Computer program for the IBM personal computer which searches for approximate matches to short oligonucleotide sequences in long target DNA sequences.

We describe a program which may be used to find approximate matches to a short predefined DNA sequence in a larger target DNA sequence. The program predicts the usefulness of specific DNA probes and sequencing primers and finds nearly identical sequences that might represent the same regulatory signal. The program is written in the C programming language and will run on virtually any computer system with a C compiler, such as the IBM/PC and other computers running under the MS/DOS and UNIX operating systems. The program has been integrated into an existing software package for the IBM personal computer (see article by Mount and Conrad, this volume). Some examples of its use are given.     

Regulation of the pro-angiogenic microenvironment by carboxyamido-triazole

Anti-angiogenic agents regulate tumor growth by inhibiting endothelial cell proliferation and invasion. Carboxyamido-triazole (CAI), an inhibitor of non-voltage-operated calcium entry and calcium influx-mediated pathways, has angiogenesis and invasion inhibitory activity. We hypothesized that CAI may express its anti-angiogenic effects through negative regulation of pro-angiogenic cytokine production and/or function. In vivo, orally administered CAI prevented A2058 human melanoma xenograft growth and concomitantly resulted in a marked reduction in circulating vascular endothelial growth factor (VEGF) and interleukin-8 (IL-8). In vitro, A2058 cell secretion of VEGF was inhibited by CAI treatment under limiting micronutrient conditions that approximate the tumor microenvironment, media restriction, and acidification to pH 6.8 (P=0.0003 and P=0.0006, respectively). VEGF and HIF-1alpha message and protein were also reduced by CAI treatment. Oral CAI treatment reduced vascular ingrowth in vivo into VEGF-containing Matrigel plugs. Commensurate with those findings, human umbilical vein endothelial cell (HUVEC) migration towards VEGF was reduced below background by exposure to CAI in the migration chamber (P<0.0001). An 88% reduction in circulating IL-8 concentration was measured in CAI-treated animals. However, IL-8 protein secretion and gene expression were increased by CAI treatment in culture (P< or =0.01), where CAI caused a dose-dependent acidification of the culture milieu (P< or =0.005). This paradox suggests that IL-8 production in vitro may be more sensitive to ambient pH than cytosolic calcium. These observations suggest that CAI inhibition of tumor cell VEGF production and endothelial cell response to VEGF results in disruption of signaling between the tumor and its microenvironment, causing a net anti-angiogenic effect.     

Development of a Learning-Oriented Computer Assisted Instruction Designed to Improve Skills in the Clinical Assessment of the Nutritional Status: A Pilot Evaluation

Computer assisted instruction (CAI) is an effective tool for evaluating and training students and professionals. In this article we will present a learning-oriented CAI, which has been developed for students and health professionals to acquire and retain new knowledge through the practice. A two-phase pilot evaluation was conducted, involving 8 nutrition experts and 30 postgraduate students, respectively. In each training session, the software developed guides users in the integral evaluation of a patient’s nutritional status and helps them to implement actions. The program includes into the format clinical tools, which can be used to recognize possible patient’s needs, to improve the clinical reasoning and to develop professional skills. Among them are assessment questionnaires and evaluation criteria, cardiovascular risk charts, clinical guidelines and photographs of various diseases. This CAI is a complete software package easy to use and versatile, aimed at clinical specialists, medical staff, scientists, educators and clinical students, which can be used as a learning tool. This application constitutes an advanced method for students and health professionals to accomplish nutritional assessments combining theoretical and empirical issues, which can be implemented in their academic curriculum.     

The blocking of capacitative calcium entry by 2-aminoethyl diphenylborate (2-APB) and carboxyamidotriazole (CAI) inhibits proliferation in Hep G2 and Huh-7 human hepatoma cells

