Comparing the performance of one-column process and four-zone simulated moving bed by computer simulation

A new one-column chromatography process, analogous to a four-zone simulated moving bed (SMB), was presented. The basic principle of the process was identical to that of a four-zone SMB. The process consisted of one chromatographic column and four tanks, instead of the four columns in the four-zone SMB (1-1-1-1), and has been used for the separation of two amino acids, phenylalanine and tryptophan, using an ion exchange resin. The operating parameters for the one-column process and four-zone SMB were obtained from equilibrium theory. Computer simulations were used to compare the performances of the new one column process to that of the general four-zone SMB, using Aspen Chromatography^™ v 11.1. The differences between the one-column and SMB processes in terms of the purities and yields of phenylalanine and tryptophan were less than 4 and about 6%, respectively. The lower purities of the one-column process were due to the loss of the developed concentration profiles in the column when the liquid was stored in tanks. The one-column process gave great flexibility, and would be useful for reconstructing an existing conventional chromatography process to one of a SMB.     

Distribution of lichen flora on South Korea

After an overview on the temporary situation of the lichenology in South Korea, localities of 95 macrolichen taxa are reported for South Korea. In this revised lichen flora of South Korea, 16 species are apparently new to the territory. Voucher specimens have been deposited in the Korean Lichen Research Institute (KoLRI) at Sunchon National University in Korea, and duplicates have also been donated to the National History Museum and Institute, in Chiba, (CBM) Japan.     

The influence of NaCl and carbonylcyanide-m-chlorophenylhydrazone on the production of extracellular proteases in a marine Vibrio strain

In general, the salinity of the ocean is close to 3.5% and marine vibrios possess the respiratory chain-linked Na+ pump. The influence of sodium chloride and the proton conductor carbonylcyanide m-chlorophenylhydrazone (CCCP) on the production of extracellular proteases in a marine Vibrio strain was examined. At the concentration of 0.5 M, sodium chloride minimally inhibited the activity of extracellular proteases by approximately 16%, whereas at the same concentration, the producton of extracellular proteases was severely inhibited. On the other hand, the production of extracellular proteases was completely inhibited by the addition of 2 microM CCCP at pH 8.5, where the respiratory chain-linked Na+ pump functions.     

Molecular characterization of the transition to malignancy in a genetically engineered mouse-based model of ductal carcinoma in situ

A transplantable model of human ductal carcinoma in situ that progresses to invasive carcinoma was developed from a genetically engineered mouse (GEM). Additional lines were established using early mammary premalignant lesions from transgenic MMTV-PyV-mT mice. These lines were verified to be premalignant and transplanted repeatedly to establish stable and predictable properties. Here, we report the first in-depth molecular analysis of neoplastic progression occurring in one premalignant transplantable GEM-derived line. Oligonucleotide microarrays showed that many genes are differentially expressed between the quiescent and prelactating mammary gland and the premalignant GEM outgrowth. In contrast, a small but consistent group of genes was associated with the transformation from premalignancy to tumor. This suggests that the majority of gene expression changes occur during the premalignant transition from normal to premalignancy, whereas many fewer changes occur during the malignant transition from premalignancy to invasive carcinoma. The premalignant transition is associated with several cell cycle-related genes and the up-regulation of oncogenes is associated with various cancers (Ccnd11, Cdk4, Myb, and Ect2). The changes identified in the malignant transition included genes previously associated with human breast cancer progression. Misregulation of the insulin-like growth factor and transforming growth factor-beta signaling pathways and the stromal-epithelial interaction were implicated. Our results suggest that this transplantable GEM-based model recapitulates human ductal carcinoma in situ at both histologic and molecular levels. With consistent tumor latency and molecular profiles, this model provides an experimental platform that can be used to assess functional genomics and molecular pharmacology and to test promising chemoprevention strategies.     

Phylogenomics of the reproductive parasite Wolbachia pipientis wMel: a streamlined genome overrun by mobile genetic elements

The complete sequence of the 1,267,782 bp genome of Wolbachia pipientis wMel, an obligate intracellular bacteria of Drosophila melanogaster, has been determined. Wolbachia, which are found in a variety of invertebrate species, are of great interest due to their diverse interactions with different hosts, which range from many forms of reproductive parasitism to mutualistic symbioses. Analysis of the wMel genome, in particular phylogenomic comparisons with other intracellular bacteria, has revealed many insights into the biology and evolution of wMel and Wolbachia in general. For example, the wMel genome is unique among sequenced obligate intracellular species in both being highly streamlined and containing very high levels of repetitive DNA and mobile DNA elements. This observation, coupled with multiple evolutionary reconstructions, suggests that natural selection is somewhat inefficient in wMel, most likely owing to the occurrence of repeated population bottlenecks. Genome analysis predicts many metabolic differences with the closely related Rickettsia species, including the presence of intact glycolysis and purine synthesis, which may compensate for an inability to obtain ATP directly from its host, as Rickettsia can. Other discoveries include the apparent inability of wMel to synthesize lipopolysaccharide and the presence of the most genes encoding proteins with ankyrin repeat domains of any prokaryotic genome yet sequenced. Despite the ability of wMel to infect the germline of its host, we find no evidence for either recent lateral gene transfer between wMel and D. melanogaster or older transfers between Wolbachia and any host. Evolutionary analysis further supports the hypothesis that mitochondria share a common ancestor with the α-Proteobacteria, but shows little support for the grouping of mitochondria with species in the order Rickettsiales. With the availability of the complete genomes of both species and excellent genetic tools for the host, the wMel–D. melanogaster symbiosis is now an ideal system for studying the biology and evolution of Wolbachia infections.     

