 activity, and cyclin A expression. Most notably, TSH and serum elicited strikingly different effects on p27 localization. TSH stimulated the nuclear accumulation of p27, whereas serum induced its nuclear export. Unexpectedly, TSH enhanced the depletion of nuclear p27 in serum-treated cells. Furthermore, only combined treatment with TSH and serum led to rapamycin-sensitive p27 turnover. Together, TSH and serum stimulated p70S6K activity that remained high through S phase. These data suggest that TSH regulates cell cycle progression, in part, by increasing the number of cycling cells through p70S6K-mediated effects on the localization of p27.     

Photocatalytic inhibition of microbial adhesion by anodized titanium

Biofouling is one of the concerns in the use of titanium for seawater cooled condensers of power plants. Earlier studies have shown that anodized titanium and its alloys with a thin film of anatase (TiO(2)) on its surface can inhibit attachment of Pseudomonas sp. when illuminated with near-UV light (350 - 380 nm). In the present study, a comparison of the photocatalytic inhibition of microbial attachment on titanium surfaces anodized at different voltages was carried out. Thin films of anatase of varying thickness were produced on titanium grade-2 by anodizing in dilute orthophosphoric acid solution at 30 V, 50 V and 100 V. The photocatalytic efficiency of these anodized surfaces was measured by the methylene blue degradation method. The anodised surfaces were exposed to liquid cultures of Gram-negative Pseudomonas sp., Gram-positive Micrococcus sp. and to a mixed algal culture. Photocatalytic inhibition of microbial attachment was maximum on the titanium surface anodized at 30 V, followed by the surface anodized at 50 V and then at 100 V. The photocatalytic inhibition of microbial attachment was also found to be dependent on the cell wall characteristics of the organism. The Gram-negative Pseudomonas sp. with a lipoproteinaceous outer membrane was the most susceptible to the photocatalytic effect, while the Gram-positive Micrococcus sp. with peptidoglycan cell wall showed moderate susceptibility and the algae with siliceous cell wall showed no susceptibility at all.     

Efficacy of three microbiological monitoring methods in a ventilated cage rack

The use of individually ventilated caging (IVC) to house mice presents new challenges for effective microbiological monitoring. Methods that exploit the characteristics of IVC have been developed, but to the authors' knowledge, their efficacy has not been systematically investigated. Air exhausted from the IVC rack can be monitored, using sentinels housed in cages that receive rack exhaust air as their supply air, or using filters placed on the exhaust air port. To aid laboratory animal personnel in making informed decisions about effective methods for microbiological monitoring of mice in IVC, the efficacy of air monitoring methods was compared with that of contact and soiled bedding sentinel monitoring. Mice were infected with mouse hepatitis virus (MHV), mouse parvovirus (MPV), murine rotavirus (agent of epizootic diarrhea of mice [EDIM]), Sendai virus (SV), or Helicobacter spp. All agents were detected using contact sentinels. Mouse hepatitis virus was effectively detected in air and soiled bedding sentinels, and SV was detected in air sentinels only. Mouse parvovirus and Helicobacter spp. were transmitted in soiled bedding, but the efficacy of transfer was dependent on the frequency and dilution of soiled bedding transferred. Results were similar when the IVC rack was operated under positive or negative air pressure. Filters were more effective at detecting MHV and SV than they were at detecting MPV. Exposure of sentinels or filters to exhaust air was effective at detecting several infectious agents, and use of these methods could increase the efficacy of microbiological monitoring programs, especially if used with soiled bedding sentinels. In contemporary mouse colonies, a multi-faceted approach to microbiological monitoring is recommended.