Calcium entry is a component of the processes regulating the proliferative phenotype of some types of cancer. In non-excitable cells, capacitative calcium entry (CCE) and non-capacitative calcium entry (NCCE) are thought to be the main pathways of Ca2+ influx into cells. Thus, blocking calcium entry may prevent normal and pathological cell proliferation and there is evidence to suggest that molecules blocking calcium entry also have antiproliferative properties. Carboxyamidotriazole (CAI), a novel inhibitor of the non-voltage-dependent calcium entry has been shown to have such properties in model systems in vitro and in vivo. We used Hep G2 and Huh-7 human hepatoma cells to investigate the effects of calcium entry blockers on cell proliferation. CAI (10 microM) and 2-APB (20 microM) completely blocked CCE in thapsigargin-treated Huh-7, and CAI and 2-APB inhibited cell proliferation with IC50 of 4.5 and 43 microM, respectively. The plateau phase of the [Ca2+]i increases triggered by 10% FCS were abolished in the absence of external Ca2+ and in the presence of CAI or 2-APB. We, therefore, suggest that CCE is the main pathway involved in regulation of the processes leading to cell proliferation.     

The m-AAA Protease Processes Cytochrome c Peroxidase Preferentially at the Inner Boundary Membrane of Mitochondria

The m-AAA protease is a conserved hetero-oligomeric complex in the inner membrane of mitochondria. Recent evidence suggests a compartmentalization of the contiguous mitochondrial inner membrane into an inner boundary membrane (IBM) and a cristae membrane (CM). However, little is known about the functional differences of these subdomains. We have analyzed the localizations of the m-AAA protease and its substrate cytochrome c peroxidase (Ccp1) within yeast mitochondria using live cell fluorescence microscopy and quantitative immunoelectron microscopy. We find that the m-AAA protease is preferentially localized in the IBM. Likewise, the membrane-anchored precursor form of Ccp1 accumulates in the IBM of mitochondria lacking a functional m-AAA protease. Only upon proteolytic cleavage the mature form mCcp1 moves into the cristae space. These findings suggest that protein quality control and proteolytic activation exerted by the m-AAA protease take place preferentially in the IBM pointing to significant functional differences between the IBM and the CM.     

Rabbits fed cholesterol-enriched diets exhibit pathological features of inclusion body myositis

Sporadic inclusion body myositis (IBM) is the most common age-related muscle disease in humans; however, its etiology is unknown, there are few animal models for this disease, and effective treatments have not been identified. Similarities between pathological findings in Alzheimer's disease brain and IBM skeletal muscle include increased levels of amyloid precursor protein (APP) and amyloid beta-protein (Abeta). Moreover, there have been suggestions that elevated levels of free cholesterol might participate in the pathogenesis of Alzheimer's disease and IBM due, in part, to its role in Abeta generation. Here, we tested the hypothesis that rabbits fed cholesterol-enriched diets might faithfully exhibit human-like IBM pathological features. In skeletal muscle of one-third of the female rabbits fed cholesterol-enriched diet but not control diet, we found features of IBM, including vacuolated muscle fibers, increased numbers of mononuclear inflammatory cells, increased intramuscular deposition of Abeta, hyperphosphorylated tau, and increased numbers of muscle fibers immunopositive for ubiquitin. The cholesterol-enriched diet increased mRNA and protein levels of APP, increased the protein levels of betaAPP cleaving enzyme, and shifted APP processing in favor of Abeta production. Our study has demonstrated that increased ingestion of high levels of dietary cholesterol can result in pathological features that resemble IBM closely and thus may serve as an important new model with which to study this debilitating disorder.     

A didactic computer simulation of the environmental impact of a pollution discharge

Algorithms for simulating the response of a benthic animal communityand several abiotic environmental variables to organic pollutionare described. An IBM PC compatible computer program incorporatingthese methods has been written. The program simulates the abundanceresponse of 23 species, the redox potential depth profile andseveral other sediment variables at any distance from a pointsource of pollution. The program has been designed as a teachingtool and examples of its use to study pollution biology andbiometrics are given. Received on April 3, 1990; accepted on September 6, 1990     

The current status and portability of our sequence handling software.

I describe the current status of our sequence analysis software. The package contains a comprehensive suite of programs for managing large shotgun sequencing projects, a program containing 61 functions for analysing single sequences and a program for comparing pairs of sequences for similarity. The programs that have been described before have been improved by the addition of new functions and by being made very much easier to use. The major interactive programs have 125 pages of online help available from within them. Several new programs are described including screen editing of aligned gel readings for shotgun sequencing projects; a method to highlight errors in aligned gel readings, new methods for searching for putative signals in sequences. We use the programs on a VAX computer but the whole package has been rewritten to make it easy to transport it to other machines. I believe the programs will now run on any machine with a FORTRAN77 compiler and sufficient memory. We are currently putting the programs onto an IBM PC XT/AT and another micro running under UNIX.     