Molecular signatures of proliferation and quiescence in hematopoietic stem cells

Stem cells resident in adult tissues are principally quiescent, yet harbor enormous capacity for proliferation to achieve self renewal and to replenish their tissue constituents. Although a single hematopoietic stem cell (HSC) can generate sufficient primitive progeny to repopulate many recipients, little is known about the molecular mechanisms that maintain their potency or regulate their self renewal. Here we have examined the gene expression changes that occur over a time course when HSCs are induced to proliferate and return to quiescence in vivo. These data were compared to data representing differences between naturally proliferating fetal HSCs and their quiescent adult counterparts. Bioinformatic strategies were used to group time-ordered gene expression profiles generated from microarrays into signatures of quiescent and dividing stem cells. A novel method for calculating statistically significant enrichments in Gene Ontology groupings for our gene lists revealed elemental subgroups within the signatures that underlie HSC behavior, and allowed us to build a molecular model of the HSC activation cycle. Initially, quiescent HSCs evince a state of readiness. The proliferative signal induces a preparative state, which is followed by active proliferation divisible into early and late phases. Re-induction of quiescence involves changes in migratory molecule expression, prior to reestablishment of homeostasis. We also identified two genes that increase in both gene and protein expression during activation, and potentially represent new markers for proliferating stem cells. These data will be of use in attempts to recapitulate the HSC self renewal process for therapeutic expansion of stem cells, and our model may correlate with acquisition of self renewal characteristics by cancer stem cells.     

Evaluation of Antigen Retrieval Buffer Systems

The introduction of antigen retrieval (AR) techniques has dramatically improved the sensitivity of immunohistochemical detection of various antigens in formalin-fixed, paraffin-embedded tissues. The microwave-heating and pressure-cooking procedures are the most effective AR methods reported to date. Although extensive efforts have been made to optimize AR procedures using these two methods, previous studies have not led to a standard protocol applicable to all antibodies derived from different clones. In this study we have investigated the optimal AR buffer conditions for 29 antibodies that are in common use for diagnostic purposes in hospitals worldwide. Borate (pH 8.0) and Tris buffer (pH 9.5) yielded the highest retrieved antigen immunoreactivity against most antibodies as compared to other buffers tested. In addition, the microwave pressure-cooking gave better results than microwave-heating alone. Therefore, borate (pH 8.0) or Tris (pH 9.5) buffer used in conjunction with the pressure-cooking procedure is strongly recommended for standard routine use.     

Analysis of substance P in rat brain by means of immunoaffinity capture and matrix-assisted laser desorption/ionization time-of-flight mass-spectrometry

Interest in the analysis of low abundance neuropeptides particularly using matrix-assisted laser desorption/ionization time-of-flight mass-spectrometry (MALDI-TOF-MS) is increasing because these neuropeptides are essential to the mechanism of transportation and the metabolism. This article describes an immunoprecipitation procedure that is suitable for MALDI-MS analysis of substance P (SP), a neuropeptide, in rat brain tissues. Substance P was precipitated from brain tissue extracts by immunoprecipitation with antibodies directed against SP, and are analyzed by MALDI-TOF-MS. Mass spectrometric analysis showed a singly charged [M+H]+ ion peak that corresponded to the SP molecular mass and was observed with a detection error of 1.6%. The average mass errors between the observed and theoretical molecular mass were within the 0.11 Da range. Capillary zone electrophoresis analysis was subsequently performed, and the effects of the different separation parameters were examined. Beginning with milligram quantities of brain tissue, picomole quantities of SP could be detected using this method.     

Automated apparatus for quantitation of social approach behaviors in mice

Mouse models of social dysfunction, designed to investigate the complex genetics of social behaviors, require an objective methodology for scoring social interactions relevant to human disease symptoms. Here we describe an automated, three chambered apparatus designed to monitor social interaction in the mouse. Time spent in each chamber and the number of entries are scored automatically by a system detecting photocell beam breaks. When tested with the automated equipment, juvenile male C57BL/6J mice spent more time in a chamber containing a stranger mouse than in an empty chamber (sociability), similar to results obtained by the observer scored method. In addition, automated scoring detected a preference to spend more time with an unfamiliar stranger than a more familiar conspecific (preference for social novelty), similar to results obtained by the observer scored method. Sniffing directed at the wire cage containing the stranger mouse correlated significantly with time spent in that chamber, indicating that duration in a chamber represents true social approach behavior. Number of entries between chambers did not correlate with duration of time spent in the chambers; entries instead proved a useful control measure of general activity. The most significant social approach behavior took place in the first five minutes of both the sociability and preference for social novelty tests. Application of these methods to C57BL/6J, DBA/2J and FVB/NJ adult males revealed that all three strains displayed tendencies for sociability and preference for social novelty. To evaluate the importance of the strain of the stranger mouse on sociability and preference for social novelty, C57BL/6J subject mice were tested either with A/J strangers or with C57BL/6J strangers. Sociability and preference for social novelty were similar with both stranger strains. The automated equipment provides an accurate and objective approach to measuring social tendencies in mice. Its use may allow higher-throughput scoring of mouse social behaviors in mouse models of social dysfunction.