Effects of carboxyamidotriazole on in vitro models of imatinib-resistant chronic myeloid leukemia

Although imatinib mesylate (IM) has revolutionized the treatment of chronic myeloid leukemia (CML), some patients develop resistance with progression of leukemia. Alternative or additional targeting of signaling pathways deregulated in bcr-abl-driven CML cells may provide a feasible option for improving clinical response and overcoming resistance. In this study, we show that carboxyamidotriazole (CAI), an orally bioavailable calcium influx and signal transduction inhibitor, is equally effective in inhibiting the proliferation and bcr-abl dependent- and independent-signaling pathways in imatinib-resistant CML cells. CAI inhibits phosphorylation of cellular proteins including STAT5 and CrkL at concentrations that induce apoptosis in IM-resistant CML cells. The combination of imatinib and CAI also down-regulated bcr-abl protein levels. Since CAI is already available for clinical use, these results suggest that it may be an effective addition to the armamentarium of drugs for the treatment of CML.     

Evidence that the species barrier of human immunodeficiency virus-1 does not extend to uptake by the blood--brain barrier: comparison of mouse and human brain microvessels

HIV-1 within the CNS produces a neuroAIDS syndrome and may act as a reservoir for reinfection of the peripheral tissues. Study of how HIV-1 crosses the blood-brain barrier (BBB) has been hampered by the lack of nonprimate animal models. However, BBB transport of HIV-1 does not involve any of the known steps conferring species specificity, including binding to CD4 receptors. In vivo and in vitro studies show that HIV-1 and its glycoprotein coat, gp120, are taken up and transported across the BBB of the mouse. Here, we compared the ability of gp120 and HIV-1 to be taken up by isolated brain microvessels (IBM) freshly isolated from mice, from post-mortem human brain, and from mice that had been treated in a manner analogous to the human material (mouse post-mortem). Freshly isolated mouse IBM took up more gp120 and HIV-1 than the human or mouse post-mortem cells. We found no difference between the ability of mouse post-mortem and human IBM to take up either gp120 or HIV-1. Wheatgerm agglutinin has been previously shown to stimulate gp120 and HIV-1 uptake by the BBB; here, it stimulated the uptake of gp120 and of HIV-1 by both mouse post-mortem and human IBM, although stimulated uptake was greatest for fresh mouse IBM. These results show that the mouse can be used to study the initial phases of HIV-1 uptake by the BBB.     

Comprehensive sequence analysis of translation termination sites in various eukaryotes

Recent investigations into the translation termination sites of various organisms have revealed that not only stop codons but also sequences around stop codons have an effect on translation termination. To investigate the relationship between these sequence patterns and translation as well as its termination efficiency, we analysed the correlation between strength of consensus and translation efficiency, as predicted according to Codon Adaptation Index (CAI) value. We used RIKEN full-length mouse cDNA sequences and ten other eukaryotic UniGene datasets from NCBI for the analyses. First, we conducted sequence profile analyses following translation termination sites. We found base G and A at position +1 as a strong consensus for mouse cDNA. A similar consensus was found for other mammals, such as Homo sapiens, Rattus norvegicus and Bos taurus. However, some plants had different consensus sequences. We then analysed the correlation between the strength of consensus at each position and the codon biases of whole coding regions, using information content and CAI value. The results showed that in mouse cDNA, CAI value had a positive correlation with information content at positions +1. We also found that, for positions with strong consensus, the strength of the consensus is likely to have a positive correlation with CAI value in some other eukaryotes. Along with these observations, biological insights into the relationship between gene expression level, codon biases and consensus sequence around stop codons will be discussed.     

综合分离分析的微机程序设计及其应用

基于单基因遗传模型,我们在IBM PC/XT微型计算机上,用BASIC语言设计了综合分离分析程序。并应用该计算机程序分析了133个用家族法和家族史法调查的寻常性鱼鳞癣核心家系,98个仅用家族法调查的寻常性鱼鳞癣核心家系和101个原因不明的父母表型正常的感音神经性耳聋核心家系。结果表明,该计算机程序可用于遗传方式分析和遗传咨询。     

Predict7, a program for protein structure prediction

We describe a program for protein sequence analysis which runs in IBM PC computers. Protein sequences are loaded from files in Mount-Conrad and Lipman-Pearson format. Seven features are analyzed: hydrophilicity, hydropathy, surface probability, side chain flexibility, antigenicity, secondary structure and N-glycosylation sites. Numeric results can be shown, printed or stored in files exportable to other programs. Graphics of up to four predictions can be displayed on the screen, printed out or plotted, with several definable options. This program has been designed to be fast, user-friendly and to be shared with the scientific community.     

Footwear Decreases Gait Asymmetry during Running

Previous research on elderly people has suggested that footwear may improve neuromuscular control of motion. If footwear does in fact improve neuromuscular control, then such an influence might already be present in young, healthy adults. A feature that is often used to assess neuromuscular control of motion is the level of gait asymmetry. The objectives of the study were (a) to develop a comprehensive asymmetry index (CAI) that is capable of detecting gait asymmetry changes caused by external boundary conditions such as footwear, and (b) to use the CAI to investigate whether footwear influences gait asymmetry during running in a healthy, young cohort. Kinematic and kinetic data were collected for both legs of 15 subjects performing five barefoot and five shod over-ground running trials. Thirty continuous gait variables including ground reaction forces and variables of the hip, knee, and ankle joints were computed for each leg. For each individual, the differences between the variables for the right and left leg were calculated. Using this data, a principal component analysis was conducted to obtain the CAI. This study had two main outcomes. First, a sensitivity analysis suggested that the CAI had an improved sensitivity for detecting changes in gait asymmetry caused by external boundary conditions. The CAI may, therefore, have important clinical applications such as monitoring the progress of neuromuscular diseases (e.g. stroke or cerebral palsy). Second, the mean CAI for shod running (131.2 ± 48.5; mean ± standard deviation) was significantly lower (p = 0.041) than the CAI for barefoot running (155.7 ± 39.5). This finding suggests that in healthy, young adults gait asymmetry is reduced when running in shoes compared to running barefoot, which may be a result of improved neuromuscular control caused by changes in the afferent sensory feedback.     

Phytopak

PHYTOPAK, a series of computer programs that facilitates the handling and analysis of large sets of phytosociological data is described. The programs are written in FORTRAN IV, and have been implemented on both IBM and CDC computers. The programs have been developed for use in the National Vegetation Classification, currently being undertaken in Britain, where a requirement for handling very large data-sets (up to 35 000 relevés with a total of over 3000 species represented) was the major requirement.The work described here forms part of the National Vegetation Classification supported by the Nature Conservancy Council. The financial support of J. P. H. by this project is gratefully acknowledged. We are particularly grateful to the University of Cambridge Computing Service for providing computing facilities and friendly advice. The conversion of the IBM version of PHYTOPAK into a CDC version was implemented by B.H. at the University of Manchester Regional Computer Centre, with the assistance of Mr. Cliff Stone. Dr. A. J. C. Malloch kindly provided us with his program for renumbering species code numbers which formed the basis of RENUM. We are grateful to Dr. P. Adam, Dr. M. P. Austin, Dr. M. B. Dale, and Mr. M. O. Hill for helpful comments and suggestions.     

Vascular Occlusion Training for Inclusion Body Myositis: A Novel Therapeutic Approach

Inclusion body myositis (IBM) is a rare idiopathic inflammatory myopathy. It is known to produces remarkable muscle weakness and to greatly compromise function and quality of life. Moreover, clinical practice suggests that, unlike other inflammatory myopathies, the majority of IBM patients are not responsive to treatment with immunosuppressive or immunomodulatory drugs to counteract disease progression¹. Additionally, conventional resistance training programs have been proven ineffective in restoring muscle function and muscle mass in these patients^2,3. Nevertheless, we have recently observed that restricting muscle blood flow using tourniquet cuffs in association with moderate intensity resistance training in an IBM patient produced a significant gain in muscle mass and function, along with substantial benefits in quality of life⁴. Thus, a new non-pharmacological approach for IBM patients has been proposed. Herein, we describe the details of a proposed protocol for vascular occlusion associated with a resistance training program for this population.Download video file.^{(106M, mp4)}     

用于质量性状通径分析微机程序设计及其应用

基于质量性状通径分析模型,我们在IBM PC/XT微型计算机上,用BASIC语言设计了通径分析程序PATH。并用该程序分析了315个原因不明精神发育迟滞(MR)的核心家系。分析结果表明,轻度、中度和重度MR的遗传度分别为0.79、0.88和0.90。仅在轻度MR中发现有教养传递作用。该程序可用于教养和生物遗传分析。     

Transport activity of the sodium bicarbonate cotransporter NBCe1 is enhanced by different isoforms of carbonic anhydrase

Transport metabolons have been discussed between carbonic anhydrase II (CAII) and several membrane transporters. We have now studied different CA isoforms, expressed in Xenopus oocytes alone and together with the electrogenic sodium bicarbonate cotransporter 1 (NBCe1), to determine their catalytic activity and their ability to enhance NBCe1 transport activity. pH measurements in intact oocytes indicated similar activity of CAI, CAII and CAIII, while in vitro CAIII had no measurable activity and CAI only 30% of the activity of CAII. All three CA isoforms increased transport activity of NBCe1, as measured by the transport current and the rate of intracellular sodium rise in oocytes. Two CAII mutants, altered in their intramolecular proton pathway, CAII-H64A and CAII-Y7F, showed significant catalytic activity and also enhanced NBCe1 transport activity. The effect of CAI, CAII, and CAII mutants on NBCe1 activity could be reversed by blocking CA activity with ethoxyzolamide (EZA, 10 μM), while the effect of the less EZA-sensitive CAIII was not reversed. Our results indicate that different CA isoforms and mutants, even if they show little enzymatic activity in vitro, may display significant catalytic activity in intact cells, and that the ability of CA to enhance NBCe1 transport appears to depend primarily on its catalytic activity.     

COMPUTER SUBTRACTION OF BACKGROUND IN AUTORADIOGRAPHY WITH TRITIATED THYMIDINE

An iterative method is suggested to estimate true labelling index even when background and experimental (background + true labelling) grain count distributions appear to be incompatible, generally a result of unavoidably insufficient counting in routine work. A computer program applying the iterative method has been written in Fortran IV for an IBM 1130 machine. This program, available on request, displays grain count distributions before as well as after correction and computes the labelling index and average grain count per labelled cell, with respective variance and standard errors.     

Glycoconjugate expression on the cell wall of tps1/tps1 trehalose-deficient Candida albicans strain and implications for its interaction with macrophages

The yeast Candida albicans has developed a variety of strategies to resist macrophage killing. In yeasts, accumulation of trehalose is one of the principal defense mechanisms under stress conditions. The gene-encoding trehalose-6-phosphate synthase (TPS1), which is responsible for trehalose synthesis, is induced in response to oxidative stress, as in phagolysosomes. Mutants unable to synthesize trehalose are sensitive to oxidative stress in vitro. In mice, the TPS1-deficient strain, tps1/tps1, displays a lower infection rate than its parental strain (CAI4). We have previously demonstrated the reduced binding capacity of tps1/tps1 and its lower resistance to macrophages. At the same time, its outer cell wall layer was seen to be altered. In this study, we show that depending on the culture conditions, the tps1/tps1 strain regulates the carbohydrate metabolism in a different way to CAI4, as reflected by the enhanced β-mannosylation of cell wall components, especially at the level of the 120 kDa glycoprotein species, accessible at the cell surface of tps1/tps1 when cultured in liquid medium, but not on solid medium. This leads to changes in its surface properties, as revealed by decreased hydrophobicity, and the lower levels of ERK1/2 phosphorylation and tumor necrosis factor-α (TNF-α) production in macrophages, thus increasing the resistance to these cells. In contrast, in solid medium, in which over-glycosylation was less evident, tps1/tps1 showed similar macrophage interaction properties to CAI4, but was less resistant to killing, confirming the protective role of trehalose. Thus, the lack of trehalose is compensated by an over-glycosylation of the cell wall components in the tps1/tps1 mutant, which reduces susceptibility to killing.